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ENVIRONMENTAL MICROBIOLOGY

Page number
Unit Name

Unit -1 FUNDAMENTALS OF MICROBIOLOGY 2

Unit 2 - MICROBIAL DIVERSITY AND NUTRIENT 41

TURNOVER

UNIT 3 – METABOLISM OF MICROORGANISM 83

UNIT 4 – MICROBIOLOGY OF WASTEWATER 131

TREATMENT SYSTEM

169
UNIT -5 – TOXICOLOGY

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UNIT -1 – FUNDAMENTALS OF MICROBIOLOGY

DEFINITION OF MICROORGANISM:
An organism that can be seen only through a microscope. Microorganisms include bacteria,
protozoa, algae, and fungi. Although viruses are not considered living organisms, they are
sometimes classified as microorganisms.
CLASSIFICATION OF MICROORGANISM:
A living being is considered when it has the ability to reproduce, respond to environmental
stimuli, and constitutes a living organism. All living organisms need sustenance to survive,
hence they require nutrient supplements. To obtain food mobility plays a major role in an
organism.

The majority of the organisms are visible to us. In these, few are extremely small and can only
be visible to us through a microscope. These are known as microorganisms and are single-
celled or unicellular. These are present in every habitat and are ubiquitous. These play a major
role in maintaining the ecological balance. These dwell inside the body and around every
organism. These are helpful in the maintenance of the health of organisms and even clean
the outside environment. These participate in food production also.

On the other hand, microorganisms are responsible for various dreadful diseases. For
example, we witness the growth of fungus on bread when left unused for a couple of days.
The reason being moist in the bread, making it one of the most favourable conditions for the
breeding of microbes.

Classification of Microorganisms

Microorganisms are categorized into four major groups:

 Bacteria: These are microscopic, single-celled organisms that grow in diverse

environments. These organisms can live in soil, the ocean, and also in the human gut.
These act in a positive way by participating in curdling milk into yoghurt and helping
in digestion.

 Fungi: These are a group of microorganisms that are eukaryotic in nature and these
consist of microorganisms such as yeasts, moulds and mushrooms. These are grouped
into a kingdom and are different from other eukaryotic life kingdoms of plants and
animals.

 Algae: These are simple, non-flowering, and typically aquatic plants of a huge group
that have seaweeds and many single-celled forms included in them. Algae has
chlorophyll in it but lacks true stems, roots, leaves, and vascular tissue. Algology or
Phycology is the study of algae and it has a range of photosynthetic organisms and
many are not closely related. They are a polyphyletic group.

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 Protozoa: Protozoa (also protozoan, plural protozoans) is an informal term for single-
celled eukaryotes, either free-living or parasitic, which feed on organic matter such as
other microorganisms or organic tissues and debris.

Apart from the above microorganisms, viruses are microscopic but differ in their reproduction
aspects as these reproduce only in cells of their hosts. The host organisms are animals,
bacteria, or plants.

Useful Microorganisms

 These are mainly used in the baking industry for the preparation of cakes, bread,
pastry, etc.

 Mainly used in the production of milk products. Example: In the formation of curd
from the milk, lactobacillus bacteria is used

 These are used in the production of alcohol.

 This is used in the preparation of organic acids such as citric acid, lactic acid, fumaric
acid, gluconic acids.

 Steroids are prepared by using microorganisms.

 Vitamins are also produced with the microorganisms such as Vitamin B complex or
Riboflavin by Ashbya gossypii, Eremothecium ashbyii and Clostridium bytyricum. A
known Vitamin C or Ascorbic acid is produced by species Acetobacter.

 Enzymes such as lipase, lactase, amylase, pectinase, penicillinase are synthesized by

microorganisms.

 The soil fertility is increased by the microorganisms and by fixing nitrogen.

 Used in pest control.

Harmful Microorganisms

 Microorganisms grow on food and spoil it.

 Some pathogens are transmitted directly from an infected person.

 Through air, water and food, the pathogens can enter our bodies easily

 Pathogens cause diseases in living beings

 Diseases Caused By Microorganisms

Prokaryotes

Prokaryotes are organisms made up of cells that lack a cell nucleus or any membrane-
encased organelles. This means the genetic material DNA in prokaryotes is not bound within
a nucleus. In addition, the DNA is less structured in prokaryotes than in eukaryotes: in
prokaryotes, DNA is a single loop while in Eukaryotes DNA is organized into chromosomes.
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Most prokaryotes are made up of just a single cell (unicellular) but there are a few that are
made of collections of cells (multicellular).

Scientists have divided the prokaryotes into two groups, the Bacteria, and the Archaea.
Some bacteria, including E Coli, Salmonella, and Listeria, are found in foods and can cause
disease; others are actually helpful to human digestion and other functions. Archaea were
discovered to be a unique life form which is capable of living indefinitely in extreme
environments such as hydrothermal vents or arctic ice.

A typical prokaryotic cell might contain the following parts:

 Cell wall: the membrane surrounding and protecting the cell

 Cytoplasm: all of the material inside a cell except the nucleus
 Flagella and pili: protein-based filaments found on the outside of some prokaryotic
cells
 Nucleoid: a nucleus-like region of the cell where genetic material is kept
 Plasmid: a small molecule of DNA that can reproduce independently

What is the prokaryotic cell structure?

Prokaryotic cells are much smaller than eukaryotic cells, have no nucleus, and lack
organelles. All prokaryotic cells are encased by a cell wall. Many also have a capsule or
slime layer made of polysaccharide. Prokaryotes often have appendages (protrusions) on
their surface.

Eukaryotic Cell Definition

“Eukaryotic cells are the cells that contain a membrane bound nucleus and organelles.”
Table of Contents

 Explanation
 Characteristics
 Structure
 Diagram
 Cell Cycle
 Examples

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What is a Eukaryotic Cell?

Eukaryotic cells have a nucleus enclosed within the nuclear membrane and form large and
complex organisms. Protozoa, fungi, plants, and animals all have eukaryotic cells. They are
classified under the kingdom Eukaryota.
They can maintain different environments in a single cell that allows them to carry out
various metabolic reactions. This helps them grow many times larger than the prokaryotic
cells.
Also refer: Difference between Prokaryotic and Eukaryotic Cells

Characteristics of Eukaryotic Cells

The features of eukaryotic cells are as follows:
1. Eukaryotic cells have the nucleus enclosed within the nuclear membrane.
2. The cell has mitochondria.
3. Flagella and cilia are the locomotory organs in a eukaryotic cell.
4. A cell wall is the outermost layer of the eukaryotic cells.
5. The cells divide by a process called mitosis.
6. The eukaryotic cells contain a cytoskeletal structure.
7. The nucleus contains a single, linear DNA, which carries all the genetic information.

Structure Of Eukaryotic Cell

The eukaryotic cell structure comprises the following:

Plasma Membrane

 The plasma membrane separates the cell from the outside environment.
 It comprises specific embedded proteins, which help in the exchange of substances
in and out of the cell.
 Cell Wall

 A cell wall is a rigid structure present outside the plant cell. It is, however, absent in
animal cells.
 It provides shape to the cell and helps in cell-to-cell interaction.
 It is a protective layer that protects the cell from any injury or pathogen attacks.
 It is composed of cellulose, hemicellulose, pectins, proteins, etc.

Cytoskeleton
The cytoskeleton is present inside the cytoplasm, which consists of microfilaments,
microtubules, and fibres to provide perfect shape to the cell, anchor the organelles, and
stimulate the cell movement.

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Endoplasmic Reticulum
It is a network of small, tubular structures that divides the cell surface into two parts:
luminal and extraluminal.
Endoplasmic Reticulum is of two types:
 Rough Endoplasmic Reticulum contains ribosomes.
 Smooth Endoplasmic Reticulum that lacks ribosomes and is therefore smooth.

Nucleus

 The nucleoplasm enclosed within the nucleus contains DNA and proteins.
 The nuclear envelop consists of two layers- the outer membrane and the inner
membrane. Both the membranes are permeable to ions, molecules, and RNA
material.
 Ribosome production also takes place inside the nucleus.

Golgi Apparatus

 It is made up of flat disc-shaped structures called cisternae.

 It is absent in red blood cells of humans and sieve cells of plants.
 They are arranged parallel and concentrically near the nucleus.
 It is an important site for the formation of glycoproteins and glycolipids.

Ribosomes
These are the main site for protein synthesis and are composed of proteins and ribonucleic
acids.

Mitochondria

 These are also known as “powerhouse of cells” because they produce energy.
 It consists of an outer membrane and an inner membrane. The inner membrane is
divided into folds called cristae.
 They help in the regulation of cell metabolism.

Lysosomes
They are known as “suicidal bags” because they possess hydrolytic enzymes to digest
protein, lipids, carbohydrates, and nucleic acids.

Plastids
These are double-membraned structures and are found only in plant cells. These are of
three types:
 Chloroplast that contains chlorophyll and is involved in photosynthesis.

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 Chromoplast that contains a pigment called carotene that provides the plants
yellow, red, or orange colours.
 Leucoplasts that are colourless and store oil, fats, carbohydrates, or proteins.

Eukaryotic Cell Diagram

Eukaryotic cell diagram mentioned below depicts different cell organelles present in
eukaryotic cells. The nucleus, endoplasmic reticulum, cytoplasm, mitochondria, ribosomes,
lysosomes are clearly mentioned in the diagram.
Explore more about Cell organelles

Eukaryotic Cell Diagram illustrated above shows the presence of a true nucleus.

Eukaryotic Cell Cycle

The eukaryotic cells divide during the cell cycle. The cell passes through different stages
during the cycle. There are various checkpoints between each stage.

Quiescence (G0)
This is known as the resting phase, and the cell does not divide during this stage. The cell
cycle starts at this stage. The cells of the liver, kidney, neurons, and stomach all reach this
stage and can remain there for longer periods. Many cells do not enter this stage and divide
indefinitely throughout their lives.

Interphase
In this stage, the cells grow and take in nutrients to prepare them for the division. It consists
of three
checkpoints:
Gap 1 (G1) – Here the cell enlarges. The proteins also increase.
Synthesis (S) – DNA replication takes place in this phase.

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Gap 2 (G2) – Ther cells enlarge further to undergo mitotic division.

Mitosis
Mitosis involves the following stages:
 Prophase
 Prometaphase
 Metaphase
 Anaphase
 Telophase
 Cytokinesis
On division, each daughter cell is an exact replica of the original cell.

Examples of Eukaryotic Cells

Eukaryotic cells are exclusively found in plants, animals, fungi, protozoa, and other complex
organisms. The examples of eukaryotic cells are mentioned below:

Plant Cells
The cell wall is made up of cellulose, which provides support to the plant. It has a large
vacuole which maintains the turgor pressure. The plant cell contains chloroplast, which aids
in the process of photosynthesis.

Fungal Cells
The cell wall is made of chitin. Some fungi have holes known as septa which allow the
organelles and cytoplasm to pass through them.

Animal Cells
These do not have cell walls. Instead, they have a cell membrane. That is why animals have
varied shapes. They have the ability to perform phagocytosis and pinocytosis.

Protozoa
Protozoans are unicellular organisms. Some protozoa have cilia for locomotion. A thin layer
called pellicle provides supports to the cell.
For more information on Eukaryotic Cells, its definition, characteristics, structure, and
examples, keep visiting BYJU’S website or download BYJU’S app for further reference.

Are eukaryotic cells unicellular or multicellular?

Eukaryotic cells may be unicellular or multicellular. Paramecium, Euglena, Trypanosoma,
Dinoflagellates are unicellular eukaryotes. Plants and animals are multicellular eukaryotes.

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What is the most important characteristic of eukaryotic cells that distinguishes it from
prokaryotic cells?
Eukaryotic cells have a membrane-bound nucleus. On the contrary, prokaryotic cells lack a
true nucleus, i.e., they have no nuclear membrane. Unlike eukaryotic cells, the prokaryotic
cells do not have mitochondria, chloroplast and endoplasmic reticulum.

Are viruses eukaryotes?

Viruses are neither eukaryotes nor prokaryotes. Since viruses are a link between living and
non-living they are not considered in either category.

What are the salient features of a eukaryotic cell?

A eukaryotic cell has the following important features:

 A eukaryotic cell has a nuclear membrane.

 It has mitochondria, Golgi bodies, cell wall.
 It also contains locomotory organs such as cilia and flagella.
 The nucleus has a DNA that carries all the genetic information.

How does a eukaryotic cell divide?

A eukaryotic cell divides by the process of mitosis. It undergoes the following stages during
cell division:

 Prophase
 Metaphase
 Anaphase
 Telophase
 Cytokinesis

When did the first eukaryotic cell evolve?

The first eukaryotic cells evolved about 2 billion years ago. This is explained by the
endosymbiotic theory that explains the origin of eukaryotic cells by the prokaryotic
organisms. Mitochondria and chloroplasts are believed to have evolved from symbiotic
bacteria.

What is the evidence for endosymbiotic theory?

The first evidence in support of the endosymbiotic theory is that mitochondria and
chloroplast have their own DNA and this DNA is similar to the bacterial DNA. The organelles
use their DNA to produce several proteins and enzymes to carry out certain activities.

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Prokaryotic Cell Definition

“Prokaryotic cells are the cells that do not have a true nucleus and membrane-bound
organelles.”

Table of Contents

 Explanation
 Characteristics
 Structure
 Diagram
 Components
 Reproduction
 Examples

What is a Prokaryotic Cell?

Prokaryotic cells are single-celled microorganisms known to be the earliest on earth.
Prokaryotes include Bacteria and Archaea. The photosynthetic prokaryotes include
cyanobacteria that perform photosynthesis.
A prokaryotic cell consists of a single membrane and therefore, all the reactions occur
within the cytoplasm. They can be free-living or parasites.

Characteristics of Prokaryotic Cell

Prokaryotic cells have different characteristic features. The characteristics of the prokaryotic
cells are mentioned below.
1. They lack a nuclear membrane.
2. Mitochondria, Golgi bodies, chloroplast, and lysosomes are absent.
3. The genetic material is present on a single chromosome.
4. The histone proteins, the important constituents of eukaryotic chromosomes, are
lacking in them.
5. The cell wall is made up of carbohydrates and amino acids.
6. The plasma membrane acts as the mitochondrial membrane carrying respiratory
enzymes.
7. They divide asexually by binary fission. The sexual mode of reproduction involves
conjugation.

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Prokaryotic Cell Structure

A prokaryotic cell does not have a nuclear membrane. However, the genetic material is
present in a region in the cytoplasm known as the nucleoid. They may be spherical, rod-
shaped, or spiral. A prokaryotic cell structure is as follows:
1. Capsule– It is an outer protective covering found in the bacterial cells, in addition to
the cell wall. It helps in moisture retention, protects the cell when engulfed, and
helps in the attachment of cells to nutrients and surfaces.
2. Cell Wall– It is the outermost layer of the cell which gives shape to the cell.
3. Cytoplasm– The cytoplasm is mainly composed of enzymes, salts, cell organelles and
is a gel-like component.
4. Cell Membrane– This layer surrounds the cytoplasm and regulates the entry and exit
of substances in the cells.
5. Pili– These are hair-like outgrowths that attach to the surface of other bacterial cells.
6. Flagella– These are long structures in the form of a whip, that help in the locomotion
of a cell.
7. Ribosomes– These are involved in protein synthesis.
8. Plasmids– Plasmids are non-chromosomal DNA structures. These are not involved in
reproduction.
9. Nucleoid Region– It is the region in the cytoplasm where the genetic material is
present.
A prokaryotic cell lacks certain organelles like mitochondria, endoplasmic reticulum, and
Golgi bodies.

Prokaryotic Cell Diagram

The prokaryotic cell diagram given below represents a bacterial cell. It depicts the absence
of a true nucleus and the presence of a flagellum that differentiates it from a eukaryotic cell.

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Prokaryotic Cell Diagram illustrates the absence of a true nucleus

Components of Prokaryotic Cells

The prokaryotic cells have four main components:
Plasma Membrane- It is an outer protective covering of phospholipid molecules which
separates the cell from the surrounding environment.
Cytoplasm- It is a jelly-like substance present inside the cell. All the cell organelles are
suspended in it.
DNA- It is the genetic material of the cell. All the prokaryotes possess a circular DNA. It
directs what proteins the cell creates. It also regulates the actions of the cell.
Ribosomes- Protein synthesis occurs here.
Some prokaryotic cells possess cilia and flagella which helps in locomotion.

Reproduction in Prokaryotes
A prokaryote reproduces in two ways:
 Asexually by binary fission
 Sexually by conjugation

Binary Fission
1. The DNA of an organism replicates and the new copies attach to the cell membrane.
2. The cell wall starts increasing in size and starts moving inwards.
3. A cell wall is then formed between each DNA, dividing the cell into two daughter
cells.

Recombination

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In this process, genes from one bacteria are transferred to the genome of other bacteria. It
takes place in three ways-conjugation, transformation, transduction.
 Conjugation is the process in which genes are transferred between two bacteria
through a protein tube structure called a pilus.
 Transformation is the mode of sexual reproduction in which the DNA from the
surroundings is taken by the bacterial cell and incorporated in its DNA.
 Transduction is the process in which the genetic material is transferred into the
bacterial cell with the help of viruses. Bacteriophages are the virus that initiates the
process.

Examples of Prokaryotic Cells

The examples of the prokaryotic cells are mentioned below:

Bacterial Cells
These are unicellular organisms found everywhere on earth from soil to the human body.
They have different shapes and structures.
The cell wall is composed of peptidoglycan that provides structure to the cell wall.
Bacteria have some unique structures such as pili, flagella and capsule.
They also possess extrachromosomal DNA known as plasmids.
They have the ability to form tough, dormant structures known as endospores that helps
them to survive under unfavourable conditions. The endospores become active when the
conditions are favourable again.

Archaeal Cells
Archaebacteria are unicellular organisms similar to bacteria in shape and size.
They are found in extreme environments such as hot springs and other places such as soil,
marshes, and even inside humans.
They have a cell wall and flagella. The cell wall of archaea does not contain peptidoglycan.
The membranes of the archaea have different lipids with a completely different
stereochemistry.
Just like bacteria, archaea have one circular chromosome. They also possess plasmids.
For more information on Prokaryotic Cells, its definition, structure, characteristics and
examples, keep visiting BYJU’S Biology website or download BYJU’S app for further
reference.

What are the structural features of prokaryotic cells?

The prokaryotic cell structure is composed of:

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 Cell wall
 Cell membrane
 Capsule
 Pili
 Flagella
 Ribosomes
 Plasmids

How is the prokaryotic cell structure different from that of the eukaryotic cell?
Prokaryotic cells lack a true nucleus. The nucleus is devoid of the nuclear membrane. On the
contrary, the nucleus of the eukaryotic cells is enclosed by a nuclear membrane. A
prokaryotic cell also lacks mitochondria and chloroplast, unlike a eukaryotic cell.

How does a prokaryotic cell divide?

Prokaryotic cells undergo asexual reproduction. Most prokaryotic cells divide by binary
fission, where the cells divide into two daughter cells.

Why is the process of cell division in prokaryotic cells different from that in eukaryotes?
Prokaryotic cells are simpler than eukaryotic cells. They do not have a nuclear membrane
surrounding their DNA, therefore, cell division is different than that in eukaryotes.

When did the prokaryotic cells evolve?

The first prokaryotic cells evolved around 3.5 billion years ago. The eukaryotic cells were
formed after the prokaryotic cells and are believed to have evolved from them.
AIR, SOIL & WATERBORNE MICROORGANISMS IN FOODS

Introduction

Microorganisms are present everywhere in our environment, in soil, air, food and water.
Also called microbes, microorganisms are living organisms, generally observable only
through a microscope. Bacteria live in almost any warm, moist environment, and there are
thousands of different kinds. They are single-cell organisms that can reproduce very quickly
and can spread through food, air or blood.

The transfer of contamination through the airborne route, soil and water body is one of the
most significant areas of high-care food production. The food industry specially the
manufacturing of the chilled meat products strive for lower levels of the air contamination,
therefore lot of experimental and numerical studies considers the concentration of airborne
particulate contaminants, such as different species of food spoilage microorganisms. The

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risk is higher when air is contaminated with eventually foodborne pathogen microorganisms
and spores.

Airborne microbes are biological airborne contaminants (also known as bio aerosols) like
bacteria, viruses or fungi as well as airborne toxins passed from one victim to the next
through the air, without physical contact, causing irritation at the very least. This usually
happens when an infected subject sneezes, coughs, or just plain breathes. It is hard to
prevent such a method of transmission. Airborne microbes are a major cause of respiratory
ailments. Mainly Bacillus, Micrococcus, and Staphylococcus are the microorganisms which
are present in air.

Soil and water are common sources of important pathogenic and spoilage microorganisms,
which is why it is important to thoroughly wash raw foods with good quality water. Air and
dust are important sources of microorganisms during food processing and can influence
food quality. Bacillus subtilis is mostly present in soil. Apart from this nitrogen-fixing
bacteria, nitrifying bacteria, denitrifying bacteria and actinomycetes are also present in a
huge quantity in soil.
Although most soil and water borne microbes will contaminate plants, very few types
actually persist on them. Those that persist, such as lactic acid bacteria and some yeasts,
must be able to adhere to the plant material and to utilize it for growth. Everyone has their
own natural microorganisms that live on, in and around their own bodies. These bacteria
are known as natural flora and our own bodies recognize that they are good for us. We, as
humans, would not survive without such creatures.

Airborne microorganisms:

Bacteria have no active mechanisms for becoming airborne. They are dispersed on dust
particles disturbed by physical agencies, in minute droplets of water generated by any
process which leads to the formation of an aerosol. Many actinomycetes, especially those in
the genus Streptococcus undergo this process. Air is mainly transport medium for
microorganisms. They occur in small numbers in air when compared with soil or water.
The bacteria in the air consist both "good" (non pathogenic) and "bad" (pathogenic)
bacteria, but most of them are good bacteria, and the levels of bad bacteria are low. The
microflora of air can be studied under two headings; outdoor and indoor microflora.
Environmental factors that affect air microflora include atmospheric temperature (There is a
progressive increase in the death rate with an increase in temperature from -18°C to 49°C),
humidity (Low and high relative humidity cause the death of most microorganisms) and air
current.

The majority of the airborne bacteria belonged to the genera Bacillus, Micrococcus, and
Staphylococcus, but a total of 37 different genera were identified in the air. These results
suggest that anthropogenic sources are major contributors to airborne bacteria:

Gram positive bacilli include:

Clostridium species
Cornybacterium species

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Gram negative bacilli include:

Salmonella species
Escherichia species
Pseudomonas species
Bacteroides species

Various Micrococcus like:

Micrococcus antarcticus; M. luteus; M. lylae; M. roseus; M. sp.

Concentration of airborne microorganisms:

The air around us is filled with microbes. Bacteria, fungi, algae, protozoa, and viruses float in
air currents. The numbers of microbes in the air range from 10 to 10,000 per cubic meter.
They are found easily up to 3000 meters and as high as six miles into the air. According to a
study, in Marseilles, concentrations of airborne viable microorganisms averaged 791 ± 598
bacteria m−3 (with a geometric mean of 536 ± 103 bacteria m−3) and 92 ± 92 fungi m−3
(with a geometric mean of 63 ± 15 fungi m−3). Airborne microflora, which increased a log-
normal distribution in Marseilles, was 3 shown to have a large variability. Airborne bacteria
increase with temperature and wind velocity whereas airborne fungi increase with
temperature and varied with wind direction in urban and natural areas. Partial identification
of bacteria in Marseilles Island showed that geographical location Outdoor airborne
microflora was investigated in urban and natural areas, the city of Marseilles had qualitative
as well as quantitative influence on airborne microflora, this was illustrated by an increase
of global airborne microorganisms, and more particularly Gram negative bacteria, in the
urban area.

Morphology:

The word bacillus may be used to describe any rod-shaped bacterium, and such bacilli are
found in many different taxonomic groups of bacteria. Bacilli are usually solitary, but can
combine to form diplobacilli, streptobacilli, and palisades.
Micrococci have Gram-positive spherical cells ranging from about 0.5 to 3 micrometers in
diameter and typically appear in tetrads. They are catalase positive, oxidase positive, indole
negative and citrate negative. Micrococcus has a substantial cell wall, which may comprise
as much as 50% of the cell mass. The genome of Micrococcus is rich in guanine and cytosine
(GC), typically exhibiting 65 to 75% GC-content.
Micrococci can grow well in environments with little water or high salt concentrations. Most
are mesophiles; some, like Micrococcus antarcticus (found in Antarctica) are psychrophiles.
Though not a spore former, Micrococcus cells can survive for an extended period of time:
unprotected cultures of soil micrococci have been revived after storage in a refrigerator for
10 years. Recent work by Greenblat et al. demonstrates that Micrococcus luteus has
survived for at least 34,000 to 170,000 years on the basis of 16S rRNA analysis, and possibly
much longer.

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Soil borne microorganisms:

There is enough evidence in the literature to believe that microorganisms were the earliest
of the living things that existed on this planet. Man depends on crop plants for his existence
and crop plants in turn depend on soil and soil microorganisms for their nutrition. Scientists,
from the beginning, studied the microorganisms from water, air, soil etc. and recognized the
role of microorganisms in natural processes and realized the importance of soil
microorganisms in growth and development of plants. Soil is considered to be the main
source of scavenging the organic wastes through microbial action and is also a rich store
house for industrial micro flora of great economic importance.
Soil microbiology emerged as a distinct branch of soil science during first half of the 19th
century. Some of the notable contributions made by several scientists in field of soil
microbiology are highlighted. J. B. Boussingault (1838) showed that leguminous plants can
fix atmospheric nitrogen 4 and increase nitrogen content in the soil. S. N. Winogradsky
discovered the autotrophic mode of life among bacteria and established the microbiological
transformation of nitrogen and sulphur. Isolated for the first time nitrifying bacteria and
demonstrated role of these bacteria in nitrification (l890), further he demonstrated that
free-living Clostridium pasteuriamum could fix atmospheric nitrogen (1893). He is known as
“Father of soil microorganism”. B. Frank, i) discovered (1880) an actinomycetes “Frankia”
(Actinorhizal symbiosis) inducing root nodules in non-legumes tress of genera Alnus sp and
Casurina growing in temperate forests, ii) coined (1885) the term " Mycorrhiza" to denote
association of certain fungal symbionts with plant roots (Mycorrhiza-A symbiotic association
between a fungus and roots of higher plants. Renamed the genus Bacillus as Rhizobium
(1889).

Bacteria in soil

Various microorganisms present in soil:

Nitrogen-fixing bacteria form symbiotic associations with the roots of legumes like clover
and lupine, and trees such as alder and locust. Visible nodules are created where bacteria
infect a growing root hair. The plant supplies simple carbon compounds to the bacteria, and
the bacteria convert nitrogen (N2) from air into a form the plant host can use. When leaves
or roots from the host plant decompose, soil nitrogen increases in the surrounding area. For
example:- Cyanobacteria, Rhizobia.
Nitrifying bacteria change ammonium (NH4+) to nitrite (NO2-) then to nitrate (NO3-), a
preferred form of nitrogen for grasses and most row crops. Nitrate is leached more easily
from the soil, so some farmers use nitrification inhibitors to reduce the activity of one type
of nitrifying bacteria. Nitrifying bacteria are suppressed in forest soils, so that most of the
nitrogen remains as ammonium. Nitrifying bacteria involves, Nitrosomonas, Nitrobacter,
Nitrosococcus.
Denitrifying bacteria convert nitrate to nitrogen (N2) or nitrous oxide (N2O) gas.
Denitrifiers are anaerobic, meaning they are active where oxygen is absent, such as in
saturated soils or inside soil aggregates.For example:- Thiobacillus denitrificans, Micrococcus
denitrificans, Paracoccus denitrificans.

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Actinomycetes are a large group of bacteria that grow as hyphae like fungi . They are
responsible for the characteristically "earthy" smell of freshly turned, healthy soil.
Actinomycetes decompose a wide array of substrates, but are especially important in
degrading recalcitrant (hard-to-decompose) compounds, such as chitin and cellulose, and
are
5 active at high pH levels. Fungi are more important in degrading these compounds at low
pH. A number of antibiotics are produced by actinomycetes such as Streptomyces.

Example of nitrogen fixing bacteria

Rhizobia are one of the groups of microorganisms living in soil. They are single celled
bacteria, approximately one thousandth of a millimetre in length. Rhizobia belong to a
family of bacteria called Rhizobiaceae. There are a number of groups of bacteria in this
family.
Rhizobia belong to a specific group of bacteria that form a mutually beneficial association,
or symbiosis, with legume plants. These bacteria take nitrogen from the air (which plants
cannot use) and convert it into a form of nitrogen called ammonium (NH4+), which plants
can use. The nitrogenase enzyme controls the process, called nitrogen fixation, and these
bacteria are often called "nitrogen fixers".
Rhizobia are found in soils of many natural ecosystems. They may also be present in
agricultural areas where they are associated with both crop legumes (like soybean) and
pasture legumes (like clover). Usually, the rhizobia in agricultural areas have been
introduced at sowing by applying an inoculum to the exterior of the seeds as liquid
formations or pellets
The presence of numerous genera of spoilage bacteria, yeasts and molds, and an occasional
pathogen on fresh produce has been recognized for many years. Several outbreaks of
human gastroenteritis have been linked to the consumption of contaminated fresh
vegetables and, to a lesser extent, fruits. Salads containing raw vegetables have been
identified as vehicles of traveler's diarrhea, an illness sometimes experienced by visitors to
developing countries. Enterotoxigenic Escherichia coli is the most common cause of this
illness. Outbreaks of salmonellosis in humans have been attributed to consumption of
contaminated tomatoes, mustard cress, bean sprouts, cantaloupe, and watermelon.

Fungi in soil:

Soil fungi are considered to be an important food source for earthworms. Fungal species
(Cladosporium cladosporioides, Rhizoctonia solani, Mucor sp., Trichoderma viride, Fusarium
nivale, Phlebia radiata, Glaeophyllum trabeum, Coniophora puteana, Coriolus versicolor),
followed by fast-growing species such as Mucor sp. and R. solan.

Most natural antibiotics come from soil fungi and bacteria:

Many microorganisms have been playing a significant role long before they were discovered
by man. Today, soil is considered to be the main source of scavenging the organic wastes
through microbial action and is also a rich store house for industrial microflora of great
economic importance. Soil bacteria and fungi live by digesting and recycling dead plant
material such as leaves and seed. This 6 material is typically carbon-rich and nitrogen-poor.

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Most common antibiotics are carbon-rich polymers made by enzymes that strongly
resemble those that normally make saturated fats. The building blocks of these polymers
are often exactly the same as those used to make saturated fats. Antibiotics are not that
easy to find in microbes. Bacteria of the Bacillus genus occur mainly in soil and produce
many widely studied antibiotic compounds. For example, Bacillus subtilus produces more
than seventy-five known antibiotics consisting predominantly of small, cyclic peptides, but
also including phospholipid, lipopeptide, and aminosugar antibiotics. It was hypothesized
that antibiotic-producing bacteria would be of the genera Bacillus or Streptomyces and that
the antibiotics would be peptides that inhibited the growth of Gram-positive bacteria.

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Waterborne microorganisms:

There are many types of watery environments ranging from freshwater ponds, streams,
puddles, lakes, rivers, and swamps to salty seas with three times the salt concentration of
the ocean. Microbes live in overgrown slime, on pipes and in open oceans with few
nutrients to support microbial life. Microbes thrive in streams with lots of oxygen to murky
bogs that have no oxygen. In ponds there is a rich thriving ecosystem of microbial life
including green and purple bacteria and algae, sulfate reducers, methane producers, and
others. Many microbes live in the bottom of lakes and rivers in sediments. Many microbes
cannot survive except in the presence of high concentrations of salt.
The largest watery place on earth is the ocean. Oceans cover 71% of the Earth's surface and
are responsible for producing about half of the world's organisms, which includes the plants,
animals, fungi, and microbes. Most life in the oceans lives at the sunlit ocean surface. Below
25 meters there is little light in the ocean, and life productivity decreases. As well as little
light, deeper waters are cooler, which supports less life. Below 50 meters, the temperature
is less than 10 degrees Celsius.

Microorganisms found in water:- water is also a habitat for various types of

microorganisms, such as, bacteria, viruses, fungi and protozoans. Campylobacter, Cholera,
Cryptosporidium, Escherichia coli, Giardia, Hepatitis, Legionella, Protozoan parasite,
Salmonella, Shigella are commonly found in water.

Morphology:-

Campylobacter: These are gram negative, non-spore forming, microaerophillic bacteria.

They are found in spiral and coccoid form and are distinguished from others due to their
darting motility. Spiral form is found in young cultures and coccoid form is found in old
cultures.eg: C. jejuni, C.coli, C. laridis. 7

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Vibrio cholera: It is a gram- negative, , comma-shaped bacterium.V. cholerae is facultatively

anaerobic and has a flagellum at one cell pole. V. cholerae was first isolated as the cause of
cholera by Italian anatomist Filippo Pacini in 1854.

Cryptospordium: There are two species which are morphologically indistinguishable by light
microscope examination namely Cryptosporidium hominis, Cryptosporidium parvum.
Oocysts size ranges from 4-6 μm. It is round or oval in shape.

Giardia: It has two morphological stages: the trophozoite and the cyst.
The trophozoite is pear shaped, with a broad anterior and much attenuated posterior. It is
10-12μm long and 5-7μm wide, bilaterally symmetrical, and has two nuclei. It is also
relatively flattened, with a large sucking disk on the anterior ventral side, which serves as
the parasite’s method of attachment to the mucosa of the host. The trophozoite also has
two median bodies and four pairs of flagella (anterior, caudal, posterior and ventral).
The G. lamblia cyst is egg-shaped, and measures 8-14μm by 7-10μm. After encystation,
each organelle duplicates, so each cyst contains four nuclei, four median bodies, eight pairs
of flagella--although these organelles are not arraigned in any clear pattern. Upon
excystation, each cyst produces two trophozoites.

Hepatitis: Among the smallest and structurally simplest of the RNA animal viruses. The
virion is non enveloped and, with a diameter of 27-32 nm, it is composed entirely of viral
protein and RNA. Electron microscopy analyses show particles with icosahedral symmetry
although no structural details could be discerned. Morphologically, Hepatitis A virus
particles are indistinguishable from other picornaviruses.

Fungi: These are a diverse group of organisms belonging to the kingdom Eumycota. This
kingdom comprises five phyla, namely Ascomycota, Basidiomycota, Zygomycota,
Chytridiomycota, and Glomeromycota. As a practical approach to classification, fungi have
been divided into groups, such as the filamentous fungi, also called moulds, the yeasts, and
the mushrooms. Some fungi are primarily adapted to aquatic environments, and will,
therefore, naturally be found in water. These fungi are zoosporic, and many belong in phyla
Chytridiomycota. Fungi belonging to the other phyla in Eumycota are primarily adapted to
terrestrial environments.
5.1. Morphology:
Fungi are composed of filaments called hyphae; their cells are long and thread-like and
connected end-to-end. Because of this diffuse association of their cells, the body of the
organism is given the special name mycelium, a term which is applied to the whole body of
any fungus. When 8

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reproductive hyphae are produced, they form a large organized structure called a
sporocarp, or mushroom. This is produced solely for the release of spores, and is not the
living, growing portion of the fungus.
In addition to being filamentous, fungal cells often have multiple nuclei. In the chytrids and
zygomycetes, the cells are coenocytic, with no distinction between individual cells. Another
feature of fungi is the presence of chitin in their cell walls. The chitin adds rigidity and
structural support to the thin cells of the fungus, and makes fresh mushrooms crisp. Most
members of the kingdom Fungi lack flagella; the structures are completely absent in all
stages of their life cycle.

Ascomycota: Ascomycota is phylum of the kingdom fungi and, together with the
Basidiomycota, form the sub kingdom Dikarya. Its members are commonly known as the sac
fungi. They are the largest phylum of Fungi, with over 64,000 species. The defining feature
of this fungal group is the "ascus", a microscopic in which nonmotile spores, called
ascospores, are formed. The ascomycetes are a monophyletic group.

Zygomycota: The unique characteristic of the Zygomycota is the zygospore. Zygospores are
formed within a zygosporangium after the fusion of specialized hyphae called gametangia
during the sexual cycle. Single zygospore is formed per zygosporangium. Because of this
one-to-one relationship, the terms are often used interchangeably. The mature zygospore is
often thick-walled and undergoes an obligatory dormant period before germination. Most
Zygomycota are thought to have a zygotic or haplontic life cycle. Thus, the only diploid
phase takes place within the zygospore. Nuclei within the zygospore are believed to
undergo meiosis during germination, but this has only been demonstrated genetically within
the model eukaryote Phycomyces blakesleeanus.

Afterword
Microorganisms play a key role in maintaining life on earth, fixing gases and breaking down
dead plant and animal matter into simpler substances that are used at the beginning of the
food chain. Biotechnologists can also exploit the activities of microbes to benefit humans,
such as in the production of medicines, enzymes and food. They are also used to breakdown
sewage and other toxic wastes into safe matter. However, the above discussed pathogens
and the types of disease caused by them makes clear that they are also agents of different
diseases and these infectious agents not only spread through contaminated water and
spoiled food but also through the omnipresent air and people with weak immune system
becomes the first victims of these diseases. Control of these air, soil and water borne
pathogens is 9 important, and today many organisations like WHO is taking care of it. Many
resistant bugs are found and controlling them is a new challenge for science. Strict adhesion
to regulations and other standards set for basic hygiene maintenance and antibiotic use is a
necessity

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An airborne disease is caused by droplets of pathogens which are expelled into the air by
coughing, sneezing or talking. The relevant pathogens may be viruses, bacteria, or fungi.
Many common infections can spread by airborne transmission are tuberculosis, influenza,
small pox. Waterborne diseases are caused by pathogenic microorganisms and most
commonly transmitted through contaminated fresh water. Infections can be spread by
bathing, washing, drinking, in the preparation of food, or the consumption of food thus
infected.

TRANSMISSIBLE DISEASE

A communicable disease is any disease that passes between people or animals. People
sometimes refer to communicable diseases as “infectious” or “transmissible” diseases.

Pathogens, including bacteria, viruses, fungi, and protists, cause communicable diseases.

A person may develop a communicable disease after becoming infected by the pathogen.
This may happen through:

direct contact with a person carrying the pathogen

contact with contaminated fluids, such as blood, mucus, or saliva

inhaling contaminated droplets from another person’s cough or sneeze

receiving a bite from an animal or insect carrying the pathogen

consuming contaminated water or foods

Once a pathogen has entered a person’s body, it will begin replicating. The individual may
then begin to experience symptoms.

Some symptoms are a direct result of the pathogen damaging the body’s cells. Others are
due to the body’s immune response to the infection.

Communicable diseases are usually mild, and symptoms pass after a few days. However,
some can be serious and potentially life threatening.

Types and symptoms

Four main types of pathogens cause infection: Viruses, bacteria, fungi, and protists.

Viruses

Viruses are tiny pathogens that contain genetic material. Unlike other pathogens, they lack
the complex structure of a cell. To replicate, they must enter the cells of other living beings.
Once inside, they use the cell’s machinery to make copies of themselves.

Some different viruses include:

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Rhinoviruses

Rhinoviruses are a group of viruses that are responsible for the common cold. Symptoms of
a cold may include:

a stuffy or runny nose

sore throat

headache

A person can catch a rhinovirus by inhaling contaminated droplets from the cough or sneeze
of another person.

Similarly, rhinoviruses spread by people touching their nose, eyes, or mouth after touching
items or surfaces that have come into contact with the virus.

Influenza

Influenza viruses are infections that attack the respiratory system. Some potential
symptoms include:

fever or chills

stuffy or runny nose

sore throat

cough

headaches

muscle or body aches

fatigue

A person can catch influenza viruses in the same way they may catch rhinoviruses.

HIV

HIV attacks the immune system of its host. This makes the person vulnerable to other
infections and diseases.

A person can contract HIV as a result of contact with blood or other body fluids containing
the virus.

The symptoms of HIV may develop gradually and in stages. They can include:

fever

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chills

rash

mouth sores

sore throat

swollen lymph nodes

night sweats

muscle aches

fatigue

The only way a person can be certain they have HIV is to have an HIV test.

Although there is no cure for HIV, medications can help to keep the virus under control.
Without such treatment, HIV can develop into AIDS.

Bacteria

Bacteria are microscopic, single celled organisms. They exist in almost every environment on
earth, including inside the human body.

Many bacteria are harmless, and some help the body to function. However, bacteria can
also cause infections that damage the body.

Some different types of bacterial infection include:

Salmonella and Escherichia coli

Salmonella and Escherichia coli (E. coli) are two different types of bacteria that can infect
the digestive system.

They typically spread through contaminated foods, such as uncooked meats, and unwashed
fruits and vegetables.

Some symptoms of these infections include:

abdominal cramps

diarrhea

fever

headache

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Tuberculosis

Tuberculosis (TB) is a bacterial infection that primarily attacks the lungs. It may cause the
following symptoms:

a cough continuing for more than 3 weeks

loss of appetite

unintentional weight loss

fever

chills

night sweats

A person can catch TB by inhaling tiny droplets or “aerosols” from the cough or sneeze of a
person who has the infection. However, the American Lung Association state that while TB
is contagious, it does not easily spread from person to person.

Fungi

Fungi are a type of organism that includes yeasts, molds, and mushrooms. There are millions
of different fungi, but only around 300 cause harmful illnesses.

Fungal infections can occur anywhere in the body, but they commonly affect the skin and
mucus membranes. Some different types of fungal infection include:

Ringworm

Ringworm is a common fungal infection of the skin. The characteristic symptom of ringworm
is a red or silver ring shaped rash. It may be dry, scaly, or itchy.

People may contract ringworm in the following ways through close contact with a person
who has ringworm. Alternatively, they can catch it from sharing towels, bedding, or other
personal items with a person who has ringworm.

Without treatment, ringworm may spread to other parts of the body.

Athlete’s foot

Athlete’s foot is a common fungal infection that affects the skin on the feet. It typically
causes sore or itchy white patches between the toes.

People can contract athlete’s foot through direct contact with someone who has the fungus,
or surfaces that have been in contact with the fungus.

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For example, an individual might contract athlete’s foot after walking barefoot in locker
rooms, showers, or swimming pools.

Protists

Protists are microscopic organisms that typically consist of a single cell.

Some protists are parasitic, meaning they live on or inside another organism and use the
organism’s nutrients for their own survival. Parasitic protists can cause various diseases.

The protist Plasmodium causes the tropical disease malaria. The parasite can pass from
person to person through mosquito bites.

Malaria causes symptoms such as:

fever and chills

headaches

vomiting

diarrhea

muscle pains

Without proper treatment, malaria can be life threatening.

How to prevent transmission

People can reduce their risk of contracting or transmitting disease causing pathogens by
following the steps below:

washing their hands thoroughly and regularly

disinfecting surfaces at home often, especially doorknobs and food areas

practicing good hygiene when preparing and handling food

avoiding eating spoiled food

avoiding touching wild animals

receiving available vaccinations

taking antimalarial medications when traveling where there is a malaria risk

Treatment

Some communicable diseases cause only mild symptoms that disappear without treatment.
Others may cause severe symptoms, or potentially life threatening complications.

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The treatment for such diseases depends on whether they are bacterial, viral, or fungal.

Viral infections

Vaccines are a highly effective method for preventing specific viral infections.

When a person receives a vaccine, they are receiving a dead or inactive form of the virus.
The immune system responds by producing antibodies capable of killing an active form of
the virus in the future.

If a person already has a virus, they may require antiviral medications to keep the virus
under control.

Bacterial infections

A person who has a bacterial infection may require a course of antibiotics to help control
the infection. These drugs work by killing off the bacteria or preventing them from
replicating.

Fungal infections

A severe or chronic fungal infection may require over-the-counter or prescription antifungal

medications. These are available in both oral and topical forms.

Summary

Communicable diseases are diseases that can pass from person to person. The pathogens
that cause these diseases can spread in various ways, such as through the air, contact with
contaminated substances or surfaces, or from animal and insect bites.

Many communicable diseases cause mild symptoms that go away without treatment.
Others require treatment to prevent them from becoming more serious.

There are steps a person can take to reduce their risk of contracting and transmitting
disease causing pathogens. These include receiving available vaccinations, practicing regular
hand washing, and maintaining good hygiene at home.

What Are Protozoa?

Protozoa are broken down into different classes:

 Sporozoa (intracellular parasites)

 Flagellates (with tail-like structures that flap around to move them)
 Amoeba (which move using temporary cell body projections called pseudopods)
 Ciliates (which move by beating multiple hair-like structures called cilia)

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Infections caused by protozoa can spread through ingesting cysts (the dormant life stage),
sexual transmission, or through insect vectors (insects that transmit diseases through bites
or stings).

Protozoa cause some common and some uncommon infections. Some of these infections
cause illness in millions of people each year; other diseases are rare.

Protozoan Diseases

Common infectious diseases caused by protozoans include:

 Malaria
 Giardia
 Toxoplasmosis

These infections arise in very different parts of the body. For example, malaria infections
start in the blood, giardia begins in the gut, and toxoplasmosis can infect lymph nodes, the
eye, and the brain.

Detecting Infections

Unlike other pathogens, cultures do not identify protozoa. However, sometimes you can see
them under a microscope inside red blood cells (as in malaria) or in the stool (as in giardia
and E. histolytica).

In addition, rapid blood tests for antibodies or antigens and PCR tests can detect their
genetic material.

Treatment

Treatment options depend on what protozoa are infecting you. Some are a lot more
successful than others.

For example, malaria is a common illness worldwide that has straightforward treatment.
However, the treatment depends on the type of malaria (Plasmodium
falciparum, Plasmodium knowlesi, Plasmodium malariae, Plasmodium ovale, and
Plasmodium vivax).

Treatment also depends on whether the protozoa are drug-resistant. P. falciparum

especially has grown resistant to some essential drugs over the last few decades.

Summary

Protozoa are single-celled organisms that can sometimes cause diseases. Common
protozoan diseases include malaria, giardia, and toxoplasmosis. Diagnosing protozoan illness
may involve blood tests, stool tests, or biopsies, depending on which protozoa a healthcare
provider suspects. Treatment varies based on the cause

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Helminths

Parasitic worms, also known as helminths,[1] are large macroparasites; adults can generally
be seen with the naked eye. Many are intestinal worms that are soil-transmitted and infect
the gastrointestinal tract. Other parasitic worms such as schistosomes reside in blood
vessels.

Some parasitic worms, including leeches and monogeneans, are ectoparasites – thus, they
are not classified as helminths, which are endoparasites.

Parasitic worms live in and feed in living hosts. They receive nourishment and protection
while disrupting their hosts' ability to absorb nutrients. This can cause weakness and disease
in the host, and poses a global health and economic problem.[2] Parasitic worms cannot
reproduce entirely within their host's body; they have a life cycle that includes some stages
that need to take place outside of the host.[3] Helminths are able to survive in their
mammalian hosts for many years due to their ability to manipulate the host's immune
response by secreting immunomodulatory products.[4] All parasitic worms produce eggs
during reproduction. These eggs have a strong shell that protects them against a range of
environmental conditions. The eggs can therefore survive in the environment for many
months or years.

Many of the worms referred to as helminths are intestinal parasites. An infection by a

helminth is known as helminthiasis, helminth infection, or intestinal worm infection. There
is a naming convention which applies to all helminths: the ending "-asis" (or in veterinary
science: "-osis") is added at the end of the name of the worm to denote the infection with
that particular worm.[citation needed] For example, Ascaris is the name of a type of helminth, and
ascariasis is the name of the infection caused by that helminth.

Helminths are a group of organisms which share a similar form but are not necessarily
related as part of evolution. The term "helminth" is an artificial term.[5][6] There is no real
consensus on the taxonomy (or groupings) of the helminths, particularly within the
nematodes.[7] The term "helminth" contains a number of phyla, many of which are
completely unrelated. However, for practical considerations the term is currently used to
describe four phyla with superficial similarities: Annelida (ringed or segmented worms),
Platyhelminthes (flatworms), Nematoda (roundworms), and Acanthocephala (thorny-
headed worms).[7] The phylum Platyhelminthes includes two classes of worms of particular
medical significance: the cestodes (tapeworms) and the trematodes (flukes and blood
flukes), depending on whether or not they have segmented bodies.[1][8]

There may be as many as 300,000 species of parasites affecting vertebrates,[9] and as many
as 300 affecting humans alone.[10]

Helminths of importance in the sanitation field are the human parasites, and are classified
as Nemathelminthes (nematodes) and Platyhelminthes, depending on whether they possess
a round or flattened body, respectively.[8]

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Ringworm (dermatophytosis) is actually caused by various fungi and not by a parasitic

worm.

Control of microorganisms

Controling microorganisms can either be positive or negative:

POSITIVE control - you want to make them grow: Industrial Fermentations; beer, wine and
bread making

NEGATIVE control - you want to destroy them by (1) physical or chemical means or (2)
antibiotics

Usually we mean negative control and the rest of this discussion relates to the destruction
or inhibition of microbes:

CONTROL OF MICROORGANISM is essential in order to prevent the transmission of diseases

and infection, stop decomposition and spoilage, and prevent unwanted microbial
contamination. Microorganisms are controlled by means of physical agents and chemical
agents. Physical agents include such methods of control as high or low temperature,
desiccation, osmotic pressure, radiation, and filtration. Control by chemical agents refers to
the use of disinfectants, antiseptics, antibiotics, and chemotherapeutic antimicrobial
chemicals.

In this unit we will concentrate on the chemical control of microbial growth with a special
emphasis on the antibiotics and chemotherapeutic antimicrobial chemicals used in treating
bacterial infections. Control of microorganisms by means of physical agents will be covered
in Lab 18 and control by means of disinfectants, antiseptics, and sanitizers will be discussed
in Lab 19.

The basis of chemotherapeutic control of bacteria is selective toxicity. Selective toxicity

means that the chemical being used should inhibit or kill the intended pathogen without
seriously harming the host. A broad spectrum agent is one generally effective against a
variety of Gram-positive and Gram-negative bacteria; a narrow spectrum agent generally
works against just Gram-positives, Gram-negatives, or only a few bacteria. As mentioned
above, such agents may be cidal or static in their action. A cidal agent kills the organism
while a static agent inhibits the organism's growth long enough for body defenses to
remove it.

There are two categories of antimicrobial chemotherapeutic agents: antibiotics and

synthetic drugs. Antibiotics are metabolic products of one microorganism that inhibit or kill
other microorganisms. Chemotherapeutic synthetic drugs are antimicrobial drugs
synthesized by chemical procedures in the laboratory. Many of today's antibiotics are now
actually semi-synthetic and some are even made synthetically.

Antibiotics are metabolic products of one microorganism that inhibit or kill other
microorganisms. Why then do bacteria produce antibiotics? There is growing support for
multiple actions for microbial antibiotic production:

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 If produced in large enough amounts, antibiotics may be used as a weapon to inhibit

or kill other microbes in the vicinity to reduce competition for food.
 Antibiotics produced in sublethal quantities may function as interspecies quorum
sensing molecules enabling a number of different bacteria to form within a common
biofilm where metabolic end products of one organism may serve as a substrate for
another. All the organisms are protected within the same biofilm.
 Antibiotics produced in sublethal quantities may function as interspecies quorum
sensing molecules enabling some bacteria to manipulate others to become motile
and swim away thus reducing the competition for food.
 Antibiotics action may result in the degradation of bacterial cell walls or DNA and
these products can act as cues that trigger other bacteria to produce a protective
biofilm.
 Antibiotics produced in sublethal quantities may trigger intraspecies quorum
sensing. Exposure to low concentrations of an antibiotic may trigger bacteria to
produce quorum sensing molecules that trigger the population to produce a
protective biofilm. The biofilm then protects the population from greater
concentrations of the antibiotic.

Basic terms used in discussing the control of microorganisms include:

 1. Sterilization
Sterilization is the process of destroying all living organisms and viruses. A sterile
object is one free of all life forms, including bacterial endospores, as well as viruses.
 2. Disinfection
Disinfection is the elimination of microorganisms, but not necessarily endospores,
from inanimate objects or surfaces.
 3. Decontamination
Decontamination is the treatment of an object or inanimate surface to make it safe
to handle.
 4. Disinfectant
A disinfectant is an agents used to disinfect inanimate objects but generally to toxic
to use on human tissues.
 5. Antiseptic
An antiseptic is an agent that kills or inhibits growth of microbes but is safe to use on
human tissue.
 6. Sanitizer
A sanitizer is an agent that reduces microbial numbers to a safe level.
 7. Antibiotic
An antibiotic is a metabolic product produced by one microorganism that inhibits or
kills other microorganisms.
 8. Chemotherapeutic synthetic drugs
Synthetic chemicals that can be used therapeutically.
 9. Cidal
An agent that is cidal in action will kill microorganisms and viruses.

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 10. Static
An agent that is static in action will inhibit the growth of microorganisms.

PRESERVATION OF MICROORGANISM

Storage in liquid nitrogen is one of the best methods for preserving all microorganisms. For
some viruses, it may be better to freeze the sample rapidly, but slow freezing with a
cryoprotective agent is desirable for all other organisms to retain maximum viability or
infectivity.

DNA replication

DNA replication is the process by which a double-stranded DNA molecule is copied to

produce two identical DNA molecules. Replication is an essential process because,
whenever a cell divides, the two new daughter cells must contain the same genetic
information, or DNA, as the parent cell.

The replication process relies on the fact that each strand of DNA can serve as a template
for duplication. DNA replication initiates at specific points, called origins, where the DNA
double helix is unwound. A short segment of RNA, called a primer, is then synthesized and
acts as a starting point for new DNA synthesis. An enzyme called DNA polymerase next
begins replicating the DNA by matching bases to the original strand. Once synthesis is
complete, the RNA primers are replaced with DNA, and any gaps between newly
synthesized DNA segments are sealed together with enzymes.

DNA replication is a crucial process; therefore, to ensure that mistakes, or mutations, are
not introduced, the cell proofreads the newly synthesized DNA. Once the DNA in a cell is
replicated, the cell can divide into two cells, each of which has an identical copy of the
original DNA.

What is Replication?

DNA replication is the process in which a double-stranded DNA molecule splits into two
identical daughter strands. That is why during cell division each daughter cell contains the
same genetic information as the parent cell. The main enzyme responsible for DNA
replication is DNA polymerase. It adds new nucleotides to each strand.

What is Transcription?

Transcription is the conversion of DNA molecule into RNA. RNA polymerase uses one of the
DNA strands as a template to synthesise a complementary RNA molecule. The RNA molecule
that is synthesised is known as the transcript

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Conclusion

Both DNA replication and Transcription involve the generation of a new copy of the DNA in a
cell. DNA transcription is involved in replicating the DNA into RNA, while the DNA replication
makes another copy of DNA. Both the process is involved in the production of new nucleic
acids- DNA or RNA. The newly produced nucleic acids have some similarities but vary in their
functions. i.e one involved in cell division and the other is involved in gene expression.

A technique mainly used to change the phenotype of an organism (host) when a genetically
altered vector is introduced and integrated into the genome of the organism. So, basically,
this process involves the introduction of a foreign piece of DNA structure into the genome
which contains our gene of interest. This gene which is introduced is the recombinant gene
and the technique is called the recombinant DNA technology.
There are multiple steps, tools and other specific procedure followed in the recombinant
DNA technology, which is used for producing artificial DNA to generate the desired product.
Let’s understand each step more in detail.
Table of Contents

 Explanation
 Tools
 Process
 Application
 DNA Cloning
 Applications Of Gene Cloning

What is Recombinant DNA Technology?

The technology used for producing artificial DNA through the combination of different
genetic materials (DNA) from different sources is referred to as Recombinant DNA
Technology. Recombinant DNA technology is popularly known as genetic engineering.
The recombinant DNA technology emerged with the discovery of restriction enzymes in the
year 1968 by Swiss microbiologist Werner Arber,
Inserting the desired gene into the genome of the host is not as easy as it sounds. It involves
the selection of the desired gene for administration into the host followed by a selection of
the perfect vector with which the gene has to be integrated and recombinant DNA formed.
Thus the recombinant DNA has to be introduced into the host. And at last, it has to be
maintained in the host and carried forward to the offspring.
Also Refer- Genes

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Tools Of Recombinant DNA Technology

The enzymes which include the restriction enzymes help to cut, the polymerases- help to
synthesize and the ligases- help to bind. The restriction enzymes used in recombinant DNA
technology play a major role in determining the location at which the desired gene is
inserted into the vector genome. They are two types, namely Endonucleases and
Exonucleases.
The Endonucleases cut within the DNA strand whereas the Exonucleases remove the
nucleotides from the ends of the strands. The restriction endonucleases are sequence-
specific which are usually palindrome sequences and cut the DNA at specific points. They
scrutinize the length of DNA and make the cut at the specific site called the restriction site.
This gives rise to sticky ends in the sequence. The desired genes and the vectors are cut by
the same restriction enzymes to obtain the complementary sticky notes, thus making the
work of the ligases easy to bind the desired gene to the vector.
The vectors – help in carrying and integrating the desired gene. These form a very important
part of the tools of recombinant DNA technology as they are the ultimate vehicles that carry
forward the desired gene into the host organism. Plasmids and bacteriophages are the most
common vectors in recombinant DNA technology that are used as they have a very high
copy number. The vectors are made up of an origin of replication- This is a sequence of
nucleotide from where the replication starts, a selectable marker – constitute genes which
show resistance to certain antibiotics like ampicillin; and cloning sites – the sites recognized
by the restriction enzymes where desired DNAs are inserted.

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Host organism – into which the recombinant DNA is introduced. The host is the ultimate
tool of recombinant DNA technology which takes in the vector engineered with the desired
DNA with the help of the enzymes.
There are a number of ways in which these recombinant DNAs are inserted into the host,
namely – microinjection, biolistics or gene gun, alternate cooling and heating, use of calcium
ions, etc.

Process of Recombinant DNA Technology

The complete process of recombinant DNA technology includes multiple steps, maintained
in a specific sequence to generate the desired product.
Step-1. Isolation of Genetic Material.
The first and the initial step in Recombinant DNA technology is to isolate the desired DNA in
its pure form i.e. free from other macromolecules.
Step-2.Cutting the gene at the recognition sites.
The restriction enzymes play a major role in determining the location at which the desired
gene is inserted into the vector genome. These reactions are called ‘restriction enzyme
digestions’.
Step-3. Amplifying the gene copies through Polymerase chain reaction (PCR).
It is a process to amplify a single copy of DNA into thousands to millions of copies once the
proper gene of interest has been cut using the restriction enzymes.
Step-4. Ligation of DNA Molecules.
In this step of Ligation, joining of the two pieces – a cut fragment of DNA and the vector
together with the help of the enzyme DNA ligase.
Step-5. Insertion of Recombinant DNA Into Host.
In this step, the recombinant DNA is introduced into a recipient host cell. This process is
termed as Transformation. Once after the insertion of the recombinant DNA into the host
cell, it gets multiplied and is expressed in the form of the manufactured protein under
optimal conditions.
As mentioned in Tools of recombinant DNA technology, there are various ways in which this
can be achieved. The effectively transformed cells/organisms carry forward the recombinant
gene to the offspring.

Application of Recombinant DNA Technology

 DNA technology is also used to detect the presence of HIV in a person.

 Gene Therapy – It is used as an attempt to correct the gene defects which give rise
to heredity diseases.
 Clinical diagnosis – ELISA is an example where the application of recombinant

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 Recombinant DNA technology is widely used in Agriculture to produce genetically-

modified organisms such as Flavr Savr tomatoes, golden rice rich in proteins, Bt-
cotton to protect the plant against ball worms and lot more.
 In the field of medicines, Recombinant DNA technology is used for the production of
Insulin.

DNA Cloning
A clone is a cluster of individual entities or cells that are descended from one progenitor.
Clones are genetically identical as the cell simply replicates producing identical daughter
cells every time. Scientists are able to generate multiple copies of a single fragment of DNA,
a gene which can be used to create identical copies constituting a DNA clone. DNA
cloning takes place through the insertion of DNA fragments into a tiny DNA molecule. This
molecule is made to replicate within the living cell, for instance, a bacterium. The tiny
replicating molecule is known as the carrier of the DNA vector
Yeast cells, viruses, Plasmids are the most commonly used vectors. Plasmids are the circular
DNA molecules that are introduced from bacteria. They are not part of the main cellular
genome. It carries genes, which provide the host cell with beneficial properties such as
mating ability, drug resistance. They can be conveniently manipulated as they are small
enough and they are capable of carrying extra DNA which is weaved into them.

Applications Of Gene Cloning

Listed below are the applications of gene cloning:

 Gene Cloning plays an important role in the medicinal field. It is used in the
production of hormones, vitamins and antibiotics.
 Gene cloning finds its applications in the agricultural field. Nitrogen fixation is carried
out by cyanobacteria wherein desired genes can be used to enhance the productivity
of crop and improvement of health. This practice reduces the use of fertilizers hence
chemical-free produce is generated
 It can be applied to the science of identifying and detecting a clone containing a
particular gene which can be manipulated by growing in a controlled environment
 It is used in gene therapy where a faulty gene is replaced by insertion of a healthy
gene. Medical ailments such as leukaemia and sickle cell anaemia can be treated
with this principle

RNA VIRUS REPLICATION - GENERAL

STRATEGIES

RNA viruses that do not have a DNA phase

Viruses that replicate via RNA intermediates need an RNA-dependent RNA-polymerase to

replicate their RNA, but animal cells do not seem to possess a suitable enzyme. Therefore,
this type of animal RNA virus needs to code for an RNA-dependent RNA polymerase.

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No viral proteins can be made until viral messenger RNA is available; thus, the nature of the
RNA in the virion affects the strategy of the virus:

Plus-stranded RNA viruses

In these viruses, the virion (genomic) RNA is the same sense as mRNA and so functions as
mRNA. This mRNA can be translated immediately upon infection of the host cell

Examples:

 poliovirus (picornavirus)
 togaviruses
 flaviviruses

Negative-stranded RNA viruses

The virion RNA is negative sense (complementary to mRNA) and must therefore be copied
into the complementary plus-sense mRNA before proteins can be made. Thus, besides
needing to code for an RNA-dependent RNA-polymerase, these viruses also need to package
it in the virion so that they can make mRNAs upon infecting the cell.

Examples:

 influenza virus (orthomyxovirus)

 measles virus, mumps virus, (paramyxoviruses)
 rabies virus (rhabdovirus)

Double-stranded RNA viruses

The virion (genomic) RNA is double stranded and so cannot function as mRNA; thus these
viruses also need to package an RNA polymerase to make their mRNA after infection of the
host cell.

Example:

 rotaviruses (belong to reovirus family)

RNA viruses that copy their RNA to DNA

 These are the retroviruses. In this case, their virion RNA, although plus-sense, does
not function as mRNA immediately on infection since it is not released from the
capsid into the cytoplasm. Instead, it serves as a template for reverse transcriptase
and is copied into DNA. Reverse transcriptase is not available in the cell, and so these
viruses need to code for this enzyme and package it in virions.

GENOME SIZE OF RNA VIRUSES

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RNA viruses tend to have a relatively small genome (although virion size may not necessarily
be small). This is probably because the lack of RNA error correction mechanisms puts a limit
on the size of RNA genomes.

The result of having a small genome is that RNA viruses tend to code for only a few proteins.
These will include a polymerase which can copy RNA into a complementary nucleic acid
(either RNA or, as in the case of retroviruses, DNA) and a viral attachment protein

POSITIVE STRAND RNA VIRUSES

Examples:

 picornaviruses
 togaviruses
 flaviviruses

PICORNAVIRUSES (PICORNAVIRIDAE)

Properties

These are small (28nm), naked icosahedral viruses (figure 1) (pico=very small). The RNA is
single-stranded, plus sense, polyadenylated. It functions as mRNA immediately upon
infection
Prototype member: poliovirus (figure 1 and 2)

Adsorption and penetration

A viral protein recognizes a receptor on the host cell membrane (this is important in the
tropism of virus).
It seems that binding to the receptor alters capsid structure in some way, a channel forms
across the cell membrane and the RNA is released into cytoplasm. The mRNA is now
available for translation.

Synthesis of viral proteins

Poliovirus virion RNA functions as an mRNA but does not have the methylated cap structure
typical of eucaryotic mRNAs - it has a "ribosome landing pad" (known as the internal
ribosome entry site or IRES) which enables ribosomes to bind without having to recognize a
5' methylated cap structure (figure 3).

Picornaviruses often interfere with host cell methylated cap recognition. Most host cell
translation is cap-dependent, so this inhibits a lot of host protein synthesis but not viral

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protein synthesis - one way in which these viruses can modify the host cell to their
advantage.

RNA replication

RNA replication is the process by which new copies of genome-length RNAs are made (figure
8).
RNA replication occurs in the cytoplasm and is carried out by the viral RNA polymerase.
The full length plus strand is coated with nucleocapsid protein as it is made (mRNAs are not
coated with this protein, which would interfere with the host protein translation
machinery).

The new positive strand is copied into full length minus strand, which is also coated with
nucleocapsid protein as it is made. (Note: since the viral RNA polymerase
synthesizes mRNAs (transcription) and full-length RNA (replication), it is also sometimes
called a transcriptase or a replicase, such names just focus on the different aspects of the
polymerase activity.)

New negative strands may:

i. be used as templates for the synthesis of more full length plus strands
ii. be used as templates for the synthesis of more mRNAs
iii. be packaged into virions

Assembly

The virus consists of two "modules" - the envelope and the nucleocapsid:

Envelope
Transmembrane proteins are made on ribosomes bound to the endoplasmic reticulum.
They are inserted into the endoplasmic reticulum membrane as they are made, glycosylated
in the endoplasmic reticulum and pass through the Golgi body where substantial
modification of the carbohydrate chains occurs. They are then transported, in vesicles, to
the appropriate cell membrane; in the case of vesicular stomatitis virus, this is the plasma
membrane

Nucleocapsid
Synthesis of the nucleocapsid was described above. The viral RNA polymerase complex
associates with the nucleocapsids as they are formed. Nucleocapsids bud out through
modified areas of membrane which contain G and M proteins (figure 10). The M (matrix)
protein is involved in assembly - it interacts with patches of G in the membrane and with
nucleocapsids.

Note:
The entire life cycle occurs in the cytoplasm
RNA polymerase and RNA modification enzymes are virally-coded and present in the virus
particle (virion)

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UNIT 2 – MICROBIAL DIVERSITY AND NUTRIENT TURNOVER

Distribution of microorganisms?

Microbial biogeography is a subset of biogeography, a field that concerns the distribution of

organisms across space and time. Although biogeography traditionally focused on plants
and larger animals, recent studies have broadened this field to include distribution patterns
of microorganisms.

In what type of environments can a microorganisms be found?

Microbes live in every kind of habitat (terrestrial, aquatic, atmospheric, or living host) and
their presence invariably affects the environment in which they grow. Their diversity
enables them to thrive in extremely cold or extremely hot environments.

Microbes and Environment

Abstract
Microbes are omnipresent in the biosphere, and their presence invariably affects the
environment in which they grow. The effects of microbes on their environment can be
beneficial or harmful or inapparent with regard to human measure or observation. The most
significant effect of the microbes on earth is their ability to recycle the primary elements
that make up all living systems, especially carbon, oxygen, and nitrogen (N). Primary
production involves photosynthetic organisms which take up CO2 from the atmosphere and
convert it to organic (cellular) material. The process is also called CO2 fixation, and it
accounts for a very large portion of organic carbon available for synthesis of cell material.
Decomposition or biodegradation results in the breakdown of complex organic materials to
other forms of carbon that can be used by other organisms. There is no naturally occurring
organic compound that cannot be degraded by some microbe, although some synthetic
compounds such as Teflon, plastics, insecticides, and pesticides are broken down very slowly
or not at all. Through the microbial metabolic processes of fermentation and respiration,
organic molecules are eventually broken down to CO2 which is returned to the atmosphere
for continuous process of primary production. Biological nitrogen fixation is a process found
only in some bacteria which remove N2 from the atmosphere and converts it to ammonia
(NH3), for use by the plants and animals. Nitrogen fixation also results in replenishment of
soil nitrogen removed by agricultural processes. Thus along with all these benefits, microbes
greatly contribute in maintaining sustainability of environment. This chapter mainly focuses
on beneficial and harmful impacts of microbes on environment and their role to maintain
quality, health, and sustainability of environment.

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Keywords

 Microbe

 Environment

 Species interaction

 Nutrient cycle

 Bioremediation

 Pathogen

 Disease

Introduction
The Earth is known as a “closed system” where materials cycle between lithosphere (rocks),
atmosphere (air), hydrosphere (water), and biosphere (organism) (Fig. 3.1). Together, they
make up all the components of our planet, both living and nonliving. Earth produces
everything it needs to ensure the survival and growth of its residents. Environment is
defined as the circumstances or conditions that surround an organism or group of
organisms. Environment is the complex of social or cultural conditions that affects an
individual or community. Since humans inhabit the natural world as well as the built or
technological, social, and cultural world, all constitute an important part of our
environment.

Exchange of materials between different spheres of earth

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Environmental studies need to understand the life processes at the microscopic level and
ecologist levels from species to ecosystem. Species refer to organisms of the same kind that
are genetically similar enough to breed in nature and produce live, fertile offspring.
Population consists of all members of the same species living in a given area in the same
time. All the populations of organism living and interacting in a particular area make up a
biological community. An ecological system or ecosystem is composed of a biological
community and its physical environment. The environment includes abiotic factor such as
climate, water, minerals, and sunlight as well as biotic factors such as organisms, their
products, and effect in a given area.

Photosynthesis is the basis of energy economy of all, but a few specific ecosystem and
ecosystem dynamics are based in how organisms share food resources. In fact one of the
major properties of an ecosystem is its productivity, the amount of biomass produced in a
given area during a given period of time. The rate of production of food creates a linked
series of feeding known as food chain, whereas when individual food chains become
interconnected, they form food web. An organism’s feeding status in an ecosystem can be
expressed as trophic level shown in Fig. 3.2.

Environmental Communities and Their Factors

The groups of similar species create population which results in a community. Microbial
community means all microbial populations in a habitat. The activities of complex
communities of microbes affect biogeochemical transformations in natural, managed, and
engineered ecosystem. Microbial communities are very important for the rigorous progress
in the field of agriculture which increases the rate of crop production. Microbial community
may be terrestrial or aquatic.

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Terrestrial Communities

A community of microbes and their environment that occurs on the landmasses of

continents and islands form a terrestrial microenvironment. Terrestrial microenvironment is
distinguished from the aquatic microenvironment ecosystems by the lower availability of
water and the consequent importance of water as a limiting factor. Rain forests are the
most diverse and productive terrestrial microenvironment, but their soil is nutrient deficient
due to extensive leaching by rainwater.

Soil
Soil formation is a slow process that involves physicochemical weathering and biological
processes over millions of years. Microbes play an important role in soil aggregate
formation and soil stability that confer fertility and productivity to soil. The soil microbes
participate in these processes through many ways, e.g., filamentous microbes assemble clay
particles using extensive network of hyphae resulting into soil aggregates. Additionally,
some microbes secrete exopolysaccharides or cause compaction of clay particles that
promote soil aggregation. The surface soil is always rich in indigenous population of bacteria
(including actinomycetes), fungi, algae, and protozoans. Additionally, human and animal
activities also introduce specific microbes in the soil by several ways. Human activity directly
adds bacteria as biodegradative agents or applying sewage sludge to agricultural fields.
Animals introduce microbes through bird dropping or excretion.

Air
The atmosphere is an inhospitable climate for microbes because of stress due to
dehydration. This results in a limited time frame for microbes to be active; however, some
microbes get resistance to these stresses through specific mechanisms promoting loss of
their biological activity. Spore-forming bacteria, molds, fungi, and cyst-forming protozoans
all have specific mechanisms through which they are protected from these harsh gaseous
environments. Therefore, viability is highly dependent on the environment, time they spend
in the environment, and type of microbes. However, many other factors also influence the
viability of microbes such as humidity, temperature, oxygen content, specific ions, UV
radiation, various pollutants, and other air-associated factors (AOFs).

Relative Humidity
The relative humidity or relative water content of the air is critical for survival of airborne
microbes. Most of the gram-negative bacteria associated with aerosols are able to survive
for longer period at low relative humidity, whereas in contrast gram-positive bacteria
remain viable longer in association with high relative humidity. The ability of microbes to
survive in aerosol is related to the organism’s surface biochemistry. One possible
explanation of this fact could be a structural change in lipid bilayers of the cell membrane in
response to very low humidity. During loss of water, the cell membrane bilayer changes
from the typical crystalline structure to a gel phase and affects the surface protein

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configuration resulting into inactivation of the cell. The viruses with enveloped
nucleocapsids (e.g., influenza virus) have longer airborne survival in low relative humidity
below 50 %, whereas viruses without nucleocapsids (e.g., enteric viruses) are able to survive
in high relative humidity above 50 %.

Temperature
Temperature is a critical factor influencing the activity of microbes. In general, high
temperature leads to inactivation due to desiccation and protein denaturation, whereas
lower temperature promotes longer survival rates. At very low temperature, some microbes
lose viability because of ice crystal formation on their surface due to freezing.

Radiation
Mostly radiation at low wavelengths, e.g., UV radiation and ionic radiation (X-rays), is
harmful for microbes causing DNA damage. These radiations target DNA by producing single
or double strand breaks and changing the structure of nucleic acid bases. UV radiation
causes damage by forming intra-strand thymidine dimers causing inhibition of biological
activity such as replication of genome, transcription, and translation. Several mechanisms
including association of microbes with large airborne particles, pigments or carotenoids,
high relative humidity, cloud cover, etc. protect microbes from these harmful radiations.
However, many microbes (e.g., Deinococcus radiodurans) have evolved mechanisms to
repair DNA damage caused by UV radiation.

Oxygen, OAF, and Ions

Oxygen, open-air factors (OAF), and ions combine to inactivate many species of airborne
microbes. Some reactive forms of oxygen including superoxide radicals, hydrogen peroxide,
and hydroxide radicals are produced due to lighting, UV radiation, or pollution and cause
DNA damage by producing mutations. Similarly, OAFs (mixture of ozone and hydrocarbons)
also cause inactivation of microbes by damaging nucleic acids and enzymes. In addition to
these factors, positive ions cause only physical decay, e.g., inactivation of cell surface
proteins, whereas negative ions confer both physical and biological damages such as DNA
damage.

Aquatic Communities

Aquatic microenvironments occupy more than 70 % of the earth’s surface including mostly
ocean but also others such as estuaries, harbors, river, lakes, wetlands, streams, springs,
aquifers, etc. The microbiota, living in aquatic environment, are the primary producers
(responsible for approximately half of all primary production on earth) and primary
consumers as well. A large variety of microbial communities live in aquatic environments
such as the planktonic, sediment, microbial mat, biofilm communities, etc. Planktons refer
to photoautotrophic microbial community including both eukaryotes (algae) and
prokaryotes (cyanobacteria) and heterotrophic community including bacteria

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(bacterioplankton) and protozoans (zooplankton). Phytoplanktons are the primary

producers in the food web using their ability to fix CO2 into organic matter through
photosynthesis. Aquatic microenvironment is further classified into three
microenvironments, occupied by microbes living in freshwater, brackish water, and marine
water.

Freshwater
The study of freshwater microenvironment is known as micro-limnology. There are two
types of freshwater environment: standing water or lentic habitats (e.g., lakes, ponds, bogs)
and running water or lotic habitats including springs, rivers, and streams. Lentic habitats are
dominated by phytoplankton, forming distinct community gradients based upon the
wavelength and the amount of light that penetrates to a depth, e.g., Chlorobium.
Chlorobium can utilize longer wavelength than other phototrophs and survive with little or
no oxygen by consuming H2S instead of H2O for photosynthesis. In freshwater environment,
two types of lakes are present: eutrophic and oligotrophic lakes. Oligotrophic lakes have
higher rate (20–120 mg carbon/m3/day) than eutrophic lakes (1–30 mg carbon/m3/day)
because eutrophic lakes have much higher levels of organic matter causing turbidity and
interfering with light penetration. However, in terms of secondary productivity, eutrophic
lakes have much higher rates (190–220 mg carbon/m3/day) as compared to oligotrophic
lakes (1–80 mg carbon/m3/day).

Brackish Water
Brackish water environment is more saline than freshwater but less saline than marine
water environment. An estuary, a part of river that meets with sea, is the best example of
brackish water environment. Estuaries are highly variable environments because salinity
changes drastically over a relatively short distance. Despite this, estuaries are highly
productive environments, e.g., mangrove swamps in the Everglades of Florida, USA.
Estuaries are generally turbid due to the large amount of organic matter brought by rivers
and the mixing action of tides; therefore, light penetration is poor. Primary producers vary
from 100 to 107organisms/ml and in relation to depth and proximity to littoral zones.
Despite the low primary productivity, substrate availability is not limited, and heterotrophic
activity is high ranging from 150 to 230 mg carbon/m3/day.

Marine Water
Marine water environments are highly diverse and contain 33–37 % salinity. The ocean is
divided into two zones on the basis of light availability: photic zone, where light can
penetrate, and aphotic zone with lower light. Marine microenvironment is further divided
into four habitats: neuston, pelagic, epibiotic, and endobiotic. Habitat at the surface of sea
(air-water interface) is termed as neuston. On the basis of the precise depth, pelagic habitat
is subdivided into epipelagic and benthopelagic zones. Epipelagic zone is found in upper 100
m of the water column, and a large proportion of organisms living in it are photosynthetic,
whereas benthopelagic zone is sea-sediment interface. The third major habitat is the

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epibiotic habitats referring to surfaces on which attachment of communities occur, while

the fourth is the endobiotic habitat with organisms (e.g., Epulopiscium) found within the
tissues of other larger organisms such as fish.

Extremophilic Communities

The organisms living in physically or geochemically extreme conditions that are detrimental
to most life on earth are termed as extremophiles. Most of the extremophiles are microbes
and belong to the domain Archaea. Here are some extreme environmental conditions
where extremophiles survive.

High Temperature
Environments with high temperature (>70 °C) including terrestrial and submarine springs
with a temperature of 100 °C, hydrothermal vents with a temperature more than 300 °C are
inhospitable for most forms of life except some bacteria and archaebacteria, e.g., Thermus,
Methanobacterium, Sulfolobus, Pyrodictium, and Pyrococcus. Pyrodictium and Pyrococcus
are capable of surviving at temperature >100 °C. Another example of such renowned
extremophiles is Thermus aquaticus having thermotolerant DNA polymerase, which is
widely used in the polymerase chain reaction (PCR). These thermophiles have developed
such characteristic mechanisms facilitating proteins in folded state even at high
temperatures due to increased salt bridges (cations that bridge charges between amino acid
residues).

High Solute
Some organisms require salt concentrations substantially higher than that found in seawater
for their growth, and they are known as halotolerant. Halobacterium and Halanaerobium
are two examples of halotolerant bacteria; however, some algae and fungi also exhibit
halotolerance feature. The main mechanism of salt tolerance operates by internal
sequestration of high balancing solute (K+ in bacteria and glycerol in halotolerant
eukaryotes) equal to external salt concentration. A second mechanism of salt tolerance
involves proteins with acidic and low proportion of nonpolar amino acids. These proteins
require high salt concentrations to balance their charge for their optimal activity. Therefore,
some obligate halophiles are unable to survive in the environment lacking high salt
concentration due to these macromolecular modifications.

Microbes
There is a wide range of microbes present in our biosphere depending on their physical and
other characteristics. Microbes fall into two groups, prokaryotes and eukaryotes, depending
upon whether they have nucleus or not. Prokaryotes lack this membrane around their
genetic material, and this group includes viruses, bacteria, and related archaea. The other
category of microbes includes algae, fungi, protists, and other microscopic animals, having
cell nucleus.

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Bacteria and Archaea

Bacteria and archaea are the smallest free-living, unicellular organisms present on the earth.
Their cell sizes typically range from 0.5 to 1.0 μm in diameter. Both exist in various cell
shapes, e.g., cocci, rods, or spirals, and some soil bacteria form branching filaments, e.g.,
actinomycetes. Their DNA is found free in the cell cytoplasm and lack a true nuclear
envelope, and the genome is mainly composed of single double-stranded DNA molecule
with smaller DNA elements known as plasmids. The size of bacterial genome typically ranges
from four to six million nucleotides in length and enable to code 3,000–4,000 genes. A
bacterial cell envelope is composed of two layers, the inner layer is cell membrane made of
phospholipids and the outer layer is cell wall made of proteins, carbohydrates, and lipids,
but its composition varies based on the type of organism. Most of the microbes move
through flagella (whiplike extensions from the cell) and file filaments, e.g., pili. The pili
enable them to attach with each other or to soil particles. Additionally these pili are also
involved in transfer of genetic material between bacterial cells, known as conjugation. These
microbes usually reproduce asexually, e.g., binary fission, resulting in the formation of two
genetically identical bacterial cells. On the basis of gram staining, bacteria are of two types:
gram positive and gram negative; both vary in cell structure and physiology (Fig. 3.3).
Bacteria and archaea both require carbon as building blocks of their cellular materials and
energy to drive the reactions involved in cell biosynthesis and metabolism. Most of the
bacteria utilize oxygen, whereas some bacteria and archaea grow anaerobically by using
alternative electron acceptors, e.g., nitrate and sulfate.

Difference in gram-positive and gram-negative bacteria

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Basically microbes are classified into autotrophs and heterotrophs. Autotrophs utilize
sunlight or inorganic compounds such as Fe2+, nitrate, or nitrite as energy source to fix
atmospheric carbon dioxide to produce carbohydrates, fats, and proteins. However,
heterotrophic bacteria use organic compounds as a source of carbon and energy. Archaea
were originally known to be found in extreme environments and termed as
“extremophiles,” but now they are widely distributed and are found in many environments
including soil.

It is hard to distinguish both archaea and bacteria on the basis of their morphology. But
most recently their classification using molecular phylogenetic tools based on a comparison
of 16S ribosomal rRNA sequences has revealed three separate domains of life: eukaryotes,
bacteria, and archaea. Archaea are closely related to eukaryotes (all multicellular organisms)
than the bacteria (Woese et al. 1990).

Fungi

Fungi belong to eukaryotes and, therefore, are more closely related to plants and animals
than bacteria or archaea. Fungal cell consists of membrane-bound nucleus with
chromosomes containing genetic material, e.g., DNA, membrane-bound organelles, e.g.,
mitochondria, and a cell wall composed of glucans and chitin. Fungi are basically
heterotroph organisms meaning thereby that they derive their food from nonliving organic
sources, e.g., saprophytic fungi, which feed on dead or decaying organic materials. Few
fungi also exist as unicellular organisms, e.g., yeast, which grow through cylindrical
threadlike structures (2–10 cm in diameter) known as hyphae. These hyphae may be either
septate, e.g., compartmentalized through cross walls, or nonseptate. The hypha is a main
part of fungus and constitutes a mycelium. Finely branched mycelium occupies a large
surface area in the soil and produces a range of enzymes acting on soil organic matter to
produce nutrients and energy required for fungal growth. Fungi can reproduce by both
sexually, e.g., through spores, and asexually, e.g., budding or binary fission. Fungi are highly
diverse and play a wide range of role in their surrounding environment such as
decomposers, mutualists, endophytes of plants, pathogens, and predators. Fungal hyphae
are the basic components of soil food webs since they constitute a food source for soil
biota, whereas fungal sporocarps provide food for larger animals.

Protists

Protists are unicellular eukaryotes having characteristic organelles but lacking cell wall.
Protists may be free living, parasitic, or opportunistic based on environmental changes. The
size of protists varies from 2 μm to several centimeters.

These are classified into four groups: flagellates (Mastigophora), amoebae (Sarcodina),
sporozoans (Sporozoa), and ciliates (Ciliophora). Flagellates are characterized by the
presence of flagella as a locomotive tool, multiplication by binary fission, and having both
heterotrophic and autotrophic feeding mechanisms.

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Flagellates are further classified into two major divisions: photosynthesizing

(Phytomastigophora) and non-photosynthesizing (Zoomastigophora). Autotrophic flagellates
possess chloroplast and synthesize their own food or nutrients by photosynthesis such as
Euglena, whereas heterotrophic flagellates are parasitic and take their nutrients from their
host, for example, Leishmania and Giardia. Amoebae have pseudopodia as their locomotive
tool. Pseudopodia help in ingestion of food materials and provide an extended basis of
further classification.

The Rhizopoda move by a fluid endoplasm, whereas the Actinopoda have a spikelike
pseudopodium for their movement and feeding. These organisms are vacuolated and
covered in a shell-like outer layer known as “test.” These shells may be composed of
proteinaceous, siliceous, or calcareous substances and have either a single or multiple
chambers. Some testate amoebae are also found in soil, building their shells by excreting
substances capable of aggregating soil particles. Some important free-living soil amoebae
are grouped in the family Vahikampfia. Entamoeba is well-known parasitic amoeboid
causing dysentery in humans. The ciliates have hairlike structures in an ordered array
surrounding the cell known as cilia that divides by transverse fission.

Cilia help in their locomotion and feeding. Ciliates are generally free living such as
Paramecium, but some species are adapted to parasitic life cycles. Sporozoans are mostly
parasitic and form spores. Few members are adapted in a symbiotic relationship and have
no locomotive organ; therefore, they rely on vectors or direct contact with susceptible host
to continue their growth and replication. For example, some species have evolved to enable
digestion in the gut of domestic livestock. However, parasitic members are totally
dependent on host for their nutrition, for example, Toxoplasma, Isospora, and Plasmodium.

Most of parasitic protists are of obvious public health concern causing deadly diseases such
as malaria, sleeping sickness, Chagas disease, leishmaniasis, giardiasis, cryptosporidiosis,
etc. These parasites have adapted themselves for surviving and reproducing in their hosts by
evading host immune responses. Many flagellated protists are capable of forming cysts that
are known to survive conventional methods of disinfection and can be transmitted to their
host via a water route.

Entamoeba histolytica is a parasitic amoeba and causes diarrhea and dysentery. Naegleria
is a free-living amoeba in freshwater, causing infections in nasal passage of humans and
capable of invading brain tissues. Toxoplasma is an invasive protist that causes blindness
and serious illness or death in unborn fetuses. Cryptosporidium is responsible for a number
of epidemics including the largest US waterborne outbreak in Milwaukee. Plasmodium is a
main causative agent of mosquito-borne disease, malaria. Domestic animals are also at risk
of serious illness and death from parasitic protozoans, for example, Histomonas,
Trichomonas, etc.

Viruses

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Viruses are small, obligate, intracellular pathogens that require a host cell for their growth
and replication. They can survive outside the host cell but not multiply without a host.

Viruses are generally species specific and can infect all types of life forms: bacteria, plants,
and animals. Public focus is most often on (1) plant viruses affecting important crops such as
tobacco, potatoes, and tomatoes or (2) animal viruses causing deadly diseases: herpes,
smallpox, rabies, mumps, measles, meningitis, hepatitis, encephalitis, influenza, diarrhea,
yellow and dengue fever, etc. The basic virus structure includes a capsid protein coat and an
internal nucleic acid (RNA or DNA), but some viruses may also have protein and lipid
envelopes, glycoprotein spikes, or more complex tail and sheath structures.

A number of protein capsomers held together by non-covalent bonds form a capsid coast
surrounding the nucleic acid molecule. The size of capsids ranges from 18 μm, as in small
parvovirus of animals, to several hundred nanometers, as in some filamentous plant viruses.
This outer coat protects and shields the viral nucleic acid and harbors specific receptor sites
for attachment on hosts. The viral capsids have two types of symmetrical organization:
helical and icosahedral. The helical viruses look like a spiral or a helix with a cylindrical
shape, whereas icosahedral viruses adopt a 20-triangular-sided spherical shape when
viewed with an electron microscope. Viruses have either RNA or DNA in the double-
stranded or single-stranded form. Viral nucleic acid length varies from 1.7 to over 200 kb,
and it encodes 4–200 genes.

Microbial Diversity
Early studies on diverse soil bacteria and fungi are mainly focused on what could be easily
cultured from soils, but the fact is that less than 10 % of the soil bacteria could be cultured,
suggesting the requirement of other approaches.

Norman Pace and colleagues in 1980 found that microorganisms could be identified in
naturally occurring microbial populations without culturing them (Hugenholtz et al. 1998),
but this process requires the PCR amplification of the rRNA genes using rRNA specific
primers and RNA extracted directly from the cells present in soil. These specific primers may
differentiate among various microbial communities at level of different domains such as
Bacteria, Eukarya, and Archaea or phylum (e.g., Actinobacteria or Bacteroidetes) (Fig. 3.4).
However, a range of approaches could be adopted in order to separate and sequence the
rRNA genes.

The advanced high-throughput DNA sequencing now allows the identification of each
individual in thousands of samples within a short duration (Caporaso et al. 2012). Once we
compare these rRNA gene sequences from cultivated species using various online
databases, e.g., GenBank, allowing identification of evolutionary (phylogenetic)
relationships among various unknown and known organisms, it displays an estimate of the
genetic diversity of organisms in a particular community. It is easy to speculate about the

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organism’s characteristics and its closest cultivated relative on the basis of sequencing
details.

Phylogenetic information sometimes also provides details about the physiology, e.g., all
cyanobacteria constitute a monophyletic group in a similar way as sulfate-reducing bacteria,
halophiles, and methanogenic archaea do.

Phylogenetic classification of living world based on 16S and 18S rRNA gene sequences

Bacterial Phyla

The composition of in situ environmental bacterial communities has been investigated using
various molecular tools. The relative abundance of the major phyla varies among diverse
soils and environmental conditions such as some members of the phyla, i.e., Proteobacteria,
Acidobacteria, and Actinobacteria, are abundant and widely distributed; however, members
belonging to Verrucomicrobia, Bacteroidetes, Firmicutes, Chloroflexi, Planctomycetes, and
Gemmatimonadetes are comparatively less prevalent.

The Proteobacteria is a diverse group of organisms among various subphyla out of which, α-,
β-, γ-, and δ-Proteobacteria are most commonly found in soil. The members belonging to α-,
β-, and γ-subphyla are copiotrophs (an organism able to grow in nutrient-rich environments
particularly carbon in contrast to oligotrophs, those found in environments with much lower
carbon concentration). The Proteobacteria are more prevalent in those area rich in resource
availability, e.g., rhizosphere soils.

The α-Proteobacteria consist of various metabolically diverse heterotrophic and autotrophic

bacteria, e.g., heterotrophic bacteria such as Sphingomonas, that are able to degrade
various toxic compounds including pentachlorophenol and polyaromatic hydrocarbons and

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also involved in weathering of minerals. Some heterotrophs that are also able to fix
atmospheric nitrogen by forming symbiotic relationships with legumes belong to family
Rhizobiaceae such as Rhizobium, Mesorhizobium, and Bradyrhizobium. The autotrophic α-
Proteobacteria also include soil methane oxidizers, e.g., Methylobacter and Methylophilus,
nitrite oxidizers, e.g., Nitrospira and Nitrobacter, and phototrophs, e.g., Rhodospirillum and
Rhodobacter.

The β-Proteobacteria is also found into three groups: heterotrophs, autotrophs, and
methanotrophs. Some of the known heterotrophs found in soil belong to the genera
Burkholderia, Alcaligenes, and Acidovorax. Out of these, Burkholderia spp. is metabolically
diverse and uses simple amino acids, sugars, and recalcitrant aromatic and phenolic
compounds as a source of carbon and therefore plays a major role in carbon turnover.
Burkholderia spp. is also known to promote plant growth by fixing atmospheric nitrogen.
Some examples of heterotrophic β-Proteobacteria are Collimonas, able to degrade live
hyphae by producing chitinase, and autotrophic β-Proteobacteria are Nitrosospira
(ammonia oxidizer), Thiobacillus (iron oxidizer), Methylomonas (methane producer),
Rhodocyclus (phototroph), etc.

The γ-Proteobacteria are also categorized into heterotrophs, lithotrophs, and phototrophs.
Pseudomonas and Xanthomonas are well-known heterotrophs. Pseudomonas spp. has a
remarkable nutritional versatility since most of them are able to grow on more than 50 or
100 different substrates including sugars, amino acids, fatty acids, alcohols, and
hydrocarbons. The γ-Proteobacteria also include various photolithotrophs, e.g., Thiocapsa
and Chromatium, that utilize sulfide or elemental S as an electron donor and CO 2 as a source
of carbon under anaerobic conditions in light.

The δ-Proteobacteria consist mainly of SO4 −2- and Fe-reducing bacteria. Desulfovibrio, a
sulfate-reducing bacteria, grow anaerobically by utilizing lactate or ethanol as carbon
sources, found in oxygen-depleted soil. This group also includes a parasitic bacteria named
as Bdellovibrio.

Acidobacteria are widely distributed and found in abundance in the soil with low pH. Since
they are poorly present in soil culture collections, a little knowledge is available about their
metabolic capabilities. The complete genome sequencing of cultured soil Acidobacteria, e.g.,
Acidobacterium capsulatum, implies that they may be oligotrophs and able to metabolize a
range of simple and complex carbon sources. These bacteria are also found in low nutrient
conditions, tolerant to changes in soil moisture. They also play a role in nitrate and nitrite
reduction but not in denitrification or nitrogen fixation.

Verrucomicrobia, also found commonly in soil, are oligotrophic in nature; however, the
ecology of Verrucomicrobia is poorly understood. The majority of bacteria of this group are
Chthoniobacter flavus and Opitutus terrae. Genome sequencing of a free-living heterotroph
bacteria found in aerobic soil, e.g., Chthoniobacter flavus, suggests that it is able to

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metabolize plant polysaccharides but not amino acids except pyruvate. However, genome
sequencing of a verrucomicrobium from rice paddy soil, e.g., Opitutus terrae, implies that it
is an anaerobic bacterium, capable of producing propionate through fermentation of
polysaccharides from plants.

Gram-positive bacteria are abundant in soil culture collections and categorized into two
groups: Actinobacteria and Firmicutes. The Actinobacteria are commonly found in soil and
further classified into three subphyla: Actinobacteridae, Acidimicrobidae, and
Rubrobacteridae. The abundance of Actinobacteridae in soil relatively increases with
addition of labile carbon sources. The Actinobacteridae includes metabolically diverse
aerobic heterotrophs, e.g., Arthrobacter, Rhodococcus, Streptomyces, and Mycobacterium.
Streptomyces is a well-known bacterium producing antimicrobial compounds. The
Rubrobacteridae includes two genera not represent in soil culture collections, e.g.,
Rubrobacter and Solirubrobacter. Acidimicrobium ferrooxidans is an acid-tolerant ferrous
iron oxidizer bacterium and has been detected in soil culture collections among few
members of Acidimicrobidae.

The Firmicutes include bacteria that are able to form endospores such as Bacillus and
Clostridium, and because of endospore production, they are able to survive longer in the soil
during dry periods. Bacillus is capable of degrading various carbon sources, including
polysaccharides from plants, whereas Clostridium may ferment sugars, starch, pectin, and
cellulose. The addition of recalcitrant C compounds in soil favors the growth of Clostridiales.

Planctomycetes multiply through budding and lack peptidoglycan in their cell walls. These
bacteria are also involved in ammonium oxidation (Anammox) in soil under anaerobic
conditions. Planctomycetes, Verrucomicrobia, and Chlamydia are important from
evolutionary point of view since they share numerous features, e.g., presence of
membrane-coat-like proteins and condensed DNA, rarely found in bacteria but more
common in Archaea and Eukaryotes. Sphingobacteria are known to be involved in aerobic
degradation of plant materials present in soil and complex organic molecules, e.g., starch,
proteins, cellulose, and chitin. Members belonging to genus Chitinophaga are filamentous
and chitinolytic and exhibit gliding movement.

Archaeal Phyla

Archaea are known to be widely distributed in the soil on the basis of 16S rRNA gene
sequences (Bates et al. 2011), and additionally these gene sequences suggest that members
belonging to the phylum Crenarchaeota are found in abundance in the marine environment.
All cultured Crenarchaeota are thermophilic or hyperthermophilic organisms: having the
ability to survive at a temperature up to 113 °C. Crenarchaeota is sulfur-dependent
extremophile; one of the best-characterized members is Sulfolobus solfataricus. This
organism was originally isolated from geothermally heated sulfuric springs in Italy and grows
at 80 °C and pH of 2–4. These organism stains are gram negative and are morphologically

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diverse having rod-, cocci-, filamentous-, and oddly shaped cells. The Crenarchaea play a
role in ammonia oxidation in the soil. Mesophilic ammonia-oxidizing archaea (AOA) is
abundant in a diverse range of marine environments, including the deep ocean, as revealed
by the quantification of the archaeal amoA gene encoding the alpha-subunit of the
ammonia monooxygenase. Nitrososphaera viennensis is a most recent ammonia-oxidizer
Crenarchaea that was extracted from garden soil, and subsequent phylogenetic analysis
confirmed its taxonomic affiliation.

Methanogens are strict soil anaerobes and grow in association with bacteria that participate
in the anaerobic food chain and convert complex organic molecules to methane (CH4) and
CO2. Methanogens generate methane through various pathways. They display various
activities such as reduction of carbon dioxide and methanol, cleavage of acetate, and
methane production from methylated compounds. Members belonging to genera
Methanosarcina, Methanosaeta, and Methanocella are widely distributed in the
environment, and both Methanosarcina and Methanosaeta produce methane through
acetate reduction.

Fungal Phyla

Seven fungal phyla are currently recognized: Chytridiomycota, Blastocladiomycota,

Neocallimastigomycota, Glomeromycota, Ascomycota, Basidiomycota, and parasitic
endobionts, e.g., Microsporidia. Among these fungal phyla, the first three have flagellated
cells at least during one stage of their life cycle and thereby turned into terrestrial organisms
in contrast to higher fungi that lack motile cells. Among these phyla, Chytridiomycota is a
basal group and may degrade chitin and keratin. Batrachochytrium dendrobatidis is a known
pathogenic unicellular chytrid of many amphibian species resulting into decline of
worldwide amphibian population. Blastocladiomycota differ from the Chytridiomycota in
reproduction since they exhibit different form of meiosis.

Glomeromycota exhibit some features identical to “lower” fungi, e.g., they have
multinucleate aseptate mycelia and most of them have no known sexual stages. They
reproduce through large thick-walled asexual spores, commonly found in soils, and may
germinate in the presence of a plant root. This phylum includes arbuscular mycorrhizal (AM)
fungi that form obligate biotrophic symbioses with mosses, approximately 80 % of all land
plants, and cyanobacterium Nostoc forming cyano-lichens. These higher fungal phyla have a
characteristic feature having two compatible nuclei in a hyphal cell also known as dikaryon.

Ascomycota is one of the largest fungal phyla with 64,000 or more number of species. The
members of this phylum have a characteristic spore-bearing saclike structure also known as
asci, produced in large numbers during sexual reproduction. Ascomycetes mostly reproduce
asexually and are rarely found to reproduce through sexual mating. Ascomycetes have a
typical haploid mycelium with septate hyphae and cell wall made up of chitin and β-glucans.
Few macroscopic ascomycetes exhibit well-known reproductive structures such as morels,

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truffles, etc. However, many ascomycetes are microscopic unicellular organisms, e.g., yeast
or Saccharomyces, and filamentous fungi, e.g., Aspergillus. Most of the Ascomycetes are
saprobes and have various enzymes to degrade complex substrates such as cellulose,
keratin, and collagen. Therefore, ascomycetes are known to play a critical role in
decomposing and nutrient recycling. Approximately 18,000 species of ascomycetes form
lichens through symbiotic relationship with green algae and cyanobacteria. However, other
Ascomycota constitute ectomycorrhizal and/or ectendomycorrhizal associations through
symbiosis with woody plants. Some ascomycetes are known plant parasites and predators.
The family Orbiliaceae includes carnivorous fungi having specialized hyphae to trap prey
including a range of soil mesofauna, protists, nematodes, and arthropods.

Members belonging to Basidiomycota are also known as club fungi and produce spores
during sexual reproduction on club-like stalks known as basidia. These microscopic basidia
are basically clustered on specialized structures also known as sporocarps. Numerous
haploid spores are produced after meiosis and released in the environment resulting into a
new haploid mycelium after germination

Microbial Application to Environment

Contribution of Microbes to Nutrient Cycling

Microbes in Carbon Cycle

Microbes play a critical role in carbon cycle on the global scale that is a key constituent of all
living organisms (Fig. 3.5). Microorganisms avail carbon for living organisms and for
themselves as well through extracting it from nonliving sources. In aquatic habitats,
microbes convert carbon anaerobically, present at oxygen-free zones such as deep mud of
lakes and ponds.

Carbon dioxide (CO2) is the most common form of carbon that enters into a carbon cycle.
CO2 is a water-soluble gas present in the atmosphere. Plants and photosynthetic alga use
CO2 during photosynthesis to synthesize carbohydrates. Additionally chemoautotrophs such
as archaea and bacteria also utilize CO2 to synthesize sugars.

This carbon, present in the form of sugar, is further processed through a chain of reactions
during respiration known as tricarboxylic acid cycle resulting into energy. Microbes may also
use carbon under anaerobic conditions to produce energy through a process called as
fermentation.

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Role of microbes in carbon cycle

Plants are the primary producers in a terrestrial ecosystem; however, free-living planktons,
cyanobacteria, and symbionts such as lichens also contribute in fixing carbon in some
ecosystems. Nonliving organic material is recycled by heterotrophic bacteria and fungi,
whereas saprobes utilize organic material and produce CO2 during respiration, thereby
contributing to carbon cycle. However, higher animals, e.g., herbivores and carnivores, also
digest organic materials to obtain energy using gut microbiota residing in their intestinal
tracts; the process is known as decomposition, resulting into inorganic products such as CO 2,
ammonia, and water.

Actinobacteria and Proteobacteria are capable of degrading soluble organic compounds,

e.g., organic acids, amino acids, and sugars (Eilers et al. 2010). Similarly, bacteroidetes are
also involved in degradation of more recalcitrant carbon compounds, e.g., cellulose, lignin,
and chitin, and utilize higher level of available nitrogen that help in production of
extracellular and transport enzymes (Treseder et al. 2011). On the contrary, bacteria living
in low-nitrogen environments are more able to metabolize nitrogen-rich organic
compounds, e.g., amino acids. The abundance of α-Proteobacteria and Bacteroidetes favors
carbon mineralization, whereas Acidobacteria oppose it (Fierer et al. 2007).

Microbes in Methane Production

Some microbes execute anaerobic or fermentative degradation of organic compounds into
organic acids and some gases, e.g., hydrogen and CO2. Methanogens are able to use that
hydrogen to reduce CO2 into methane under strict anaerobic conditions (Fig. 3.6). In order

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to complete cycle, methane-oxidizing bacteria, e.g., methanotrophs, transform methane to

CO2, water, and energy under aerobic conditions.

Production of methane by microbes

Other microbes such as green and purple sulfur bacteria participate in carbon cycle by
degrading hydrogen sulfide (H2S) into compounds having carbon during energy production
(see in reaction). Some bacteria, e.g., Thiobacillus ferrooxidans, derive energy from
oxidation of ferrous iron to ferric iron and thereby contribute to carbon cycle. Few microbes
such as Bacteroides succinogenes, Clostridium butyricum, and Syntrophomonas spp. make a
collaborative effort (also known as interspecies hydrogen transfer) for anaerobic
degradation of carbon to produce CO2 and methane in bulk. The following reaction shows
anaerobic photoautotrophism in purple sulfur bacteria:

Microbes in Nitrogen Cycle

Nitrogen is an essential element present in protein and nucleic acid structure.
Microorganisms play a critical role in nitrogen cycle through various processes such as
nitrogen fixation, nitrate reduction, nitrification, denitrification, etc. (Fig. 3.7). The microbial
processes limit the productivity of an ecosystem because nitrogen availability is a limiting
factor for plant biomass production. Ammonification involves decomposition of organic
nitrogen into ammonia. Both bacteria and archaea are capable of fixing atmospheric
nitrogen through reduction into ammonium (Fig. 3.7). Nitrogenase is an oxygen-sensitive
enzyme that catalyzes nitrogen fixation under low oxygen environment. N-fixation requires
energy in form of ATP (16 mol) per mole of fixed nitrogen.

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Role of microbes in nitrogen cycle

The free-living microbes such as Azotobacter, Burkholderia, and Clostridium have an ability
to fix nitrogen, and few of them form a symbiotic relationship with the rhizosphere of plants
such as Rhizobium, Mesorhizobium, and Frankia. Sophora and Clianthus are native legumes
and form a symbiotic relationship with Mesorhizobium or Rhizobium leguminosarum. The
symbiotic rhizobia are able to fix nitrogen by two or three orders of magnitude higher than
free-living soil bacteria.

Nitrification involves two steps: first, ammonia is oxidized to nitrite and then to nitrate. The
oxidation of ammonia to nitrite is carried out by few soil bacteria, e.g., Nitrosospira,
Nitrosomonas, Crenarchaeum, or Nitrososphaera, and thereafter nitrite is oxidized to nitrate
by some bacteria, e.g., Nitrobacter and Nitrospira (Fig. 3.7). Nitrification also changes the
ionic state of soil from positive to negative through oxidation of ammonia to nitrite and
release of energy, which is used by nitrifying microbes to assimilate CO2.

Denitrification involves sequential reduction of nitrate (NO3 −), nitrite (NO2 −), and nitric
oxide (NO) to the greenhouse gas nitrous oxide (N2O) or benign nitrogen gas (N2). Since this
process requires limiting oxygen, therefore, it occurs mostly in waterlogged areas that
provide anaerobic environment. Nitrogen cycle involves denitrification process through
which fixed nitrogen returns back to the atmosphere from soil and water in order to
complete the nitrogen cycle. Denitrification involves a range of soil microbiota belonging to
Proteobacteria, Actinobacteria, and Firmicutes and other soil eukaryotes. Most of the
bacteria lack single or multiple enzymes involved in denitrification and known to be
incomplete denitrifier, for example, most of the fungi and bacteria lack nitrous oxide
reductase and thereby produce N2O as a final product. Therefore, incomplete denitrification
results into emission of greenhouse gases.

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Microbes in Sulfur Cycle

Sulfur is an important component of a couple of vitamins and essential metabolites, and it is
found in two amino acids, cysteine and methionine. In spite of its paucity in cells, it is an
absolutely essential element for living systems. Like nitrogen and carbon, the microbes can
transform sulfur from its most oxidized form (sulfate or SO4) to its most reduced state
(sulfide or H2S) (Fig. 3.8). The sulfur cycle, in particular, involves some unique groups of
prokaryotes. Two unrelated groups of prokaryotes oxidize H2S to S and S to SO4. The first is
the anoxygenic photosynthetic purple and green sulfur bacteria that oxidize H 2S as a source
of electrons for cyclic photophosphorylation. The second is the “colorless sulfur bacteria”
(now a misnomer because the group contains many archaea) which oxidize H2S and S as
sources of energy. In either case, the organisms can usually mediate the complete oxidation
of H2S to SO4.

Role of microbes in sulfur cycle

Sulfur-oxidizing prokaryotes are frequently thermophiles found in hot (volcanic) springs and
near deep-sea thermal vents that are rich in H2S. They may be acidophiles as well, because
they acidify their own environment by the production of sulfuric acid. Since SO 4 and S may
be used as electron acceptors for respiration, sulfate-reducing bacteria produce H2S during a
process of anaerobic respiration analogous to denitrification. The use of SO4 as an electron
acceptor is an obligatory process that takes place only in anaerobic environments. The
process results in the distinctive odor of H2S in anaerobic bogs, soils, and sediments where it
occurs. Sulfur is assimilated by bacteria and plants as SO4 for use and reduction to sulfide.
Animals and bacteria can remove the sulfide group from proteins as a source of S during
decomposition. These processes complete the sulfur cycle.

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Microbes in Phosphorus Cycle

Phosphorus is a critical element of various building blocks such as nucleic acids, e.g., DNA
and RNA, ADP, ATP, and phospholipids. Phosphorus is a rare element in the environment
because of its tendency to precipitate in the presence of divalent and trivalent cations at
neutral and alkaline pH.

Microorganisms (bacteria and fungi) mineralize organic phosphate in the form of phosphate
esters into inorganic phosphate through a process driven by phosphatase enzymes (Fig. 3.9).
Additionally, they also convert insoluble phosphorus into soluble form by a reaction with
resulting byproducts such as organic acids. Mycorrhizal fungi help plants to overcome
phosphorus limitation through its mobilization from insoluble mineral form by producing
oxalate, e.g., various ectomycorrhizal basidiomycetous fungi express phosphate
transporters in their extraradical hyphae during phosphorus deficiency in surrounding
environments.

Role of microbes in phosphorus cycle

Contribution of Microbes in Recycling Wastes and Detoxification

Bacteria and fungi are able to biodegrade or detoxify substances through various ways;
thereby, microbial processes are extensively used for bioremediation.

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Biodegradation
Bioremediation/biotransformation is a waste management tool that involves naturally
occurring organisms to remove or neutralize hazardous waste into less toxic or nontoxic
substances. The most commonly used microorganisms are Flavobacterium,
Arthrobacterium, and Azotobacter. Bioremediation focuses on different sources and hence
is called with different names:

$$ \begin{array}{l}\mathrm{Plant}\to \mathrm{Phytoremediation}\hfill \\
{}\mathrm{Fungi}\to \mathrm{Mycoremediation}\hfill \end{array} $$
Biotechnological treatment of waste management involves use of microorganisms to
detoxify air, water, and soil pollutants and carried out at lower temperature and pressure;
therefore, it requires less energy than the conventional physicochemical treatment method.
Depending upon the types of contaminants’ site of monitoring and favorable environmental
conditions, bioremediation may be carried out either in situ or ex situ

Bioremediation and its types

Biostimulation and bioaugmentation processes promote the rate of degradation of organic
and inorganic pollutants . These treatment technologies have been found eco-friendly and
cost-effective means of pollution control leading to increased public acceptance and
compliance with environmental legislation.

Overview of bioremediation

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Heterotrophic microbes such as Pseudomonas, Sphingomonas, and Mycobacterium are

known to be involved in oil degradation. Pseudomonas is one of well-studied bacteria
capable of degrading alkanes, monoaromatics, naphthalene, and phenanthrene under
aerobic conditions. The hydrocarbon-degrading bacteria are dominant in soil contaminated
with oil; however, higher concentration of hydrocarbons may deplete available nitrogen and
phosphorus in that area since these elements are assimilated during biodegradation.

Microbes (bacteria and fungi) are able to degrade a range of biodegradable pesticides such
as atrazine, which is degraded by a bacterium, e.g., Arthrobacter nicotinovorans, and related
derivatives such as simazine, propazine, and cyanazine (Aislabie et al. 2005). Non-
biodegradable pesticides, e.g., DDT (dichlorodiphenyltrichloroethane), are not readily
degraded and still persist in the soil. Some fungi having ability to degrade lignin, such as
Phanerochaete chrysosporium, are able to degrade various contaminants such as
pentachlorophenol and dioxin, and the best example are Zygomycetes that degraded
various contaminants during wood-treating operation in Whakatane (Thwaites et al. 2006).

Biodegradation of a contaminant depends upon its chemical structure and physical state
since various contaminants, e.g., oil are readily degradable, but synthetic contaminants, e.g.,
DDT and aldrin, are nondegradable and persist in the environment. The ability for
degradation also depends upon rare and novel structures and water solubility since less
soluble compounds are difficult to degrade. Additionally, poorly water-soluble or
hydrophobic contaminants also readily bind to clay particles and, therefore, are easily
available to microbes present in soil. These soil microbes utilize these contaminants as
energy source, present at higher concentration, and these could be toxic for them, resulting
into slow biodegradation. Biodegradation also involves a contact between contaminants
and microbes. Some microbes, e.g., chemotactic bacteria and fungi, have an ability to sense
and move toward them.

Metal Detoxification
The microbial ability to withstand metal toxicity and their physiological adaptation to metal
stress has an important significance. Indeed, the expression and activity of proteins involved
in metal uptake are crucial for metal resistance, and different bacteria adapt distinct
complements of these systems. Bacteria have evolved some regulatory mechanisms to
control membrane transporter activity that take up metals, and some of these activities are
determined by regulators that bind to metal ions with femtomolar activities. In response to
metal exposure, some microorganisms upregulate the expression of extracellular polymers
or siderophores containing functional groups that are capable of coordinating metal ions
and may be subject to reduced uptake or increased efflux by membrane transporters upon
binding to toxic metals. Many microbes have ability to precipitate metals as metal oxides,
metal sulfides, metal protein aggregates, or metal crystals forming particulates in close
association with cytoplasmic membranes. In addition, some microbes can use cytoplasmic
proteins, e.g., bacterioferritin and metallothioneins, to bind, sequester, or store metals
(Carrondo 2003). Some microbes use metals in specific redox and covalent reactions that

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convert toxic metal species into less toxic forms either oxidative or reductive metabolism
(Fig. 3.11).

Oxidative and reductive biodegradation of environmental pollutants

Few microbes have evolved detoxification mechanisms during their exposure to heavy
metals, e.g., copper, mercury, lead, zinc, cadmium, etc. One of the known examples is
cadmium accumulation in agricultural soils in New Zealand due to extensive use of
superphosphate fertilizer (Loganathan et al. 2003). Due to metal toxicity, microbes have
evolved few defense mechanisms such as metal sequestration, detoxification, and efflux of
ions.

Bacteria sequester heavy metals through their binding with cell membrane, cell wall, and
extracellular polysaccharides (Harrison et al. 2007). Microbes may also detoxify toxic metals
through reduction using various cellular enzymes, e.g., mercury oxidase reduces Hg+2 to Hg,
which has a low evaporation point and, therefore, diffuses from cell (Nies 1999).

Few gram-negative bacteria, e.g., Alcaligenes eutrophus, have evolved a mechanism to fight
with metal toxicity by expelling them from cytoplasm to external environment through
cation/proton antiporter, present at cell membrane (Silver and Phung 1996). Nowadays, the

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microbial ability to transform heavy metals is being extensively used as a tool for
bioremediation

Aerosol

Aerosol is defined as a suspension system of solid or liquid particles in a gas. An aerosol

includes both the particles and the suspending gas, which is usually air. [1] Meteorologists
usually refer them as particle matter - PM2.5 or PM10, depending on their size.[6] Frederick
G. Donnan presumably first used the term aerosol during World War I to describe an aero-
solution, clouds of microscopic particles in air. This term developed analogously to the term
hydrosol, a colloid system with water as the dispersed medium.[7] Primary aerosols contain
particles introduced directly into the gas; secondary aerosols form through gas-to-particle
conversion.[8]

Key aerosol groups include sulfates, organic carbon, black carbon, nitrates, mineral dust,
and sea salt, they usually clump together to form a complex mixture.[6] Various types of
aerosol, classified according to physical form and how they were generated, include dust,
fume, mist, smoke and fog.[9]

There are several measures of aerosol concentration. Environmental science and

environmental health often use the mass concentration (M), defined as the mass of
particulate matter per unit volume, in units such as μg/m3. Also commonly used is the
number concentration (N), the number of particles per unit volume, in units such as number
per m3 or number per cm3.[10]

Particle size has a major influence on particle properties, and the aerosol particle radius or
diameter (dp) is a key property used to characterise aerosols.

Aerosols vary in their dispersity. A monodisperse aerosol, producible in the laboratory,

contains particles of uniform size.

Most aerosols, however, as polydisperse colloidal systems, exhibit a range of particle sizes.[8]
Liquid droplets are almost always nearly spherical, but scientists use an equivalent diameter
to characterize the properties of various shapes of solid particles, some very irregular.

The equivalent diameter is the diameter of a spherical particle with the same value of some
physical property as the irregular particle.[11] The equivalent volume diameter (de) is defined
as the diameter of a sphere of the same volume as that of the irregular particle. [12] Also
commonly used is the aerodynamic diameter,

Generation and applications

People generate aerosols for various purposes, including:

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 as test aerosols for calibrating instruments, performing research, and testing

sampling equipment and air filters;[41]
 to deliver deodorants, paints, and other consumer products in sprays;[42]
 for dispersal and agricultural application
 for medical treatment of respiratory disease;[43] and
 in fuel injection systems and other combustion technology.[44]

Some devices for generating aerosols are:[2]

 Aerosol spray
 Atomizer nozzle or nebulizer
 Electrospray
 Electronic cigarette
 Vibrating orifice aerosol generator (VOAG)

Stability of generated aerosol particles

Stability of nanoparticle agglomerates is critical for estimating size distribution of

aerosolized particles from nano-powders or other sources. At nanotechnology workplaces,
workers can be exposed via inhalation to potentially toxic substances during handling and
processing of nanomaterials.

Nanoparticles in the air often form agglomerates due to attractive inter-particle forces, such
as van der Waals force or electrostatic force if the particles are charged. As a result, aerosol
particles are usually observed as agglomerates rather than individual particles. For exposure
and risk assessments of airborne nanoparticles, it is important to know about the size
distribution of aerosols. When inhaled by humans, particles with different diameters are
deposited in varied locations of the central and periphery respiratory system.

Particles in nanoscale have been shown to penetrate the air-blood barrier in lungs and be
translocated into secondary organs in the human body, such as the brain, heart and liver.
Therefore, the knowledge on stability of nanoparticle agglomerates is important for
predicting the size of aerosol particles, which helps assess the potential risk of them to
human bodies.

Different experimental systems have been established to test the stability of airborne
particles and their potentials to deagglomerate under various conditions. A comprehensive
system recently reported is able to maintain robust aerosolization process and generate
aerosols with stable number concentration and mean size from nano-powders.[45] The
deagglomeration potential of various airborne nanomaterials can be also studied using
critical orifices.[46] In addition, an impact fragmentation device was developed to investigate
bonding energies between particles.[47]

A standard deagglomeration testing procedure could be foreseen with the developments of

the different types of existing systems. The likeliness of deagglomeration of aerosol particles
in occupational settings can be possibly ranked for different nanomaterials if a reference
method is available. For this purpose, inter-laboratory comparison of testing results from

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different setups could be launched in order to explore the influences of system

characteristics on properties of generated nanomaterials aerosols.

Detection

Aerosol can either be measured in-situ or with remote sensing techniques.

In situ observations

Some available in situ measurement techniques include:

 Aerosol mass spectrometer (AMS)

 Differential mobility analyzer (DMA)
 Electrical aerosol spectrometer (EAS)
 Aerodynamic particle sizer (APS)
 Aerodynamic aerosol classifier (AAC)
 Wide range particle spectrometer (WPS)
 Micro-Orifice Uniform Deposit Impactor(MOUDI)
 Condensation particle counter (CPC)
 Epiphaniometer
 Electrical low pressure impactor (ELPI)
 Aerosol particle mass-analyser (APM)
 Centrifugal Particle Mass Analyser (CPMA)

Remote sensing approach

Remote sensing approaches include:

 Sun photometer
 Lidar
 Imaging spectroscopy

Size selective sampling

Particles can deposit in the nose, mouth, pharynx and larynx (the head airways region),
deeper within the respiratory tract (from the trachea to the terminal bronchioles), or in the
alveolar region.[48] The location of deposition of aerosol particles within the respiratory
system strongly determines the health effects of exposure to such aerosols.[48] This
phenomenon led people to invent aerosol samplers that select a subset of the aerosol
particles that reach certain parts of the respiratory system.[49] Examples of these subsets of
the particle-size distribution of an aerosol, important in occupational health, include the
inhalable, thoracic, and respirable fractions. The fraction that can enter each part of the
respiratory system depends on the deposition of particles in the upper parts of the
airway.[50] The inhalable fraction of particles, defined as the proportion of particles originally
in the air that can enter the nose or mouth, depends on external wind speed and direction
and on the particle-size distribution by aerodynamic diameter.[51] The thoracic fraction is the
proportion of the particles in ambient aerosol that can reach the thorax or chest region. [52]
The respirable fraction is the proportion of particles in the air that can reach the alveolar
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region.[53] To measure the respirable fraction of particles in air, a pre-collector is used with a
sampling filter. The pre-collector excludes particles as the airways remove particles from
inhaled air. The sampling filter collects the particles for measurement. It is common to use
cyclonic separation for the pre-collector, but other techniques include impactors, horizontal
elutriators, and large pore membrane filters.[54]

Two alternative size-selective criteria, often used in atmospheric monitoring, are PM 10 and
PM2.5. PM10 is defined by ISO as particles which pass through a size-selective inlet with a 50%
efficiency cut-off at 10 μm aerodynamic diameter and PM2.5 as particles which pass through
a size-selective inlet with a 50% efficiency cut-off at 2.5 μm aerodynamic diameter. PM10
corresponds to the "thoracic convention" as defined in ISO 7708:1995, Clause 6; PM 2.5
corresponds to the "high-risk respirable convention" as defined in ISO 7708:1995, 7.1.[55]
The United States Environmental Protection Agency replaced the older standards for
particulate matter based on Total Suspended Particulate with another standard based on
PM10 in 1987[56] and then introduced standards for PM2.5 (also known as fine particulate
matter) in 1997

ATMOSPHERIC

Several types of atmospheric aerosol have a significant effect on Earth's climate: volcanic,
desert dust, sea-salt, that originating from biogenic sources and human-made. Volcanic
aerosol forms in the stratosphere after an eruption as droplets of sulfuric acid that can
prevail for up to two years, and reflect sunlight, lowering temperature.

Desert dust, mineral particles blown to high altitudes, absorb heat and may be responsible
for inhibiting storm cloud formation. Human-made sulfate aerosols, primarily from burning
oil and coal, affect the behavior of clouds.[58]

Although all hydrometeors, solid and liquid, can be described as aerosols, a distinction is
commonly made between such dispersions (i.e. clouds) containing activated drops and
crystals, and aerosol particles. The atmosphere of Earth contains aerosols of various types
and concentrations, including quantities of:

 natural inorganic materials: fine dust, sea salt, or water droplets

 natural organic materials: smoke, pollen, spores, or bacteria
 anthropogenic products of combustion such as: smoke, ashes or dusts

Aerosols can be found in urban ecosystems in various forms, for example:

 Dust
 Cigarette smoke
 Mist from aerosol spray cans

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 Soot or fumes in car exhaust

The presence of aerosols in the earth's atmosphere can influence its climate, as well as
human health.

Effects

 Volcanic eruptions release large amounts of sulphuric acid, hydrogen sulfide and
hydrochloric acid into the atmosphere. These gases represent aerosols and
eventually return to earth as acid rain, having a number of adverse effects on the
environment and human life.[59]
 Aerosols interact with the Earth's energy budget in two ways, directly and indirectly.

E.g., a direct effect is that aerosols scatter and absorb incoming solar radiation.[60]
This will mainly lead to a cooling of the surface (solar radiation is scattered back to
space) but may also contribute to a warming of the surface (caused by the
absorption of incoming solar energy).[61] This will be an additional element to the
greenhouse effect and therefore contributing to the global climate change.[62]

The indirect effects refer to the aerosols interfering with formations that interact
directly with radiation. For example, they are able to modify the size of the cloud
particles in the lower atmosphere, thereby changing the way clouds reflect and
absorb light and therefore modifying the Earth's energy budget.[59]

There is evidence to suggest that anthropogenic aerosols actually offset the effects
of greenhouse gases in some areas, which is why the Northern Hemisphere shows
slower surface warming than the Southern Hemisphere, although that just means
that the Northern Hemisphere will absorb the heat later through ocean currents
bringing warmer waters from the South.[63] On a global scale however, aerosol
cooling decreases greenhouse-gases-induced heating without offsetting it
completely. [64]

 When aerosols absorb pollutants, it facilitates the deposition of pollutants to the

surface of the earth as well as to bodies of water.[62] This has the potential to be
damaging to both the environment and human health.

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 Aerosols in the 20 μm range show a particularly long persistence time in air

conditioned rooms due to their "jet rider" behaviour (move with air jets,
gravitationally fall out in slowly moving air);[65] as this aerosol size is most effectively
adsorbed in the human nose,[66] the primordial infection site in COVID-19, such
aerosols may contribute to the pandemic.

 Aerosol particles with an effective diameter smaller than 10 μm can enter the
bronchi, while the ones with an effective diameter smaller than 2.5 μm can enter as
far as the gas exchange region in the lungs,[67] which can be hazardous to human
health.

Biogeochemical cycle,

any of the natural pathways by which essential elements of living matter are circulated. The
term biogeochemical is a contraction that refers to the consideration of the biological,
geological, and chemical aspects of each cycle.

Carbon cycle
The generalized carbon cycle.

Elements within biogeochemical cycles flow in various forms from the nonliving (abiotic)
components of the biosphere to the living (biotic) components and back. In order for the

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living components of a major ecosystem (e.g., a lake or a forest) to survive, all the chemical
elements that make up living cells must be recycled continuously.

Each biogeochemical cycle can be considered as having a reservoir (nutrient) pool—a

larger, slow-moving, usually abiotic portion—and an exchange (cycling) pool—a smaller but
more-active portion concerned with the rapid exchange between the biotic and abiotic
aspects of an ecosystem.

Nitrogen cycle
The nitrogen cycle.

Biogeochemical cycles can be classed as gaseous, in which the reservoir is the air or
the oceans (via evaporation), and sedimentary, in which the reservoir
is Earth’s crust. Gaseous cycles include those of nitrogen, oxygen, carbon,
and water; sedimentary cycles include those of iron, calcium, phosphorus, sulfur, and other
more-earthbound elements.

Gaseous cycles tend to move more rapidly than do sedimentary ones and to adjust more
readily to changes in the biosphere because of the large atmospheric reservoir. Local
accumulations of carbon dioxide (CO2), for example, are soon dissipated by winds or taken
up by plants. Extraordinary disturbances (such as global warming) and more-frequent
local disturbances (such as wildfires and storm-driven events) can, however, seriously affect
the capacity for self-adjustment.

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Hydrologic cycle
This diagram shows how, in the hydrologic cycle, water is transferred between the land
surface, the ocean, and the atmosphere.

Sedimentary cycles vary from one element to another, but each cycle consists
fundamentally of a solution (or water-related) phase and a rock (or sediment) phase. In the
solution phase, weathering releases minerals from Earth’s crust in the form of salts, some of
which dissolve in water, pass through a series of organisms, and ultimately reach the deep
seas, where they settle out of circulation indefinitely. In the rock phase, other salts deposit
out as sediment and rock in shallow seas, eventually to be weathered and recycled.

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Phosphorus cycle

Phosphorus, which cycles primarily through the terrestrial and aquatic environments, is one
of the most-important elements influencing the growth of plants.

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Sulfur cycle
Major sulfur-producing sources include sedimentary rocks, which release hydrogen sulfide
gas, and human sources, such as smelters and fossil-fuel combustion, both of which release
sulfur dioxide into the atmosphere.

Plants and some animals obtain their nutrient needs from solutions in the environment.
Other animals acquire the bulk of their needs from the plants and animals that they
consume. After the death of an organism, the elements fixed in its body are returned to
the environment through the action of decomposers (decay organisms such
as bacteria, insects, and fungi) and become available to other living organisms again.

oxygen cycle
The generalized oxygen cycle.

Biosafety in the Laboratory

is a concise set of practical guidelines for handling and disposing of biohazardous material.
The consensus of top experts in laboratory safety, this volume provides the information
needed for immediate improvement of safety practices. It discusses high- and low-risk
biological agents (including the highest-risk materials handled in labs today), presents the
"seven basic rules of biosafety," addresses special issues such as the shipping of dangerous
materials, covers waste disposal in detail, offers a checklist for administering laboratory
safety--and more.

Contents

 Committee on Hazardous Biological Substances in the Laboratory

 Board on Chemical Sciences and Technology
 Commission on Physical Sciences, Mathematics, and Resources
 Preface
 1. Introduction, Overview, and Recommendations
o A INTRODUCTION
o B OVERVIEW

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oC RECOMMENDATIONS
 2. Descriptive Epidemiology of Occupational Infections of Laboratory Workers
o A INTRODUCTION
o B THE EPIDEMIOLOGIC TRIAD
o C LABORATORY-ASSOCIATED INFECTIONS
 3. Safe Handling of Infectious Agents
o A— GUIDELINES FOR HANDLING PATHOGENIC MICROORGANISMS
o B ORGANISMS POSING SPECIAL RISKS
o C HAZARDS FROM VERTEBRATE ANIMALS AND INSECTS IN THE LABORATORY
o D PRIMARY AND CONTINUOUS CELL CULTURES
o E HANDLING OF NECROPSY AND SURGICAL SPECIMENS
o F GOOD LABORATORY PRACTICES
o G TRANSPORTATION AND SHIPMENT OF SPECIMENS
o H LABELING OF SPECIMENS WITHIN THE LABORATORY
o I PREVENTION OF AEROSOL AND DROPLET GENERATION
o J CONTAINMENT EQUIPMENT
o K BIOSAFETY IN LARGE-SCALE PRODUCTION
o L BIOSAFETY IN PHYSICIANS' OFFICE LABORATORIES AND OTHER SMALL
VOLUME CLINICAL LABORATORIES
 4. Safe Disposal of Infectious Laboratory Waste
o A INTRODUCTION
o B INFECTIOUS POTENTIAL OF LABORATORY WASTE
o C CHARACTERISTICS OF INFECTIOUS LABORATORY WASTE
o D RESPONSIBILITY FOR THE SAFE HANDLING AND DISPOSAL OF INFECTIOUS
WASTE
o 1 Generators of Infectious Waste
o E WASTE HANDLING AND TREATMENT METHODS
o F INFECTIOUS WASTE REQUIRING SPECIAL CONSIDERATION
 5. Safety Management
o A ADMINISTRATIVE ORGANIZATION AND RESPONSIBILITIES
o B FACILITIES
o C OPERATIONS
o D MEDICAL PROGRAM
o E EMERGENCIES
o F REGULATION AND ACCREDITATION
o G TEACHING BIOSAFETY IN ACADEMIC SETTINGS

Extreme environment

An extreme environment is a habitat that is considered very hard to survive in due to its
considerably extreme conditions such as temperature, accessibility to different energy
sources or under high pressure. For an area to be considered an extreme environment, it
must contain certain conditions and aspects that are considered very hard for other life
forms to survive. Pressure conditions may be extremely high or low; high or low content of
oxygen or carbon dioxide in the atmosphere; high levels of radiation, acidity, or alkalinity;
absence of water; water containing a high concentration of salt or sugar; the presence of
sulphur, petroleum, and other toxic substances.[1]

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Examples of extreme environments include the geographical poles, very arid deserts,
volcanoes, deep ocean trenches, upper atmosphere, outer space, and the environments of
every planet in the Solar System except the Earth. Any organisms living in these conditions
are often very well adapted to their living circumstances, which is usually a result of long-
term evolution. Physiologists have long known that organisms living in extreme
environments are especially likely to exhibit clear examples of evolutionary adaptation
because of the presumably intense past natural selection they have experienced

Types

Among extreme environments are places that are alkaline, acidic, or unusually hot or cold or
salty, or without water or oxygen. There are also places altered by humans, such as mine
tailings or oil impacted habitats. [3][4]

 Alkaline: broadly conceived as natural habitats above pH 9 whether persistently, or

with regular frequency or for protracted periods of time.
 Acidic: broadly conceived as natural habitats below pH 5 whether persistently, or
with regular frequency or for protracted periods of time.
 Extremely cold: broadly conceived habitats periodically or consistently below -17 °C
either persistently, or with regular frequency or for protracted periods of time.
Includes montane sites, polar sites, and deep ocean habitats.
 Extremely hot: broadly conceived habitats periodically or consistently in excess of
40 °C either persistently, or with regular frequency or for protracted periods of time.
Includes sites with geothermal influences such as Yellowstone and comparable
locations worldwide or deep-sea vents.
 Hypersaline: environments with salt concentrations greater than that of seawater,
that is, >3.5%. Includes salt lakes.
 Under pressure: broadly conceived as habitats under extreme hydrostatic pressure –
i.e. aquatic habitats deeper than 2000 meters and enclosed habitats under pressure.
Includes habitats in oceans and deep lakes.
 Radiation: broadly conceived as habitats exposed to abnormally high radiation or of
radiation outside the normal range of light. Includes habitats exposed to high UV and
IR radiation.
 Without water: broadly conceived as habitats without free water whether
persistently, or with regular frequency or for protracted periods of time. Includes hot
and cold desert environments, and some endolithic habitats
 Without oxygen: broadly conceived as habitats without free oxygen – whether
persistently, or with regular frequency, or for protracted periods of time. Includes
habitats in deeper sediments.
 Altered by humans: i.e. anthropogenically impacted habitats. Includes mine tailings,
oil impacted habitats, and pollution by heavy metals or organic compounds.
 Without light: deep ocean environments and habitats such as caves.
 Void of food: areas on earth that lack an abundance of food such as the vast ocean,
desert and high country.
 Extreme pressure: deep ocean areas

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Extreme habitats

Many different habitats can be considered extreme environments, such as the polar ice
caps, the driest spots in deserts, and abysmal depths in the ocean. Many different places on
the Earth demand that species become highly specialized if they are to survive. In particular,
microscopic organisms that can't be seen with the naked eye often thrive in surprising
places.[5]

Polar regions

Owing to the dangerously low temperatures, the number of species that can survive in
these remote areas is very slim. Over years of evolution and adaptation to this extremely
cold environment, both microscopic and larger species have survived and thrived no matter
what conditions they have faced.[6] By changing their eating patterns and due to their dense
pelt or their body fat, only a few species have been capable of adapting to such harsh
conditions and have learned how to thrive in these cold environments.[7]

Deserts

A desert is known for its extreme temperatures and extremely dry climate. The type of
species that reside in this area have adapted to these harsh conditions over years and years.
Species that are able to store water and have learned how to protect themselves from the
Sun's harsh rays are the only ones that are capable of surviving in these extreme
environments.[8]

Oceans

The oceans depths and temperatures contains some of the most extreme conditions for any
species to survive. The deeper one travels, the higher the pressure and the lower the
visibility gets, causing completely blacked out conditions.[9] Many of these conditions are
too intense for humans to travel to, so instead of sending humans down to these depths to
collect research, scientists are using smaller submarines or deep sea drones to study these
creatures and extreme environments.[10]

Types of species in extreme environments

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The origin of each species

There are many different species that are either commonly known or not known amongst
many people. These species have either adapted over time into these extreme
environments or they have resided their entire life no matter how many generations. The
different species are able to live in these environments because of their flexibility with
adaptation. Many can adapt to different climate conditions and hibernate, if need be, to
survive.

The following list contains only a few species that live in extreme environments.

Examples

 Giant kangaroo rat

 Certain species of frogs
 Thermotolerant worms (Alvinella pompejana)
 Devil worms, Halicephalobus mephisto
 Greenland shark
 Marine microorganism
 Bdelloidea
 Tardigrade (waterbear)
 Himalayan jumping spider, Euophrys omnisuperstes
 Cockroach

Extreme environment examples

 Antarctica

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 Dead Sea
 Mammoth Hot Springs
 Mariana Trench
 Mono Lake
 Mount Everest
 Sahara

Archaebacteria

are known to be the oldest living organisms on earth. They belong to the kingdom Monera
and are classified as bacteria because they resemble bacteria when observed under a
microscope. Apart from this, they are completely distinct from prokaryotes. However, they
share slightly common characteristics with the eukaryotes.

These can easily survive under very harsh conditions such as the bottom of the sea and the
volcanic vents and are thus known as extremophiles.

Archaebacteria

Characteristics of Archaebacteria

Following are the important characteristics of archaebacteria:

 Archaebacteria are obligate or facultative anaerobes, i.e., they flourish in the

absence of oxygen and that is why only they can undergo methanogenesis.
 The cell membranes of the Archaebacteria are composed of lipids.
 The rigid cell wall provides shape and support to the Archaebacteria. It also protects
the cell from bursting under hypotonic conditions.

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 The cell wall is composed of Pseudomurein, which prevents archaebacteria from the
effects of Lysozyme. Lysozyme is an enzyme released by the immune system of the
host, which dissolves the cell wall of pathogenic bacteria.
 These do not possess membrane-bound organelles such as nuclei, endoplasmic
reticulum, mitochondria, lysosomes or chloroplast. Its thick cytoplasm contains all
the compounds required for nutrition and metabolism.
 They can live in a variety of environments and are hence called extremophiles. They
can survive in acidic and alkaline aquatic regions, and also in temperature above
boiling point.
 They can withstand a very high pressure of more than 200 atmospheres.
 Archaebacteria are indifferent towards major antibiotics because they contain
plasmids which have antibiotic resistance enzymes.
 The mode of reproduction is asexual, known as binary fission.
 They perform unique gene transcription.
 The differences in their ribosomal RNA suggest that they diverged from both
prokaryotes and eukaryotes.

Types of Archaebacteria

Archaebacteria are classified on the basis of their phylogenetic relationship. The major types
of Archaebacteria are discussed below:

Crenarchaeota

The Crenarchaeota are Archaea, which exist in a broad range of habitats. They are tolerant
to extreme heat or high temperatures. They have special proteins that help them to
function at temperatures as high as 230 degrees Celsius. They can be found in deep-sea
vents and hot springs, regions with superheated water. These include thermophiles,
hyperthermophiles, and thermoacidophiles.

Euryarchaeota

These can survive under extremely alkaline conditions and have the ability to produce
methane, unlike any other living being on earth. These include methanogens and halophiles.

Korarchaeota

They possess the genes common with Crenarchaeota and Euryarchaeota. All three are
believed to have descended from a common ancestor. These are supposed to be the oldest
surviving organism on earth. These include hyperthermophiles.

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Thaumarchaeota

These include archaea that oxidize ammonia.

Nanoarchaeota

This is an obligate symbiont of archaea belonging to the genus Ignicoccus.

Importance of Archaebacteria

The importance of archaebacteria can be understood from the following points:

 Archaebacteria have compelled the scientists to reconsider the common definition

of species. Species are a group with gene flow within its members. The
archaebacteria exhibit gene flow across its species.
 The Archaebacteria are methanogens, i.e., they are capable of producing methane.
They act on the organic matter and decompose it to release methane which is then
used for cooking and lighting.
Examples of Archaebacteria

Following are the important examples of archaebacteria:

Lokiarcheota

It is a thermophilic archaebacterium found in deep-sea vents known as the Loki’s castle. It

has a unique genome. Some of the genes of the genome are involved in phagocytosis. They
also possess the eukaryotic genes that are used by the eukaryotes to control their shapes. It
is believed that Lokiarcheota and eukaryotes shared a common ancestor several billion
years ago.

Methanobrevibacter smithii

It is a methane-producing bacteria found in the human gut. It helps in the breakdown of

complex plant sugars and extracts energy from the food consumed by us. Some help to
protect against colon cancer. People suffering from colon cancer and obesity have very high
levels of Euryarchaeota bacteria in their gut.

The Archaebacteria cannot perform photosynthesis and show high levels of gene transfer
between lineages. The discovery of Archaebacteria has made scientists believe that life can
exist even in extreme environmental conditions.

Nutrients cycling in soil

Microorganisms are responsible for the degradation of organic matter, which controls the
release of plant nutrients, but is also important for the maintenance of soil structure and
sustainability of soil quality for plant growth

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The role of microorganisms bacteria and fungi in nutrient cycling?

Soil microbes play a vital role in the sustained growth of plants. They decompose and
recycling nutrients bound in organic materials. They help access minerals in rocks large and
small. And, they can even refine nitrogen from the air into a useful form for plants!

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UNIT 3 – METABOLISM OF MICROORGANISM

Microbial metabolism is the means by which a microbe obtains the energy and nutrients
(e.g. carbon) it needs to live and reproduce. Microbes use many different types of metabolic
strategies and species can often be differentiated from each other based on metabolic
characteristics. The specific metabolic properties of a microbe are the major factors in
determining that microbe's ecological niche, and often allow for that microbe to be useful in
industrial processes or responsible for biogeochemical cycles.

Types

Flow chart to determine the metabolic characteristics of microorganisms

All microbial metabolisms can be arranged according to three principles:

1. How the organism obtains carbon for synthesizing cell mass:[1]

 autotrophic – carbon is obtained from carbon dioxide (CO2)

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 heterotrophic – carbon is obtained from organic compounds

 mixotrophic – carbon is obtained from both organic compounds and by fixing carbon
dioxide

2. How the organism obtains reducing equivalents (hydrogen atoms or electrons) used
either in energy conservation or in biosynthetic reactions:

 lithotrophic – reducing equivalents are obtained from inorganic compounds

 organotrophic – reducing equivalents are obtained from organic compounds

3. How the organism obtains energy for living and growing:

 phototrophic – energy is obtained from light[2]

 chemotrophic – energy is obtained from external chemical compounds[3]

In practice, these terms are almost freely combined. Typical examples are as follows:

 Chemolithoautotrophs obtain energy from the oxidation of inorganic compounds

and carbon from the fixation of carbon dioxide. Examples: Nitrifying bacteria, sulfur-
oxidizing bacteria, iron-oxidizing bacteria, Knallgas-bacteria[4]
 Photolithoautotrophs obtain energy from light and carbon from the fixation of
carbon dioxide, using reducing equivalents from inorganic compounds. Examples:
Cyanobacteria (water (H
2O) as reducing equivalent = hydrogen donor), Chlorobiaceae, Chromatiaceae
(hydrogen sulfide (H
2S) as hydrogen donor), Chloroflexus (hydrogen (H
2) as reducing equivalent donor)
 Chemolithoheterotrophs obtain energy from the oxidation of inorganic compounds,
but cannot fix carbon dioxide (CO2). Examples: some Thiobacilus, some Beggiatoa,
some Nitrobacter spp., Wolinella (with H
2 as reducing equivalent donor), some Knallgas-bacteria, some sulfate-reducing
bacteria[citation needed]
 Chemoorganoheterotrophs obtain energy, carbon, and hydrogen for biosynthetic
reactions from organic compounds. Examples: most bacteria, e. g. Escherichia coli,
Bacillus spp., Actinomycetota
 Photoorganoheterotrophs obtain energy from light, carbon and reducing
equivalents for biosynthetic reactions from organic compounds. Some species are
strictly heterotrophic, many others can also fix carbon dioxide and are mixotrophic.
Examples: Rhodobacter, Rhodopseudomonas, Rhodospirillum, Rhodomicrobium,
Rhodocyclus, Heliobacterium, Chloroflexus (alternatively to photolithoautotrophy
with hydrogen)

Fermentation

Fermentation is a specific type of heterotrophic metabolism that uses organic carbon

instead of oxygen as a terminal electron acceptor. This means that these organisms do not

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use an electron transport chain to oxidize NADH to NAD+

and therefore must have an alternative method of using this reducing power and
maintaining a supply of NAD+
for the proper functioning of normal metabolic pathways (e.g. glycolysis).

As oxygen is not required, fermentative organisms are anaerobic. Many organisms can use
fermentation under anaerobic conditions and aerobic respiration when oxygen is present.

These organisms are facultative anaerobes. To avoid the overproduction of NADH,

obligately fermentative organisms usually do not have a complete citric acid cycle. Instead
of using an ATP synthase as in respiration, ATP in fermentative organisms is produced by
substrate-level phosphorylation where a phosphate group is transferred from a high-energy
organic compound to ADP to form ATP.

As a result of the need to produce high energy phosphate-containing organic compounds

(generally in the form of Coenzyme A-esters) fermentative organisms use NADH and other
cofactors to produce many different reduced metabolic by-products, often including
hydrogen gas (H2).

These reduced organic compounds are generally small organic acids and alcohols derived
from pyruvate, the end product of glycolysis. Examples include ethanol, acetate, lactate, and
butyrate. Fermentative organisms are very important industrially and are used to make
many different types of food products. The different metabolic end products produced by
each specific bacterial species are responsible for the different tastes and properties of each
food.

Not all fermentative organisms use substrate-level phosphorylation. Instead, some

organisms are able to couple the oxidation of low-energy organic compounds directly to the
formation of a proton motive force or sodium-motive force and therefore ATP synthesis.
Examples of these unusual forms of fermentation include succinate fermentation by
Propionigenium modestum and oxalate fermentation by Oxalobacter formigenes. These
reactions are extremely low-energy yielding. Humans and other higher animals also use
fermentation to produce lactate from excess NADH, although this is not the major form of
metabolism as it is in fermentative microorganisms

Aerobic respiration

Aerobic metabolism occurs in Bacteria, Archaea and Eucarya. Although most bacterial
species are anaerobic, many are facultative or obligate aerobes. The majority of archaeal
species live in extreme environments that are often highly anaerobic. There are, however,
several cases of aerobic archaea such as Haiobacterium, Thermoplasma, Sulfolobus and
Yymbaculum. Most of the known eukaryotes carry out aerobic metabolism within their
mithchondria which is an organelle that had a symbiogenesis origin from prokarya . All
aerobic organisms contain oxidases of the cytochrome oxidase super family, but some

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members of the Proteobacteria (E. coli and Acetobacter) can also use an unrelated
cytochrome bd complex as a respiratory terminal oxidase.[6]

Anaerobic respiration

While aerobic organisms during respiration use oxygen as a terminal electron acceptor,
anaerobic organisms use other electron acceptors. These inorganic compounds have a lower
reduction potential than oxygen,[3] meaning that respiration is less efficient in these
organisms and leads to slower growth rates than aerobes. Many facultative anaerobes can
use either oxygen or alternative terminal electron acceptors for respiration depending on
the environmental conditions.

Most respiring anaerobes are heterotrophs, although some do live autotrophically. All of the
processes described below are dissimilative, meaning that they are used during energy
production and not to provide nutrients for the cell (assimilative). Assimilative pathways for
many forms of anaerobic respiration are also known.

Denitrification – nitrate as electron acceptor

Denitrification

Denitrification is the utilization of nitrate (NO−

3) as a terminal electron acceptor. It is a widespread process that is used by many members
of the Proteobacteria. Many facultative anaerobes use denitrification because nitrate, like
oxygen, has a high reduction potential. Many denitrifying bacteria can also use ferric iron
(Fe3+
) and some organic electron acceptors. Denitrification involves the stepwise reduction of
nitrate to nitrite (NO−
2), nitric oxide (NO), nitrous oxide (N
2O), and dinitrogen (N
2) by the enzymes nitrate reductase, nitrite reductase, nitric oxide reductase, and nitrous
oxide reductase, respectively. Protons are transported across the membrane by the initial
NADH reductase, quinones, and nitrous oxide reductase to produce the electrochemical
gradient critical for respiration. Some organisms (e.g. E. coli) only produce nitrate reductase
and therefore can accomplish only the first reduction leading to the accumulation of nitrite.
Others (e.g. Paracoccus denitrificans or Pseudomonas stutzeri) reduce nitrate completely.
Complete denitrification is an environmentally significant process because some
intermediates of denitrification (nitric oxide and nitrous oxide) are important greenhouse
gases that react with sunlight and ozone to produce nitric acid, a component of acid rain.
Denitrification is also important in biological wastewater treatment where it is used to
reduce the amount of nitrogen released into the environment thereby reducing
eutrophication. Denitrification can be determined via a nitrate reductase test.

Nutrients of Human Metabolism

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Carbohydrates, lipids, and proteins are the major constituents of foods and serve as fuel
molecules for the human body. The digestion (breaking down into smaller pieces) of these
nutrients in the alimentary tract and the subsequent absorption (entry into the
bloodstream) of the digestive end products make it possible for tissues and cells to
transform the potential chemical energy of food into useful work.

The major absorbed end products of food digestion are monosaccharides, mainly glucose
(from carbohydrates); monoacylglycerol and long-chain fatty acids (from lipids); and small
peptides and amino acids (from protein). Once in the bloodstream, different cells can
metabolize these nutrients. We have long known that these three classes of molecules are
fuel sources for human metabolism, yet it is a common misconception (especially among
undergraduates) that human cells use only glucose as a source of energy. This
misinformation may arise from the way most textbooks explain energy metabolism,
emphasizing glycolysis (the metabolic pathway for glucose degradation) and omitting fatty
acid or amino acid oxidation. Here we discuss how the three nutrients (carbohydrates,
proteins, and lipids) are metabolized in human cells in a way that may help avoid this
oversimplified view of the metabolism.

Growth phase in microbiology?

The bacterial growth curve represents the number of live cells in a bacterial population over
a period of time. There are four distinct phases of the growth curve: lag, exponential (log),
stationary, and death. The initial phase is the lag phase where bacteria are metabolically
active but not dividing.

PHASE OF THE BACTERIAL GROWTH

Bacteria are prokaryotic organisms that most commonly replicate by the asexual process of
binary fission. These microbes reproduce rapidly at an exponential rate under favorable
conditions. When grown in culture, a predictable pattern of growth in a bacterial population
occurs. This pattern can be graphically represented as the number of living cells in a
population over time and is known as a bacterial growth curve. Bacterial growth cycles in a
growth curve consist of four phases: lag, exponential (log), stationary, and death.

Bacteria require certain conditions for growth, and these conditions are not the same for all
bacteria. Factors such as oxygen, pH, temperature, and light influence microbial growth.
Additional factors include osmotic pressure, atmospheric pressure, and moisture
availability. A bacterial population's generation time, or time it takes for a population to
double, varies between species and depends on how well growth requirements are met.

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Phases of the Bacterial Growth Cycle

In nature, bacteria do not experience perfect environmental conditions for growth. As such,
the species that populate an environment change over time. In a laboratory, however,
optimal conditions can be met by growing bacteria in a closed culture environment. It is
under these conditions that the curve pattern of bacterial growth can be observed.

The bacterial growth curve represents the number of live cells in a bacterial population over
a period of time.

 Lag Phase: This initial phase is characterized by cellular activity but not growth. A
small group of cells are placed in a nutrient rich medium that allows them to
synthesize proteins and other molecules necessary for replication. These cells
increase in size, but no cell division occurs in the phase.

 Exponential (Log) Phase: After the lag phase, bacterial cells enter the exponential or
log phase. This is the time when the cells are dividing by binary fission and doubling
in numbers after each generation time. Metabolic activity is high as DNA, RNA, cell
wall components, and other substances necessary for growth are generated for
division. It is in this growth phase that antibiotics and disinfectants are most
effective as these substances typically target bacteria cell walls or the protein
synthesis processes of DNA transcription and RNA translation.

 Stationary Phase: Eventually, the population growth experienced in the log phase
begins to decline as the available nutrients become depleted and waste products
start to accumulate. Bacterial cell growth reaches a plateau, or stationary phase,
where the number of dividing cells equal the number of dying cells.

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 This results in no overall population growth. Under the less favorable conditions,
competition for nutrients increases and the cells become less metabolically active.
Spore forming bacteria produce endospores in this phase and pathogenic bacteria
begin to generate substances (virulence factors) that help them survive harsh
conditions and consequently cause disease.

 Death Phase: As nutrients become less available and waste products increase, the
number of dying cells continues to rise. In the death phase, the number of living cells
decreases exponentially and population growth experiences a sharp decline.
 As dying cells lyse or break open, they spill their contents into the environment
making these nutrients available to other bacteria. This helps spore producing
bacteria to survive long enough for spore production. Spores are able to survive the
harsh conditions of the death phase and become growing bacteria when placed in an
environment that supports life.

Aerobic and Anaerobic respiration.

Cellular respiration is a process that takes place inside the cells where energy is released by
the breakdown of glucose molecules. The process can be conveniently divided into two
categories based on the usage of oxygen, namely aerobic and anaerobic respiration.

Table of Contents

 Differences
o Aerobic
o Anaerobic
 Conclusion

Let us have a look at the major difference between aerobic and anaerobic respiration.

Difference Between Aerobic and Anaerobic Respiration

The primary difference between aerobic and anaerobic respiration is the presence or
absence of oxygen during the processes. More detailed differences are between the two are
as follows:

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Aerobic Respiration Anaerobic Respiration

Oxygen is present when this form of respiration Oxygen is absent when this form of respiration
takes place. takes place.

Gases are not exchanged in this form of

Gases are exchanged in this form of respiration.
respiration.

It can be found in the cytoplasm and the

It can be found only in the cytoplasm.
mitochondria.

Glucose breaks down into carbon dioxide and Glucose breaks down into ethyl alcohol, carbon
water. dioxide and energy.

Lower organisms such as bacteria and yeast

All higher organisms such as mammals have
use this type. In other organisms, it occurs
this type of respiration.
during heavy activities.

However, it is a misconception that humans and other multicellular organisms use only
aerobic respiration. This is disproven by the fact that our muscles, during vigorous exercises,
undergo anaerobic respiration, where lactic acid is produced as the waste-byproduct
instead of carbon dioxide.

Aerobic and Anaerobic Respiration

What is Aerobic Respiration?

As already stated, cellular respiration can be of two types: aerobic and anaerobic. Aerobic
means “with air”. Therefore, aerobic respiration is the process of cellular respiration that uses
oxygen to produce energy from food. This type of respiration is common in most of the plants
and animals, including humans, birds and other mammals.

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While breathing, we inhale air that contains oxygen and we exhale air rich in carbon dioxide.
As we breathe in, the oxygen-rich air is transported to all the parts of our body and ultimately
to each cell. Inside the cell, the food, which contains glucose, is broken down into carbon
dioxide and water with the help of oxygen. The process of breaking down the food particles
releases energy, which is then utilized by our body. The energy released via aerobic
respiration helps plants and animals, including us, grow.

The process can be simply explained with the help of the following equation:

Glucose + Oxygen → Carbon dioxide + Water + Energy

Aerobic respiration is a continuous process and it happens all the time inside the cells of
animals and plants.

What is Anaerobic Respiration?

Anaerobic means “without air”. Therefore, this type of cellular respiration does not use
oxygen to produce energy. Sometimes there is not enough oxygen around for some
organisms to respire, but they still need the energy to survive. Due to lack of oxygen, they
carry out respiration in the absence of oxygen to produce the energy they require, which is
referred to as anaerobic respiration. Anaerobic respiration usually occurs in lower plants and
microorganisms. In the absence of oxygen, the glucose derived from food is broken down into
alcohol and carbon dioxide along with the production of energy.

Glucose → Alcohol + Carbon dioxide + Energy

 Anaerobic respiration is also used by multi-cellular organisms, like us, as a temporary

response to oxygen-less conditions. During heavy or intensive exercise such as
running, sprinting, cycling or weight lifting, our body demands high energy. As the
supply of oxygen is limited, the muscle cells inside our body resort to anaerobic
respiration to fulfil the energy demand.
 How do you feel when you exercise too much? Have you ever wondered why you get
those muscle cramps when you run very fast? Anaerobic respiration is the culprit to

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be blamed. Cramps occur when muscle cells respire anaerobically. Partial breakdown
of glucose, due to lack of oxygen, produces lactic acid and the accumulation of lactic
acid causes muscle cramps. That is why a hot shower after heavy sports relieve the
cramps as it improves blood circulation in the body, which in turn enhances the supply
of oxygen to the cells.

Glucose → Lactic acid + Energy

Anaerobic respiration produces a relatively lesser amount of energy as compared to aerobic

respiration, as glucose is not completely broken down in the absence of oxygen.

Conclusion

The fundamental difference between aerobic and anaerobic respiration is the usage of
oxygen in the process of cellular respiration. Aerobic respiration, as the name suggests, is
the process of producing the energy required by cells using oxygen. The by-product of this
process produces carbon dioxide along with ATP – the energy currency of the cells.
Anaerobic respiration is similar to aerobic respiration, except, the process happens without
the presence of oxygen. Consequently, the by-products of this process are lactic acid and
ATP.

Contrary to popular belief, multicellular organisms, including humans, use anaerobic

respiration to produce energy, though this only happens when the muscles do not get
adequate oxygen due to extremely vigorous activities.

What is Respiration?

Respiration is a biochemical process which is common in all living organisms. In this process,
there is the movement of air in and out of the lungs.

List out the different types of Respiration?

There are two types of Respiration:

1. Aerobic Respiration — Takes place in the presence of oxygen.

2. Anaerobic Respiration –Takes place in the absence of oxygen.

What is the overall equation of aerobic cellular respiration?

The equation for aerobic cellular respiration is:

C6H12O6 + 6O2 ————–> 6CO2 + 6H2O + ATP

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List out the different types of Anaerobic Respiration?

There are two main types of anaerobic respiration:

1. Alcoholic fermentation
2. Lactic acid fermentation.

Name the different stages of Aerobic Respiration?

The three stages of Aerobic Cellular Respiration are:

1. Glycolysis
2. The Krebs cycle
3. Oxidative phosphorylation.

Where does the aerobic and anaerobic respiration occur in the cell?

In the cell, Aerobic respiration occurs within the mitochondria of a cell, and the anaerobic
respiration occurs within the cytoplasm of a cell.

DIFFERENCE BETWEEN GLYCOLYSIS & KREBS CYCLE

Respiration is an essential process that occurs in all living beings in which oxygen is utilised
and carbon dioxide is released from the body. The mechanism of cellular respiration
involves the following mechanism:

 Glycolysis – Partial breakdown of Glucose to Pyruvic acid (Anaerobic)

 Krebs Cycle – Complete oxidation of Pyruvate to release Carbon dioxide (Aerobic
respiration)
 Electron Transport system – Oxidation of NADH and FADH2 to generate ATP

Here, in the article, let us discuss the difference between the Krebs Cycle and glycolysis but
first let us take a look at what each of these terms means.

Glycolysis – It is an anaerobic process, which occurs in the cytoplasm of the cell. In

glycolysis, partial oxidation of glucose occurs, which yields two molecules of pyruvic acid.

Krebs Cycle – It is an aerobic process that takes place in the mitochondria of the cell. It gives
Carbon dioxide after complete oxidation of pyruvic acid formed during glycolysis.

Given below in a tabular column are the differences between glycolysis and Krebs Cycle.

Glycolysis vs Krebs Cycle

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Glycolysis Krebs Cycle

1. It is the first step of respiration yielding Krebs Cycle is the second step of aerobic
two molecules of pyruvic acid after the respiration in which pyruvate is oxidised
partial breakdown of a glucose molecule in completely into inorganic substances
a set of enzymatic processes forming carbon dioxide

2. Occurs in all the living organisms Occurs in aerobes

3. Occurs inside the cytoplasm Occurs inside the mitochondria

4. No carbon dioxide evolved Carbon dioxide evolved

5. Oxygen not required for glycolysis Oxygen is required for Krebs Cycle

One ATP or GTP molecule is produced

6. Four ATP molecules are produced in the
by substrate-level phosphorylation in
glycolysis for each glucose molecule
each turn of the Kreb’s cycle

7. Consumes 2 molecules of ATP for initial

Doesn’t consume ATP
phosphorylation of substance molecules

8. Net gain of two molecules of ATP and two Each turn of the Krebs cycle yields three
molecules of NADH gained for every molecules of NADH and two molecules of
molecule of glucose broken down FADH2

9. Occurs as a linear sequence Occurs as a cyclic sequence

Both glycolysis and the Krebs cycle are enzyme-mediated and are under constant regulation
based on the energy requirement of the cell/organism. The rates of these processes vary
under various conditions such as the well-fed state, fasting state, exercised state and
starvation state.

Fermentation

Fermentation is a metabolic process that produces chemical changes in

organic substrates through the action of enzymes. In biochemistry, it is narrowly defined as
the extraction of energy from carbohydrates in the absence of oxygen.

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History of Fermentation

 The term "ferment" comes from the Latin word fervere, which means "to boil."
Fermentation was described by late 14th century alchemists, but not in the modern
sense. The chemical process of fermentation became a subject of scientific
investigation about the year 1600. Fermentation is a natural process. People applied
fermentation to make products such as wine, mead, cheese, and beer long before
the biochemical process was understood.

 In the 1850s and 1860s, Louis Pasteur became the first zymurgist or scientist to
study fermentation when he demonstrated fermentation was caused by living cells.
However, Pasteur was unsuccessful in his attempts to extract the enzyme
responsible for fermentation from yeast cells. In 1897, German chemist Eduard
Buechner ground yeast, extracted fluid from them, and found the liquid could
ferment a sugar solution. Buechner's experiment is considered the beginning of the
science of biochemistry, earning him the 1907 Nobel Prize in chemistry.

Examples of Products Formed by Fermentation

Most people are aware of food and beverages that are fermentation products, but may not
realize many important industrial products results from fermentation.

 Beer
 Wine
 Yogurt
 Cheese
 Certain sour foods containing lactic acid, including sauerkraut, kimchi, and pepperoni
 Bread leavening by yeast
 Sewage treatment
 Some industrial alcohol production, such as for biofuels
 Hydrogen gas

Ethanol Fermentation

Yeast and certain bacteria perform ethanol fermentation where pyruvate (from glucose
metabolism) is broken into ethanol and carbon dioxide. The net chemical equation for the
production of ethanol from glucose is:

C6H12O6 (glucose) → 2 C2H5OH (ethanol) + 2 CO2 (carbon dioxide)

Ethanol fermentation has used the production of beer, wine, and bread. It's worth noting
that fermentation in the presence of high levels of pectin results in the production of small
amounts of methanol, which is toxic when consumed.

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Lactic Acid Fermentation

The pyruvate molecules from glucose metabolism (glycolysis) may be fermented into lactic
acid. Lactic acid fermentation is used to convert lactose into lactic acid in yogurt production.
It also occurs in animal muscles when the tissue requires energy at a faster rate than oxygen
can be supplied. The next equation for lactic acid production from glucose is:

C6H12O6 (glucose) → 2 CH3CHOHCOOH (lactic acid)

The production of lactic acid from lactose and water may be summarized as:

C12H22O11 (lactose) + H2O (water) → 4 CH3CHOHCOOH (lactic acid)

Hydrogen and Methane Gas Production

The process of fermentation may yield hydrogen gas and methane gas.

Methanogenic archaea undergo a disproportionation reaction in which one electron is

transferred from a carbonyl of a carboxylic acid group to a methyl group of acetic acid to
yield methane and carbon dioxide gas.

Many types of fermentation yield hydrogen gas. The product may be used by the organism
to regenerate NAD+ from NADH. Hydrogen gas may be used as a substrate by sulfate
reducers and methanogens. Humans experience hydrogen gas production from intestinal
bacteria, producing flatus.

Fermentation Facts

 Fermentation is an anaerobic process, meaning it does not require oxygen in order

to occur. However, even when oxygen is abundant, yeast cells prefer fermentation
to aerobic respiration, provided a sufficient supply of sugar is available.
 Fermentation occurs in the digestive system of humans and other animals.
 In a rare medical condition called gut fermentation syndrome or auto-brewery
syndrome, fermentation in the human digestive tract leads to intoxication by ethanol
production.1
 Fermentation occurs in human muscle cells. Muscles can expend ATP faster than
oxygen can be supplied. In this situation, ATP is produced by glycolysis, which does
not use oxygen.
 Although fermentation is a common pathway, it is not the only method used by
organisms to obtain energy anaerobically. Some systems use sulfate as the final
electron acceptor in the electron transport chain.

Glycolysis

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Glycolysis is the process by which one molecule of glucose is converted into two molecules of
pyruvate, two hydrogen ions and two molecules of water. Through this process, the ‘high
energy’ intermediate molecules of ATP and NADH are synthesised. Pyruvate molecules then
proceed to the link reaction, where acetyl-coA is produced. Acetyl-coA then proceeds to
the TCA cycle.

In this article, we will look at the steps of glycolysis, its regulation and consider some clinical
conditions related to glycolysis.
Overview
Glycolysis is the metabolism of glucose into two pyruvate molecules, with the net generation
of two molecules of ATP and two molecules of NADH.

It is regulated at the entry to the pathway and at the irreversible steps (1, 3 and 10). This will
be discussed in more detail below.

Glycolysis is an anaerobic reaction, and in low oxygen conditions is the cell’s sole source of
ATP. You can read more about anaerobic respiration here.
Entry Points
Substrates can enter the glycolysis pathway via three different ways, which are referred to as
‘entry points’. These are:

1. Dietary glucose – glucose is directly absorbed into the blood stream from
the gastrointestinal tract and enters the pathway.
2. Glycogenolysis – glucose is released from hepatic stores of glycogen and enters the
pathway.
3. Other monosaccharides – galactose and fructose enter the glycolysis pathway at
various levels via common intermediates.

Glycogen in skeletal muscle cannot be fully broken down into glucose. This means it cannot
leave the cell, therefore can only feed into glycolysis within the individual skeletal muscle cells
it is stored in.

Transport into Cells

In order for circulating glucose to be used by cells, it needs to pass from the extracellular
space (bloodstream) into the intracellular space. Various transporters (GLUT 1-4) transport
glucose into cells. They have different kinetics and methods of regulation depending on the
purpose of glycolysis in that cell.

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GLUT
Key feature Location Reason
transporter

Regulates basal uptake even

Low Km, so highly
GLUT-1 All cells at low glucose levels, ensures a
active
constant supply of energy for survival

Acts as a glucose sensor – increases

Concentration-
Liver, pancreatic uptake in high glucose levels for
GLUT-2 dependent, due to
islets storage. Also regulates insulin release
high Km
from the pancreas

Muscle, adipose, Increases uptake in presence of

GLUT-4 Insulin-dependent
heart insulin (i.e. after meals) for storage

Phases of Glycolysis
Glycolysis can be considered as a two part process. Firstly, energy is consumed to generate
high energy intermediates, which then go on to release their energy during the second phase.

 Energy investment phase – requires two ATP molecules to produce high energy
intermediates.
 Energy pay out phase – The intermediate is metabolised, producing four ATP molecules and
two NADH molecules.

Energy Investment Phase

Reaction 1

Adapted from work by Thomas Shafee (Own work) [CC BY 4.0 (http://creativecommons.org/
licenses/by/4.0)], via Wikimedia Commons

Figure 1 – Reaction 1 of glycolysis

Glucose is phosphorylated by hexokinase to form glucose-6-phosphate (G6P). The negative

charge effectively traps G6P in the cell as it cannot pass through the membrane.

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This reaction consumes a molecule of ATP, so is spontaneous and irreversible. It is regulated

by product inhibition; higher concentrations of G6P inhibit hexokinase and slow the reaction.
In the liver, glucokinase also catalyses this reaction. It has a higher Km than hexokinase, and
therefore works at greater concentrations of serum glucose.

Galactose can enter glycolysis here through its conversion into G6P, via galactose-1-
phosphate and glucose-1-phosphate.

Reaction 2

In reaction two, G6P is converted into fructose-6-phosphate by glucose isomerase.

This provides an entry point for fructose into glycolysis.

Reaction 3

Adapted from work by Thomas Shafee (Own work) [CC BY 4.0 (http://creativecommons.org/
licenses/by/4.0)], via Wikimedia Commons

Fig 2 – Reaction 3 of glycolysis

Fructose-6-phosphate is phosphorylated by phosphofructokinase into fructose-1,6-

bisphosphate. This creates an unstable molecule that will split spontaneously to form two 3
carbon molecule and consumes the second molecule of ATP.

This is a key regulatory step of glycolysis. It is allosterically inhibited by ATP and activated by
AMP. Furthermore, phosphofructokinase is inhibited by glucagon, whilst insulin activates the
enzyme. This ensures that when there is high blood glucose, and therefore high circulating
insulin, the speed of glycolysis increases.

This is also the step of commitment to glycolysis. Once fructose-1,6-bisphosphate has been
formed glycolysis has to occur, as the molecule cannot enter other metabolic pathways.

Reaction 4

By reaction 4, the energy consumption of the ‘investment phase’ is complete and two ATP
molecules have been consumed.

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Here, fructose-1,6-bisphosphate is converted into two triose sugars by fructose-bisphosphate

aldolase. Namely, these triose sugars are glyceraldehyde-3-phosphate (GA3P) and
dihydroxyacetone phosphate (DHAP).

Reaction 5

Here, DHAP is converted into a second molecule of GA3P.

Both molecules of GA3P then enter the second stage of glycolysis, the payout phase.

Energy Payout Phase

In the payout phase, a molecule of NADH and two molecules of ATP are produced per
molecule of GA3P entering the pathway. As our first molecule of glucose has generated two
molecules of GA3P, the total payout from the payout phase is 2 NADH + 4 ATP.

As we used 2 ATP in the investment phase, the net gain from our first molecule of glucose is 2
NADH and 2 ATP.

Reaction 6

In reaction 6, GA3P is converted into 1,3-bisphosphoglycerate (1,3-BPG) by glyceraldehyde

phosphate dehydrogenase.

This yields a molecule of NADH, formed by the reduction of NAD+.

Reaction 7

Here, 1,3-BPG is converted into 3-phosphoglycerate (3PG) by phosphoglycerate kinase.

This generates a molecule of ATP.

Reaction 8

3PG is converted into 2PG by phosphoglycerate mutase.

Reaction 9

2PG is converted into phosphenolpyruvate by enolase.

Reaction 10

Adapted from work by Thomas Shafee (Own work) [CC BY 4.0 (http://creativecommons.org/
licenses/by/4.0)], via Wikimedia Commons

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Fig 3 – Reaction 10 of glycolysis

Phosphenolpyruvate is converted into pyruvate by pyruvate kinase, which yields our second
molecule of ATP. This is irreversible, and is therefore another key regulatory step.

Fates of Pyruvate

Pyruvate is a versatile molecule which feeds into numerous pathways. After glycolysis, it can
be converted to acetyl-CoA, which has numerous metabolic destinations, including the TCA
cycle. It can also be converted into lactate, which enters the Cori cycle in absence of
mitochondria or oxygen.

Other Important Pathway Interactions

DHAP, an intermediate of glycolysis, can be converted to glycerol phosphate in the liver and
adipose tissue. This can feed into biosynthetic pathways, such as triglyceride and
phospholipid biosynthesis, which also recycles NADH. 1,3-BPG can also be converted to 2,3-
BPG in red blood cells to alter the affinity of haemoglobin for O2.

Citric Acid Cycle(KREBS CYCLE)

The citric acid cycle is a key component of the metabolic pathway by which all aerobic
organisms generate energy.

Key Points

 The Krebs cycle is a series of chemical reactions used by all aerobic organisms to
generate energy through the oxidization of acetate—derived from carbohydrates,
fats, and proteins —into carbon dioxide.
 Theoretically there are several alternatives to the TCA cycle, but the TCA cycle
appears to be the most efficient.
 The citric acid cycle consumes acetate (in the form of acetyl-CoA) and water, reduces
NAD+ to NADH, and produces carbon dioxide.

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Key Terms

 acetyl CoA: Acetyl coenzyme A or acetyl-CoA is an important molecule in metabolism,

used in many biochemical reactions. Its main function is to convey the carbon atoms
within the acetyl group to the citric acid cycle (Krebs cycle) to be oxidized for energy
production.
 citric acid cycle: An alternative name for the Krebs cycle.
 glycolysis: The cellular degradation of the simple sugar glucose to yield pyruvic acid
and ATP as an energy source.

The citric acid cycle, shown in —also known as the tricarboxylic acid cycle (TCA cycle) or the
Krebs cycle—is a series of chemical reactions used by all aerobic organisms to generate
energy through the oxidation of acetate—derived from carbohydrates, fats, and proteins—
into carbon dioxide. The cycle provides precursors including certain amino acids as well as
the reducing agent NADH that is used in numerous biochemical reactions. Its central
importance to many biochemical pathways suggests that it was one of the earliest
established components of cellular metabolism; it may have originated abiogenically.

The name of this metabolic pathway is derived from citric acid, a type of tricarboxylic acid
that is first consumed and then regenerated by this sequence of reactions to complete the
cycle. The cycle consumes acetate (in the form of acetyl-CoA) and water, reduces NAD+ to
NADH, and produces carbon dioxide. The NADH generated by the TCA cycle is fed into the
oxidative phosphorylation pathway. The net result of these two closely linked pathways is
the oxidation of nutrients to produce usable energy in the form of ATP.

Components of the TCA cycle were derived from anaerobic bacteria, and the TCA cycle itself
may have evolved more than once. Theoretically there are several alternatives to the TCA
cycle, however the TCA cycle appears to be the most efficient. If several alternatives
independently evolved, they all rapidly converged to the TCA cycle.

The citric acid cycle is a key component of the metabolic pathway by which all aerobic
organisms generate energy. Through the catabolism of sugars, fats, and proteins, a two
carbon organic product acetate in the form of acetyl-CoA is produced. Acetyl-CoA along with
two equivalents of water (H2O) are consumed by the citric acid cycle, producing two
equivalents of carbon dioxide (CO2) and one equivalent of HS-CoA. In addition, one
complete turn of the cycle converts three equivalents of nicotinamide adenine dinucleotide
(NAD+) into three equivalents of reduced NAD+ (NADH), one equivalent of ubiquinone (Q)
into one equivalent of reduced ubiquinone (QH2), and one equivalent each of guanosine
diphosphate (GDP) and inorganic phosphate (Pi) into one equivalent of guanosine
triphosphate (GTP). The NADH and QH2 that is generated by the citric acid cycle is used by
the oxidative phosphorylation pathway to generate energy-rich adenosine triphosphate
(ATP).

One of the primary sources of acetyl-CoA is sugars that are broken down by glycolysis to
produce pyruvate that, in turn, is decarboxylated by the enzyme pyruvate dehydrogenase.
This generates acetyl-CoA according to the following reaction scheme:

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CH3C(=O)C(=O)O– (pyruvate) + HSCoA + NAD+ → CH3C(=O)SCoA (acetyl-CoA) + NADH + H+ +

CO2

The product of this reaction, acetyl-CoA, is the starting point for the citric acid cycle.

The Citric Acid Cycle: The citric acid cycle, or Krebs cycle, is a series of chemical reactions used
by all aerobic organisms to generate energy through the oxidization of acetate—derived from
carbohydrates, fats, and proteins—into carbon dioxide. In addition, the cycle provides
precursors including certain amino acids as well as the reducing agent NADH that is used in
numerous biochemical reactions.

The Kreb cycle in simple terms?

a sequence of reactions in the living organism in which oxidation of acetic acid or acetyl
equivalent provides energy for storage in phosphate bonds (as in ATP) — called also citric
acid cycle, tricarboxylic acid cycle.

The process of Krebs cycle?

The Krebs cycle is a series of chemical reactions used by all aerobic organisms to generate
energy through the oxidization of acetate—derived from carbohydrates, fats, and proteins
—into carbon dioxide.
 Krebs cycle Steps
o Oxidative Decarboxylation of pyruvate to Acetyl CoA
o Step 1: Condensation of acetyl CoA with oxaloacetate

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o Step 2: Isomerization of citrate into isocitrate

o Step 3: Oxidative decarboxylations of isocitrate
o Step 4: Oxidative decarboxylation of α-ketoglutarate
o Step 5: Conversion of succinyl-CoA into succinate
o Step 6: Dehydration of succinate to fumarate
o Step 7: Hydration of fumarate to malate
o Step 8: Dehydrogenation of L-malate to oxaloacetate

the Krebs cycle and what is its purpose?

The tricarboxylic acid (TCA) cycle, also known as the Krebs or citric acid cycle, is the main
source of energy for cells and an important part of aerobic respiration. The cycle harnesses
the available chemical energy of acetyl coenzyme A (acetyl CoA) into the reducing power of
nicotinamide adenine dinucleotide (NADH).

Hexose Monophosphate Pathway

Introduction
The hexose monophosphate shunt, also known as the pentose phosphate pathway, is a
unique pathway used to create products essential in the body for many reasons. The HMP
shunt is an alternative pathway to glycolysis and is used to produce ribose-5-phosphate and
nicotinamide adenine dinucleotide phosphate (NADPH). This pathway occurs in the
oxidative and non-oxidative phases, each comprising a series of reactions. The HMP shunt
also has significance in the medical world, as enzyme or co-factor deficiencies can have
potentially fatal implications on the affected patients.

Function
The HMP shunt is parallel to the glycolysis pathway and takes place in the cytoplasm. A 6-
carbon sugar, glucose, may enter the glycolytic pathway or enter the alternative HMP shunt
depending on the cell’s individual needs at the time. Once the glucose enters the HMP
shunt, it undergoes a series of reactions, broken down into the oxidative(irreversible) and
non-oxidative phases (reversible).
The oxidative phase is responsible for converting the intermediate glucose-6-phosphate to
6-phosphogluconate, using the glucose-6-phosphate dehydrogenase (G6PD) enzyme. The
by-product of this reaction is the important molecule NADPH. 6-phosphogluconate then
converts into ribulose-5-phosphate, and NADPH gets produced again as a by-product.
The non-oxidative phase of the HMP shunt involves the conversion of ribulose-5-phosphate
to ribose-5-phosphate (R-5-P) through a series of independent reactions. It is important to

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note that no NADPH molecules get created in this part of the HMP shunt. R-5-P in this
reaction can be returned to the glycolytic pathway as fructose-6-phosphate. This step
requires the transketolase enzyme with the presence of the thiamine co-factor. Thiamine
also participates in a plethora of other metabolic reactions throughout the body. It is used
by enzyme alpha-ketoglutarate in the Krebs cycle, for the enzyme pyruvate dehydrogenase
as well as branch-chained ketoacid dehydrogenase.[3]
The HMP shunt pathway is under the regulation of the demands of NADPH in the respective
tissue. The rate-limiting enzyme is G6PD and has allosteric inhibition directed by the
presence of NADPH and allosteric activation via the presence of NADP+. Consequently, the
activity of G6PD activity also increases in a fed state with a high carbohydrate diet, and
conversely, decreases in a starving or a diabetic state.[3][4]
Go to:

Mechanism
The importance of the HMP shunt is R-5-P and NADPH molecules that generated in the
reaction. R-5-P undergoes a series of reactions to create the different ribose sugars that
comprise the DNA and RNA molecules, required to carry genetic information. Moreover, R-
5-P also converts into erythrose-4-phosphate for the synthesis of aromatic amino acids. As
such, this pathway is an essential source of these ribose sugars.[5]
NADPH is also an important molecule as it has a plethora of functions throughout the body.
It takes part in anabolism processes such as that of synthesis steroids and fatty acids.
NADPH is also important in the respiratory burst process, an essential component of the
immune response within phagolysosomes. The NADPH is used to reduce oxygen into an
oxygen radical that converts into hydrogen peroxide, which in turn, transforms into bleach.
Bleach in the phagolysosomes is introduced to offending pathogens to cause death. NADPH
is also required to reduce the molecule glutathione. Using glutathione reductase and
NAPDH, oxidized glutathione is converted into its reduced form, and can detoxify free
radicals and peroxides; this limits the possibility of any free radical injury in tissues with
NADPH present, namely that the red blood cells.[5][6]
Since this pathway creates NADPH, a reducing agent utilized in many anabolic processes, the
HMP shunt is common in more tissues than others. These tissues are those with significant
anabolic functions such as adrenal cortex (fatty acid and steroid synthesis), mammary gland
(milk production), liver, and red blood cells.[6]
Go to:

Clinical Significance
Interruptions in the HMP shunt can have drastic effects on an individual. A pathology that
can arise is the deficiency of the enzyme glucose-6-phosphate dehydrogenase, the enzyme
required to produce NADPH in the initial step of the HMP shunt. The consequences are
mostly seen in red blood cells as the cells can no longer defend against the constant
introduction of oxidizing agents. This condition of glucose-6-phosphate dehydrogenase
deficiency is an X-linked recessive disorder, most commonly seen in African Americans. The
deficiency leads to hemolytic anemia in response to any of the offending agents such as fava

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beans, medications like nitrofurantoin, or anti-malarial drugs. Infections are the most likely
offending agents to exacerbate this condition as the inflammation introduces free radicals
into red blood cells that cannot detoxify those radicals, and oxidative damage ensues. The
damage occurs as the injury causes protein and globin chains to denature within the red
blood cells. The globin chains aggregate and form the characteristic Heinz bodies seen in
G6PD deficient patients. Consequently, the spleen phagocytoses the aggregated portion of
the RBC and leaves the remnant bite cells in circulation to create degmacytes, or bite cells,
as seen in a peripheral smear of these patients. Although detrimental to the health, this
condition does confer a degree of resistance to malaria as the parasites require the reduced
form of glutathione to proliferate while these patients only have the oxidized form in the
red blood cells.
Thus, the evolutionary advantage maintains a deficiency in the African American
population.[7][8][9] Another use of the HMP shunt in medicine is its use in diagnosing
patients with thiamine (vitamin B1) deficiency.
Thiamine deficient patients present with a wide array of symptoms, ranging from Wernicke-
Korsakoff syndrome to dry or wet beriberi. Wernicke-Korsakoff is characterized by a triad of
ataxia, ophthalmoplegia, and confabulations. Wet beriberi can result in cardiac failure and
dilated cardiomyopathies, while dry beriberi results in sequelae of neurological symptoms.
As thiamine deficiency can lead to so many complications and potentially become fatal, it is
important to detect the deficiency promptly. The deficiency is tested by administering
thiamine to a suspected patient and detecting the activity of the transketolase enzyme of
HMP shunt within red blood cells.
If transketolase activity increases, it confirms the diagnosis of thiamine deficiency. As such,
thiamine, as well as dextrose, is administered since thiamine deficiency causes impaired
glucose breakdown.[10][11]
Definition HMP: It is an alternative minor pathway for glucose oxidation. Does not produce
ATP nor utilize it. Producing NADPH+H+ (Biosyn Lipids) and Ribose (Synthesis of NA).

Intracellular site and tissue distribution:

Cytosolic in tissues namely: liver, Adipose tissues, Lactating mammary gland, RBCs,
Suprarenal cortex, Thyroid and testis. Not active in non-lactating mammary gland, and in
skeletal muscles. Reactions of the Pentose Phosphate Pathway
The pathway are divided into two phases:
1. Phase I : Oxidative irreversible phase
2. Phase II : Non-oxidative reversible phase.
Reactions of phase I (oxidative irreversible phase): In the first phase, glucose-6-phosphate
undergoes dehydrogenation and decarboxylation to give pentose, ribulose-5-phosphate
with generation of NADPH

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Regulation of HMP shunt: -

The key regulatory enzymes are G-6-PD and 6-phospho-gluconate dehydrogenases.

They are activated by fed state, glucose, insulin, thyroxine and NADP.

But….

They are inhibited during starvation, diabetes mellitus and with high NADPH.H+ /NADP
ratio.

Functions and metabolic importance of HMP shunt:

Production of pentoses: Tissues must satisfy their own requirement of pentoses

since dietary pentoses are not utilizable and ribose is not a significant constituent
of systemic blood.

Pentoses are used for:

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1. Nucleic acids, Ribose for RNA and Deoxyribose for DNA.

2. Coenzymes synthesis, e.g., NAD+, FAD, CoASH
3. Free nucleotide Coenzymes, e.g., ATP, GTP
4. Synthesis of certain vitamins, e.g., B2 and B12

HMP pathway is the major human source for production of NADPH.H+ required
for:

1. Fatty acid synthesis (lipogenesis) and fatty acid desaturation.

2. Cholesterol and other steroid synthesis.

3. Synthesis of sphingosine and cerebrosides.

4. Synthesis of non-essential amino acids, e.g., glutamate and tyrosine from

phenylalanine.

5. Regeneration of reduced glutathione (G-S-H). 6. Metabolic hydroxylation with

cytp450.

Electron transport chain

The electron transport chain ?

The electron transport chain is a series of four protein complexes that couple redox
reactions, creating an electrochemical gradient that leads to the creation of ATP in a
complete system named oxidative phosphorylation. It occurs in mitochondria in both
cellular respiration and photosynthesis.

The three main steps in the electron transport chain are:

 Generation of a proton gradient across the mitochondrial membrane. Proton accumulation
occurs in the intermembrane space of mitochondria.
 Reduction of molecular oxygen and formation of water. ...
 ATP synthesis by chemiosmosis.

The electron transport chain and what is its function?

The electron transport chain is a series of electron transporters embedded in the inner
mitochondrial membrane that shuttles electrons from NADH and FADH2 to molecular

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oxygen. In the process, protons are pumped from the mitochondrial matrix to the
intermembrane space, and oxygen is reduced to form water.
The electron transport chain in simple terms?
The electron transport chain is a cluster of proteins that transfer electrons through a
membrane within mitochondria to form a gradient of protons that drives the creation of
adenosine triphosphate (ATP).

Example of electron transport chain?

Electrons may enter an electron transport chain at the level of a mobile cytochrome or
quinone carrier. For example, electrons from inorganic electron donors (nitrite, ferrous
iron, electron transport chain) enter the electron transport chain at the cytochrome level.

The 5 steps of the electron transport chain?

 Delivery of electrons by NADH and FADH 2start subscript, 2, end subscript. ...
 Electron transfer and proton pumping. ...
 Splitting of oxygen to form water. ...
 Gradient-driven synthesis of ATP.


Why is oxygen important in the electron transport chain?

Oxygen is the final electron acceptor in the electron transport chain, which allows for
oxidative phosphorylation. Without oxygen, the electrons will be backed up, eventually
causing the electron transport chain to halt.

Is electron transport aerobic or anaerobic?

Both aerobic and anaerobic respiration use electron-transport chain-reaction pathways in
energy production. However, aerobic respiration produces much more energy, or ATP
molecules, than anaerobic respiration.

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The major events of the electron transport chain?

The events of the electron transport chain involve NADH and FADH, which act as electron
transporters as they flow through the inner membrane space. In complex I, electrons are
passed from NADH to the electron transport chain, where they flow through the remaining
complexes. NADH is oxidized to NAD in this process.

How many complexes does the electron transport chain have?

There are four protein complexes (labeled complex I-IV) in the electron transport chain,
which are involved in moving electrons from NADH and FADH2 to molecular oxygen.
Complex I establishes the hydrogen ion gradient by pumping four hydrogen ions across the
membrane from the matrix into the intermembrane space.

The complex of electron transport chain?

The ETC proteins in a general order are complex I, complex II, coenzyme Q, complex III,
cytochrome C, and complex IV. Coenzyme Q, also known as ubiquinone (CoQ), is made up
of quinone and a hydrophobic tail. Its purpose is to function as an electron carrier and
transfer electrons to complex III.

Complex 1 made up of?

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Complex I is composed of flavin mononucleotide (FMN) and an enzyme containing iron-

sulfur (Fe-S). FMN, which is derived from vitamin B2 (also called riboflavin), is one of several
prosthetic groups or co-factors in the electron transport chain

The name of complex 1 in electron transport chain?

NADH oxidoreductase
Complex I of the electron transport chain, also known as NADH oxidoreductase or NADH
dehydrogenase, is a very large, L-shaped structure that functions to accept high energy
electrons from NADH molecules.

Complex 2 do in the electron transport chain?

Complex II of the electron transport chain, also known as succinate reductase, is involved in
the citric acid cycle. It contains the enzyme called succinate dehydrogenase that was used
by the citric acid cycle to transform succinate into fumarate and in the process form FADH2.

Complex 3 in the electron transport chain?

Complex III of the electron transport chain, also known as Q-cytochrome c oxidoreductase
or simply cytochrome reductase, is a multi-subunit structure that functions to accept
electrons from ubiquinol and transfer them onto another electron carrier called
cytochrome c.

Complex 4 in the electron transport chain?

Complex IV of the electron transport chain, also known as cytochrome c oxidase, is a

multiunit structure that functions to transfer electrons form cytochrome c to
oxygen and in the process form water and help generate a proton gradient.

Initiation of Electron Transport Chain:

Once the NADH has been made from a metabolite in the citric acid cycle inside of the
mitochondria, it interacts with the first complex 1 enzyme, known as NADH reductase. This
complex 1 contains a coenzyme flavin mononucleotide (FMN) which is similar to FAD.

Why are electron acceptors important?

The electron acceptor contributes to overcome the potential losses existing on the
cathode, thus it is one of the major factors influencing power generation in MFCs.

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The electron transport chain in the mitochondrion is the site of oxidative

phosphorylation in eukaryotes. The NADH and succinate generated in the citric acid
cycle are oxidized, which releases the energy of oxygen to power ATP synthase.

Photosynthetic electron transport chain of the thylakoid membrane.

An electron transport chain (ETC[1]) is a series of protein complexes and other molecules
that transfer electrons from electron donors to electron acceptors via redox reactions
(both reduction and oxidation occurring simultaneously) and couples this electron transfer
with the transfer of protons (H+ ions) across a membrane. Many of the enzymes in the
electron transport chain are membrane-bound.
The flow of electrons through the electron transport chain is an exergonic process. The
energy from the redox reactions creates an electrochemical proton gradient that drives the
synthesis of adenosine triphosphate (ATP). In aerobic respiration, the flow of electrons
terminates with molecular oxygen as the final electron acceptor that provides most of the

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energy.[2] In anaerobic respiration, other, lower-energy electron acceptors are used, such
as sulfate.
In an electron transport chain, the redox reactions are driven by the difference in the Gibbs
free energy of reactants and products. The free energy released when a higher-energy
electron donor and acceptor convert to lower-energy products, while electrons are
transferred from a lower to a higher redox potential, is used by the complexes in the
electron transport chain to create an electrochemical gradient of ions. It is this
electrochemical gradient that drives the synthesis of ATP via coupling with oxidative
phosphorylation with ATP synthase.[3]
In eukaryotic organisms the electron transport chain, and site of oxidative phosphorylation,
is found on the inner mitochondrial membrane. The energy of oxygen released in its
reaction with reduced compounds such as cytochrome c and (indirectly) NADH and FADH is
used by the electron transport chain to pump protons into the intermembrane space,
generating the electrochemical gradient over the inner mitochondrial membrane. In
photosynthetic eukaryotes, the electron transport chain is found on the thylakoid
membrane. Here, light energy drives electron transport through a proton pump and the
resulting proton gradient causes subsequent synthesis of ATP. In bacteria, the electron
transport chain can vary between species but it always constitutes a set of redox reactions
that are coupled to the synthesis of ATP through the generation of an electrochemical
gradient and oxidative phosphorylation through ATP synthase.[4]

Contents

 1Mitochondrial electron transport chains

o 1.1Mitochondrial redox carriers
 1.1.1Complex I
 1.1.2Complex II
 1.1.3Complex III
 1.1.4Complex IV
o 1.2Coupling with oxidative phosphorylation
o 1.3Reverse electron flow
 2Bacterial electron transport chains
o 2.1Electron donors
o 2.2Complexes I and II
o 2.3Quinone carriers
o 2.4Proton pumps
o 2.5Cytochrome electron carriers
o 2.6Terminal oxidases and reductases
o 2.7Electron acceptors
 3Photosynthetic

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Mitochondrial electron transport chains[edit]

Most eukaryotic cells have mitochondria, which produce ATP from reactions of oxygen with
products of the citric acid cycle, fatty acid metabolism, and amino acid metabolism. At
the inner mitochondrial membrane, electrons from NADH and FADH2 pass through the
electron transport chain to oxygen, which provides the energy driving the process as it is
reduced to water.[5] The electron transport chain comprises an enzymatic series of electron
donors and acceptors. Each electron donor will pass electrons to an acceptor of higher
redox potential, which in turn donates these electrons to another acceptor, a process that
continues down the series until electrons are passed to oxygen, the most energy-rich[2] and
terminal electron acceptor in the chain.
Each reaction releases energy because a higher-energy donor and acceptor convert to
lower-energy products. Via the transferred electrons, this energy is used to generate
a proton gradient across the mitochondrial membrane by "pumping" protons into the
intermembrane space, producing a state of higher free energy that has the potential to do
work.
This entire process is called oxidative phosphorylation since ADP is phosphorylated to ATP
by using the electrochemical gradient that the redox reactions of the electron transport
chain have established driven by the energy of oxygen.
Mitochondrial redox carriers[edit]
Energy associated with the transfer of electrons down the electron transport chain is used
to pump protons from the mitochondrial matrix into the intermembrane space, creating an
electrochemical proton gradient (ΔpH) across the inner mitochondrial membrane.
This proton gradient is largely but not exclusively responsible for the
mitochondrial membrane potential (ΔΨM).[6] It allows ATP synthase to use the flow of
H+ through the enzyme back into the matrix to generate ATP from adenosine
diphosphate (ADP) and inorganic phosphate. Complex I (NADH coenzyme Q reductase;
labeled I) accepts electrons from the Krebs cycle electron carrier nicotinamide adenine
dinucleotide (NADH), and passes them to coenzyme Q (ubiquinone; labeled Q), which also
receives electrons from Complex II (succinate dehydrogenase; labeled II).
Q passes electrons to Complex III (cytochrome bc1 complex; labeled III), which passes them
to cytochrome c (cyt c). Cyt c passes electrons to Complex IV (cytochrome c oxidase; labeled
IV), which uses the electrons and hydrogen ions to release the energy of molecular oxygen
as it is reduced to water.
Four membrane-bound complexes have been identified in mitochondria. Each is an
extremely complex transmembrane structure that is embedded in the inner membrane.
Three of them are proton pumps.

The structures are electrically connected by lipid-soluble electron carriers and water-soluble
electron carriers. The overall electron transport chain can be summarized as follows:

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NADH+H+ → Complex I → Q

↑
Complex II
↑
Succinate → Complex III → cytochrome c → Complex IV → H2O
↑
Complex II
↑
Succinate

Reverse electron flow

Reverse electron flow is the transfer of electrons through the electron transport chain
through the reverse redox reactions. Usually requiring a significant amount of energy to
be used, this can reduce the oxidized forms of electron donors. For example, NAD + can
be reduced to NADH by Complex I.[16]

There are several factors that have been shown to induce reverse electron flow.
However, more work needs to be done to confirm this. One example is blockage of ATP
synthase, resulting in a build-up of protons and therefore a higher proton-motive force,
inducing reverse electron flow.[17]

Bacterial electron transport chains

In eukaryotes, NADH is the most important electron donor. The associated electron
transport chain is NADH → Complex I → Q → Complex III → cytochrome c → Complex
IV → O2 where Complexes I, III and IV are proton pumps, while Q and cytochrome c are
mobile electron carriers.

The electron acceptor providing the energy for this process is molecular oxygen.
In prokaryotes (bacteria and archaea) the situation is more complicated, because there
are several different electron donors and several different electron acceptors. The
generalized electron transport chain in bacteria is:

Donor Donor Donor

↓ ↓ ↓
dehydrogenase → quinone → bc1 → cytochrome
↓ ↓
oxidase(reductase) oxidase(reductase)
↓ ↓

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Acceptor Acceptor

Electrons can enter the chain at three levels: at the level of a dehydrogenase, at the
level of the quinone pool, or at the level of a mobile cytochrome electron carrier.
These levels correspond to successively more positive redox potentials, or to
successively decreased potential differences relative to the terminal electron acceptor.
In other words, they correspond to successively smaller Gibbs free energy changes for
the overall redox reaction.
Individual bacteria use multiple electron transport chains, often simultaneously.
Bacteria can use a number of different electron donors, a number of different
dehydrogenases, a number of different oxidases and reductases, and a number of
different electron acceptors.
For example, E. coli (when growing aerobically using glucose and oxygen as an energy
source) uses two different NADH dehydrogenases and two different quinol oxidases, for
a total of four different electron transport chains operating simultaneously.
A common feature of all electron transport chains is the presence of a proton pump to
create an electrochemical gradient over a membrane. Bacterial electron transport
chains may contain as many as three proton pumps, like mitochondria, or they may
contain two or at least one.

Electron donors
In the current biosphere, the most common electron donors are organic molecules.
Organisms that use organic molecules as an electron source are called organotrophs.
Chemoorganotrophs (animals, fungi, protists) and photolithotrophs (plants and algae)
constitute the vast majority of all familiar life forms.
Some prokaryotes can use inorganic matter as an electron source. Such an organism is
called a (chemo)lithotroph ("rock-eater"). Inorganic electron donors include hydrogen,
carbon monoxide, ammonia, nitrite, sulfur, sulfide, manganese oxide, and ferrous iron.

Lithotrophs have been found growing in rock formations thousands of meters below the
surface of Earth. Because of their volume of distribution, lithotrophs may actually
outnumber organotrophs and phototrophs in our biosphere.
The use of inorganic electron donors such as hydrogen as an energy source is of
particular interest in the study of evolution. This type of metabolism must logically have
preceded the use of organic molecules and oxygen as an energy source.

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Electron acceptors[edit]
Just as there are a number of different electron donors (organic matter in organotrophs,
inorganic matter in lithotrophs), there are a number of different electron acceptors,
both organic and inorganic. If oxygen is available, it is invariably used as the terminal
electron acceptor in aerobic bacteria and facultative anaerobes, because it generates
the greatest Gibbs free energy change and produces the most energy.[2]
In anaerobic environments, different, lower-energy electron acceptors are used,
including nitrate, nitrite, ferric iron, sulfate, carbon dioxide, and small organic molecules
such as fumarate.

Photosynthetic[edit]
Further information: Light-dependent reaction and Photosynthetic reaction center
In oxidative phosphorylation, electrons are transferred from a moderate-energy
electron donor such as NADH to a high-energy acceptor such as O2 through an electron
transport chain, releasing the energy. In photophosphorylation, the energy of sunlight is
used to create a high-energy electron donor which can subsequently reduce oxidized
components and couple to ATP synthesis via proton translocation by the electron
transport chain.[10]
Photosynthetic electron transport chains, like the mitochondrial chain, can be
considered as a special case of the bacterial systems. They use mobile, lipid-soluble
quinone carriers (phylloquinone and plastoquinone) and mobile, water-soluble carriers
(cytochromes). They also contain a proton pump. The proton pump in all photosynthetic
chains resembles mitochondrial Complex III. The commonly-held theory
of symbiogenesis proposes that both organelles descended from bacteria.
Oxidative phosphorylation

The process of oxidative phosphorylation uses the energy of the proton gradient established
by the electron transport system as a means of phosphorylating ADP to make ATP. The
establishment of the proton gradient is dependent upon electron transport. If electron
transport stops or if the inner mitochondrial membrane’s impermeability to protons is
compromised, oxidative phosphorylation will not occur because without the proton
gradient to drive the ATP synthase, there will be no synthesis of ATP.

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The main purpose of the oxidative phosphorylation process?

Oxidative phosphorylation is a highly efficient method of producing large amounts of ATP,
the basic unit of energy for metabolic processes. During this process electrons are
exchanged between molecules, which creates a chemical gradient that allows for the
production of ATP.

The major steps of oxidative phosphorylation in mitochondria include:

1. Delivery of Electrons by NADH and FADH2. Reduced NADH and FADH2 transfer their
electrons to molecules near the beginning of the transport chain. ...
2. Electron Transport and Proton Pumping. ...
3. Splitting of Oxygen to form Water. ...
4. ATP Synthesis.
Why it is called oxidative phosphorylation?
In the mitochondrion, what the proton gradient does is facilitate the production of ATP
from ADP and Pi. This process is known as oxidative phosphorylation, because the
phosphorylation of ADP to ATP is dependent on the oxidative reactions occurring in
the mitochondria.

The types of oxidative phosphorylation?

3.1 NADH-coenzyme Q oxidoreductase (complex I)
 3.2 Succinate-Q oxidoreductase (complex II)
 3.3 Electron transfer flavoprotein-Q oxidoreductase.
 3.4 Q-cytochrome c oxidoreductase (complex III)
 3.5 Cytochrome c oxidase (complex IV)
 3.6 Alternative reductases and oxidases.
 3.7 Organization of complexes.

The output of oxidative phosphorylation?

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Oxidative phosphorylation is the process by which ATP is synthesised when electrons are
transported from the energy precursors produced in the citric acid cycle through various
enzyme complexes to molecular oxygen. The input is NADH, FADH2, O2 and ADP. The output
is NAD+, FAD+, H2O and ATP.

Another name for oxidative phosphorylation?

Oxidative phosphorylation, also known as electron transport-linked phosphorylation, refers

to the metabolic pathway in which the energy released by nutrients during oxidation is
utilized to generate ATP through electrical transport chain.

Oxidative phosphorylation regulated?

Oxidative phosphorylation is regulated by the energy needs of cells, primarily the levels of
ADP compared to ATP, following Le Chatelier's Principle of chemical equilibria.

Happens when oxidative phosphorylation is inhibited?

Substrate-level phosphorylation is a supplementary pathway for ATP synthesis, which
directly generates ATP and NADH without the electron transport chain. Therefore, blocking
or restraining oxidative phosphorylation can effectively decrease ATP concentrations in the
cell.

ATP synthase

The protein complex harvesting energy from the proton gradient and using it to make ATP
from ADP is an enzyme that has several names - Complex V, PTAS (Proton Translocating ATP
Synthase), and ATP synthase (Figure 5.29). Central to its function is the movement of
protons through it (from the intermembrane space back into the matrix). Protons will only
provide energy to make ATP if their concentration is greater in the intermembrane space
than in the matrix and if ADP is available.

It is possible, in some cases, for the concentration of protons to be greater inside the matrix
than outside of it. When this happens, the ATP synthase can run backwards, with protons
moving from inside to out, accompanied by conversion of ATP to ADP + Pi. This is usually not
a desirable circumstance and there are some controls to reduce its occurrence.

Normally, ATP concentration will be higher inside of the mitochondrion and ADP
concentration be higher outside the mitochondrion. However, when the rate of ATP
synthesis exceeds the rate of ATP usage, then ATP concentrations rise outside the
mitochondrion and ADP concentrations fall everywhere.

This may happen, for example, during periods of rest. It has the overall effect of reducing
transport and thus lowering the concentration of ADP inside the matrix. Reducing ADP
concentration in the matrix reduces oxidative phosphorylation and has effects on
respiratory control (see HERE).

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Another important consideration is that when ATP is made in oxidative phosphorylation, it is

released into the mitochondrial matrix, but must be transported into the cytosol to meet
the energy needs of the rest of the cell.

This is accomplished by action of the adenine nucleotide translocase, an antiport that moves
ATP out of the matrix in exchange for ADP moving into the matrix. This transport system is
driven by the concentrations of ADP and ATP and ensures that levels of ADP are maintained
within the mitochondrion, permitting continued ATP synthesis.

One last requirement for synthesis of ATP from ADP is that phosphate must also be
imported into the matrix. This is accomplished by action of the phosphate translocase,
which is a symport that moves phosphate into the mitochondrial matrix along with a proton.

There is evidence that the two translocases and ATP synthase may exist in a complex, which
has been dubbed the ATP synthasome.

In summary, the electron transport system charges the battery for oxidative
phosphorylation by pumping protons out of the mitochondrion. The intact inner membrane
of the mitochondrion keeps the protons out, except for those that re-enter through ATP
Synthase. The ATP Synthase allows protons to re-enter the mitochondrial matrix and
harvests their energy to make ATP.

ATP synthase mechanism

The ATP Synthase itself is an amazing nanomachine that makes ATP using a gradient of
protons flowing through it from the intermembrane space back into the matrix. It is not easy
to depict in a single image what the synthase does. Figure 5.31 and Figure 5.32 illustrate the
multi-subunit nature of this membrane protein, which acts like a turbine at a hydroelectric
dam. The movement of protons through the ATP Synthase c-ring causes it and the γ-ε stalk
attached to it to turn. It is this action that is necessary for making ATP.

In ATP Synthase, the spinning components, or rotor, are the membrane portion (c ring) of
the F0 base and the γ-ε stalk, which is connected to it. The γ-ε stalk projects into the F1 head

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of the mushroom structure. The F1 head contains the catalytic ability to make ATP. The F1
head is hexameric in structure with paired α and β proteins arranged in a trimer of dimers.
ATP synthesis occurs within the β subunits.

Enzymes
Enzymes are proteins that help speed up chemical reactions in our bodies. Enzymes are
essential for digestion, liver function and much more. Too much or too little of a certain
enzyme can cause health problems. Enzymes in our blood can also help healthcare providers
check for injuries and diseases.

What are enzymes?

Enzymes are proteins that help speed up metabolism, or the chemical reactions in our
bodies. They build some substances and break others down. All living things have enzymes.

Our bodies naturally produce enzymes. But enzymes are also in manufactured products and
food.

What do enzymes do?

One of the most important roles of enzymes is to aid in digestion. Digestion is the process of
turning the food we eat into energy. For example, there are enzymes in our saliva, pancreas,
intestines and stomach. They break down fats, proteins and carbohydrates. Enzymes use
these nutrients for growth and cell repair.

Enzymes also help with:

 Breathing.
 Building muscle.
 Nerve function.
 Ridding our bodies of toxins.

What are the different types of enzymes?

There are thousands of individual enzymes in the body. Each type of enzyme only has one
job. For example, the enzyme sucrase breaks down a sugar called sucrose. Lactase breaks
down lactose, a kind of sugar found in milk products.

Some of the most common digestive enzymes are:

 Carbohydrase breaks down carbohydrates into sugars.

 Lipase breaks down fats into fatty acids.
 Protease breaks down protein into amino acids.

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PARTS OF ENZYMES

What are the parts of an enzyme?

Each enzyme has an “active site.” This area has a unique shape. The substance an enzyme
works on is a substrate. The substrate also has a unique shape. The enzyme and the
substrate must fit together to work.

How do temperature and pH affect enzymes?

Enzymes need the right conditions to work. If conditions aren’t right, enzymes can change
shape. Then, they no longer fit with substrates, so they don’t work correctly.

Each enzyme has an ideal temperature and pH:

 pH: Enzymes are sensitive to acidity and alkalinity. They don’t work properly if an
environment is too acidic or basic. For example, an enzyme in the stomach called
pepsin breaks down proteins. If your stomach doesn’t have enough acid, pepsin
can’t function optimally.
 Temperature: Enzymes work best when your body temperature is normal, about
98.6°F (37°C). As temperature increases, enzyme reactions increase. But if the
temperature gets too high, the enzyme stops working. That’s why a high fever can
disrupt bodily functions.

COMMON CONDITIONS & DISORDERS

What health conditions can enzyme problems cause?

Metabolic disorders are often the result of not having enough of a certain enzyme. Parents
can pass them to their children through genes (inherited). Some examples of inherited
metabolic disorders include:

 Fabry disease prevents body from making enzymes (alpha-galactosidase A) that

break down fat (lipids).
 Krabbe disease (globoid cell leukodystrophy) affects enzymes needed for the
protective covering (myelin) on nerve cells (Central Nervous System).
 Maple syrup urine disease affects enzymes needed to break down certain branch
chain amino acids.

Other health conditions related to enzyme imbalances include:

 Crohn’s disease an imbalance of the bacteria in your gut (gut microbiome) may
influence an autoimmune response of the intestinal tract. This may play a role in
presentation and severity of Crohn’s disease.

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 Exocrine pancreatic insufficiency (EPI) is a condition where your pancreas doesn’t

have enough digestive enzymes. You can’t break down food or absorb nutrients.
Chronic pancreatitis, pancreatic cancer, diabetes or cystic fibrosis can lead to EPI.
 Lactose intolerance is a shortage of the enzyme needed to digest sugars in milk
(lactose) and dairy.

How are enzyme tests used to diagnose health conditions?

Your healthcare provider can use a variety of enzyme and protein blood tests to check for
certain health conditions. For example, elevated liver enzymes could be a sign of liver
disease.

CARING FOR YOUR ENZYMES

Do I need to take enzyme supplements?

People without chronic health conditions can usually get the enzymes they need from a
healthy diet. But, if you have certain health conditions, your healthcare provider may
recommend taking enzyme supplements. For instance, many people with EPI may take a
digestive enzyme before they eat. This helps their bodies absorb nutrients from food. Talk
to your healthcare provider before taking any type of enzyme supplement.

Can medications affect enzyme levels?

Some medications affect enzyme levels. For example, antibiotics can kill certain bacteria
needed for some enzymes to work their best. This is the reason antibiotics may cause
diarrhea. To kill the bacteria making you sick, they also wipe out important good bacteria
that aid in digestion.

Statins (medications that lower cholesterol) can raise liver enzymes and muscle enzymes.
They may increase the risk of damage to the liver or muscles.

When should I contact my doctor about an enzyme problem?

You won’t know if you have an enzyme problem without a blood test. Contact your doctor if
you experience any of the following problems:

 Abdominal pain.
 Bloating or gas.
 Diarrhea.
 Fatigue.
 Nausea and vomiting.
 Unexplained weight loss.
 Low red blood counts (anemia).
 Gastrointestinal bleeding.

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Bioenergetics

is the branch of biochemistry that focuses on how cells transform energy, often by
producing, storing or consuming adenosine triphosphate (ATP). Bioenergetic processes,
such as cellular respiration or photosynthesis, are essential to most aspects of cellular
metabolism, therefore to life itself

Example of bioenergetics?
Glycogenesis, gluconeogenesis, and citric acid cycle are examples of bioenergetic
processes.

Bioenergetics in human body?

What is Human Bioenergetics? Human Bioenergetics is the multidisciplinary study of how
energy is transferred in cells, tissues, and organisms. The manner in which the body
regulates energy transfer pathways and processes has a fundamental influence on health.

The principles of bioenergetics?

There are two laws of bioenergetics.
1) Energy cannot be created or destroyed, but can be changed from one form to another.
2) Energy transfer will always proceed in the direction of increased entropy, and the release
of “free energy”.

The characteristics of bioenergetics?

The first fundamental characteristic of bioenergetics consists in the fact that organisms are
open systems, which function only under conditions of constant exchange of materials
and energy with the surrounding medium. The thermodynamics of such a system is
essentially different from classical thermodynamics.

Bioenergetics useful to humans?

Through bioenergetics studies biochemists are able to understand how energy released
during cellular metabolism can be used to “do work” that is useful for the cells e.g.
movement of molecules, cells or organs or heating within a body.

Bioenergetics used for?

Bioenergetics is the flow of energy a biological system, and describes the conversion of
macronutrients (carbs, protein, fat, which all contain chemical energy) into micronutrients
(glucose, ATP) that can be used for energy.

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Bioenergetic testing?
Core Specific Bioenergetic Testing analyzes hundreds of foods against various parts of your
body. We also provide a list of triggers and your body's intolerances.

Bioenergetic dysfunction?
The factor or factors that initiate inflammation in AD remain elusive. One factor that may
substantially contribute to chronic inflammation is bioenergetic dysfunction, which may
arise at the level of the mitochondria.

Biomeridian test?
A: Biomeridian testing is a noninvasive method used to assess the energy meridians
(channels) of the body, and it can tell you: The functional status of your meridians ant their
related organs, systems, or functions, by determining if they are stressed, balanced, or
weakened, and by how much.

Metabolic disruption
A metabolic disorder occurs when abnormal chemical reactions in your body disrupt this
process. When this happens, you might have too much of some substances or too little of
other ones that you need to stay healthy.

BIODEGRADATION OF ORGANIC POLLUTANTS

Introduction

Biodegradation is defined as the biologically catalyzed reduction in complexity of chemical

compounds [1]. Indeed, biodegradation is the process by which organic substances are
broken down into smaller compounds by living microbial organisms [2]. When
biodegradation is complete, the process is called "mineralization". However, in most cases
the term biodegradation is generally used to describe almost any biologically mediated
change in a substrate

The microbial organisms transform the substance through metabolic or enzymatic

processes. It is based on two processes: growth and cometabolism. In growth, an organic
pollutant is used as sole source of carbon and energy. This process results in a complete
degradation (mineralization) of organic pollutants

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Bioremediation of polluants utilizing biodegradation abilities of microorganisms include the

natural attenuation, although it may be enhanced by engineered techniques, either by
addition of selected microorganisms (bioaugmentation) or by biostimulation, where
nutrients are added. Genetic engineering is also used to improve the biodegradation
capabilities of microorganisms by GEM. Nevertheless, there are many factors affecting the
efficiency of this process and risks associated to the use of GEM in the field.

Role of microorganisms in biodegradation of pollutants

In this chapter, biodegradation is described associated with environmental bioremediation.

Therefore, biodegradation is nature's way of recycling wastes, or breaking down organic
matter into nutrients that can be used and reused by other organisms. In the
microbiological sense, "biodegradation" means that the decaying of all organic materials is
carried out by a huge assortment of life forms comprising mainly bacteria, yeast and fungi,
and possibly other organisms.

Bioremediation and biotransformation methods endeavour to harness the astonishing,

naturally occurring, microbial catabolic diversity to degrade, transform or accumulate a
huge range of compounds including hydrocarbons (e.g. oil), polychlorinated biphenyls
(PCBs), polyaromatic hydrocarbons (PAHs), radionuclides and metals [12].

2.1. Some biodegradable pollutants

 HYDROCARBONS
 POLYCYCLIC AROMATIC HYDROCARBONS (PAHs)
 Polychlorinated biphenyls (PCBs)
 Pesticides:
 Dyes:
 Radionuclides:

Factors affecting microbial degradation

Microorganisms can degrade numerous of organic pollutants owing to their metabolic

machinery and to their capacity to adapt to inhospitable environments. Thus,

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microorganisms are major players in site remediation. However, their efficiency depends on
many factors, including the chemical nature and the concentration of pollutants, their
availability to microorganisms, and the physicochemical characteristics of the environment
[111]. So, factors that influence the rate of pollutants degradation by microorganisms are
either related to the microorganisms and their nutritional requirements (biological factors)
or associated to the compounds in which electrons are transferred to various more oxidized
compounds referred

The process of biodegradation?

In biodegradation processes, a chemical compound is transformed or eliminated by the
biological action of living organisms. In general terms, biodegradability is the tendency of a
lubricant to be ingested and metabolized by microorganisms.

Biodegradable organic pollutants?

Biodegradable pollutants: These pollutants are natural organic substances which can be
decomposed or consumed by natural microbial or biological processes and converted into
CO2, water, or simple organic molecules.

Some examples of organic pollutants?

Organic Pollutant
 Pesticide.
 Graphene.
 Toxicity.
 Trichloroethylene.
 Polychlorinated Biphenyl.
 Polycyclic Aromatic Hydrocarbon.
 Dioxin.
 Bacterium
 TWO types of biodegradation?
The breakdown of materials by microorganisms when oxygen is present is aerobic digestion,
and the breakdown of materials when oxygen is not present is anaerobic digestion.

The study of biodegradation of pollutants?

Biodegradation is defined as the action of microbes on any complex organic compounds
that are found in any environmental matrix.

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The factors affecting biodegradation?

Biodegradation rates are known to be affected by such factors as the availability of
inorganic nutrients, the presence of multiple substrates, the redox environment, substrate
concentration, temperature, water activity, and the adaptive response of the
microorganisms (7-9)

The role of microbes in biodegradation?

Microorganisms use their metabolic pathways for the biodegradation of organic pollutants
into inorganic compounds, carbon dioxide, and water after their partial or complete
mineralization. Organisms use HCs as their energy source.

Important is biodegradation to the environment?

What Makes Biodegradation Important? Biodegradation is nature's waste management and
recycling system. It breaks down everything from yard waste to crude oil. It is a natural
process necessary to keep our planet clean and healthy.

Biodegradation is helpful in environment cleanup?

Bacteria and archaebacteria species have bioremediation potential which could help break
them to less harmful substances. Recent studies have shown the potential of
microorganisms to decrease the environmental impact of landfills which are suffocating the
land near metropolises.

Organic pollutants: Types and properties

Organic contaminants are carbon-containing chemical substances that have a demonstrably

harmful impact on one or more environmental elements.Three wide groups can include
organic pollutants: (i) hydrocarbons, (ii) oxygen, phosphorous and nitrogen compounds and
(iii) organometallic compounds. Hydrocarbons and associated substances that include such
substances as Dichloro-diphenyl trichloroethane (DDT), dioxins, and polycyclic aromatic
hydrocarbons (PAHs) are the primary group of agricultural pollutes (Goodhead& Tyler 2009).
These substances include carbon and hydrogen materials, including those that produce
chlorine and oxygen. The number of chemical bonds that are mainly C-H, C-C, C-Cl, C = C and
C = C (aromatic) are small. Both these bonds are reasonably stable and have minimal
polarity and so the associated compounds are given this domain.

DEGRADATION OFORGANIC POLLUTANTS:

In the world, organic compounds are exposed to many physical, biochemical, and biological
processes that interconnect in environmental mechanisms to decide the ultimate fate of the
substance. A great deal of acid is used when neutralized by chemical methods, which is
neither economical nor healthy and poses significant health risks.(Sarnaik and Kanekar
1995). There are several mechanisms for organic pollutant degradation, of which, some are
described below:

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Physical processes-

Physical methods have been employed for decades of organic degradation, and could
involve different processes such as photocatalytic degradation by utilizing the Ag-modified
Zn2GeO4 nanorods, the nano-composite graphene oxide hydrogels, the organic Silica, etc.
Decomposition by catalytic / photocatalytic oxidation of these organic compounds is known
to be the greenest approach for plastic waste management. Visible light reaction
semiconductors have drawn attention as effective European Journal of Molecular & Clinical
Medicine ISSN 2515-8260 Volume 07, Issue 10 , 2020 3556 photo catalysts from several
researchers (Sun et al 2010). Many catalysts are used to degrade organic compounds photo
catalytically.

TiO2 has been used as a photo catalyst owing to its low expense, chemical stability and
non-toxicity. TiO2is favored since is a good photo-oxidation catalyst with a powerful photo-
induced photo-oxidation ability. Many researchers paired TiO2 with small strip-band semi-
conductors, which increased the isolation of images by creating heterogeneous joints.
Researchers changed the Ag PO surface using sol gel phase TiO2 (Yao et al 2012). Ag3PO4
was deposited on TiO2 for heterostructure. Photoelectrodes utilizing a parallel chemical
deposition and UV-reducing process of Ag /Ag3PO4 / TiO2 hetero-structure.

The TiO2nanoparticles as a photocatalyst are one successful tool for degrading organic
compounds because of their non-toxicity, low expense, physical and chemical stability and
high reactivity (Wang et al 2009).

Chemical Processes-

Bioremediation approaches require electrical chlorinated benzene dehalogenation, such

that chlorine is separated step by step from the heavily chlorinated benzene such order to
create less chlorinated benzene and eventually converted to benzene. Chlorobenzenes and
the cathodic route of reaction for hexachlorobenzenes were analyzed as follows:
hexachlorobenzene, pentachlorobenzenes, 1,2,3,5 tetrachlorobenzene, 1,2,4-
trichlorobenzene1, 4-dichlorobenzene andmonochlorobenzene. There are two catalytic
degradation of organic modules by MnO2 nanostructure, and numerous community
mineralization’s have been recorded for different organic compounds / dyes, for example,
Rhodamine, MB, Benzyl alcohol at high temperatures with strong oxidants

Biological processes-

The bioremediation of polluted soil allows cost-competitive care for multiple locations
facing expensive incineration or prolonged liabilities for land degradation at present.
Technology was found to be cost-effective in the field under the circumstances of complete
site remediation. Bio stimulation, bio-stimulation and bioaugmentation involve various
forms of biological processes.

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1. Bio-attenuation (Natural Attenuation): The toxins are converted into less noxious types
or immobilized types of bio-attenuation. These mechanisms of transformation and
immobilization are primarily attributable to biodegradation by microorganisms and, to some
degree, reactions with natural chemicals and geologic medial sorption(Smet and Pritchard
2003).
2. Bio-stimulation: Bio-stimulation is a method of decontamination of degraded soils in
which microbial development by altering the atmosphere is encouraged. The concentrations
of microbial transformation of chemical contaminants are heavily influenced by the
provision and supply of nutrients, such as carbon, nitrogen, potassium, oxygen required,
appropriate pH, redox capacity, as well as organic pollutant sort and concentration.
Nutrients in the form of manure, sluggish releases and oleophilic are applied to promote
microbial degradation (Nikolopoulou and kalogerakis 2008).
3. Bio-augmentation: - Bio-enhancement is the incorporation of bacterial crops to improve
the pace of contaminant depletion. The polluted soil sediments produce very well-equipped
microflora for large levels of organic contaminants. The remediation of European Journal of
Molecular & Clinical Medicine ISSN 2515-8260 Volume 07, Issue 10 , 2020 3557 soils freshly
contaminated by hydrocarbons may involve microorganisms extracted from infected soil
sediments. The 2% bio-remediated soil priming has been found to promote the
biodegradation of soil PAH components treated with fuel oil
PROCESS/ MECHANISM OF BIODEGRADATION OF ORGANIC POLLUTANTS
Enzymes are present in the soil in large, three dimensional assemblies of mineral and
organic particles that inhibit and influence their mobility

CONCLUSION:

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The best way to treat contaminated organic pollutant sites via bioremediation is because it
is not only sustainable but also eco-friendly. In addition, there is no collateral habitat loss
quality. The microorganism in our environment offers greater potential to transform toxic
compounds into less toxic by-products. Microbial enzymes play a major and vital role in the
biodegradation of organically contaminated soils such as diesel, petroleum or PAHs etc. In
future, these enzymes will be opened upwhich are likely to open a new era of microbiology
to support different technologies for cleaning up the environment.

UNIT 4 – MICROBIOLOGY OF WASTEWATER TREATMENT SYSTEM

Biological treatment process:

Biological treatments rely on bacteria, nematodes, or other small organisms to break
down organic wastes using normal cellular processes. Wastewater typically contains a
buffet of organic matter, such as garbage, wastes, and partially digested foods.

Three main categories, biological wastewater treatment can be:

1. aerobic, when microorganisms require oxygen to break down organic matter to

carbon dioxide and microbial biomass

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2. anaerobic, when microorganisms do not require oxygen to break down organic

matter, often forming methane, carbon dioxide, and excess biomass
3. anoxic, when microorganisms use other molecules than oxygen for growth, such as
for the removal of sulfate, nitrate, nitrite, selenate, and selenite

Aerobic wastewater treatment technologies

Activated sludge was first developed in the early 1900s in England and has become the
conventional biological treatment process widely used in municipal applications but can also
be used in other industrial applications. Wastewaters from the primary treatment phase
enter an aeration tank where it is aerated in the presence of suspended (freely floating)
aerobic microorganisms. The organic material is broken down and consumed, forming
biological solids which flocculate into larger clumps, or flocs. The suspended flocs enter a
settling tank and are removed from the wastewater by sedimentation. Recycling of settled
solids to the aeration tank controls levels of suspended solids, while excess solids are
wasted as sludge. Activated sludge treatment systems typically have larger space
requirements and generate large amounts of sludge, with associated disposal costs, but
capital and maintenance costs are relatively low, compared to other options.
Fixed-bed bioreactors, or FBBRs, developed as forced-air industrial treatment systems in
the 1970s and 80s, consist of multiple-chambered tanks in which the chambers are packed
tight with porous ceramic, porous foam, and/or plastic media; the wastewater passes
through the immobilized bed of media. Of all biological treatment systems, FBBRs can hold
the most contaminant-eating microbes in the smallest volume, which makes FBBRs space-
saving and energy-efficient technologies ideal for treating wastewaters from medium to
medium-high BOD feed levels down to very low effluent levels. The media is engineered to
have a high enough surface area to encourage a robust biofilm formation with long solids
lifespan, resulting in low sludge formation and lowest sludge disposal costs. A well-
engineered fixed-bed will allow wastewater to flow through the system without channeling
or plugging. Chambers can be aerobic and still have anoxic zones to achieve aerobic
carbonaceous removal and full anoxic denitrification at the same time. More advanced
biological processes can be facilitated with these systems (for example, nitrification,
denitrification, deselenation, sulfide-reduction, and anammox), by having unique bacterial
populations colonize the biofilm media in separate tank chambers, which can be uniquely
configured to treat your facility’s specific wastewater constituents.
Moving bed bioreactors, or MBBRs, invented in the late 1980s in Norway, already has been
applied in over 800 applications in more than 50 countries, with approximately half treating
domestic wastewater and half treating industrial wastewater. MBBRs typically consist of
aeration tanks filled with small moving polyethylene biofilm carriers held within the vessel
by media retention sieves.

Today the plastic biofilm carriers come from many vendors in many sizes and shapes, are
typically half- to one-inch diameter cylinders or cubes and are designed to be suspended
with their immobilized biofilm throughout the bioreactor by aeration or mechanical mixing.
Because of the suspended moving bio-film carriers, MBBRs allow high BOD wastewaters to
be treated in a smaller area with no plugging. MBBRs are typically followed by a secondary

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clarifier, but no sludge is recycled to the process; excess sludge settles, and a slurry removed
by vacuum truck, or settled solids are filter pressed and disposed as a solid waste.
MBBRs are often used to remove the bulk of BOD load upstream of other biological
treatment processes or used in situations where effluent quality is less important; they are
not used for polishing BOD to low effluent levels. They are used for treating wastewaters
produced in food and beverage facilities, meat processing and packing plants, petrochemical
facilities, and refineries.
Membrane bioreactors, or MBRs, came into common use in the 1990s once membrane
modules were submerged directly in the aeration tank, and air scour was implemented to
keep the membranes from fouling. MBRs are advanced biological wastewater treatment
technologies that combine conventional suspended-growth activated sludge with
membrane filtration, rather than sedimentation, to separate and recycle the suspended
solids. As a result, MBRs operate with much higher mixed-liquor suspended solids (MLSS)
and longer solids residence times (SRTs), producing a significantly smaller footprint with a
much higher quality effluent compared to conventional activated sludge.
MBRs primarily target BOD and total suspended solids (TSS). MBR system design varies
depending on the nature of the wastewater and the treatment goals, but a typical MBR
might consist of aerobic (or anaerobic) treatment tanks, an aeration system, mixers, a
membrane tank, a clean-in-place system, and either a hollow fiber or flat sheet
ultrafiltration membrane. As a result of its many parts and cleaning processes, MBRs are
known for high capital, high operating, and high maintenance costs.
Biological trickling filters are used to remove organic contaminants from both air and
wastewater. They work by passing air or water through a media designed to collect a biofilm
on its surfaces. The biofilm may be composed of both aerobic and anaerobic bacteria which
breakdown organic contaminants in water or air. Some of the media used for these systems
include gravel, sand, foam, and ceramic materials. The most popular application of this
technology is municipal wastewater treatment and air remediation to remove H2S at
municipal sewer plants, but they can be used in many situations where odor control is
important.

Anaerobic Wastewater Treatment Technologies

Upflow anaerobic sludge blankets, or UASBs, useanaerobic bacteria to, as mentioned in the
intro of this article,breakdown organics without the use of oxygen, resulting with a
combustible methane-bearing biogas, treated effluent, and anaerobic sludge. With UASB
systems, the general idea is that wastewaters are pumped into the base of the system,
where the organics in the wastewater flow through a blanket of sludge before entering the
upper gas-liquid-solids (GLS) separator, where collection hoods capture the biogas while
allowing the suspended solids to settle and return to the lower reaction zone, while the
cleaned effluent overflows out of the top of the system. The biogas (methane and carbon
dioxide) is either flared or used to generate steam or electricity for use in other processes at
the facility.
The UASB process creates less sludge than aerobic biosystems and therefore needs to be
cleaned out and emptied less than other biological treatment systems, but they require
skilled operators to maintain optimal hydraulic and anaerobic conditions for UASBs to

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operate properly. Expanded granular sludge beds, or EGSBs, are a similar process, but EGSBs
use a stronger upward force to encourage more wastewater-to-sludge contact.
Anaerobic digesters also useanaerobic bacteria to break down organic waste without
oxygen and produce biogas, mostly for sewage treatment, and there are a variety of
anaerobic digesters available. They each perform the same process in slightly different
ways. Examples include covered lagoons, fixed film, suspended and submerged media, and
continuous stirred tank reactors.

ALPHA OXIDATION

What is Alpha Oxidation?

 Alpha – oxidation is defined as the oxidation of fatty acid (methyl group at beta carbon)
with the removal of one carbon unit adjacent to the α-carbon from the carboxylic end.
 The carbon unit is removed in the form of CO2.
 Alpha oxidation occurs in those fatty acids that have a methyl group (-CH3) at the beta-
carbon, which blocks beta oxidation.
 There is no production of ATP.

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Location

Peroxisomes are the cellular sites for α-oxidation.

 Substrate: Phytanic acid, which is present in milk or derived from phytol present in
chlorophyll and animal fat.
 Phytanic acid is a 20-carbon, branched-chain fatty acid.

The Pathway

 Branched chain fatty acids are oxidized at the α-carbon (mainly in brain and other
nervous tissue), and the carboxyl carbon is released as CO2. Branches can interfere with
the normal β-oxidation pathway, most often at the acyl-CoA dehydrogenase step.

 The fatty acid is thus degraded by one carbon initially, and then two carbons at a time.
Both acetyl-CoA and propionyl-CoA are products if the branches are methyl groups.

Steps of alpha oxidation

1. Activation of phytanic acid

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2. Hydroxylation
3. Removal of formyl CoA (CO2)
4. Oxidation of Pristanal
5. Beta-oxidation of pristanic acid

Reactions Involved

1. Phytanic acid is first attached to CoA to form phytanoyl-CoA.

2. Phytanoyl-CoA is oxidized by phytanoyl-CoA dioxygenase, in a process using Fe2+and O2,
to yield 2-hydroxyphytanoyl-CoA.
3. 2-hydroxyphytanoyl-CoA is cleaved by 2-hydroxyphytanoyl-CoA lyase in a TPP-
dependent reaction to form pristanal and formyl-CoA (in turn later broken down
into formate and eventually CO2).
4. Pristanal is oxidized by aldehyde dehydrogenase to form pristanic acid (which can then
undergo beta-oxidation).
5. Propionyl-CoA is released as a result of beta oxidation when the beta carbon is
substituted.

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Significance of alpha oxidation

 α- oxidation is important in the catabolism of branched-chain fatty acids.

 Oxidation of methylated fatty acid.
 Production of cerebronic acid which synthesizes cerebroside and sulfatides.
 Production of odd chain fatty acids.
 The reaction is also a route for the synthesis of hydroxy fatty acids. The α-hydroxy fatty
acid can be further oxidized and decarboxylated to a fatty acid one carbon shorter than
the original. Thus, if an odd-chain-length compound is used initially, an even-chain-
length acid is produced that can be further oxidized by β-oxidation.

Associated Diseases

 Refsum’s disease is the only defect in peroxisomal α-oxidation currently known.

 The Refsum’s disease is an autosomal recessive disorder and is the deficiency of
phytanoyl-CoA hydroxylase.
 The clinical characteristics of Refsum’s disease include peripheral neuropathy and
ataxia, retinitis pigmentosa, and abnormalities of skin and bones.

Beta-oxidation
What do you mean by beta-oxidation?
Inside mitochondria beta oxidation of fatty acids takes place in which two carbon atoms are
removed in the form of acetyl-CoA from acyl-CoA at the carboxyl terminal. The bond is
broken between the second carbon/beta carbon and the third carbon/gamma carbon,
hence the name beta oxidation.

What is Beta Oxidation?

Beta oxidation is a catabolic process in which fatty acid molecules are broken down inside the
cytosol of prokaryotes and in the mitochondria in eukaryotes generating acetyl CoA, NADH,
and FADH2. This acetyl CoA will enter the citric acid cycle. The NADH and FADH2 produced
here act as co-enzymes that are useful in the electron transport chain.

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Why does beta-oxidation occur?

Fatty acid β-oxidation is the process by which fatty acids are broken down to produce
energy. Fatty acids primarily enter a cell via fatty acid protein transporters on the cell
surface. Once inside, FACS adds a CoA group to the fatty acid. CPT1 then converts the long-
chain acyl-CoA to long-chain acylcarnitine.

What are the products of beta-oxidation?

The products of beta-oxidation are:
 acetyl CoA.
 FADH2, NADH and H. +

Beta Oxidation Steps

Beta oxidation takes place in four steps: dehydrogenation, hydration, oxidation and

thyolisis. Each step is catalyzed by a distinct enzyme.

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Dehydrogenation

In the first step, acyl-CoA is oxidized by the enzyme acyl CoA dehydrogenase. A double bond
is formed between the second and third carbons (C2 and C3) of the acyl-CoA chain entering
the beta oxidation cycle; the end product of this reaction is trans-Δ2-enoyl-CoA (trans-delta
2-enoyl CoA). This step uses FAD and produces FADH2, which will enter the citric acid cycle
and form ATP to be used as energy. (Notice in the following figure that the carbon count
starts on the right side: the rightmost carbon below the oxygen atom is C1, then C2 on the
left forming a double bond with C3, and so on.)

Hydration

In the second step, the double bond between C2 and C3 of trans-Δ2-enoyl-CoA is hydrated,
forming the end product L-β-hydroxyacyl CoA, which has a hydroxyl group (OH) in C2, in
place of the double bond. This reaction is catalyzed by another enzyme: enoyl CoA
hydratase. This step requires water.

Oxidation

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In the third step, the hydroxyl group in C2 of L-β-hydroxyacyl CoA is oxidized by NAD+ in a
reaction that is catalyzed by 3-hydroxyacyl-CoA dehydrogenase. The end products are β-
ketoacyl CoA and NADH + H. NADH will enter the citric acid cycle and produce ATP that will
be used as energy.

Thiolysis

Finally, in the fourth step, β-ketoacyl CoA is cleaved by a thiol group (SH) of another
CoA molecule (CoA-SH). The enzyme that catalyzes this reaction is β-ketothiolase. The
cleavage takes place between C2 and C3; therefore, the end products are an acetyl-CoA
molecule with the original two first carbons (C1 and C2), and an acyl-CoA chain two carbons
shorter than the original acyl-CoA chain that entered the beta oxidation cycle.

End of Beta Oxidation

In the case of even-numbered acyl-CoA chains, beta oxidation ends after a four-carbon acyl-
CoA chain is broken down into two acetyl-CoA units, each one containing two carbon atoms.
Acetyl-CoA molecules enter the citric acid cycle to yield ATP.

In the case of odd-numbered acyl-CoA chains, beta oxidation ensues in the same way except
for the last step: instead of a four-carbon acyl-CoA chain being broken down into two acetyl-
CoA units, a five-carbon acyl-CoA chain is broken down into a three-carbon propionyl-CoA
and a two-carbon acetyl-CoA. Another chemical reaction then converts propionyl-CoA to

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succinyl-CoA (see the figure below), which enters the citric acid cycle to produce ATP.

Energy Yield and End Products

Each beta oxidation cycle yields 1 FADH2, 1 NADH and 1 acetyl-CoA, which in terms of
energy is equivalent to 17 ATP molecules:

 1 FADH2 (x 2 ATP) = 2 ATP

 1 NADH (x 3 ATP) = 3 ATP
 1 acetyl-CoA (x 12 ATP) = 12 ATP
 Total = 2 + 3 + 12 = 17 ATP
However, the theoretical ATP yield is higher than the real ATP yield. In reality, the
equivalent of about 12 to 16 ATPs is produced in each beta oxidation cycle.

Besides energy yield, the fatty acyl-CoaA chain becomes two carbons shorter with each
cycle. In addition, beta oxidation yields great amounts of water; this is beneficial for
eukaryotic organisms such as camels given their limited access to drinkable water.

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NITRIFICATIION & DENITRIFICATION

Nitrification Definition

Nitrification is a biological process of oxidation of ammonia into nitrite which is then

followed by the oxidation of nitrite into nitrate.
 Nitrification is a part of the nitrogen cycle where living beings oxidize ammonia present
in the soil into useful forms of nitrogen that can then be consumed by various
organisms.

 In the nitrogen cycle, nitrogen is converted into many forms successively passing from
atmosphere to soil to organisms and back to the atmosphere.
 Nitrification is an aerobic process that occurs in soil by various aerobic microorganisms
like bacteria and some archaea.

 The bacteria oxidizing ammonia are termed ammonia-oxidizing bacteria (AOB), and the
archaea oxidizing archaea are termed ammonia-oxidizing archaea (AOA).

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 The reactions involved in nitrification are the following:

2NH4+ + 3O2 → 2NO2– + 4H+ + 2H2O
–
2NO2 + O2 → 2NO3–

 Because this process consists of two reaction steps, two different groups of
microorganisms are involved in nitrification.

 The first step of oxidation of ammonia is brought about by microbes in the soil which
includes bacteria of the genus Nitrosomonas and Nitrosococcus, and arceae like
Nitrosopumilus maritimus and Nitrososphaera viennensis. The conversion of ammonia
into nitrite is the rate-limiting step of nitrification.

 The second reaction is performed by bacteria of the genus Nitrobacter and Nitrospira.

 All of these microorganisms are chemoautotrophs that utilize the energy from the
reaction to produce organic carbon compounds.
 Nitrification is important in many organisms as it is the only process of obtaining
nitrogen source for some microorganisms present in the soil.

 These organisms convert ammonia into nitrates which is more soluble than ammonia
and thus can be taken into the system more conveniently.

 Besides, it is also important in agricultural systems where ammonia is used as a

fertilizer. The ammonia is then converted into nitrate which facilitates nitrogen leaching
into the plants.
 Nitrifying bacteria also play an important role in wastewater treatment where
different nitrogen compounds are converted into nitrates and then nitrogen before
removing the gas out of the water.
 The nitrification process is controlled by a number of factors like the availability of
oxygen, soil moisture, and the availability of ammonia.
 The activity of the nitrifying bacteria also decreases in acidic conditions and at a
temperature above 35°C.

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Denitrification Definition

Denitrification is a biological process of reduction of nitrate into nitrite, which is then

followed by the reduction of nitrate into nitrogen gas that usually results in the removal of
nitrogen gas into the air.
 Denitrification, like nitrification, is a microbial process that is performed by various
groups of microorganisms.
 It is also an important step in the nitrogen cycle where nitrogen is released back into
the atmosphere from the ground.
 In this case, the oxidized products of nitrogen are reduced to its gaseous forms, mainly
nitrous oxide (N2O) and nitrogen gas (N2).
 Denitrification, unlike nitrification, is performed by facultative anaerobes that perform
denitrification as anaerobic respiration to reduce oxidized forms into gases.
 Denitrification takes place at about 10% or less concentration of oxygen and organic
carbon compounds.


 The process of denitrification takes place through a set of half-reactions, which are:
NO3 + 2H+ + 2e– → NO2– + H2O
–

NO2− + 2 H+ + e− → NO + H2O
2NO + 2 H+ + 2 e− → N2O + H2O
N2O + 2 H+ + 2 e− → N2 + H2O
 The overall reaction can be represented as:
2NO3− + 10e− + 12H+ → N2 + 6H2O

 The process is primarily performed by heterotrophic bacteria like Paracoccus

denitrificans and some species of Pseudomonas, but some autotrophic denitrifiers
like Thiobacillus denitrificans are also present.

 Denitrification is an important microbiological process that is performed naturally in

both terrestrial and marine environments.
 Besides, denitrification follows nitrification in wastewater treatments to convert
nitrogen-rich compounds into nitrogen gas before being released into the atmosphere.

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 However, sometimes denitrification can be disadvantageous by removing the

NO3– present in the soil, thus reducing the extent of leaching.
 Denitrification is controlled by various factors like the concentration of oxygen and
carbons, even though some aerobic bacteria of the genus Proteobacteria, might
facilitate denitrification even in the presence of oxygen.

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Eutrophication

What Is Eutrophication?
Eutrophication is the process in which a water body becomes overly enriched with nutrients,
leading to plentiful growth of simple plant life. The excessive growth (or bloom) of algae and
plankton in a water body are indicators of this process. Eutrophication is considered to be a
serious environmental concern since it often results in the deterioration of water quality and
the depletion of dissolved oxygen in water bodies. Eutrophic waters can eventually become
“dead zones” that are incapable of supporting life.

Table of Content

 Definition of Eutrophication
 Causes of Eutrophication
 Classification of Eutrophication
 Effects of Eutrophication
 FAQs on Eutrophication

Definition of Eutrophication
Eutrophication may be defined as the inorganic nutrient enrichment of natural waters,
leading to an increased production of algae and macrophytes.
Many lakes are naturally eutrophic and in some cases there is a progressive eutrophication
as the lake matures. The term Eutrophication is more widely known in relation to human
activities where the artificial introduction of plant nutrients has led to community changes
and a deterioration of water quality in many freshwater systems. This aspect has become
increasingly important with increases in human population and more extensive
development of agriculture and eutrophication now ranks with other major anthropogenic
effects such as deforestation, global warming depletion of the ozone layer and large scale
environmental disturbance in relation to its potentially harmful effect on natural
ecosystems.

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An image detailing the excessive growth of algae in a eutrophic water body is provided below.
Aquatic ecosystems are home to several plant and animal life forms – both simple and
complex. The process of eutrophication destroys the balance in these ecosystems by
favouring the growth of simple plant life. This greatly decreases the biodiversity of the
ecosystem by killing off several desirable species.

Causes of Eutrophication
The availability of nutrients such as nitrogen and phosphorus limits the growth of plant life in
an ecosystem. When water bodies are overly enriched with these nutrients, the growth of
algae, plankton, and other simple plant life is favoured over the growth of more complex plant
life.

How do Water Bodies Become Overly Enriched?

Phosphorus is considered one of the primary limiting factors for the growth of plant life in
freshwater ecosystems. Several sources also claim that the availability of nitrogen is an
important limiting factor for the growth of algae.
Phosphates tend to stick to the soil and are transported along with it. Therefore, soil erosion
is a major contributor to the phosphorus enrichment of water bodies. Some other
phosphorus-rich sources that enrich water bodies with the nutrient include:

 Fertilizers
 Untreated sewage
 Detergents containing phosphorus
 Industrial discharge of waste.
Among these sources, the primary contributors to eutrophication include agriculture and
industrial wastes.

What Happens to the Huge Biomass of Algae in Eutrophic Waters?

The excessive growth of algae in eutrophic waters is accompanied by the generation of a large
biomass of dead algae. These dead algae sink to the bottom of the water body where they
are broken down by bacteria, which consume oxygen in the process.

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Process of Eutrophication

The overconsumption of oxygen leads to hypoxic conditions (conditions in which the

availability of oxygen is low) in the water. The hypoxic conditions at the lower levels of the
water body lead to the suffocation and eventual death of larger life forms such as fish.

Classification of Eutrophication
The process of eutrophication can be categorized into two types based on its root cause. Both
these types are explained in this subsection.

Anthropogenic Eutrophication
Anthropogenic eutrophication is caused by human activity – Agricultural farms, golf courses,
lawns, etc. are supplied with nutrients by humans in the form of fertilizers. These fertilizers
are washed away by rains and eventually find their way into water bodies such as lakes and
rivers.
When introduced to an aqueous ecosystem, the fertilizers supply plentiful nutrients to algae
and plankton, resulting in the eutrophication of the water body.
Overpopulation places a huge demand on industrial and agricultural expansion, which in turn
leads to deforestation. When this occurs, the soil erodes more easily, resulting in increased
soil deposits in water bodies.

If the soil is rich in phosphorus, it can lead to eutrophication and severely damage the
ecosystem in and around the water body.

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When sewage pipes and industrial wastes are directed to water bodies, the nutrients present
in the sewage and other wastes increase the rate at which eutrophication occurs.

Natural Eutrophication
Natural eutrophication refers to the excessive enrichment of water bodies via natural events.
For example, the nutrients from the land can be washed away in a flood and deposited into a
lake or a river. These water bodies become overly enriched with nutrients, enabling the
excessive growth of algae and other simple plant life.
The process of natural eutrophication is much slower when compared to the process of
anthropogenic eutrophication. This process is also somewhat dependant on the temperature
of the environment. It may even be complemented by the temperature changes brought on
by global warming.

Effects of Eutrophication
Primarily, the adverse effects of eutrophication on aquatic bodies include a decrease in
biodiversity, increase in toxicity of the water body, and change in species dominance. Some
other important effects of this process are listed below.

 Phytoplanktons grow much faster in such situations. These phytoplankton species are
toxic and are inedible.
 Gelatinous zooplankton blooms fast in these waters.
 Increased biomass of epiphytic and benthic algae can be observed in eutrophic waters.
 Significant changes arise in the species composition of macrophytes and the biomass.
 The water loses its transparency and develops a bad smell and colour. The treatment
of this water becomes difficult.
 Depletion of dissolved oxygen in the water body.
 Frequent fish kill incidents occur and many desirable fish species are removed from
the water body.
 The populations of shellfish and harvestable fish are lowered.

The aesthetic value of the water body diminishes significantly.

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NUTRIENT REMOVAL – BOD, NITROGEN, PHOSPHATE

Why is nutrient removal important?

• Excess nutrients are harmful to the environment

• They can lead to eutrophication in rivers, lakes and estuaries

• Oxygen dead zones

• Fish kills

• Harmful Algal Blooms (HABs)

• Water Resource Recovery Facilities (WRRFs) are required to remove nutrients from
wastewater

How are nutrients removed?

Biological Nutrient Removal (BNR)

• Definition: The removal of nitrogen and phosphorus by the use of, proliferation and
selection of certain microbial populations (“bugs”)

• Different wastewater processes create the proper environment to select and enhance the
growth of the desired bacteria

• Example: Aerobic, anoxic, and anaerobic environments

• These wastewater processes are arranged in many different configurations to achieve the
desired treatment

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Phosphorus Removal

Ortho-Phosphate

• Technologies: Wet chemistry analyzer

• Usually requires a filter

• Total Phosphorus (TP) not usually necessary for control

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Nitrogen Removal

Dissolved Oxygen

• Electrochemical

 Faster response

 Larger range

• Optical  Simple operation

Ammonia-Nitrogen

• ISE (probe)

 Large measuring range

 Fast response time

• Wet chemistry analyzer (cabinet)

 Low concentration measurement*

Nitrate-Nitrogen

• ISE (probe)

• Wet chemistry analyzer

• Optical (probe)

 Simple operation

How are nutrients removed from wastewater?

Biological nutrient removal (BNR) removes total nitrogen (TN) and total phosphorus (TP)
from wastewater through the use of microorganisms under different environmental
conditions in the treatment process

Important to remove nitrogen and phosphorus from wastewater?

The removal of nitrogen and phosphorus from wastewater has become an emerging
worldwide concern because these compounds cause eutrophication in natural water.
Moreover, nitrate is a risk to human health, especially as a possible cause of infant
methaemoglobinaemia

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Sewage microbiology

• Sewage is 99.9% water and 0.02-0.04% solids and sewage sludge is formed at the
wastewater sewage treatment plants by being evacuated through the sewage system.
It contains mineral, organic, and biological impurities and microorganisms in soluble,
insoluble and colloidal forms.

Sludge microbiology:
Activated sludge (AS) is composed of aerobic and anaerobic microorganisms such as
bacteria, archaea, fungi, and protists. It is capable of degrading organic compounds,
including petroleum products, toluene, and benzopyrene

Indicators of water quality

What organisms are most commonly used as indicators of water quality?

Historically, the most commonly used indicator organisms are called “coliform” bacteria-
particularly Escherichia coli (better known as E. coli), from which the larger group gets its
name.

What are indicator organisms provide suitable examples?

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The indicators generally employed worldwide are total coliforms, fecal coliforms, and/or
Escherichia coli. Total coliforms constitute a group of closely related bacteria in the family
Enterobacteriaceae that are usually not pathogenic and are widespread in ambient water.

What is indicator organism of water pollution?

The most common indicators are total coliforms, fecal coliforms, E. coli, and enterococci.
The presence of bacteria commonly found in human feces, termed coliform bacteria (e.g. E.
coli), in surface water is a common indicator of faecal contamination.

Coliforms
What is coliforms in water?
Coliform bacteria are organisms that are present in the environment and in the feces of all
warm-blooded animals and humans. Coliform bacteria will not likely cause illness. However,
their presence in drinking water indicates that disease-causing organisms (pathogens)
could be in the water system.

How do you treat coliform in water?

How can I treat drinking water that contains coliforms? When coliform bacteria are found in
a public water supply, the supplier is required to notify users within 24 hours, and will
immediately begin disinfecting the water supply by feeding chlorine into the water system.

What causes high coliform count in water?

The primary sources of fecal coliform bacteria to freshwater are wastewater treatment
plant discharges, failing septic systems, and animal waste. Bacteria levels do not
necessarily decrease as a watershed develops from rural to urban. Instead, urbanization
usually generates new sources of bacteria.

What are coliform bacteria, and how can they enter drinking water?

Coliform bacteria are microscopic organisms that originate in the intestinal tract of
warmblooded animals and are also present in soil and vegetation. Total coliform bacteria
are generally harmless; however, their presence in drinking water indicates the possibility
that diseasecausing bacteria, viruses or parasites (pathogens) are also present in the water.
Bacterial pollution can result from runoff from woodlands, pastures and feedlots, septic
tanks and sewage plants, and animals and wildfowl. Most coliform bacteria enter natural
streams by direct deposition of waste in the water and runoff from areas with high
concentrations of animals or humans. Domesticated animals contribute heavily to the
bacterial population.

Because most intestinal pathogens cannot survive outside of a host body, they are not
common in ground water (unless the ground water is close to a source of contamination).
Most wells obtain their water at a depth at which pathogens are no longer present;

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however, an improperly constructed or damaged well, or one that is too shallow, may
become contaminated.

What are health concerns from coliforms in drinking water?

Most strains of coliform bacteria are harmless and live in the intestines of healthy humans
and animals. Coliforms indicate that water may be contaminated with sewage or similar
wastes. Diseases which may be present in water that tests positive for coliform bacteria
include typhoid fever, cholera, hepatitis, dysentery, diarrhea, giardiasis, and hemolytic
uremic syndrome.

How can I treat drinking water that contains coliforms?

When coliform bacteria are found in a public water supply, the supplier is required to notify
users within 24 hours, and will immediately begin disinfecting the water supply by feeding
chlorine into the water system. If fecal coliform bacteria or a specific type of bacteria (E.
coli) have been found, the water supplier must issue a “Boil Order” notice.

Decontaminating and re-testing the water supply will likely take several days, and during
that period you should boil all water for several minutes before using it for drinking,
cooking, brushing teeth, making ice, or ingesting it in any way. Always follow the
instructions provided by the water supplier until the “Boil Order” is rescinded.

Private well owners are responsible for disinfection of their water supply. The first thing to
do is to determine if the contamination is in the well or in the water system inside the
home. Test the water at the well itself (or as close as possible). If it contains coliforms then
the well is contaminated. Otherwise, the source of contamination is somewhere in the
plumbing system itself and not the well.

If the water at the well returns an uncontaminated test result, but the water at the tap is
contaminated, the contamination source may be an inadequately maintained treatment
system. Many point-of-use systems, such as activated charcoal filters, at the faucet or in a
refrigerator water system, can harbor bacteria. These filters should be maintained and
replaced according to the manufacturer’s directions. Other possible sources include cross
connection with irrigation water or back siphonage from garden hoses.

If tests show that your well water is contaminated with bacteria, you must determine how
the bacteria are entering the well and correct that problem. Consider the possible sources
of the contamination, which might include a faulty septic system, storm runoff, or livestock
waste runoff. If practical, correct any of these problems before installing permanent water
treatment equipment.

A contaminated water well can be treated by “shock chlorination,” in which a strong

chlorine solution is introduced into the well and allowed to circulate through the water
system. See the shaded box at the end of this fact sheet. “Shock chlorination” may also

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result in removal of films of biological material in the plumbing system that can harbor
coliform bacteria.

A well can become contaminated in a number of ways:

o A shallow or dug well can be contaminated by surface water.

o An improperly sealed well casing or liner can allow bacteria from upper soil
layers to enter the well. An old well casing may also rust through.

o The well casing may become cracked during an earthquake, from subsidence
or from impact damage from farm machinery or snowplows.

o If there is no watertight seal on the well casing, surface water may enter the
well during a flood or storm runoff.

o Bacteria can enter a well when it is drilled, or when a pump is installed or

serviced. These bacteria may come from contaminated drilling equipment, or
from pipe or cable laid out on the ground prior to installation.

o Lack of a backflow device on a pump can allow contaminated water to be

siphoned back into a well.

If contamination persists, the next option is to install a continuous water treatment system
to provide disinfection. Continuous water treatment methods include:

Chlorination. Chlorine is fed into the water after it has left the pump and before it enters
the holding tank.

Ultraviolet Radiation (UV). Light is used to kill microorganisms such as bacteria (viruses,
cysts and worms may be unaffected).

Ozonation. Ozone is more effective against bacteria and viruses than chlorine, but requires
an on-site ozone generator. Ozonation is generally more expensive than other water
treatment methods.
Will a water filter remove coliform?
Biological contaminants such as coliform bacteria are most effectively eliminated through
chlorine disinfection, filtration, ultraviolet irradiation, and ozonation

TOTAL COLIFORMS

Total coliforms include bacteria that are found in the soil, in water that has been
influenced by surface water, and in human or animal waste. Fecal coliforms are the group

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of the total coliforms that are considered to be present specifically in the gut and feces of
warm-blooded animals.

What are acceptable levels of total coliform?

Regarding the primary drinking water standards or health based standards, the
concentration of total coliform bacteria and E. coli should be Absent, Negative, or < 1
colony-forming unit per 100 mls ( cfu/100 ml).

What is a high total coliform count?

Coliforms count is a hygienic indicator and high level of coliform counts generally
indicates unsanitary condition or poor hygiene practices during or after food production.

Is total coliform harmful?

Total coliforms are a group of related bacteria that are (with few exceptions) not harmful to
humans. A variety of bacteria, parasites, and viruses, known as pathogens, can potentially
cause health problems if humans ingest them. EPA considers total coliforms a useful
indicator of other pathogens for drinking water.

TOTAL COLLIFORMS BACTERIA IN DRINKING WATER:

Total coliform bacteria is all around us. They are in the soil and vegetation throughout our
environment and are generally harmless. Total coliform bacteria in drinking water typically
doesn’t have a health risk associated with it and if water testing only detects it, the source is
probably environmental and not fecal contamination.

Total coliform bacteria is often considered an indicator there may be something more
serious contaminating a drinking water system, specifically E. coli bacteria. Total coliform
bacteria are colorless, odorless, and tasteless and the only way it can be detected in drinking
water is through submitting a sample for laboratory testing.

Bacterial contamination can result from a number of sources. These sources include surface
runoff containing animal waste from feedlots, dog runs, or other locations where animal
waste is deposited or piled. Human waste can also be a bacterial contamination source,
most often from a failing onsite wastewater system such as residential septic tanks, laterals,
mounds systems, or lagoons. Additional contamination sources include insects, rodents, or
animals that may get trapped in a well and die, thus introducing bacteria to the well water.
Flooding events where wellheads are submerged by floodwaters that commonly contain
high levels of bacteria are yet another source of contamination.

As said earlier, not all bacteria present a health risk, but some do. E. coli is a subgroup of the
coliform bacteria group. Most E. coli bacteria are harmless and exist in the intestines of
people and warm-blooded animals. However, some strains can cause illness. If E. coli
bacteria is detected in a water sample, it is usually an indicator of recent fecal

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contamination of the water. That means there is a greater risk that water-borne pathogens
are present.

Proper well location, construction, and maintenance are key in reducing well vulnerability to
bacterial contamination. It is recommended that private wells be tested for total coliform
bacteria after the well is initially drilled and annually thereafter. Testing should also happen
if a well has been unused for one or more years, after repairs are made to any of the well
components, if the well is inundated by floodwaters or surface runoff or if contamination is
suspected due to prolonged illness. Treatment is achieved through disinfection, most
commonly by shock chlorination. A Nebraska-licensed water well professional can assist well
owners with decontaminating a private water well system.

E-COLI:

What is E. Coli?

E. coli (Escherichia coli), is a type of bacteria that normally lives in your intestines. It’s also found
in the gut of some animals.
Most types of E. coli are harmless and even help keep your digestive tract healthy. But some
strains can cause diarrhea if you eat contaminated food or drink fouled water.

How Do You Get Infected?

You can become infected when you swallow even a small amount of E. coli bacteria. Among the
ways this can happen:

 Ground meat: You eat ground meat that carries E. coli, and the meat wasn’t cooked
enough to kill the bacteria. When meat is processed, sometimes bacteria from the
animals’ intestines make their way into the meat. This happens more with ground meat
because it comes from more than one animal.
 Untreated milk: You drink unpasteurized milk, which hasn’t been heated to kill bacteria.
E. coli can get into the milk from the cow’s udder or from milking equipment.
 Vegetables and fruit: You might eat fresh vegetables or fruit that’s been tainted by
water that has the bacteria. This happens most often when manure from nearby
animals mixes with the water supply.
 Other foods and beverages: You might also get E. coli from unpasteurized fruit juices
and yogurt and cheese made from raw milk.
 Water: You swallow water that contains E. coli, perhaps while swimming in a pool, lake,
or pond.

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 Other people: You might get E. coli from another person who has it, such as a child. The
bacteria can be passed to you if you clean up after an infected person and then
don’t wash your hands really well before you touch your mouth.
 Animals: It can be found at petting zoos or animal exhibits at fairs.

Symptoms

You’ll probably start to feel ill 2 to 5 days after you’ve taken in the E. coli bacteria. The most
common symptoms are:

 Abdominal cramps
 Diarrhea, which may be bloody
 Nausea
 Constant fatigue

You may not have a fever. If you do, it may be slight.

Treatments

The only way your doctor can know for sure if you have an E. coli infection is to send a
sample of your stool to a lab to be analyzed.
Fortunately, the infection usually goes away on its own.
For some types of E.coli associated with diarrhea, such as the watery travelers’
diarrhea, antibiotics can shorten the length of time you have symptoms and might be used
in moderately severe cases.
But if you have fever or bloody diarrhea or if your doctor suspects Shiga toxin-producing E.
coli, antibiotics should not be taken. They can actually increase the production of Shiga toxin
and worsen your symptoms.
It’s important to rest and get plenty of fluids to replace what your body is losing through
vomiting or diarrhea.
Don’t take over-the-counter medications that fight diarrhea. You don’t want to slow down
your digestive system, because that will delay your body’s shedding of the infection.
When you start to feel better, stick to low-fiber foods at first such as:

 Crackers
 Toast
 Eggs
 Rice

Dairy products and foods that are high in fat or fiber can make your symptoms worse.

Prevention

One of the most important things you can do to protect yourself and your family against E.
coli is wash your hands, particularly in these situations:

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 Before you prepare food

 Before preparing bottles or food for infants or toddlers
 Before touching anything, such as a pacifier, that goes into a small child’s mouth.
 After you’ve used the bathroom or changed a diaper
 After you’ve had contact with animals, even your own pets
 After handling raw meat

You can also prevent E. coli infections by being careful about the foods that carry the greatest
chance of contamination:

 Cook hamburgers until they’re 160 F inside.

 Drink only pasteurized milk, juice, and cider.
 Wash all of your produce before you eat it. Be especially careful to get dirt off
leafy greens such as lettuce and spinach.

In your kitchen, a couple of simple rules will help keep you safe:
Wash: Clean knives, counters, and cutting boards with hot, soapy water after raw meat has
touched them.
Keep raw and cooked separate: Use different cutting boards for food that you eat raw, such as
vegetables and fruit. Don’t put cooked meat back on the same plate you used for raw meat
without washing the plate first.
When you're swimming, try not to swallow the water, whether it's a pool, a lake, or the ocean. It
may be tainted with E. coli from feces.
Streptococcus
INTRODUCTION

Streptococcus species refer to the gram-positive streptococci that are not pneumococci.
There are now approximately 50 species of Streptococci, however, only five cause disease in
humans. These include: Group A—beta-hemolytic, S. pyogenes; Group B—beta hemolytic, S.
agalactiae; Group C—beta-hemolytic; Group D—Enterococcus; and, Group F—alpha-
hemolytic, S. viridans. These account for approximately 1% of adult pneumonias, and most
are due to Group A, beta-hemolytic streptococcus. Group B streptococci are a major cause
of neonatal pneumonia.

Classification

Streptococci belong to the genus Streptococcus. Classification is based on their hemolytic

effect on blood agar, antigenic composition, growth characteristics, genetic component, and
biochemical reactions. Hemolytic effect on blood agar allows differentiation into those
causing partial hemolysis [alpha (α)-hemolysis], complete hemolysis [beta (β)-hemolysis],
and those that do not cause hemolysis [gamma (γ)-hemolysis]. Pneumococci are alpha
hemolytic. They are different from all the other streptococci in having a
thick polysaccharide capsule. The capsule is composed of repeating oligosaccharides.
Antigenic differences in these polysaccharides are the basis of their identification into about

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90 different serotypes Musher (2000). Recognition of these serotypes provides the basis for
the production of the 23-valent and 7-valent vaccines.

Streptococcal Infections

Streptococci have been isolated from hamsters with pneumonia. Streptococcal pneumonias
are relatively uncommon and oftentimes are associated with stress. In a survey of naturally
occurring diseases of hamsters, diplococcus (Streptococcus pnuemoniae) was the cause of
pneumonia from one of 14 laboratories (Renshaw et al., 1975). S. agalactiae has been
reported to cause acute pleuropneumonia and septicemia in hamsters (Kummeneje et al.,
1975). Diplococci and Streptococcus sp. have been reported to cause pneumonia and
cervical lymphadenitis in hamsters (Frisk, 1987). The course of the disease is short (3 days),
the route of infection being aerosol. Clinical signs include depression, anorexia, nasal and
ocular discharge, dehydration, and weight loss. Diagnosis is made by clinical signs, lesions,
and culture. Treatment may be attempted with antibiotics that are specifically effective
against the isolated organism.

COMMENTS

Streptococci are common causes of bacterial infection of the eye and its adnexa.
Most streptococci are sensitive to the β-lactam family of antibiotics. Emerging strains of
penicillin-resistant organisms pose a threat to the treatment of streptococcal infections on
an empiric basis. A specimen for culture and in vitro antimicrobial susceptibility testing
should be obtained whenever possible. Because resistance patterns of streptococci causing
ocular infections change over time, outcomes of studies dating back more than 10 years,
may not be applicable to the current prevalence of infecting agents. Also, resistance
between regions and countries may vary considerably. Up-to-date, local characteristics and
resistance patterns of the causative bacteria should always be taken into account when
choosing antibiotic treatment. Local health authorities and other relevant bodies may advise
against prescribing certain antibiotics in order to restrict the development of bacterial
resistance and reserve these drugs for more serious infections.

CLOSTRIDIUM

What is the function of Clostridium?

Clostridium species are chemoorganotrophic bacteria. They can ferment a variety of
nutrients, like carbohydrate, protein, organic acid and other organics, to produce acetic
acid, propionic acid, butyric acid, and some solvents, such as acetone and butanol.

What are the characteristics of Clostridium?

CHARACTERISTICS: Clostridium is a genus of gram-positive, spore-forming bacteria
belonging to the family Clostridiaceae. Vegetative cells are rod shaped and arranged in
pairs or short chains. The majority of species are obligate anaerobes; however, some
species can grow under aerobic conditions or are aerotolerant.

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What are the 4 species of Clostridium?

The four clinically important species are C. tetani, C. botulinum, C. perfringens, and C.

What is the common name of Clostridium?

Clostridium perfringens (formerly known as C. welchii, or Bacillus welchii) is a Gram-
positive, rod-shaped, anaerobic, spore-forming pathogenic bacterium of the genus
Clostridium.
Where is Clostridium found in the body?
clostridium, (genus Clostridium), any of a genus of rod-shaped, usually gram-positive
bacteria, members of which are found in soil, water, and the intestinal tracts of humans
and other animals.
What antibiotics treat Clostridium?
Antibiotics are the mainstay to treat C. difficile infection. Commonly used antibiotics
include: Vancomycin (Vancocin HCL, Firvanq)

What foods contain clostridia?

Clostridium perfringens is most commonly found in meat, poultry, cooked dried beans and
gravies. Because the bacteria also live in the soil, contamination from unwashed vegetables
is also possible.

How can you reduce Clostridium?

Clean household surfaces such as countertops, sinks, faucets, bathroom doorknobs, and
toilets regularly using warm/hot water with any household soap or any bleach-containing
household cleaning product. Do not apply undiluted bleach directly to surfaces

What is a clostridial infection?

clostridial infection, any of several infectious conditions in animals and humans resulting
from Clostridium species, bacteria that are found in soil and that enter the body via
puncture wounds or contaminated food. These bacteria synthesize and release poisonous
substances called exotoxins.

Where does Clostridium come from?

Clostridium perfringens bacteria are one of the most common causes of foodborne illness
(food poisoning). CDC estimates these bacteria cause nearly 1 million illnesses in the United

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States every year. C. perfringens can be found on raw meat and poultry, in the intestines of
animals, and in the environment.

Treatment
In general, the treatment of clostridial infection is high-dose penicillin G, to which the
organism has remained susceptible.[19] Clostridium welchii and Clostridium tetani respond
to sulfonamides.[20] Clostridia are also susceptible
to tetracyclines, carbapenems (imipenem), metronidazole, vancomycin,
and chloramphenicol.[21]
The vegetative cells of clostridia are heat-labile and are killed by short heating at
temperatures above 72–75 °C. The thermal destruction of Clostridium spores requires
higher temperatures (above 121.1 °C, for example in an autoclave) and longer cooking times
(20 min, with a few exceptional cases of > 50 min recorded in the
literature). Clostridia and Bacilli are quite radiation-resistant, requiring doses of about
30 kGy, which is a serious obstacle to the development of shelf-stable irradiated foods for
general use in the retail market.[22] The addition of lysozyme, nitrate, nitrite and propionic
acid salts inhibits clostridia in various foods.[23][24][25]
Fructooligosaccharides (fructans) such as inulin, occurring in relatively large amounts in a
number of foods such as chicory, garlic, onion, leek, artichoke, and asparagus, have
a prebiotic or bifidogenic effect, selectively promoting the growth and metabolism of
beneficial bacteria in the colon, such as bifidobacteria and lactobacilli, while inhibiting
harmful ones, such as clostridia, fusobacteria, and bacteroides

Pathogenesis and classification

Clostridium contains around 250 species that include common free-living bacteria, as well as
important pathogens.[10][11] The main species responsible for disease in humans are:[12]

 Clostridium botulinum can produce botulinum toxin in food or wounds and can
cause botulism. This same toxin is known as Botox and is used in cosmetic surgery to
paralyze facial muscles to reduce the signs of aging; it also has numerous
other therapeutic uses.
 Clostridium perfringens causes a wide range of symptoms, from food
poisoning to cellulitis, fasciitis, necrotic enteritis[13] and gas gangrene.[14]
 Clostridium tetani causes tetanus.
 Clostridium sordellii (now Paeniclostridium) can cause a fatal infection in exceptionally
rare cases after medical abortions.[15]
Bacillus and Clostridium are often described as Gram-variable, because they show an
increasing number of gram-negative cells as the culture ages.[16]
Clostridium and Bacillus are both in the phylum Firmicutes, but they are in different classes,
orders, and families. Microbiologists distinguish Clostridium from Bacillus by the following
features:[2]

 Clostridium grows in anaerobic conditions; Bacillus grows in aerobic conditions.

 Clostridium forms bottle-shaped endospores; Bacillus forms oblong endospores.

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 Clostridium does not form the enzyme catalase; Bacillus secretes catalase to destroy
toxic byproducts of oxygen metabolism.
Clostridium and Desulfotomaculum are both in the class Clostridia and order Clostridiales,
and they both produce bottle-shaped endospores, but they are in different
families. Clostridium can be distinguished from Desulfotomaculum on the basis of the
nutrients each genus uses (the latter requires sulfur).
Glycolysis and fermentation of pyruvic acid by Clostridia yield the end products butyric
acid, butanol, acetone, isopropanol, and carbon dioxide.[16]
The Schaeffer-Fulton stain (0.5% malachite green in water) can be used to distinguish
endospores of Bacillus and Clostridium from other microorganisms.[17] There is a
commercially available polymerase chain reaction (PCR) test kit (Bactotype) for the
detection of C. perfringens and other pathogenic bacteria.[18]

BIOLEECHING

BIOLEACHING DEFINITION & PROCESS

What is bioleaching or biomining?

Bioleaching (or biomining) is a process in mining and biohydrometallurgy (natural processes

of interactions between microbes and minerals) that extracts valuable metals from a low-
grade ore with the help of microorganisms such as bacteria or archaea.

Bioleaching techniques are often more effective than traditional mining applications and
can even be used to clean mine tailings sites.

Metals extracted from bioleaching include:

 Gold
 Copper
 Silver
 Cobalt
 Uranium
 Zinc
 Nickel

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How does the bioleaching process work?

There are many types of bioleaching processes, and copper is the most common. A few of
the most popular types of bioleaching extract metals from ore by retrieving sulfide minerals
using bacteria that receive energy from non-carbon compounds.

Below are a few approaches to bioleaching:

Direct v. Indirect Bioleaching

Direct bioleaching uses minerals that are easily receptive to oxidation to create a direct
enzymatic strike using the microorganisms to separate the metal and the ore.

In indirect bioleaching, microorganisms are not in direct contact with minerals during the
process. However, leaching agents are created by microbes, which still oxidise the ore.

Some advantages of bioleaching include:

 Bioleaching can stabilise sulphate toxins from the mine without causing harm to the
environment.
 Poisonous sulfur dioxide emissions harm the environment and can cause health problems
for miners, and bioleaching avoids this process entirely.
 Bioleaching is more cost-effective than smelting processes.
 Some Bioleaching offers a different way to extract valuable metals from low-grade ores
that have already been processed.

The Commercial Process

Commercial metal extraction is a quicker process that can be optimized through humidity,
potential hydrogen (pH), temperature, and chemical elements.

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The 3 most common commercial biomining processes are:

1. Slope Leaching

Fine ore is kept in a large, slope-shaped dump. During slope leaching, a water solution made
of inoculum is continuously sprayed over the ore. After that, the leach liquor (or remaining
liquid) is gathered at the bottom and processed for supplemental metal recovery.

2. Heap Leaching

In this technique, the ore is arranged in large heaps. During heap leaching, an aqueous
mixture of microorganisms is sprinkled over the leach pile. Then, the solution is collected
and processed to help recover even more metal.

3. In-situ Leaching

The ore remains in its natural state while the leaching process takes place. Water that
contains thiobacillus is pushed through drilled passageways within the ore. The leach fluid is
then stored until it is time for metal recovery.

What's next for bioleaching?

The world of tomorrow relies on how we produce the global demands of today. As
technology continues to progress, there will be more room for improvements in how the
industry works.

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UNIT -5 – TOXICOLOGY

TOXICOLOGY
Ecotoxicology – toxicants and toxicity, Factors influencing toxicity, Effects – acute, chronic,
Test organisms – toxicity testing, Bioconcentration – Bioaccumulation, biomagnification,
bioassay, biomonitoring, bioleaching.

Ecotoxicology

Ecotoxicology is the study of the effects of toxic chemicals on biological organisms, especially
at the population, community, ecosystem level. Ecotoxicology is a multidisciplinary field,
which integrates toxicology and ecology.

A toxicant is any toxic substance made by humans or introduced into the environment by
human activity. Toxicants are poisonous. Examples of toxicants are man-made chemicals such
as insecticides, bisphenol, and millions of other industrial chemicals.
“Toxin” is used when poisons are produced biologically or from living organisms

Examples of toxins are the venom which comes from snakes as well as viruses which can harm
humans because these viruses can be poisonous to humans
TOXIXITY:
Toxicity is the degree to which a substance can damage an organism. Toxicity can refer to the
effect on a whole organism, such as an animal, bacterium, or plant, as well as the effect on a
substructure of the organism, such as a cell (cytotoxicity) or an organ such as the liver
(hepatotoxicity).
By extension, the word may be metaphorically used to describe toxic effects on larger and
more complex groups, such as the family unit or society at large. Sometimes the word is more
or less synonymous with poisoning in everyday usage

There are generally four types of toxic entities; chemical, biological, physical and radiation:

 Chemical toxicants include inorganic substances such as, lead, mercury, hydrofluoric
acid, and chlorine gas, and organic compounds such as methyl alcohol, most
medications, and poisons from living things. While some weakly radioactive
substances, such as uranium, are also chemical toxicants, more strongly radioactive
materials like radium are not, their harmful effects (radiation poisoning) being caused
by the ionizing radiation produced by the substance rather than chemical interactions
with the substance itself.

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 Disease-causing microorganisms and parasites are toxic in a broad sense, but are
generally called pathogens rather than toxicants. The biological toxicity of pathogens
can be difficult to measure because the "threshold dose" may be a single organism.
Theoretically one virus, bacterium or worm can reproduce to cause a serious infection.

 However, in a host with an intact immune system the inherent toxicity of the organism
is balanced by the host's ability to fight back; the effective toxicity is then a
combination of both parts of the relationship. In some cases, e.g. cholera, the disease
is chiefly caused by a nonliving substance secreted by the organism, rather than the
organism itself. Such nonliving biological toxicants are generally called toxins if
produced by a microorganism, plant, or fungus, and venoms if produced by an animal.

 Physical toxicants are substances that, due to their physical nature, interfere with
biological processes. Examples include coal dust, asbestos fibers or finely divided
silicon dioxide, all of which can ultimately be fatal if inhaled. Corrosive chemicals
possess physical toxicity because they destroy tissues, but they're not directly
poisonous unless they interfere directly with biological activity.

 Water can act as a physical toxicant if taken in extremely high doses because the
concentration of vital ions decreases dramatically if there's too much water in the
body. Asphyxiant gases can be considered physical toxicants because they act by
displacing oxygen in the environment but they are inert, not chemically toxic gases.

 Radiation can have a toxic effect on organisms.

In some instances, individuals can have unpredictable reactions, or idiosyncratic responses,

to a drug or other substance. An idiosyncratic response is uncommon, and it is sometimes
impossible to understand whether it is the result of a genetic predisposition or has some other
cause such as the status of the immune system. It could result in an abnormally small or short,
or abnormally large or long response to the drug or other substance. Or, the response could
be qualitatively different than what has been observed in most other individuals.

The toxicity of a substance usually depends on the following factors:

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 Form and innate chemical activity

 Dosage, especially dose-time relationship
 Exposure route
 Species
 Life stage, such as infant, young adult, or elderly adult
 Gender
 Ability to be absorbed
 Metabolism
 Distribution within the body
 Excretion
 Health, including organ function, of the individual. Also, pregnancy involves
physiological changes that could influence toxicity.
 Nutritional status
 Presence of other chemicals
 Circadian rhythms (the time of day a drug or other substance is administered)
 Acute exposure
 A single exposure to a toxic substance which may result in severe biological harm or
death; acute exposures are usually characterized as lasting no longer than a day.
 Chronic exposure
 Continuous exposure to a toxin over an extended period of time, often measured in
months or years; it can cause irreversible side effects.

Toxicity is the ability of a substance to cause harmful health effects. These effects can
strike a single cell, a group of cells, an organ system, or the entire body.
 A toxic effect may be visible damage, or a decrease in performance or function
measurable only by a test.
 All chemicals can cause harm at a certain level. When a small amount can be harmful,
the chemical is considered toxic. When only a very large amount of the chemical can
cause damage, the chemical is considered to be relatively non-toxic.

The toxicity of a substance depends on three factors: its chemical structure, the
extent to which the substance is absorbed by the body, and the body’s ability to
detoxify the substance (change it into less toxic substances) and eliminate it from the
body.

Factors that influence toxicity

There are many factors that can increase or decrease the toxicity of toxicants or stressors,
making interpretation of test results difficult. These can be chemical, biological, or
toxicological.

Chemical factors

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Water chemistry plays an important role in the toxicity of certain toxicants. This includes pH,
salinity, water hardness, conductivity, temperature, and amounts of dissolved organic carbon
(DOC) For instance, the toxicity of copper is decreased with increasing amounts of DOC, as
described by the biotic ligand model (BLM).[5]

Biological factors

Chronic toxicity will vary with differences in organisms, including species, size, and age.
Certain species are more susceptible to toxic effects, as shown in species sensitivity
distributions (SSDs). Certain life stages are more susceptible to adverse effects, which is why
early life stage (ELS) toxicity tests are performed for certain aquatic species.
In addition, other physical factors, like organism size, can lead to differences in response to
toxicants.
The toxicity of a substance is the potential of that substance to cause harm, and is only one
factor in determining whether a hazard exists.

The hazard of a chemical is the practical likelihood that the chemical will cause harm. A
chemical is determined to be a hazard depending on the following factors:

 toxicity:
 how much of the substance is required to cause harm,
 route of exposure:how the substance enters your body,
 dose:how much enters your body,
 duration:the length of time you are exposed,
 multiple exposures:other chemicals you are exposed to, and individual susceptibility:
 how your body reacts to the substance, compared to other individuals.

 In general, the greater the amount of a substance that enters your body, the greater
is the effect on your body. This connection between amount and effect is called the
dose-response relationship
.
 For example, solvents such as toluene, acetone, and trichloroethylene all affect the
brain in the same way, but to different degrees at different doses. The effects of these
solvents are similar to those which result from drinking alcoholic beverages. At a low
dose, you may feel nothing or a mild, sometimes pleasant (“high”) sensation. A larger
dose may cause dizziness or headache. With an even larger dose you may feel as if
you are drunk, pass out, or even stop breathing.When you inhale a toxic chemical, the
dose you receive depends on four factors:

 the level (concentration) of chemical in the air,

 how hard (fast and deep) you are breathing, which depends on your degree of physical
exertion,
 how much of the chemical that is inhaled stays in your lungs or is absorbed into your
bloodstream, and

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 how long the exposure lasts.

When a toxic substance causes damage at the point where it first contacts the body, that
damage is called a local effect. The most common points at which substances first contact the
body are the skin, eyes, nose, throat, and lungs.

Many toxic substances can also enter the body and travel in the bloodstream to internal
organs.
Effects that are produced this way are called systemic.

The internal organs most commonly affected are the liver, kidneys, heart, nervous system
(including the brain), and reproductive system.

A toxic chemical may cause local effects, systemic effects, or both. For example, if ammonia
gas is inhaled, it quickly irritates the lining of the respiratory tract (nose, throat, and lungs).
Almost no ammonia passes from the lungs into the blood. Since damage is caused only at the
point of initial contact, ammonia is said to exert a local effect. An epoxy resin is an example
of a substance with local effects on the skin. On the other hand, if liquid phenol contacts the
skin, it irritates the skin at the point of contact (a local effect) and can also be absorbed
through the skin, and may damage the liver and kidneys (systemic effects).

Sometimes, as with phenols, the local effects caused by a chemical provide a warning that
exposure is occurring. You are then warned that the chemical may be entering your body and
producing systemic effects which you can’t yet see or feel. Some chemicals, however, do not
provide much warning, so they are particularly hazardous. For example, some toxic solvents
can pass through the skin and cause serious internal damage without producing any
observable effect on the skin.

Effects
Acute exposure
A single exposure to a toxic substance which may result in severe biological harm or
death; acute exposures are usually characterized as lasting no longer than a day.

Chronic exposure
Continuous exposure to a toxin over an extended period of time, often measured in
months or years; it can cause irreversible side effects.

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Acute toxicity describes the adverse effects of a substance that result either from a single
exposure or from multiple exposures in a short period of time (usually less than 24 hours). To
be described as acute toxicity, the adverse effects should occur within 14 days of the
administration of the substance.

Acute toxicity is distinguished from chronic toxicity, which describes the adverse health
effects from repeated exposures, often at lower levels, to a substance over a longer time
period (months or years).

It is widely considered unethical to use humans as test subjects for acute (or chronic) toxicity
research. However, some information can be gained from investigating accidental human
exposures (e.g., factory accidents). Otherwise, most acute toxicity data comes from animal
testing or, more recently, in vitro testing methods and inference from data on similar
substances.

There are also different values based on the method of entry of the compound (oral, dermal,
or inhalation).

 Threshold limit value-time-weighted-average: The maximum concentration to which

a worker can be exposed every work day (8 hours) and experience no adverse health
effects.
 Short-Term Exposure Limit, STEL or Threshold limit value-short-term exposure limit,
TLV-STEL: The concentration which no person should be exposed to for more than 15
minutes during an 8-hour work day.
 Ceiling value, CV or Threshold limit value-ceiling, TLV-C: The concentration which no
person should ever be exposed to.

Responses and treatments

When a person has been exposed to an acutely toxic dose of a substance, they can be treated
in a number of ways in order to minimize the harmful effects. Obviously, the severity of the
response is related to the severity of the toxic response exhibited. These treatment methods
include (but are not limited to):

 Emergency showers used for removing irritating or hazardous chemicals from the skin.
 Emergency eye washes used for removing any irritating or hazardous chemicals from
the eyes.
 Activated charcoal used to bind and remove harmful substances consumed orally. This
is used as an alternative to conventional stomach pumping.

Chronic toxicity, the development of adverse effects as a result of long term exposure to a
contaminant or other stressor, is an important aspect of aquatic toxicology.[1] Adverse effects

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associated with chronic toxicity can be directly lethal but are more commonly sublethal,
including changes in growth, reproduction, or behavior. Chronic toxicity is in contrast to acute
toxicity, which occurs over a shorter period of time to higher concentrations.

Various toxicity tests can be performed to assess the chronic toxicity of different
contaminants, and usually last at least 10% of an organism’s lifespan.[2] Results of aquatic
chronic toxicity tests can be used to determine water quality guidelines and regulations for
protection of aquatic organisms.

Chronic toxicity tests are performed to determine the long term toxicity potential of toxicants
or other stressors, commonly to aquatic organisms. Examples of common aquatic chronic
toxicity test organisms, durations, and endpoints include:

 Fathead minnow, Pimephales promelas, larval survival and growth

 Daphnia, Daphnia magna, 21-d survival and reproduction
 Green algae, Raphidocelis subcapitata, 72-h growth
 Amphipod, Hyalella azteca, 42-d survival, growth, and reproduction

Application of chronic toxicity test results

Results from chronic toxicity tests can be used to calculate values that can be used for
determining water quality standards. These include:

NOEC/LOEC

The no observed effects concentration (NOEC) is determined as the highest tested

concentration that shows no statistically significant difference from the control. The lowest
observed effects concentration (LOEC) is the lowest concentration of those tested that
produced a statistically significant difference from the control. NOECs and LOECs can be
derived from both acute and chronic tests and are used by agencies to set water quality
standards.

MATC/CV

The maximum acceptable toxicant concentration (MATC) is calculated as the geometric mean
of the NOEC and LOEC. MATC is sometimes called the chronic value (CV) and defined as “the
concentration (threshold) at which chronic effects are first observed”.[3]

Challenges with chronic toxicity testing

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The chronic toxicity of toxicants is useful information to know in determining water quality
guidelines, but this information is not always easily obtained. Chronic toxicity tests can be
costly and difficult, due to challenges in keeping control organisms alive, maintaining water
quality, retaining constant chemical exposures, and the sheer time required for tests. Because
of this, acute toxicity tests are more commonly employed, and ACRs and AFs are used to
estimate chronic toxicity of toxicants to organisms.

Toxicity tests on organisms - For all new chemicals before being released on the market and
also for the reevaluation of the safety of chemicals already used. - It is necessary to unify them
all around the world to avoid excessive use of laboratory organisms and to decrease the price
of testing. Once a test is done according to approved methods in one country, it can be
accepted after a short evaluation and without further testing also in other countries. –
The rule of three R: Reduction (less laboratory animals) Replacement (replace with
evolutionary lower organisms and if possible, by tissue cultures, cells, computer modeling)
Refinement (the animal should be stressed and harmed as little as possible, welfare of
laboratory animal is absolutely necessary)

In toxicology, the median lethal dose, LD50 (abbreviation for "lethal dose, 50%"), LC50 (lethal
concentration, 50%) or LCt50 is a measure of the lethal dose of a toxin, radiation, orpathogen.
The value of LD50 for a substance is the dose required to kill half the members of a tested
population after a specified test duration. LD50 figures are frequently used as a general
indicator of a substance's acute toxicity. A lower LD50 is indicative of increased toxicity.

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Whole Effluent Toxicity (WET) refers to the aggregate toxic effect to aquatic organisms from
all pollutants contained in a facility's wastewater (effluent). It is one way we implement the
Clean Water Act's prohibition of the discharge of toxic pollutants in toxic amounts. WET tests
measure wastewater's effects on specific test organisms' ability to survive, grow and
reproduce.
LD50 (lethal dose which causes death of 50 % of tested animals)
LC50 (lethal concentration which causes death of 50 % of tested animals)
NOEC (no observed effect concentration – the highest concentration tested in which no
adverse effect was detected)
LOEC (lowest observed effect concentration – the lowest concentration tested in which an
adverse effect was detected)
Based on the value of LD/LC/LOEC assessed, chemical is classified and described by Risk and
Safety Statements, also known as R/S statements or R/S sentences, a system of hazard codes
and phrases.

Toxicology testing, also known as safety assessment, or toxicity testing, is conducted to

determine the degree to which a substance can damage a living or non-living organisms. It is
often conducted by researchers using standard test procedures to comply with governing
regulations, for example for medicines and pesticides. Much toxicology is considered to be
part of the field of preclinical development. Stages of in vitro and in vivo research are
conducted to determine safe doses of exposure in humans before a first-in-man study.
Toxicology testing may be conducted by the pharmaceutical
industry, biotechnology companies or contract research organizations.

Bio concentration is a related but more specific term, referring to uptake and accumulation
of a substance from water alone. By contrast, bioaccumulation refers to uptake from all
sources combined (e.g. water, food, air, etc.)

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Bioaccumulation

Bioaccumulation is defined as the increase in concentration of a substance(s) in an organism

or a part of that organism. Toxic substances are lipophilic or fat-loving, the reason why these
substances are deposited and concentrated in the fat tissues of the organisms. The affected
organism has a higher concentration of the substance than the concentration in the
organism's surrounding environment.
The toxic substances are very slowly metabolized or excreted so if the organism keeps on
consuming prey or food contaminated with toxic substances, the concentration of the
substance will further increase in its body, hence, bioaccumulation results. When a certain
threshold level is reached, measured in parts per million (ppm), symptoms due to the type of
toxin are manifested.
Examples of toxic substances that bioaccumulate are lead, mercury,
dichlorodiethyltrichloroethane (DDT), among others. Lead exposure can be serious for young
children because their growing bodies absorb lead more easily than adults. They are more
susceptible to its harmful effects.
Even low level of lead exposure can harm the intellectual development, behavior, growth,
and hearing of infants. High concentrations of lead in pregnant mothers can cause
miscarriages and stillbirths. Mercury, as mentioned above, can cause paralysis. DDT at
concentration above 236 mg per kg of body weight, can cause death among humans.
Concentration of 6-10 mg/kg leads to such symptoms as headache, nausea, vomiting,
confusion, and tremors.
or

Bioaccumulation refers to the accumulation of substances, such as pesticides, or other

chemicals in an organism. Bioaccumulation occurs when an organism absorbs a -
possibly toxic - substance at a rate faster than that at which the substance is lost
by catabolism and excretion.

Thus, the longer the biological half-life of a toxic substance the greater the risk of chronic
poisoning, even if environmental levels of the toxin are not very high. Bioaccumulation, for
example in fish, can be predicted by models.

Hypotheses for molecular size cutoff criteria for use as bioaccumulation potential indicators
are not supported by data. Biotransformation can strongly modify bioaccumulation of
chemicals in an organism.

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Biomagnification
Biomagnification is also called Bioamplification. It is simply the increase in concentration of a
substance in a food chain, not an organism. Persistent organic pollutants (POPs) are
compounds that biomagnify. Persistent organic pollutants (POPs) are chemical substances
that persist in the environment.
These substances bioaccumulate through the food web and pose risk not only to humans but
also other living organisms because of their adverse effects. These pollutants consists of
pesticides (such as DDT), industrial chemicals (such as polychlorinated biphenyls, PCBs) and
unintentional by-products of industrial processes (such as dioxins and furans).

In essence, bio magnification is similar to bioaccumulation but is descriptive of higher level

biological processes, not individual.

Adequate care must therefore be exercised in choosing what to ingest. Foods may be laced
with toxic substances that are harmful to the body when consumed in large quantities.
BIOASSAY
Bioassay ( biological assessment), or biological standardization is a type of
scientific experiment. A bioassay involves the use of live animal or plant (in vivo) or tissue or
cell (in vitro) to determine the biological activity of a substance, such as a hormone or drug.
Bioassays are typically conducted to measure the effects of a substance on a
living organism and are essential in the development of new drugs and in monitoring
environmental pollutants.
Both are procedures by which the potency or the nature of a substance is estimated by
studying its effects on living matter. A bioassay can also be used to determine the
concentration of a particular constitution of a mixture that may cause harmful effects on
organisms or the environment.

BIOMONITORING

BIOLOGICAL MONITORING, OR BIOMONITORING, is the use of biological responses to assess

changes in the environment, generally changes due to anthropogenic causes. Biomonitoring
programs may be qualitative, semi-quantitative, or quantitative. Biomonitoring is a valuable
assessment tool that is receiving increased use in water quality monitoring programs of all
types.
There are two types of biomonitoring. One type of biomonitoring is surveillance before and
after a project is complete or before and after a toxic substance enters the water. The other
type of biomonitoring is to ensure compliance with regulations or guidelines or to ensure
water quality is maintained.

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Biomonitoring involves the measurement of chemicals in humans (and other species) and is
rapidly advancing our ability to detect and quantify potentially hazardous exposure to a wide
range of chemicals. Biomonitoring studies now routinely yield novel, and sometimes
surprising, information about the accumulation of chemicals in living receptors. How best to
enhance, interpret, and utilize biomonitoring is an important policy issue.

Biomonitoring may have its greatest value in dealing with chemicals that alter the
development of living things. In recent years, it has become clear that many anthropogenic
chemicals, including some that are widely used, can interfere with the cellular signaling
mechanisms controlling development, even at the low levels of environmental exposure
experienced by the general population. Both animal and human studies link altered cellular
signaling to developmental impairments.

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Bioleaching or Microbial ore leaching is the extraction of metals from their ores through the
use of living organisms. This is much cleaner than the traditional heap leaching using cyanide.
Bioleaching is one of several applications within biohydrometallurgy and several methods are
used to recover copper, zinc, lead, arsenic, antimony, nickel, molybdenum, gold, silver,
and cobalt.
 Bioleaching is cheaper than chemical extraction, safer for the environment, and more
efficient in extracting metals with low concentration in ores.

 It is performed by iron and sulfide oxidizing bacteria or acid producing fungus.

 Bacteria recycle the major leaching reagent, like ferric iron, and perform
further oxidation steps while gaining energy from the electron transfer.

 lower the production costs.

 cause less environmental pollution in comparison to the traditional leaching methods.

 very efficiently extract metals when their concentration in the ore is low.

How is bioaccumulation differentiated from biomagnification? Are these two concepts

similar?

Bioaccumulation refers to the accumulation of a toxic chemical in the tissue of a particular

organism. Biomagnification refers to the increased concentration of a toxic chemical the
higher an animal is on the food chain.
University questions :-
2marks
What are the objectives of bioassay test ?
What are Toxicants ?/ Define Toxicants
What is bio leaching
16marks :-
What is ecotoxicology ?write about factors influencing toxicity
Define bio concentration. What is the significance of biomonitering ?
Illustrate using examples that effect of acute and chronic in the tested organisms

Write short notes on the following Bioaccumulation ii. Biomonitoring iii. Biomagnification iv
.Bioleaching

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