Basic Microbiology Notes

Prepared by: Arnevynce Laurel

Basic Microbiology - Lecture Notes

I. History and Scope of Microbiology

Microbiology is the study of microorganisms, which are living organisms too
small to be seen with the naked eye. These include bacteria, viruses, fungi,
protozoa, algae, and prions. Microbiology plays a significant role in various
fields, such as medicine, agriculture, industry, and environmental science.

A. Historical Background

1. Early Discoveries

 Antonie van Leeuwenhoek (1673-1723)

o Known as the "Father of Microbiology."

o Developed simple microscopes and was the first to observe and

describe microorganisms, which he called "animalcules."

 Robert Hooke (1665)

o Coined the term "cell" after observing cork under a microscope.

o Published Micrographia, documenting microscopic structures.

2. Spontaneous Generation vs. Biogenesis

 Spontaneous Generation Theory

o Suggested that life could arise spontaneously from non-living

matter.

o Advocated by scientists like Aristotle and supported until the

17th century.

 Disproving Spontaneous Generation

o Francesco Redi (1668): Demonstrated that maggots only form

in meat when flies can lay eggs on it.
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o Louis Pasteur (1861): Used swan-neck flask experiments to
show that microorganisms come from the air, not spontaneous
generation.

3. Germ Theory of Disease

 Proposed that microorganisms are the cause of diseases.

 Louis Pasteur (1822-1895): Demonstrated that microbes cause

fermentation and spoilage. Developed vaccines for rabies and anthrax.

 Robert Koch (1843-1910): Formulated Koch’s postulates to establish

the link between a specific microorganism and a disease. Discovered
the causative agents of tuberculosis, anthrax, and cholera.

4. Advances in Microbiology

 Edward Jenner (1796): Developed the first vaccine (smallpox

vaccine) using cowpox material.

 Alexander Fleming (1928): Discovered penicillin, the first antibiotic,

which revolutionized medicine.

B. Branches of Microbiology

Microbiology is a broad field with specialized branches, including:

1. Medical Microbiology

o Study of pathogens and diseases they cause.

o Focuses on diagnosis, treatment, and prevention of infectious

diseases.

2. Environmental Microbiology

o Study of microbes in natural environments, including soil, water,

and air.

o Explores roles in nutrient cycling, pollution control, and

bioremediation.

3. Industrial Microbiology

o Use of microbes in industries for the production of antibiotics,

enzymes, biofuels, and fermented foods.
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4. Agricultural Microbiology

o Study of microorganisms that impact agriculture.

o Includes soil fertility, pest control, and symbiotic relationships

(e.g., nitrogen-fixing bacteria in legumes).

5. Food Microbiology

o Study of microbes in food production and spoilage.

o Ensures food safety and enhances fermentation processes.

6. Virology

o Study of viruses, their structure, function, and interactions with

host organisms.

7. Mycology

o Study of fungi, including yeasts, molds, and mushrooms.

8. Parasitology

o Study of protozoa and parasitic worms that cause diseases.

C. Importance of Microbiology

1. In Medicine

o Development of vaccines, antibiotics, and diagnostic tools.

o Understanding disease mechanisms to improve public health.

2. In Agriculture

o Enhances crop production through biofertilizers and pest control.

o Protects plants from microbial diseases.

3. In Industry

o Produces alcohol, organic acids, vitamins, and biofuels.

o Advances in biotechnology for sustainable production.

4. In Environmental Science

o Addresses pollution through bioremediation.

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o Supports ecosystems by recycling nutrients.

5. In Research

o Provides insights into genetic engineering, cell biology, and

immune responses.

D. Scope of Microbiology

Microbiology continues to evolve with technological advancements,

expanding its scope into fields like:

1. Molecular Biology and Biotechnology

o Genetic engineering of microorganisms to produce insulin,

hormones, and vaccines.

2. Immunology

o Study of immune responses to microbial infections.

o Development of immunotherapies and vaccines.

3. Astrobiology

o Exploration of microorganisms' potential to survive in

extraterrestrial environments.

4. Nanotechnology

o Using microbes in nanomaterial production and biosensors.

5. Public Health

o Study of epidemiology and microbial surveillance to control

pandemics.

E. Key Contributions to Microbiology

 Louis Pasteur: Pasteurization, vaccines, fermentation.

 Robert Koch: Germ theory, Koch's postulates.

 Antonie van Leeuwenhoek: Microscopy and discovery of

microorganisms.

 Alexander Fleming: Penicillin, the first widely used antibiotic.

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II. Taxonomy
Taxonomy is the science of classifying and organizing living organisms based
on their characteristics, relationships, and evolutionary history. It provides a
structured framework to identify, name, and group organisms in a way that
reflects their biological relationships.

A. Taxonomy vs. Systematics

1. Taxonomy

o Deals with the identification, classification, and naming of

organisms.

o Focuses on organizing organisms into categories (e.g., genus,

species).

o Provides a static framework for grouping organisms.

2. Systematics

o Broader than taxonomy; includes evolutionary relationships.

o Focuses on reconstructing the phylogenetic relationships among

organisms.

o Uses data from morphology, genetics, and molecular biology to

understand evolutionary history.

Example:

 Taxonomy: Classifying a bacterium as Escherichia coli.

 Systematics: Understanding how Escherichia coli is related to other

bacteria in evolutionary terms.

B. Components of Taxonomy

Taxonomy consists of three interrelated components: classification,

identification, and nomenclature.

i. Classification
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 Definition: The process of arranging organisms into groups (taxa)
based on shared characteristics.

 Purpose: To organize diversity and reveal evolutionary relationships.

 Hierarchy of Classification (Taxonomic Ranks):

o Domain (e.g., Bacteria, Archaea, Eukarya)

o Kingdom

o Phylum

o Class

o Order

o Family

o Genus

o Species

Example:

o Domain: Bacteria

o Kingdom: Eubacteria

o Phylum: Proteobacteria

o Class: Gammaproteobacteria

o Order: Enterobacterales

o Family: Enterobacteriaceae

o Genus: Escherichia

o Species: Escherichia coli

ii. Identification

 Definition: The process of determining the identity of an unknown

organism by comparing its traits with known organisms.

 Methods of Identification:

o Morphological Characteristics: Shape, size, and structure.

o Biochemical Tests: Enzyme activity and metabolic capabilities.

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o Molecular Techniques: DNA sequencing and polymerase chain
reaction (PCR).

o Immunological Tests: Detecting specific antigens or

antibodies.

Example: Identifying a bacterium as Staphylococcus aureus based on its

gram-positive staining, cluster arrangement, and coagulase activity.

iii. Nomenclature

 Definition: The process of assigning names to organisms according to

established rules.

 Purpose: To ensure a standardized system of naming organisms.

 Rules of Nomenclature:

o Governed by the International Code of Nomenclature of

Bacteria (ICNB) for prokaryotes and the International Code of
Zoological Nomenclature (ICZN) for animals.

o Scientific names are binomial:

 Genus name: Capitalized and italicized (e.g., Homo).

 Species name: Lowercase and italicized (e.g., sapiens).

Example:

 Scientific Name: Escherichia coli

o Escherichia (Genus)

o coli (Species)

III. Classification of Microorganisms

Classification is the process of organizing microorganisms into groups (taxa)
based on their shared characteristics and evolutionary relationships. The
classification system helps scientists study, identify, and understand
microorganisms systematically.

A. Hierarchical Classification of Microorganisms

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Microorganisms are classified into a hierarchy of taxonomic ranks, starting
from the broadest category (domain) to the most specific (species). The main
taxonomic ranks are:

1. Domain

o The highest taxonomic rank, grouping organisms based on

fundamental differences in cell structure and genetics.

o Three domains:

 Bacteria: Prokaryotic, single-celled organisms without a

nucleus.

 Archaea: Prokaryotic organisms similar to bacteria but

with distinct genetic and biochemical traits, often found in
extreme environments.

 Eukarya: Eukaryotic organisms with cells that have a

nucleus and membrane-bound organelles.

2. Kingdom

o Divides domains into broad categories.

Examples:

 Bacteria Domain: Single kingdom (Bacteria).

 Eukarya Domain: Includes kingdoms like Protista, Fungi,

Plantae, and Animalia.

3. Phylum

4. Class

5. Order

6. Family

7. Genus

8. Species

o The most specific rank, representing organisms that can

interbreed or share a high degree of similarity.

Example Classification: Escherichia coli

 Domain: Bacteria
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 Kingdom: Bacteria

 Phylum: Proteobacteria

 Class: Gammaproteobacteria

 Order: Enterobacterales

 Family: Enterobacteriaceae

 Genus: Escherichia

 Species: coli

B. Types of Microorganisms

Microorganisms are broadly classified into the following groups:

1. Bacteria

 Prokaryotic, single-celled organisms with a simple cell structure.

 Lack a nucleus and membrane-bound organelles.

 Reproduce asexually by binary fission.

 Shapes: Cocci (spherical), Bacilli (rod-shaped), Spirilla (spiral).

 Example: Escherichia coli (E. coli).

2. Archaea

 Prokaryotic organisms with unique genetic and metabolic traits.

 Thrive in extreme environments (e.g., high temperature, salinity, or

acidity).

 Do not have peptidoglycan in their cell walls.

 Example: Methanogens (produce methane gas).

3. Fungi

 Eukaryotic organisms that can be unicellular (yeasts) or multicellular

(molds, mushrooms).

 Have a cell wall made of chitin.

 Reproduce sexually or asexually via spores.

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 Example: Saccharomyces cerevisiae (baker's yeast).

4. Protozoa

 Unicellular, eukaryotic organisms that are mostly motile.

 Lack a cell wall but have a flexible cell membrane.

 Use flagella, cilia, or pseudopodia for movement.

 Example: Amoeba proteus.

5. Algae

 Eukaryotic, photosynthetic microorganisms found in aquatic

environments.

 Can be unicellular or multicellular.

 Contain chlorophyll and perform oxygenic photosynthesis.

 Example: Chlorella (unicellular green algae).

6. Viruses

 Acellular (non-living) entities composed of genetic material (DNA or

RNA) enclosed in a protein coat (capsid).

 Require a host cell to replicate.

 Example: Influenza virus.

7. Prions

 Infectious protein particles that lack nucleic acids.

 Cause neurodegenerative diseases by inducing misfolding of normal

proteins.

 Example: Prion causing Creutzfeldt-Jakob disease.

C. Basis of Microbial Classification

Microorganisms are classified based on the following criteria:

1. Morphological Characteristics

o Shape, size, arrangement, and structure (e.g., cocci, bacilli).

2. Physiological and Biochemical Properties

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o Nutritional requirements, metabolic pathways, and enzyme
production.

3. Genetic and Molecular Analysis

o DNA and RNA sequencing to determine evolutionary

relationships.

4. Cell Structure

o Prokaryotic vs. Eukaryotic.

o Presence or absence of cell wall and its composition (e.g.,

peptidoglycan in bacteria).

5. Ecological Role

o Autotrophs (producers) vs. Heterotrophs (consumers).

o Habitat and ecological niche.

6. Pathogenicity

o Ability to cause disease in humans, animals, or plants.

D. Importance of Microbial Classification

1. Organized Study: Helps scientists understand the diversity and

relationships among microorganisms.

2. Medical Applications: Identifies pathogens for diagnosis, treatment,

and prevention of diseases.

3. Industrial Use: Identifies beneficial microbes for biotechnological

applications.

4. Environmental Impact: Aids in studying microbial roles in

ecosystems and biogeochemical cycles.

IV. Microbial Diversity / Survey of the

Microbial World
Microbial diversity refers to the vast range of microorganisms, including
cellular and acellular forms. Microorganisms inhabit almost every
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environment on Earth and are categorized based on their cellular structure,
function, and reproduction.

A. Cellular Microorganisms – The Three Domains

Cellular microorganisms are living organisms with cell-based structures. They

are classified into three domains: Archaea, Bacteria, and Eukarya.

i. Archaea

 Characteristics:

o Prokaryotic organisms (no nucleus or membrane-bound

organelles).

o Unique cell membranes composed of ether-linked lipids.

o Cell walls lack peptidoglycan.

o Thrive in extreme environments (extremophiles), such as hot

springs, salt lakes, and deep-sea hydrothermal vents.

 Types:

o Methanogens: Produce methane gas (e.g., Methanobacterium).

o Halophiles: Thrive in high-salt environments (e.g.,

Halobacterium).

o Thermoacidophiles: Live in hot, acidic conditions (e.g.,

Sulfolobus).

ii. Bacteria

 Characteristics:

o Prokaryotic organisms with simple structures.

o Cell walls often contain peptidoglycan.

o Found in diverse environments, including soil, water, and living

hosts.

o Play roles in decomposition, nutrient cycling, and pathogenesis.

 Shapes:

o Cocci (spherical): e.g., Staphylococcus aureus.

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o Bacilli (rod-shaped): e.g., Escherichia coli.

o Spirilla (spiral-shaped): e.g., Spirillum volutans.

 Examples of Roles:

o Beneficial: Nitrogen-fixing bacteria like Rhizobium.

o Pathogenic: Tuberculosis-causing Mycobacterium tuberculosis.

iii. Eukarya

 Characteristics:

o Eukaryotic organisms (cells with a nucleus and membrane-bound

organelles).

o Larger and more complex than prokaryotes.

o Includes fungi, protists, plants, and animals.

a. Fungi

 Characteristics:

o Eukaryotic organisms with cell walls made of chitin.

o Can be unicellular (yeasts) or multicellular (molds, mushrooms).

o Heterotrophic: Obtain nutrients by absorbing organic matter.

 Examples:

o Yeasts: Saccharomyces cerevisiae (used in baking and brewing).

o Molds: Aspergillus (important in biotechnology).

o Mushrooms: Agaricus (edible fungi).

 Roles:

o Decomposers in ecosystems.

o Pathogens in plants and animals.

o Producers of antibiotics like penicillin.

b. Protists

 Characteristics:

o Eukaryotic, mostly unicellular organisms.

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o Include protozoa (animal-like), algae (plant-like), and slime molds
(fungus-like).

o Found in aquatic and moist environments.

 Examples:

o Protozoa: Amoeba proteus (moves with pseudopodia).

o Algae: Chlamydomonas (photosynthetic).

o Slime Molds: Physarum (important in nutrient cycling).

B. Acellular Microorganisms

Acellular microorganisms lack cellular structures and are not considered

living organisms. They include viruses, viroids, and prions.

i. Viruses

 Characteristics:

o Acellular entities composed of genetic material (DNA or RNA)

enclosed in a protein coat (capsid).

o Some have an additional lipid envelope.

o Obligate intracellular parasites: Require a host cell to replicate.

 Examples:

o Influenza virus (causes flu).

o HIV (causes AIDS).

o Bacteriophages (viruses that infect bacteria).

 Structure:

o Simple: Nucleic acid and protein coat.

o Complex: Additional structures like tails (in bacteriophages).

ii. Viroids

 Characteristics:

o Small, circular RNA molecules without a protein coat.

o Infect plants and disrupt cellular processes.

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o Do not encode proteins but replicate within host cells.

 Example: Potato spindle tuber viroid (PSTVd).

iii. Prions

 Characteristics:

o Infectious protein particles that lack genetic material.

o Cause neurodegenerative diseases by inducing misfolding of

normal proteins.

o Resistant to heat and chemical treatments.

 Examples:

o Creutzfeldt-Jakob Disease (CJD) in humans.

o Bovine Spongiform Encephalopathy (BSE or mad cow disease) in

cattle.

Summary

Microbial diversity encompasses both cellular and acellular forms. Cellular

microorganisms include members of the three domains (Archaea, Bacteria,
and Eukarya), while acellular microbes (viruses, viroids, and prions) are
simpler entities that require host cells to carry out biological functions. This
diversity reflects the immense adaptability and ecological significance of
microbes in nature.

V. Anatomy of Cellular Microorganisms

The anatomy of cellular microorganisms refers to the structural components
and features that define their form and function. These structures determine
how they interact with their environment, obtain nutrients, and reproduce.

A. Archaea and Bacteria

1. Shared Features (Prokaryotic Structure)

 Cell Shape:

o Cocci (spherical), Bacilli (rod-shaped), Spirilla (spiral), Vibrio

(comma-shaped), and Pleomorphic (variable shape).
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 Cell Wall:

o Provides structural support and protection.

o Bacteria: Cell wall contains peptidoglycan (Gram-positive: thick

layer; Gram-negative: thin layer with an outer membrane).

o Archaea: Cell wall lacks peptidoglycan; may contain

pseudopeptidoglycan or protein layers.

 Plasma Membrane:

o Semi-permeable lipid bilayer that regulates transport.

o Archaea: Ether-linked lipids; highly stable.

o Bacteria: Ester-linked lipids.

 Cytoplasm:

o Contains enzymes, nutrients, and genetic material.

o Includes ribosomes (70S) for protein synthesis.

 Nucleoid:

o Region containing circular DNA.

o No membrane-bound nucleus.

 Flagella:

o Hair-like structures used for motility.

o Powered by a rotary motor in bacteria.

 Pili/Fimbriae:

o Short, hair-like structures for attachment or conjugation (transfer

of DNA).

2. Unique Features of Archaea

 Thrive in extreme environments.

 Methanogenesis: Unique metabolic process.

 Membrane lipids with branched chains for stability.

B. Fungi
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1. General Features

 Eukaryotic organisms with diverse morphologies.

 Can be unicellular (yeasts) or multicellular (molds and mushrooms).

 Cell walls contain chitin (rigid polysaccharide).

 Heterotrophic: Absorb nutrients from organic matter.

2. Anatomy

 Hyphae:

o Filamentous structures in multicellular fungi.

o Form a network called mycelium.

 Spores:

o Reproductive structures dispersed for propagation.

 Cell Structure:

o Nucleus: Contains genetic material.

o Mitochondria: Sites of energy production.

o Vacuoles: Store nutrients and waste.

o Plasma Membrane: Phospholipid bilayer beneath the cell wall.

3. Example Structures

 Yeast (e.g., Saccharomyces cerevisiae): Oval-shaped single cells.

 Mold (e.g., Rhizopus): Long hyphae and sporangia for spore

production.

C. Algae

1. General Features

 Eukaryotic, photosynthetic organisms found in aquatic environments.

 Can be unicellular (e.g., Chlorella) or multicellular (e.g., kelp).

 Contain chlorophyll and other pigments for photosynthesis.

2. Anatomy
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 Cell Wall:

o Made of cellulose or silica (in diatoms).

 Chloroplasts:

o Contain pigments like chlorophyll a, b, or c for photosynthesis.

 Plasma Membrane:

o Regulates exchange of substances.

 Flagella (in motile species):

o Used for movement.

 Pyrenoids:

o Structures in chloroplasts that store starch.

3. Examples of Algae

 Unicellular: Chlamydomonas (motile, green algae).

 Multicellular: Macrocystis (giant kelp).

D. Protozoans

1. General Features

 Eukaryotic, unicellular organisms lacking cell walls.

 Found in aquatic and moist environments.

 Heterotrophic, feeding on bacteria or organic matter.

2. Anatomy

 Plasma Membrane:

o Flexible boundary surrounding the cytoplasm.

 Nucleus:

o Stores genetic material (some have a macronucleus and

micronucleus, e.g., Paramecium).

 Cytoplasm:
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o Contains organelles like mitochondria, food vacuoles, and
contractile vacuoles.

 Locomotory Structures:

o Pseudopodia: Temporary cytoplasmic extensions (e.g.,

Amoeba).

o Cilia: Hair-like structures for movement and feeding (e.g.,

Paramecium).

o Flagella: Long, whip-like structures for swimming (e.g., Giardia).

3. Life Cycle Stages

 Trophozoite: Active, feeding stage.

 Cyst: Dormant, resistant stage for survival in harsh conditions.

Comparison of Cellular Microorganisms

Feature Archaea Bacteria Fungi Algae Protozoans

Cell Type Prokaryotic Prokaryotic Eukaryotic Eukaryotic Eukaryotic

Pseudopeptidoglyc Peptidoglyc Cellulose or

Cell Wall Chitin Absent
an an silica

Locomotio Flagella Pseudopodia,

Flagella (distinct) Flagella Non-motile
n (some) cilia, flagella

Metabolis Heterotroph Photosynthet

Extremophiles Varied Heterotrophic
m ic ic

VI. Microbial Growth Requirements

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Microbial growth refers to an increase in the number of cells rather than their
size. Microorganisms require specific nutrients and environmental conditions
to grow and reproduce effectively. This section explores the key factors
influencing microbial growth.

A. Microbial Nutrition

Microorganisms require nutrients to build cellular components, produce

energy, and carry out metabolic activities. These nutrients can be classified
into two main groups:

1. Macronutrients

 Required in large amounts for cell structure and metabolism.

 Include:

o Carbon (C): Building block for organic molecules (proteins,

lipids, carbohydrates).

o Nitrogen (N): Essential for proteins, nucleic acids, and ATP.

o Phosphorus (P): Needed for nucleic acids, ATP, and membrane

phospholipids.

o Sulfur (S): Component of amino acids (cysteine and

methionine) and vitamins.

o Hydrogen (H), Oxygen (O): Found in water and organic

molecules.

2. Micronutrients

 Required in trace amounts for enzyme function and structural stability.

 Include elements like iron (Fe), magnesium (Mg), zinc (Zn), copper
(Cu), and manganese (Mn).

3. Types of Nutritional Modes

 Autotrophs:

o Obtain carbon from carbon dioxide (CO₂).

o Examples: Photosynthetic bacteria like Cyanobacteria.

 Heterotrophs:
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o Obtain carbon from organic compounds.

o Examples: Most fungi and protozoa.

 Phototrophs:

o Use light as an energy source.

o Examples: Algae, photosynthetic bacteria.

 Chemotrophs:

o Use chemical compounds (organic or inorganic) as energy

sources.

o Examples: Sulfur-oxidizing bacteria.

B. Physical Factors Affecting Growth

Microbial growth is influenced by physical environmental factors. Each

microorganism has an optimal range for these factors.

1. Temperature

 Microorganisms are categorized based on their temperature

preferences:

o Psychrophiles: Thrive at low temperatures (-5°C to 20°C).

o Mesophiles: Grow best at moderate temperatures (20°C to

45°C). Most pathogens are mesophiles.

o Thermophiles: Thrive at high temperatures (45°C to 80°C).

o Hyperthermophiles: Grow at extremely high temperatures

(above 80°C).

2. pH

 Microorganisms have specific pH ranges:

o Acidophiles: Thrive in acidic environments (pH < 6).

o Neutrophiles: Grow in neutral pH environments (pH 6–8).

o Alkaliphiles: Thrive in alkaline environments (pH > 8).

3. Oxygen Requirements
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 Microorganisms are classified based on their oxygen needs:

o Obligate Aerobes: Require oxygen for growth.

o Obligate Anaerobes: Cannot tolerate oxygen.

o Facultative Anaerobes: Can grow with or without oxygen.

o Microaerophiles: Require low levels of oxygen.

o Aerotolerant Anaerobes: Do not use oxygen but can survive in

its presence.

4. Osmotic Pressure

 Microorganisms adapt to varying salt concentrations:

o Halophiles: Thrive in high salt environments.

o Non-halophiles: Prefer low salt conditions.

o Osmotolerant: Can grow across a range of salt concentrations.

5. Light

 Light influences the growth of photosynthetic microorganisms (e.g.,

algae).

6. Water Availability

 Water is essential for microbial metabolism. Dehydration or extreme

desiccation can inhibit growth.

C. Growth and Reproduction

Microbial growth involves both cell division and population increase. The rate
of growth depends on nutrient availability and environmental conditions.

1. Growth Phases in Bacteria (Batch Culture)

 Lag Phase:

o Period of adaptation; cells prepare for division.

 Log (Exponential) Phase:

o Rapid cell division; maximum growth rate.

 Stationary Phase:
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o Growth rate slows as nutrients are depleted and waste
accumulates.

 Death Phase:

o Cells die due to unfavorable conditions.

2. Reproduction

 Bacteria: Reproduce asexually through binary fission (one cell divides

into two identical daughter cells).

 Fungi:

o Yeasts: Budding (e.g., Saccharomyces cerevisiae).

o Molds: Spore formation (e.g., Penicillium).

 Algae and Protozoa:

o Asexual reproduction (binary fission, budding).

o Some reproduce sexually under specific conditions.

 Archaea: Binary fission, budding, or fragmentation.

VII. Microbial Metabolism

Microbial metabolism refers to the chemical processes that occur within
microorganisms to sustain life. These processes involve the conversion of
nutrients into energy and cellular components. Metabolism is divided into
two types: catabolism (breaking down molecules to produce energy) and
anabolism (building complex molecules using energy).

A. Aerobic Metabolism

Aerobic metabolism occurs in the presence of oxygen and is a highly efficient

way of producing energy.

1. Overview

 Involves the complete oxidation of organic molecules like glucose.

 Generates energy in the form of ATP through three main stages:

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o Glycolysis: Breaks down glucose into pyruvate, producing ATP
and NADH.

o Krebs Cycle (Citric Acid Cycle): Pyruvate is further oxidized,

generating NADH and FADH₂.

o Electron Transport Chain (ETC): NADH and FADH₂ donate

electrons to the ETC, where oxygen acts as the final electron
acceptor, forming water.

2. Energy Yield

 Glucose + Oxygen → Carbon Dioxide + Water + ATP

 Produces up to 38 ATP per glucose molecule in prokaryotes (slightly

less in eukaryotes).

3. Microorganisms

 Many bacteria (e.g., Escherichia coli under aerobic conditions) and

fungi use aerobic metabolism.

B. Anaerobic Metabolism

Anaerobic metabolism occurs in the absence of oxygen. Microorganisms can

use alternative electron acceptors or rely on fermentation.

i. Anaerobic Respiration

 Definition:

o Similar to aerobic respiration but uses a molecule other than

oxygen as the final electron acceptor (e.g., nitrate, sulfate, or
carbon dioxide).

 Examples of Electron Acceptors:

o Nitrate (NO₃⁻): Reduced to nitrogen gas (N₂).

o Sulfate (SO₄²⁻): Reduced to hydrogen sulfide (H₂S).

o Carbon Dioxide (CO₂): Reduced to methane (CH₄) by

methanogens.

 Energy Yield:
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o Less ATP than aerobic respiration because alternative electron
acceptors are less efficient.

 Examples of Microorganisms:

o Pseudomonas (denitrification).

o Desulfovibrio (sulfate reduction).

o Methanogens (methane production in Archaea).

ii. Fermentation

 Definition:

o A metabolic process that converts organic molecules (e.g.,

glucose) into energy without using an electron transport chain.

 Pathway:

o Begins with glycolysis, producing pyruvate.

o Pyruvate is then converted into various end products depending

on the microorganism.

 Energy Yield:

o Produces only 2 ATP per glucose molecule.

 End Products:

o Lactic Acid Fermentation: Produces lactic acid (e.g., in

Lactobacillus and muscle cells).

o Alcoholic Fermentation: Produces ethanol and carbon dioxide

(e.g., in Saccharomyces cerevisiae).

o Mixed Acid Fermentation: Produces a mixture of acids (e.g., in

Escherichia coli).

 Applications:

o Used in food production (yogurt, bread, beer, wine).

C. Photosynthesis
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Photosynthesis is the process by which certain microorganisms use light
energy to synthesize organic molecules (e.g., glucose) from carbon dioxide
and water.

1. Types of Photosynthetic Microorganisms

 Cyanobacteria: Perform oxygenic photosynthesis (produce oxygen).

 Purple and Green Sulfur Bacteria: Perform anoxygenic

photosynthesis (do not produce oxygen).

2. Process of Photosynthesis

 Light-Dependent Reactions:

o Occur in the thylakoid membranes or equivalents.

o Light energy is captured by pigments like chlorophyll (oxygenic)

or bacteriochlorophyll (anoxygenic).

o This energy drives the production of ATP and NADPH.

 Light-Independent Reactions (Calvin Cycle):

o Use ATP and NADPH to fix carbon dioxide into organic molecules
like glucose.

3. Oxygenic vs. Anoxygenic Photosynthesis

Oxygenic
Feature Anoxygenic Photosynthesis
Photosynthesis

Pigments Chlorophyll Bacteriochlorophyll

Hydrogen sulfide (H₂S) or other

Electron Donor Water (H₂O)
donors

By-Products Oxygen (O₂) Sulfur or other compounds

Example Cyanobacteria, algae,

Purple and green sulfur bacteria
Organisms plants
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VIII. Microbial Genetics

Microbial genetics studies how microorganisms inherit, express, and transfer
genetic material. This knowledge underpins advancements in biotechnology,
medicine, and environmental sciences.

A. Structures of DNA, RNA, and Proteins

1. DNA (Deoxyribonucleic Acid)

 Structure:

o Double helix of two complementary strands.

o Composed of nucleotides with three components:

 Deoxyribose Sugar

 Phosphate Group

 Nitrogenous Bases: Adenine (A), Thymine (T), Cytosine

(C), Guanine (G).

o Base pairing: A-T and G-C via hydrogen bonds.

 Function:

o Stores genetic information.

o Template for replication and transcription.

2. RNA (Ribonucleic Acid)

 Structure:

o Single-stranded molecule.

o Contains ribose sugar and nitrogenous bases: Adenine (A), Uracil

(U), Cytosine (C), Guanine (G).

o Base pairing: A-U and G-C (when applicable).

 Types and Functions:

o mRNA (Messenger RNA): Carries genetic code from DNA to

ribosomes.
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o tRNA (Transfer RNA): Brings amino acids to ribosomes for
protein synthesis.

o rRNA (Ribosomal RNA): Structural and functional component

of ribosomes.

3. Proteins

 Structure:

o Polymers of amino acids linked by peptide bonds.

o Have four structural levels:

 Primary (sequence of amino acids).

 Secondary (alpha helices and beta sheets).

 Tertiary (3D folding).

 Quaternary (interaction of multiple polypeptides).

 Function:

o Enzymes, structural components, signaling, and transport.

B. Central Dogma

The Central Dogma describes the flow of genetic information in cells:

DNA → RNA → Protein

1. Replication: DNA copies itself during cell division.

2. Transcription: DNA is transcribed into mRNA.

3. Translation: mRNA is translated into a protein using ribosomes, tRNA,

and amino acids.

C. Genetic Variation

i. Mutation

 Definition: Permanent changes in the DNA sequence.

 Types of Mutations:

o Point Mutation: Single nucleotide change (e.g., substitution).

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o Insertion/Deletion: Addition or removal of nucleotides.

o Frameshift Mutation: Alters the reading frame of the genetic

code.

 Effects:

o Silent (no change in protein function).

o Missense (changes one amino acid).

o Nonsense (introduces a stop codon).

ii. Genetic Recombination

 Definition: Exchange of genetic material between DNA molecules,

leading to new combinations of genes.

 Mechanisms:

o Homologous Recombination: Exchange between similar

sequences.

o Site-Specific Recombination: Exchange at specific DNA sites.

iii. Modes of Gene Transfer

 Transformation: Uptake of naked DNA from the environment.

 Conjugation: Direct transfer of DNA between bacteria through a pilus.

 Transduction: Transfer of DNA by bacteriophages (viruses infecting

bacteria).

D. Epigenetic Factors

 Definition: Heritable changes in gene expression without altering the

DNA sequence.

 Mechanisms in Microorganisms:

o DNA Methylation: Addition of methyl groups to DNA, affecting

gene activity.

o Histone Modifications (in eukaryotes): Changes in chromatin

structure influencing gene accessibility.

 Functions:
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o Regulate gene expression.

o Respond to environmental changes.

E. Applications in Biotechnology

Microbial genetics has revolutionized biotechnology with applications in

medicine, agriculture, and industry.

1. Recombinant DNA Technology:

o Inserting foreign genes into microorganisms for protein

production (e.g., insulin synthesis using Escherichia coli).

2. CRISPR-Cas Systems:

o Gene editing tool derived from microbial immune systems.

o Used for precise genome modifications.

3. Gene Therapy:

o Delivering therapeutic genes into humans using viral vectors.

4. Bioremediation:

o Engineering microbes to degrade pollutants.

5. Industrial Applications:

o Production of biofuels, enzymes, and food additives.

6. Agriculture:

o Development of genetically modified organisms (GMOs) for pest

resistance and improved yields.

IX. Acellular Infectious Agents

Acellular infectious agents are non-living entities that rely on host cells for
replication and survival. Unlike cellular organisms, they lack the structures
necessary for independent metabolism or reproduction. These agents include
viruses, viroids, and prions.
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A. Viruses

1. Characteristics

 Structure:

o Nucleic Acid Core: Either DNA or RNA (single-stranded or

double-stranded).

o Capsid: Protein coat that encloses the nucleic acid, made of

subunits called capsomeres.

o Envelope (Optional): Lipid layer derived from the host cell

membrane; contains viral proteins for host recognition.

 Size: Typically 20–300 nanometers.

 Host Specificity: Infect specific hosts or tissues within a host,

determined by surface proteins.

2. Classification

 Based on:

o Type of nucleic acid (DNA or RNA).

o Capsid symmetry (icosahedral, helical, or complex).

o Presence of an envelope.

o Mode of replication.

3. Life Cycle

 Lytic Cycle:

o Virus enters the host, replicates, and causes cell lysis to release
new virions.

 Lysogenic Cycle:

o Viral DNA integrates into the host genome (as a prophage) and
replicates with the host without causing immediate damage.

 Examples:

o Influenza virus, HIV (Human Immunodeficiency Virus),

bacteriophages (viruses that infect bacteria).
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4. Medical Importance

 Causes diseases such as the common cold, influenza, COVID-19, and

AIDS.

 Basis for vaccines and antiviral therapies.

B. Viroids

1. Characteristics

 Smallest infectious agents.

 Structure:

o Composed of short, circular, single-stranded RNA.

o No protein coat (capsid).

 Size: ~250–400 nucleotides.

 Host Range: Infects plants, causing significant agricultural damage.

2. Mechanism of Action

 The RNA interacts with the host's cellular machinery, disrupting normal
functions and leading to disease.

 Does not encode proteins but may affect gene expression.

3. Examples

 Potato Spindle Tuber Viroid (PSTVd): Affects potato and tomato

crops.

 Coconut Cadang-Cadang Viroid: Infects coconut palms.

4. Importance

 Significant economic impact in agriculture.

 Studied for RNA biology and gene expression.

C. Prions

1. Characteristics

 Infectious proteins that lack nucleic acids (DNA or RNA).

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 Structure:

o Misfolded versions of normal cellular proteins.

o Resistant to heat, radiation, and chemical agents.

 Host Range: Mainly affects mammals.

2. Mechanism of Action

 Prions induce normal proteins in the host to misfold into the prion form,
leading to accumulation in neural tissues.

 Causes progressive neurodegenerative diseases.

3. Examples of Prion Diseases

 Human Diseases:

o Creutzfeldt-Jakob Disease (CJD).

o Kuru (linked to cannibalism in specific cultures).

 Animal Diseases:

o Bovine Spongiform Encephalopathy (BSE or “mad cow disease”).

o Scrapie (in sheep).

4. Importance

 Challenged traditional concepts of infectious agents by showing that

proteins alone can transmit disease.

 Highlighted the role of protein misfolding in neurodegenerative

diseases.

XI. Control and Destruction of

Microorganisms
The control and destruction of microorganisms are essential in microbiology,
medicine, food safety, and industry. Various physical and chemical agents
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are used to eliminate or inhibit microbial growth, preventing infections,
spoilage, and contamination.

A. Physical Agents

Physical methods of microbial control rely on environmental factors such as

temperature, pressure, and radiation to kill or inhibit microorganisms.

1. Heat

 Moist Heat: More effective than dry heat due to water’s ability to
transfer heat efficiently.

o Boiling (100°C, 10–30 minutes): Kills most bacteria, fungi,

and viruses but does not destroy endospores.

o Autoclaving (121°C, 15 psi, 15–30 minutes): Uses

pressurized steam to achieve sterilization, killing even bacterial
endospores.

o Pasteurization (63°C for 30 minutes or 72°C for 15

seconds): Reduces microbial load in liquids like milk and juice
without destroying nutrients.

 Dry Heat: Less effective than moist heat but used for materials that
cannot be exposed to moisture.

o Hot Air Sterilization (160–170°C for 2 hours): Used for

glassware, metal instruments.

o Incineration (800–1000°C): Destroys contaminated materials

completely.

2. Low Temperature

 Refrigeration (0–4°C): Slows microbial growth but does not kill

microbes.

 Freezing (-20°C or lower): Stops microbial activity but may not kill
all microbes.

3. Filtration

 Physically removes microbes from liquids or air.

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 Membrane Filters (0.22 µm–0.45 µm pore size): Used for
sterilizing heat-sensitive liquids like vaccines and antibiotics.

 HEPA Filters (High-Efficiency Particulate Air): Remove airborne

microorganisms and particles.

4. Radiation

 Ultraviolet (UV) Radiation (260 nm): Damages DNA by causing

thymine dimers, used for surface sterilization.

 Ionizing Radiation (Gamma rays, X-rays): Generates free radicals

that destroy microbial DNA, used for sterilizing medical equipment and
food preservation.

5. Desiccation (Drying)

 Removal of water to prevent microbial growth. Used in food

preservation (e.g., dried fruits, meats).

6. Osmotic Pressure

 High concentrations of sugar or salt draw water out of microbial cells,

inhibiting growth (e.g., pickling, salting meats).

B. Chemical Agents

Chemical methods involve the use of disinfectants, antiseptics, sterilizing

agents, and chemotherapeutic drugs to control or destroy microorganisms.

i. Disinfectants, Antiseptics, and Sterilizing Agents

Type Definition Examples Uses

Chemicals used on Bleach, phenol, Cleaning floors,

Disinfectan
non-living surfaces hydrogen peroxide, tables, and medical
ts
to kill microbes. alcohols. equipment.

Chemicals applied Used for wound

Iodine, alcohol,
to living tissue to cleaning, hand
Antiseptics chlorhexidine,
reduce microbial sanitizers, and
hydrogen peroxide.
load. surgical scrubs.

Sterilizing Kill all forms of Ethylene oxide gas, Used for heat-
Agents microbial life, formaldehyde, sensitive medical
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Type Definition Examples Uses

instruments and
including spores. glutaraldehyde.
surgical tools.

Common Disinfectants and Their Mechanisms:

 Alcohols (Ethanol, Isopropanol, 60-90%) – Denature proteins and

disrupt membranes.

 Halogens (Chlorine, Iodine) – Oxidize cellular components, effective

against bacteria, viruses, and fungi.

 Phenolics (Lysol, Triclosan) – Disrupt cell membranes and proteins,

remain active on surfaces for long periods.

 Quaternary Ammonium Compounds (Quats) – Disrupt

membranes, effective against Gram-positive bacteria.

 Aldehydes (Formaldehyde, Glutaraldehyde) – Cross-link proteins

and nucleic acids, used in sterilization.

ii. Chemotherapeutic Agents

Chemotherapeutic agents are antimicrobial drugs used to treat infections

within a living organism.

1. Antibiotics

 Naturally produced by bacteria and fungi to inhibit or kill other

microorganisms.

 Examples:

o Penicillin – Inhibits cell wall synthesis.

o Tetracycline – Inhibits protein synthesis.

o Ciprofloxacin – Inhibits DNA replication.

2. Synthetic Antimicrobials

 Chemically synthesized drugs used to target microbes.

 Examples:

o Sulfonamides – Inhibit folic acid synthesis.

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o Fluoroquinolones – Inhibit bacterial DNA gyrase.

3. Antiviral Drugs

 Target viral replication processes.

 Examples:

o Acyclovir (for herpes virus).

o Oseltamivir (Tamiflu) – Inhibits influenza virus neuraminidase.

4. Antifungal Drugs

 Target fungal cell membranes or cell walls.

 Examples:

o Amphotericin B – Disrupts fungal cell membranes.

o Azoles (Fluconazole) – Inhibits ergosterol synthesis.

5. Antiparasitic Drugs

 Used to treat protozoal and helminthic infections.

 Examples:

o Metronidazole – Treats amoebiasis and giardiasis.

o Mebendazole – Treats helminth infections.

XII. Microbial Applications in

Environmental Science
Microorganisms play essential roles in maintaining ecological balance and
sustainability. They contribute significantly to geochemical cycles, waste
degradation, and bioremediation processes. These applications are
crucial in agriculture, industry, and environmental conservation.

A. Role in Geochemical Cycles

Microbes drive the cycling of essential elements such as carbon, nitrogen,

sulfur, and phosphorus. These cycles are fundamental to life on Earth, as
they regulate the availability of nutrients in ecosystems.
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1. Carbon Cycle

Microorganisms play a crucial role in carbon cycling by facilitating the

transformation of carbon compounds.

 Photosynthetic Microbes (Carbon Fixation): Cyanobacteria, algae,

and some bacteria (e.g., Prochlorococcus) fix atmospheric CO₂ into
organic matter through photosynthesis.

 Decomposers: Bacteria and fungi break down dead organic matter,

releasing CO₂ through respiration.

 Methanogens (Methane Production): Archaea like

Methanobacterium convert organic matter into methane (CH₄) in
anaerobic environments (e.g., swamps, rice paddies).

 Methanotrophs (Methane Utilization): Bacteria like Methylococcus

consume methane and convert it into CO₂, reducing greenhouse gas
emissions.

2. Nitrogen Cycle

Microorganisms help convert nitrogen between different chemical forms,

making it available for plants and animals.

 Nitrogen Fixation: Bacteria like Rhizobium (symbiotic with legumes)

and Azotobacter (free-living) convert atmospheric nitrogen (N₂) into
ammonia (NH₃), which plants can use.

 Nitrification: Nitrosomonas bacteria convert ammonia into nitrites

(NO₂⁻), while Nitrobacter converts nitrites into nitrates (NO₃⁻), a form
usable by plants.

 Denitrification: Bacteria such as Pseudomonas denitrificans reduce

nitrates back to nitrogen gas (N₂), returning it to the atmosphere.

3. Sulfur Cycle

 Sulfur Oxidation: Thiobacillus and Beggiatoa convert hydrogen

sulfide (H₂S) into sulfuric acid (H₂SO₄).

 Sulfur Reduction: Sulfate-reducing bacteria (SRB) like Desulfovibrio

reduce sulfates (SO₄²⁻) to hydrogen sulfide (H₂S).

4. Phosphorus Cycle
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 Phosphate Solubilization: Bacteria such as Bacillus and
Pseudomonas help dissolve insoluble phosphates, making phosphorus
available for plants.

B. Role in Biodegradation of Solid Wastes and Liquid Wastes

Microorganisms are used in waste management to degrade organic matter

and detoxify pollutants.

1. Solid Waste Biodegradation

Solid waste includes organic and synthetic materials. Microbial degradation is

used in composting, bioremediation, and waste recycling.

a. Composting

 Uses microorganisms such as bacteria (Bacillus, Pseudomonas) and

fungi (Aspergillus, Penicillium) to break down organic waste (food
scraps, leaves, manure) into nutrient-rich compost.

 Produces humus, which improves soil fertility.

b. Plastic Biodegradation

 Some microbes degrade plastics and synthetic polymers.

 Ideonella sakaiensis can break down polyethylene terephthalate (PET),

a common plastic.

 Pseudomonas putida degrades styrene and other industrial pollutants.

c. Landfill Bioremediation

 Microbes help in the degradation of landfill waste by breaking down

organic materials into methane (biogas) and carbon dioxide.

 Methanogenic archaea convert waste into usable energy.

2. Liquid Waste Biodegradation

Liquid waste includes industrial effluents, sewage, and oil spills. Microbial
treatments help remove contaminants.

a. Wastewater Treatment

Microorganisms play a key role in sewage and industrial wastewater

treatment.
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 Primary Treatment: Settling and removal of solids.

 Secondary Treatment (Biological Treatment):

o Aerobic Digestion: Bacteria like Nitrosomonas and Nitrobacter

break down organic matter.

o Activated Sludge Process: Microbial communities degrade

organic waste.

o Trickling Filters: Biofilms of bacteria break down contaminants.

 Tertiary Treatment: Further purification using algae and microbial

filtration.

b. Bioremediation of Oil Spills

Certain microbes degrade petroleum hydrocarbons.

 Alcanivorax and Pseudomonas species break down crude oil into non-
toxic compounds.

 Used in cleaning oil spills in marine environments.

c. Heavy Metal Bioremediation

 Some bacteria can remove toxic heavy metals from wastewater.

 Pseudomonas and Shewanella reduce toxic metals like arsenic, lead,

and mercury into less harmful forms.

Basic Microbiology – Laboratory Notes

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I. Microscopy
Microscopy is the technique used to observe microorganisms and other small
structures that are invisible to the naked eye. It is an essential tool in
microbiology for studying cell structure, morphology, and motility.

A. Types of Microscopes

1. Light Microscopes (Optical Microscopes)

Use visible light to magnify specimens.

 Bright-Field Microscope:

o Most commonly used in microbiology laboratories.

o Produces a dark image against a bright background.

o Used for stained and unstained specimens.

 Dark-Field Microscope:

o Produces a bright image of the specimen against a dark

background.

o Used for observing live, unstained specimens (e.g., Treponema

pallidum, the syphilis-causing bacterium).

 Phase-Contrast Microscope:

o Enhances contrast by detecting differences in refractive index.

o Used for observing live, unstained cells and internal structures.

 Fluorescence Microscope:

o Uses ultraviolet (UV) light to excite fluorescent dyes attached to

the specimen.

o Used in immunofluorescence techniques to detect specific

microbes.

2. Electron Microscopes

Use beams of electrons instead of light, providing much higher resolution.

 Transmission Electron Microscope (TEM):

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o Produces highly detailed images of thin sections of specimens.

o Used to study internal structures of cells and viruses.

 Scanning Electron Microscope (SEM):

o Produces three-dimensional images of the specimen’s surface.

o Used to study the morphology of microorganisms.

B. Parts of a Light Microscope and Their Functions

Part Function

Ocular Lens
Magnifies the image (typically 10×).
(Eyepiece)

Provide different levels of magnification (4×, 10×, 40×,

Objective Lenses
100× oil immersion).

Stage Holds the slide in place.

Condenser Focuses light onto the specimen.

Controls the amount of light passing through the

Diaphragm (Iris)
specimen.

Coarse
Moves the stage up and down for rough focusing.
Adjustment Knob

Fine Adjustment
Provides fine focusing for a sharp image.
Knob

Light Source Provides illumination for viewing the specimen.

C. Magnification and Resolution

 Magnification: The process of enlarging an image. It is the product of

the ocular lens and objective lens magnification.

o Example: If the ocular lens is 10× and the objective lens is 40×,
the total magnification is 400×.

 Resolution (Resolving Power): The ability of the microscope to

distinguish two closely spaced objects as separate.
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o The higher the resolution, the clearer the image.

o Electron microscopes have much higher resolution than light

microscopes.

D. Microscopy Techniques in Microbiology

1. Staining Techniques

Microorganisms are usually colorless, so stains are used to enhance contrast.

 Simple Staining: Uses a single stain (e.g., methylene blue) to

highlight cell shape and size.

 Differential Staining: Uses multiple stains to distinguish between

different types of bacteria.

o Gram Staining: Differentiates bacteria into Gram-positive

(purple) and Gram-negative (pink).

o Acid-Fast Staining: Identifies acid-fast bacteria like

Mycobacterium tuberculosis.

 Special Stains:

o Capsule Stain: Highlights bacterial capsules.

o Endospore Stain: Identifies bacterial spores.

2. Wet Mount and Hanging Drop Techniques

Used to observe living microorganisms and their motility.

 Wet Mount: A drop of liquid culture is placed on a slide and covered

with a coverslip.

 Hanging Drop Method: A drop of culture is suspended on a coverslip

over a concave slide, allowing observation of motility.

II. Aseptic Techniques

Aseptic techniques are essential procedures in microbiology that prevent
contamination of cultures, specimens, and laboratory environments. These
techniques are crucial for obtaining accurate experimental results and
ensuring the safety of both researchers and the public.
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A. Study of Microorganisms from the Environment

Microorganisms are present everywhere, including the human body, air,

water, and various surfaces. Studying these microbes helps understand their
role in health, disease, and the environment.

i. Normal Body Flora (Microbiota)

Normal body flora consists of microorganisms that naturally inhabit the

human body without causing disease under normal conditions. They play a
role in digestion, immunity, and protection against harmful microbes.

Examples of Normal Flora in Different Body Sites:

Body Site Common Microorganisms

Staphylococcus epidermidis,
Skin
Propionibacterium acnes

Mouth Streptococcus mutans, Lactobacillus

Gastrointestinal
Escherichia coli, Bacteroides, Lactobacillus
Tract

Streptococcus pneumoniae, Haemophilus

Respiratory Tract
influenzae

Urogenital Tract Lactobacillus (in females), Corynebacterium

ii. Common Items

Everyday objects are covered with microbes, including pathogenic and non-
pathogenic species. Studying microorganisms from these items can help
assess hygiene and contamination risks.

Common Items Studied for Microbial Contamination:

 Air: Bacteria and fungal spores (Bacillus, Aspergillus).

 Water: Bacteria such as E. coli (indicator of fecal contamination).

 Surfaces (Tables, Phones, Doorknobs): Staphylococcus aureus,

Pseudomonas spp.

 Food: Salmonella, Listeria, Bacillus cereus (foodborne pathogens).

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 Currency (Money, Coins): Staphylococcus, Escherichia coli,
Klebsiella (potential pathogens).

B. Key Aseptic Techniques in Microbiology

1. Hand Hygiene and Personal Protective Equipment (PPE)

 Proper handwashing with soap or alcohol-based sanitizers.

 Wearing gloves, lab coats, and masks to prevent contamination.

2. Sterilization and Disinfection

 Sterilization: Kills all forms of microbial life (e.g., autoclaving, dry

heat, filtration).

 Disinfection: Reduces microbial load on surfaces (e.g., alcohol,

bleach, UV light).

3. Aseptic Handling of Cultures and Samples

 Using sterile instruments (loops, pipettes) to handle microbial cultures.

 Flaming the mouth of culture tubes before and after transfer.

4. Proper Disposal of Microbial Waste

 Contaminated materials must be discarded in biohazard waste

containers.

 Autoclaving or chemical disinfection before disposal.

5. Working in a Sterile Environment

 Performing microbial transfers in a laminar flow hood.

 Avoiding unnecessary exposure of cultures to air.

III. Culture Media

Culture media are nutrient-rich substances used to support the growth of
microorganisms in laboratory conditions. Proper preparation and sterilization
of culture media are essential to ensure uncontaminated and reliable
microbial cultures.
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A. Preparation of Culture Media

1. Components of Culture Media

Culture media typically contain the following components:

 Water – Serves as a solvent for all nutrients.

 Carbon Source – Provides energy (e.g., glucose, lactose).

 Nitrogen Source – Required for protein synthesis (e.g., peptones,

amino acids).

 Minerals and Salts – Essential for enzymatic activities (e.g., NaCl,

Mg²⁺, K⁺).

 Growth Factors – Some microbes require vitamins or blood

components.

 Agar (for Solid Media) – A polysaccharide from red algae used to

solidify the medium.

2. Types of Culture Media Based on Physical State

Type Description Examples

Liquid
No solidifying agent; used for growing Nutrient Broth, Tryptic
(Broth)
large numbers of microbes. Soy Broth (TSB)
Media

Semi-Solid Contains a small amount of agar (0.3- SIM (Sulfide Indole

Media 0.5%); used for motility tests. Motility) Medium

Contains 1.5-2% agar; used for Nutrient Agar,

Solid Media
isolating and growing colonies. MacConkey Agar

3. Types of Culture Media Based on Function

Type Function Examples

General Supports the growth of a

Nutrient Agar, Tryptic Soy Agar
Purpose wide variety of
(TSA)
Media microorganisms.

Contains inhibitors to MacConkey Agar (selects Gram-

Selective
suppress the growth of negative), Mannitol Salt Agar
Media
unwanted microbes. (selects Staphylococcus)
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Type Function Examples

Distinguishes bacteria
Differential Blood Agar (hemolysis detection),
based on metabolic
Media Eosin Methylene Blue (EMB)
properties.

Contains additional
Enriched
nutrients to support Blood Agar, Chocolate Agar
Media
fastidious organisms.

Transport Maintains the viability of

Stuart’s Medium, Amies Medium
Media microbes during transport.

B. Sterilization of Culture Media

Sterilization is necessary to eliminate contaminants before media use.

1. Methods of Sterilization

Method Principle Application

Uses steam under pressure Most culture media,

Autoclaving (121°C, 15 psi, 15-20 minutes) to glassware, biohazard
kill all microorganisms. waste

Passes liquid through a membrane Used for heat-sensitive

Filtration filter (0.22 µm) to remove solutions (e.g., antibiotic-
bacteria. containing media)

Uses high temperatures (160-

Dry Heat Glassware, metal
170°C for 2 hours) to destroy
Sterilization instruments
microbes.

Radiation Used for plastic petri

Destroys DNA of microorganisms.
(UV, Gamma) dishes, syringes

2. Aseptic Dispensing of Sterile Media

 Pouring sterilized agar into petri dishes inside a laminar flow hood.

 Cooling agar to ~45-50°C before pouring to prevent condensation.

 Avoiding exposure of media to air for extended periods.

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IV. Microscopic Examination of

Microorganisms
Microscopic examination is essential in microbiology for identifying and
studying microorganisms. It involves different techniques, including wet
mounts, hanging drop preparations, and various staining methods.

A. Wet Mount and Hanging Drop Techniques

These methods allow the observation of living microorganisms in their

natural state, helping to determine their motility, shape, and
arrangement.

1. Wet Mount Technique

 A simple and quick method where a drop of liquid culture or water

containing the specimen is placed on a slide and covered with a
coverslip.

 Advantages:

o Allows observation of living cells.

o Easy to prepare.

 Disadvantages:

o Cells dry out quickly.

o Bacteria are hard to see without staining.

2. Hanging Drop Technique

 A drop of culture is suspended from a coverslip over a concave slide.

 Used for observing motility of bacteria (e.g., Proteus, Pseudomonas).

 Advantages:

o Reduces drying, allowing longer observation.

o Shows bacterial movement clearly.

 Disadvantages:

o Requires a special slide with a concave depression.

Basic Microbiology Notes
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B. Staining Techniques

Staining improves contrast, making bacteria more visible under the

microscope. It also helps differentiate microorganisms based on their
structural characteristics.

i. Gram Stain (Differential Stain)

The Gram stain differentiates bacteria into Gram-positive (purple) and

Gram-negative (pink) based on cell wall composition.

Gram Staining Procedure:

1. Crystal violet (Primary stain) – stains all bacteria purple.

2. Iodine (Mordant) – binds with crystal violet to form a complex.

3. Alcohol (Decolorizer) – removes stain from Gram-negative bacteria.

4. Safranin (Counterstain) – stains Gram-negative bacteria pink.

Bacterial Color After

Cell Wall Composition Examples
Group Gram Stain

Gram- Staphylococcus
Thick peptidoglycan Purple
Positive aureus, Bacillus

Gram- Thin peptidoglycan + Escherichia coli,

Pink
Negative Outer membrane Pseudomonas

ii. Special Stains (For Specific Structures)

Some bacteria have unique structures that require specialized staining

techniques.

a. Spore Stain (Schaeffer-Fulton Method)

Used to detect endospores in bacteria like Bacillus and Clostridium.

 Primary Stain: Malachite green (stains spores green).

 Decolorizer: Water (removes stain from vegetative cells).

 Counterstain: Safranin (stains vegetative cells pink).

 Result:
Basic Microbiology Notes
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o Spores = Green

o Vegetative cells = Pink

b. Capsule Stain

Detects capsules, which are protective outer layers found in some bacteria
(Klebsiella pneumoniae, Streptococcus pneumoniae).

 Uses negative staining (e.g., India ink or Congo red).

 Capsules appear as a clear halo around cells against a dark

background.

c. Flagellar Stain

Used to visualize bacterial flagella, which are normally too thin to be seen.

 A mordant (e.g., tannic acid) thickens the flagella, making them visible
under a light microscope.

 Example of Flagellated Bacteria: Proteus vulgaris, Pseudomonas

aeruginosa

V. Factors Affecting Microbial Growth

Microbial growth is influenced by various physical and chemical factors.
Understanding these factors helps in controlling microbial populations in
different environments, including laboratories, industries, and medical
settings.

A. Temperature

Temperature affects microbial metabolism, enzyme activity, and membrane

stability. Microorganisms are classified based on their optimal temperature
for growth:

Optimal
Category Examples
Temperature

Polaromonas vacuolata (found in deep

Psychrophiles 0–15°C
oceans, glaciers)

Psychrotrophs 15–30°C Listeria monocytogenes (causes

Basic Microbiology Notes
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Optimal
Category Examples
Temperature

foodborne illness in refrigerated foods)

Escherichia coli, Staphylococcus aureus

Mesophiles 20–45°C
(common in human body)

Bacillus stearothermophilus (found in hot

Thermophiles 45–80°C
springs)

Hyperthermoph Thermococcus, Pyrolobus fumarii (found

80–120°C
iles in hydrothermal vents)

 High temperatures denature proteins and destroy cell membranes.

 Low temperatures slow metabolism and can cause ice crystal

formation, damaging cells.

B. pH

pH affects enzyme activity and cell membrane stability. Microorganisms have

specific pH ranges for optimal growth:

Optimal
Category Examples
pH

Acidophile Lactobacillus acidophilus (found in yogurt),

pH < 5.5
s Helicobacter pylori (causes stomach ulcers)

Neutrophil pH 6.5– Escherichia coli, Staphylococcus aureus (common

es 7.5 human pathogens)

Alkaliphile Bacillus alcalophilus (found in soda lakes, alkaline

pH > 8.5
s soils)

 Extreme pH values denature proteins and disrupt cell membranes.

 Most human pathogens are neutrophiles since body fluids are near
pH 7.4.

C. Oxygen (O₂) Requirement

Basic Microbiology Notes
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Microorganisms differ in their ability to use oxygen for metabolism. Oxygen
can be toxic due to the formation of reactive oxygen species (ROS) like
superoxide radicals (O₂⁻) and hydrogen peroxide (H₂O₂).

Category Oxygen Requirement Example

Obligate Require oxygen; use aerobic Mycobacterium

Aerobes respiration. tuberculosis

Can grow with or without oxygen;

Facultative Escherichia coli,
prefer aerobic respiration but can
Anaerobes Staphylococcus aureus
ferment.

Obligate Oxygen is toxic; use anaerobic

Clostridium botulinum
Anaerobes respiration or fermentation.

Aerotolerant Do not use oxygen but can

Lactobacillus
Anaerobes tolerate it.

Require low oxygen (2–10%) for

Microaerophiles Helicobacter pylori
growth.

 Obligate aerobes need oxygen for ATP production.

 Obligate anaerobes lack enzymes like superoxide dismutase (SOD)

and catalase, making oxygen toxic.

D. Desiccation (Drying)

Water is essential for microbial metabolism, so drying affects microbial

survival. Some microbes can withstand desiccation better than others:

Resistance to
Category Examples
Desiccation

Bacillus, Clostridium (endospores),

Highly Form spores or
Mycobacterium tuberculosis (waxy
Resistant protective structures.
cell wall)

Can survive dry

Moderately Staphylococcus aureus (thick cell
conditions for some
Resistant wall), Enterococcus
time.

Sensitive to Require moisture to Neisseria gonorrhoeae (causes

Basic Microbiology Notes
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Resistance to
Category Examples
Desiccation

gonorrhea), Treponema pallidum

Desiccation survive.
(causes syphilis)

 Endospores protect bacteria from extreme dryness.

 Mycobacteria have a waxy mycolic acid layer that reduces water loss.

E. Media Components

The composition of culture media influences microbial growth.

Component Function Examples

Carbon Provides energy and building

Glucose, lactose, peptone
Source blocks.

Nitrogen Required for protein and nucleic Ammonium salts, amino

Source acid synthesis. acids

Essential for enzyme activity and

Minerals Magnesium, potassium, iron
structure.

Growth Vitamins, blood, NAD (for

Needed by fastidious bacteria.
Factors Haemophilus)

Agar Solidifies media. Extracted from red algae

 Fastidious bacteria like Haemophilus influenzae require enriched

media (e.g., Chocolate Agar with NAD and hemin).

 Selective media contain inhibitors to suppress unwanted microbes.

VI. Antimicrobial Susceptibility Testing

(AST)
Antimicrobial Susceptibility Testing (AST) is used to determine the
effectiveness of antibiotics, antifungals, and other antimicrobial agents
against specific microorganisms. It is crucial in clinical microbiology to
guide proper treatment and prevent antimicrobial resistance (AMR).
Basic Microbiology Notes
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A. Common Methods of AST

Method Principle Example

Used in routine
Disk Diffusion
Antibiotic-impregnated disks create clinical labs to test
(Kirby-Bauer
zones of inhibition on an agar plate. bacterial
Test)
susceptibility.

Determines the minimum inhibitory Used for precise

Broth Dilution
concentration (MIC) and minimum quantification of
(MIC/MBC
bactericidal concentration (MBC) of antibiotic
Test)
an antibiotic. effectiveness.

E-test Uses a strip with a gradient of Often used for

(Epsilometer antibiotic concentrations to difficult-to-treat
Test) determine the MIC. infections.

Machines like VITEK or Phoenix Used in large

Automated
analyze microbial growth in response hospitals for rapid
Systems
to antibiotics. AST.

B. Interpretation of AST Results

 Susceptible (S): The microorganism is inhibited by the antibiotic.

 Intermediate (I): The antibiotic may be effective at higher doses.

 Resistant (R): The microorganism is not affected by the antibiotic.

VII. Food and Water Bacteriology

Food and water microbiology is essential for public health and safety,
ensuring that food and drinking water are free from pathogenic
microorganisms.

A. Common Tests in Food and Water Bacteriology

Test Purpose Example

Used to assess
Total Plate Count Measures the total number of viable
overall microbial
(TPC) bacteria in food/water.
load.
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Test Purpose Example

Detects Escherichia coli and related

Used in water
Coliform Test bacteria, indicating fecal
quality testing.
contamination.

Common in
Most Probable Estimates bacterial contamination in
drinking water
Number (MPN) water.
analysis.

Used in bottled
Heterotrophic Measures bacteria that grow in low-
water and food
Plate Count (HPC) nutrient conditions.
safety.

B. Common Foodborne Pathogens

 Salmonella – Found in poultry and eggs.

 Listeria monocytogenes – Can grow in refrigerated foods.

 Escherichia coli O157:H7 – Found in undercooked beef.

 Vibrio cholerae – Causes cholera in contaminated water.

VIII. Fermentation
Fermentation is the anaerobic conversion of carbohydrates into
alcohol, acids, or gases by microorganisms like bacteria and yeast. It is
widely used in food and beverage production.

A. Wine Making

Microorganism: Saccharomyces cerevisiae (yeast)

Process:

1. Crushing & Pressing: Grapes are crushed to extract juice.

2. Fermentation: Yeast converts sugars to ethanol and carbon

dioxide.

3. Aging & Bottling: The wine matures for better flavor.

B. Yogurt Production

Microorganisms: Lactobacillus bulgaricus and Streptococcus thermophilus

Process:
Basic Microbiology Notes
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1. Milk Pasteurization: Destroys unwanted microbes.

2. Fermentation: Lactic acid bacteria convert lactose into lactic acid,

thickening the milk.

3. Cooling & Storage: Yogurt is stored at cold temperatures to maintain

texture.

C. Kimchi or Sauerkraut Production

Microorganisms: Lactobacillus spp. (lactic acid bacteria)

Process:

1. Preparation: Cabbage or vegetables are mixed with salt and spices.

2. Fermentation: Lactic acid bacteria produce acid, preserving the food.

3. Storage: The product is kept under controlled conditions to develop

flavor.