Isolation and Characterization of Potential

Phyllosphere Bioinoculants from Coriandrum

sativum
Avinash K
Sri Shakthi Institute of Engineering and Technology
Suresh Vikram S
Sri Shakthi Institute of Engineering and Technology
Vivan K S
Sri Shakthi Institute of Engineering and Technology
Amaana Shirine S
Sri Shakthi Institute of Engineering and Technology
Gopinath B (  [email protected] )
Sri Shakthi Institute of Engineering and Technology

Research Article

Keywords: Bioinoculants, Phyllosphere, Coriandrum sativum, Sustainable

Posted Date: October 5th, 2023

DOI: https://doi.org/10.21203/rs.3.rs-3394427/v1

License:   This work is licensed under a Creative Commons Attribution 4.0 International License.
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Abstract
Coriandrum sativum, also known as Coriander, is an annual herb in the apiaceae family. It is a common
leguminous plant that grows abundantly in India. It has also been used for intercropping with short
irrigation intervals. This plant is well-known for its ability to thrive in low-phosphate and drought-prone
soils. This study aims to identify isolates from the phyllosphere region that have potential to be used as
bioinoculants. Phyllosphere sample was collected from the native farmland and neighbourhood and
screened for bioinoculants. A total of 98 bacterial strains were isolated and tested for production of
ammonia. The nitrogen fixing strains were chosen based on their ammonia production. The selected 28
isolates were tested for various PGP factors such as HCN production, IAA production and Phosphate
solubilization and also tested for extracellular enzymatic activities such as Amylase production, Protease
degradation, Gelatinase activity, Catalytic activity and Lipase production, based on the results, few
exhibited potent antagonistic activity against Colletotrichum capscici and Rhizoctonia solani. Based on
the results of extracellular activities and antagonistic assay, five potential strains were subjected to
germination study. One best PGPP was identified through amplification and sequencing of the 16S rRNA
gene. The research uncovered a previously unknown diversity of plant growth promoting bacteria with
enormous potential for use as a bioinoculant in agriculture. This study emphasises the importance of
Phyllosphere microbiota for plant growth, in addition to identifying promising candidates for
bioinoculation.

1. Introduction
Agriculture is a major field that has attained a very advanced level when compared to the last few
decades. Scientific developments have a vital role in the improvement of agriculture in many ways such
as mechanical inventions, irrigation techniques, chemical fertilizers, pesticides, insecticides, etc. In 2050,
the world would need a 65% higher production of food products when compared to the current market
production rate. However, that may be achieved by excessive input of chemical fertilizer, but the biological
and physicochemical characterization of soil may degrade eventually. Dumping more unbalanced
amounts of chemical fertilizer may increase the plant yield and productivity for a single crop cultivation
cycle, the non-absorbed chemical or synthetic fertilizer may remain in the soil and can acidify soil nature
(Sharma and Singhvi, 2017).

Current research studies have shown that the degradation of soil can be prevented and also restored by
the application of bio-fertilizer, which can be considered a greater and safer replacement for chemical
fertilizers. On the formation of biofertilizer, the plant symbiotic rhizosphere- and phyllosphere- bacteria
had shown plant growth-promoting activity. Based on traits, isolates were screened for antagonistic
activity among themselves and also with the plant to prevent any future accidents (Ghosh., 2004).
Microbes that were used in the formation of biofertilizer can be bacteria, fungi, algae, etc. Those were
generally described as bio-inoculants. Bio-inoculants were substances derived from microbes or live
microbes that could be applied to plants as an additive source of nutrients and could also increase the
plant growth parameters.
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Despite the importance of phyllosphere bacteria with different PGP characteristics in sustainable
agriculture, little is known about their impact on shaping plant performance traits. For better survival in
the phyllosphere, bacteria have developed mechanisms such as ultraviolet variation (UV) resistance,
biofilm formation, and aggregation.

Coriander (Coriandrum sativum L.), an annual herb, is commonly used for seasoning. Its plant seeds,
leaves, and roots are edible, with distinct flavors and applications. The flavor of the herb is light and
refreshing. Coriander can be used as a flavoring agent in various food preparations due to the perishable
nature of the leaves and seeds are used to increase the palatability of ripe fruits (seeds). Coriander is
used in cooking in its entirety, primarily for fresh leaves and ripe fruits. Coriander leaves have a different
flavor than coriander seeds, with citrus undertones (Kumar, J., et al, 2021). The Coriander plant is a rich
reservoir of micronutrients and nutritional elements and is luxurious in growth, so we decided to isolate
potential phyllobacteria from it. Coriander is a good source of tocopherol and vitamin K, low in saturated
fat but high in linoleic acid. Plant leaves are high in vitamins, while seeds are high in polyphenols and
essential oils.

In general, phyllosphere bacteria originate primarily in soil, seed, and air and adapt to life on leaf tissue,
where several factors such as environmental parameters shape their community composition. The
phyllosphere hosts a variety of bacterial isolates at the genus and species levels, depending on plant
genotype and climate. The capabilities of plant-microbe interactions in the phyllosphere can contribute to
our comprehension of how plants benefit from their microbial partners. As a result, having more
information on phyllosphere bacteria from various geographical locations in terms of their PGP traits
may make it possible to find qualified applicants for organic amendments (Stone et al., 2018).

The primary objectives of this study were to isolate and characterize the leaf-colonizing bacteria with the
potential for plant growth–promoting activities from Coriandrum sativum in a village region of
Coimbatore district, Tamil Nadu State, India. The bacteria were screened in vitro for PGP traits and the
most promising isolate was identified by sequencing of 16S rRNA genes. We also examined the
germination property of the selected isolate compared with the control under the greenhouse effect (Lau,
S.-E., et al, 2022).

2. Materials and methods

2.1. Isolation of Phyllobacteria:
Coriandrum sativum leaves were collected from agricultural fields, which were not subjected to any
chemical fertilizer or synthetic pesticides and herbicides. On employment to enrichment technique to
increase the papulation of symbiotic bacteria leaves were inoculated in nutrient broth for overnight. Serial
dilution plate method was adapted for isolating microorganisms from enriched broth culture. After
incubation for 24 hours, unique colonies were selected based on colour, shape, elusion and growth
(Schlegel and Jannasch, 1967).

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2.2. Characterization of PGPR:
2.2.1. Production of ammonia:
It was done by growing the isolates in peptone water at 30°C for 4 days. To each culture tube, 1mL of
Nessler’s reagent was added. Development of faint yellow colour indicates small amounts of ammonia
and deep yellow to brownish colour indicates maximum amount of ammonia production (Tsegave. Z et
al.,2019).

2.2.2. Determination of IAA:

Luria Bertani (LB) broth medium (25 mL) was used to enrich isolates in which 50 µg/mL tryptophan,
incubated for 24 h at 28°C on rotary shaker. Cultures were centrifuged at 10,000 rpm for 15 min with
addition of two drops of ortho phosphoric acid and 4 ml of Salkowsky reagent were mixed with
supernatant (2 mL). Development of pink colour was achieved by incubating for 25 min at room
temperature confirms qualitative IAA production (Tsegave. Z et al., 2019).

2.2.3. HCN Production:

A 4.4 g/l glycine amended King’s B medium was used to streak isolates. Whatman No.1 filter paper discs
were dipped in 0.5% picric acid in 2% sodium carbonate solution. On lid of each petriplate the discs were
placed and sealed. Plates were incubated for 4 days at 28 ± 2℃. Production of HCN was recorded from
colour change of the filter paper from deep yellow to orange and orange to brown (Bharucha U.D., et al,
2013).

2.3. Extra cellular enzyme production:

2.3.1. Phosphate solubilization:
To check phosphate solubilisation, isolates were streaked on Pikovaskya’s (Pikovaskya, 1948) agar
medium and incubated for 4 days at 28°C. Phosphate solubilising activity was confirmed by clearing
zones around the bacterial colony plates.

2.3.2. Catalase activity:

Catalase test was performed by growing bacterial culture for 48 hr on trypticase soy agar medium
followed by adding three to four drops of H2O2 on bacterial culture which was grown. The prescence of
effervescence indicates Catalase activity (Kumar, et al, 2012).
2.3.3. Protease Production:
Protease production qualitative assay was performed on sterile skim milk agar plates containing casein
5.0 g, Yeast extract 2.5 g, Glucose 1.0 g, Agar 15.0 g, Distilled water 1000 mL, Skim milk 7% was added as
inducer. Isolates were inoculated and zone of clearance around the colony was observed by incubation at
30⁰C and indicating the enzymatic degradation of protease.

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2.3.4. Production of lipase:
Nutrient agar amended with egg yolk was used to test the bacterial isolates. Saturated aqueous solution
of copper sulphate (CuSO4) was used to flood agar medium and kept for 10–15 min after 48 h of growth.
The excess reagent was poured off. Formation of greenish blue colour zones around the colony indicated
the production of lipase (Malleswari D and Bagyanarayana G., 2013).

2.3.5. Production of amylase (starch hydrolysis):

Amylase assay medium comprises Beef extract 3.0, Peptone 5.0, Soluble Starch 2.0, Agar 15.0, Distilled
water 1lit and inoculated isolates incubated at 30°C for 48 h., The plates were flooded with iodine
solution at the end of incubation period, kept for a minute and then poured off. Iodine reaction with starch
forms a blue colour compound. This blue colour fades rapidly. The production of amylase is indicated by
the colour less zone surrounding colonies (Malleswari D and Bagyanarayana G., 2013).

2.3.6. Gelatinase test:

The culture was inoculated into gelatine containing tubes followed by incubation for 5 to 7 days. Upon
refrigeration of the tubes for an hour confirms if gelatinase present, failing of liquid medium to solidify
upon refrigeration (Geetha K., et al, 2014).

2.4. Salt tolerance:

Isolates were tested for their salt tolerance ability by spot inoculating the isolates on nutrient agar plates
containing different concentrations of NaCl (2%, 4%, 6%, 8% and 10%), incubated at 32°C for 5 days.
Observe the growth (Geetha K., et al, 2014).

2.5. Antagonistic test:

Selected five potential strains were screened for antagonism against Colletotrichum capscici and
Rhizoctonia solani using potato dextrose agar (PDA) medium by using potato dextrose agar medium,
placing the pathogen and bioagent in opposite direction and allowing them to grow for 7 days (Geetha K.,
et al, 2014).

2.6. Germination test:

Green gram seeds were surface sterilized with 0.1% HgCl2 for 3 min, followed by successive washing with
sterile distilled water and then the water was decanted. The seeds were kept for 10 min in 48 h old
cultures. The seeds were shown on paper cup filled with sterilized sand collect from and incubated at
30℃ for 2–7 days. Seeds were treated with sterilized medium was treated as control. The root and shoot
lengths were recorded on 7th days (Geetha K., et al, 2014).

2.7. Morphological characterization of isolates:

The selected bacterial isolates were examined on Nutrient Agar plates for their morphological features.
After 3 days of incubation, different colony characteristics such as Gram reaction, shape, motility,
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elevation, surface, etc. were recorded (Mącik, M., et al, 2020).

2.8. Seed Germination and Seedling Growth:

Test Seed germination tests were carried out to determine the effect of inoculated rhizobacterial species
or strains on the rates of seed germination and seedling growth. For this, green gram seeds were used as
plant materials. Healthy seeds were surface sterilized with 70% alcohol for 3 min and followed with 1%
sodium hypochlorite for 5 minutes and rinsed 5 times with sterile distilled water. Five PGPR species or
strains were grown in nutrient broth on shaking incubator (180 rpm) at 30°C for 24 h. The surface-
sterilized seeds of green gram were inoculated in broth culture of the PGPR species or strain cultures for
30 min including normal water (C) as control. 25 inoculated seeds of each treatment were placed in
separate plastic pot filled with sterilized red soil and the pots were incubated at room temperature for 7
days. Percentages of germination and seedling growth were calculated and evaluated the effect of single
seed bacterization on a shoot and root growth of green gram crops. Vigor index of seedling was
measured at the end of the experiment according to the formula. Seed vigor index = (Mean shoot length +
Mean root length) × germination %.

2.9. Identification of bacterial strains using 16SrRNA gene

sequencing:
The DNA template was prepared by picking an individual colony of each bacterial strain, and
amplification of the 16S rRNA gene was carried out by the PCR. PCR amplification of DNA was performed
following Katsivela et al. (1999) using universal primers (27F: 5′́- ' AGAGTTTGATCTGGCTCAG-3′; 1492R:
5′́-TACGGTACCTTGTTACGACTT-3′́) in a reaction mixture (25 µL). The amplification program for the full-
length 16S rRNA gene consisted of an initial denaturing at 94°C for 2 min, followed by 30 cycles of
denaturation at 94°C for 2 min, primer annealing at 55°C for 1 min and primer extension at 72°C for 2
min, followed by a final extension at 72°C for 10 min, in a thermocycler. Amplified PCR products of the
16S ribosomal gene were separated on 1% agarose gel in 0.5× TE (Tris-EDTA) buffer containing 2 µL
ethidium bromide (20 mg/ml). The purification process for PCR product was done by using Montage PCR
Clean up kit (Millipore). Sequencing reactions were performed using a ABI PRISM® BigDyeTM Terminator
Cycle Sequencing Kits with AmpliTaq® DNA polymerase (FS enzyme) (Applied Biosystems). Single-pass
sequencing was performed on each template using below 16s rRNA universal primers. The fluorescent-
labeled fragments were purified from the unincorporated terminators with an ethanol precipitation
protocol. The samples were resuspended in distilled water and subjected to electrophoresis in an ABI
3730xl sequencer (Applied Biosystems).

The 16s rRNA sequence was blast using NCBI blast similarity search tool. The phylogeny analysis of
query sequence with the closely related sequence of blast results was performed followed by multiple
sequence alignment. The program MUSCLE 3.7 was used for multiple alignments of sequences (Edgar
2004). The resulting aligned sequences were cured using the program Gblocks 0.91b. This Gblocks
eliminates poorly aligned positions and divergent regions (removes alignment noise) (Talavera and
Castresana 2007). Finally, the program PhyML 3.0 aLRT was used for phylogeny analysis and HKY85 as

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Substitution model. PhyML was shown to be at least as accurate as other existing phylogeny programs
using simulated data, while being one order of magnitude faster. PhyML was shown to be at least as
accurate as other existing phylogeny programs using simulated data, while being one order of magnitude
faster. The program Tree Dyn 198.3 was used for tree rendering. (Dereeper et al., 2008).

3. Experimental Results
Over all 98 bacterial strains were isolated from the phyllosphere region of coriander, from enrichment
technique and followed by serial dilution method. The Phyllobactera have been screened for ammonia
production to identify nitrogen fixative bacteria. Among the 98 isolates, only 28 isolates were positive for
ammonia production test. Isolates CS02, CS04, CS07, CS10, CS13, CS16, CS19, CS22, CS24, CS28, CS32,
CS38, CS39, CS41, CS43, CS46, CS48, CS49, CS50, CS56, CS61, CS63, CS66, CS70, CS76, CS79, CS82,
and CS85 exhibited significant ammonia production. Isolate CS10, CS24, and CS61 has shown high level
of ammonia production. All the 28 nitrogen fixative strains were tested for growth promotion traits. Plant
growth promotion was assessed by qualitative determination of Indole Acetic Acid (IAA), ammonia
production and Hydrocyanic acid production.

Among 28 nitrogen fixative, only 3 (CS10, CS24, CS61) were the best producers of ammonia, High
production of IAA was associated with CS10, CS24, CS28, CS56, and CS82 and whereas isolates CS02,
CS07, CS43, CS50, and CS66 were the minimum indole acetic acid producers. Overall, 14 isolates were
positive for HCN production (Sharma, K., et al, 2007). Isolate CS31, CS32 and CS49 were the maximum
producer of Hydrocyanic acid.

Table I: Production of NH 3 , IAA and HCN by PGP isolates from phyllosphere of Coriander.

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S. No Isolate Name NH3 Production IAA Production HCN Production

01. CS02 ++ + +

02. CS04 ++ - -

03. CS07 ++ + ++

04. CS10 +++ +++ ++

05. CS13 ++ + +

06. CS16 ++ ++ +

07. CS19 ++ ++ +

08. CS22 + - -

09. CS24 +++ +++ -

10. CS28 ++ +++ +

11. CS32 ++ - +

12. CS38 + ++ +

13. CS39 + ++ +

14. CS41 +++ ++ -

15. CS43 ++ + -

16. CS46 + ++ -

17. CS48 + + +

18. CS49 ++ + -

19. CS50 + + ++

20. CS54 + - -

21. CS56 ++ +++ +

22. CS61 +++ ++ -

23. CS63 ++ + -

24. CS66 + + -

25. CS70 ++ - +

26. CS76 + ++ -

27. CS79 + - -

- = No production; + = Weak production; ++ = medium production; +++ = high production

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 S. No Isolate Name NH3 Production IAA Production HCN Production

28. CS82 +++ +++ -

- = No production; + = Weak production; ++ = medium production; +++ = high production

Production of extracellular enzymes by microorganisms plays an important role in the management of

plant pathogens as well as holds enormous economic potential. In view of the significance of
extracellular enzymes all the 28 PGP isolates were tested for their extracellular enzyme production like
phosphatases, lipase, catalase, lipase, amylase, and gelatinase activity. The results of extracellular
activities were listed on below table (Table II).

Table II: Extracellular enzyme activity of selected phyllobacterial isolates from Coriandra phyllosphere.

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S. Isolate P.Solibiliz Catalase Protease Lipase Amylase Gelatinase
No Name activity
-ation activity activity activity activity

01. CS02 0.5 ++ - 0.6 0.2 ++

02. CS04 - + - - 0.4 ++

03. CS07 0.3 +++ - 0.4 - ++

04. CS10 - + - 0.4 0.2 ++

05. CS13 0.6 + 2.7 - 0.4 ++

06. CS16 0.6 - 2.5 0.4 0.6 ++

07. CS19 1.2 - 1.9 0.4 0.9 ++

08. CS22 0.5 ++ 1.1 0.7 0.5 ++

09. CS24 1.1 + 2.4 - 0.3 -

10. CS28 0.8 +++ 0.8 1.5 0.9 +++

11. CS32 0.4 + 2.2 0.3 0.5 ++

12. CS38 1.1 - 0.5 - 0.5 ++

13. CS39 0.7 ++ - - 0.5 ++

14. CS41 0.2 ++ 0.5 - - +++

15. CS43 0.3 + 1.2 - 0.4 -

16. CS46 0.7 ++ 2.1 - - ++

17. CS48 0.6 ++ - 0.3 - ++

18. CS49 1.2 - 1.5 - 0.3 -

19. CS50 - + - 0.4 0.5 +++

20. CS54 0.9 - 1.5 0.5 - -

21. CS56 - + 2.1 0.3 0.9 ++

22. CS61 - - - - 0.5 ++

23. CS63 0.4 - 1.2 - 0.4 ++

24. CS66 0.7 - 1.6 0.4 0.2 -

25. CS70 0.8 - 2.2 0.3 0.9 -

26. CS76 - + 1.9 - 0.5 ++

27. CS79 0.2 + - 0.3 - ++

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 S. Isolate P.Solibiliz Catalase Protease Lipase Amylase Gelatinase
No Name activity
-ation activity activity activity activity

28. CS82 0.6 - - 0.4 0.4 -

- = No production; + = Weak production; ++ = medium production; +++ = high production; numerical values
represent the radius of zone formed in centimeter.

Based on PGP and enzyme activities of five isolates viz., CS02, CS13, CS19, CS28, CS38 were selected for
further studies. The colony morphology, growth at different concentrations of NaCl was studied for all the
five selected isolates. These isolates are showing motility. They are salt tolerant at 2% and 4% and
mesophilic i.e., all the selected isolates were able to grow at 20–50°C and at 10°C, 60°C the growth was
not observed (Table III).

Table III: Morphological characterization of isolates and growth on different NaCl concentration and
different temperature.

Isolate Gram Shape of Motility Growth at NaCl (%) Growth at temperature

code staining bacteria (0C)

2 4 6 8 10 10 20 30 40 50

CS02 (-)ve Rod + ++ + - - - - ++ + + +

CS13 (+)ve Cocci + + + - - - - ++ ++ + +

CS19 (-)ve Rod + + + - - - - + + + +

CS28 (-)ve Rod + ++ + + - - - ++ ++ + +

CS38 (-)ve Rod + + + - - - - ++ + + +

-=No Growth; + = Medium growth; ++ = moderate growth

All 5 isolates were tested for antagonistic activity against fungal pathogens Rhizoctonia solani (Otten
and Gilligan, 1998) and Colletotrichum capscici (Naziya et al., 2019). The isolates CS19 inhibited R.
solani and zone of inhibition was found to be 0.35 cm. The isolate CS02 showed antagonistic activity
against Colletotrichum capscici with an inhibition zone of 0.26 cm and no inhibition were found on other
isolates (Whipps, J.M., 2001).

Potential chosen five strains were applied for germination test and growth rate percentage. Overall study
indicates the higher growth of plant shoot and root region when compared to control or non-treated
seeds. Seeds treated with CS19 and CS28 isolates shown 92% of seed germination (Table IV). Maximum
root length and shoot length was observed from seeds treated with CS28 isolate (Biswas et al., 2000).

Table IV: Plant growth promoting potential of selected isolates by germination study.

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 S. Isolate Germination RL Average SL Average Moisture Dry
No Name percentage (%) (cm) (cm) Weight (g) Weight
(g)

1. Control 68 6.3 12.2 3.97 0.94

2. CS02 80 14.1 21.6 8.71 1.52

3. CS13 88 10.5 20.5 6.89 1.55

4. CS19 96 9.5 19.7 6.83 1.16

5. CS28 96 20.9 20.9 8.06 1.89

6. CS38 84 20.4 20.4 6.88 1.01

4. Discussion
A wide array of beneficial phyllosphere bacteria have been categorized as PGPR including mainly
diazotrophs, bacilli, pseudomonads and rhizobia. PGPR may induce plant growth promotion through
different direct or indirect modes of action. In the present study, Isolation of bacterial cultures from
phyllosphere samples of Coriandrum sativum from rural regions of Coimbatore district. The phyllosphere
region supported a total of 98 PGP isolates with varied characteristics. Among 98 isolates 28 Bacterial
strains were selected based on nitrogen fixative ability. All the isolates were screened for their plant
growth promoting activities viz., Indole acetic acid production (IAA, Phosphate solubilization, HCN
production, other lytic enzymes like Catalase, Protease, Lipase, Amylase, Gelatinase, Antagonistic
activities and Seed germination parameters. Characterization of selected phyllobacterial isolates by using
conventional methods like morphological characters, cultural characteristics on agar plate, growth on
broth media, growth on NaCl, and growth at different temperature was done as described in Bergey’s
Manual of Systematic Bacteriology. All isolates are gram positive, motile and rod shaped. Most of the
PGP’s are known to produce IAA. Another important trait of PGP isolates, that may indirectly influence the
plant growth, is the production of ammonia. All the selected isolates were positive for ammonia
production. A secondary metabolite produced commonly by rhizosphere pseudomonads is Hydrogen
Cyanide (HCN), a gas known to negatively affect root metabolism and root growth and is a potential and
environmentally compatible mechanism for biological control of weeds. Phosphorous is a major
essential macronutrient for biological growth and development. In our study, from 28 bacterial isolates 22
were able to solubilize phosphate in the plate-based assay, by showing a clear halo zone around the
colony. Among which CS19, CS24, CS38, CS49 showed more production then the other isolates. Fungal
plant diseases are one of the major concerns to agricultural production all worldwide. Antagonistic
microorganisms are a potential non-chemical mean of plant disease control. Many strains of Bacillus
have been shown to be potential biocontrol agent against fungal pathogens. In the present study, 28
isolates were selected for Coriandrum plant growth promoting and bio protecting activity which was
originally isolated from Coriandrum phyllosphere.

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Most of the rhizobacteria showed multiple traits for plant growth promotion and disease suppression.
The predominant trait observed was IAA production, HCN production, phosphate solubilization followed
by Ammonium production. Bacterial inoculants are able to increase plant growth and germination rate,
improve seedling emergence, responses to external stress factors and protect plants from disease. In the
present study it was observed that the length of root and shoot were enhanced by 70% compared to the
control due to seed treatment with PGPR. This might be due to the production of growth hormones like
IAA, gibberellins, auxins by bacteria. More than 80% of isolates showed positive response towards
germination. The highest response for seed germination was observed with isolate CS19 and CS28
showing 96%. Gelatinase and protease activity was detected in 20 and 17 isolates. Among 28 isolates 11
isolates not shown catalase activity. 16 isolates shown lipase activity. Salinity is an ever-increasing
problem in many regions of the world, particularly in arid and semi-arid regions. Salinity stress decreases
crop growth and productivity due to salt-induced reduction in the photosynthetic activity. In this study, the
ability of the isolates to withstand high salt concentration was a unique feature which may facilitate
competitiveness in soil ecosystem, where these isolates could be used. Selected five isolates were tested
for salt tolerance capabilities and observed that all the 5 isolates were observed to grow at high salt
concentration 2% and 4%. While only CS28 strain was capable of growing at 6% of NaCl, none of the
isolates were able to show growth at 6% and at 8%, 10% of NaCl concentrations. All the selected isolates
were able to grow at 200C − 50°C, 60°C and at 10°C the growth was not observed.

Based on the results of above mention test results, CS28 isolate was selected for molecular identification
through 16S rRNA Sequencing. A BLAST search of nucleotide sequences was done using multiple
sequencing alignment method and a common similarity were found between two genus
Stenotrophomonas maltophilia as 93.58% and geniculate as 93.44%. The program Tree Dyn 198.3 was
used for tree rendering.

Declarations

CONFLICTS OF INTEREST:
The authors declare that they have no conflict of interest.

FUNDING:
This research received no external funding.

ACKNOWLEDGEMENT
We are thankful to Yaazh Xenomics, Coimbatore for their kind help in our research work and we are also
thankful to Mrs. Sivakami K for providing the Organic Coriandrum sativum sample. We also thankful to

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Head Department of Biotechnology, Sri Shakthi Institute of Engineering and Technology, Coimbatore for
providing necessary laboratory facilities.

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Figures

Figure 1

Comparsion of shoot and root length of model plant (Green gram) between control and treated plant.

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Figure 2

Comparison of moisture and dry weight of model plant (Green gram) between control and treated plant.

Figure 3

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BLAST interaction shown similarities with sequences from the database.

Figure 4

Phylogenetic tree constructed from the data obtained from 16S rRNA Sequencing.

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