The Microbiology and the Integrated Pest Management (IPM) group focuses on research, development and dissemination of ecofriendly technologies through innovative delivery models to promote sustainable agricultural practices. The research focus is technology specific (eg. biofertilisers, biopesticides, and bio control agents) and need based to address problems encountered at farm hold level. The group also focuses on extensive demonstrations and trainings in the areas ofbio-pesticides, biofertilisers, soil testing, and integrated nutrient and disease management to support to the farmers. Some of these demystified technologies have also been set up as community managed enterprise. Soil analysis is also undertaken as part of the soil health assessment by analyzing the chemical and biological properties of the soil and a soil health card has been developed.

Screening for novel microbes and their beneficial functions

Bioprospecting for saline tolerant, nitrogen fixing, phosphate solubilising and plant growth promoting microbes from the pristine agriculture and mangrove ecosystems to be used as potential biofertilisers and biocontrol agents in agriculture practice in hear, normal and stressed soils.

Diversity of Pseudomonas from the saline Agroecosystems: The diversity of Pseudomonas populations isolated from three different plant rhizospheres, namely paddy, pearl millet and cotton grown in saline soils along the coastline of Southern India was studied. The genetic diversity was analyzed using molecular tools, Random Amplified Polymorphic DNA (RAPD) and Polymerase chain reaction Restriction Fragment Length polymorphism (PCR-RFLP). The biochemical characterization, antibiotic resistance assay and RAPD profiles revealed the presence of a largely homogenous group and the PCR-RFLP restriction showed two groups predominantly present in all the three rhizospheres, while the minor group consisted of salt-sensitive isolates restricted to the paddy rhizosphere alone. Increasing salinity caused a predominant selection of salt tolerant species, in particular Pseudomonas pseudoalcaligenes and Pseudomonas alcaligenes, irrespective of the host rhizosphere. The diversity of Pseudomonas associated with the finger millet rhizosphere was studied by amplification of the 16S rDNA using Pseudomonas genus-specific and GTG(S)primers. Free tree V1.6.6. using Nei and Li/Dice similarity matrix calculation and clustering was done by UPGMA algorithm.

Diversity of Azospirillum from the saline rice ecosystems: The diversity of the Azospirillum associated with rice fields of the coastal agro ecosystem was estimated by PCR RFLP of the 16S rONA. Among the 302 isolates tested 19 restriction pattern were observed. The frequency of occurrence of salt tolerant spp. of A. brasilense was high than A. lipojernum, which was sensitive to salinity, thereby indicating high association between the salinity and distribution of the Azospirillum in the rice ecoysytems.

Genetic Diversity of Bradyrhizobia isolated from different crops: The genetic diversity of Bradyrhizobial strains associated with blackgram and cowpea grown in two different agricultural soils (non-saline and saline) along the coastline of Tamil Nadu has been analyzed. DNA polymorphism of phenotypically indistinguishable isolates was analyzed using RAPD of 16S rONA and nif D, which showed considerable genetic diversity. Soil pH and salinity had an effect on the selection of natural populations as revealed by PCR-RFLP of 16S rONA. A group of strains and a type strain sharing three different 16S PCR-RFLP types were shown to have the same set of symbiotic genes as inferred from the PCR-RFLP pattern of nif D. The diversity of bradyrhizobia associated with the Soybean (Glycine max Merrill.) was studied using restriction fragment length polymorphism (RFLP)and phylogenetic analysis. Cluster analysis of16S rONA restriction patterns with seven tetrameric endonucleases grouped these isolates with Bradyrhizobium spp. resolving two genotypes within these Bradyrhizobia and the intergenic spacers (IGS) and RFLPs with three restriction enzymes formed six genotypes. The IGS sequences of the strains of Indian origin have very low similarity to those of the type strains of validly described species of Bradyrhizobiwn from other legumes and also from soybean. The RFLPs of symbiotic genes (nif H and nod C) and phylogeny based on the nif H DNA sequence delineated all isolates into two biovarieties. It clearly showed the existence of the potential new biovariety among soybean nodulating bradyrhizobia. One strain designated M6 can be proposed as a novel sp. in the genus Bradyrhizobium.

Novel genus Swaminathania salitolerans gen. nov. spp. nov. from the mangroves: Screening for microbial strains, from mangrove-associated wild rice (Porteresia coarctata- Tateoka), resulted in the isolation of a number of strains belonging to the taxa Swaminathania, Vibrio, Bacillus, Enterobacter and Azospirillum. Novel salt tolerant strain Swaminathania salitolerans gen. nov., sp. nov. isolated from P. coaractata were found to fix nitrogen and solubilize phosphate and showed intensive colonization in Pokkali than in P. caroctata and Ponni. Phylogenetic analysis based on their 16S rRNA genes showed homology to the genus Acidomonas, Asaia, Acetobacter, Gluconacetobacter, Gluconobacter and Kozakia in the Acetobacteraceae. Like wise a diazotrophic strain MSSRF40T was identified as new genus in the family Enterobacteriaceae for which Mangroveibacter plantisponsor gen. nov. sp. nov., was assigned.

Novel Vibrios associated with the mangroves

Four novel species of the genus Vibrio sola ted from the mangrove rhizosphere were named as Vibrio rhizosphaerae sp. nov., Vibrio porteresiae sp.nov., Vibrio mangrovi sp.nov and Vibrio plantisponsor sp.nov is proposed. Following the polyphasic taxonomic approach. Multilocus sequence and phylogenetic analysis using 168 rRNA, recA, pyrH, 'poA and nifH gene sequences, PCR based genomic fingerprinting using PCR-RFLP of 168 rRNA, ribotyping and repetitive extragenic palindrome (rep)-PCR using GTGs and BOX primers. Fatty acid analysis, biochemical characterization, G+C content analysis and whole genomic DNADNA hybridization, has identified these Vibrio strains and also defined a new ecological niche for vibrios as a rhizosphere-associated heterotrophic diazotroph.

Diversity of actinomycetes from different ecological niches

The diversity of the actinomycetes from different ecosystems have been isolated and efficient phosphate solubilising strains have been identified.

The isolation of DNA has been standardized and the amplification of the 16 S rDNA was carried using fD1 (5'-AGTTTGATCCTGGC TCAG-3') position7- 26 and r P2 (5'ACGGCTACCTTGTTACGAC TT-3')- position 1513 to 1494 primers. The initial genetic diversity analysis of antagonistic actinomycetes was performed using GTGs primer, which showed 4 different genotypes of actinomycetes. A further study on the characterization of these genotypes is under progress.

Metagenomic of the unculturables from the mangroves: The diversity of unculturable microbes associated with the rhizosphere of five different mangrove spp. viz., P.coarctata, Rhizophora mucornata, Avicennia marina, Sonnerratia apetla and Excoecaria agallocha collected from Pichavaram mangroves, South India was analyzed using molecular tools like RFLP, Amplified Ribosomal DNA Restriction Analysis (ARDRA), and Single-Strand Conformation Polymorphisms (SSCP). The DNA from the soil was extracted using Soil Master™ and the 16S rRNA was amplified using Coml and Com2-Ph primers. Single stranded DNA was obtained by lambda exonuclease digestion. The SSCP pattern revealed that majority of the dominant populations were closely related to previously described soil bacteria (i.e., Planococcus, Planomicrobium, Bacillus, and low G+ C Grampositive bacterium, and glacial ice bacterium).

Biological control of Phytopathogens using PGPRs: Different group of organisms viz., Pseudomonas, Bacillus and actinomycetes were screened for their antagonistic activity against Rhizoctonia solani; Macrophomina phaseolina, Pyricularia oryzae and Xanthomonas oryzae etc. to isolate efficient strains with antifungal and antibacterial activity has been identified. Antagonistic Pseudomonas was selected using Pseudomonas primers. The diversity of the antibiotic coding genes for DAPG, PLT and PRNT were determined using specific primers. About 48% of the Pseudomonas strains showed positive amplification for DAPG gene of size with DAPG primers Phl2a and Phl2b and 16% of the strains show positive amplification for HeN primers PM2 and PM7..

Bioprospecting for metabolites from microbes collected from the Eastern Ghats

The demand for new effective antibiotics to combat the development of resistance by microbes to the existing drugs. Discovering novel biomolecules using the untapped resources has gained momentum and the modern biodiversity prospecting integrates the systematic search for the new sources of biomolecules, genes and other economically valuable natural products. The project involves nine institute and Nicholas Piramal India Limited. This project envisages a mega scale screening for microbes from different ecological niches. In the present study screening for metabolites from microbes isolated from soil samples collected from different ecological regions of the Eastern Ghats and the mangrove ecosystem will be in 30 different media and a total of 1000 cultures/ month (total of 13, 000 cultures) were isolated to purity and the colony characteristics viz., colony size, configuration, margin, elevation, colour, mucilage, biofilm formation and pigment production were recorded. The cultures were stab inoculated and dispatched to Nicholas Piramal Limited (NPL) Mumbai where the isolates were screened for novel biomolecules with anti-cancer, anti-diabetes, anti-inflammatory and anti- infectivity activity by HITS (High infectivity throughput screening tests). Two sets of glycerol vials were stored in cry preservation vials at -80°C. And another set of stab was sent to National Centre for Cell Sciences (NCCS) for storage.

Development of biological software for good agricultural practices

Trials were conducted in the coastal agriecosystems in farmer's field in Chidambaram to study the performance of the phosphate solubilizers and nitrogen fixers under field conditions. The phosphobacterial strains, viz., PS-5 and PS-10, nitrogen fixers MSA- 274 and MSA-160 and S. salitolerans strains, PA-6 and PA-51 were used as these strains were proved efficient in the green house conditions. The trial was conducted in four different locations covering 207 acres. Biofertilizers formulated in vermiculite were used. Three treatments viz., seed bacterization, root dipping and foliar application (in intervals of 30 days) were given. These treatments resulted in an increase in the yield parameters
of 20-30% compared to the chemical control. The Pseudomonas strains MSP-393, MSP27; MSP147, MSP573 were tested for the control of bacterial blight disease and sheath blight disease. Toxicological analysis of the strain MSP 393 proved it to be non toxic by HBAT.

Talc and Liquid based formulations: Talc and liquid formulation of the biofertiliser and the biocontrol agents were standardized. Third party testing of the talc formulations in the field was carried by Elbitech. The liquid formulations were standardized using ifferent cell protectants and preservatives and the shelf life of the product was stable up to six months of storage and the CFU was maintained upto 4x109 and hence proved to be superior to the talc based formulations. The performance of the liquid formulations in the field is being evaluated by a trial at the Biocentre in Puducherry.

Ecoenterprrise development

One of the innovation areas is the demystification of technologies and establishing low cost units for the production of the biological software's for the promotion of good agriculture practices as part of promotion of ecoagriculture and organic farming. The ecoenterprises like Trichoderma viride, Trichogramma and Pseudomonas fluorescens and biofertilisers like Azospirillum and phosphobacteria are successfully management by the women SHGs. To improve their technical skills representatives from the Jhansi Rani WSHG and Manimegalai WSHG were given hands on training in the production of Azospirillum and phosphobacteria in the microbiology lab at MSSRF. Two decentralized units have been set up one at Kannivadi, Dindigul district and the other a Kezhsathamangalam in Puducherry. The project interventions helped to enhance the awareness among men and women farmers of the region on different kind of biofertilizers and its crop specificity over a period of time

Integrated Pest Management

IPM group is involved in bioprospecting, mass multiplication of promising biopesticides, field testing and establishing micro enterprises and demystification of technologies.

Bioprospecting

Biological control of Helicoverpa armigera using Cipadessa baccifera, Clausena dentata, Dodonaea angustifolia and Melia dubia (at laboratory &field level) was carried out. Different solvent as well as aqueous extracts of the traditional plants were tested against different stages of H. armigera. The adult longevity, eggs laid and hatchability was recorded in the adult, while the pupation, larval duration, pupal weight and adult emergence were recorded in the larvae. The adult life span of H. armigera was reduced by petroleum ether and chloroform extracts of D. angustifolia, acetone extract of M. dubia and chloroform extract of Clausena dentata. Among the test plants, D. angustifolia reduced the fecundity compared to others. The egg hatchability was suppressed by many of the treatments, despite the induction in egg laying.

Pupation was least and malformed adults were high in petroleum ether extract of Cipadessa baccifera, acetone extract of Clausena dentata, hexane extract of D. angustifolia and chloroform extract of M. ubia. Life span was less in adults emerged from acetone and petroleum ether extracts and solvents (Cipadessa baccifera), hexane extract further fed with normal diet (Clausena dcntata), acetone extract (D. angustifolia) and chloroform extract (M. dubia).

Efficacy of D. angustifolia extract against H. annigera on rose and Earias vitella in cotton under field conditions

There was a substantial decrease in the eggs, number of larvae and percent damage of buds and flowers after 4th spray of extract. Hexane and acetone extracts were highly effective, and on par with NPV. The compound responsible for insecticidal activity against H. armigera in leaf extract of Argemone mexicana was identified as 3-hydroxy-2- (15-methylhexadecyl)-1-cyclopentanone and from Clausena dentata as ((23E)-23, 24,27- trimeth yl-octocos- 23-enoicacid -[(2E)-4-methyl-pen t-2-enoil-oxy -methyl ester).

Mass multiplication of biopesticide

The multiplication of three species of Trichogramma namely T. chilonis, T. japonicum and T. brassicae has been standardised for crop specific problems. The group has developed low cost technology for mass multiplication of entomopathogens, Beauveria bassiana, Metarhizium anisopliae, Verticillium lecanii, Nomureae rileyi and fungus Paecilomyces lilacinus. Different substrates viz., rice, sorghum, barley, groundnut, potato, beetroot, carrot, sawdust and paddy straw were used for the mass production of the entomopathogens. The addition of 1% (w/ w) yeast extract (YE) in all the substrates enhanced the sporulation of B. bassiana as against the control. Among the grains tested, the maximum sporulation of B. bassiana was observed on rice supplemented with 1%YE (117.0 x 108spores/ g).

Field testing and Establishment of micro enterprises (Extension)

Trichogramma units at Kannivadi (2 nos.), Puducherry (1 no.) and Pudukottai (3 nos.) and mushroom units at Puducherry (2 nos.) has been established.

Mass production technology of Trichogramma has been demistified and transferred the technology to WSHG. The crop calendar based production coupled with quality has been a positive step to manage these notorious pests. The parasitoid production centers are successfully run by seven groups.

The trainees have now become trainers and the horizontal transfer of knowledge and technology through well-planned objectives and execution has promoted capacity building, leadership quality, employment generation, reduced pesticide usage, higher cost benefit ratio and sustainable agriculture in the villages.

MSSRF has imparted training to one woman SHG at Kannivadi, Dindigul District, in the different technologies involved in rPM, based on the cropping pattern and the pest incidence in their area. After one year of initiation, the concept has spread into other places and the pioneer group members started training six groups and now each group run this as a micro enterprise through which they reach the farmers in and around their area and create awareness about this particular parasitoid and its control. They have proved this as an economically and environmentally viable enterprise, indirectly reduced the usage of chemical pesticides by 35% in their area.

Edible mushroom is a healthy food and its production is promoted as a micro enterprise. Six women and three youth SHGs benefited from the training imparted by the IPM group. Other groups who were restricting to savings were also attracted towards such training and expressed their willingness to participate in the future.

Technologies taken to the field from us and adopted as practiced by others: Use of Trichogramma chilonis in the management of a wide range of pests; Mushroom production (oyster and milky); Integrated pest management packages such as multiple cropping, crop rotation, use of light traps, mechanical removal of pests etc; Spraying botanicals as a preventive tool against sucking pests, defoliators and borers and use of entomopathogens in the management of field problems at Thonimalai are among the technologies taken to the field.

Demonstration of the efficacy of entomopathogens against various insect pests at Thonimalai under organic farming.

Support to organic farming at Thonimalai by way of field survey, identification of pests and diseases were provided with guidance to application of IPM packages suitable to the cropping system. The major pests and diseases observed and the IPM practices followed at Thonimalai are given below.

Banana pseudostem weevil: Among the 245 trees in the 2 field, 85 trees were infested with banana pseudostem weevil. Prophylactic pseudostem injection of neem oil coupled with soil application of neem cake was carried out to prevent the incidence of banana pseudo stem weevil. 146 pseudo stem traps + 2.6 Kg of B. bassiana pasted traps were installed.

Surveillance of insect pests of Morinda citrifolia in the west coast of Kerala

Periodical visits to west coast of Kerala and Karnataka are made to conduct survey in Morinda citrifolia and M. tinctoria (natural habitat as well as farms). The pest population if any on the above species in different seasons is recorded and documented. The major pests recorded till date are Membracid bugs, Coccinellid, Valley carpenter beetle, Plant hopper, Grasshopper, White fly, Green plant bug, Predatory bug, Mantis, Fruit fly, Weaver ants, leaf hopper.

Demystification of technology: Parasitoids and predators

The mass multiplication procedure of parasitoids, Trichogramma chilonis, and Tetrastichus sp.; predators, green lacewing, Chrysoperla carnea, and aphid predator,Cryptolaemus montrouzieri were demystified. The objective is to reduce the cost, simplify the methodology and develop a user friendly model so that it can be converted into a micro enterprise.

Soil analysis

Soil testing helps to diagnose soil health and evolve soil specific and crop specific solution. Soil testing is the basis for scientific management of fertilizer requirements. The basic objective of the soil-testing programme is to educate farmers on judicious and economic use of fertilizers and better soil management practices for increasing agricultural production based on the soil nature. Soil health cards have been developed and are being promoted in all sites especially in the watershed sites and places where good agriculture practices are being promoted. GIS based tools are being used to map the soil profiles and as monitoring tools. So far @ 5000 soil samples from different sites have been analysed for pH, EC, organic carbon, macro nutrients (N, P and K), exchangeable cations (Ca, Mg, Na & K) and micro nutrients (Cu, Zn, Fe & Mn). 67 water samples from Jeypore and Puducherry were analyzed for pH, Electrical conductivity, anions, cations, SAR and RSC. Developed a soil health card for the benefit of farmers.

Biological properties of the soil

The microbial population plays an important role in the maintaining the soil health. The microbial population and the different enzyme activity related to the microbial activity has also been estimated. 691 samples from project sites were analysed for dehydrogenase and phosphatase activity. 180 soil samples from Pudukottai, Puducherry and Jeypore were analyzed for microbial load. Six phosphate solubilizers and 12 nitrogen fixing positive strains were isolated and sub cultured for future studies.