BIOFERTILIZERS AND BIOPESTICIDES DR. Mrs. REKHA GULVE ASSO. PROF.MICROBIOLOGY MRS. K.S.K.COLLEGE, BEED.

BIOFERTILIZERS Definition: Biofertilizers can be defined as a live microbial preparation, when applied to seed ,root or soil, colonizes the rhizosphere or the interior of the plant, and promotes growth and releasing growth promoting substances by increasing the availability of primary nutrients. Biofertilizers also called ‘Microbial Inoculants’. E.g. - Rhizofertilizer, Azofertilizers, Azospirillum fertilizers, Azolla fertilizers.

Advantages of Biofertilizers Biofertilizers are cost effective. They are eco-friendly . Increase crop yield by 20-30%. Improve soil texture and water holding capacity and restore natural soil fertility. Add nutrients to soil and make nutrients available to crop. Supplement to fertilizers, replace chemical nitrogen and phosphorous by 25%.

N2 fixing Biofertilizers Groups Examples N2  fixing Biofertilizers 1. Free-living Azotobacter, Beijerinkia, Clostridium, Klebsiella, Anabaena, Nostoc,  2. Symbiotic Rhizobium, Frankia, Anabaena ,Azollae 3. Associative Symbiotic Azospirillum P Solubilizing Biofertilizers Bacteria Bacillus megaterium var. phosphaticum, Bacillus subtilis Bacillus circulans, Pseudomonas striata Fungi Penicillium sp, Aspergillus awamori P Mobilizing Biofertilizers Arbuscular mycorrhiza Glomus sp. ,Gigaspora sp.,Acaulospora sp., Scutellospora sp. & Sclerocystis sp. Ectomycorrhiza Laccaria sp., Pisolithus sp., Boletus sp., Amanita sp. Ericoid mycorrhizae Pezizella ericae 4. Orchid mycorrhiza Rhizoctonia solani Biofertilizers for Micro nutrients Silicate and Zinc solubilizers Bacillus sp. Plant Growth Promoting Rhizobacteria Pseudomonas Pseudomonas fluorescens

1) Rhizofertilizers : Rhizofertilizers contain different species belonging to Rhizobium genus. Rhizobia are Gram negative, motile rods, aerobic in nature, present in soil. They form root nodules in leguminous plants and fix nitrogen symbiotically. Effective nodules are large, pink, and firm and contain Rhizobia as bacteroids. Pink color of root nodules is because of leghemoglobin. Bacteroids Growth of Rhizobium on Congored yest extract mannitol agar

R. leguminosarum nodules Pea Plant Pink color is leghaemoglobin a protein that carries oxygen to the bacteroids

Nodulation Process: The bacteria enter in the root nodule through root hairs, the interaction is highly specific and result in nodule formation. Inside the nodules many bacteria are presents in bacteroid form (undividing stage) which produce ‘nitrogenase enzyme’. The enzyme reduces the atmospheric nitrogen into ammonia this processes is called nitrogen fixation. The Rhizobium species are host specific means one Rhizobial species able to nodulate or infect a group of specific leguminous plant species, is called ‘cross inoculation group’. For ex. Rhizobium leguminosarum biovar viciae infect or form nodule in pea group. The species of Rhizobium are differ in their ability to fix nitrogen. Screening is essential to select the efficient nitrogen fixing species for preparation of biofertilizers. The estimated nitrogen fixation by Rhizobium species is 50 to150 Kg/Ha or even more nitrogen in field trial in India. 10 to 15% crop yield increase when rhizofertilizers use.

Host Group Rhizobium Species Crops Pea group Rhizobium leguminosarum Green pea, Lentil Soybean group R.japonicum Soybean Lupini Group R. lupine orinthopus Lupinus Alfafa grp.Group R.mellilotiMedicago Trigonella Melilotus Beans group R. phaseoli Phaseoli Clover group R. trifoli Trifolium Cowpea group R. species Moong, Redgram, Cowpea, Groundnut Cicer group Bengal gram

Isolation of Rhizobium Strain An undamaged pink colored root nodule is selected The nodule is surface sterilized with 10% HgCl2 For few minutes. The nodule is washed with sterile distilled water. The washed nodules are crushed in physiological saline. The suspension is streaked on Congored yest extract mannitol agar( CRYEMA) plates. The plates are incubated at 270C for 3-7 days.

Starter culture of a Rhizobial inoculant A colony of efficient Rhizobial isolate is transferred to sterile YEM broth in the 250 ml capacity flask. The flask is incubated on rotary shaker at 28-30 0 C for 4-5 days. The culture is called starter culture or mother culture. The starter culture attains a cell density of 10 5- 106 cells/ml, this is called ‘seed’ and is used for charging large fermenters.

Mass production of Rhizobial Inoculant For mass production jacketed stainless steel fermentor equipped with various control devices is used. In fermentor YEM broth is used. Various parameters controlled during fermentation include : Aeration(1lit gas/min/20lit medium); agitation, temperature(28-30 0 C) and pH 5-7. 1% inoculum is used. A population density of 5x 10 9 /ml will be obtained in 96 hours. Broth is taken out from the fermentor and mixed with suitable carrier.