Senior Research Fellow
Department of Biotechnology, Chandigarh University
Agro- chemicals and synthetic fertilizers have played a substantial role in making our country self-sufficient in food grain production. Six decades of imprudent use of chemical fertilizers has rendered the soil on the verge of infertility. The nature has endowed various microorganisms with the innate potential to augment plant growth through various mechanisms. These microorganisms naturally inhibit the roots of plants and symbiotically help them in their growth. Different species of bacteria (Pseudomonas, Enterobacter, Klebsiella, Acetobacter, Corynebacterium, Gluconacetobacter, Erwinia, Escherichia, Ralstonia, Flavobacterium, Serratia, Bacilli, Rhizobium and Agrobacterium) and fungi (Candida krissii, Mucor ramosissimus and Penicillium expansum) have been reported to enhance the crop yield by secreting chemical compounds that help the plant to withstand abiotic and biotic stress conditions. These microorganisms also aid the plants to acquire nutrients from the soil.
There is a huge untapped potential for the use of these microorganism for the benefit of agriculture. Microbes can be used to tolerate crop stress conditions like salinity, disease susceptibility, pest attack and nutrient deficiency. Application of microbes has shown 12% enhancement of paddy yield, 30% increase in yield of banana, 70% improvement of beans’ yield and 74% increment in yield of wheat. With advancements in genomic techniques, molecular tools, next generation sequencing methods and omics tools, isolation and identification of agriculturally important microbes has become relatively easier. These bacteria are generally found in association with plant roots and thus termed as rhizobacteria. The crop specific rhizobacteria can be isolated and multiplied to generate millions of copies for crop improvement programs.
These microbes can be delivered either through seed treatment, direct soil application or foliar spray. The bio formulations of live or latent rhizobacteria that has the potential of augmenting plant growth is termed as biofertilizer. Biofertilizers can be carrier-based, liquid or encapsulated. When the biofertilizer is prepared by mixing the bacterial cells with some carrier material like clay, coconut shell powder, peat, talc, perlite, vermiculite, rice bran, wheat bran, it is called a carrier based biofertilizer. The carrier material ensures the survivability and efficiency of bacteria during storage. Liquid biofertilizers are broth cultures containing dormant forms of desired microorganisms along with required nutrients, minerals and organic oils. However, the viability of bacteria in both carrier-based and liquid biofertilizer is under scrutiny due to high contamination rate and low viability during environmental stress conditions. Encapsulated biofertilizers are projected towards development of novel bioformulations, making use of polymer solutions to encapsulate bacteria into a biodegradable matrix ensuring high viability, low contamination and prolonged slow release.
Steps involved in the biofertilizer development
1. Isolation of rhizobacteria with plant growth promoting potential- The first step in the making of biofertilizer involves isolation of a potential rhizobacteria from the specific crop for which the intended biofertilizer is made. The special microbiological media are available for the isolation of the bacteria of interest. For example, for the isolation of phosphate solubilising bacteria, one can use Pikovskaya’s agar media. The rhizobacteria showing clear zones around the colony ensure its phosphate solubilisation phenotype. Similarly, one can isolate bacteria of interest using different available media. The screened bacteria are then maintained as pure cultures in 20% glycerol stocks at -80°C for further use.
2. Studying the Plant Growth Promoting potential (PGP) of isolates – The isolated rhizobacteria are further screened for various PGP potential like plant hormone production, secretion of antimicrobial compounds, siderophore production and metabolite production ensuring stress tolerance in plants. The bacteria with the ability to produce antimicrobial compounds ensure plant protection to pest and pathogens while the production of phytohormones is crucial for plant growth and development. The rhizobacteria with the maximum of all these traits is the prime candidate for biofertilizer. Biosafety assessment of the rhizobacteria is a crucial step here, as it ensures that the microbe being used in the biofertilizer is not pathogenic to the humans.
3. Molecular tools for enhanced PGPR activity- A single microbe is endowed with one or few PGP traits only. There is no single fit biofertilizer that works for all crops and climatic conditions or that provides complete nutrient supply to plants. However, these rhizobacteria can be engineered to augment indole acetic acid production, disease resistance, chitinases activity, nitrogen fixation and phosphorus solubilisation potential. Advances in gene editing tools and omics technologies have eased the process of gene manipulation in bacteria allowing non-PGPR strains to work as PGPR inoculants in rhizosphere.
With the advancements in molecular biology, synthetic biology and hands on genome editing techniques, there will be vast opportunities to design a microbe that can provide all the three essential macronutrients (nitrogen, phosphorous, potassium) to the plants regardless of the crop.
4. Encapsulation- Once you have the desired microbe, the next step is encapsulating the live microbe in a biodegradable matrix that provides protection, ensures viability during long storage and prevents contamination. The microbe can be encapsulated in natural (sodium alginate, agarose, lignin, biochar, gum arabic) and synthetic (olystyrene, polyacrylamides, polyurethane, poly (alkylene oxides), poly(vinyl acetate), polyvinylpyrrolidone (PVP) and polyethylene glycol) polymers. The matrix acts as a ‘mini-fermenter’ in which low biomass can maintain their metabolic activity for a long time.
5. In-vitro testing and field trials- The prepared biofertilizer is then studied for it’s effectiveness on test plant, encapsulation efficiency, microbe release rate in different storage conditions and soil behaviour in pot experiments under controlled conditions. If the desired results are obtained in lab conditions, the prepared biofertilizer undergoes field trials for its possible beneficial effects on the test crop. Each trial is planned in triplicates to minimize the chances of error. After the success of field trials, the biofertilizer can be scaled up for market use. Post market surveillance is an importance aspect which should not be overlooked. This involves processes and activities used to assess the quality and standard after distribution of the biofertilizer to the farmers in an uncontrolled environment.
Conclusion – The increasing concern regarding chemical fertilizers and the urge to promote organic farming has led researchers to look for bacterial disposition of plant growth promotion. In this regard, numerous bacteria and fungi have been isolated, screened and studied for their potential to act as key players in enhancing plant growth and combating biotic and abiotic stress. Still, acceptance of bio fertilizers for large scale agriculture is a grave concern due to difference in performance as affected by time and space. Encapsulating the bacteria in a polymer matrix and applying it in fields in the form of beads, for better delivery to the roots can be a better alternative to liquid or carrier based bioformulations. Field trials for different crops and at different areas need to be performed to determine their potential for commercial usage.