Lata Nain

Identification of multi-trait PGPR isolates and evaluating their potential as inoculants for wheat

Development of an effective plant growth promoting rhizobacteria (PGPR) inoculant necessitates the presence of a diverse set of traits that can help its colonization of the rhizosphere and survival under varying environmental conditions. In our investigation, a set of 100 bacterial isolates from the rhizosphere of wheat plants were screened initially on the basis of a seed germination assay; ten bacterial isolates (AW1–AW10) were selected. These isolates were then tested in vitro for specific PGPR traits, such as the production of IAA, siderophore, ammonia, HCN, P solubilization, ACC deaminase activity, acetylene reduction assay and antifungal activity. Of the ten isolates, AW5 was found to be promising for all PGP attributes. An experiment undertaken in the controlled conditions of the National Phytotron Facility revealed the potential of three isolates (AW1, AW5 and AW7) in enhancing the growth parameters of wheat plants. Characterization of these isolates using polyphasic approaches involving both phenotypic and genotypic attributes led to their identification as Bacillus sp. (AW1), Providencia sp. (AW5), and Brevundimonas diminuta (AW7), respectively. These strains could prove effective PGPR inoculants as they possess a number of traits useful for their establishment and proliferation in soil. The genus Providencia is reported for the first time for its PGP potential, using cultural as well as functional attributes to show its suitability as an inoculant for wheat crop.

Evaluation of fungicidal activity of extracellular filtrates of cyanobacteria--possible role of hydrolytic enzymes.

A set of seventy axenised and unicyanobacterial isolates belonging to the genus Anabaena were evaluated for biocidal activity against a set of phytopathogenic fungi. Among them, 35 Anabaena strains showed zone of inhibition against one or more fungi. The extracellular filtrates from 4 and 8 weeks old cultures of these Anabaena strains were further evaluated in terms of hydrolytic enzymes, proteins and IAA employing standard methods. Significant differences were also observed among the strains in terms of their FPase, chitosanase and xylanase activity, while low and relatively similar values of CMCase, cellobiase and protease activity were recorded in the strains analyzed. IAA production was also observed in all the strains. Comparative evaluation of activity of hydrolytic enzymes and antifungal activity revealed that such enzymes may contribute to the fungicidal activity of the cyanobacterial strains, besides other bioactive compounds, including IAA, which are established promising traits for biocontrol agents. This study is a first time report on the production of hydrolytic enzymes by these oxygenic photosynthetic prokaryotes, which can be potential candidates for the development of biocontrol agent(s) against selected phytopathogenic fungi. (© 2008 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

Novel perspectives for evolving enzyme cocktails for lignocellulose hydrolysis in biorefineries

The unstable and uncertain availability of petroleum sources as well as rising cost of fuels have shifted global efforts to utilize renewable resources for the production of greener energy and a replacement which can also meet the high energy demand of the world. Bioenergy routes suggest that atmospheric carbon can be cycled through biofuels in carefully designed systems for sustainability. Significant potential exists for bioconversion of biomass, the most abundant and also the most renewable biomaterial on our planet. However, the requirements of enzyme complexes which act synergistically to unlock and saccharify polysaccharides from the lignocellulose complex to fermentable sugars incur major costs in the overall process and present a great challenge. Currently available cellulase preparations are subject to tight induction and regulation systems and also suffer inhibition from various end products. Therefore, more potent and efficient enzyme preparations need to be developed for the enzymatic saccharification process to be more economical. Approaches like enzyme engineering, reconstitution of enzyme mixtures and bioprospecting for superior enzymes are gaining importance. The current scenario, however, also warrants the need for research and development of integrated biomass production and conversion systems.

Pretreatment of paddy straw with Trametes hirsuta for improved enzymatic saccharification

Delignification of paddy straw with the white-rot fungus, Trametes hirsuta under solid state fermentation, for enhanced sugar recovery by enzymatic saccharification was studied. T. hirsuta MTCC136 showed high ligninase and low cellulase activities. Solid state fermentation of paddy straw with T. hirsuta enhanced carbohydrate content by 11.1% within 10 days of incubation. Alkali extracts of Trametes pretreated paddy straw showed high absorbance at 205 nm indicating high lignin break down. The amount of value-added lignin recovered from the Trametes pretreated paddy straw was much higher than controls. Enzymatic hydrolysis of the Trametes pretreated paddy straw yielded much higher sugars than controls and yields increased till 120 h of incubation. Saccharification efficiency of the biologically pretreated paddy straw with Accelerase®1500 was 52.69% within 72 h and was higher than controls. Thus, the study brings out the delignification potential of T. hirsuta for pretreatment of lignocellulosic substrate and facilitating efficient enzymatic digestibility of cellulose.

Cyanobacteria mediated plant growth promotion and bioprotection against Fusarium wilt in tomato

Cyanobacteria - phytopathogenic fungi - tomato plant interactions were evaluated for developing suitable biological options for combating biotic stress (Fusarium wilt) and enhancing plant vigour. Preliminary evaluation was undertaken on the fungicidal and hydrolytic enzyme activity of the cyanobacterial strains (Anabaena variabilis RPAN59, A. laxa RPAN8) under optimized environmental/nutritional conditions, followed by amendment in compost-vermiculite. Such formulations were tested against Fusarium wilt challenged tomato plants, and the Anabaena spp. (RPAN59/8) amended composts significantly reduced mortality in fungi challenged treatments, besides fungal load in soil. Cyanobacteria amended composts also led to an enhancement in soil organic C, nitrogen fixation, besides significant improvement in growth, yield, fruit quality parameters, N, P and Zn content. The tripartite interactions also enhanced the activity of defence and pathogenesis related enzymes in tomato plants. A positive correlation (r = 0.729 to 0.828) between P content and pathogenesis/defense enzyme activity revealed their role in enhancing the resistance of the plant through improved nutrient uptake. Light and scanning electron microscopy (SEM) revealed cyanobacterial colonization, which positively correlated with reduced fungal populations. The reduced disease severity coupled with improved plant growth/ yields, elicited by cyanobacterial treatments, illustrated the utility of such novel formulations in integrated pest and nutrient management strategies for Fusarium wilt challenged tomato crop.

Phylogenetic Diversity and Characterization of Novel and Efficient Cellulase Producing Bacterial Isolates from Various Extreme Environments

A set of 300 bacterial strains isolated from various extreme environments were screened for the presence of cellulase activity on CMC agar plates. Phylogenetic analysis of the positive strain, based on 16S rRNA gene sequences indicated that the isolates were clustered within Firmicutes and Actinobacteria. A majority (17) of the isolates were identified as Bacillus, Paenibacillus, and Lysinibacillus sp., and the remaining three were identified as Arthobacter, Rhodococcus, and Bhargavaea cecembensis. Among the 20 positive isolates, 6 were evaluated for the production of cellulases on five different cellulosic substrates. Two isolates, B. cecembensis and Bacillus sp., based on maximum enzyme production on all cellulosic substrates, especially CMC and rice straw, were evaluated in terms of enzyme properties and kinetics. The enzymes of these two isolates are found to be active over broad range of pH and temperature. Such thermostable enzymes facilitate the development of efficient and cost-effective forms of the simultaneous saccharification and fermentation process converting lignocellulosic biomass into biofuels and value-added products.

Development of cyanobacterium-based biofilms and their in vitro evaluation for agriculturally useful traits.

The ability of cyanobacteria to be useful as matrices for agriculturally important bacteria was evaluated. Biofilms were generated with the selected strain Anabaena torulosa after co-culturing with Azotobacter chroococcum, Pseudomonas striata, Serratia marcescens, and Mesorhizobium ciceri. The biochemical attributes were compared with individual bacterial and cyanobacterial cultures. The biofilms were characterized in terms of proteins, chlorophyll, IAA production, acetylene-reducing activity, phosphate solubilization, and antagonism towards selected phytopathogenic fungi. An enhancement in the population counts was recorded in A. torulosa–S. marcescens and A. torulosa–P. striata biofilms. The A. torulosa–A. chroococcum and A. torulosa–M. ciceri biofilms were also able to utilize new saccharides as compared to the individual cultures. Such novel biofilms with agriculturally useful traits can provide additional advantages including the broader spectrum of activity and the presence or formation of biologically active compounds; they also suggest the way to effective inoculants for sustainable and environment friendly agriculture.

Electrophoretic fabrication of chitosan-zirconium-oxide nanobiocomposite platform for nucleic acid detection.

The present work describes electrophoretic fabrication of nanostructured chitosan-zirconium-oxide composite (CHIT-NanoZrO(2)) film (180 nm) onto indium-tin-oxide (ITO)-coated glass plate. This nanobiocomposite film has been explored as immobilization platform for probe DNA specific to M. Tuberculosis as model biomolecule to investigate its sensing characteristics. It is revealed that pH-responsive behavior of CHIT and its cationic skeleton is responsible for the movement of CHIT-NanoZrO(2) colloids toward cathode during electrophoretic deposition. The FT-IR, SEM, TEM, and EDX techniques have been employed for the structural, morphological, and composition analysis of the fabricated electrodes. The morphological studies clearly reveal uniform inter-linking and dispersion of hexagonal nanograins of ZrO(2) (30-50 nm) into the chitosan matrix, resulting in homogeneous nanobiocomposite formation. Electrochemical response measurements of DNA/CHIT-NanoZrO(2)/ITO bioelectrode, carried out using cyclic voltammetry and differential pulse voltammetry, reveal that this bioelectrode can specifically detect complementary target DNA up to 0.00078 μM with sensitivity of 6.38 × 10(-6) AμM(-1).

Streptomyces griseorubens mediated delignification of paddy straw for improved enzymatic saccharification yields.

Biological pretreatment of paddy straw was carried out using an actinomycete isolate, identified as Streptomyces griseorubens ssr38, for delignification under solid state fermentation and enhanced sugar recovery by enzymatic saccharification. After 10 days incubation, the inoculated paddy straw was extracted with mild alkali and high absorbance at 205 nm was shown by the extracts indicating the ability of S. griseorubens ssr38 to depolymerize/solubilize lignin to a high extent. Also, almost 25% of depolymerized lignin could be recovered as value-added acid-precipitable polymeric lignin (APPL) as compared to controls. Enrichment in carbohydrate content of inoculated paddy straw following delignification led to a high saccharification efficiency of 97.8% upon enzymatic hydrolysis with Accelerase®1500. The study, therefore, proves the potential of actinomycetes, besides the conventionally used white-rot fungi, for biological pretreatment, in the biomass to bioethanol process, with respect to the high extent of delignification, lignin recovery, cellulose enrichment and very high saccharification efficiency.

Developing PGPR consortia using novel genera Providencia and Alcaligenes along with cyanobacteria for wheat

A pot experiment was undertaken using unsterile soil supplemented with the recommended dose of chemical fertilizers in order to investigate the effect of indole-3-acetic acid (IAA)-producing proteo- and cyanobacterial strains on the growth and yield of wheat (Triticum aestivum variety PBW343). Two proteobacterial (WRB4 Providencia sp. and WRB10 Alcaligenes sp.) and two cyanobacterial (WRC3 Anabaena oscillarioides and WRC4 Anabaena torulosa) strains were used individually and in combination. The treatment in which proteobacteriumWRB4 (Providencia sp.) was inoculated showed significantly higher values in comparison with controls for various plant-growth parameters recorded, i.e. shoot length, root length, shoot weight, root weight and crop biomass, followed by the treatment in which the WRC3 (Anabaena oscillarioides) was used. A positive interaction among the proteo- and cyanobacterial strains, in particular WRC3 and WRB4, was also observed by way of enhancement of plant-growth parameters. Significant enhancement in soil microbiological activities such as fluorescein diacetate (FDA) hydrolysis and dehydrogenase activity were recorded in the treatments, particularly in those inoculated with cyanobacterial strains, when compared with fertilizer controls. This is a first-time report on the potential of selected combinations of proteobacterial genera such as Providencia and Alcaligenes and cyanobacteria such as Anabaena as plant growth-promoting organisms in wheat crop.