Core microbiota of wheat rhizosphere under Upper Indo-Gangetic plains and their response to soil physicochemical properties.

Wheat is widely cultivated in the Indo-Gangetic plains of India and forms the major staple food in the region. Understanding microbial community structure in wheat rhizosphere along the Indo-Gangetic plain and their association with soil properties can be an important base for developing strategies for microbial formulations. In the present study, an attempt was made to identify the core microbiota of wheat rhizosphere through a culture-independent approach. Rhizospheric soil samples were collected from 20 different sites along the upper Indo-Gangetic plains and their bacterial community composition was analyzed based on sequencing of the V3-V4 region of the 16S rRNA gene. Diversity analysis has shown significant variation in bacterial diversity among the sites. The taxonomic profile identified Proteobacteria, Chloroflexi, Actinobacteria, Bacteroidetes, Acidobacteria, Gemmatimonadetes, Planctomycetes, Verrucomicrobia, Firmicutes, and Cyanobacteria as the most dominant phyla in the wheat rhizosphere in the region. Core microbiota analysis revealed 188 taxa as core microbiota of wheat rhizosphere with eight genera recording more than 0.5% relative abundance. The order of most abundant genera in the core microbiota is Roseiflexus> Flavobacterium> Gemmatimonas> Haliangium> Iamia> Flavisolibacter> Ohtaekwangia> Herpetosiphon. Flavobacterium, Thermomonas, Massilia, Unclassified Rhizobiaceae, and Unclassified Crenarchaeota were identified as keystone taxa of the wheat rhizosphere. Correlation studies revealed, pH, organic carbon content, and contents of available nitrogen, phosphorus, and iron as the major factors driving bacterial diversity in the wheat rhizosphere. Redundancy analysis has shown the impact of different soil properties on the relative abundance of different genera of the core microbiota. The results of the present study can be used as a prelude to be developing microbial formulations based on core microbiota.

Influence of host genotype in establishing root associated microbiome of indica rice cultivars for plant growth promotion.

Rice plants display a unique root ecosystem comprising oxic-anoxic zones, harboring a plethora of metabolic interactions mediated by its root microbiome. Since agricultural land is limited, an increase in rice production will rely on novel methods of yield enhancement. The nascent concept of tailoring plant phenotype through the intervention of synthetic microbial communities (SynComs) is inspired by the genetics and ecology of core rhizobiome. In this direction, we have studied structural and functional variations in the root microbiome of 10 indica rice varieties. The studies on α and ß-diversity indices of rhizospheric root microbiome with the host genotypes revealed variations in the structuring of root microbiome as well as a strong association with the host genotypes. Biomarker discovery, using machine learning, highlighted members of class Anaerolineae, α-Proteobacteria, and bacterial genera like Desulfobacteria, Ca. Entotheonella, Algoriphagus, etc. as the most important features of indica rice microbiota having a role in improving the plant's fitness. Metabolically, rice rhizobiomes showed an abundance of genes related to sulfur oxidation and reduction, biofilm production, nitrogen fixation, denitrification, and phosphorus metabolism. This comparative study of rhizobiomes has outlined the taxonomic composition and functional diversification of rice rhizobiome, laying the foundation for the development of next-generation microbiome-based technologies for yield enhancement in rice and other crops.

Genetic Diversity and Anti-Oxidative Potential of Streptomyces spp. Isolated from Unexplored Niches of Meghalaya, India.

Streptomyces is genetically and functionally diverse genus known to produce a wide array of phenolics and flavonoids with significant biotechnological applications. 52 isolates belonging to 26 species of Streptomyces collected from Meghalaya, India were analyzed for their genetic diversity using BOX-PCR. Significant inter- and intra- generic diversity was observed among the Streptomyces isolates especially those belonging to S. cacaoi, S. lavendulae, S. olivochromogenes, S. aureus, S. flavovirens. During bioactivity screening of the isolates, S. rectiviolaceus MJM72 recorded the highest DPPH activity (77.13 ± 0.91%) whereas S. antimycoticus MSCA162 showed excellent ABTS radical scavenging activity (99.65 ± 0.41%). On the other hand, S. novaecaesareae MJM58 had the highest (756.4 ± 7.38 µg GAE g-1 fresh weight) phenolic content while S. rectiviolaceus MJM72 was recorded with the highest flavonoid content (69.3 ± 0.12 µg QE g-1 fresh weight). As compared to total flavonoid content, total phenolic content had a stronger correlation with antioxidant activities. HPLC analysis of five selected isolates showed presence of gallic acid and pyrocatechol as predominant phenolics. In case of flavonoids, three isolates showed presence of rutin with S. rochei MSCA130 having the highest rutin content (0.95 µg g-1 fresh weight). The results of this study showed high genetic diversity and antioxidant potential among the Streptomyces isolates.

Bacterial chitinases: genetics, engineering and applications.

Chitinases are a group of enzymes that catalyze chitin hydrolysis and are present in all domains of life. Chitinases belong to different glycosyl hydrolase families with great diversity in their sequences. Microorganisms such as bacteria and fungi produce chitinases for nutrition, and energy, and to parasitize the chitinous hosts. But chitinases from bacteria are of special interest due to their ubiquitous nature and ability to perform under extreme conditions. Chitinases produced by bacteria have been explored for their use in agriculture and industry. In agriculture, their main role is to control chitin-containing insect pests, fungal pathogens, and nematodes. In the seafood industry, they found their role in the management of processing wastes which are mainly chitinous substances. Chitinases are also used to synthesize low molecular weight chitooligomers which are proven bioactive compounds with activities such as anti-tumour, antimicrobial, and immunity modulation. Considering their importance in ecology and biotechnological applications, several bacterial chitinases have been studied in the last two decades. Despite their potential, bacterial chitinases have a few limitations such as low production and lack of secretion systems which make the wild-type enzymes unfit for their applications in industries and other allied sectors. This review is an attempt to collate significant works in bacterial chitinases and their application in various industries and the employment of various tools and techniques for improvement to meet industrial requirements.

Bio-priming with a consortium of Streptomyces araujoniae strains modulates defense response in chickpea against Fusarium wilt.

Wilt caused by Fusarium oxysporum f. sp. ciceris (Foc) is one of the major diseases of chickpea affecting the potential yield significantly. Productivity and biotic stress resilience are both improved by the association and interaction of Streptomyces spp. with crop plants. In the present study, we evaluated two Streptomyces araujoniae strains (TN11 and TN19) for controlling the wilt of chickpea individually and as a consortium. The response of Foc challenged chickpea to inoculation with S. araujoniae TN11 and TN19 individually and as a consortium was recorded in terms of changes in physio-biochemical and expression of genes coding superoxide dismutase (SOD), peroxidase, and catalase. Priming with a consortium of TN11 and TN19 reduced the disease severity by 50-58% when challenged with Foc. Consortium primed-challenged plants recorded lower shoot dry weight to fresh weight ratio and root dry weight to fresh weight ratio as compared to challenged non-primed plants. The pathogen-challenged consortium primed plants recorded the highest accumulation of proline and electrolyte leakage. Similarly, total chlorophyll and carotenoids were recorded highest in the consortium treatment. Expression of genes coding SOD, peroxidase, and catalase was up-regulated which corroborated with higher activities of SOD, peroxidase, and catalase in consortium primed-challenged plants as compared to the challenged non-primed plants. Ethyl acetate extracts of TN11 and TN19 inhibited the growth of fungal pathogens viz., Fusarium oxysporum f. sp. ciceris. Macrophomina phaseolina, F. udum, and Sclerotinia sclerotiarum by 54-73%. LC-MS analyses of the extracts showed the presence of a variety of antifungal compounds like erucamide and valinomycin in TN11 and valinomycin and dinactin in TN19. These findings suggest that the consortium of two strains of S. araujoniae (TN11 and TN19) can modulate defense response in chickpea against wilt and can be explored as a biocontrol strategy.

Diversity of Bacterial Endophytes of Maize (Zea mays) and Their Functional Potential for Micronutrient Biofortification.

Microorganisms due, to their immense metabolic diversity, have the potential to augment the uptake of iron and zinc in addition to other important nutrients in plants. In the present work, 129 different strains of endophytic bacteria were retrieved from stems and leaves of maize. Qualitative screening of these endophytes showed that 24.5% of these isolates were siderophore producers, while 14% could solubilize insoluble zinc compounds and 33% of them had phytase activity. Based on zinc solubilization efficiency and siderophore production ability, 10 isolates each from zinc solubilizers and siderophore producers were selected. Molecular identification indicated that the selected bacteria belonged to diverse genera Microbacterium, Pseudonocardia, Bacillus, Cellulosimicrobium, Staphylococcus, Luteimonas, Bordetella, Brevundimonas, Streptomyces, Cupriavidus, Sphingomonas, Ralstonia, Ochrobactrum, Conyzicola, Paenibacillus and Leifsonia. Quantitative analyses of Zn solubilization using Atomic absorption spectrophotometry (AAS) revealed that Microbacterium hydrothermale M10 and M. proteolyticum B2 were potential solubilizers of different forms of insoluble zinc compounds viz. ZnCO3 (56.63-89.88 ppm), ZnO (106.38-120.08 ppm) and ZnS (3.62-5.56 ppm). Similarly, quantitative estimation of siderophore production activity revealed two endophytes viz. Bacillus altitudinis C7 (97.25% siderophore units) and Pseudonocardia alni M29 (92.05% siderophore units) as potential siderophore producers. These endophytes with potential to produce siderophores and phytases and ability to solubilize zinc can be an important starting material for trials on field to improve Fe and Zn content in edible portion of food crops.

Genomic Diversity of Pigeon Pea (Cajanus cajan L. Millsp.) Endosymbionts in India and Selection of Potential Strains for Use as Agricultural Inoculants.

Pigeon pea (Cajanus cajan L. Millsp. ) is a legume crop resilient to climate change due to its tolerance to drought. It is grown by millions of resource-poor farmers in semiarid and tropical subregions of Asia and Africa and is a major contributor to their nutritional food security. Pigeon pea is the sixth most important legume in the world, with India contributing more than 70% of the total production and harbouring a wide variety of cultivars. Nevertheless, the low yield of pigeon pea grown under dry land conditions and its yield instability need to be improved. This may be done by enhancing crop nodulation and, hence, biological nitrogen fixation (BNF) by supplying effective symbiotic rhizobia through the application of "elite" inoculants. Therefore, the main aim in this study was the isolation and genomic analysis of effective rhizobial strains potentially adapted to drought conditions. Accordingly, pigeon pea endosymbionts were isolated from different soil types in Southern, Central, and Northern India. After functional characterisation of the isolated strains in terms of their ability to nodulate and promote the growth of pigeon pea, 19 were selected for full genome sequencing, along with eight commercial inoculant strains obtained from the ICRISAT culture collection. The phylogenomic analysis [Average nucleotide identity MUMmer (ANIm)] revealed that the pigeon pea endosymbionts were members of the genera Bradyrhizobium and Ensifer. Based on nodC phylogeny and nod cluster synteny, Bradyrhizobium yuanmingense was revealed as the most common endosymbiont, harbouring nod genes similar to those of Bradyrhizobium cajani and Bradyrhizobium zhanjiangense. This symbiont type (e.g., strain BRP05 from Madhya Pradesh) also outperformed all other strains tested on pigeon pea, with the notable exception of an Ensifer alkalisoli strain from North India (NBAIM29). The results provide the basis for the development of pigeon pea inoculants to increase the yield of this legume through the use of effective nitrogen-fixing rhizobia, tailored for the different agroclimatic regions of India.

Halomonas icarae sp. nov., a moderately halophilic bacterium isolated from beach soil in India.

A moderately halophilic, Gram-stain-negative, aerobic bacterium, strain D1-1T, belonging to the genus Halomonas, was isolated from soil sampled at Pentha beach, Odisha, India. Phylogenetic trees reconstructed based on 16S rRNA genes and multilocus sequence analysis of gyrB and rpoD genes revealed that strain D1-1T belonged to the genus Halomonas and was most closely related to Halomonas alimentaria YKJ-16T (98.1 %) followed by Halomonas ventosae Al12T (97.5 %), Halomonas sediminicola CPS11T (97.5 %), Halomonas fontilapidosi 5CRT (97.4 %) and Halomonas halodenitrificans DSM 735T (97.2 %) on the basis of 16S rRNA gene sequence similarity. Sequence identities with other species within the genus were lower than 97.0 %. The digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) values of 22.4-30 % and 79.5-85.4 % with close relatives of H. halodenitrificans DSM 735T, H. alimentaria YKJ-16T, H. ventosae Al12T and H. fontilapidosi 5CRT were lower than the threshold recommended for species delineation (70 % and 95-96 % for dDDH and ANI, respectively). Further, strain D1-1T formed yellow-coloured colonies; cells were rod-shaped, motile with optimum growth at 30 °C (range, 4-45 °C) and 2-8 % NaCl (w/v; grew up to 24 % NaCl). The major fatty acids were summed feature 8 (C18 : 1 ω7c/C18 : 1 ω6c), summed feature 3 (C16 : 1 ω7c/C16 : 1 ω6c) and C16 : 0 and the main respiratory quinone was ubiquinone Q-9 in line with description of the genus. Based on its chemotaxonomic and phylogenetic characteristics and genome uniqueness, strain D1-1T represents a novel species in the genus Halomonas, for which we propose the name Halomonas icarae sp. nov., within the family Halomonadaceae. The type strain is D1-1T (=JCM 33602T=KACC 21317T=NAIMCC-B-2254T).

Spatial distribution and identification of bacteria in stressed environments capable to weather potassium aluminosilicate mineral.

In the present study, we studied the distribution of silicate mineral weathering bacteria (SWB) in stressed environments that release potassium from insoluble source of mineral. Out of 972 isolates, 340 isolates were positive and mineral weathering potential ranged from 5.55 to 180.05%. Maximum abundance of SWB occurred 44.71% in saline environment followed by 23.53% in low temperature and 12.35% each in high temperature and moisture deficit. Among isolates, silicate mineral weathering efficiency ranged from 1.9 to 72.8 µg mL-1 available K in liquid medium. The phylogenetic tree of SWB discriminated in three clusters viz. Firmicutes, Proteobacteria and Actinobacteria. This is the first report on SWB in stressed environments and identified 27 genera and 67 species which is not reported earlier. Among them Bacillus was the predominant genera (58.60%) distantly followed by Pseudomonas (6.37%), Staphylococcus (5.10%) and Paenibacillus (4.46%). These bacterial strains could be developed as inoculants for biological replenishment of K in stressed soils. Graphical abstract.

Sustainable use of the spent mushroom substrate of Pleurotus florida for production of lignocellulolytic enzymes.

Spent mushroom substrate (SMS), a major byproduct of the mushroom industry, is a lignocellulosic biomass, which contains approximately 57-74.3% of holocellulose fraction. This study was aimed at utilizing SMS of Pleurotus florida for recovery of lignocellulolytic enzymes and sugars and also as a substrate for production of cellulolytic enzymes using different isolates of Trichoderma and Aspergillus under solid-state fermentation (SSF). SMS of P. florida extracts contained significant amounts of laccase (3,015.8 ± 29.5 U/g SMS) and xylanase (1,187.9 ± 12 U/g SMS) activity. Crystallinity pattern and chemical changes in SMS revealed that SMS had a lower crystallinity index (34.2%) as compared with the raw biomass (37.8%), which, in turn, helps in enhancing the accessibility of cellulolytic enzymes to holocellulose. Among the isolates, Trichoderma longibrachiatum A-01 showed maximum activity of endoglucanase (220.4 ± 5.9 U/mg), exoglucanase (78.5 ± 3.2 U/mg) and xylanase (1,550.4 ± 11.6 U/mg) while Aspergillus aculeatus C-08 showed maximum activity of cellobiase (113.9 ± 3.9 U/mg). Extraction with sodium citrate buffer (pH 4.8) showed maximum cellulolytic enzyme activity as compared with other solvents tested. Partial purification of endoglucanase, exoglucanase, xylanase, and cellobiase resulted in 56.3% (1,112.5 U/mg), 48.4% (212.5 U/mg), 44% (4,492.3 U/mg), and 62% (705.0 U/mg) yield with an increase by 5.2-, 4.5-, 4.1-, and 5.0-fold as compared with crude extract. The results reveal that SMS from P. florida could be a potential and cost-effective substrate for production of cellulolytic enzymes from T. longibrachiatum A-01 and A. aculeatus C-08.

Interaction between arbuscular mycorrhizal fungi and Bacillus spp. in soil enhancing growth of crop plants.

Soil microorganisms play an important role in enhancing soil fertility and plant health. Arbuscular mycorrhizal fungi and plant growth promoting rhizobacteria form a key component of the soil microbial population. Arbuscular mycorrhizal fungi form symbiotic association with most of the cultivated crop plants and they help plants in phosphorus nutrition and protecting them against biotic and abiotic stresses. Many species of Bacillus occurring in soil are also known to promote plant growth through phosphate solubilization, phytohormone production and protection against biotic and abiotic stresses. Synergistic interaction between AMF and Bacillus spp. in promoting plant growth compared to single inoculation with either of them has been reported. This is because of enhanced nutrient uptake, protection against plant pathogens and alleviation of abiotic stresses (water, salinity and heavy metal) through dual inoculation compared to inoculation with either AMF or Bacillus alone.

Mutagenicity, genotoxicity and oxidative stress induced by pesticide industry wastewater using bacterial and plant bioassays.

Atomic absorption spectrophotometer and gas chromatography analysis revealed the presence of heavy metals, organochlorine and organophosphate pesticides in industrial wastewater. XAD, Dichloromethane and n-Hexane extracted wastewater were analysed for genotoxic potential using Ames Salmonella/mammalian microsome test. The XAD concentrated sample displayed remarkable mutagenic activity compared to solvent assisted liquid-liquid extraction. Strain TA98 was found utmost sensitive towards all extracts. Wastewater induced chromosomal aberrations in roots of Allium cepa showed significant (p < 0.05) decrease in mitotic index. Seeds of Vigna radiata germinated on soft agar plates treated with different concentration of wastewater showed significant reduction in germination (52 %), seedling vigor index (76 %), radicle length (56 %), plumule length (47 %), biomass of radicle (64 %) and plumule (57%) at highest wastewater concentration. Propidium iodide stained V. radiata roots showed oxidative stress induced by wastewater under CLS microscopy. Further, genotoxicity of wastewater was confirmed by plasmid nicking assay using pBR322 plasmid.

Draft Genome Sequence of Halotolerant Bacterium Chromohalobacter salexigens ANJ207, Isolated from Salt Crystal Deposits in Pipelines.

Chromohalobacter salexigens ANJ207 was isolated from a salt crystal and is known to tolerate up to 30% NaCl concentration. Here, we report the de novo draft assembly of C. salexigens ANJ207. The genome was determined to have 3.66 Mb represented in 13 scaffolds, with a total of 3,406 genes predicted.

Kitazin-pea interaction: understanding the fungicide induced nodule alteration, cytotoxicity, oxidative damage and toxicity alleviation by Rhizobium leguminosarum.

Realizing the severity of fungicidal toxicity to legumes and importance of fungicide tolerant rhizobia in legume production, kitazin tolerant (2400 µg mL-1) strain RP1 was recovered from pea nodules and was identified as Rhizobium leguminosarum (accession no. KY940047). R. leguminosarum produced indole acetic acid (80.5 ± 2.5 mL-1), siderophores: salicylic acid (54 ± 7.3 µg mL-1) and 2,3-dihydoxybenzoic acid (31.9 ± 2.7 µg mL-1), α-ketobutyrate (51 ± 3.2 per mg per protein per hour), solubilized insoluble phosphate (29.5 ± 1.8 µg mL-1) and secreted 29.5 + 2.6 µg mL-1 exopolysaccharides, which, however, decreased consistently with gradually increasing kitazin concentrations. Beyond the tolerance level, kitazin caused structural damage and altered membrane integrity of RP1, as revealed under scanning (SEM) and confocal (CLSM) electron microscopy. Phytotoxicity of kitazin to peas was obvious under both in vitro and in vivo conditions. A significant reduction of 23, 68, 57 and 50% in germination, seedling vigor index, plumule length and radicle length was found at 2× kitazin compared to the control. Cellular damage and cytotoxicity induced by kitazin in membrane altered root cells was detected with acridine orange/propidium iodide (AO/PI) and Evans blue dye. A maximum increase of 1.72, 5.2, 9.3 and 1.72, 5.2, 9.3-fold in red and blue fluorescence was quantified at 1×, 2×, and 3× doses of kitazin, respectively. In contrast, application of R. leguminosarum RP1 alleviated toxicity and enhanced the length of plant organs, dry biomass, symbiotic attributes, photosynthetic pigments, nutrient uptake and grain features of peas comparatively uninoculated and fungicide-treated plants. Additionally, strain RP1 expressively reduced the antioxidant enzymes peroxidase, ascorbate peroxidase, guaiacol peroxidase, catalase and malondialdehyde contents by 10, 2.2, 11, 20 and 4% compared to stressed plants raised at 192 µg kg-1 soil. Moreover, a decline of 19, 21 and 20% in proline content extracted from roots, shoots and grains, respectively was recorded for R. leguminosarum inoculated pea plants grown with 96 µg kg-1 kitazin. Also, the SEM and CLSM of roots revealed the bacterial colonization. In conclusion, R. leguminosarum tolerated a higher level of kitazin, secreted plant growth promoting (PGP) bioactive molecules even under fungicide stress and significantly increased the performance of peas while reducing the levels of proline and antioxidant enzymes. So, it can safely be suggested to legume growers that RP1 strain could inexpensively be explored as an efficient biofertilizer for enhancing the production of legumes especially peas while growing even under fungicide (kitazin) enriched soils.

First De Novo Draft Genome Sequence of the Pathogenic Fungus Fusarium udum F02845, Associated with Pigeonpea (Cajanus cajan L. Millspaugh) Wilt.

Fusarium udum F02845 is a destructive fungal pathogen which causes pigeonpea (Cajanus cajan L. Millspaugh) wilt. Here we report the first de novo draft assembly of Fusarium udum F02845, isolated from an infected pigeonpea stem. The genome was determined to be 56.38 Mb in size, with a G+C content of 42.44%, and predicted to have 712 scaffolds with a total number of 11,829 genes.

Bacterial xylanases: biology to biotechnology.

In this review, a comprehensive discussion exclusively on bacterial xylanases; their gene organization; different factors and conditions affecting enzyme yield and activity; and their commercial application have been deliberated in the light of recent research findings and extensive information mining. Improved understanding of biological properties and genetics of bacterial xylanase will enable exploitation of these enzymes for many more ingenious biotechnological and industrial applications.

Cloning and expression of dnaK gene from Bacillus pumilus of hot water spring origin.

A set of thermotolerant strains isolated from hot springs of Manikaran and Bakreshwar (India) were selected with an aim to isolate dnak gene which encodes DnaK protein. The gene dnaK along with its flanking region was successfully amplified from 5 different strains (4 from Bakreshwar and one from Manikaran). Restriction fragment length polymorphism (RFLP) revealed that amplicons were almost identical in sequence. The dnak gene from one representative, Bacillus pumilus strain B3 isolated from Bakreshwar hot springs was successfully cloned and sequenced. The dnaK gene was flanked by gene grpE on one side. The dnaK gene was 1842 bp in length encoding a polypeptide of 613 amino acid residues. Calculated molecular weight and pI of the protein were 66,128.36 Da and 4.72 respectively. The deduced amino acid sequence of this gene shared high sequence homology with other DnaK proteins and its homologue Hsp 70 from other microorganisms, but possessed 36 substitutions and two insertions, as compared to DnaK protein of Bacillus subtilis. The dnaK gene of B. pumilus was successfully expressed in Escherichia coli BL 21 (DE3) using pET expression systems. Heterologous expression of dnaK of B. pumilus in E. coli BL 21 (DE3) allowed for the growth of E. coli up to 50 °C and survival up to 60 °C for 16 h, suggesting that dnak from B. pumilus imparts tolerance to host cells under high temperature. This novel gene can be an important component for possible utilization in abiotic stress management of plants.