Stalk rot species diversity and molecular phylogeny associated with diseased maize in India.

Stalk rot disease is a major constraint in maize production and till date reported to be caused by two to three species of phytopathogenic fungi but, in our present study, we disclose the first report of stalk rot is caused by complex species of phytopathogens, which belongs to five different genera. Therefore, to substantiate these findings, a total of 105 diseased samples of maize were collected from 21 different locations in six different geographical locations of India from which 48 isolates were used for the research study. Morphological features such as pigmentation, colony color, type of mycelium and pattern of mycelium was examined using macro and microscopic methods. A total of 11 different spp. of pathogens belonging to the five different genera: Fusarium verticillioides (56.25%), F. equiseti (14.5%), F. andiyazi (6.25%), F. solani (2.08%), F. proliferatum (2.08%), F. incarnatum (2.08%), Lasidioplodia theobrame (6.25%), Exserohilum rostrtum (4.16%), Nigrospora spp. (4.16%). and Schizophyllum commune (2.08%) were identified by different housekeeping genes (ITS, TEF-1α, RPB2 and Actin). Fusarium verticillioides, F. equiseti and F. andiyazi were major pathogens involved in stalk rot. This is the first report on F. proliferatum, F. solani, F. incarnatum, Lasidioplodia theobrame, Exserohilum rostrtum, Nigrospora spp. and Schizophyllum commune causing stalk rot of maize and their distribution in the different states of India. Studies on population dynamics of PFSR will enhance the understanding of pathogen behavior, virulence, or its association with different pathogens across India, which will facilitate the development of resistant maize genotypes against the PFSR.

Rice-Magnaporthe transcriptomics reveals host defense activation induced by red seaweed-biostimulant in rice plants.

Red seaweed extracts have been shown to trigger the biotic stress tolerance in several crops. However, reports on transcriptional modifications in plants treated with seaweed biostimulant are limited. To understand the specific response of rice to blast disease in seaweed-biostimulant-primed and non-primed plants, transcriptomics of a susceptible rice cultivar IR-64 was carried out at zero and 48 h post inoculation with Magnaporthe oryzae (strain MG-01). A total of 3498 differentially expressed genes (DEGs) were identified; 1116 DEGs were explicitly regulated in pathogen-inoculated treatments. Functional analysis showed that most DEGs were involved in metabolism, transport, signaling, and defense. In a glass house, artificial inoculation of MG-01 on seaweed-primed plants resulted in the restricted spread of the pathogen leading to the confined blast disease lesions, primarily attributed to reactive oxygen species (ROS) accumulation. The DEGs in the primed plants were defense-related transcription factors, kinases, pathogenesis-related genes, peroxidases, and growth-related genes. The beta-D-xylosidase, a putative gene that helps in secondary cell wall reinforcement, was downregulated in non-primed plants, whereas it upregulated in the primed plants indicating its role in the host defense. Additionally, Phenylalanine ammonia-lyase, pathogenesis-related Bet-v-I family protein, chalcone synthase, chitinases, WRKY, AP2/ERF, and MYB families were upregulated in seaweed and challenge inoculated rice plants. Thus, our study shows that priming rice plants with seaweed bio-stimulants resulted in the induction of the defense in rice against blast disease. This phenomenon is contributed to early protection through ROS, protein kinase, accumulation of secondary metabolites, and cell wall strengthening.

Bacillus velezensis (strains A6 & P42) as a potential biocontrol agent against Klebsiella variicola, a new causal agent of soft rot disease in carrot.

Bacterial soft rot is one of the most devastating diseases and a major constraint encountered during carrot farming. Biological agents are the best eco-friendly alternatives to agrochemicals to manage soft rot disease to ensure environmental sustainability. In this study, about eight isolates of bacterial pathogen causing soft rot in carrots were collected from Karnataka, India. Based on the 16S rRNA sequencing the pathogen isolates causing soft rot of carrot were identified as Klebsiella variicola. The morphological characteristics of K. variicola was investigated under scanning electron microscopy. The pathogenicity assay showed that all eight isolates were pathogenic to the carrot. An in vitro and in planta assay of two novel strains of Bacillus velezensis (A6 and P42) against K. variicola indicated that both strains had strong antagonistic activity against all the pathogen strains. Furthermore, the volatile bioactive compounds produced by A6 and P42 strains were analyzed in GC-MS, which revealed the presence of 10 and 6 bioactive compounds in their culture filtrate, respectively, with antibacterial and antifungal properties. The present study suggests that both A6 and P42 strains of B. velezensis were antagonistic to K. variicola and can be used as biocontrol agents to manage soft rot diseases of carrot under field conditions.

Red-seaweed biostimulants differentially alleviate the impact of fungicidal stress in rice (Oryza sativa L.).

Red seaweed-derived biostimulants facilitate plant health and impart protection against abiotic stress conditions by their bioactive compounds and plant nutrients. The potency of red seaweed biostimulants (LBS6 and LBD1) on rice cv. IR-64 in response to fungicides induced stress was investigated in this study. Foliar application of LBS6 maintained the stomatal opening and leaf temperature under the fungicidal stress condition. Reactive Oxygen Species (ROS) such as hydrogen peroxide and superoxide radicals were significantly reduced in LBS6-treated stressed plants. After applying seaweed biostimulants, ROS production was stabilized by antioxidants viz., CAT, APX, SOD, POD, and GR. LBS-6 application increased the Ca+ and K+ levels in the stressed plants, which perhaps interacted with ROS and stomatal opening signalling systems, respectively. In the rice plants, fungicidal stress elevated the expression of stress-responsive transcriptional factors (E2F, HSFA2A, HSFB2B, HSFB4C, HSFC1A, and ZIP12). A decline in the transcript levels of stress-responsive genes was recorded in seaweed treated plants. For the first time, we present an integrative investigation of physicochemical and molecular components to describe the mechanism by which seaweed biostimulants in rice improve plant health under fungicidal stress conditions.

Diversity and biopotential of Bacillus velezensis strains A6 and P42 against rice blast and bacterial blight of pomegranate.

Bacillus velezensis is widely known for its inherent biosynthetic potential to produce a wide range of bio-macromolecules and secondary metabolites, including polyketides (PKs) and siderophores, as well as ribosomally and non-ribosomally synthesized peptides. In the present study, we aimed to investigate the bio-macromolecules, such as proteins and peptides of Bacillus velezensis strains, namely A6 and P42 by whole-cell sequencing and highlighted the potential application in controlling phytopathogens. The bioactive compounds, specifically secondary metabolites, were characterized by whole-cell protein profiling, Thin-Layer Chromatography, Infra-Red Spectroscopy, Nuclear Magnetic Resonance, Gas Chromatograph and Electro Spray Liquid Chromatography. Gas Chromatography analysis revealed that the A6 and P42 strains exert different functional groups of compounds, such as aromatic ring, aliphatic, alkene, ketone, amine groups and carboxylic acid. Whole-cell protein profiling of A6 and P42 strains of B. velezensis by nano-ESI LC-MS/MS revealed the presence of 945 and 5303 proteins, respectively. The in vitro evaluation of crude extracts (10%) of A6 and P42 significantly inhibited the rice pathogen, Magnaporthe oryzae (MG01), whereas the cell-free culture filtrate (75%) of strain P42 showed 58.97% inhibition. Similarly, in vitro evaluation of crude extract (10%) of P42 strain inhibited bacterial blight of pomegranate pathogen, Xanthomonas axonopodis pv. punicae, which eventually resulted in a higher inhibition zone of 3 cm, whereas the cell-free extract (75%) of the same strain significantly suppressed the growth of the pathogen with an inhibition zone of 1.48 cm. From the results obtained, the crude secondary metabolites and cell-free filtrates (containing bio-macromolecules) of the strains A6 and P42 of B. velezensis can be employed for controlling the bacterial and fungal pathogens of crop plants.

Comparative metagenomic analysis of rice soil samples revealed the diverse microbial population and biocontrol organisms against plant pathogenic fungus Magnaporthe oryzae.

Intensive cropping degrades soil quality and disrupts the soil microbiome. To understand the effect of rice monocropping on soil-microbiome, we used a comparative 16S rRNA metagenome sequencing method to analyze the diversity of soil microflora at the genomic level. Soil samples were obtained from five locations viz., Chamarajnagara, Davangere, Gangavathi, Mandya, and Hassan of Karnataka, India. Chemical analysis of soil samples from these locations revealed significant variations in pH (6.00-8.38), electrical conductivity (0.17-0.69 dS m-1), organic carbon (0.51-1.29%), available nitrogen (279-551 kg ha-1), phosphorous (57-715 kg ha-1) and available potassium (121-564 kg ha-1). The 16S metagenome analysis revealed that the microbial diversity in Gangavathi soil samples was lower than in other locations. The soil sample of Gangavathi showed a higher abundance of Proteobacteria (85.78%) than Mandya (27.18%). The Firmicutes were more abundant in Chamarajnagar samples (36.01%). Furthermore, the KEGG pathway study revealed enriched nitrogen, phosphorus, and potassium metabolism pathways in all soil samples. In terms of the distribution of beneficial microflora, the decomposers were more predominant than the nutrient recyclers such as nitrogen fixers, phosphorous mineralizers, and nitrifiers. Furthermore, we isolated culturable soil microbes and tested their antagonistic activity in vitro against a fungal pathogen of rice, Magnaporthe oryzae strain MG01. Six Bacillus sp. and two strains of Trichoderma harzianum showed higher antagonistic activity against MG01. Our findings indicate that metagenome sequencing can be used to investigate the diversity, distribution, and abundance of soil microflora in rice-growing areas. The knowledge gathered can be used to develop strategies for maintaining soil quality and crop conservation to increase crop productivity. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02783-y.

Metagenome sequencing of fingermillet-associated microbial consortia provides insights into structural and functional diversity of endophytes.

Endophytes confer unique ecological advantages to their host plants. In this study, we have characterized the diversity of endophytic consortia associated with the GPU-28 (GPU) and Udurumallige (UM) finger millet varieties, which are resistant and susceptible to the blast disease, respectively. Whole genome metagenome sequencing of GPU and UM helped to identify 1029 species (includes obligate endophytes) of microbiota. Among them, 385 and 357 species were unique to GPU and UM, respectively. Remaining 287 species were common to both the varieties. Actinobacteria and other plant-growth promoting bacteria were abundant in GPU as compared to UM. Functional annotation of genes predicted from genomes of endophytes associated with GPU variety showed that many genes had functional role in stress response, secondary metabolism, aromatic compounds, glutathione, and cysteine synthesis pathways as compared to UM. Based on in vitro and in planta studies, Bacillus cereus and Paenibacillus spp. were found to be effective in suppressing the growth of blast disease pathogen Magnaporthe grisea (strain MG03). In the future, these strains could serve as potential biocontrol agents to reduce the incidence of blast disease in finger millet crop.

Exploring the Potentiality of Novel Rhizospheric Bacterial Strains against the Rice Blast Fungus Magnaporthe oryzae.

Rice blast caused by Magnaporthe oryzae is a major disease. In the present study, we aimed to identify and evaluate the novel bacterial isolates from rice rhizosphere for biocontrol of M. oryzae pathogen. Sixty bacterial strains from the rice plant's rhizosphere were tested for their biocontrol activity against M. oryzae under in vitro and in vivo. Among them, B. amyloliquefaciens had significant high activity against the pathogen. The least disease severity and highest germination were recorded in seeds treated with B. amyloliquefaciens UASBR9 (0.96 and 98.00%) compared to untreated control (3.43 and 95.00%, respectively) under in vivo condition. These isolates had high activity of enzymes in relation to growth promoting activity upon challenge inoculation of the pathogen. The potential strains were identified based on 16S rRNA gene sequencing and dominance of these particular genes were associated in Bacillus strains. These strains were also confirmed for the presence of antimicrobial peptide biosynthetic genes viz., srfAA (surfactin), fenD (fengycin), spaS (subtilin), and ituC (iturin) related to secondary metabolite production (e.g., AMPs). Overall, the results suggested that application of potential bacterial strains like B. amyloliquefaciens UASBR9 not only helps in control of the biological suppression of one of the most devastating rice pathogens, M. grisea but also increases plant growth along with a reduction in application of toxic chemical pesticides.