Comparative seasonal analysis of Eri silkworm (Samia ricini Donovan) gut composition: implications for lignocellulose degradation.

Conversion of biomass such as lignocelluloses to an alternative energy source can contribute to sustainable development. Recently, biomass-degrading enzymes are reported to be common resources in insect-microbe interacting systems. Northeast India harbors ample sericigenous insect resources which are exploited for their silk products. Samia ricini Donovan is an economically important poly-phytophagous silkmoth capable of digesting foliage from different plant species, suggesting the versatility of a robust gut system. Here, a gut bacterial profile was determined by 16S rRNA gene characterization across the holometabolous life cycle during the summer and winter seasons, revealing 3 phyla, 13 families, and 22 genera. Comparative analysis among the seasonal gut isolates revealed a high diversity in summer, predominated by the genus Bacillus due to its high occurrence in all developmental stages. Shannon's diversity index demonstrated the second and fourth instars of summer as well as the fifth instar of winter to be relatively better developmental stages for gut bacteria assembly. Bacterial community shifts in concert to host developmental changes were found to be apparent between early instars and late instars in summer, which differed from those of winter. Forty-three and twenty-nine gut bacterial isolates were found to be cellulolytic and xylanolytic enzyme producers, respectively. The present results illustrate the gut microbiota of S. ricini over the seasons and support the holometabolous life cycle effect as the most likely factor shaping the gut bacterial microbiota. These findings may provide leads for the development of new cleaner and environmentally friendly lignocellulose-degrading enzymes.

Occurrence, identification and characterization of diazotrophic bacteria from aerial roots of Rhynchostylis retusa (L.) Blume for plant growth-promoting activity.

In this study, the diversity of diazotrophic bacteria of orchid Rhynchostylis retusa (L.) Blume and its potential application in plant growth promotion were evaluated. About 183 nitrogen-fixing bacteria were isolated to screen various plant growth-promoting traits viz. phosphate solubilization,IAA, siderophore, HCN, biofilm and ammonia production. Based on 16S rRNA gene sequencing analysis Achromobacter, Arthrobacter, Acinetobacter, Bacillus, Brevibacterium, Curtobacterium, Erwinia, Kosakonia, Lysinibacillus, Klebseilla, Microbacterium, Mixta, Pantoea, Pseudomonas and Stenotrophomonas isolates were selected and showed positive results for PGP traits. Overall, genus Pantoea, Brevibacterium, Achromobacter, Arthrobacter, Klebsiella, Mixta, Bacillus, and Pseudomonas had the most pronounced PGP characteristics and acetylene reduction among the screened isolates. BOX PCR fingerprinting analysis showed variation in polymorphic banding patterns among diazotrophic strains. PCR amplification of nifH gene and the presence of 37 kDa nitrogenase reductase enzyme band in western blot indicated presence of nitrogenase activity. Our study showed that orchid R. retusa diazotroph interaction helps orchid plant to fix nitrogen, essential nutrients, and control pathogen entry. To the best of our knowledge, this is the first report on characterization of diazotrophic bacterial community from aerial roots of R. retusa.

Immunogenicity and cross-protective efficacy of recombinant H5HA1 protein of clade 2.3.2.1a highly pathogenic H5N1 avian influenza virus expressed in E.coli.

The global spread of H5N1 highly pathogenic avian influenza virus (HPAIV) in poultry has caused great economic loss to the poultry farmers and industry with significant pandemic threat. The current study involved production of recombinant HA1 protein of clade 2.3.2.1a H5N1 HPAIV (rH5HA1) in E.coli and evaluation of its protective efficacy in chickens. Purification under denaturing conditions and refolding by dialysis against buffers containing decreasing concentrations of urea was found to preserve the biological activity of the expressed recombinant protein as assessed by hemagglutination assay, Western blot and ELISA. The Montanide ISA 71 VGA adjuvanted rH5HA1 protein was used for immunization of chickens. Humoral response was maintained at a minimum of 4log2 hemagglutination inhibition (HI) titre till 154 days post 2nd booster. We evaluated the protective efficacy of rH5HA1 protein in immunized chickens by challenging them with homologous (2.3.2.1a) and heterologous (2.3.2.1c) clades of H5N1 HPAIV. In both the groups, the HI titre significantly increased (P < 0.05) after challenge and the virus shedding significantly (P < 0.05) reduced between 3rd and 14th day post challenge. The virus shedding ratio in oro-pharyngeal swabs did not differ significantly between both the groups except on 7 days post challenge and during the entire experimental period in cloacal swabs. These results indicate that rH5HA1 was able to induce homologous and cross protective immune response in chickens and could be a potential vaccine candidate used for combating the global spread of H5N1 HPAIV threat. To our knowledge, this is the first study to report immunogenicity and protective efficacy of prokaryotic recombinant H5HA1 protein in chicken.