Yogesh S. Shouche

Mountain-associated clade endemism in an ancient frog family (Nyctibatrachidae) on the Indian subcontinent

Night frogs (Nyctibatrachidae) form a family endemic to the Western Ghats, a hill chain along the west coast of southern India. Extant members of this family are descendants of a lineage that originated on the subcontinent during its longtime isolation in the Late Cretaceous. Because the evolutionary history of Nyctibatrachidae has always been tightly connected to the subcontinent, these tropically-adapted frogs are an ideal group for studying how patterns of endemism originated and evolved during the Cenozoic in the Western Ghats. We used a combined set of mitochondrial and nuclear DNA fragments to investigate the phylogenetic relationships of 120 ingroup specimens of all known species of Nyctibatrachidae. Our analyses indicate that, although this family had an early origin on the Indian subcontinent, the early diversification of extant nyctibatrachids happened only in the Eocene. Biogeographic analyses show that dispersal across the Palghat gap and Shencottah gap was limited, which led to clade endemism within mountain ranges of the Western Ghats. It is likely that multiple biota have been affected simultaneously by these prominent geographical barriers. Our study therefore further highlights the importance of considering the Western Ghats-Sri Lanka biodiversity hotspot as an assemblage of distinct mountain regions, each containing endemism and deserving attention in future conservation planning.

Bacterial diversity in different regions of gastrointestinal tract of Giant African Snail (Achatina fulica)

The gastrointestinal (GI) tract of invasive land snail Achatina fulica is known to harbor metabolically active bacterial communities. In this study, we assessed the bacterial diversity in the different regions of GI tract of Giant African snail, A. fulica by culture‐independent and culture‐dependent methods. Five 16S rRNA gene libraries from different regions of GI tract of active snails indicated that sequences affiliated to phylum γ‐Proteobacteria dominated the esophagus, crop, intestine, and rectum libraries, whereas sequences affiliated to Tenericutes dominated the stomach library. On phylogenetic analysis, 30, 27, 9, 27, and 25 operational taxonomic units (OTUs) from esophagus, crop, stomach, intestine, and rectum libraries were identified, respectively. Estimations of the total bacterial diversity covered along with environmental cluster analysis showed highest bacterial diversity in the esophagus and lowest in the stomach. Thirty‐three distinct bacterial isolates were obtained, which belonged to 12 genera of two major bacterial phyla namely γ‐Proteobacteria and Firmicutes. Among these, Lactococcus lactis and Kurthia gibsonii were the dominant bacteria present in all GI tract regions. Quantitative real‐time polymerase chain reaction (qPCR) analysis indicated significant differences in bacterial load in different GI tract regions of active and estivating snails. The difference in the bacterial load between the intestines of active and estivating snail was maximum. Principal component analysis (PCA) of terminal restriction fragment length polymorphism suggested that bacterial community structure changes only in intestine when snail enters estivation state.

Diversified diazotrophs associated with the rhizosphere of Western Indian Himalayan native red kidney beans (Phaseolus vulgaris L.)

Red kidney beans (RKBs) are one of the major components in the human diet of Western Indian Himalaya (WIH). Their cultivation in these habitats is strongly influenced by various biotic and abiotic stresses and therefore, there must be a selection of RKB associated microorganisms that are adapted to these harsh conditions. Seven cold adaptive diazotrophs from the same rhizosphere were isolated in our previous study to reveal the low-temperature associated proteins and mechanisms. However, the diversity and phylogenetic affiliations of these rhizosphere diazotrophs are still unknown. In this study, RKB rhizospheric soil from two different agro-ecosystems of WIH namely S1 (Chhiplakot, 30.70°N/80.30°E) and S2 (Munsyari, 30.60°N/80.20°E) were explored for the assessment of nitrogenase reductase gene (nifH) diversity by plating respective clone libraries SN1 and SN2. The RKB rhizosphere diazotroph assemblage was very diverse and apparently consists mainly of the genera Rhizobium, followed by unknown diazotrophic microorganisms. Deduced amino acid sequence analysis revealed the presence of diverse nifH sequences, affiliated with a wide range of taxa, encompassing members of the Proteobacteria, Actinobacteria and Firmicutes. Members of cyanobacteria, methanotrophs and archaea were also detected. To the best of our knowledge, this is the first major metagenomic effort that revealed the presence of diverse nitrogen-fixing microbial assemblages in indigenous RKB rhizospheric soil which can further be explored for improved crop yield/productivity.

ε-Caprolactam Utilization by Proteus sp. and Bordetella sp. Isolated From Solid Waste Dumpsites in Lagos State, Nigeria, First Report

The ε-caprolactam is the monomer of the synthetic non-degradable nylon-6 and often found as nonreactive component of nylon-6 manufacturing waste effluent. Environmental consequences of its toxicity to natural habitats and humans pose a global public concern. Soil samples were collected from three designated solid waste dumpsites, namely, Abule-Egba, Olusosun and Isheri-Igando in Lagos State, Nigeria. Sixteen bacteria isolated from these samples were found to utilize the ε-caprolactam as a sole source of carbon and nitrogen at concentration ofxa0≤20xa0gxa0l−1. The isolates were characterized using their 16S rRNA gene sequence and showed similarity with Pseudomonas sp., Proteus sp., Providencia sp., Corynebacterium sp., Lysinibacillus sp., Leucobacter sp., Alcaligenes sp. and Bordetella sp. Their optimal growth conditions were found to be at temperature range of 30 to 35xa0°C and pH range of 7.0–7.5. High Performance liquid chromatography analysis of the ε-caprolactam from supernatant of growth medium revealed that these isolates have potential to remove 31.6–95.7xa0% of ε-caprolactam. To the best of our knowledge, this study is first to report the ability of Proteus sp. and Bordetella sp. for ε-caprolactam utilization.

A new species of Scytonema isolated from Bilaspur, Chhattisgarh, India

Filamentous cyanobacterium (strain 10C‐PS) isolated from a fresh water body of Bilaspur, Chhattisgarh, India is being described as new species of the polyphyletic genus Scytonema. Phenotypic, molecular and phylogenetic characterization was performed and the combined results validated the strain as a new species. Careful observations of the filaments, presence of a distinctly textured sheath throughout the length of the trichome, differences in the shape and dimensions of the vegetative cells, and heterocytes provided reliable morphological signals that the strain differed from rest of the closely related species. Sequencing of the 16S rRNA gene showed 96.89% sequence similarity with Scytonema hofmanni PCC 7110 while rbcl and psbA sequencing showed 95% and 92% similarities with Scytonema hofmanni PCC 7110 and Nostoc sp. PCC 7524 respectively while the nifD gene sequence similarity was found to be 96% with Scytonema hofmanni PCC 7110. The PC‐IGS region was sequenced and concatenated cpcB, IGS and cpcA regions indicated 97% closest similarity with Scytonema sp. PCC 7110 and Scytonema bohnerii Ind24. Subsequent phylogenetic analyses gave a strong pattern of distinct clustering in case of all the molecular markers. The phenotypic, genetic and phylogenetic observations prove conclusively that the strain 10C‐PS is a new species in the genus Scytonema with the name proposed being Scytonema bilaspurensis.

Genomic and physiological analyses of an indigenous strain, Enterococcus faecium 17OM39

The human gut microbiome plays a crucial role in human health and efforts need to be done for cultivation and characterisation of bacteria with potential health benefits. Here, we isolated a bacterium from a healthy Indian adult faeces and investigated its potential as probiotic. The cultured bacterial strain 17OM39 was identified as Enterococcus faecium by 16S rRNA gene sequencing. The strain 17OM39 exhibited tolerance to acidic pH, showed antimicrobial activity and displayed strong cell surface traits such as hydrophobicity and autoaggregation capacity. The strain was able to tolerate bile salts and showed bile salt hydrolytic (BSH) activity, exopolysaccharide production and adherence to human HT-29 cell line. Importantly, partial haemolytic activity was detected and the strain was susceptible to the human serum. Genomics investigation of strain 17OM39 revealed the presence of diverse genes encoding for proteolytic enzymes, stress response systems and the ability to produce essential amino acids, vitamins and antimicrobial compound Bacteriocin-A. No virulence factors and plasmids were found in this genome of the strain 17OM39. Collectively, these physiological and genomic features of 17OM39 confirm the potential of this strain as a candidate probiotic.

Cultivable microbial diversity associated with cellular phones

A substantial majority of global population owns cellular phones independently to demographic factors like age, economic status, and educational attainment. In this study, we investigated the diversity of microorganisms associated with cellular phones of 27 individuals using cultivation-based methods. Cellular phones were sampled using cotton swabs and a total of 554 isolates representing different morphotypes were obtained on four growth media. Matrix-assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry could generate protein profiles for 527 isolates and species-level identification was obtained for 415 isolates. A dendrogram was constructed based on the protein profiles of the remaining isolates, to group 112 isolates under 39 different proteotypes. The representative strains of each group were selected for 16S rRNA gene and ITS region sequencing based identification. Staphylococcus, Bacillus, Micrococcus, and Pseudomonas were the most frequently encountered bacteria, and Candida, Aspergillus, Aureobasidium, and Cryptococcus were in case of fungi. At species-level the prevalence of Micrococcus luteus, Staphylococcus hominis, Staphylococcus epidermidis, Staphylococcus arlettae, Bacillus subtilis, and Candida parapsilosis was observed, most of these species are commensal microorganisms of human skin. UPGMA dendrogram and PCoA biplot generated based on the microbial communities associated with all cellular phones exhibited build-up of specific communities on cellular phones and the prevalence of objectionable microorganisms in some of the cellular phones can be attributed to the poor hygiene and sanitary practices. The study also revealed the impact of MALDI-TOF MS spectral quality on the identification results. Overall MALDI-TOF appears a powerful tool for routine microbial identification and de-replication of microorganisms. Quality filtering of MALDI-TOF MS spectrum, development of better sample processing methods and enriching the spectral database will improve the role of MALDI-TOF MS in microbial identifications.

A new species of Scytonema isolated from Bilaspur, Chhattisgarh, India using the polyphasic approach

A false branching cyanobacterium (strain 1F-PS) isolated from a fresh water body of Bilaspur (Chhattisgarh, India) is described here as a new species of the polyphyletic genus Scytonema. Morphological, ecological, molecular and phylogenetic evidence validated the strain as a new species. Observations of the filaments in different phases of growth, different levels of microscopic studies, the presence of a textured thin sheath throughout the length of the trichome, differences in the shape and dimensions of the vegetative cells and heterocytes and ecological attributes show that the strain differed from the rest of the closely related species. Sequencing of the 16S rRNA gene resulted in 99% similarity with Scytonema bilaspurensis and 97.07% sequence similarity with Scytonema hofmannii PCC 7110, while rbcl and psbA sequences showed 99 and 97% similarities with S. bilaspurensis, respectively. Phylogenetic assessments indicated a large phylogenetic distance and separate clustering of the strain 1F-PS for all the molecular markers.

Comparative genome analysis reveals key genetic factors associated with probiotic property in Enterococcus faecium strains

BackgroundEnterococcus faecium though commensal in the human gut, few strains provide a beneficial effect to humans as probiotics while few are responsible for the nosocomial infection. Comparative genomics of E. faecium can decipher the genomic differences responsible for probiotic, pathogenic and non-pathogenic properties. In this study, we compared E. faecium strain 17OM39 with a marketed probiotic, non-pathogenic non-probiotic (NPNP) and pathogenic strains.ResultsE. faecium 17OM39 was found to be closely related with marketed probiotic strain T110 based on core genome analysis. Strain 17OM39 was devoid of known vancomycin, tetracycline resistance and functional virulence genes. Moreover, E. faecium 17OM39 genome was found to be more stable due to the absence of frequently found transposable elements. Genes imparting beneficial functional properties were observed to be present in marketed probiotic T110 and 17OM39 strains. Genes associated with colonization and survival within gastrointestinal tract was also detected across all the strains.ConclusionsBeyond shared genetic features; this study particularly identified genes that are responsible for imparting probiotic, non-pathogenic and pathogenic features to the strains of E. faecium. Higher genomic stability, absence of known virulence factors and antibiotic resistance genes and close genomic relatedness with marketed probiotics makes E. faecium 17OM39 a potential probiotic candidate. The work presented here demonstrates that comparative genome analyses can be applied to large numbers of genomes, to find potential probiotic candidates.

Rare Biosphere in Human Gut: A Less Explored Component of Human Gut Microbiota and Its Association with Human Health

With the advent of next-generation sequencing technologies, we are able to ascertain the vast microbial diversity of various habitats. These studies have led to the discovery of presence and persistence of microbial communities that are extremely low in the abundance. These rare microbial taxa in soil and marine environments are known to act as seed-bank for the organisms that are waiting for the favorable conditions to make their move. The human body is one of the diverse ecosystems known to us supporting the growth of myriads of microbes in or on our body. Studies in the last decade or so have significantly contributed to our understanding of the involvement of these human-associated microbes with our health and diseases. So far, the studies on human gut microbiota have focused largely on dominated microbial species, and the information about the rare taxa associated with the human body is still lurking. This chapter provides evidence that microbes from the three domains of life (bacteria, archaea, and eukarya) existing in the gut have the rare components relevant to human health. Hence, we advocate the need for more studies concerning the rare taxa present in the human gut and their association with the general well-being.