Strain-Dependent Adhesion Variations of Shouchella clausii Isolated from Healthy Human Volunteers: A Study on Cell Surface Properties and Potential Probiotic Benefits.

The probiotic potential of Shouchella clausii is widely recognized, but little is known about its adhesive properties. Hence, this study aims to investigate the adhesion potential and cell surface properties of four human-origin S. clausii strains (B619/R, B603/Nb, B106, and B637/Nm). We evaluated epithelial adhesion, Extracellular Matrix (ECM) binding, aggregation ability, and cell surface hydrophobicity and used genome analysis for validation. Our results demonstrate that adhesion capability is a strain-specific attribute, with significant variations observed among the four strains. B619/R, B603/Nb, and B106 displayed stronger adhesion properties than B637/Nm. Supplementary adhesion assays showed that B637/Nm displayed high hydrophobicity, significant auto-aggregation, and significant mucin-binding abilities. Conversely, B619/R, B603/Nb, and B106 had mildly hydrophobic surfaces and low aggregation abilities. Genome annotation revealed the presence of various adhesion proteins in four strains. Notably, the reduced adhesion potential of B637/Nm was supported by the absence of the cell wall surface anchor family protein (LPxTG motif), which is crucial for interactions with intestinal epithelial cells or mucus components. Further, docking studies provided insights into the interaction of adhesion proteins with gut mucins. These findings contribute to a better understanding of how S. clausii strains interact with the gut environment, facilitating the development of probiotic formulations tailored for improved gut health and well-being.

Unveiling the Probiotic Potential of Streptococcus thermophilus MCC0200: Insights from In Vitro Studies Corroborated with Genome Analysis.

Streptococcus thermophilus is widely used as a starter culture in the dairy industry and has garnered attention as a beneficial bacterium owing to its health-promoting functionalities in humans. In this study, the probiotic potential of S. thermophilus MCC0200 isolated from a dairy product was investigated through a combinatorial approach of in vitro and in silico studies. MCC0200 demonstrated the ability to survive harsh gastrointestinal (GI) transit, adhere to intestinal mucosa and exert health-promoting traits in in vitro studies. These findings were corroborated with in silico evidence, wherein, MCC0200 genome harboured genes associated with tolerance to GI conditions, intestinal adhesion and colonization. Genome mapping also highlighted the ability of MCC0200 to produce compounds advantageous for the host (folate, bacteriocins), to release antioxidant enzymes that can quench the free radicals (superoxide dismutase, NADH peroxidase), and to metabolize food components that can be harmful to sensitive people (lactose). MCC0200 also demonstrated a positive effect on reducing cholesterol levels, proving to be a potential candidate for food and pharmaceutical applications. The absence of transmissible antibiotic resistance genes and virulence genes underscored the generally regarded as safe (GRAS) nature of MCC0200. This study explored the potential of Streptococcus thermophilus for its probable applications as a probiotic beyond the dairy industry.

Phylogenetically and physiologically diverse methanogenic archaea inhabit the Indian hot spring environments.

Mesophilic and thermophilic methanogens belonging to the hydrogenotrophic, methylotrophic, and acetotrophic groups were isolated from Indian hot spring environments using BY and BCYT growth media. Following initial Hinf I-based PCR-RFLP screening, 70 methanogens were sequenced to ascertain their identity. These methanogens were phylogenetically and physiologically diverse and represented different taxa distributed across three physiological groups, i.e., hydrogenotrophs (53), methylotrophs (14) and acetotrophs (3). Overall, methanogens representing three families, five genera, and ten species, including two putative novel species, were recognized. The highest number and diversity of methanogens was observed at 40 â„ƒ, dominated by Methanobacterium (10; 3 species), Methanosarcina (9; 3 species), Methanothermobacter (7; 2 species), Methanomethylovorans (5; 1 species) and Methanoculleus (3; 1 species). Both putative novel methanogen species were isolated at 40 â„ƒ and belonged to the genera Methanosarcina and Methanobacterium. At 55 â„ƒ, limited diversity was observed, and resulted in the isolation of only two genera of methanogens, i.e., Methanothermobacter (28; 2 species) and Methanosarcina (4; 1 species). At 70 â„ƒ, only members of the genus Methanothermobacter (5; 2 species) were isolated, whereas no methanogen could be cultured at 85 â„ƒ. Ours is the first study that documents the extensive range of cultivable methanogenic archaea inhabiting hot springs across various geothermal provinces of India.

Description of Sporanaerobium hydrogeniformans gen. nov., sp. nov., an obligately anaerobic, hydrogen-producing bacterium isolated from Aravali hot spring in India.

An obligately anaerobic bacterium XHS1971T, capable of degrading cellulose and xylan, was isolated from a sediment sample of Aravali hot spring, Ratnagiri, India. Cells of strain XHS1971T were Gram-stain-negative, spore-forming, motile, long-rods. Growth was observed at temperatures 30-50 °C (optimum 40-45 °C), pH 5.0-10.0 (optimum pH 8.0) and NaCl concentrations 0-0.5% (optimum 0%). Generation time of strain XHS1971T was 5 h under optimised growth conditions. Strain XHS1971T showed the ability to metabolise different complex and simple sugars constituting lignocellulosic biomass. Glucose was fermented majorly into hydrogen, formic acid, acetic acid, and ethanol, whereas carbon dioxide, butyric acid, lactic acid and succinic acid were produced in traces. 16S rRNA gene analysis of strain XHS1971T revealed < 94.5% homology with Cellulosilyticum lentocellum DSM5427T followed by Cellulosilyticum ruminicola JCM14822T, identifying strain as a distinct member of family Lachnospiraceae. The major cellular fatty acids (> 5%) were C14:0, C16:0, C18:0, and C16:1 ω7c. The genome size of the strain was 3.74 Mb with 35.3 mol% G + C content, and genes were annotated to carbohydrate metabolism, including genes involved in the degradation of cellulose and xylan and the production of hydrogen, ethanol and acetate. The uniqueness of strain was further validated by digital DNA-DNA hybridisation (dDDH), Average Nucleotide Identity (ANI), and Average Amino Acid Identity (AAI) values of 22%, 80%, and 63%, respectively, with nearest phylogenetic affiliates. Based on the detailed analyses, we propose a new genus and species, Sporanaerobium hydrogeniformans gen. nov., sp. nov., for strain XHS1971T (= MCC3498T = KCTC15729T = JCM32657T) within family Lachnospiraceae.

Pseudodesulfovibrio thermohalotolerans sp. nov., a novel obligately anaerobic, halotolerant, thermotolerant, and sulfate-reducing bacterium isolated from a western offshore hydrocarbon reservoir in India.

OBJECTIVE: Characterization and documentation of strain MCM B-1480T, a novel sulfate-reducing bacterium isolated from produced water of India's western offshore hydrocarbon reservoir. METHOD: Strain MCM B-1480T was unequivocally identified using a polyphasic approach routinely followed in bacterial systematics. The morphological and biochemical characterization of strain MCM B-1480T was carried out using standard microbiological techniques. RESULTS: MCM B-1480T was a Gram-stain-negative, motile, non-spore-forming, curved-rod-shaped bacterium. MCM B-1480T could grow at temperatures between 20 and 60 °C (optimum 37 °C), pH 6-8 (optimum 7), and required 1-6% NaCl (optimum 3%) for growth. Strain MCM B-1480T was reducing sulfate to produce hydrogen sulfide during growth. This strain used lactate and pyruvate as prominent electron donors, whereas sulfate, sulfite, thiosulfate, and nitrate served as electron acceptors. MCM B-1480T shared maximum 16S rRNA gene sequence homology of 98.65% with the members of the genus Pseudodesulfovibrio. The G + C content of the 3.87 Mb MCM B-1480T genome was 60.39%. Digital DDH (27.7%) and average nucleotide identity (ANI 84%) with the closest phylogenetic affiliate (less than 70% and 95%, respectively) reaffirmed its distinctiveness. The major cellular fatty acids components, namely iso-C15:0, anteiso-C15:0, C16:0, and anteiso-C17:0, differentiated strain MCM B-1480T from other species of Pseudodesulfovibrio. Genome annotation revealed the presence of genes encoding dissimilatory sulfate reduction and nitrate reduction in strain MCM B-1480T. CONCLUSION: The polyphasic studies, including SSU rRNA gene sequencing, average nucleotide identity, Digital DNA-DNA hybridization, cell wall fatty acids analysis, etc., identified strain MCM B-1480T as a novel taxon and Pseudodesulfovibrio thermohalotolerans sp. nov. was proposed (= JCM 39269T = MCC 4711T).

Actinomyces ruminis sp. nov., an obligately anaerobic bacterium isolated from the rumen of cattle.

An obligately anaerobic, rod-shaped, Gram-stain-positive, non-spore-forming, non-motile bacterial strain; designated as CtC72T was isolated from the rumen of cattle. The 16S rRNA gene sequence similarity of less than 98.65% revealed the strain as a member of the genus Actinomyces, nearest to but distinct from Actinomyces qiguomingii DSM 106201T, Actinomyces ruminicola DSM 27982T, Actinomyces procaprae JCM 33484T, Actinomyces succiniciruminis TISTR 2317, Actinomyces glycerinitolerans TISTR 2318. The low values of digital DNA-DNA hybridization (< 70%) and average nucleotide identity (< 95%) further highlighted the distinctive nature of strain CtC72T from its closest relatives. The strain CtC72T could grow at temperatures between 30 and 50 °C (optimum 40 °C), pH between 6.0 and 9.0 (optimum 7.5-8.0), and NaCl between 0 and 1.5% (optimum 0%). The strain hydrolysed cellulose and xylan and utilised a range of mono-, di-, and oligo-saccharides as a source of carbon and energy. Glucose fermentation resulted in acetic acid and formic acid as major metabolic products, while propionic acid, lactic acid, and ethanol as minor products along with CO2 production. The DNA G + C content of strain CtC72T was 68.40 (mol%, Tm) and 68.05 (%, digital). Major cellular fatty acids (> 10%) were C16:0, C18:1 ω9c, and C18:1 ω9c DMA. Based on these data, we propose that strain CtC72T be classified as a novel species, Actinomyces ruminis sp. nov., under the genus Actinomyces. The type strain is CtC72T (= KCTC 15726T = JCM 32641T = MCC 3500T).

Genomic architecture of three newly isolated unclassified Butyrivibrio species elucidate their potential role in the rumen ecosystem.

One cellulose-degrading strain CB08 and two xylan-degrading strains XB500-5 and X503 were isolated from buffalo rumen. All the strains were designated as putative novel species of Butyrivibrio based on phylogeny, phylogenomy, digital DNA-DNA hybridization, and average nucleotide identity with their closest type strains. The draft genome length of CB08 was ~3.54 Mb, while X503 and XB500-5 genome sizes were ~3.24 Mb and ~3.27 Mb, respectively. Only 68.28% of total orthologous clusters were shared among three genomes, and 40-44% of genes were identified as hypothetical proteins. The presence of genes encoding diverse carbohydrate-active enzymes (CAZymes) exhibited the lignocellulolytic potential of these strains. Further, the genome annotations revealed the metabolic pathways for monosaccharide fermentation to acetate, butyrate, lactate, ethanol, and hydrogen. The presence of genes for chemotaxis, antibiotic resistance, antimicrobial activity, synthesis of vitamins, and essential fatty acid suggested the versatile metabolic nature of these Butyrivibrio strains in the rumen environment.

Genomics and simulated laboratory studies reveal Thermococcus sp. 101C5 as a novel hyperthermophilic archaeon possessing a specialized metabolic arsenal for enhanced oil recovery.

Laboratory evaluation of hyperthermophiles with the potential for Enhanced Oil Recovery (EOR) is often hampered by the difficulties in replicating the in situ growth conditions in the laboratory. In the present investigation, genome analysis was used to gain insights into the metabolic potential of a hyperthermophile to mobilize the residual oil from depleting high-temperature oil reservoirs. Here, we report the 1.9 Mb draft genome sequence of a hyperthermophilic anaerobic archaeon, Thermococcus sp. 101C5, with a GC content of 44%, isolated from a high-temperature oil reservoir of Gujarat, India. 101C5 possessed the genetic arsenal required for adaptation to harsh oil reservoir conditions, such as various heat shock proteins for thermo-adaptation, Trk potassium uptake system proteins for osmo-adaptation, and superoxide reductases against oxidative stress. Microbial Enhanced Oil Recovery (MEOR) potential of the strain was established by ascertaining the presence of genes encoding enzymes involved in the production of the metabolites such as hydrogen, bio-emulsifier, acetate, exopolysaccharide, etc. Production of these metabolites which pressurize the reservoir, emulsify the crude oil, lower the viscosity and reduce the drag, thus facilitating mobilization of the residual oil was experimentally confirmed. Also, the presence of crude oil degradative genes highlighted the ability of the strain to mobilize heavy residual oil, which was confirmed under simulated conditions in sand-pack studies. The obtained results demonstrated additional oil recoveries of 42.1% and 56.5% at 96 °C and 101 °C, respectively, by the strain 101C5, illustrating its potential for application in high-temperature oil reservoirs. To our best knowledge, this is the first report of genome analysis of any microbe assessed for its suitability for MEOR from the high-temperature oil reservoir.

Genome sequencing and analysis of a psychrotrophic methanogen Methanosarcina sp. nov. MSH10X1 cultured from methane hydrate deposits of Krishna Godavari Basin of India.

Methanosarcina sp. strain MSH10X1, a psychrotrophic methanogen, was isolated from sub-seafloor methane hydrate deposits of Krishna Godavari Basin on India's east coast. The strain could grow from 5 to 40 °C following all three i.e. methylotrophic, acetoclastic, and hydrogenotrophic modes of methanogenesis utilizing different substrates like methanol, trimethylamine, H2/CO2 (80/20), acetate, valerate, isobutyrate, isopropanol, and isobutanol. The genome sequencing and analysis of this strain revealed a circular chromosome of 3,557,383 bp length having 42.47 mol% G + C content, which consisted of 3110 coding genes, 58 tRNA genes, and 3 rRNA operons. The KEGG analysis highlighted the presence of genes responsible for all three modes of methanogenesis. The presence of genes like mtaB, mtaC, and mttB in the genome provided evidence for possible adaptation of strain MSH10X1 in the deep sea's low-temperature conditions.

Xylanimonas oleitrophica sp. nov., a novel petroleum hydrocarbon degrading bacterium isolated from an Indian oil reservoir.

A novel Gram-stain-positive, aerobic, non-spore-forming, and rod-shaped bacterium designated as PW21was isolated from produced water sampled from an oil reservoir in Limbodara, Gujarat, India. Growth occurred at 30-45 °C (optimum 37 °C), at pH 6.5-9.5 (optimum pH 8.5) and in the presence of 0.5-5.0% (w/v) NaCl [optimum, 3% (w/v)]. Phylogenetic analysis based on the 16S rRNA gene sequences showed that PW21T belonged to the class Actinobacteria, order Actinomycetales, family Promicromonosporaceae, and genus Xylanimonas, with the highest sequence similarity to Isoptericola cucumis AP38 (97.95% with 98.8% completeness), followed by Xylanimonas pachnodae NBRC 107,786T (97.82%) and Xylanimonas allomyrinae 2JSPR-7T(97.75%) with 100% completeness for the mentioned strains of Xylanimonas. The genome size of PW21T was 3.4 Mbp with a G+C content of 73.0 mol% (draft genome sequence). The average nucleotide identity (ANI) value of the draft genomes between PW21T and related species were found in between 76.99 and 78.93%. Major cellular fatty acids (> 10% of total fatty acids) of PW21Twere iso-C15:0, anteiso-C15:0, and C16:0. Phylogenetic, physiological, and biochemical characterisation identified strain PW21T (= KCTC 49,338T = JCM 33,795T = MCC 3936T) as a novel species of the genus Xylanimonas, and hence, name Xylanimonas oleitrophica sp. nov. PW21T is proposed.

Xylanolytic and Ethanologenic Potential of Gut Associated Yeasts from Different Species of Termites from India.

Xylophagous termites are capable of degrading lignocellulose by symbiotic gut microorganisms along with the host's indigenous enzymes. Therefore, the termite gut might be a potential niche to obtain natural yeasts with celluloytic, xylanolytic and ethanologenic traits required for bioethanol production from lignocellulosic biomass. In this study, we cultured 79 yeasts from three different termites viz. Coptotermes heimi, Odontotermes javanicus and Odontotermes obesus. After suitable screening methods, we identified 53 yeasts, which belonged to 10 genera and 16 different species of both ascomycetous and basidiomycetous yeasts. Most yeasts in the present study represent their first-ever isolation from the termite gut. Representative strains of identified yeasts were evaluated for their cellulolytic, xylanolytic, and ethanologenic abilities. None of the isolates showed cellulase activity; 22 showed xylanolytic activity, while six produced substantial quantities of ethanol. Among xylanolytic cultures, Pseudozyma hubeiensis STAG 1.7 and Hannaella pagnoccae STAG 1.14 produced 1.31 and 1.17 IU of xylanase. Among ethanologenic yeasts, the strains belonging to genera Candida and Kodamaea produced high amount of ethanol. Overall, highest ethanol level of 4.42 g/L was produced by Candida tropicalis TS32 using 1% glucose, which increased up to 22.92 g/L at 35 °C, pH 4.5 with 5% glucose. Fermentation of rice straw hydrolysate gave 8.95 g/l of ethanol with a yield of 0.42 g/g using the strain TS32. Our study highlights the gut of wood-feeding termites as a potential source of diverse yeasts that would be useful in the production of xylanase and bioethanol.

A novel, enantioselective, thermostable recombinant hydantoinase to aid the synthesis of industrially valuable non-proteinogenic amino acids.

Overexpression of a novel hydantoinase (hyuH) from P. aeruginosa (MCM B-887) in E. coli yielded optically pure carbamoyl amino acids. The use of optically pure carbamoyl amino acids as substrates facilitates the synthesis of non-proteinogenic amino acids. The enzyme hyuH shared a maximum of 92 % homology with proven hydantoinase protein sequences from the GenBank database, highlighting its novelty. Expression of hydantoinase gene was improved by >150 % by overexpressing it as a fusion protein in specialized E. coli CODON + host cells, providing adequate machinery for effective translation of the GC-rich gene. The presence of distinct residues in the substrate binding and active site of MCM B-887 hydantoinase enzyme explained its unique and broad substrate profile desirable for industrial applications. The purified enzyme, with a specific activity of 53U/mg of protein, was optimally active at 42 °C and pH 9.0 with a requirement of 2 mM Mn2+ ions. Supplementation of 500 mM of Na-glutamate enhanced the thermostability of the enzyme by more than 200 %.

Seven new Neocallimastigomycota genera from wild, zoo-housed, and domesticated herbivores greatly expand the taxonomic diversity of the phylum.

We isolated and characterized 65 anaerobic gut fungal (AGF; Neocallimastigomycota) strains from fecal samples of five wild (W, axis deer, white-tailed deer, Boer goat, mouflon, and Nilgiri tahr), one zoo-housed (Z, zebra), and three domesticated (D,  horse, sheep, and goat) herbivores in the US states of Texas (TX) and Oklahoma (OK), Wales (WA), and the Indian states of Kerala (KE) and Haryana (HA). Phylogenetic assessment using the D1-D2 regions of the large subunit (28S) rDNA and internal transcribed spacer 1 (ITS1) identified seven monophyletic clades that are distinct from all currently recognized AGF genera. All strains displayed monocentric thalli and produced exclusively or predominantly monoflagellate zoospores, with the exception of axis deer strains, which produced polyflagellate zoospores. Analysis of amplicon-based AGF diversity surveys indicated that zebra and horse strains are representatives of uncultured AL1 group, whereas domesticated goat and sheep strains are representatives of uncultured AL5 group, previously encountered in fecal and rumen samples of multiple herbivores. The other five lineages, all of which were isolated from wild herbivores, have not been previously encountered in such surveys. Our results significantly expand the genus-level diversity within the Neocallimastigomycota and strongly suggest that wild herbivores represent a yet-untapped reservoir of AGF diversity. We propose seven novel genera and eight novel Neocallimastigomycota species to comprise these strains, for which we propose the names Agriosomyces longus (mouflon and wild Boer goat), Aklioshbomyces papillarum (white-tailed deer), Capellomyces foraminis (wild Boar goat), and C. elongatus (domesticated goat), Ghazallomyces constrictus (axis deer), Joblinomyces apicalis (domesticated goat and sheep), Khoyollomyces ramosus (zebra-horse), and Tahromyces munnarensis (Nilgiri tahr).

Illustration of the microbial community selected by optimized process and nutritional parameters resulting in enhanced biomethanation of rice straw without thermo-chemical pretreatment.

Effects of different process and nutritional parameters on microbial community structure and function were investigated to enhance the biomethanation of rice straw without any thermochemical pre-treatment. The study was performed in a mesophilic anaerobic digester with cattle dung slurry as inoculum. The highest methane yield of 274 ml g-1 volatile solids was obtained from particulate rice straw (1 mm size, 7.5% solids loading rate) at 37 °C, pH-7, when supplemented with urea (carbon: nitrogen ratio, 25:1) and zinc as trace element (100 µM) at 21 days hydraulic retention time. The optimization of conditions selected Clostridium, Bacteroides, and Ruminococcus as dominant hydrolytic bacteria and Methanosarcina as the methanogen. Analysis of metagenome and metatranscriptome revealed wide array of bacterial lignocellulolytic enzymes that efficiently hydrolyzed the rice straw. The methane yield was >80% of the theoretical yield, making this green process a sustainable choice for efficient extraction of energy from rice straw.

A putative novel Methylobacter member (KRF1) from the globally important Methylobacter clade 2: cultivation and salient draft genome features.

Methane oxidation by methanotrophs is a very important environmental process in the mitigation of methane. Methylobacter (Mtb.) clade 2 members have been reported as dominant methane oxidisers in soils and sediments worldwide. We enriched and purified a methanotroph from a tropical rice field soil sample from India. The highly enriched culture showed the presence of motile, long and thick rods (3-5 µm × 0.9-1.2 µm) and minor presence of short, thin rods. The culture was purified on agarose medium and formed yellow colonies which showed the presence of only thick and long rods, henceforth termed as strain KRF1. Based on 16S rRNA gene sequence analysis, strain KRF1 shows close phylogenetic affiliation to Methylobacter tundripaludum SV96T (98.6% similarity). Due to the taxonomic novelty, and being the first member of Mtb. related to Mtb. tundripaludum from the tropics, the draft genome was sequenced. From the blastx analysis of the contigs, it was clear that the culture still had contamination of another organism, a Methylophilus species. The data binned in two clear bins: Mtb. related contigs and Methylophilus-related contigs. The binned draft genome of KRF1 shows features including the typical pathways for methane metabolism, denitrification and the presence of molybdenum iron and vanadium-iron nitrogenase genes. KRF1 is phylogenetically distinct from the five strains of Mtb. tundripaludum including SV96T, Lake Washington strains and OWC strains, showing ~ 26% DDH and ~ 81% ANIb values and a unique position in a phylogenomic tree. Subsequently, KRF1 has been completely purified from its methylotrophic partner and a pure culture has been established and maintained in a WDCM approved culture collection, the MACS Collection of Microorganisms (as MCM 1471). KRF1 is thus the first cultured member of a putative novel species of Methylobacter clade 2 isolated from the tropics.

Meta-omics based analyses of microbiome involved in biomethanation of rice straw in a thermophilic anaerobic bioreactor under optimized conditions.

Biomethanation of rice straw was performed at 55 °C without thermochemical pretreatment using cattle dung supplemented with Methanothermobacter thermautotrophicus strains. Methane yield of 323 ml g-1 VS obtained under optimized conditions such as particle size (1 mm), carbon to nitrogen ratio (15:1), substrate to inoculum ratio (1:1), organic loading rate (7.5% w/v) and hydraulic retention time (20 days), was one of the highest ever reported from rice straw. Metagenome analysis revealed several putative novel taxa among resident microbes. The genomes of Clostridium, Hungateiclostridium, Alkaliphilus, Anaerocolumna, Olsenella, Paenibacillus, Pseudoclostridium, Tepidanaerobacter and Turicibacter were recovered as metagenome assisted genomes. Clostridium spp. and M. thermautotrophicus were the dominant hydrolytic and methanogenic microbes, respectively. Syntrophic acetate oxidation coupled to hydrogenotrophic methanogenesis was found to be the major pathway for methane production. Efficient thermophilic biomethanation of rice straw without thermochemical pretreatment using cattle dung supplemented with M. thermautotrophicus is reported for the first time.

Purification and Characterization of an Active Principle, Lawsone, Responsible for the Plasmid Curing Activity of Plumbago zeylanica Root Extracts.

Plasmid curing is the process of obviating the plasmid encoded functions such as antibiotic resistance, virulence, degradation of aromatic compounds, etc. in bacteria. Several plasmid curing agents have been reported in literature, however, no plasmid curing agent can eliminate all plasmids from different hosts. Hence, there is always a need for novel plasmid curing agents that can be effectively used for reversal of plasmid encoded functions such as virulence, antibiotic resistance, etc. In the present study, an active principle responsible for the plasmid curing activity was purified from roots of Plumbago zeylanica by bioassay guided fractionation and identified as 2-hydroxy-1,4-naphthoquinone (lawsone), on the basis of spectral and analytical data such as NMR, GCMS, FTIR. Plasmid curing activity of lawsone was observed against reference as well as wild plasmids (pBR322, pRK2013, R136, pUPI281, and pUPI282) residing in a range of hosts. Curing of plasmid was confirmed by agarose gel electrophoresis. MICs of antibiotics against A. baumannii A24 (pUPI281) and E. coli (pRK2013) decreased significantly in presence of lawsone suggesting synergy between lawsone and antibiotics. Lawsone also inhibited transfer of plasmid pRK2013 to E. coli either by transformation or conjugation. Viability assays (MTT) revealed that lawsone was not toxic to mammalian cells. Thus, the present investigation has revealed lawsone as an effective plasmid curing agent capable of suppressing development and spread of antibiotic resistance. Further, lawsone has important application in basic research to identify phenotypes encoded by the plasmids in plasmid curing experiments. To the best of our knowledge this is the first report of plasmid curing activity of lawsone isolated from roots of P. zeylanica.

Liebetanzomycespolymorphus gen. et sp. nov., a new anaerobic fungus (Neocallimastigomycota) isolated from the rumen of a goat.

An extended incubation strategy to culture slow growing members of anaerobic fungi resulted in the isolation of a novel anaerobic fungus from the rumen of a goat after 15 days. The novel genus, represented by type strain G1SC, showed filamentous monocentric thallus development and produced uniflagellate zoospores, hence, showing morphological similarity to the genera Piromyces, Buwchfawromyces, Oontomyces and Pecoramyces. However, strain G1SC showed genetic similarity to the genus Anaeromyces, which, though produces uniflagellate zoospore, also exhibits polycentric thallus development. Moreover, unlike Anaeromyces, strain G1SC did not show hyphal constrictions, instead produced a branched, determinate and anucleate rhizoidal system. This fungus also displayed extensive sporangial variations, both exogenous and endogenous type of development, short and long sporangiophores and produced septate sporangia. G1SC utilised various complex and simple substrates, including rice straw and wheat straw and produced H2, CO2, formate, acetate, lactate, succinate and ethanol. Phylogenetic analysis, using internal transcribed spacer 1 (ITS1) and D1/D2 domain of large-subunit (LSU) rRNA locus, clearly showed a separate lineage for this strain, near Anaeromyces. The ITS1 based geographical distribution studies indicated detection of environmental sequences similar (93-96%) to this strain from cattle faeces. Based on morphological and molecular characterisation results of strain G1SC, we propose a novel anaerobic fungus Liebetanzomycespolymorphus gen. et sp. nov., in the phylum Neocallimastigomycota.

Cultivated methanotrophs associated with rhizospheres of traditional rice landraces from Western India belong to Methylocaldum and Methylocystis.

Aerobic methanotrophs associated with Indian rice plants have rarely been cultivated. In the present study, we cultured aerobic methanotrophic bacteria from the rhizosphere regions of rice plants. Rhizospheric soils from seven rice landraces traditionally grown and maintained by tribal people in Jawhar region belonging to part of the Western Ghats in India, were used. Seven methanotrophic cultures were isolated from the last positive dilution (10- 4). Methanotrophs were identified by analyzing the partial methane monooxygenase gene, pmoA gene and three of these belonged to the genus Methylocaldum (gammaproteobacterial, Type I methanotrophs) and four belonged to the genus Methylocystis (alphaproteobacterial, Type II methanotrophs). We present here the first report on the cultivation of methanotrophs from Indian traditional rice landraces originating from a biodiversity hotspot.

Cultivation of multiple genera of hydrogenotrophic methanogens from different environmental niches.

Six genera of hydrogenotrophic methanogens, namely Methanobrevibacter, Methanobacterium, Methanocorpusculum, Methanothermobacter, Methanoculleus, and Methanospirillum were cultivated from diverse environmental niches like rumen, feces, gut, and sediments using BY medium. We also report a putative novel genus and two novel species of methanogens isolated from termite, Indian star tortoise, and green iguana.