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1.  Exploration of Microbial Diversity and Community Structure of Lonar Lake: The Only Hypersaline Meteorite Crater Lake within Basalt Rock 
Lonar Lake is a hypersaline and hyperalkaline soda lake and the only meteorite impact crater in the world situated in basalt rocks. Although culture-dependent studies have been reported, a comprehensive understanding of microbial community composition and structure in Lonar Lake remains elusive. In the present study, microbial community structure associated with Lonar Lake sediment and water samples was investigated using high-throughput sequencing. Microbial diversity analysis revealed the existence of diverse, yet largely consistent communities. Proteobacteria (30%), Actinobacteria (24%), Firmicutes (11%), and Cyanobacteria (5%) predominated in the sequencing survey, whereas Bacteroidetes (1.12%), BD1-5 (0.5%), Nitrospirae (0.41%), and Verrucomicrobia (0.28%) were detected in relatively minor abundances in the Lonar Lake ecosystem. Within the Proteobacteria phylum, the Gammaproteobacteria represented the most abundantly detected class (21–47%) within sediment samples, but only a minor population in the water samples. Proteobacteria and Firmicutes were found at significantly higher abundance (p ≥ 0.05) in sediment samples, whereas members of Actinobacteria, Candidate division TM7 and Cyanobacteria (p ≥ 0.05) were significantly abundant in water samples. Compared to the microbial communities of other hypersaline soda lakes, those of Lonar Lake formed a distinct cluster, suggesting a different microbial community composition and structure. Here we report for the first time, the difference in composition of indigenous microbial communities between the sediment and water samples of Lonar Lake. An improved census of microbial community structure in this Lake ecosystem provides a foundation for exploring microbial biogeochemical cycling and microbial function in hypersaline lake environments.
doi:10.3389/fmicb.2015.01553
PMCID: PMC4722114  PMID: 26834712
16S rRNA gene; microbial diversity; soda lake; basalt rock; Lonar Lake; India
2.  Frankixalus, a New Rhacophorid Genus of Tree Hole Breeding Frogs with Oophagous Tadpoles 
PLoS ONE  2016;11(1):e0145727.
Despite renewed interest in the biogeography and evolutionary history of Old World tree frogs (Rhacophoridae), this family still includes enigmatic frogs with ambiguous phylogenetic placement. During fieldwork in four northeastern states of India, we discovered several populations of tree hole breeding frogs with oophagous tadpoles. We used molecular data, consisting of two nuclear and three mitochondrial gene fragments for all known rhacophorid genera, to investigate the phylogenetic position of these new frogs. Our analyses identify a previously overlooked, yet distinct evolutionary lineage of frogs that warrants recognition as a new genus and is here described as Frankixalus gen. nov. This genus, which contains the enigmatic ‘Polypedates’ jerdonii described by Günther in 1876, forms the sister group of a clade containing Kurixalus, Pseudophilautus, Raorchestes, Mercurana and Beddomixalus. The distinctiveness of this evolutionary lineage is also corroborated by the external morphology of adults and tadpoles, adult osteology, breeding ecology, and life history features.
doi:10.1371/journal.pone.0145727
PMCID: PMC4720377  PMID: 26790105
3.  Draft Genome Sequences of Yersinia pestis Strains from the 1994 Plague Epidemic of Surat and 2002 Shimla Outbreak in India 
Indian Journal of Microbiology  2014;54(4):480-482.
We report the first draft genome sequences of the strains of plague-causing bacteria, Yersinia pestis, from India. These include two strains from the Surat epidemic (1994), one strain from the Shimla outbreak (2002) and one strain from the plague surveillance activity in the Deccan plateau region (1998). Genome size for all four strains is ~4.49 million bp with 139–147 contigs. Average sequencing depth for all four genomes was 21x.
doi:10.1007/s12088-014-0475-7
PMCID: PMC4186941  PMID: 25320451
IonTorrent; NGS, next generation sequencing; Plague; Shimla; Surat
4.  Dynamics of Midgut Microflora and Dengue Virus Impact on Life History Traits in Aedes aegypti 
Acta tropica  2014;140:151-157.
Significant morbidity and potential mortality following dengue virus infection is a re-emerging global health problem. Due to the limited effectiveness of current disease control methods, mosquito biologists have been searching for new methods of controlling dengue transmission. While much effort has concentrated on determining genetic aspects to vector competence, paratransgenetic approaches could also uncover novel vector control strategies. The interactions of mosquito midgut microflora and pathogens may play significant roles in vector biology. However, little work has been done to see how the microbiome influences the host's fitness and ultimately vector competence. Here we investigated the effects of the midgut microbial environment and dengue infection on several fitness characteristics among three strains of the primary dengue virus vector mosquito Aedes aegypti. This included comparisons of dengue infection rates of females with and without their normal midgut flora. According to our findings, few effects on fitness characteristics were evident following microbial clearance or with dengue virus infection. Adult survivorship significantly varied due to strain and in one strain varied due to antibiotic treatment. Fecundity varied in one strain due to microbial clearance by antibiotics but no variation was observed in fertility due to either treatment. We show here that fitness characteristics of Ae. aegypti vary largely between strains, including varying response to microflora presence or absence, but did not vary in response to dengue virus infection.
doi:10.1016/j.actatropica.2014.07.015
PMCID: PMC4253069  PMID: 25193134
Aedes aegypti; midgut microflora; fitness; dengue; life history trait; vector competence
5.  Quinolone resistance mutations in the faecal microbiota of Swedish travellers to India 
BMC Microbiology  2015;15:235.
Background
International travel contributes to the spread of antibiotic resistant bacteria over the world. Most studies addressing travel-related changes in the faecal flora have focused on specific mobile resistance genes, or depended on culturing of individual bacterial isolates. Antibiotic resistance can, however, also spread via travellers colonized by bacteria carrying chromosomal antibiotic resistance mutations, but this has received little attention so far. Here we aimed at exploring the abundance of chromosomal quinolone resistance mutations in Escherichia communities residing in the gut of Swedish travellers, and to determine potential changes after visiting India. Sweden is a country with a comparably low degree of quinolone use and quinolone resistance, whereas the opposite is true for India.
Methods
Massively parallel amplicon sequencing targeting the quinolone-resistance determining region of gyrA and parC was applied to total DNA extracted from faecal samples. Paired samples were collected from 12 Swedish medical students before and after a 4–15 week visit to India. Twelve Indian residents were included for additional comparisons. Methods known resistance mutations were common in Swedes before travel as well as in Indians, with a trend for all mutations to be more common in the Indian sub group. There was a significant increase in the abundance of the most common amino acid substitution in GyrA (S83L, from 44 to 72 %, p = 0.036) in the samples collected after return to Sweden. No other substitution, including others commonly associated with quinolone resistance (D87N in GyrA, S80I in ParC) changed significantly. The number of distinct genotypes encoded in each traveller was significantly reduced after their visit to India for both GyrA (p = 0.0020) and ParC (p = 0.0051), indicating a reduced genetic diversity, similar to that found in the Indians.
Conclusions
International travel can alter the composition of the Escherichia communities in the faecal flora, favouring bacteria carrying certain resistance mutations, and, thereby, contributes to the global spread of antibiotic resistance. A high abundance of specific mutations in Swedish travellers before visiting India is consistent with the hypothesis that these mutation have no fitness cost even in the absence of an antibiotic selection pressure.
Electronic supplementary material
The online version of this article (doi:10.1186/s12866-015-0574-6) contains supplementary material, which is available to authorized users.
doi:10.1186/s12866-015-0574-6
PMCID: PMC4619388  PMID: 26498929
Antimicrobial resistance; Gut microbiota; 454 sequencing; Fluoroquinolones
6.  Technicalities and Glitches of Terminal Restriction Fragment Length Polymorphism (T-RFLP) 
Indian Journal of Microbiology  2014;54(3):255-261.
Terminal restriction fragment length polymorphism (T-RFLP) is a rapid, robust, inexpensive and simple tool for microbial community profiling. Methods used for DNA extraction, PCR amplification and digestion of amplified products have a considerable impact on the results of T-RFLP. Pitfalls of the method skew the similarity analysis and compromise its high throughput ability. Despite a high throughput method of data generation, data analysis is still in its infancy and needs more attention. Current article highlights the limitations of the methods used for data generation and analysis. It also provides an overview of the recent methodological developments in T-RFLP which will assist the readers in obtaining real and authentic profiles of the microbial communities under consideration while eluding the inherent biases and technical difficulties.
doi:10.1007/s12088-014-0461-0
PMCID: PMC4039717  PMID: 24891731
Impreciseness; Microbial ecology; NGS, next generation sequencing; Reproducibility; Resolution
7.  Distribution and Evolutionary Impact of Wolbachia on Butterfly Hosts 
Indian Journal of Microbiology  2014;54(3):249-254.
Wolbachia are maternally inherited endosymbiotic alpha-proteobacteria found in terrestrial arthropods and filarial nematodes. They are transmitted vertically through host cytoplasm and alter host biology by inducing various reproductive alterations, like feminization, parthenogenesis, male killing (MK) and cytoplasmic incompatibility. In butterflies, some effects especially MK and sperm-egg incompatibility are well established. All these effects skew the sex ratio towards female and subsequently favor the vertical transmission of Wolbachia. Some of the insects are also infected with multiple Wolbachia strains which may results in some complex phenomenon. In the present review the potential of Wolbachia for promoting evolutionary changes in its hosts with emphasis on recent advances in interactions of butterfly–Wolbachia is discussed. In addition to this, strain diversity of Wolbachia and its effects on various butterfly hosts are also highlighted.
doi:10.1007/s12088-014-0448-x
PMCID: PMC4039723  PMID: 24891730
Symbiosis; Wolbachia; Phenotypic effects; Butterfly; Evolutionary impact
8.  Secretor Status Is Strongly Associated with Microbial Alterations Observed during Pregnancy 
PLoS ONE  2015;10(7):e0134623.
During pregnancy there are significant changes in gut microbiota composition and activity. The impact of secretor status as determined by genotyping FUT2 (fucosyltransferase 2) gene was taken as one of the confounding factors associated with faecal microbiota changes during pregnancy. In this prospective study, we followed women during pregnancy (total = 123 of which secretors = 108, non-secretors = 15) and characterised their gut microbiota by quantitative polymerase chain reaction (qPCR), Fluorescence In situ Hybridisation (FISH), Denaturing Gradient Gel Electrophoresis (DGGE) and pyrosequencing. qPCR revealed that C. coccoides group counts decreased significantly in non-secretors in comparison to secretors (p = 0.02). Similar tendency was found by FISH analysis in Clostridium histolyticum and Lactobacillus-Enterococcus groups between the secretor and the non-secretor pregnant women. DGGE analysis showed significant decrease in richness of Clostridium sp. between secretor and non-secretor mothers during pregnancy. Pyrosequencing based analysis at phyla level showed that there is greater increase in Actinobacteria in secretors in comparison to non-secretors, whereas Proteobacteria showed more increase in non-secretors. Change in relative abundance of Clostridiaceae family from first to third trimester were significantly associated with secretor status of pregnant women (p = 0.05). Polyphasic approach for microbiota analysis points out that the host secretor status (FUT2 genotype) affects the gut microbiota during pregnancy. This may lead to altered infant gut microbiota colonization.
doi:10.1371/journal.pone.0134623
PMCID: PMC4521695  PMID: 26231005
9.  Insights into Diversity and Imputed Metabolic Potential of Bacterial Communities in the Continental Shelf of Agatti Island 
PLoS ONE  2015;10(6):e0129864.
Marine microbes play a key role and contribute largely to the global biogeochemical cycles. This study aims to explore microbial diversity from one such ecological hotspot, the continental shelf of Agatti Island. Sediment samples from various depths of the continental shelf were analyzed for bacterial diversity using deep sequencing technology along with the culturable approach. Additionally, imputed metagenomic approach was carried out to understand the functional aspects of microbial community especially for microbial genes important in nutrient uptake, survival and biogeochemical cycling in the marine environment. Using culturable approach, 28 bacterial strains representing 9 genera were isolated from various depths of continental shelf. The microbial community structure throughout the samples was dominated by phylum Proteobacteria and harbored various bacterioplanktons as well. Significant differences were observed in bacterial diversity within a short region of the continental shelf (1–40 meters) i.e. between upper continental shelf samples (UCS) with lesser depths (i.e. 1–20 meters) and lower continental shelf samples (LCS) with greater depths (i.e. 25–40 meters). By using imputed metagenomic approach, this study also discusses several adaptive mechanisms which enable microbes to survive in nutritionally deprived conditions, and also help to understand the influence of nutrition availability on bacterial diversity.
doi:10.1371/journal.pone.0129864
PMCID: PMC4465901  PMID: 26066038
10.  Microbial Culture Collection (MCC) and International Depositary Authority (IDA) at National Centre for Cell Science, Pune 
Indian Journal of Microbiology  2014;54(2):129-133.
Culture collections are valuable resources for the sustainable use of microbial diversity and its conservation. Advances in biotechnology have further increased their importance and some of these have been recognized as International Depositary Authority (IDA) for the deposition of patent cultures. Microbial Culture Collection at National Centre for Cell Science was established by the Department of Biotechnology, Government of India is country’s newest culture collection with largest holdings. It is recognized as an IDA under the Budapest Treaty and Designated National Repository under the Biodiversity Act 2002. This article describes its various service related activities.
doi:10.1007/s12088-014-0447-y
PMCID: PMC4188496  PMID: 25320411
Culture collection; International Depositary Authority; Biodiversity conservation
11.  DNA barcoding of Pentatomomorpha bugs (Hemiptera: Heteroptera) from Western Ghats of India 
Meta Gene  2014;2:737-745.
Recent studies from East Asia and Canadian National Collection of Insects have established the utility of DNA barcoding technique in identification of true bugs. The present study is an expansion of the database by adding mitochondrial cytochrome c oxidase I (mtCOI) sequences from forty three species of indigenous true bugs of India. mtCOI gene analysis of infraorder Pentatomomorpha covering a total of seventy three species that belong to five superfamilies; Pentatomoidea, Coreoidea, Pyrrhocoroidea, Lygaeoidea and Aradoidea revealed more than 3% interspecific distances in all the taxa studied except for two cases which showed barcode sharing. Less than 2% intra-specific divergence was observed in 97% of the taxa analysed and the average interspecies genetic distance was about 29 times higher than the average intraspecies genetic divergence. Distinct sequence divergence pattern at generic level and NJ clustering analysis suggests that COI barcode is an excellent molecular marker for species level identification of unknown taxa; however it may not be useful for resolving deep levels of divergence. Species identification even at nymphal stage could be achieved confirming the efficacy of this technique.
Graphical abstract
Fig. 1. NJ tree based on mtCOI from Cyclopelta siccifolia in three different life stages; egg, nymph and adult. Two Cimicomorpha taxa Rhynocoris marginatus and Rhynocoris ornatus are used as outgroups.
Highlights
•mtCOI barcoding of true bugs from Western Ghats of India is reported.•Study includes 9 families and 73 species; 43 from present work and 30 from GenBank.•mtCOI barcodes can be used for species level identification of unknown bug taxa.•Species identification even at nymphal stage could be achieved with this technique.•Present study adds 41 new mtCOI sequences to the existing global database.
doi:10.1016/j.mgene.2014.09.006
PMCID: PMC4287870  PMID: 25606457
COI, Cytochrome c oxidase I; CBOL, Consortium for the Barcode of Life; iBOL, International Barcode of Life, dNTPs, deoxynucleotide triphosphates; EtBr, Ethidium bromide; PEG, Polyethylene glycol; K2P, Kimura-2-parameter; NCBI, National Centre for Biological Information; BLAST, Basic local alignment search tool, MEGA, Molecular evolutionary genetics analysis; NJ, Neighbour joining; MP, Maximum parsimony; DNA barcoding; mtCOI; Heteroptera; Pentatomomorpha; Western Ghats; India
12.  Genome analysis of a major urban malaria vector mosquito, Anopheles stephensi 
Genome Biology  2014;15(9):459.
Background
Anopheles stephensi is the key vector of malaria throughout the Indian subcontinent and Middle East and an emerging model for molecular and genetic studies of mosquito-parasite interactions. The type form of the species is responsible for the majority of urban malaria transmission across its range.
Results
Here, we report the genome sequence and annotation of the Indian strain of the type form of An. stephensi. The 221 Mb genome assembly represents more than 92% of the entire genome and was produced using a combination of 454, Illumina, and PacBio sequencing. Physical mapping assigned 62% of the genome onto chromosomes, enabling chromosome-based analysis. Comparisons between An. stephensi and An. gambiae reveal that the rate of gene order reshuffling on the X chromosome was three times higher than that on the autosomes. An. stephensi has more heterochromatin in pericentric regions but less repetitive DNA in chromosome arms than An. gambiae. We also identify a number of Y-chromosome contigs and BACs. Interspersed repeats constitute 7.1% of the assembled genome while LTR retrotransposons alone comprise more than 49% of the Y contigs. RNA-seq analyses provide new insights into mosquito innate immunity, development, and sexual dimorphism.
Conclusions
The genome analysis described in this manuscript provides a resource and platform for fundamental and translational research into a major urban malaria vector. Chromosome-based investigations provide unique perspectives on Anopheles chromosome evolution. RNA-seq analysis and studies of immunity genes offer new insights into mosquito biology and mosquito-parasite interactions.
Electronic supplementary material
The online version of this article (doi:10.1186/s13059-014-0459-2) contains supplementary material, which is available to authorized users.
doi:10.1186/s13059-014-0459-2
PMCID: PMC4195908  PMID: 25244985
13.  Carl Woese: from Biophysics to Evolutionary Microbiology 
Indian Journal of Microbiology  2013;53(3):247-252.
This article is a tribute to Carl R. Woese, a biophysicist turned evolutionary microbiologist who passed away on December 30, 2012. We focus on his life, achievements, the discovery of Archaea and contributions to the development of molecular phylogeny. Further, the authors share their views and the lessons learnt from Woese’s life with the microbiologists in India. We also emphasize the need for interdisciplinary collaboration and interaction for the progress and betterment of science.
doi:10.1007/s12088-013-0401-4
PMCID: PMC3689406  PMID: 24426118
Woese; Molecular phylogeny; Archaea; rRNA
14.  Diversified diazotrophs associated with the rhizosphere of Western Indian Himalayan native red kidney beans (Phaseolus vulgaris L.) 
3 Biotech  2014;5(4):433-441.
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.
Electronic supplementary material
The online version of this article (doi:10.1007/s13205-014-0238-5) contains supplementary material, which is available to authorized users.
doi:10.1007/s13205-014-0238-5
PMCID: PMC4522724
Red kidney beans; Himalayan agro-ecosystems; Rhizosphere; nifH; Bacterial diazotrophic ecology
15.  Genome sequencing of multidrug resistant novel Clostridium sp. BL8 reveals its potential for pathogenicity 
Gut Pathogens  2014;6:30.
Background
The human gut microbiome is important for maintaining the health status of the host. Clostridia are key members of the human gut microbiome, carrying out several important functions in the gut environment. Hence understanding the role of different Clostridium species isolated from human gut is essential. The present study was aimed at investigating the role of novel Clostridium sp. isolate BL8 in human gut using genome sequencing as a tool.
Findings
The genome analysis of Clostridium sp. BL8 showed the presence of several adaptive features like bile resistance, presence of sensory and regulatory systems, presence of oxidative stress managing systems and presence of membrane transport systems. The genome of Clostridium sp. BL8 consists of a wide variety of virulence factors like phospholipase C (alpha toxin), hemolysin, aureolysin and exfoliative toxin A, as well as adhesion factors, proteases, Type IV secretion system and antibiotic resistance genes. In vitro antibiotic sensitivity testing showed that Clostridium sp. BL8 was resistant to 11 different tested antibiotics belonging to 6 different classes. The cell cytotoxicity assay confirmed the cytotoxic effect of Clostridium sp. BL8 cells, which killed 40% of the Vero cells after 4 hrs of incubation.
Conclusions
Clostridium sp. BL8 has adapted for survival in human gut environment, with presence of different adaptive features. The presence of several virulence factors and cell cytotoxic activity indicate that Clostridium sp. BL8 has a potential to cause infections in humans, however further in vivo studies are necessary to ascertain this fact.
doi:10.1186/1757-4749-6-30
PMCID: PMC4115488  PMID: 25076986
Clostridium; Virulence; Antibiotic resistance; Ion Torrent PGM™
16.  Characterization of bacterial community shift in human Ulcerative Colitis patients revealed by Illumina based 16S rRNA gene amplicon sequencing 
Gut Pathogens  2014;6:22.
Background
The healthy human intestine is represented by the presence of bacterial communities predominantly belonging to obligate anaerobes; however disparity and dysanaerobiosis in intestinal microflora may lead to the progression of ulcerative colitis (UC). The foremost aim of this study is to consider and compare the gut microbiota composition in patients suffering from different stages of UC.
Methods
This study represents data from the biopsy samples of six individuals suffering from UC. The samples were collected by colonoscopy and were processed immediately for isolation of DNA. Mucosal microbiota was analyzed by means of 16S rRNA gene-based Illumina high throughput sequencing. Quantitative real-time PCR (qPCR) was performed to determine total bacterial abundances.
Results
Analysis of 23,927 OTUs demonstrated a significant reduction of bacterial diversity consistently from phylum to species level (p < 0.05) for individuals suffering from severe stage of UC. Significant increase in abundance of unusual aerobes and facultative anaerobes, including members from the phylum Proteobacteria (p- = 0.031) was also observed. A 10 fold increase in the total bacterial count was detected in patients suffering from severe inflammatory stage (2.98 +/-0.49 E + 09/ml) when compared with patients with moderate (1.03+/-0.29 E + 08/ml) and mild (1.76 +/-0.34 E + 08/ml) stages of inflammation.
Conclusion
The reduction of bacterial diversity with an increase in the total bacterial count indicates a shift of bacterial communities which signifies dysbiosis and dysanaerobiosis at the mucosal level for patients suffering from UC.
doi:10.1186/1757-4749-6-22
PMCID: PMC4094692  PMID: 25018784
Inflammatory disease; Bacterial community shift; 16S rRNA gene; High through-put sequencing; QIIME analysis
18.  ε-Caprolactam Utilization by Proteus sp. and Bordetella sp. Isolated From Solid Waste Dumpsites in Lagos State, Nigeria, First Report 
Indian Journal of Microbiology  2013;53(2):221-226.
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 of ≤20 g l−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 35 °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 31.6–95.7 % 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.
doi:10.1007/s12088-013-0356-5
PMCID: PMC3626951  PMID: 24426112
ε-Caprolactam; Biodegradation; Bacteria; 16S rRNA; HPLC
19.  Acquired Genetic Mechanisms of a Multiresistant Bacterium Isolated from a Treatment Plant Receiving Wastewater from Antibiotic Production 
Applied and Environmental Microbiology  2013;79(23):7256-7263.
The external environment, particularly wastewater treatment plants (WWTPs), where environmental bacteria meet human commensals and pathogens in large numbers, has been highlighted as a potential breeding ground for antibiotic resistance. We have isolated the extensively drug-resistant Ochrobactrum intermedium CCUG 57381 from an Indian WWTP receiving industrial wastewater from pharmaceutical production contaminated with high levels of quinolones. Antibiotic susceptibility testing against 47 antibiotics showed that the strain was 4 to >500 times more resistant to sulfonamides, quinolones, tetracyclines, macrolides, and the aminoglycoside streptomycin than the type strain O. intermedium LMG 3301T. Whole-genome sequencing identified mutations in the Indian strain causing amino acid substitutions in the target enzymes of quinolones. We also characterized three acquired regions containing resistance genes to sulfonamides (sul1), tetracyclines [tet(G) and tetR], and chloramphenicol/florfenicol (floR). Furthermore, the Indian strain harbored acquired mechanisms for horizontal gene transfer, including a type I mating pair-forming system (MPFI), a MOBP relaxase, and insertion sequence transposons. Our results highlight that WWTPs serving antibiotic manufacturing may provide nearly ideal conditions for the recruitment of resistance genes into human commensal and pathogenic bacteria.
doi:10.1128/AEM.02141-13
PMCID: PMC3837729  PMID: 24038701
20.  Microbiology of Lonar Lake and other soda lakes 
The ISME Journal  2012;7(3):468-476.
Soda lakes are saline and alkaline ecosystems that are believed to have existed throughout the geological record of Earth. They are widely distributed across the globe, but are highly abundant in terrestrial biomes such as deserts and steppes and in geologically interesting regions such as the East African Rift valley. The unusual geochemistry of these lakes supports the growth of an impressive array of microorganisms that are of ecological and economic importance. Haloalkaliphilic Bacteria and Archaea belonging to all major trophic groups have been described from many soda lakes, including lakes with exceptionally high levels of heavy metals. Lonar Lake is a soda lake that is centered at an unusual meteorite impact structure in the Deccan basalts in India and its key physicochemical and microbiological characteristics are highlighted in this article. The occurrence of diverse functional groups of microbes, such as methanogens, methanotrophs, phototrophs, denitrifiers, sulfur oxidizers, sulfate reducers and syntrophs in soda lakes, suggests that these habitats harbor complex microbial food webs that (a) interconnect various biological cycles via redox coupling and (b) impact on the production and consumption of greenhouse gases. Soda lake microorganisms harbor several biotechnologically relevant enzymes and biomolecules (for example, cellulases, amylases, ectoine) and there is the need to augment bioprospecting efforts in soda lake environments with new integrated approaches. Importantly, some saline and alkaline lake ecosystems around the world need to be protected from anthropogenic pressures that threaten their long-term existence.
doi:10.1038/ismej.2012.137
PMCID: PMC3578565  PMID: 23178675
Lonar Lake; soda lakes; alkaliphiles; stable-isotope probing; microbial prospecting
21.  Functional sub-division of the Drosophila genome via chromatin looping 
Nucleus  2013;4(2):115-122.
Insulators help in organizing the eukaryotic genomes into physically and functionally autonomous regions through the formation of chromatin loops. Recent findings in Drosophila and vertebrates suggest that insulators anchor multiple loci through long-distance interactions which may be mechanistically linked to insulator function. Important to such processes in Drosophila is CP190, a common co-factor of insulator complexes. CP190 is also known to associate with the nuclear matrix, components of the RNAi machinery, active promoters and borders of the repressive chromatin domains. Although CP190 plays a pivotal role in insulator function in Drosophila, vertebrates lack a probable functional equivalent of CP190 and employ CTCF as the major factor to carry out insulator function/chromatin looping. In this review, we discuss the emerging role of CP190 in tethering genome, specifically in the perspective of insulator function in Drosophila. Future studies aiming genome-wide role of CP190 in chromatin looping is likely to give important insights into the mechanism of genome organization.
doi:10.4161/nucl.23389
PMCID: PMC3621743  PMID: 23333867
Insulators; CP190; long-range interactions; chromatin organization
22.  Ectopically tethered CP190 induces large-scale chromatin decondensation 
Scientific Reports  2014;4:3917.
Insulator mediated alteration in higher-order chromatin and/or nucleosome organization is an important aspect of epigenetic gene regulation. Recent studies have suggested a key role for CP190 in such processes. In this study, we analysed the effects of ectopically tethered insulator factors on chromatin structure and found that CP190 induces large-scale decondensation when targeted to a condensed lacO array in mammalian and Drosophila cells. In contrast, dCTCF alone, is unable to cause such a decondensation, however, when CP190 is present, dCTCF recruits it to the lacO array and mediates chromatin unfolding. The CP190 induced opening of chromatin may not be correlated with transcriptional activation, as binding of CP190 does not enhance luciferase activity in reporter assays. We propose that CP190 may mediate histone modification and chromatin remodelling activity to induce an open chromatin state by its direct recruitment or targeting by a DNA binding factor such as dCTCF.
doi:10.1038/srep03917
PMCID: PMC3905270  PMID: 24472778
23.  Midgut Microbial Community of Culex quinquefasciatus Mosquito Populations from India 
PLoS ONE  2013;8(11):e80453.
The mosquito Culex quinquefasciatus is a ubiquitous species that serves as a major vector for west nile virus and lymphatic filariasis. Ingestion of bloodmeal by females triggers a series of physiological processes in the midgut and also exposes them to infection by these pathogens. The bacteria normally harbored in the midgut are known to influence physiology and can also alter the response to various pathogens. The midgut bacteria in female Cx. quinquefasciatus mosquitoes collected over a large geographical area from India was studied. Examination of 16S ribosomal DNA amplicons from culturable microflora revealed the presence of 83 bacterial species belonging to 31 bacterial genera. All of these species belong to three phyla i.e. Proteobacteria, Firmicutes and Actinobacteria. Phylum Proteobacteria was the most dominant phylum (37 species), followed by Firmicutes (33 species) and Actinobacteria (13 species). Phylum Proteobacteria, was dominated by members of γ-proteobacteria class. The genus Staphylococcus was the largest genus represented by 11 species whereas Enterobacter was the most prevalent genus and recovered from all the field stations except Leh. Highest bacterial prevalence was observed from Bhuj (22 species) followed by Nagrota (18 species), Masimpur (18 species) and Hathigarh (16 species). Whereas, least species were observed from Leh (8 species). It has been observed that individual mosquito harbor extremely diverse gut bacteria and have very small overlap bacterial taxa in their gut. This variation in midgut microbiota may be one of the factors responsible for variation in disease transmission rates or vector competence within mosquito population. The present data strongly encourage further investigations to verify the potential role of the detected bacteria in mosquito for the transmission of lymphatic filariasis and west nile virus. To the best of our knowledge this is the first study on midgut microbiota of wild Cx. quinquefasciatus from over a large geographical area.
doi:10.1371/journal.pone.0080453
PMCID: PMC3843677  PMID: 24312223
24.  Comparative Genome Analysis of Megasphaera sp. Reveals Niche Specialization and Its Potential Role in the Human Gut 
PLoS ONE  2013;8(11):e79353.
With increasing number of novel bacteria being isolated from the human gut ecosystem, there is a greater need to study their role in the gut ecosystem and their effect on the host health. In the present study, we carried out in silico genome-wide analysis of two novel Megasphaera sp. isolates NM10 (DSM25563) and BL7 (DSM25562), isolated from feces of two healthy individuals and validated the key features by in vitro studies. The analysis revealed the general metabolic potential, adaptive features and the potential effects of these isolates on the host. The comparative genome analysis of the two human gut isolates NM10 and BL7 with ruminal isolate Megasphaera elsdenii (DSM20460) highlighted the differential adaptive features for their survival in human gut. The key findings include features like bile resistance, presence of various sensory and regulatory systems, stress response systems, membrane transporters and resistance to antibiotics. Comparison of the “glycobiome” based on the genomes of the ruminal isolate with the human gut isolates NM10 and BL revealed the presence of diverse and unique sets of Carbohydrate-Active enzymes (CAZymes) amongst these isolates, with a higher collection of CAZymes in the human gut isolates. This could be attributed to the difference in host diet and thereby the environment, consequently suggesting host specific adaptation in these isolates. In silico analysis of metabolic potential predicted the ability of these isolates to produce important metabolites like short chain fatty acids (butyrate, acetate, formate, and caproate), vitamins and essential amino acids, which was further validated by in vitro experiments. The ability of these isolates to produce important metabolites advocates for a potential healthy influence on the host. Further in vivo studies including transcriptomic and proteomic analysis will be required for better understanding the role and impact of these Megasphaera sp. isolates NM10 and BL7 on the human host.
doi:10.1371/journal.pone.0079353
PMCID: PMC3832451  PMID: 24260205
25.  A Treatment Plant Receiving Waste Water from Multiple Bulk Drug Manufacturers Is a Reservoir for Highly Multi-Drug Resistant Integron-Bearing Bacteria 
PLoS ONE  2013;8(10):e77310.
The arenas and detailed mechanisms for transfer of antibiotic resistance genes between environmental bacteria and pathogens are largely unclear. Selection pressures from antibiotics in situations where environmental bacteria and human pathogens meet are expected to increase the risks for such gene transfer events. We hypothesize that waste-water treatment plants (WWTPs) serving antibiotic manufacturing industries may provide such spawning grounds, given the high bacterial densities present there together with exceptionally strong and persistent selection pressures from the antibiotic-contaminated waste. Previous analyses of effluent from an Indian industrial WWTP that processes waste from bulk drug production revealed the presence of a range of drugs, including broad spectrum antibiotics at extremely high concentrations (mg/L range). In this study, we have characterized the antibiotic resistance profiles of 93 bacterial strains sampled at different stages of the treatment process from the WWTP against 39 antibiotics belonging to 12 different classes. A large majority (86%) of the strains were resistant to 20 or more antibiotics. Although there were no classically-recognized human pathogens among the 93 isolated strains, opportunistic pathogens such as Ochrobactrum intermedium, Providencia rettgeri, vancomycin resistant Enterococci (VRE), Aerococcus sp. and Citrobacter freundii were found to be highly resistant. One of the O. intermedium strains (ER1) was resistant to 36 antibiotics, while P. rettgeri (OSR3) was resistant to 35 antibiotics. Class 1 and 2 integrons were detected in 74/93 (80%) strains each, and 88/93 (95%) strains harbored at least one type of integron. The qPCR analysis of community DNA also showed an unprecedented high prevalence of integrons, suggesting that the bacteria living under such high selective pressure have an appreciable potential for genetic exchange of resistance genes via mobile gene cassettes. The present study provides insight into the mechanisms behind and the extent of multi-drug resistance among bacteria living under an extreme antibiotic selection pressure.
doi:10.1371/journal.pone.0077310
PMCID: PMC3812170  PMID: 24204801

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