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Chakraborty, R.

Publications and source records attributed to Chakraborty, R..

7 recordsLinked to original sources

Defining the toxicity limits on microbial range in a metal-contaminated aquifer

In extreme environments, toxic compounds restrict which microorganisms persist. However, in complex mixtures of inhibitory compounds, it is challenging to determine which specific compounds cause changes in abundance and prevent some microorganisms from growing. We focused on a contaminated aquifer in Oak Ridge, Tennessee, U.S.A. that has low pH and high concentrations of uranium, nitrate and many other inorganic ions. In the most contaminated wells, the microbial community is enriched in the Rhodanobacter genus. Rhodanobacter relative abundance is positively correlated with low pH and high concentrations of U, Mn, Al, Cd, Zn, Ni, Co, Ca, NO3-, Mg, Cl, SO42-, Sr, K and Ba and we sought to determine which of these correlated parameters are selective pressures that favor the growth of Rhodanobacter over other taxa. Using high-throughput cultivation, we determined that of the ions correlated high Rhodanobacter abundance, only low pH and high U, Mn, Al, Cd, Zn, Co and Ni (a) are selectively inhibitory of a sensitive Pseudomonas isolate from a background well versus a representative resistant Rhodanobacter isolate from a contaminated well, and (b) reach toxic concentrations in the most contaminated wells that can inhibit the sensitive Pseudomonas isolate. We prepared mixtures of inorganic ions representative of the most contaminated wells and verified that few other isolates aside from Rhodanobacter can tolerate these 8 parameters. These results clarify which toxic inorganic ions are causal factors that impact the microbial community at this field site and are not merely correlated with taxonomic shifts.

microbiology

Resource limitation modulates the fate of dissimilated nitrogen in a dual-pathway Actinobacterium

Respiratory ammonification and denitrification are two evolutionarily unrelated dissimilatory nitrogen (N) processes central to the global N cycle, the activity of which is thought to be controlled by carbon (C) to nitrate (NO3-) ratio. Here we find that Intrasporangium calvum C5, a novel menaquinone-based dual-pathway denitrifier/respiratory ammonifier, disproportionately utilizes ammonification rather than denitrification when grown under carbon or nitrate limitation, not C:NO3- ratio. Higher growth rates are promoted by ammonification and metabolite and transcriptional profiles during growth show that the bacterium produces its own formate from a fermentable carbon source (lactate) to further generate a proton motive force for the ammonification pathway. Transcript abundances encoding for nitrite reducing enzymes, NrfAH and NirK, also significantly increase in response to nitrite production. Mechanistically, our results suggest that pathway selection is driven by intracellular redox potential (redox poise), which may be lowered during resource limitation, thereby decreasing catalytic activity of upstream electron transport steps needed for denitrification enzymes. Our work advances our understanding of the biogeochemical flexibility of N-cycling organisms and pathway evolution.

microbiology

The first complete mitochondrial genome of marigold pest thrips, Neohydatothrips samayunkur (Sericothripinae) and comparative analysis

The complete mitogenomes in order Thysanoptera is limited to subfamily Thripinae heretofore. In the present study, we sequenced the first mitochondrial genome of Neohydatothrips samayunkur (15,295 bp), a member of subfamily Sericothripinae. The genome was characterized by 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNAs), two ribosomal RNA genes (rRNAs) and three control regions (CRs). This mitogenome had two overlapping regions of 4 bp and twenty four intergenic spacers accounting for 165 bp. All the tRNA had typical cloverleaf secondary structures, except for trnV and trnS which lacked DHU stem and loop. The mitogenomes of N. samayunkur was highly rearranged with many unique features as compared to other thrips mitogenomes, atp6 and nad1 were terminated with TAG and TGA stop codons respectively; location of trnL2, trnA, trnC, and trnV was rearranged; and the first control region (CR1) was upstream of nad6. The phylogenetic analysis of 13 PCGs implementing maximum likelihood and Bayesian inference showed the clustering of N. samayunkur with Scirtothrips dorsalis supporting the Scirtothrips genus-group and Sericothripinae morphology based relationships. Generation of more mitogenomes from different hierarchical level in the order Thysanoptera is required to understand the gene rearrangements, phylogeny and evolutionary relationships.

genomics

Sequencing and characterization of the complete mitochondrial genome of Critically Endangered Black Soft-shell Turtle (Nilssonia nigricans)

The complete mitochondrial genome (16796 bp) of an endangered freshwater turtle, Nilssonia nigricans was firstly sequenced and annotated. The mitogenome was encoded by 37 genes and a major non-coding Control Region (CR). The mitogenome was A+T biased (62.16%) and spread with six overlapping and 19 intergenic spacer regions. The initiation codons were exceptionally changed as ATA, ATT, and ATC in three Protein-coding genes (PCGs) and a single base (A, T, and C) incomplete termination codons in nine PCGs. The Relative synonymous codon usage (RSCU) value was consistent among all the studied species; exception with significant reduction of Serine (S) frequency in N. nigricans, N. formosa, and R. swinhoei. The secondary structure of N. nigricans showed a lack of conventional dihydrouridine (DHU) arm in trnS (GCT), as well as formed a small loop structure in the acceptor stem of both trnR (TCG) and trnH (GTG). The mitogenome of N. nigricans also revealed two unique tandem repeats (ATTAT)8, and (TATTA)20 in CR. Further, the average Ka/Ks values of 13 PCGs were indicating a strong natural selection in the studied Trionychidae species. The constructed Maximum Likelihood (ML) phylogeny by PCGs shows cohesive clustering of N. nigricans with N. formosa. The resulted phylogeny illustrated the similar topology by all studied species from different taxonomic ranks and supported the previous taxonomic classification. Moreover, further taxon sampling from different taxonomic hierarchy, and their mitogenomics study is vital to reconcile the Testudines phylogeny and assure their evolutionary relationship.

genomics

The complete mitochondrial genome of Melon thrips, Thrips palmi (Thripinae) and comparative analysis: A vector for Tospoviruses

The melon thrips, Thrips palmi is a serious pest and vector for plant viruses on a wide range of economically important crops. DNA barcoding evidenced the presence of cryptic diversity in T. palmi and that warrants exhaustive molecular data. Our present study is on decoding the first complete mitochondrial genome of T. palmi (15,333 bp) through NGS technology. The mitogenome contains 37 genes, including 13 PCGs, two rRNAs, 22 tRNAs, and two control regions. The comparative analyses were conducted for gene arrangements, nucleotide composition, codon usage and phylogenetic relationship with other thrips mitogenomes. The nucleotide composition was 78.29% AT, and 21.72% GC with positive AT skewness (0.09) and negative GC skewness (-0.06). The ATN initiation codons were observed in 12 PCGs except cox1 with unique start codon (TTG). The RSCU analysis revealed Phe, Leu, Ile, Tyr, Asn, Lys and Met were the most frequently used amino acids in all PCGs. The codon CGG (Arg) was absent in T. palmi as compared to other thrips mitogenomes. The Ka/Ks ratio ranges from 0.078 in cox1 to 0.913 in atp8. We observed the typical cloverleaf secondary structure in most of the tRNA genes with a few exceptions; absence of DHU stem and loop in trnV and trnS, absence of DHU loop in trnE, lack of T-arm and loop in trnN. The position of trnS1 (between cox3 and CR2) is unique in T. palmi among all the studied thrips mitogenomes. The mitogenome contained 24 intergenic spacer regions and 12 overlapping regions. The CR2 is 63.77% similar to CR1, indicating a possible duplication and translocation in control region. Both the ML and BI phylogenetic trees revealed the close relationships of Thrips with Scirtothrips as compared to Frankliniella. Thus, more mitogenomes on the diverse thrips species is required to understand the in-depth phylogenetic and evolutionary relationships.

genomics

Magic pools: parallel assessment of transposon delivery vectors in bacteria

Transposon mutagenesis coupled to next-generation sequencing (TnSeq) is a powerful approach for discovering the functions of bacterial genes. However, the development of a suitable TnSeq strategy for a given bacterium can be costly and time-consuming. To meet this challenge, we describe a parts-based strategy for constructing libraries of hundreds of transposon delivery vectors, which we term \"magic pools\". Within a magic pool, each transposon vector has a different combination of promoters and antibiotic resistance markers as well as a random DNA barcode sequence, which allows the tracking of each vector during mutagenesis experiments. To identify an efficient vector for a given bacterium, we mutagenize it with a magic pool and sequence the resulting insertions; we then use the best vector to generate a large mutant library. We used the magic pool strategy to construct transposon mutant libraries in five genera of bacteria, including three genera of the phylum Bacteroidetes.

genomics

Construction of soil defined media using quantitative exometabolomic analysis of soil metabolites

Exometabolomics enables analysis of metabolite utilization of low molecular weight organic substances by soil isolates. Environmentally-based defined media are needed to examine ecologically relevant patterns of substrate utilization. Here, we describe an approach for the construction of defined media using untargeted characterization of water soluble soil metabolites. To broadly characterize soil metabolites, both liquid chromatography mass spectrometry (LC/MS) and gas chromatography mass spectrometry (GC/MS) were used. With this approach, 96 metabolites were identified, including amino acids, amino acid derivatives, sugars, sugar alcohols, mono- and di-carboxylic acids, osmolytes, nucleobases, and nucleosides. From this pool of metabolites, 25 were quantified. Water soluble organic carbon was fractionated by molecular weight and measured to determine the fraction of carbon accounted for by the quantified metabolites. This revealed that, community structures, these soil metabolites have an uneven quantitative distribution, with a single metabolite, trehalose accounting for 9.9 percent of much like soil microbial the (< 1 kDa) water extractable organic carbon. This quantitative information was used to formulate two soil defined media (SDM), one containing 23 metabolites (SDM1) and one containing 46 (SDM2). To evaluate SDM for supporting the growth of bacteria found at this field site, we examined the growth of 30 phylogenetically diverse soil isolates obtained using standard R2A medium. The simpler SDM1 supported the growth of up to 13 isolates while the more complex SDM2 supported up to 25 isolates. One isolate, Pseudomonas corrugata strain FW300-N2E2 was selected for a time-series exometabolomics analysis to investigate SDM1 substrate preferences. Interestingly, it was found that this organism preferred lower-abundance substrates such as guanine, glycine, proline and arginine and glucose and did not utilize the more abundant substrates maltose, mannitol, trehalose and uridine. These results demonstrate the viability and utility of using exometabolomics to construct a tractable environmentally relevant media. We anticipate that this approach can be expanded to other environments to enhance isolation and characterization of diverse microbial communities.\n\nHighlights O_LILC/MS and GC/MS analyses of soil extracts revealed a diversity of 96 metabolites.\nC_LIO_LISoil defined media were constructed based on water extractable soil metabolomics data.\nC_LIO_LIThe defined media supported the growth of 25 out of 30 bacterial isolates.\nC_LIO_LIExometabolomics demonstrated preferential consumption of amino acids for one isolate.\nC_LIO_LIThese media can be used to understand environmentally relevant microbial substrate preferences.\nC_LI\n\nAbbreviationsDOM dissolved organic matter; SOM soil organic matter; WEOC water extractable organic carbon; LMWOS low molecular weight organic substances; SDM(1/2) soil defined media (1 and 2); R2A Reasoners 2A agar medium; ORFRC Oak Ridge Field Research Center; LC/MS liquid chromatography mass spectrometry; GC/MS gas chromatography/ mass spectrometry; TOC total organic carbon; HILIC hydrophilic interaction liquid chromatography

microbiology