Biology subjects

Das, L.

Publications and source records attributed to Das, L..

7 recordsLinked to original sources

bioRxiv2026

Hydration and H/D exchange-dependent infrared signatures of the GCN4 leucine zipper

Attenuated total reflectance Fourier-transform infrared (ATR-FTIR) spectroscopy of proteins in aqueous solution is often limited by water absorption and other optical artifacts. To overcome these limitations, we evaluated the structural features and hydrogen-deuterium exchange (HDX) kinetics of the -helical protein GCN4 in both hydrated (wet) and vacuum-dried (dry) states. While solvent heavily mask the second-derivative spectra of wet samples, vacuum drying yielded a thin, protein-rich film on the ATR crystal, significantly enhancing the signal-to-noise ratio and resolving the protein features without altering the native structure. Dry-state analysis clearly resolved the Amide I, Amide II, and deuterium-shifted Amide II (1450 cm-1) bands. Notably, second-derivative analysis of the dry spectra of the HDX samples revealed a bimodal Amide I distribution consisting of a stationary band at 1653 cm-1 from the solvent-inaccessible regions and an isotopically sensitive band shifting from 1648 cm-1 to 1644 cm-1 from solvent-accessible regions. These results demonstrate that vacuum-dried ATR-FTIR spectroscopy effectively eliminates solvent masking, providing the spectral clarity required to resolve discrete -helical sub-populations after deuteration.

biochemistry↗

bioRxiv2026

Postpartum Systemic Inflammation is Reflected in Early Distinct Fecal Microbiome Differences in Dairy Cows

A tightly regulated inflammatory response occurs during and following parturition; however, excessive or prolonged inflammation negatively affects herd health and productivity. The gut microbiota plays an important role in host immunity and metabolism and undergoes substantial changes during the transition period. However, the relationship between systemic inflammation, measured by serum acute-phase proteins, and gut microbial dynamics during early lactation remains poorly understood. We investigated fecal microbiota dynamics in relation to systemic inflammation in early postpartum dairy cows. Fecal and blood samples were collected from 71 Holstein cows on days 1 and 3 in milk (DIM). The V3-V4 region of the 16S rRNA gene was sequenced and microbial diversity, differential abundance, and microbial interaction networks were evaluated. Inflammatory status, defined by fibrinogen, haptoglobin, and their combined classification, was associated with significant alterations in fecal microbial composition during the immediate postpartum period, independent of body condition score, parity, DIM, and DNA extraction parameters. Differential abundance analyses revealed extensive taxonomic restructuring, while network analyses identified increased modularity, altered keystone taxa distribution, and greater compartmentalization of microbial interactions in animals with elevated inflammatory markers. Several taxa were consistently associated with inflammatory status across analytical approaches. Notably, Ruminococcaceae UCG-002 and Dielma were associated with inflammatory states, whereas Xylanibacter, Marvinbryantia, Akkermansia, and Oscillibacter were associated with non-inflammatory status. This study identifies an association between systemic inflammation and fecal microbiota composition in early transition dairy cows, providing a foundation for future microbiome-based biomarkers of inflammatory status. IMPORTANCESubclinical systemic inflammation during early postpartum can negatively affect dairy cow health and productivity, yet current monitoring relies on repeated blood sampling and transient inflammatory biomarkers. For the first time association of systemic inflammation, assessed using fibrinogen, haptoglobin, and their combined classification, with alterations in fecal microbial composition, microbial interaction networks, and keystone taxa distribution during the early postpartum period was established. Several bacterial taxa were consistently associated with either elevated or normal inflammatory states across differential abundance, network, and odds ratio analyses. Many of these taxa remain poorly characterized in dairy cattle, highlighting the need for future mechanistic studies. This study demonstrates that systemic inflammation during early postpartum is associated with measurable alterations in the fecal microbiota. This work provides a foundation for developing microbiome-based biomarkers for detecting and monitoring subclinical systemic inflammation in dairy cattle.

microbiology↗

bioRxiv2025

The unique role of nucS-mediated non-canonical mismatch repair in Mycobacterium tuberculosis resistance evolution

DNA surveillance mechanisms play a vital role in maintaining genome stability and minimizing mutation rates. One such mechanism, post-replicative mismatch repair (MMR), corrects replication errors that escape DNA polymerase proofreading activity. In most bacteria and eukaryotes, MMR is orchestrated by MutS and MutL proteins. However, certain archaeal and actinobacterial species, including the major human pathogen Mycobacterium tuberculosis, lack these components. Instead, they rely on the nuclease EndoMS/NucS, a structurally distinct enzyme that governs a non-canonical MMR pathway. Given that M. tuberculosis acquires drug resistance exclusively through chromosomal mutations, understanding mutation rate regulation in this pathogen is critical. Nevertheless, despite its anticipated significance, the role of NucS in drug resistance evolution remains largely unexplored in this organism. This study investigates NucS function in M. tuberculosis and uncovers a unique resistance dynamic distinct from other Actinobacteria. While nucS deletion alters the mutational spectrum, it minimally affects the emergence of rifampicin-, isoniazid-, and ethambutol-resistant mutations, in stark contrast to its role in other Actinobacteria. We demonstrated that this atypical behaviour is not attributable to the presence of a single NucS polymorphism, R144S, in the NucS sequence of the M. tuberculosis reference strain H37Rv, which differs from the NucS consensus sequence. Constructing and analysing an H37Rv variant possessing the NucS consensus sequence revealed a subtly altered mutational spectrum but unchanged mutation rates. Notably, database analysis of the R144S polymorphism in clinical isolates revealed its prevalence and significant association with ethambutol resistance. These findings challenge the established view that nucS serves as a genome stability guardian that minimizes mutation rates in M. tuberculosis, suggesting additional mismatch repair mechanisms beyond NucS or a highly efficient replication system in this pathogen.

microbiology↗

bioRxiv2025

Loss of the ESX-5 secretion locus in Mycobacterium tuberculosis reshapes the mycomembrane and enhances ESX-1 substrate secretion

The ESX-5 secretion system, uniquely found in slow-growing mycobacteria, is predicted to secrete over 150 proteins across the inner membrane of Mycobacterium tuberculosis (M.tb). Although many of these substrates are believed to promote M.tb virulence, most remain poorly characterized. Here, we use a complete locus deletion strain of ESX-5 in M.tb to examine the molecular changes caused by a broad loss in ESX-5 secretory substrates. We confirmed the selective loss of PE/PPE proteins secreted by ESX-5 into both the culture filtrate (CF) and outer mycomembrane (OMM) fractions of the M.tb {Delta}esx5 mutant. In examining other ESX systems, we found that ESX-1 substrate levels were increased in both the CF and OMM fractions of the {Delta}esx5 mutant. Conversely, the ESX-3 locus was transcriptionally repressed upon ESX-5 deletion. We noted that the {Delta}esx5 mutant had altered morphology in the form of wrinkled distortions of the bacterial surface. Likewise, we identified increased susceptibility of the {Delta}esx5 mutant to a variety of large (molecular weight >550 g/mol) antimicrobial compounds, suggesting that an intact ESX-5 system is required for M.tb to exclude such molecules. Our findings suggest that removing the ESX-5 system from M.tb fundamentally alters the properties of the mycobacterial OMM and impacts the expression and secretion activity of other ESX systems. Significance StatementMycobacterium tuberculosis (M.tb) uses the ESX-5 secretion system to export numerous proteins that shape host-pathogen interactions. Here, we found that deleting ESX-5 from M.tb not only prevented the secretion of many ESX-5 substrates but also impacted other ESX systems. The M.tb {Delta}esx5 mutant had increased ESX-1 substrate secretion but reduced ESX-3 expression. In addition, the M.tb {Delta}esx5 mutant displayed altered cell surface morphology and increased vulnerability to large antibiotic drugs, suggesting a critical role for ESX-5 for maintaining outer membrane integrity. These findings highlight ESX-5 as a central modulator of secretion and cell envelope composition with implications for drug targeting and vaccine development.

microbiology↗

bioRxiv2022

The genome wide analysis deciphered virulence factors and secondary metabolites of Chryseobacterium and description of Chryseobacterium indicum sp. nov.

This study describes a novel bacterium, Chryseobacterium strain PS-8, isolated from the skin of freshwater pufferfish (Tetraodon cutcutia). Cells are Gram-negative. The genome size is 4.72 Mb, and the G+C content is 36.4%. The in silico DDH homology, ANI, and AAI were below the cutoff value, 70% and 95% to 96%, respectively, as required for novel species delineation. The phylogenomic analysis using core and non-recombinant core genes, the strain PS-8 showed clustering with Chryseobacterium gambrini DSM 18014T. Generally, Chryseobacterium species are considered opportunistic pathogens. Prediction of the virulence genes revealed genes for adherence, biofilm and stability (glf, lpxD, clpE, cps0, clpE, IlpA), proliferation (tufA, cap8E, galE, kfic, bioB, clpP, kdtB, csgD, carB), resistance to immune response (htpB, katA, wbtl, sodB, kpsF), and host-defense evasion system (kpst, clpE, kfic, ybtQ, cap8G, clpP). The cladogram based upon the virulence genes showed a similar phylogeny amongst the Chryseobacterium species. Additionally, secondary metabolites producing gene clusters were identified, including microviridin, resorcinol and polyene, terpene, etc. Our study showed that strain PS-8 constitute a novel species for which Chryseobacterium indicum sp. nov. is proposed. The type strain is PS-8T (= TBRC 15233T = NBRC 115235T).

microbiology↗

bioRxiv2021

Pufferfish (Tetraodon cutcutia) sampled from freshwater river serves an intermediate reservoir of sucrose nonfermenting variant of Vibrio cholerae PS-4.

This study describes the genomic characteristics of Vibrio cholerae strain PS-4, which fails to ferment sucrose on thiosulfate{square}citrate{square}bile salt{square}sucrose (TCBS) agar medium. This bacterium was isolated from skin mucus of a freshwater pufferfish. In order to understand the sucrose nonfermenting phenotype, the genome of the strain PS-4 was sequenced. The gene encoding the sucrose specific phosphotransferase system IIB (sucR) was absent, resulting for the defective sucrose fermenting phenotype. In contrast, genes encoding glucose-specific transport system IIB (ptsG) and fructose specific transport system IIB (fruA) were present and showing acid production while growing with respective sugars. The overall genome relatedness indices (OGRI) such as in silico DDH, average nucleotide identity (ANI) and average amino acid identity (AAI) were above the threshold value, i.e., 70% and 95-96%, respectively. Phylogenomic analysis based on genome wide core genes and the non-recombinant core-genes, strain PS-4 clustered with Vibrio cholerae ATCC 14035T. Further, genes encoding ctx, zot, ace, tcp and rfb were absent. It showed hemolytic activity and reacted strongly to the R antibody. This is the first report on the existence of Vibrio cholerae from the pufferfish adds to our knowledge of the new ecological niche of this bacterium. IMPORTANCEVibrio cholerae, the causative agent of cholera, is a natural inhabitant of aquatic environments. It ferments sucrose, producing characteristic yellow colonies on TCBS agar. Vibrio cholerae strain PS-4 described in this study was a sucrose nonfermenting variant associated with pufferfish skin and did not produce yellow colonies on TCBS agar. Genes encoding sucrose specific phosphotransferase system IIB (sucR) was absent. The observed phenotype in the characteristic metabolic pathway indicates niche specific adaptive evolution for this bacterium. Our study suggested that nonfermenting phenotype on TCBS agar may not be considered always for Vibrio cholerae species delineation.

microbiology↗

bioRxiv2020

Phenotypic and molecular characterization of extended spectrum β-lactamase producing Escherichia coli and Klebsiella pneumoniae isolates from various samples of animal origin from Assam, India

Extended-spectrum beta-lactamase (ESBL) producing Enterobacteriaceae has become a major threat globally. Here we have characterized ESBL producing E. coli and K. pneumoniae from various sources, studied antibiogram and resistance gene profiles. Out of 385 samples, 31 (8.05%) were positive for ESBL producing E. coli. Such isolates could be recovered from 10.05, 8.33, 15.63, 6.67 and 4.35 per cent of cattle milk, curd, chicken, pork and cattle faeces samples, respectively. A total of 59 (15.32%) samples were positive for ESBL producing K. pneumoniae, which were isolated from 14.35, 6.25, 21.43 and 34.78 per cent cattle milk, chicken, beef and cattle faeces, respectively. All the 90 isolates were confirmed as ESBL producers by CDT and ESBL-E strip tests. Antibiogram revealed that 74.19% and 69.49% of the ESBL producing E. coli and K. pneumoniae isolates, respectively showed resistance to ceftizoxime, 25.81% and 23.73% to both co-trimoxazole and tetracycline, 19.35% and 25.42% to ciprofloxacin, 9.68% and 16.95% to chloramphenicol, 3.23% and 5.08% to pipercillin-tazobactam, and 3.23% and 3.39% to gentamicin. Resistance gene profiling showed blaCTX-M gene as most predominant (100%). The blaTEM gene was found in 54.84% and 55.93%, blaSHV gene in 90.32% and 77.97%, Sul 1 gene in 90.32% and 86.44% of ESBL producing E. coli and K. pneumoniae isolates, respectively. The Int1 gene was detected in 70.97% and 62.71% isolates, while qnrB gene was found in 3.23% and 10.17% of E. coli and K. pneumoniae isolates, respectively.

molecular biology↗