Biology subjects

Shukla, S. K.

Publications and source records attributed to Shukla, S. K..

6 recordsLinked to original sources

bioRxiv2024

Trace DNA from kill sites identifies predating tigers

Predation ecology and evidence-based conflict management strategies require reliable and accurate identification of individual predators. Identifying predators is, however, complex, as they are secretive and individual identification is difficult. Trace DNA that predators leave behind at kill sites might provide an effective strategy to identify them but remains poorly evaluated at scale. We use non-invasive genetic samples from kill sites to assess their utility for predator identification. We systematically investigated 198 livestock kills in two critical source tiger populations in central India: Kanha and Bandhavgarh Tiger Reserves. We collected 342 salivary swabs from carcasses, 33 scat and 395 shed hair samples as potential sources of predator DNA, and individual tigers were identified using up to 123 SNP markers. All three sample sources identified predator species with high success (>95%). We identified individuals (with at least one sample per kill site, based on >40 SNPs) at 86% of all kill sites where tigers were detected. Shed hair samples were most effective for individual identification, followed by saliva and scat. Sample source and sampling season were the primary determinants of the number of SNPs typed per sample and the success of individual identification. Based on the site and type of sample collection, we classify species and individuals into three categories: true predator (high confidence as predator), circumstantial predator (medium confidence) and predator uncertain (low confidence). Individuals were classified as a true predator at 72 sites, circumstantial predator at 34 sites and predator uncertain at 49 sites. Our protocol allowed us to differentiate between predators and scavengers, even when multiple tigers were detected at the same kill site. Surprisingly, [~]40% of Bandhavgarhs tigers were identified at at least one kill site. We suggest that when paired with systematic kill site investigation and sample collection, these methods can be effectively used to understand predation ecology better and facilitate evidence-based conflict management.

genetics↗

bioRxiv2023

DHODH inhibition enhances the efficacy of immune checkpoint blockade by increasing cancer cell antigen presentation

Pyrimidine nucleotide biosynthesis is a druggable metabolic dependency of cancer cells, and chemotherapy agents targeting pyrimidine metabolism are the backbone of treatment for many cancers. Dihydroorotate dehydrogenase (DHODH) is an essential enzyme in the de novo pyrimidine biosynthesis pathway that can be targeted by clinically approved inhibitors. However, despite robust preclinical anticancer efficacy, DHODH inhibitors have shown limited single-agent activity in phase 1 and 2 clinical trials. Therefore, novel combination therapy strategies are necessary to realize the potential of these drugs. To search for therapeutic vulnerabilities induced by DHODH inhibition, we examined gene expression changes in cancer cells treated with the potent and selective DHODH inhibitor brequinar (BQ). This revealed that BQ treatment causes upregulation of antigen presentation pathway genes and cell surface MHC class I expression. Mechanistic studies showed that this effect is 1) strictly dependent on pyrimidine nucleotide depletion, 2) independent of canonical antigen presentation pathway transcriptional regulators, and 3) mediated by RNA polymerase II elongation control by positive transcription elongation factor B (P-TEFb). Furthermore, BQ showed impressive single-agent efficacy in the immunocompetent B16F10 melanoma model, and combination treatment with BQ and dual immune checkpoint blockade (anti-CTLA-4 plus anti-PD-1) significantly prolonged mouse survival compared to either therapy alone. Our results have important implications for the clinical development of DHODH inhibitors and provide a rationale for combination therapy with BQ and immune checkpoint blockade.

cancer biology↗

bioRxiv2022

Association Between Polygenic Risk Score And Gut Microbiome Of Multiple Sclerosis

BackgroundMultiple sclerosis (MS) is a complex autoimmune disease in which both the roles of genetic susceptibility and environmental/microbial factors have been investigated. More than 200 genetic susceptibility variants have been identified along with the dysbiosis of gut microbiota, both independently have been shown to be associated with MS. We hypothesize that MS patients harboring genetic susceptibility variants along with gut microbiome dysbiosis are at a greater risk of exhibiting the disease. We investigated the polygenic risk score for MS in conjunction with gut microbiota in the same cohort of 117 relapsing remitting MS (RRMS) and 26 healthy controls. DNA samples were genotyped using Illuminas Infinium Immuno array-24 v2 chip followed by calculating polygenic risk score and the microbiota was determined by sequencing the V4 hypervariable region of the 16S rRNA gene. ResultsWe identified two clusters of MS patients, Cluster A and B both having a higher polygenic risk score than the control group. The Cluster B with the higher polygenic risk score had a distinct gut microbiota, different than the Cluster A. MS group whose microbiome was similar to that of the control group despite a higher genetic risk score than the control group. This could be due to i) the non-active state of the disease in that group of MS patients at the time of fecal sample collection and/or ii) the restoration of the gut microbiome post disease modifying therapy to treat the MS. ConclusionOur study showed that there seems to be association between polygenic risk score and gut microbiome dysbiosis in triggering the disease in a small cohort of MS patients. The MS Cluster A who have a higher polygenic risk score but microbiome profile similar to that of healthy controls could be due to the remitting phase of the disease or due to the effect of disease modifying therapies.

genetics↗

bioRxiv2017

The first recorded incidence of Deinococcus radiodurans R1 biofilm formation and its implications in heavy metals bioremediation

Radiation tolerant Deinococcus radiodurans R1 is reported to be a potential bacterium for the treatment of low level active wastes. So far there are no reports on the biofilm producing capability of D. radiodurans and heavy metal biosorption. In this study, it was observed that a recombinant D. radiodurans strain with a plasmid harbouring gfp and kanR has formed significant biofilm (~10 m thick). Analysis of biofilm matrix components produced by D. radiodurans showed that the matrix consisted primarily of proteins and carbohydrates with a little amount of extracellular DNA (eDNA). Further, studies showed that D. radiodurans biofilm formation was enhanced at higher concentrations (up to 25 mM) of Ca2+. Further studies on D. radiodurans biofilm showed that Ca2+ enhanced significant biosorption of the heavy metals (Co, Ni). In the presence of 25 mM Ca2+, the D. radiodurans (Kanr) biofilm showed 35% and 25% removal of Co2+ and Ni2+ respectively. While in the absence of Ca2+, D. radiodurans biofilm showed relatively low biosorption of Co (7%) and Ni (3%). Ca2+ also significantly enhanced exopolysaccharide (EPS) production in the biofilm matrix. This infers that EPS could have mediated the heavy metal biosorption. This study signifies the potential use of D. radiodurans biofilm in the remediation of radioactive waste components.\n\nSignificance and Impact of this StudyThis is the first ever recorded study on the Deinococcus radiodurans R1 biofilm. This organism, being the most radioresistant micro-organism ever known, has always been speculated as a potential bacterium to develop a bioremediation process for radioactive heavy metal contaminants. However, the lack of biofilm forming capability proved to be a bottleneck in developing such technology. This study records the first incidence of biofilm formation in a recombinant D. radiodurans, serendipitously, and also discusses its implications in removal of heavy-metals, such as Co and Ni.

microbiology↗

bioRxiv2017

Staphylococcus aureus Biofilm Removal by Targeting Biofilm-Associated Extracellular Proteins

AimAmong cell surface proteins, biofilm-associated protein promotes biofilm development in Staphylococcus aureus strains. Aim of this study was to investigate proteinase-mediated biofilm dispersion in different isolates of S. aureus.\n\nMethods and ResultsMicrotitre plate based biofilm assay showed that 2 g/mL proteinase K significantly inhibited biofilm development in bap-positive S. aureus V329 as well as other S. aureus strains, i.e. SA7, SA10, SA33, SA352 and but not in bap-mutant M556 and SA392 (a weak biofilm producing strain). However, proteinase K treatment on S. aureus planktonic cells showed that there was no inhibition of planktonic growth at any concentration of proteinase K when tested up to 32 g/mL. This observation ruled out the possibility of S. aureus biofilm inhibition by altering the cell viability. Proteinase K treatment upon 24 h old preformed biofilms showed an enhanced dispersion of bap-positive V329 and SA7, SA10, SA33 and SA352 biofilms, however, proteinase K did not affect the bap-mutant S. aureus M556 and SA392 biofilms. Biofilm compositions study before and after proteinase K treatment indicated that Bap might also be involved in eDNA retention in the biofilm matrix that aid in biofilm stability. When proteinase K was used in combination with antibiotics, a synergistic effect in antibiotic efficacy was observed against all biofilm forming S. aureus strains.\n\nConclusionProteinase K inhibited biofilms growth in S. aureus bovine mastitis isolates but did not affect their planktonic growth. An enhanced dispersion of preformed S. aureus biofilms was observed upon proteinase K treatment. Proteinase K treatment with antibiotics showed a synergistic effect against S. aureus biofilms.\n\nSignificance of the studyThe study suggests that dispersing S. aureus by protease can be of use while devising strategies against S. aureus biofilms. Proteinase K treatment has a wider scope for control of S. aureus biofilms.

microbiology↗

bioRxiv2017

An Improved Crystal Violet Assay for Biofilm Quantification in 96-Well Microtitre Plate

Microplates are essential tools for biofilm research since it allows high throughput screening of biofilm forming strains or in the assay of anti-biofilm drugs. However, 96 well microtitre plate based assays share the issue of \"edge effect\". The primary cause of the \"edge effect\" phenomenon is evaporation. As edge effect causes a significant increase in plate rejection rate by introducing experimental error, we improvised the classical crystal violet assay to reduce water loss from the peripheral wells. The improvised method showed a significant reduction in edge effect and minimised error in crystal violet assay

microbiology↗