A Nanopore Sequencing-based Pharmacogenomic Panel to Personalize Tuberculosis Drug Dosing.

Rationale: Standardized dosing of antitubercular drugs leads to variable plasma drug levels, which are associated with adverse drug reactions, delayed treatment response, and relapse. Mutations in genes affecting drug metabolism explain considerable interindividual pharmacokinetic variability; however, pharmacogenomic assays that predict metabolism of antitubercular drugs have been lacking. Objectives: We sought to develop a Nanopore sequencing panel and validate its performance in patients with active tuberculosis (TB) to personalize treatment dosing. Methods: We developed a Nanopore sequencing panel targeting 15 SNPs in five genes affecting the metabolism of antitubercular drugs. For validation, we sequenced DNA samples (n = 48) from the 1,000 Genomes Project and compared the variant calling accuracy with that of Illumina genome sequencing. We then sequenced DNA samples from patients with active TB (n = 100) from South Africa on a MinION Mk1C and evaluated the relationship between genotypes and pharmacokinetic parameters for isoniazid (INH) and rifampin (RIF). Measurements and Main Results: The pharmacogenomic panel achieved 100% concordance with Illumina sequencing in variant identification for the samples from the 1,000 Genomes Project. In the clinical cohort, coverage was more than 100× for 1,498 of 1,500 (99.8%) amplicons across the 100 samples. Thirty-three percent, 47%, and 20% of participants were identified as slow, intermediate, and rapid INH acetylators, respectively. INH clearance was 2.2 times higher among intermediate acetylators and 3.8 times higher among rapid acetylators, compared with slow acetylators (P < 0.0001). RIF clearance was 17.3% (2.50-29.9) lower in individuals with homozygous AADAC rs1803155 G→A substitutions (P = 0.0015). Conclusions: Targeted sequencing can enable the detection of polymorphisms that influence TB drug metabolism on a low-cost, portable instrument to personalize dosing for TB treatment or prevention.

Identification of stage-specific differentially expressed genes and SNPs in gastric cancer employing RNA-Seq based transcriptome profiling.

We analysed over 400 million reads obtained from Illumina sequencing of six pairs of libraries representing two each of stage I, II, and III gastric tumors and corresponding normal tissues to identify differentially expressed genes (DEGs), single nucleotide polymorphisms (SNPs), and transcription factors (TFs). In total, 2207 DEGs including 972 upregulated genes and 1235 downregulated genes were detected. Of these, several stage-specific signature genes were identified. The protein-protein interaction networks involving DEGs and TFs were constructed. The KEGG pathway analysis of SNP harbouring genes revealed their involvement in different cancer related pathways like apoptosis, mTOR pathway, and MAPK signaling pathway. The SNP analysis showed implication of host genes in GO categories like immune system process, regulation of signaling, response to stress, and transport. A biased chromosomal distribution of DEGs and SNP harbouring genes was observed. Our study would provide further insights into the complex regulatory mechanisms operating during gastric tumorigenesis.

Pooling Sputum Samples for Efficient Mass Tuberculosis Screening in Prisons.

BACKGROUND: Although systematic tuberculosis screening in high-risk groups is recommended by the World Health Organization (WHO), implementation in prisons has been limited due to resource constraints. Whether Xpert Ultra sputum pooling could be a sensitive and efficient approach to mass screening in prisons is unknown. METHODS: In total, 1280 sputum samples were collected from incarcerated individuals in Brazil during mass screening and tested using Xpert G4. We selected samples for mixing in pools of 4, 8, 12, and 16, which were then tested using Ultra. In each pool, a single positive sample of differing Xpert mycobacterial loads was used. Additionally, 10 pools of 16 negative samples each were analyzed as controls. We then simulated tuberculosis screening at prevalences of 0.5-5% and calculated the cost per tuberculosis case detected at different sputum pooling sizes. RESULTS: The sensitivity and specificity of sputum pooling were high (sensitivity: 94%; 95% confidence interval [CI]: 88-98; specificity: 100%, 95% CI: 84-100). Sensitivity was greater in pools in which the positive sample had a high mycobacterial load compared to those that were very low (100% vs 88%). In settings with a higher tuberculosis prevalence, pools of 4 and 8 were more efficient than larger pool sizes. Larger pools decreased the costs by 87% at low prevalences, whereas smaller pools led to greater cost savings at higher prevalence at higher prevalences (57%). CONCLUSIONS: Sputum pooling using Ultra was a sensitive strategy for tuberculosis screening. This approach was more efficient than individual testing across a broad range of simulated tuberculosis prevalence settings and could enable active case finding to be scaled while containing costs.

A Rapid Pharmacogenomic Assay to Detect NAT2 Polymorphisms and Guide Isoniazid Dosing for Tuberculosis Treatment.

Rationale: Standardized dosing of antitubercular drugs contributes to a substantial incidence of toxicities, inadequate treatment response, and relapse, in part due to variable drug concentrations achieved. SNPs in the NAT2 (N-acetyltransferase-2) gene explain the majority of interindividual pharmacokinetic variability of isoniazid (INH). However, an obstacle to implementing pharmacogenomic-guided dosing is the lack of a point-of-care assay. Objectives: To develop and test a NAT2 classification algorithm, validate its performance in predicting isoniazid clearance, and develop a prototype pharmacogenomic assay. Methods: We trained random forest models to predict NAT2 acetylation genotype from unphased SNP data using a global collection of 8,561 phased genomes. We enrolled 48 patients with pulmonary tuberculosis, performed sparse pharmacokinetic sampling, and tested the acetylator prediction algorithm accuracy against estimated INH clearance. We then developed a cartridge-based multiplex quantitative PCR assay on the GeneXpert platform and assessed its analytical sensitivity on whole blood samples from healthy individuals. Measurements and Main Results: With a 5-SNP model trained on two-thirds of the data (n = 5,738), out-of-sample acetylation genotype prediction accuracy on the remaining third (n = 2,823) was 100%. Among the 48 patients with tuberculosis, predicted acetylator types were 27 (56.2%) slow, 16 (33.3%) intermediate, and 5 (10.4%) rapid. INH clearance rates were lowest in predicted slow acetylators (median 14.5 L/h), moderate in intermediate acetylators (median 40.3 L/h), and highest in fast acetylators (median 53.0 L/h). The cartridge-based assay accurately detected all allele patterns directly from 25 µl of whole blood. Conclusions: An automated pharmacogenomic assay on a platform widely used globally for tuberculosis diagnosis could enable personalized dosing of INH.

Comprehensive RNAseq analysis for identification of genes expressed under heat stress in lentil.

Lentils are highly sensitive to abrupt increases in temperature during the mid to late reproductive stages, leading to severe biomass and seed yield reduction. Therefore, we carried out an RNAseq analysis between IG4258 (heat tolerant) and IG3973 (heat sensitive) lentil genotypes at the reproductive stage under both normal and heat stress conditions in the field. It resulted in 209,549 assembled transcripts and among these 161,809 transcripts had coding regions, of which 94,437 transcripts were annotated. The differential gene expression analysis showed upregulation of 678 transcripts and downregulation of 680 transcripts between the tolerant and sensitive genotypes at the early reproductive stage. While 76 transcripts were upregulated and 47 transcripts were downregulated at the late reproductive stage under heat stress conditions. The validation of 12 up-or downregulated transcripts through RT-PCR corresponded well with the expression analysis data of RNAseq, with a correlation of R2  = 0.89. Among these transcripts, the DN364_c1_g1_i9 and DN2218_c0_g1_i5 transcripts encoded enzymes involved in the tryptophan pathway, indicating that tryptophan biosynthesis plays a role under heat stress in lentil. Moreover, KEGG pathways enrichment analysis identified transcripts associated with genes encoding proteins/regulating factors related to different metabolic pathways including signal transduction, fatty acid biosynthesis, rRNA processing, ribosome biogenesis, gibberellin (GA) biosynthesis, and riboflavin biosynthesis. This analysis also identified 6852 genic-SSRs leading to the development of 4968 SSR primers that are potential genomic resources for molecular mapping of heat-tolerant genes in lentil.

Integrating transcriptomic and proteomic data for accurate assembly and annotation of genomes.

Complementing genome sequence with deep transcriptome and proteome data could enable more accurate assembly and annotation of newly sequenced genomes. Here, we provide a proof-of-concept of an integrated approach for analysis of the genome and proteome of Anopheles stephensi, which is one of the most important vectors of the malaria parasite. To achieve broad coverage of genes, we carried out transcriptome sequencing and deep proteome profiling of multiple anatomically distinct sites. Based on transcriptomic data alone, we identified and corrected 535 events of incomplete genome assembly involving 1196 scaffolds and 868 protein-coding gene models. This proteogenomic approach enabled us to add 365 genes that were missed during genome annotation and identify 917 gene correction events through discovery of 151 novel exons, 297 protein extensions, 231 exon extensions, 192 novel protein start sites, 19 novel translational frames, 28 events of joining of exons, and 76 events of joining of adjacent genes as a single gene. Incorporation of proteomic evidence allowed us to change the designation of more than 87 predicted "noncoding RNAs" to conventional mRNAs coded by protein-coding genes. Importantly, extension of the newly corrected genome assemblies and gene models to 15 other newly assembled Anopheline genomes led to the discovery of a large number of apparent discrepancies in assembly and annotation of these genomes. Our data provide a framework for how future genome sequencing efforts should incorporate transcriptomic and proteomic analysis in combination with simultaneous manual curation to achieve near complete assembly and accurate annotation of genomes.

Detection, survival and infectious potential of Mycobacterium tuberculosis in the environment: a review of the evidence and epidemiological implications.

Much remains unknown about Mycobacterium tuberculosis transmission. Seminal experimental studies from the 1950s demonstrated that airborne expulsion of droplet nuclei from an infectious tuberculosis (TB) patient is the primary route of transmission. However, these findings did not rule out other routes of M. tuberculosis transmission. We reviewed historical scientific evidence from the late 19th/early 20th century and contemporary studies investigating the presence, persistence and infectiousness of environmental M. tuberculosis We found both experimental and epidemiological evidence supporting the presence and viability of M. tuberculosis in multiple natural and built environments for months to years, presumably following contamination by a human source. Furthermore, several studies confirm M. tuberculosis viability and virulence in the environment using guinea pig and mouse models. Most of this evidence was historical; however, several recent studies have reported consistent findings of M. tuberculosis detection and viability in the environment using modern methods. Whether M. tuberculosis in environments represents an infectious threat to humans requires further investigation; this may represent an untapped source of data with which to further understand M. tuberculosis transmission. We discuss potential opportunities for harnessing these data to generate new insights into TB transmission in congregate settings.

Quantitative Proteomic and Phosphoproteomic Analysis of H37Ra and H37Rv Strains of Mycobacterium tuberculosis.

Mycobacterium tuberculosis, the causative agent of tuberculosis, accounts for 1.5 million human deaths annually worldwide. Despite efforts to eradicate tuberculosis, it still remains a deadly disease. The two best characterized strains of M. tuberculosis, virulent H37Rv and avirulent H37Ra, provide a unique platform to investigate biochemical and signaling pathways associated with pathogenicity. To delineate the biomolecular dynamics that may account for pathogenicity and attenuation of virulence in M. tuberculosis, we compared the proteome and phosphoproteome profiles of H37Rv and H37Ra strains. Quantitative phosphoproteomic analysis was performed using high-resolution Fourier transform mass spectrometry. Analysis of exponential and stationary phases of these strains resulted in identification and quantitation of 2709 proteins along with 512 phosphorylation sites derived from 257 proteins. In addition to confirming the presence of previously described M. tuberculosis phosphorylated proteins, we identified 265 novel phosphorylation sites. Quantitative proteomic analysis revealed more than five-fold upregulation of proteins belonging to virulence associated type VII bacterial secretion system in H37Rv when compared to those in H37Ra. We also identified 84 proteins, which exhibited changes in phosphorylation levels between the virulent and avirulent strains. Bioinformatics analysis of the proteins altered in their level of expression or phosphorylation revealed enrichment of pathways involved in fatty acid biosynthesis and two-component regulatory system. Our data provides a resource for further exploration of functional differences at molecular level between H37Rv and H37Ra, which will ultimately explain the molecular underpinnings that determine virulence in tuberculosis.

Genome-Wide Survey of Genes Under Positive Selection in Avian Pathogenic Escherichia coli Strains.

The ability to obtain bacterial genomes from the same host has allowed for comparative studies that help in the understanding of the molecular evolution of specific pathotypes. Avian pathogenic Escherichia coli (APEC) is a group of extraintestinal strains responsible for causing colibacillosis in birds. APEC is also suggested to possess a role as a zoonotic agent. Despite its importance, APEC pathogenesis still has several cryptic pathogenic processes that need to be better understood. In this work, a genome-wide survey of eight APEC strains for genes with evidence of recombination revealed that ∼14% of the homologous groups evaluated present signs of recombination. Enrichment analyses revealed that nine Gene Ontology (GO) terms were significantly more represented in recombinant genes. Among these GO terms, several were noted to be ATP-related categories. The search for positive selection in these APEC genomes revealed 32 groups of homologous genes with evidence of positive selection. Among these groups, we found several related to cell metabolism, as well as several uncharacterized genes, beyond the well-known virulence factors ompC, lamB, waaW, waaL, and fliC. A GO term enrichment test showed a prevalence of terms related to bacterial cell contact with the external environment (e.g., viral entry into host cell, detection of virus, pore complex, bacterial-type flagellum filament C, and porin activity). Finally, the genes with evidence of positive selection were retrieved from genomes of non-APEC strains and tested as were done for APEC strains. The result revealed that none of the groups of genes presented evidence of positive selection, confirming that the analysis was effective in inferring positive selection for APEC and not for E. coli in general, which means that the study of the genes with evidence of positive selection identified in this study can contribute for the better understanding of APEC pathogenesis processes.

Fimbria-Encoding Gene yadC Has a Pleiotropic Effect on Several Biological Characteristics and Plays a Role in Avian Pathogenic Escherichia coli Pathogenicity.

The extraintestinal pathogen termed avian pathogenic Escherichia coli (APEC) is known to cause colibacillosis in chickens. The molecular basis of APEC pathogenesis is not fully elucidated yet. In this work, we deleted a component of the Yad gene cluster (yadC) in order to understand the role of Yad in the pathogenicity of the APEC strain SCI-07. In vitro, the transcription level of yadC was upregulated at 41°C and downregulated at 22°C. The yadC expression in vivo was more pronounced in lungs than in spleen, suggesting a role in the early steps of the infection. Chicks infected with the wild-type and mutant strains presented, respectively, 80% and 50% mortality rates. The ΔyadC strain presented a slightly decreased ability to adhere to HeLa cells with or without the d-mannose analog compared with the wild type. Real-time PCR (RT-PCR) assays showed that fimH was downregulated (P < 0.05) and csgA and ecpA were slightly upregulated in the mutant strain, showing that yadC modulates expression of other fimbriae. Bacterial internalization studies showed that the ΔyadC strain had a lower number of intracellular bacteria recovered from Hep-2 cells and HD11 cells than the wild-type strain (P < 0.05). Motility assays in soft agar demonstrated that the ΔyadC strain was less motile than the wild type (P < 0.01). Curiously, flagellum-associated genes were not dramatically downregulated in the ΔyadC strain. Taken together, the results show that the fimbrial adhesin Yad contributes to the pathogenicity and modulates different biological characteristics of the APEC strain SCI-07.

Does Local Infiltration of Anesthesia Reduce Port-site Pain in Gynecological Laparoscopic Surgeries? A Pilot Study.

Objectives: The objectives of this study were to evaluate the visual analog scale (VAS) score in patients receiving port-site bupivacaine infiltration in gynecological laparoscopic surgeries and to compare it with those receiving placebo and to evaluate the additional analgesic requirement in the first 24 h after surgery. Materials and Methods: A prospective interventional study was conducted on 60 women scheduled for benign gynecological laparoscopic surgeries. Patients were randomized into two groups using an alternative sequential method of allocation. Approval from the Institute's Ethics Committee was sought. Informed written consent was taken from all the patients. All laparoscopic surgeries were performed under general anesthesia. Double-blinding was done. A VAS with a 10 cm vertical score ranging from "no pain" to "worst possible pain" was used to assess the postoperative pain when the patient awakened in the operating room (2 h after surgery), then after 6 and 24 h. The primary outcome measured was pain perception by the patient (as VAS scores), and the secondary outcome was the need for additional analgesia. Results: Comparison of both groups with the VAS score shows P > 0.001, i.e., nonsignificant in all the groups. Additional analgesics were required in 56% of the patients in the intervention group and 60% of the patients in the control group; however, 44% and 40% of the patients from the intervention and control groups, respectively, do not require any additional analgesic in the postoperative period. Conclusion: The local infiltration of bupivacaine does not significantly reduce the port-site postoperative pain in gynecological laparoscopic surgeries.

Better among the two for Burn Mortality Prediction in Developing Nations: Revised Baux or Modified Abbreviated Burn Severity Index?

Background: Burns is one of the leading causes of mortality in developing countries like India. Most of the major burns requiring hospital care are not triaged adequately for the use of medical resources. An efficient mortality predicting scale would not only help in better care to those who will benefit the most but also make it easy to explain to patient's attendants. Among the various tools, revised Baux (rBaux) and modified Abbreviated Burn Severity Index (ABSI) are two most commonly used scales in developed nations. We proposed this study to analyze the reliability of these two scoring scales in our burn population. Aim: This study aimed to retrospectively study the two scoring systems and analyze them for their reliability in predicting mortality compared to actual observed mortality in each case. Materials and Methods: This study was conducted on all burn patients admitted to the intensive care unit of our hospital. Data on their demographic profile, total burn surface area, thickness of burns, inhalational injury, and other comorbidities were collected from files. rBaux and modified ABSI (mABSI) were calculated. The end result in the form of survival or nonsurvival was also recorded. Appropriate statistical analysis using Mann-Whitney U-test, Chi-square test, and receiver operator characteristic curve was done to look for a better scoring system out of the two. Results: A total of 504 patients were included in the study, out of which 337 were survivors. Female gender was not a risk factor for mortality in our study. The median rBaux score in the survivor group was 100 (80-110) and in nonsurvivor group was 111 (103-123). The median mABSI score in the survivor group was 8 (7-9) and in nonsurvivor group was 10 (9-11). The area under the receiver operating characteristic curve shows mABSI having better specificity for predicting mortality. rBAUX, though more sensitive, overestimates mortality than actual observed mortality. Conclusion: mABSI predicts mortality better than rBaux. A multicentric prospective study is recommended for mABSI to be used as a standard mortality predictor in burns in India.

Natural variability and heritability of root-nodulation traits in chickpea (Cicer arietinum L.) minicore.

The existence of large variations for nodulation traits in chickpea minicore was revealed and genetic materials for beneficial biological nitrogen fixation (BNF) traits like early nodulation, high nodulation, and delayed nodule senescence were identified. Early-nodulating genotypes viz. ICC12968, ICC7867, ICC13816, ICC867, ICC15264, ICC15510, and ICC283 produced > 10 nodule number per plant (NNPP) at 15 as well as 30 days after sowing (DAS). Maximum of 36 NNPP at stage 3 i.e., 253% higher than check cultivar were observed in Iran originated ICC6874. Chickpea minicore showed large variations for nodule mass that ranged up to 850 mg/plant at 60 DAS and 2290 mg/plant at 90 DAS. Strong positive correlation was found between nodule fresh weight and specific weight at stage 3 (0.69) and stage 4 (0.76). Besides these, few slight positive significant correlations were also observed viz., nodule number per plant at stage 3 and 4 (0.45), nodule fresh weight at stage 3 and 4 (0.39). Principal component analysis (PCA) indicated that dimensions 1 (21%), 2 (17.6%), and 3 (13%) accounted for a substantial portion of the phenotypic variance, each contributing more than 10%. Accessions viz. ICC1431, ICC13599, ICC13764, and ICC13863 with pink active root nodules and high nodule biomass at later crop growth stages are considered as genetic resources to extend the BNF support in chickpea. High broad-sense heritability values of 76.43 and 90.23 were observed for early nodulation and delayed nodule senescence, respectively. Hence, the identified genotypes for early nodulation and delayed nodule senescence can be used for improving symbiotic efficiency in chickpea. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03908-1.

Green synthesis of silver nanoparticle for catalytic applications and priming study by seed germination.

Silver nanoparticles (AgNPs) have been successfully synthesized using leaf extract of Neem (Azadirachta Indica), Mint (Mentha Piperita), Tulsi (Ocimum Tenuiflorum), Bermuda grass (Cynodon Dactylon) and silver salt. As plant extracts produce best capping material for the stabilization of nanoparticles. AgNPs were characterized by UV-Vis spectroscopy in range of 200-800 nm and transmission electron microscopy TEM, XRD and FTIR. The nanoparticles synthesized were mainly in sizes between 25 and 100 nm. They appeared to be spherical, nanotriangles and irregular in shape. Catalytic application was observed for all the aqueous solution of leaves, quantity taken was 1 ml, 2 ml, 3 ml, 4 ml and 5 ml. Furthermore, prepared Ag nanoparticles are also used for seed germination.

Culture-Free Whole Genome Sequencing of Mycobacterium tuberculosis Using Ligand-Mediated Bead Enrichment Method.

Background: Direct whole genome sequencing (WGS) of Mycobacterium tuberculosis (Mtb) can be used as a tool to study drug resistance, mixed infections, and within-host diversity. However, WGS is challenging to obtain from clinical samples due to low number of bacilli against a high background. Methods: We prospectively collected 34 samples (sputum, n = 17; bronchoalveolar lavage, n = 13; and pus, n = 4) from patients with active tuberculosis (TB). Prior to DNA extraction, we used a ligand-mediated magnetic bead method to enrich Mtb from clinical samples and performed WGS on Illumina platform. Results: Mtb was definitively identified based on WGS from 88.2% (30/34) of the samples, of which 35.3% (12/34) were smear negative. The overall median genome coverage was 15.2% (interquartile range [IQR], 7.7%-28.2%). There was a positive correlation between load of bacilli on smears and genome coverage (P < .001). We detected 58 genes listed in the World Health Organization mutation catalogue in each positive sample (median coverage, 85% [IQR, 61%-94%]), enabling the identification of mutations missed by routine diagnostics. Mutations causing resistance to rifampicin, isoniazid, streptomycin, and ethambutol were detected in 5 of 34 (14.7%) samples, including the rpoB S441A mutation that confers resistance to rifampicin, which is not covered by Xpert MTB/RIF. Conclusions: We demonstrate the feasibility of magnetic bead-based enrichment for culture-free WGS of Mtb from clinical specimens, including smear-negative samples. This approach can also be integrated with low-cost sequencing workflows such as targeted sequencing for rapid detection of Mtb and drug resistance.

A multilectin affinity approach for comparative glycoprotein profiling of rheumatoid arthritis and spondyloarthropathy.

BACKGROUND: Arthritis refers to inflammation of joints and includes common disorders such as rheumatoid arthritis (RA) and spondyloarthropathies (SpAs). These diseases differ mainly in terms of their clinical manifestations and the underlying pathogenesis. Glycoproteins in synovial fluid might reflect the disease activity status in the joints affected by arthritis; yet they have not been systematically studied previously. Although markers have been described for assisting in the diagnosis of RA, there are currently no known biomarkers for SpA. MATERIALS AND METHODS: We sought to determine the relative abundance of glycoproteins in RA and SpA by lectin affinity chromatography coupled to iTRAQ labeling and LC-MS/MS analysis. We also used ELISA to validate the overexpression of VCAM-1, one of the candidate proteins identified in this study, in synovial fluid from RA patients. RESULTS AND DISCUSSION: We identified proteins that were previously reported to be overexpressed in RA including metalloproteinase inhibitor 1 (TIMP1), myeloperoxidase (MPO) and several S100 proteins. In addition, we discovered several novel candidates that were overexpressed in SpA including Apolipoproteins C-II and C-III and the SUN domain-containing protein 3 (SUN3). Novel molecules found overexpressed in RA included extracellular matrix protein 1 (ECM1) and lumican (LUM). We validated one of the candidate biomarkers, vascular cell adhesion molecule 1 (VCAM1), in 20 RA and SpA samples using ELISA and confirmed its overexpression in RA (p-value <0.01). Our quantitative glycoproteomic approach to study arthritic disorders should open up new avenues for additional proteomics-based discovery studies in rheumatological disorders.

Proteomic analysis of purified protein derivative of Mycobacterium tuberculosis.

BACKGROUND: Purified protein derivative (PPD) has been used for more than half a century as an antigen for the diagnosis of tuberculosis infection based on delayed type hypersensitivity. Although designated as "purified," in reality, the composition of PPD is highly complex and remains ill-defined. In this report, high resolution mass spectrometry was applied to understand the complexity of its constituent components. A comparative proteomic analysis of various PPD preparations and their functional characterization is likely to help in short-listing the relevant antigens required to prepare a less complex and more potent reagent for diagnostic purposes. RESULTS: Proteomic analysis of Connaught Tuberculin 68 (PPD-CT68), a tuberculin preparation generated from M. tuberculosis, was carried out in this study. PPD-CT68 is the protein component of a commercially available tuberculin preparation, Tubersol, which is used for tuberculin skin testing. Using a high resolution LTQ-Orbitrap Velos mass spectrometer, we identified 265 different proteins. The identified proteins were compared with those identified from PPD M. bovis, PPD M. avium and PPD-S2 from previous mass spectrometry-based studies. In all, 142 proteins were found to be shared between PPD-CT68 and PPD-S2 preparations. Out of the 354 proteins from M. tuberculosis-derived PPDs (i.e. proteins in either PPD-CT68 or PPD-S2), 37 proteins were found to be shared with M. avium PPD and 80 were shared with M. bovis PPD. Alignment of PPD-CT68 proteins with proteins encoded by 24 lung infecting bacteria revealed a number of similar proteins (206 bacterial proteins shared epitopes with 47 PPD-CT68 proteins), which could potentially be involved in causing cross-reactivity. The data have been deposited to the ProteomeXchange with identifier PXD000377. CONCLUSIONS: Proteomic and bioinformatics analysis of different PPD preparations revealed commonly and differentially represented proteins. This information could help in delineating the relevant antigens represented in various PPDs, which could further lead to development of a lesser complex and better defined skin test antigen with a higher specificity and sensitivity.

Non-fat milk solids attenuate hypercholesterolemic effect of conjugated linoleic acid.

The present study instigates the notion that non-fat milk has a beneficial effect on hypercholesterolemia caused by dietary conjugated linoleic acid (CLA). Two groups of mice (6 per group) were fed synthetic CLA (predominantly a mixture of t10, c12 and c9, t11 isomers) at 1 % level in the diet with or without skim milk powder (SMP). Another group (control) was fed a similar diet without CLA and SMP. The study was conducted for 60 days (d). Daily feed intake and weekly body weight were recorded. Plasma lipid profile was determined monthly. At the conclusion of experiment, animals were sacrificed and liver tissue was removed to assay cholesterol and triacylglycerols (TAG). Following 60 day feeding, the plasma total cholesterol content increased significantly in CLA group (218 mg/dl) than in control group (167 mg/dl), while in CLA + SMP group (fed non-fat milk solids along with CLA), it was similar to that in control group. The fractions of high density lipoproteins (HDL) and low density lipoprotein + very low density lipoprotein (LDL + VLDL) cholesterol as well as triacylglycerols (TAG) in plasma increased in CLA group, but in CLA + SMP group it remained similar to the control levels. Thus, it is concluded that dietary CLA increases plasma cholesterol and TAG contents in mice on normocholesterolemic diet and the inclusion of non-fat milk solids reverses this effect.

Reduced graphene Oxide/NiO based nano-composites for the efficient removal of alizarin dye, indigo dye and reduction of nitro aromatic compounds.

Removal of alizarin red S (ARS) and Indigo dye from aqueous media and reduction of nitro aromatic compounds are successfully done under mild condition by using reduced Graphene Oxide-Nickel Oxide (rGO-NiO) nanocomposite as catalyst. RGO-NiO is well characterized by different analytical techniques. Morphology, structural, and composition studies done by HRTEM, FESEM, EDX, TGA, FTIR, XPS, Raman spectroscopy, and XRD. RGO-NiO nanocomposite has high stability for the removal of ARS, Indigo dye, reduction reaction nitro aromatic compounds.

Implication of gut microbes and its metabolites in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is the third most common cancer with a significant impact on loss of life. In 2020, nearly 1.9 million new cases and over 9,35,000 deaths were reported. Numerous microbes that are abundant in the human gut benefit host physiology in many ways. Although the underlying mechanism is still unknown, their association appears to be crucial in the beginning and progression of CRC. Diet has a significant impact on the microbial composition and may increase the chance of getting CRC. Increasing evidence points to the gut microbiota as the primary initiator of colonic inflammation, which is connected to the development of colonic tumors. However, it is unclear how the microbiota contributes to the development of CRCs. Patients with CRC have been found to have dysbiosis of the gut microbiota, which can be identified by a decline in commensal bacterial species, such as those that produce butyrate, and a concurrent increase in harmful bacterial populations, such as opportunistic pathogens that produce pro-inflammatory cytokines. We believe that using probiotics or altering the gut microbiota will likely be effective tools in the fight against CRC treatment. PURPOSE: In this review, we revisited the association between gut microbiota and colorectal cancer whether cause or effect. The various factors which influence gut microbiome in patients with CRC and possible mechanism in relation with development of CRC. CONCLUSION: The clinical significance of the intestinal microbiota may aid in the prevention and management of CRC.