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.

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.

A Nanopore sequencing-based pharmacogenomic panel to personalize tuberculosis drug dosing.

Rationale: Standardized dosing of anti-tubercular (TB) 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 (PGx) assays that predict metabolism of anti-TB drugs have been lacking. Objectives: To develop a Nanopore sequencing panel and validate its performance in active TB patients to personalize treatment dosing. Measurements and Main Results: We developed a Nanopore sequencing panel targeting 15 single nucleotide polymorphisms (SNP) in 5 genes affecting the metabolism of isoniazid (INH), rifampin (RIF), linezolid and bedaquiline. For validation, we sequenced DNA samples (n=48) from the 1000 genomes project and compared variant calling accuracy with 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 INH and RIF. Results: The PGx panel achieved 100% concordance with Illumina sequencing in variant identification for the samples from the 1000 Genomes Project. In the clinical cohort, coverage was >100x for 1498/1500 (99.8%) amplicons across the 100 samples. One third (33%) of participants were identified as slow, 47% were intermediate and 20% were rapid isoniazid acetylators. Isoniazid clearance was significantly impacted by acetylator status (p<0.0001) with median (IQR) clearances of 11.2 L/h (9.3-13.4), 27.2 L/h (22.0-31.7), and 45.1 L/h (34.1-51.1) in slow, intermediate, and rapid acetylators. Rifampin clearance was 17.3% (2.50-29.9) lower in individuals with homozygous AADAC rs1803155 G>A substitutions (p=0.0015). Conclusion: Targeted sequencing can enable detection of polymorphisms influencing TB drug metabolism on a low-cost, portable instrument to personalize dosing for TB treatment or prevention.

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.

Challenges in Harnessing Shared Within-Host Severe Acute Respiratory Syndrome Coronavirus 2 Variation for Transmission Inference.

Background: The limited variation observed among severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) consensus sequences makes it difficult to reconstruct transmission linkages in outbreak settings. Previous studies have recovered variation within individual SARS-CoV-2 infections but have not yet measured the informativeness of within-host variation for transmission inference. Methods: We performed tiled amplicon sequencing on 307 SARS-CoV-2 samples, including 130 samples from 32 individuals in 14 households and 47 longitudinally sampled individuals, from 4 prospective studies with household membership data, a proxy for transmission linkage. Results: Consensus sequences from households had limited diversity (mean pairwise distance, 3.06 single-nucleotide polymorphisms [SNPs]; range, 0-40). Most (83.1%, 255 of 307) samples harbored at least 1 intrahost single-nucleotide variant ([iSNV] median, 117; interquartile range [IQR], 17-208), above a minor allele frequency threshold of 0.2%. Pairs in the same household shared significantly more iSNVs (mean, 1.20 iSNVs; 95% confidence interval [CI], 1.02-1.39) than did pairs in different households infected with the same viral clade (mean, 0.31 iSNVs; 95% CI, .28-.34), a signal that decreases with increasingly stringent minor allele frequency thresholds. The number of shared iSNVs was significantly associated with an increased odds of household membership (adjusted odds ratio, 1.35; 95% CI, 1.23-1.49). However, the poor concordance of iSNVs detected across sequencing replicates (24.8% and 35.0% above a 0.2% and 1% threshold) confirms technical concerns that current sequencing and bioinformatic workflows do not consistently recover low-frequency within-host variants. Conclusions: Shared within-host variation may augment the information in consensus sequences for predicting transmission linkages. Improving sensitivity and specificity of within-host variant identification will improve the informativeness of within-host variation.

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.

A non-invasive miRNA-based approach in early diagnosis and therapeutics of oral cancer.

Oral or mouth cancer is the 16th most common form of cancer among the world's topmost malignancies. Healthy lifestyle and control of known risk factors can reduce its incidences further. Patients succumb to oral cancer when diagnosed late and lack timely access to tertiary care. Molecular biomarkers might help in early detection of oral cancer. Recently, researchers have identified numerous microRNAs which play a crucial role in promoting and suppressing oral cancers. miRNAs are short non-coding RNA molecules (18-22 nucleotides) that play a pivotal role in regulating gene expression. Understanding the miRNA interplays in oral cancers could augment the development of potential diagnostic, prognostic, and therapeutic tools. Liquid biopsy- a non-invasive approach that has been used lately, allows the determination of miRNAs in biological fluids that play essential roles in tumor suppression and cancer promotion. Herein, we summarize an update on the role of miRNAs in the diagnosis and treatment of oral cancer.

Recent Trends in Photocatalytic Enantioselective Reactions.

Enantioselective synthesis through photocatalysis is one of the highly preferred approaches towards preparation of optically active compounds. This review elaborates and critically analyzes the different strategies of photocatalytic enantioselective reactions through H-bonding, transition metal catalysis, phase-transfer catalysis (PTC), chiral Lewis acid catalysis, N-heterocyclic carbene catalysis, and amine catalysis, and also explores ion pairs. In addition, it explains the different catalysis modes with multifunctional approaches for enantioselective photocatalytic reactions.

SARS-CoV-2 infection drives an inflammatory response in human adipose tissue through infection of adipocytes and macrophages.

Obesity, characterized by chronic low-grade inflammation of the adipose tissue, is associated with adverse coronavirus disease 2019 (COVID-19) outcomes, yet the underlying mechanism is unknown. To explore whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection of adipose tissue contributes to pathogenesis, we evaluated COVID-19 autopsy cases and deeply profiled the response of adipose tissue to SARS-CoV-2 infection in vitro. In COVID-19 autopsy cases, we identified SARS-CoV-2 RNA in adipocytes with an associated inflammatory infiltrate. We identified two distinct cellular targets of infection: adipocytes and a subset of inflammatory adipose tissue-resident macrophages. Mature adipocytes were permissive to SARS-CoV-2 infection; although macrophages were abortively infected, SARS-CoV-2 initiated inflammatory responses within both the infected macrophages and bystander preadipocytes. These data suggest that SARS-CoV-2 infection of adipose tissue could contribute to COVID-19 severity through replication of virus within adipocytes and through induction of local and systemic inflammation driven by infection of adipose tissue-resident macrophages.

Diagnostic and prognostic biomarkers in colorectal cancer and the potential role of exosomes in drug delivery.

Colorectal cancer (CRC) is the third most common cancer with the second most frequent cause of death worldwide. One fourth to one fifth of the CRC cases are detected at advance stage. Early detection of colorectal cancer might help in decreasing mortality and morbidity worldwide. CRC being a heterogeneous disease, new non-invasive approaches are needed to complement and improve the screening and management of CRC. Reliable and early detectable biomarkers would improve diagnosis, prognosis, therapeutic responses, and will enable the prediction of drug response and recurrence risk. Over the past decades molecular research has demonstrated the potentials of CTCs, ctDNAs, circulating mRNAs, ncRNAs, and exosomes as tumor biomarkers. Non-invasive screening approaches using fecal samples for identification of altered gut microbes in CRC is also gaining attention. Exosomes can be potential candidates that can be employed in the drug delivery system. Further, the integration of in vitro, in vivo and in silico models that involve CRC biomarkers will help to understand the interactions occurring at the cellular level. This review summarizes recent update on CRC biomarkers and their application along with the nanoparticles followed by the application of organoid culture in CRC.

Synthesis of N-Benzylideneaniline by Schiff base reaction using Kinnow peel powder as Green catalyst and comparative study of derivatives through ANOVA techniques.

The cheap and easy availability of the Kinnow peel waste has reported various applications due to presence of multifunctional groups. Therefore, in present study we explored its application to synthesize N-Benzylideneaniline and its derivatives based on Schiff base reaction. Kinnow peel powder is characterized by FTIR, TEM, SEM, XRD, EDX, and TGA for functional groups, morphology, surface, elements and thermal stability. Benzaldehyde, aniline, and their derivatives such as 4-methyl benzaldehyde, 4-hydroxy benzaldehyde, 4-methoxy benzaldehyde, and 4-methoxy aniline have been used to compare the efficacy of the Schiff base reaction using analysis of variance (ANOVA) and it has been observed that combination of Aniline and benzaldehyde for Schiff base reaction provided 85% yield of relative product.

Molecular markers in cancer.

Cancer remains a common health issue having significant socioeconomic burden worldwide. Despite the awareness campaigns, cancer cases continue to increase due to an aging population and lack of early detection biomarkers. Accordingly, much research has focused on non-traditional approaches which include novel imaging modalities and liquid biopsy. In addition, a considerable number of biomacromolecules as well as other biomarkers have been identified to further explore their potential use as diagnostic, prognostic and therapeutic tools. In this review, we provide an update on these new findings and explore their clinical application in cancer.

Detection of M. tuberculosis in the environment as a tool for identifying high-risk locations for tuberculosis transmission.

Tuberculosis (TB) remains a leading cause of infectious mortality globally, yet most cases cannot be epidemiologically linked even with extensive contact investigations and whole genome sequencing. Consequently, there remain major gaps in our understanding of where and when M. tuberculosis (Mtb) exposures occur. We aimed to investigate whether Mtb can be detected in environments where TB patients were recently present, which could serve as a tool for characterizing exposure risk. We collected 389 environment surface (ES) swabs from two high TB burden prisons in Brazil, sampling 41 (n = 340) cells occupied by individuals with active TB and 7 (n = 49) cells from individuals without TB. In a subset of pooled swabs (n = 6) and a swab from a cigarette lighter from the cell with active TB patients, we enriched Mtb DNA using RNA-bait hybrid capture assays and performed whole genome sequencing. In prison cells, Mtb DNA was detected in 55/340 (16 %) of ES swabs from cells occupied by active TB patients and none (0/49) from cells in which no active TB patients were present. Mtb was detected in 13/16 (81 %) prison cells occupied by the individuals with high/medium sputum Xpert Mtb load and 8/25 (32 %) with low/very low sputum Mtb load (p = 0.003). Seven hybrid capture samples had a median genomic coverage of 140×. rpoB mutations conferring high-level rifampin resistance were detected in 3/7 ES swabs. Mtb was frequently detectable in environments recently occupied by individuals with active TB. This approach could be applied in congregate environments to identify and characterize high-risk settings for Mtb exposure.

Gastrointestinal symptoms and fecal shedding of SARS-CoV-2 RNA suggest prolonged gastrointestinal infection.

Background: COVID-19 manifests with respiratory, systemic, and gastrointestinal (GI) symptoms.1, SARS-CoV-2 RNA is detected in respiratory and fecal samples, and recent reports demonstrate viral replication in both the lung and intestinal tissue.2, 3, 4 Although much is known about early fecal RNA shedding, little is known about long-term shedding, especially in those with mild COVID-19. Furthermore, most reports of fecal RNA shedding do not correlate these findings with GI symptoms.5. Methods: We analyzed the dynamics of fecal RNA shedding up to 10 months after COVID-19 diagnosis in 113 individuals with mild to moderate disease. We also correlated shedding with disease symptoms. Findings: Fecal SARS-CoV-2 RNA is detected in 49.2% [95% confidence interval, 38.2%-60.3%] of participants within the first week after diagnosis. Whereas there was no ongoing oropharyngeal SARS-CoV-2 RNA shedding in subjects at 4 months, 12.7% [8.5%-18.4%] of participants continued to shed SARS-CoV-2 RNA in the feces at 4 months after diagnosis and 3.8% [2.0%-7.3%] shed at 7 months. Finally, we found that GI symptoms (abdominal pain, nausea, vomiting) are associated with fecal shedding of SARS-CoV-2 RNA. Conclusions: The extended presence of viral RNA in feces, but not in respiratory samples, along with the association of fecal viral RNA shedding with GI symptoms suggest that SARS-CoV-2 infects the GI tract and that this infection can be prolonged in a subset of individuals with COVID-19. Funding: This research was supported by a Stanford ChemH-IMA grant; fellowships from the AACR and NSF; and NIH R01-AI148623, R01-AI143757, and UL1TR003142.

Assessing impact of ventilation on airborne transmission of SARS-CoV-2: a cross-sectional analysis of naturally ventilated healthcare settings in Bangladesh.

OBJECTIVES: To evaluate the risk of exposure to SARS-CoV-2 in naturally ventilated hospital settings by measuring parameters of ventilation and comparing these findings with results of bioaerosol sampling. STUDY DESIGN: Cross-sectional study. STUDY SETTING AND STUDY SAMPLE: The study sample included nine hospitals in Dhaka, Bangladesh. Ventilation characteristics and air samples were collected from 86 healthcare spaces during October 2020 to February 2021. PRIMARY OUTCOME: Risk of cumulative SARS-CoV-2 infection by type of healthcare area. SECONDARY OUTCOMES: Ventilation rates by healthcare space; risk of airborne detection of SARS-CoV-2 across healthcare spaces; impact of room characteristics on absolute ventilation; SARS-CoV-2 detection by naturally ventilated versus mechanically ventilated spaces. RESULTS: The majority (78.7%) of naturally ventilated patient care rooms had ventilation rates that fell short of the recommended ventilation rate of 60 L/s/p. Using a modified Wells-Riley equation and local COVID-19 case numbers, we found that over a 40-hour exposure period, outpatient departments posed the highest median risk for infection (7.7%). SARS-CoV-2 RNA was most frequently detected in air samples from non-COVID wards (50.0%) followed by outpatient departments (42.9%). Naturally ventilated spaces (22.6%) had higher rates of SARS-CoV-2 detection compared with mechanically ventilated spaces (8.3%), though the difference was not statistically significant (p=0.128). In multivariable linear regression with calculated elasticity, open door area and cross-ventilation were found to have a significant impact on ventilation. CONCLUSION: Our findings provide evidence that naturally ventilated healthcare settings may pose a high risk for exposure to SARS-CoV-2, particularly among non-COVID-designated spaces, but improving parameters of ventilation can mitigate this risk.

Variation in Severe Acute Respiratory Syndrome Coronavirus 2 Bioaerosol Production in Exhaled Breath.

We developed a simple, noninvasive mask sampling method to quantify and sequence severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from exhaled breath. We found substantial variation between individuals in SARS-CoV-2 copies exhaled over a 15-minute period, which moderately correlated with nasal swab viral load. Talking was associated with a median of 2 log10 greater exhaled viral copies. Exposure varies substantially between individuals but may be risk stratified by nasal swab viral load and whether the exposure involved conversation.

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.

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.