The multi-dimensional impact of captopril modification on human serum albumin.

Captopril is a thiol drug, widely used for the management of hypertension and cardiovascular diseases. Reactive thiols are found to covalently modify the cysteines of plasma proteins and affect their structure and function. Human serum albumin (HSA) is prone to undergo modification by various low molecular weight compounds, including drugs. Cysteine34 (Cys34) in HSA has a free thiol group with antioxidant properties, considered to be the most redox-sensitive amino acid in plasma. Through mass-spectrometric analysis, we demonstrate for the first time that captopril forms a disulfide adduct at Cys34 residue and increases the protease susceptibility of HSA to trypsin. As evidenced by our biophysical and electron microscopy studies, HSA undergoes structural alteration, aggregation and morphological changes when treated with different captopril concentrations. Molecular dynamics studies further revealed the regions of secondary structural changes in HSA due to disulfide adduct formation by captopril at Cys34. It also elucidated the residues involved in the noncovalent interactions with captopril. It is envisaged that structural change in HSA may influence the efficacy of drug delivery as well as its own biological function. These findings may thus provide significant insights into the field of pharmacology intriguing further investigation into the effects of long-term captopril treatment.

Seasonal variations of leaf ecophysiological traits and strategies of co-occurring evergreen and deciduous trees in white oak forest in the central Himalaya.

The present study investigates the seasonal variations in leaf ecophysiological traits and strategies employed by co-occurring evergreen and deciduous tree species within a white oak forest (Quercus leucotrichophora A. Camus) ecosystem in the central Himalaya. Seasonal variations in physiological, morphological, and chemical traits were observed from leaf initiation until senescence in co-occurring deciduous and evergreen tree species. We compared various parameters, including net photosynthetic capacity (Aarea and Amass), leaf stomatal conductance (gswarea and gswmass), transpiration rate (Earea and Emass), specific leaf area (SLA), mid-day water potential (Ψmd), leaf nitrogen (N) and phosphorus (P) concentration, leaf total chlorophyll concentration, photosynthetic nitrogen- and phosphorus-use efficiency (PNUE and PPUE), and water use efficiency (WUE) across four evergreen and four deciduous tree species. Our findings reveal that evergreen and deciduous trees exhibit divergent strategies in coping with seasonal changes, which are crucial for their survival and growth. Deciduous trees consistently exhibited significantly higher photosynthetic rates, transpiration rates, mass-based N and P concentrations (Nmass and Pmass), mass-based chlorophyll concentration (Chlmass), SLA, and leaf Ψmd, while maintaining lower leaf structural investments throughout the year compared to evergreen trees. These findings indicate that deciduous trees achieve greater assimilation rates per unit mass and higher nutrient-use efficiency. Physiological, morphological, and leaf N and P concentrations were higher in the summer (fully expanded leaf) than in the fall (senesced leaf). These insights provide valuable contributions to our understanding of tree species coexistence and their ecological roles in temperate forest ecosystems, with implications for forest management and conservation in the Himalayan region.

Evaluating therapeutic potential of AYUSH-64 constituents against omicron variant of SARS-CoV-2 using ensemble docking and simulations.

The COVID-19 pandemic in the later phase showed the presence of the B.1.1.529 variant of the SARS-CoV-2 designated as Omicron. AYUSH-64 a poly herbal drug developed by Central Council for Research in Ayurvedic Sciences (CCRAS) has been recommended by Ministry of Ayush in asymptomatic, mild to moderate COVID-19 patients. One of the earlier, in-silico study has shown the binding of the constituents of AYUSH-64 to the main protease (Mpro) of the SARS-CoV-2. This study enlisted four phytochemicals of AYUSH-64, which were found to have significant binding with the Mpro. In continuation to the same, the current study proposes to understand the binding of these four phytochemicals to main protease (Mpro) and receptor binding domain (RBD) of spike protein of the Omicron variant. An enhanced molecular docking methodology, namely, ensemble docking has been used to find the most efficiently binding phytochemical. Using molecular dynamics (MD) simulations and clustering approach it was observed that the Mpro and RBD Spike of Omicron variant of SARS-CoV-2 in complex with human ACE2 tends to attain 4 and 8 conformational respectively. Based on the docking studies, the best binding phytochemical of the AYUSH-64, akummicine N-oxide was selected for MD simulations. MD simulations of akummicine N-oxide bound to omicron variant of Mpro and RBD Spike-ACE complex was performed. The conformational, interaction and binding energy analysis suggested that the akummicine N-oxide binds well with Mpro and RBD Spike-ACE2 complex. The interaction between RBD Spike and ACE2 was observed to weaken in the presence of akummicine N-oxide. Hence, it can be inferred that, these phytochemicals from AYUSH-64 formulation may have the potential to act against the Omicron variant of SARS-CoV-2.

In-silico evaluation of Oroxylum indicum vent compounds in the plausible treatment and prevention of nasopharyngeal cancer.

BACKGROUND: Shyonaka (Oroxylum indicum Vent) is widely used in Ayurveda and in ethnomedical practice for the treatment of inflammation, pain, diarrhea, non-healing ulcers, and cancer. Owing to the high prevalence of Epstein-Barr virus (EBV) infection in Nasopharyngeal carcinoma (NPC) patients, simultaneous targeting of proteins involved in both EBV replication and NPC proliferation might help to manage the disease effectively. OBJECTIVES: This study is designed to identify potential dual targeting inhibitors from Oroxylum indicum having the potential to inhibit both EBV and NPC. This study also attempted quantitative analysis of Shyonaka Bark Decoction (SBD) to confirm the presence of Baicalein and Chrysin which are predominant marker compounds of Shyonaka. METHODOLOGY: The HPLC analysis of stem bark and root bark of Oroxylum indicum was done to estimate the presence of marker compounds Baicalein and Chrysalin. The in-silico analysis included ADMET analysis followed by molecular docking of known compounds from Oroxylum indicum (retrieved from IMPPAT database) onto the target proteins of EBV (BHRF1, NEC1, dUTPase, Uracil DNA glycosylase) and NPC (COX-2, EGFR, and MDM2) using DOCK6 tool. Further validations were done using the molecular dynamics simulations of top screened molecules onto the selected target proteins using AMBER20 package and their corresponding MMGBSA binding free-energy values were calculated. RESULTS: The molecular docking revealed that the key molecules from the plant, scutellarein 7-rutinoside (S7R), scutellarin (SCU) and 6-hydroxyluteolin, Baicalein and 5,7-Dihydroxy-2-phenyl-6-[3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxychromen-4-one (57D) are effectively intervening with the target proteins of EBV, one of the key causative factors of NPC and the NPC specific targets which have the potential to reduce tumor size and other consequences of NPC. The molecular dynamics simulations of S7R, Baicalein and 57D, Baicalein with MDM-2 protein and dUTPase protein, respectively, showed stable interactions between them which were further assessed by the binding energy calculations. CONCLUSION: Overall, the in-silico evaluation of these phytochemicals with target proteins indicates their potential to inhibit both EBV and NPC which needs further in-vitro and in-vivo validations.

Antibiotic Susceptibility and Biofilm Production among Coagulase Negative Staphylococci Isolated from Clinical Samples at Tertiary Care Hospital.

BACKGROUND: Coagulase Negative Staphylococci have been widely associated with medical device implant treatment and immune-compromised patients. Despite having increasing interest in Coagulase Negative Staphylococci, few studies from Nepal have reported the association of these organisms with urinary tract infections, conjunctivitis, high vaginal swabs, and cerebrospinal fluid. This study was carried out to determine antibiotic susceptibility pattern and biofilm production among Coagulase Negative Staphylococci isolated from clinical samples at tertiary care hospital. METHODS: This study was a hospital based cross-sectional study in which 3690 clinical samples were included. Isolation and identification of isolates was done following standard microbiological protocol. Coagulase Negative Staphylococci were identified phenotypically on the basis of gram staining, slide and tube coagulase test and by various sugar fermentation tests. Antibiotic susceptibility test was done following Kirby Bauer disk diffusion method (Clinical and Laboratory Standards Institute 2020). Biofilm production was determined by Tissue Culture Plate technique. RESULTS: A total of 113 isolates of Coagulase Negative Staphylococci were detected. Among them S. epidermidis (45.1%), S. saprophyticus (23.9%), S. haemolyticus (16.8%), S. hominis (5.3%), S. capitis (2.7%), -----S. cohini (1.8%), S. lugdunensis (1.8%) and S. sciuri (2.7%) were identified phenotypically. All isolates were found to be resistant against Ampicillin and 111 (98.2%) were sensitive against Linezolid.23.9% of CoNS were strong biofilm producers, 19.5% moderate and 56.6 % were non/weak biofilm producers. CONCLUSIONS: It requires susceptibility test for prescribing antibiotics against Coagulase Negative Staphylococci in hospital and the misuse of antibiotics should be prevented.

Topography-mediated light environment regulates intra-specific seasonal and diurnal patterns of photosynthetic plasticity and plant ecophysiological adaptation strategies.

Due to substantial topographic variations in the Himalaya, incident solar radiation in the forest canopy is highly unequal. This results in significant environmental differences at finer scales and may lead to considerable differences in photosynthetic productivity in montane forests. Therefore, local-scale ecophysiological investigations, may be more effective and instructive than landscape-level inventories and models. We investigated leaf ecophysiological differences and related adaptations between two Quercus semecarpifolia forests in aspect-mediated, significantly varying light regimes in the same mountain catchment. Seasonal and diurnal rates of photosynthesis (A) were significantly higher in south aspect (S) than the north (N). Although temperature was a key contributor to seasonal fluctuations in photosynthetic physiology, photoperiod significantly determined intraspecific variations in seasonal and diurnal plasticity of leaf ecophysiological traits between the two topography-mediated light environments. The regression model for A as a function of stomatal conductivity (gsw) explained the critical role of gsw in triggering photosynthetic plasticity as an adaptive function against varying environmental stresses due to seasonal solar differences. We also examined, modifications in chlorophyll content between the two light regimes across seasons to determine the chlorophyll adaptation strategy. The N aspect had higher leaf chl a, b, and chl a + b and a lower chl-allocation ratio (a/b) than S, which helped to optimize the required light reception in the photoreaction centers for improved photosynthetic performance. The leaf light response curves for A and gsw were observed against varying incident photosynthetic photon flux densities (0-2000 mol.m2 s-1 PPFD) for both aspects. We found that the same species developed significantly distinct light response strategies and photosynthetic capacities in S than in N for the given magnitudes of PPFD. Such acquired ecophysiological adaptations owing to varying light environments may provide significant clues for understanding the impact of future climate change on Himalayan tree species.

Studying early structural changes in SOS1 mediated KRAS activation mechanism.

KRAS activation is known to be modulated by a guanine nucleotide exchange factor (GEF), namely, Son of Sevenless1 (SOS1). SOS1 facilitates the exchange of GDP to GTP thereby leading to activation of KRAS. The binding of GDP/GTP to KRAS at the REM/allosteric site of SOS1 regulates the activation of KRAS at CDC25/catalytic site by facilitating its exchange. Different aspects of the allosteric activation of KRAS through SOS1 are still being explored. To understand the SOS1 mediated activation of KRAS, molecular dynamics simulations for a total of nine SOS1 complexes (KRAS-SOS1-KRAS) were performed. These nine systems comprised different combinations of KRAS-bound nucleotides (GTP/GDP) at REM and CDC25 sites of SOS1. Various conformational and thermodynamic parameters were analyzed for these simulation systems. MMPBSA free energy analysis revealed that binding at CDC25 site of SOS1 was significantly low for GDP-bound KRAS as compared to that of GTP-bound KRAS. It was observed that presence of either GDP/GTP bound KRAS at the REM site of SOS1 affected the activation related changes in the KRAS present at CDC25 site. The conformational changes at the catalytic site of SOS1 resulting from GDP/GTP-bound KRAS at the allosteric changes may hint at KRAS activation through different pathways (slow/fast/rare). The allosteric effect on activation of KRAS at CDC25 site may be due to conformations adopted by switch-I, switch-II, beta2 regions of KRAS at REM site. The effect of structural rearrangements occurring at allosteric KRAS may have led to increased interactions between SOS1 and KRAS at both the sites. The SOS1 residues involved in these important interactions with KRAS at the REM site were R694, S732 and K735. Whereas the ones interacting with KRAS at CDC25 site were S807, W809 and K814. This may suggest the crucial role of these residues in guiding the allosteric activation of KRAS at CDC25 site. The conformational shifts observed in the switch-I, switch-II and alpha3 regions of KRAS at CDC25 site may be attributed to be a part of allosteric activation. The binding affinities, interacting residues and conformational dynamics may provide an insight into development of inhibitors targeting the SOS1 mediated KRAS activation.

Leaf and tree age-related changes in leaf ecophysiological traits, nutrient, and adaptive strategies of Alnus nepalensis in the central Himalaya.

Leaf ecophysiological traits are known to change with leaf and tree age. In the present study, we measured the effect of leaf and tree age on leaf ecophysiological and morphological traits of nitrogen-fixing Alnus nepalensis (D. Don) which is a pioneer tree species in degraded lands. Three naturally occurring A. nepalensis forest stands, namely young (5-8 years old), mature (40-55 years old), and old (130-145 years old), were considered in this study. We also investigated the seasonal variations in leaf ecophysiological and morphological traits during leaf flushing, fully expanded, and leaf senescence phenological stages. The ecophysiological and morphological traits were compared between leaf and tree ages using a linear mixed-effect model (LMM) and Tukey's HSD test. Fully expanded leaves and young trees demonstrate ecophysiological traits consistent with acquisitive resource-use strategies. Our results revealed that net photosynthetic capacity (Aarea and Amass), leaf stomatal conductance (gswarea and gswmass), transpiration rate (Earea and Emass), specific leaf area (SLA), predawn and midday water potential (Ψ), leaf total chlorophyll concentration, photosynthetic N- and P-use efficiency (PNUE and PPUE) were higher in younger trees than mature and old trees. We found lower wateruse efficiency (WUE) and intrinsic water-use efficiency (WUEi) in young trees than in mature and old ones. Mass-based net photosynthetic capacity (Amass) was positively correlated with PNUE, PPUE, transpiration rate, stomatal conductance, SLA and chlorophyll concentrations but negatively correlated with WUE and WUEi. However, mass-based leaf nitrogen (N) and phosphorus (P) concentrations were the highest in fully expanded leaves and did not vary with tree age despite N concentration being negatively correlated with SLA. Overall, this study provides valuable insights into the age-related changes in leaf ecophysiological traits of A. nepalensis. The findings underscore the importance of considering tree age when studying plant ecophysiology and highlight the acquisitive resource-use strategies employed by young trees for rapid growth and establishment.

MolToxPred: small molecule toxicity prediction using machine learning approach.

Different types of chemicals and products may exhibit various health risks when administered into the human body. For toxicity reasons, the number of new drugs entering the market through the conventional drug development process has been reduced over the years. However, with the advent of big data and artificial intelligence, machine learning techniques have emerged as a potential solution for predicting toxicity and ensuring efficient drug development and chemical safety. An ML model for toxicity prediction can reduce experimental costs and time while addressing ethical concerns by drastically reducing the need for animals and clinical trials. Herein, MolToxPred, an ML-based tool, has been developed using a stacked model approach to predict the potential toxicity of small molecules and metabolites. The stacked model consists of random forest, multi-layer perceptron, and LightGBM as base classifiers and Logistic Regression as the meta classifier. For training and validation purposes, a comprehensive set of toxic and non-toxic molecules is curated. Different structural and physicochemical-based features in the form of molecular descriptors and fingerprints were employed. MolToxPred utilizes a comprehensive feature selection process and optimizes its hyperparameters through Bayesian optimization with stratified 5-fold cross-validation. In the evaluation phase, MolToxPred achieved an AUROC of 87.76% on the test set and 88.84% on an external validation set. The McNemar test was used as the post-hoc test to determine if the stacked models' performance was significantly different compared to the base learners. The developed stacked model outperformed its base classifiers and an existing tool in the literature, reaffirming its better performance. The hypothesis is that the incorporation of a diverse set of data, the subsequent feature selection, and a stacked ensemble approach give MolToxPred the edge over other methods. In addition to this, an attempt has been made to identify structural alerts responsible for endpoints of the Tox21 data to determine the association of a molecule with a plausible downstream pathway of action. MolToxPred may be helpful for drug discovery and regulatory pipelines in pharmaceutical and other industries for in silico toxicity prediction of small molecule candidates.

Recurrence of pulmonary tuberculosis in India: Findings from the 2019-2021 nationwide community-based TB prevalence survey.

Recurrent Tuberculosis patients contribute to a significant proportion of TB burden in India. A nationwide survey was conducted during 2019-2021 across India among adults to estimate the prevalence of TB. A total of 322480 individuals were screened and 1402 were having TB. Of this, 381 (27.1%) had recurrent TB. The crude prevalence (95% CI) of recurrent TB was 118 (107-131) per 100,000 population. The median duration between episodes of TB was 24 months. The proportion of drug resistant TB was 11.3% and 3.6% in the recurrent group and new TB patients respectively. Higher prevalence of recurrent TB was observed in elderly, males, malnourished, known diabetics, smokers, and alcohol users. (p<0.001). To prevent TB recurrence, all treated tuberculosis patients must be followed at least for 24 months, with screening for Chest X-ray, liquid culture every 6 months, smoking cessation, alcohol cessation, nutritional interventions and good diabetic management.

Prevalence and factors associated with tuberculosis infection in India.

BACKGROUND: The risk of tuberculosis (TB) disease is higher in individuals with TB infection. In a TB endemic country like India, it is essential to understand the current burden of TB infection at the population level. The objective of the present analysis is to estimate the prevalence of TB infection in India and to explore the factors associated with TB infection. METHODS: Individuals aged > 15 years in the recently completed National TB prevalence survey in India who were tested for TB infection by QuantiFERON-TB Gold Plus (QFT-Plus) assay were considered for this sub-analysis. TB infection was defined as positive by QFT-Plus (value >0.35 IU/ml). The estimates for prevalence, prevalence ratio (PR) and adjusted risk ratio (aRR) estimates with 95% confidence intervals (CIs) were calculated. RESULTS: Of the 16864 individuals analysed, the prevalence of TB infection was 22.6% (95% CI:19.4 -25.8). Factors more likely to be associated with TB infection include age > 30 years (aRR:1.49;95% CI:1.29-1.73), being male (aRR:1.26; 95%CI: 1.18-1.34), residing in urban location (aRR:1.58; 95%CI: 1.03-2.43) and past history of TB (aRR:1.49; 95%CI: 1.26-1.76). CONCLUSION: About one fourth (22.6%) of the individuals were infected with TB in India. Individuals aged > 30 years, males, residing in urban location, and those with past history of TB were more likely to have TB infection. Targeted interventions for prevention of TB and close monitoring are essential to reduce the burden of TB in India.

Probing the structure of human tRNA3Lys in the presence of ligands using docking, MD simulations and MSM analysis.

The tRNA3Lys, which acts as a primer for human immunodeficiency virus type 1 (HIV-1) reverse transcription, undergoes structural changes required for the formation of a primer-template complex. Small molecules have been targeted against tRNA3Lys to inhibit the primer-template complex formation. The present study aims to understand the kinetics of the conformational landscape spanned by tRNA3Lys in apo form using molecular dynamics simulations and Markov state modeling. The study is taken further to investigate the effect of small molecules like 1,4T and 1,5T on structural conformations and kinetics of tRNA3Lys, and comparative analysis is presented. Markov state modeling of tRNA3Lys apo resulted in three metastable states where the conformations have shown the non-canonical structures of the anticodon loop. Based on analyses of ligand-tRNA3Lys interactions, crucial ion and water mediated H-bonds and free energy calculations, it was observed that the 1,4-triazole more strongly binds to the tRNA3Lys compared to 1,5-triazole. However, the MSM analysis suggest that the 1,5-triazole binding to tRNA3Lys has brought rigidity not only in the binding pocket (TΨC arm, D-TΨC loop) but also in the whole structure of tRNA3Lys. This may affect the easy opening of primer tRNA3Lys required for HIV-1 reverse transcription.

Novel alkyl-substituted 4-methoxy benzaldehyde thiosemicarbazones: Multi-target directed ligands for the treatment of Alzheimer's disease.

Alzheimer's disease (AD) is a devastating neurodegenerative disorder affecting mental ability and interrupts neurocognitive functions. Treating multifactorial conditions of AD with a single-target-directed drug is highly difficult. Thus, a multi-target-directed ligand (MTDL) development strategy has been developed as a promising approach for the treatment of AD. Herein, we have synthesized two novel thiosemicarbazones as MTDLs and reported their bioactivities against diverse neuropathological events involved in AD. In vitro studies revealed that both compounds exhibited promising anticholinesterase activity (AChE, IC50 = 15.98 µM, MZET and IC50 = 30.23 µM, MZMT), well supported by a detailed computational study. Both analogs have shown good thermodynamic behaviour and stability through interactions with characteristic amino acid residues throughout simulation of 100 ns against acetylcholinesterase enzyme. In an electrophysiology assay, these analogs have shown a characteristic inhibitory response against the GluN1-1a + GluN2B subunit of N-methyl-D-aspartate receptors. Pre-treatment of BV-2 microglial cells with MZET effectively decreased nitrite production compared to nitrite produced by lipopolysaccharide-treated cells alone. Further, the effect of MZMT and MZET on autophagy regulation was determined using stably transfected SH-SY5Y neuroblastoma cells. MZET significantly enhanced the autophagy flux in neuroblastoma cells. A significant decrease in copper-catalysed oxidation of amyloid-ß in presence of synthesized thiosemicarbazones was also observed. Collectively, our findings indicated that these analogs have potential as effective anti-AD candidates and can be used as a prototype to develop more safer multi-targeted anti-AD drugs.

Experience of "One Stop TB Diagnostic Solution" Model in Engaging a Private Laboratory for End-to-End Diagnostic Services in the National TB Elimination Program in Hisar, India.

The complete diagnostic evaluation of tuberculosis based on its drug-resistance profile is critical for appropriate treatment decisions. The TB diagnostic landscape in India has been transformed with the scaling-up of WHO-recommended diagnostics, but challenges remain with specimen transportation, completing diagnostic assessment, turnaround time (TAT), and maintaining laboratories. Private laboratories have demonstrated efficiencies for specimen collection, transportation, and the timely testing and issue of results. A one-stop TB diagnostic model was designed to assess the feasibility of providing end-to-end diagnostic services in the Hisar district of Haryana state, India. A NTEP-certified private laboratory was engaged to provide the services, complementing the existing public sector diagnostic services. A total of 10,164 specimens were collected between May 2022 and January 2023 and these were followed for the complete diagnostic assessment of Drug-Susceptible TB (DS-TB) and Drug-Resistant TB (DR-TB) and the time taken for issuing results. A total of 2152 (21%) patients were detected with TB, 1996 (93%) Rifampicin-Sensitive and 134 (6%) with Rifampicin-Resistant TB. Nearly 99% of the patients completed the evaluation of DS-TB and DR-TB within the recommended TAT. The One-Stop TB/DR-TB Diagnostic Solution model has demonstrated that diagnostic efficiencies could be enhanced through the strategic purchase of private laboratory services.

Community health workers to reduce unmet surgical needs in an urban slum in India: an implementation study.

BACKGROUND: The Surgical Accredited & Trained Healthcare Initiative (SATHI) project demonstrates how community healthcare workers (CHWs) with merely 8 y of formal schooling and training for a short period can reduce unmet surgical needs. METHODS: A pilot study was carried out in the slums of a metropolitan city in India to know the effectiveness of a SATHI in reducing the burden of unmet surgical needs. In total, 12 730 people from 3000 households were included in the study for a duration of 6 months. RESULTS: We found 10% surgical needs (n=293) out of which 57% had unmet surgical needs. Out of total surgical needs, about half of the needs were cataract and abdominal, followed by extremities and chest conditions. SATHIs were able to convert 99 patients (60%) from unmet to met needs, who underwent surgery/treatment. The conversion from unmet to met among all surgery needs was highest for abdominal conditions (29%) followed by cataracts (17%). CONCLUSIONS: SATHIs with short training can reduce the burden of unmet surgical needs. SATHIs were able to convert a significant proportion of unmet to met needs by trust building, facilitating access to healthcare and ensuring post-operative adherence. Scaling up could help in the achievement of equitable healthcare across India.

Nutritional supplementation to prevent tuberculosis incidence in household contacts of patients with pulmonary tuberculosis in India (RATIONS): a field-based, open-label, cluster-randomised, controlled trial.

BACKGROUND: In India, tuberculosis and undernutrition are syndemics with a high burden of tuberculosis coexisting with a high burden of undernutrition in patients and in the population. The aim of this study was to determine the effect of nutritional supplementation on tuberculosis incidence in household contacts of adults with microbiologically confirmed pulmonary tuberculosis. METHODS: In this field-based, open-label, cluster-randomised controlled trial, we enrolled household contacts of 2800 patients with microbiologically confirmed pulmonary tuberculosis across 28 tuberculosis units of the National Tuberculosis Elimination Programme in four districts of Jharkhand, India. The tuberculosis units were randomly allocated 1:1 by block randomisation to the control group or the intervention group, by a statistician using computer-generated random numbers. Although microbiologically confirmed pulmonary tuberculosis patients in both groups received food rations (1200 kcal, 52 grams of protein per day with micronutrients) for 6 months, only household contacts in the intervention group received monthly food rations and micronutrients (750 kcal, 23 grams of protein per day with micronutrients). After screening all household contacts for co-prevalent tuberculosis at baseline, all participants were followed up actively until July 31, 2022, for the primary outcome of incident tuberculosis (all forms). The ascertainment of the outcome was by independent medical staff in health services. We used Cox proportional hazards model and Poisson regression via the generalised estimating equation approach to estimate unadjusted hazard ratios, adjusted hazard ratios (aHRs), and incidence rate ratios (IRRs). This study is registered with CTRI-India, CTRI/2019/08/020490. FINDINGS: Between Aug 16, 2019, and Jan 31, 2021, there were 10 345 household contacts, of whom 5328 (94·8%) of 5621 household contacts in the intervention group and 4283 (90·7%) of 4724 household contacts in the control group completed the primary outcome assessment. Almost two-thirds of the population belonged to Indigenous communities (eg, Santhals, Ho, Munda, Oraon, and Bhumij) and 34% (3543 of 10 345) had undernutrition. We detected 31 (0·3%) of 10 345 household contact patients with co-prevalent tuberculosis disease in both groups at baseline and 218 (2·1%) people were diagnosed with incident tuberculosis (all forms) over 21 869 person-years of follow-up, with 122 of 218 incident cases in the control group (2·6% [122 of 4712 contacts at risk], 95% CI 2·2-3·1; incidence rate 1·27 per 100 person-years) and 96 incident cases in the intervention group (1·7% [96 of 5602], 1·4-2·1; 0·78 per 100 person-years), of whom 152 (69·7%) of 218 were patients with microbiologically confirmed pulmonary tuberculosis. Tuberculosis incidence (all forms) in the intervention group had an adjusted IRR of 0·61 (95% CI 0·43-0·85; aHR 0·59 [0·42-0·83]), with an even greater decline in incidence of microbiologically confirmed pulmonary tuberculosis (0·52 [0·35-0·79]; 0·51 [0·34-0·78]). This translates into a relative reduction of tuberculosis incidence of 39% (all forms) to 48% (microbiologically confirmed pulmonary tuberculosis) in the intervention group. An estimated 30 households (111 household contacts) would need to be provided nutritional supplementation to prevent one incident tuberculosis. INTERPRETATION: To our knowledge, this is the first randomised trial looking at the effect of nutritional support on tuberculosis incidence in household contacts, whereby the nutritional intervention was associated with substantial (39-48%) reduction in tuberculosis incidence in the household during 2 years of follow-up. This biosocial intervention can accelerate reduction in tuberculosis incidence in countries or communities with a tuberculosis and undernutrition syndemic. FUNDING: Indian Council of Medical Research-India TB Research Consortium.

Nutritional support for adult patients with microbiologically confirmed pulmonary tuberculosis: outcomes in a programmatic cohort nested within the RATIONS trial in Jharkhand, India.

BACKGROUND: Undernutrition is a common comorbidity of tuberculosis in countries with a high tuberculosis burden, such as India. RATIONS is a field-based, cluster-randomised controlled trial evaluating the effect of providing nutritional support to household contacts of adult patients with microbiologically confirmed pulmonary tuberculosis in Jharkhand, India, on tuberculosis incidence. The patient cohort in both groups of the trial was provided with nutritional support. In this study, we assessed the effects of nutritional support on tuberculosis mortality, treatment success, and other outcomes in the RATIONS patient cohort. METHODS: We enrolled patients (aged 18 years or older) with microbiologically confirmed pulmonary tuberculosis across 28 tuberculosis units. Patients received nutritional support in the form of food rations (1200 kcal and 52 g of protein per day) and micronutrient pills. Nutritional support was for 6 months for drug-susceptible tuberculosis and 12 months for multidrug-resistant tuberculosis; patients with drug-susceptible tuberculosis could receive an extension of up to 6 months if their BMI was less than 18·5 kg/m2 at the end of treatment. We recorded BMI, diabetes status, and modified Eastern Cooperative Oncology Group (ECOG) performance status at baseline. Clinical outcomes (treatment success, tuberculosis mortality, loss to follow-up, and change in performance status) and weight gain were recorded at 6 months. We assessed the predictors of tuberculosis mortality with Poisson and Cox regression using adjusted incidence rate ratios (IRRs) and adjusted hazard ratios (HRs). The RATIONS trial is registered with the Clinical Trials Registry of India (CTRI/2019/08/020490). FINDINGS: Between Aug 16, 2019, and Jan 31, 2021, 2800 patients (mean age 41·5 years [SD 14·5]; 1979 [70·7%] men and 821 [29·3%] women) were enrolled. At enrolment, 2291 (82·4%) patients were underweight (BMI <18·5 kg/m2), and 480 (17·3%) had a BMI of less than 14 kg/m2. The mean weight and BMI were 42·6 kg (SD 7·8) and 16·4 kg/m2 (2·6) in men and 36·1 kg (7·3) and 16·2 kg/m2 (2·9) in women. During the 6-month follow-up, treatment was successful in 2623 (93·7%) patients, 108 (3·9%) tuberculosis deaths occurred, 28 (1·0%) patients were lost to follow-up, and treatment failure was experienced by five (0·2%) patients. The median weight gain was 4·6 kg (IQR 2·8-6·8), but 1441 (54·8%) of 2630 patients remained underweight. At 2 months, 1444 (54·0%) of 2676 patients gained at least 5% of baseline weight. Baseline weight (adjusted IRR 0·95, 95% CI 0·90-0·99), BMI (0·88, 0·76-1·01), poor performance status (ECOG categories 3-4; 5·33, 2·90-9·79), diabetes (3·30, 1·65-6·72), and haemoglobin (0·85, 0·71-1·00) were predictors of tuberculosis mortality. A reduced hazard of death (adjusted HR 0·39, 95% CI 0·18-0·86) was associated with a 5% weight gain at 2 months. INTERPRETATION: In this study, nutritional support was provided to a cohort with a high prevalence of severe undernutrition. Weight gain, particularly in the first 2 months, was associated with a substantially decreased hazard of tuberculosis mortality. Nutritional support needs to be an integral component of patient-centred care to improve treatment outcomes in such settings. FUNDING: India Tuberculosis Research Consortium, Indian Council of Medical Research.

Design of LNA Analogues Using a Combined Density Functional Theory and Molecular Dynamics Approach for RNA Therapeutics.

Antisense therapeutics treat a wide spectrum of diseases, many of which cannot be addressed with the current drug technologies. In the quest to design better antisense oligonucleotide drugs, we propose five novel LNA analogues (A1-A5) for modifying antisense oligonucleotides and establishing each with the five standard nucleic acids: adenine (A), guanine (G), cytosine (C), thymine (T), and uracil (U). Monomer nucleotides of these modifications were considered for a detailed Density Functional Theory (DFT)-based quantum chemical analysis to determine their molecular-level structural and electronic properties. A detailed MD simulation study was done on a 14-mer ASO (5'-CTTAGCACTGGCCT-3') containing these modifications targeting PTEN mRNA. Results from both molecular- and oligomer-level analysis clearly depicted LNA-level stability of the modifications, the ASO/RNA duplexes maintaining stable Watson-Crick base pairing preferring RNA-mimicking A-form duplexes. Notably, monomer MO isosurfaces for both purines and pyrimidines were majorly distributed on the nucleobase region in modifications A1 and A2 and in the bridging unit in modifications A3, A4, and A5, suggesting that A3/RNA, A4/RNA, and A5/RNA duplexes interact more with the RNase H and solvent environment. Accordingly, solvation of A3/RNA, A4/RNA, and A5/RNA duplexes was higher compared to that of LNA/RNA, A1/RNA, and A2/RNA duplexes. This study has resulted in a successful archetype for creating advantageous nucleic acid modifications tailored for particular needs, fulfilling a useful purpose of designing novel antisense modifications, which may overcome the drawbacks and improve the pharmacokinetics of existing LNA antisense modifications.

Estimating the Burden of Tuberculosis in India: A Modelling Study.

Background: Measurements TB incidence and mortality are crucial for monitoring progress towards SDG goals for TB. Until recently, WHO estimated TB burden in India with applied simple, transparent equilibrium models to data from Gujarat, an Indian state where the first state-level prevalence survey was conducted in 2011. However, since then there has been several interventions in India including national TB prevalence survey, infection survey, sub-national survey & certification which gives opportunity for national and sub-national estimates for TB incidence and mortality. Methods: We developed a model is a compartmental, deterministic framework, taking account of TB natural history, as well as India's healthcare system including health care seeking from public and private sector. To address changes in TB burden owing to COVID disruptions, we followed same model that used by WHO in the global TB Report 2022 with additional impact of delta wave in 2021. Major sources of data included National TB Prevalence survey, trends in caseloads in public and private sector including their contribution and mortality information. Results: We estimated total TB incidence of 2.77 million in the year 2022 as against 2.97 in the year 2015 and corresponding TB mortality of 0.32 and 0.36 million respectively. In terms of rate per 1,00,000 TB incidence in 2022 was 196 as compared to 225 in the year 2015 and mortality was 23 and 27 respectively. TB incidence estimates are similar to what was estimated by WHO, while mortality estimates appear different in our estimates due to different calibration targets depending on in-country published data. Conclusion: Even if TB burden is infeasible to measure directly, a range of data can nonetheless offer indirect evidence for its estimation: mathematical modelling can be a helpful tool for bringing together these diverse sources of evidence, and deriving estimates that are consistent with them all. While the RGI reported mortality is an important source of information, its quality and coverage for medically certified cause of deaths requires improvement in India.

The potential impact of vaccination on tuberculosis burden in India: A modelling analysis.

Background & objectives: Vaccination will play an important role in meeting the end tuberculosis (TB) goals. While certain vaccine candidates in advanced stages of clinical trials raise hope for the future availability of new tools, in the immediate term, there is also increasing interest in Bacille Calmette-Guérin revaccination among adults and adolescents as a potential strategy. Here, we sought to estimate the potential epidemiological impact of TB vaccination in India. Methods: We developed a deterministic, age-structured, compartmental model of TB in India. Data from the recent national prevalence survey was used to inform epidemiological burden while also incorporating a vulnerable population who may be prioritized for vaccination, the latter consistent with the burden of undernutrition. Using this framework, the potential impact on incidence and mortality of a vaccine with 50 per cent efficacy was estimated, if rolled out in 2023 to cover 50 per cent of the unvaccinated each year. Simulated impacts were compared for disease- vs. infection-preventing vaccines, as well as when prioritizing vulnerable groups (those with undernutrition) rather than the general population. A sensitivity analyses were also conducted with respect to the duration, and efficacy, of vaccine immunity. Results: When rolled out in the general population, an infection-preventing vaccine would avert 12 per cent (95% Bayesian credible intervals (Crl): 4.3-28%) of cumulative TB incidence between 2023 and 2030, while a disease-preventing vaccine would avert 29 per cent (95% Crl: 24-34%). Although the vulnerable population accounts for only around 16 per cent of India's population, prioritizing this group for vaccination would achieve almost half the impact of rollout in the general population, in the example of an infection-preventing vaccine. Sensitivity analysis also highlights the importance of the duration and efficacy of vaccine-induced immunity. Interpretation & conclusions: These results highlight how even a vaccine with moderate effectiveness (50%) could achieve substantial reductions in TB burden in India, especially when prioritized for the most vulnerable.