ABOUT THE PROJECT

Tuberculosis (TB) persists as a significant global health concern, affecting an estimated 10 million individuals worldwide in 2023. Delving into the intricate molecular mechanisms employed by Mycobacterium tuberculosis (Mtb) to adapt within the host environment, particularly those governed post-transcriptionally by small noncoding RNA (sRNA), holds promise for developing more efficacious treatment modalities. While the regulatory role of sRNAs in the pathogenesis of enteric bacteria is well-established, their functions within Mtb remain largely unexplored.
Building upon our prior investigations into Mtb's transcriptomic responses during effective treatment, our findings pointed towards a potential significance of MTS2823 in maintaining Mtb's infectious capability. This observation has spurred our current hypothesis that MTS2823 plays a pivotal role in TB infection, with alterations in its expression pattern potentially impacting Mtb's ability to invade new host cells. To put this hypothesis to the test, we propose employing a targeted repression strategy utilizing the CRISPR-dCAS9 technique to elucidate the importance of MTS2823 in Mtb growth and infection in vitro.
By undertaking this study, we aim to shed light on the biological role of MTS2823 in Mtb pathogenesis and establish a toolkit for investigating other regulatory sRNAs within mycobacteria.

SPECIFIC OBJECTIVES

• Optimize the CRISPRi system for precise repression of sRNA in Mycobacterium tuberculosis.
• Determine the significance of sRNA MTS2823 for Mtb survival, growth, and in vitro infection within macrophages.

ABOUT THE PROJECT

In 2020, a study of vitamin D in DRTB cases and contacts was conducted. Estimation of Vitamin D levels was undertaken among cases and household and non-household controls. Active TB / LTBI was ruled out among contacts by symptom screening, chest X-ray and Interferon-Gamma Release Assays (IGRA). Individuals who had low Vitamin D levels were administered oral Vitamin D. No preventive treatment was given to either of the contacts.
Among the 180 HH contacts and 82 NHH contacts enrolled in the study, 65 and 26 respectively were detected to be IGRA positive.
This study was designed to assess the progress of the contacts during this period.

Study Objectives

• To assess development of active tuberculosis among the contacts.
• To determine the role IGRA in predicting tuberculosis among contacts.
• To assess the effect of Vitamin D on development of tuberculosis among contacts.

Methodology

HH and non HH contacts were contacted, written consent was taken and consenting individuals were interviewed using a questionnaire to obtain information regarding TB symptoms, tests and treatment taken during the period. Symptomatic individuals at the time of interview were assessed for active TB. Co-morbidities such as diabetes mellitus, hypertension, history of Covid-19 and HIV were recorded and response to treatment recorded.

KEY FINDINGS/ACHIEVEMENTS

• Among the 262 contacts (180 household and 82 non-household), 34.73% had latent TB.
• Of 219 contacts which were interviewed, three (2 household and 1 non-household) developed active TB, with a crude incidence rate of 4.64 per 1000 people.
• Baseline Vitamin D deficiency was prevalent in 75.3% of contacts (including the three active TB cases).
• Vitamin D supplementation showed a non-significant trend in reducing TB risk (OR = 0.56), p = 0.492).
• IGRA status did not significantly predict TB development.

PUBLICATION

• Dholakia, Y., Govekar, L., & Mistry, N. (2024). Long-term follow-up of contacts of drug-resistant tuberculosis cases in high-burden areas of Mumbai, India. Indian J Tuberc. 2024;71 Suppl 1: S86-S90.

ABOUT THE PROJECT

The National Strategic Plan (NSP) for Tuberculosis (TB) Elimination in India 2017-25 included Private Provider Engagement (PPE) as a key strategy for TB elimination. In the last 5 years various TB management interventions have been undertaken by the National TB Elimination Program, Government of India. The present study aims to evaluate the impact of PPSA Intervention under Private Sector Engagement for TB control in the country.

The study is being conducted in five districts which are representative of different levels of implementation of PPE Strategy. The study districts have been selected in consultation between BMGF, FMR, and CTD. A sample size of 250 TB patients per site i.e., a total of 1250 patients would be enrolled and followed up for duration of six months under the study. The study methodology is prospective cohort study and is observational in design. Inclusion criteria for the study include Drug Sensitive - Pulmonary TB patients, new or retreated between 18-70 years of age. Drug Resistant TB Patients and Extra Pulmonary TB patients will be excluded from the study.

The primary objective of study is to determine the level of exposure to PPE strategy of an averagely treated patient in selected five districts in India. It aims to measure the patient level outcomes in terms of enhanced access, correct diagnosis, shorter care pathways, patient retention and adherence as well as relief from excessive costs. It also aims to evaluate the strength of association between different components of the private engagement strategy and successful patient outcomes.

Scoping visits undertaken

Scoping visits were undertaken as part of the preparatory stage in the Madhubani and Pune sites. Madhubani has an active GOI patient-provider support agency (PPSA, since February 2022), which transitioned from the joint effort for elimination of TB (JEET) PPSA. Shortcomings observed with the program included a weak linkage of PPSA with private labs/chemists and lack of capacity-building workshops being organized for the private providers. Scoping visit also highlighted the need for improvement in service delivery from the program's side, as reflected by the district's scarcity of Cartridge based Nucleic Acid Amplification Tests (CB-NAAT) diagnostic cartridges and anti-TB drugs. Scoping in the Pune Municipal Corporation (PMC), a site not having an active PPSA currently, showed considerable efforts from the district TB team to engage and serve private sector patients. GOI PPSA is expected to be functional soon in PMC area.

ABOUT THE PROJECT

Whole genome sequencing (WGS) of Mycobacterium tuberculosis (MTB) has been demonstrated to reliably replace current methods, which take months to generate a drug resistance profile. Treatment of patients becomes extensively delayed leading to clinical deterioration and death. Direct-from-sputum WGS of MTB DNA can significantly reduce the turn-around-time (TAT) of universal DST from months to days and enable its wider use. This project aims to develop a low-cost method for the enrichment of MTB DNA directly from sputum for WGS as also the determination of the drug susceptibility profile.

ABOUT THE PROJECT

Bedaquiline (BDQ) is a relatively new drug introduced in the treatment of MDR TB, and the presence of clinical TB isolates having high MIC of BDQ in the CRyPTIC repository (BDQ naïve cases) is concerning. In this pilot project, we plan to revive selected clinical TB isolates (drug-susceptible, MDR and XDR) with high MIC of BDQ from the CRyPTIC repository. We will then study the growth kinetics of the revived clinical isolates using Mycobacterial Growth Indicator Tubes (MGIT). A comparison of growth of these isolates (between various drug susceptibility categories) will be performed to understand strain fitness in isolates with high MIC of BDQ.

KEY ACHIEVEMENTS

• MDR -TB isolates irrespective of their BDQ susceptibility showed a marginally reduced lag phase (~61 h) in comparison to drug susceptible (DS) (~70 h) and pre-extensively drug resistant (pre-XDR) isolates (~68 h)
• The DS TB isolates irrespective of BDQ susceptibility showed higher stochasticity in the doubling time (31-124 h) as compared to MDR (51-99 h) and pre-XDR (45-93 h) isolates
• The average revival time for BDQ monoresistant TB isolates was lower (~142 h) than the drug susceptible isolates (~272 h). Also, the average revival time for pre-XDR isolates with BDQ resistance was lower (~132 h) than BDQ susceptible pre-XDR isolates (~320 h)
• Reduced revival time of DS and pre-XDR isolates with BDQ resistance indicate potential fitness benefit of BDQ resistance in clinical TB isolates

ABOUT THE PROJECT

The study over three phases evaluated the anti-tubercular activity of a proprietary industrial compound for potential use in TB and MDR-TB treatment. The study in the initial phase established the anti-tubercular activity on drug sensitive and drug resistant strains of TB by rezasurin microtitre assay and agar proportion method. Through this, the minimum inhibitory concentration was defined. Subsequently the study evaluated the kill kinetics and synergism with standard anti-TB drugs for its potential use as a combination therapy with standard anti-TB drugs. The possible mechanism of action was studied by scanning electron microscope. In addition, the study also established the compound’s potential to target and kill intracellular TB bacteria through a macrophage cell line infection model.

OUTCOMES

A patent has been filed as co-inventor along with the study’s sponsor.

ABOUT THE PROJECT

The long duration and high cost of multi-drug resistant TB (MDR-TB) treatment motivate efforts to develop cost-effective preventive and adjunctive therapies including, Vit D supplementation. Vit D confers multi-modal protective approaches to human hosts and encourages better clinical responses.

This project was undertaken to expand the evidence base for the use of Vit D supplementation in the course of treatment of drug resistant TB as well as prevention of disease progression in household contacts exposed to TB infection. This study involved measurement of Vit D levels in both randomly selected DR TB patients within two months of starting treatment and two of their household contacts. The latter was tested to ascertain whether they are latently infected with TB. Additionally, they were evaluated for their food intake and frequency with a focus on intake of micro and macronutrients. The case-control study also assessed TB infection levels amongst household contacts of DR TB patients. This project was designed to provide guidance to support a larger intervention trial of Vit D supplementation for prevention and adjunct treatment of DR TB disease.

KEY FINDINGS/ACHIEVEMENTS

• Low levels of Vit D were identified as an independent risk factor for active MDR-TB compared to household and non-household controls.
• There was no association between Vit D status and IGRA positivity, an indicator of sub-clinical TB infection.

PUBLICATIONS

• Shukla, A., Bromage, S., Dholakia, Y…Mistry, N. et al. (2022). Case-control study of vitamin D status and adult multidrug-resistant pulmonary tuberculosis in Mumbai, India. Int J Tuberc Lung Dis, 26:826-834.
• Mistry, N.F., Hemler, E.C., Dholakia, Y., Shukla, A., Dev, P., Govekar, L…. Fawzi, W. et al. (2020). Protocol for a case-control study of vitamin D status, adult multidrug-resistant tuberculosis disease and tuberculosis infection in Mumbai, India. BMJ Open,10: 1-8.

ABOUT THE PROJECT

As a part of developing contemporary molecular epidemiology capacity, FMR invested early on in developing the whole genome sequencing (WGS) techniques for understanding TB disease transmission and drug resistance. The work led to the publication of one of the first reports on the application for tracking transmission and diagnosis of drug resistance in India. This led to the invitation by the University of Oxford to participate in their multi-country global project CRyPTIC. The aim of the CRyPTIC project was to achieve sufficiently accurate genetic prediction of resistance to 14 anti-tuberculosis drugs from whole genome sequencing to replace slow, cumbersome, culture-based drug susceptibility testing (DST) for Mycobacterium tuberculosis complex (MTBC). This would enable rapid-turnaround near-to-patient assays to revolutionize drug-resistant TB identification and management. In the first phase of the project (CRyPTIC-BMGF), 14-drug microtiter DST was validated for 1,000 isolates from India (FMR and Hinduja Hospital). The second phase of the CRyPTIC project (CRyPTIC-WT) was in continuation of the CRyPTIC-BMGF, which used large-scale global and clade-representative genome sequences (>15,000 isolates sequenced from 27 countries during the project) to build-up on the drug resistance catalogue. India (FMR and Hinduja Hospital) contributed to 6,500 isolates in this project. The project identified genomic variants with precision for improving statistical methods to detect associations between variants and DST, and the predicted resistant variants were validated.

INDIA KEY FINDINGS/ACHIEVEMENTS

• Largest contribution of TB sequences to the global repository
• Drug resistance profile and strain lineage determined for all the study isolates.
• Transmission clusters of Beijing determined in Mumbai Metropolitan Region
• Demonstrated through local genome analysis the drastic increase of fluoroquinolone drug resistance, as well as emerging bedaquiline (new drug) resistance, which endangers the success of newly endorsed MDR-TB treatment regimens

PUBLICATIONS

• CRyPTIC Consortium (2024). Quantitative measurement of antibiotic resistance in Mycobacterium tuberculosis, reveals genetic determinants of resistance and susceptibility in a target gene approach. Nat Commun, 15(1): 488. Res Sq [Preprint]. rs.3.rs-3378915. doi.org/1021203/rs.3.rs-3378915/v1
• Sonnenkalb, L., Carter, J. J., Spitaleri, A, et al. (The CRyPTIC Consortium) (2023). Bedaquiline and clofazimine resistance resistance in Mycobacterium tuberculosis: an in vitro and in silico data analysis. Lancet Microbe, 4: e358–368.
• The CRyPTIC Consortium. (2022). A data compendium associating the genomes of 12,289 Mycobacterium tuberculosis isolates with quantitative resistance phenotypes to 13 antibiotics. PLoS Biol, 20: e 3001721. bioRxiv preprint doi:10.1101/2021.09.14.460274
• The CRyPTIC Consortium. (2022). Genome-wide association studies of global Mycobacterium tuberculosis resistance to 13 antimicrobials in 10,228 genomes identify new resistance mechanisms. PLoS Biol, 20: e3001755. bioRxiv preprintdoi:10.1101/2021.09.14.460272
• Fowler, P.W., Wright, C., Spiers, H. et al. (The CRyPTIC Consortium). A crowd of BashTheBug volunteers reproducibly and accurately measure the minimum inhibitory concentrations of 13 antitubercular drugs from photographs of 96-well broth microdilution plates. Elife, 11: e75046. bioRxiv preprint doi:10.1101/2021.07.20.453060
• The CRyPTIC Consortium. (2022). Epidemiological cutoff values for a 96-well broth microdilution plate for high-throughput research antibiotic susceptibility testing of M. tuberculosis.Eur Resp J, 60(4):2200239.
• Hunt, M., Letcher, B., Malone, K. M., et al. (CRyPTIC consortium) (2022). Minos: variant adjudication and joint genotyping of cohorts of bacterial genomes. Genome Biol, 23(1): 147.
• Walker, T.M., Miotto, P., Köser, C.U. et al. (The CRyPTIC Consortium) (2022). The 2021 WHO catalogue of Mycobacterium tuberculosis complex mutations associated with drug resistance: A new global standard for molecular diagnostics. Lancet Microbe, 3: 265-273.
• Yang, Y., Walker, T.M., Kouchaki, S., et al. (CRyPTIC Consortium) (2021). An end-to-end heterogeneous graph attention network for Mycobacterium tuberculosis drug-resistance prediction. Brief Bioinform, 22(6): bbab299.
• Adams, O., Deme, J.C., Parker, J.L,…. et al. (The CRyPTIC Consortium) (2021). Cryo-EM structure and resistance landscape of M. tuberculosis MmpL3: An emergent therapeutic target. Structure, 29:1
• Horter, S., Daftary, A., Keam, T.,…Mistry, N. et al. (2021). Person-centred care in tuberculosis. Int J Tuber and Lung Dis, 25: 784-787.
• The CRyPTIC Consortium, Lachapelle, A.C. (2021). A generalisable approach to drug susceptibility prediction for M. tuberculosis using machine learning and whole-genome sequencing.
• The CRyPTIC Consortium, Carter, J.J. (2021). Quantitative measurement of antibiotic resistance in Mycobacterium tuberculosis reveals genetic determinants of resistance and susceptibility in a target gene approach. bioRxiv preprint doi.org/10.1101/2021.09.14.460353
• Sonnenkalb, L., Carter, J., Spitaleri,…Nilgiriwala, K., et al. (The CRyPTIC Consortium) (2021). Deciphering bedaquiline and clofazimine resistance in tuberculosis: An evolutionary medicine approach. bioRxiv preprint doi.org/10.1101/2021.03.19.436148
• Dreyer, V., Mandal, A., Dev, P…. Nilgiriwala, K., et al. (The CRyPTIC Consortium) (2022). High fluoroquinolone resistance proportions among multidrug resistant tuberculosis driven by dominant L2 Mycobacterium tuberculosis clones in the Mumbai Metropolitan Region. Genome Med, 14: 1-16. bioRxiv preprint doi:10.1101/2021.02.02.429364.
• Kouchaki, S., Yang, Y., Lachapelle, A. et al. (CRyPTIC Consortium) (2020). Multi-label random forest model for tuberculosis drug resistance classification and mutation ranking. Front Microbiol, 11: 667.
• Wilson, D.J., The CRyPTIC Consortium. (2020). GenomegaMap within-species genome-wide d _N/d _S estimation from over 10,000 genomes. Mol Biol Evol, 37: 2450-2460
• Yang, Y., Walker, T.M., Walker, A.S.,… et al. (The CRyPTIC Consortium) (2019). DeepAMR for predicting co-occurrent resistance of Mycobacterium tuberculosis. Bioinformatics, 35: 3240-3249.
• Kouchaki, S., Yang, Y., Walker, T.M.,… et al. (The CRyPTIC Consortium) (2018). Application of machine learning techniques to tuberculosis drug resistance analysis. Bioinformatics, 35: 2276–2282.
• Rancoita, P.M.V., Cugnata, F., Gibertoni, Cruz A.L.,… et al. (The CRyPTIC Consortium) (2018). Validating a 14-drug microtiter plate containing bedaquiline and delamanid for large scale research susceptibility testing of Mycobacterium tuberculosis. Antimicrob. Agent Chemother, 62: e00344-18.
• Sagili, K.D., Muniyandi, M., Nilgiriwala, K.S. et al. (2018). Cost-effectiveness of GeneXpert and LED-FM for diagnosis of pulmonary tuberculosis: A systematic review. PLOS One, 13: e0205233.
• CRyPTIC Consortium and the 100,000 Genomes Project (2018). Prediction of susceptibility to first-line tuberculosis drugs by DNA sequencing: The CRyPTIC Consortium and the 100,000 Genomes Project. N Engl J Med, 379: 1403-1415.

CONFERENCE PRESENTATIONS

• Nilgiriwala, K. High minimum inhibitory of Bedaquiline naive clinical isolates of Mycobacterium tuberculosis from Mumbai, India. Oral Presentation. UNION/CDC Late-breaker session on “Tuberculosis, TB-HIV and diabetes and lung health”, organized by UNION/CDC, Hyderabad, 2nd November 2019.
• Mistry N. Harnessing political support and inter-sectoral Co-ordination for TB elimination by 2025. Invited Panelist. Meeting at 50th Union World Conference on Lung Health,organized by Global Coalition Against TB, Hyderabad,31st October 2019.
• Nilgiriwala, K. Next generation DNA sequencing and its application in TB. Oral Presentation. Meeting on “NGS for DR-TB” organized by Medecins Sans Frontiers, Mumbai, 9th May 2019.
• Nilgiriwala, K. Application of WGS for Public Health. Oral Presentation. Truth seekers meeting on “Application of WGS for Public Health” Organized by Tata Institute of Social Sciences, Mumbai, 20th July 2017.
• Mistry, N. Highlighting issues likely to affect application and outcomes of WGS in India for TB control. Oral Presentation. Workshop on “Building capacity for Whole genome sequencing of mycobacterial strains” organized by National Institute for Research in Tuberculosis, Chennai, 19th February 2017.
• Mistry, N. Insight into Applications of whole shotgun sequencing of M. tuberculosis in the high endemic region of Mumbai. Poster Discussion. 47th Union World Conference on “Lung Health” organized by The Union, Liverpool (UK), 26th -29th October 2016.

ABOUT THE PROJECT

A pilot study aimed to test the feasibility of portable Nanopore sequencing (using MinION – a USB-based portable sequencing device) on DNA isolated from sputum containing Mycobacterium tuberculosis (M.tb) was explored. Nanopore sequencing is a unique, simple to use technology that enables direct, real-time analysis of long DNA or RNA fragments. It has fast turnaround time making it an ideal technology for field use. In this study, Nanopore sequencing of sputum DNA samples was conducted by single plex and by multiplexing of 6-8 samples in one flow cell to check the feasibility of creating a cost-effective point-of-care test for TB.

Subsequently an additional study (Phase II) entitled Rapid diagnosis of TB from sputum DNA (isolation by UBB protocol) using Nanopore sequencing platform was undertaken in which the Ultimate Boiling Buffer (UBB) protocol standardized by Oxford for isolation of Mycobacterium tuberculosis (M.tb) DNA was tested on 30 clinical sputum samples. The lower limit of detection of M.tb in sputum required for prediction of DST and strain lineage was determined. Validation of Nanopore sequencing data was conducted by GeneXpert, Illumina sequencing of sputum & culture DNA, and culture DST. Multiplexing (of up to 5 samples in one flow cell) to design a cost-effective workflow for potential diagnosis of TB from sputum in future was tested

KEY FINDINGS/ACHIEVEMENTS

PUBLICATIONS

• Votintseval, A.A., Bradley, P., Pankhurst, L., Loose, M., Del Ojo, E.C., Nilgiriwala, K., Chatterjee, A. et al. (2016). Same-day diagnostic and surveillance data for tuberculosis via whole genome sequencing of direct respiratory samples. JCM, 55: 1285-1298.

CONFERENCE PRESENTATIONS

• Nilgiriwala, K. Diagnosis of tuberculosis by whole genome sequencing of DNA from sputum. Oral Presentation. International Virtual Conference “London Calling: Dedicated to Nanopore DNA/RNA sequencing” organized by Oxford Nanopore Technologies Limited from 17th to 19th June 2020.
• Nilgiriwala, K. Towards Point-of-Care Diagnosis of Pulmonary Tuberculosis and Drug Susceptibility Testing by Whole Genome Sequencing of DNA Isolated from Sputum. Poster presentation. Conference on “2017 Next Gen Genomics, Biology, Bioinformatics, Technology”, organized by SciGenom Research Foundation, Bhubaneswar, 2nd-4th October 2017.

ABOUT THE PROJECT

The Foundation invested in a survey of health facilities in high-burden TB wards of Mumbai and in the training of the facility staff in adopting infection control (IC) measures. Personal practice, infrastructural and administrative aspects of IC were recorded at these facilities and outcomes were conveyed to the local RNTCP and MCGM. The introduction of germicidal UV lights in in-patient and out-patient care was advocated in both Delhi and Mumbai. This initiative was successfully taken up at scale by the National Institute of TB and Respiratory Diseases, New Delhi, with around 32 units installed in various locations. The staff of NITRD were repeatedly trained in maintenance and measurement of lamp and fixture efficacy. The latter was undertaken with help from CDC, USA expert, a Fogarty Fellow and Mr. Bill Palmer (Aeromed) who supplied and supervised the installation for correctness and safety.

ABOUT THE PROJECT

The aim of the study was to examine and validate critical genes associated with the acquisition of multi-drug resistance in longitudinal clinical isolates and in vitro generated resistant strains by quantitative real-time PCR

KEY FINDINGS/ACHIEVEMENTS

• The study identified five genes associated with drug resistance that were not previously known to be associated with the development of drug resistance
• Among the five genes identified, the levels of ppsD, a critical gene involved in the production of lipids present on the surface of the bacteria, appeared to reflect treatment response in patients and hence could be considered as a potential biomarker for the prediction of poor treatment response in patients undergoing first-line TB treatment

PUBLICATIONS

• Sriraman, K.,Kekane, R., Shah, D., Saranath, D. & Mistry, N. (2019). Expression of ppsD a gene involved in synthesis of Mycobacterium tuberculosis virulence factor PDIM, reflects treatment response in pulmonary tuberculosis patients. bioRxiv, 576470.
• Sriraman,K., Nilgiriwala, K., Saranath, D., Chatterjee, A. & Mistry, N.(2017). Deregulation of genes associated with alternate drug resistance mechanism of Mycobacterium tuberculosis. CurrMicrobiol, 75: 394-400.

CONFERENCE PRESENTATIONS

• Sriraman K. ppsD –a potential marker for treatment responses in tuberculosis infection. Poster presentation. Keystone Symposia on Molecular & Cellular Biology on “Tuberculosis: Translating Scientific Findings for Clinical and Public Health Impact(X7)”, organized by Keystone Symposia, Canada, 15th - 19th April 2018.
• Sriraman K. Transcriptional analysis of genes associated with rapid acquisition of multidrug resistance in Mycobacterium tuberculosis. Oral Presentation. 47th Union World Conference on “Lung Health” organized by The Union, Liverpool (UK), on 26th -29th October 2016
• Sriraman, K. Transcriptional analysis of gene associated with rapid acquisition of MDR in MTB. Oral Presentation. Joint meeting of stakeholders on “Progress and Plans of TB Control Activities in Mumbai” organized by Municipal Corporation of greater Mumbai, Central TB Division and Maharashtra state, Mumbai, 16th August 2016.

ABOUT THE PROJECT

A private sector engagement model for control of TB, the public private interface agency (PPIA) was attempted in response to the National Strategic Plan in Mumbai and Patna for drug sensitive (DS) and drug resistant (DR) TB patients. The former was evaluated over 2 years after maturity to examine effect on reduction of patient pathways and retention. The model engaged private providers, diagnostic facilities and pharmacies into an effective network providing free diagnostic tests and treatment.

KEY FINDINGS/ACHIEVEMENTS

Drug Sensitive TB Patients

• Patients first accessing engaged facilities at first point of care had shorter pathways from symptoms to initiation of treatment.
• The NTEP in Mumbai provided early diagnosis compared to private providers engaged with PPIA.
• Good retention of patients post diagnosis was experienced in Mumbai and Patna, but a hub-spoke model in Mumbai hampered retention emphasizing effects of local design. Multiple diagnosis in patients were a common feature. On average DS-TB patients accessed 2-3 providers during the pathway.

Drug Resistant – TB patients

• Total pathway from symptom to treatment initiation in new/primary DR cases was twice that of treatment patients (190 vs. 69 days)
• Qualitative research highlighted the role of awareness action gap in patients, the extensive distance that DR patients had to travel to avail diagnosis treatment (89-650 kms). The case histories were vastly heterogenous implying that a wide breadth of case features (comprehensive care would need to be implemented)
• Uncertainties and bewilderment plague the pathway when on an average 7-9 providers are seen comprising both public and private providers

The Project undertaken on initiation by the BMGF spanning 5 years constituted the longest national field study undertaken by the Foundation. The FMR was considerably strengthened in field work measures through this experience and was able to contribute meaningfully with recommendations for improving patient care.

PUBLICATIONS

• Shah, S., Shah, S., Rangan, S., Rai, S., Lobo, E., Kamble, S., Dholakia, Y. & Mistry, N.F. (2020). Effect of public-private interface agency in Patna and Mumbai, India: Does it alter durations and delays in care seeking for drug-sensitive pulmonary tuberculosis? Gates Open Research, 4: 1-15.
• Bhattacharya Chakravarty, A., Rangan, S., Rai, S., Kamble, S.,Raste, T., Shah, S., Dholakia, Y. Shah, S.& Mistry, N.(2019). Such a long journey: What health seeking pathways of patients with drug resistant tuberculosis in Mumbai tell us?PLOS One, 14: e0209924.
• Lobo, E.,Shah, S., Rangan, S., Dholakia, Y. & Mistry, N.(2018). Pathway to care for drug resistant tuberculosis cases during a retrospective study conducted in high TB burden wards in Mumbai. Gates Open Research, 2: 1-6.
• Mistry, N., Lobo, E., Rangan, S. & Dholakia, Y. (2017). Pulmonary tuberculosis in Patna India: duration delays and health care seeking behavior among patients identified through household survey. J. Epidemiology & Global Health, 7: 241-248.
• Dholakia, Y., Mistry, N., Lobo, E. &Rangan, S. (2016). Use of standardized patients to assess quality of tuberculosis care. Lancet Infect Dis, 16: 23.
• Mistry, N., Rangan, S., Dholakia, Y., Lobo, E., Shah, S. & Patil, A. (2016). Duration and delay in care seeking, diagnosis and treatment initiation in uncomplicated pulmonary tuberculosis patients in Mumbai. PLOS One, 11: e0152287.

CONFERENCE PRESENTATIONS

• Mistry, N. Methods and results from patient pathway analysis. Oral Presentation. Brainstorm/planning workshop to plan “Possible studies that can evaluate the impact of the private sector engagement in India”, organized by Bill and Melinda Gates Foundation, Hyderabad, 3rd November 2019.
• Dholakia, Y. Lessons from Public Private Interface Agency (PPIA) intervention in TB care: Mumbai, Patna, India”. Poster Presentation. 50th Union World Conference on Lung Health on “Ending the emergency: Science leadership action”, organized by The Union, Hyderabad, 2nd November 2019.
• Dev, P. Footfalls and days: Journey of pulmonary drug resistant TB patients in Mumbai. Poster Presentation. 50th Union World Conference on Lung Health on “Ending the emergency: Science leadership action”, organized by The Union, Hyderabad, 31st October 2019.
• Mistry, N. Optimizing private engagement to streamline patient pathways. Panel Discussion. Global Coalition Against TB Expert Group Meeting on “Patient pathways and challenges that TB care seekers face in navigating a complex ecosystem and adapting existing interventions to improving access to care while ensuring affordable care which is agnostic to sector in which it is sought” organized by Global Health Strategies, New Delhi, 28th March 2019.
• Dholakia, Y. TB care seeking in Mumbai: Duration, movements and retention in care. Oral Presentation. 73rd NATCON conference on “Tuberculosis and chest diseases” organized by Indira Gandhi Govt. Medical college and TB association of Maharashtra, Nagpur, from 4th - 6th January 2019.
• Mistry, N. DS-PTB patient Retention / Movement in Mumbai. Panel Discussion on “ Patient Retention and Tuberculosis” organized by Harvard T.H. Chan School of Public Health and Foundation for Medical Research, Mumbai, 29th August 2018.
• Mistry, N. Patient pathways to care in TB. Invited speaker. Winter Symposium on “Advances in Tuberculosis Research”, organized by Christian Medical College Vellore, RePORT and Indo US consortium, Vellore, 11th to 13th February 2016.

ABOUT THE PROJECT

This project aimed at exploitation of RNA interference (RNAi) approach for reducing the intramacrophage load of Mycobacterium tuberculosis. This was proposed to be accomplished through siRNA-loaded nanoplexes targeting intramacrophage proteins required for the survival of the bacteria. The role of FMR was to test the specificity of siRNA through checking for the expression of host bfl-1 posttreatment with the generated siRNA’s using quantitative real-time PCR and determine the bactericidal potential of the siRNA particles through use of routinely practiced in-vitro macrophage model.

KEY FINDINGS/ACHIEVEMENTS

• The study showed the chitosan nanoplexes developed by ICT, Mumbai, were found to be significantly (99.1%) inhibitory to TB bacteria at a modest concentration of 500 μg/ mL and non-toxic to host cells (macrophages)
• The study was one of the first reports demonstrating the inhibitory effect of water-soluble chitosan oligosaccharides nanoplexes against TB bacteria and that it could be potentially employed for the development of effective drug delivery vehicles or systems against TB bacteria
• Further, the study demonstrated that chitosan nanoplexes could efficiently deliver siRNAs directed against host proteins required for TB survival into host cells and therefore can be potentially used as a treatment option for TB.

PUBLICATIONS

• Koli, U., Nilgiriwala, K., Sriraman, K., Jain R, Dandekar P. Targeting tuberculosis infection in macrophages using chitosan oligosaccharide nanoplexes. 2019. J Nanopart Res 21, 200.