Prevalence and Geographical Distribution of Various Nontuberculous Mycobacterial (NTM) Species/Subspecies in India: Early Findings of Indian Council of Medical Research (ICMR), New Delhi Sponsored Multicentre Research Project Between 2021-2024

Abstract Rationale: Prevalence of nontuberculous mycobacterial (NTM) disease is increasing globally. In high-TB-burden countries because of similarities with TB, NTM disease is usually not considered in the differential diagnosis and treated as TB. A prospective multicentre study was conducted at six sites in five states of India between 2021-2024 to determine prevalence of NTM disease and geographic variation of NTM species/subspecies among TB-suspects. Methods: The study was performed on GeneXpert MTB/RIF (Cepheid, USA) negative pulmonary and extrapulmonary specimens applying inclusion and exclusion criteria. All participating centres obtained ethical clearance. Written informed consent was obtained from all study patients. The confirmed NTM isolates (negative on SD-MPT64TB Ag immunochromatography kit, Standard Diagnostics, South Korea) were subjected to GenoType®-mycobacterium common mycobacteria/additional species and GenoType®-NTM-DR line probe assay (LPA) kits (Hain Lifescience, Germany) for NTM species/subspecies identification and were later validated by whole genome sequencing (WGS, Illumina-MiSeq sequencing using Nextra XT-DNA Library Prep protocol, USA). Demographic and other clinical details were carefully recorded. American Thoracic Society 2007 and 2020 guidelines were followed for NTM diagnosis. Results: NTM were isolated from 226 patients [NTM-pulmonary-disease (NTM-PD)=187; extrapulmonary-NTM (EPNTM)=39] among 71,143 TB-suspects with a prevalence of 0.32%. Of these 138(61%) were male. The median time to diagnose NTM was 119.7 days (IQR=57.1-132.1). The mean age of patients was 46.1 years (range=12-73). The mean body mass index (BMI) was 23.1 kg/m2(SD=3.1) and 39.8% (90/226) patients had BMI <18.5 kg/m2. Overall isolated NTM species included: M.avium complex (MAC) (n=58; M.intracellulare=38, M.avium=20), M.abscessus (n=56; M.abscessus subsp. abscessus=29, M.abscessus subsp. massiliense=25, M.abscessus subsp. bolletii=2), M.fortuitum (n=46), M. kansasii (n=45), M. simiae (n=11), M. gordonae (n=7) and one each of M.lentiflavum, M.senegalense and M.mucogenicum. LPA showed 97.6% (122/125) concordance with WGS results in NTM species/subspecies identification. One clarithromycin-resistant M.abscessus subsp. abscessus isolate [T28 sequevar of erm (41) gene] was identified by LPA and WGS. In NTM-PD, species isolated were MAC (27.8%;52/187), M.kansasii (24.1%;45/187), M.abscessus (21.4%; 40/187), M.fortuitum (16%;30/187), M.simiae (5.9%;11/187), M.gordonae (3.7%;7/187), M.mucogenicum (0.5%;1/187) and M.lentiflavum (0.5%;1/187). In EPNTM, M.abscessus (41%;16/39), M.fortuitum (41%;16/39), MAC (15.4%;6/39) and M.senegalense (2.6%;1/39) were commonest species. Following regional variations of NTM species were noted: MAC was predominantly isolated from north and south Indian states, M.kansasii, and M.abscessus from western and central states respectively. Conclusion: This is the first comprehensive Indian study describing prevalence and geographical variation of NTM species/subspecies among TB-suspects.LPA can be safely recommended for NTM diagnosis in high-TB-burden countries.