A Genome-Focused Investigation Reveals the Emergence of a Mycobacterium tuberculosis Strain Related to Multidrug-Resistant Tuberculosis in the Amazon Region of Brazil.

A previous study in Pará, Northern Brazil, described a strain of Mycobacterium tuberculosis with a unique genotype (SIT2517/T1) associated with multidrug-resistant tuberculosis (MDR-TB). To improve our understanding of MDR-TB transmission dynamics of these strains within this region, we performed phenotypic and genotypic drug susceptibility testing (pDST/gDST), 24-loci mycobacterial interspersed repetitive units (MIRU-VNTR) genotyping, whole-genome sequencing (WGS) and geo-epidemiology analysis. Of the 28 SIT2517/T1 isolates, 19 (67.9%) could be genotyped by 24-loci MIRU-VNTR and 15 by WGS. All belonged to sublineage 4.1.1.3, distinct from other representative Lineage 4 isolates identified in Brazil. The MDR phenotype determined by pDST was confirmed by gDST, the latter also demonstrating the presence of additional mutations conferring pre-extensively drug-resistance (pre-XDR). Discrepancies between gDST and pDST were observed for pyrazinamide and fluoroquinolones. Thirteen out of 15 isolates analyzed by WGS were clustered when applying a 12 single nucleotide polymorphisms (SNPs) cutoff. The SIT2517/T1 isolates were distributed across the metropolitan regions of Belém and Collares municipalities, showing no geographic clustering. WGS-transmission network analysis revealed a high likelihood of direct transmission and the formation of two closely linked transmission chains. This study highlights the need to implement TB genomic surveillance in the Brazilian Amazon region.

New insights on tuberculosis transmission dynamics and drug susceptibility profiles among the prison population in Southern Brazil based on whole-genome sequencing.

BACKGROUND: The rate of tuberculosis (TB) infection among the prison population (PP) in Brazil is 28 times higher than that in the general population, and prison environment favors the spread of TB. OBJECTIVE: To describe TB transmission dynamics and drug resistance profiles in PP using whole-genome sequencing (WGS). METHODS: This was a retrospective study of Mycobacterium tuberculosis cultivated from people incarcerated in 55 prisons between 2016 and 2019; only one isolate per prisoner was included. Information about movement from one prison to another was tracked. Clinical information was collected, and WGS was performed on isolates obtained at the time of TB diagnosis. RESULTS: Among 134 prisoners included in the study, we detected 16 clusters with a total of 58 (43%) cases of M. tuberculosis. Clusters ranged from two to seven isolates with five or fewer single nucleotide polymorphism (SNP) differences, suggesting a recent transmission. Six (4.4%) isolates were resistant to at least one anti-TB drug. Two of these clustered together and showed resistance to rifampicin, isoniazid, and fluoroquinolones, with 100% concordance between WGS and phenotypic drug-susceptibility testing. Prisoners with clustered isolates had a high amount of movement between prisons (two to eight moves) during the study period. CONCLUSIONS: WGS demonstrated the recent transmission of TB within prisons in Brazil. The high movement among prisoners seems to be related to the transmission of the same M. tuberculosis strain within the prison system. Screening for TB before and after the movement of prisoners using rapid molecular tests could play a role in reducing transmission.

New insights on tuberculosis transmission dynamics and drug susceptibility profiles among the prison population in Southern Brazil based on whole-genome sequencing

ABSTRACT Background: The rate of tuberculosis (TB) infection among the prison population (PP) in Brazil is 28 times higher than that in the general population, and prison environment favors the spread of TB. Objective: To describe TB transmission dynamics and drug resistance profiles in PP using whole-genome sequencing (WGS). Methods: This was a retrospective study of Mycobacterium tuberculosis cultivated from people incarcerated in 55 prisons between 2016 and 2019; only one isolate per prisoner was included. Information about movement from one prison to another was tracked. Clinical information was collected, and WGS was performed on isolates obtained at the time of TB diagnosis. Results: Among 134 prisoners included in the study, we detected 16 clusters with a total of 58 (43%) cases of M. tuberculosis. Clusters ranged from two to seven isolates with five or fewer single nucleotide polymorphism (SNP) differences, suggesting a recent transmission. Six (4.4%) isolates were resistant to at least one anti-TB drug. Two of these clustered together and showed resistance to rifampicin, isoniazid, and fluoroquinolones, with 100% concordance between WGS and phenotypic drug-susceptibility testing. Prisoners with clustered isolates had a high amount of movement between prisons (two to eight moves) during the study period. Conclusions: WGS demonstrated the recent transmission of TB within prisons in Brazil. The high movement among prisoners seems to be related to the transmission of the same M. tuberculosis strain within the prison system. Screening for TB before and after the movement of prisoners using rapid molecular tests could play a role in reducing transmission.

Xpert MTB/RIF performance to diagnose tuberculosis and rifampicin resistance in a reference centre in southern Brazil.

Effective treatment of tuberculosis (TB) remains a serious public health problem in many countries, including Brazil, especially when considering drug-resistant disease. Xpert MTB/RIF has been implemented in many countries to reduce the time to TB diagnosis and to rapidly detect rifampicin resistance. The study aimed to describe and evaluate Xpert MTB/RIF performance in diagnosing pulmonary TB and rifampicin resistance in a tertiary healthcare facility in Brazil. A cross-sectional study was performed, which included all isolates of confirmed pulmonary TB patients from 2015 to 2018. Both Xpert MTB/RIF and GenoType MTBDRplus assays were performed to detect rifampicin and isoniazid resistance. In addition, isolates with detected resistance to rifampicin and/or isoniazid were analysed by phenotypic testing using MGIT-960 SIRE kit and whole-genome sequencing (WGS) using Illumina MiSeq Sequencing System. 2148 respiratory specimens tested with Xpert MTB/RIF were included: n=1556 sputum, n=348 bronchoalveolar lavage and n=244 gastric washing. The overall Xpert MTB/RIF sensitivity in sputum was 94% and the overall specificity was 98%. The negative predictive value in sputum of all the patients was 99% with a positive predictive value of 89%. The concordance between Xpert MTB/RIF and phenotypic susceptibility test was 94.1%, while its concordance with WGS was 78.9%. Xpert MTB/RIF is a rapid and accurate diagnostic strategy for pulmonary TB, which can contribute to improvement in TB control. However, detection of rifampicin resistance might be associated with false-positive results.

Accuracy of whole genome sequencing versus phenotypic (MGIT) and commercial molecular tests for detection of drug-resistant Mycobacterium tuberculosis isolated from patients in Brazil and Mozambique.

BACKGROUND: The fast and accurate diagnosis of drug-resistant tuberculosis (DR-TB) is critical to reducing the spread of disease. Although commercial genotypic drug-susceptibility tests (DST) are close to the goal, they are still not able to detect all relevant DR-TB related mutations. Whole genome sequencing (WGS) allows better comprehension of DR-TB with a great discriminatory power. We aimed to evaluate WGS in M. tuberculosis isolates compared with phenotypic and genotypic DST. METHODS: This cross-sectional study evaluated 30 isolates from patients with detected DR-TB in Brazil and Mozambique. They were evaluated with phenotypic (MGIT-SIRE™) and genotypic (Xpert-MTB/RIF™, Genotype-MTBDRplus™, and MTBDRsl™) DST. Isolates with resistance to at least one first- or second-line drug were submitted to WGS and analyzed with TB profiler database. RESULTS: WGS had the best performance among the genotypic DST, compared to the phenotypic test. There was a very good concordance with phenotypic DST for rifampicin and streptomycin (89.6%), isoniazid (96.5%) and ethambutol (82.7%). WGS sensitivity and specificity for detection resistance were respectively 87.5 and 92.3% for rifampicin; 95.6 and 100% for isoniazid; 85.7 and 93.3% for streptomycin while 100 and 77.2% for ethambutol. Two isolates from Mozambique showed a Val170Phe rpoB mutation which was neither detected by Xpert-MTB/RIF nor Genotype-MTBDRplus. CONCLUSION: WGS was able to provide all the relevant information about M. tuberculosis drug susceptibility in a single test and also detected a mutation in rpoB which is not covered by commercial genotypic DST.