SAM-TB: a whole genome sequencing data analysis website for detection of Mycobacterium tuberculosis drug resistance and transmission.

Whole genome sequencing (WGS) can provide insight into drug-resistance, transmission chains and the identification of outbreaks, but data analysis remains an obstacle to its routine clinical use. Although several drug-resistance prediction tools have appeared, until now no website integrates drug-resistance prediction with strain genetic relationships and species identification of nontuberculous mycobacteria (NTM). We have established a free, function-rich, user-friendly online platform for MTB WGS data analysis (SAM-TB, http://samtb.szmbzx.com) that integrates drug-resistance prediction for 17 antituberculosis drugs, detection of variants, analysis of genetic relationships and NTM species identification. The accuracy of SAM-TB in predicting drug-resistance was assessed using 3177 sequenced clinical isolates with results of phenotypic drug-susceptibility tests (pDST). Compared to pDST, the sensitivity of SAM-TB for detecting multidrug-resistant tuberculosis was 93.9% [95% confidence interval (CI) 92.6-95.1%] with specificity of 96.2% (95% CI 95.2-97.1%). SAM-TB also analyzes the genetic relationships between multiple strains by reconstructing phylogenetic trees and calculating pairwise single nucleotide polymorphism (SNP) distances to identify genomic clusters. The incorporated mlstverse software identifies NTM species with an accuracy of 98.2% and Kraken2 software can detect mixed MTB and NTM samples. SAM-TB also has the capacity to share both sequence data and analysis between users. SAM-TB is a multifunctional integrated website that uses WGS raw data to accurately predict antituberculosis drug-resistance profiles, analyze genetic relationships between multiple strains and identify NTM species and mixed samples containing both NTM and MTB. SAM-TB is a useful tool for guiding both treatment and epidemiological investigation.

China's tuberculosis epidemic stems from historical expansion of four strains of Mycobacterium tuberculosis.

A small number of high-burden countries account for the majority of tuberculosis cases worldwide. Detailed data are lacking from these regions. To explore the evolutionary history of Mycobacterium tuberculosis in China-the country with the third highest tuberculosis burden-we analysed a countrywide collection of 4,578 isolates. Little genetic diversity was detected, with 99.4% of the bacterial population belonging to lineage 2 and three sublineages of lineage 4. The deeply rooted phylogenetic positions and geographic restriction of these four genotypes indicate that their populations expanded in situ following a small number of introductions to China. Coalescent analyses suggest that these bacterial subpopulations emerged in China around 1,000 years ago, and expanded in parallel from the twelfth century onwards, and that the whole population peaked in the late eighteenth century. More recently, sublineage L2.3, which is indigenous to China and exhibited relatively high transmissibility and extensive global dissemination, came to dominate the population dynamics of M. tuberculosis in China. Our results indicate that historical expansion of four M. tuberculosis strains shaped the current tuberculosis epidemic in China, and highlight the long-term genetic continuity of the indigenous M. tuberculosis population.

Have compensatory mutations facilitated the current epidemic of multidrug-resistant tuberculosis?

Compensatory mutations have been suggested to promote multidrug-resistant tuberculosis (MDR-TB) transmission, but their role in facilitating the recent transmission of MDR-TB is unclear. To investigate the epidemiological significance of compensatory mutations, we analyzed a four-year population-based collection of MDR-TB strains from Shanghai (the most populous city in China) and 1346 published global MDR-TB strains. We report that MDR-TB strains with compensatory mutations in the rpoA, rpoB, or rpoC genes were neither more frequently clustered nor found in larger clusters than those without compensatory mutations. Our results suggest that compensatory mutations are not a major contributor to the current epidemic of MDR-TB.

Internal migration and transmission dynamics of tuberculosis in Shanghai, China: an epidemiological, spatial, genomic analysis.

BACKGROUND: Massive internal migration from rural to urban areas poses new challenges for tuberculosis control in China. We aimed to combine genomic, spatial, and epidemiological data to describe the dynamics of tuberculosis in an urban setting with large numbers of migrants. METHODS: We did a population-based study of culture-positive Mycobacterium tuberculosis isolates in Songjiang, Shanghai. We used whole-genome sequencing to discriminate apparent genetic clusters of M tuberculosis sharing identical variable-number-tandem-repeat (VNTR) patterns, and analysed the relations between proximity of residence and the risk of genomically clustered M tuberculosis. Finally, we used genomic, spatial, and epidemiological data to estimate time of infection and transmission links among migrants and residents. FINDINGS: Between Jan 1, 2009, and Dec 31, 2015, 1620 cases of culture-positive tuberculosis were recorded, 1211 (75%) of which occurred among internal migrants. 150 (69%) of 218 people sharing identical VNTR patterns had isolates within ten single-nucleotide polymorphisms (SNPs) of at least one other strain, consistent with recent transmission of M tuberculosis. Pairs of strains collected from individuals living in close proximity were more likely to be genetically similar than those from individuals who lived far away-for every additional km of distance between patients' homes, the odds that genotypically matched strains were within ten SNPs of each other decreased by about 10% (OR 0·89 [95% CI 0·87-0·91]; p<0·0001). We inferred that transmission from residents to migrants occurs as commonly as transmission from migrants to residents, and we estimated that more than two-thirds of migrants in genomic clusters were infected locally after migration. INTERPRETATION: The primary mechanism driving local incidence of tuberculosis in urban centres is local transmission between both migrants and residents. Combined analysis of epidemiological, genomic, and spatial data contributes to a richer understanding of local transmission dynamics and should inform the design of more effective interventions. FUNDING: National Natural Science Foundation of China, National Science and Technology Major Project of China, and US National Institutes of Health.

Assessment of tuberculosis contact investigation in Shanghai, China: An 8-year cohort study.

BACKGROUND: Tuberculosis (TB) contact investigation has been observed as a useful programmatic tool in active case finding. We collected data of contact cases to evaluate the effectiveness of TB contact investigation programme in Shanghai, China. METHODS: Since 2009, we screened and followed up the close contacts of bacteria-positive TB cases in Songjiang, Shanghai and calculated the incidence of TB in close contacts and confirmed the transmission by genotyping and sequencing. RESULTS: A total of 4584 close contacts of 1765 contagious TB index cases were followed up for an average of 4 years. About 62 contacts (333/100 000, 95% CI: 256-428) developed TB excluding 6 co-prevalent cases. The contact cases consisted 1.50% (39/2592) of all the bacteria-positive cases in population. Transmission links were confirmed in 60% (9/15) familial contacts and 22% (2/9) in non-familial contacts. Source cases come from more than close contacts and both index and contact cases created other secondary cases in community. CONCLUSIONS: Familial contacts are more likely to acquire TB from the index, indicating the priority of family members in TB contact investigation in China. However, most non-familial contacts were infected from sources in the community and contact cases attributed little to case finding in the TB-prevalent setting. Thus, active case finding should be strengthened in general population.

Transmission is a Noticeable Cause of Resistance Among Treated Tuberculosis Patients in Shanghai, China.

It is generally believed that drug resistance among treated tuberculosis (TB) patients is as a result of acquired drug resistance due to inappropriate treatment. Previous studies have shown that primary drug resistance caused by transmission also plays a role among treated cases. Differentiating the two types of drug resistance will help in developing appropriate strategies for control of drug resistant tuberculosis. In this study, we tested the hypothesis that drug resistance among treated TB patients is mainly caused by primary resistance rather than acquired resistance. Defining resistance profiles by molecular drug susceptibility test, we used Unit Variable Number Tandem Repeats (VNTR) to genotype and Whole Genome Sequencing (WGS) to confirm the accordance of the first and last Mycobacterium tuberculosis isolates from treated pulmonary TB patients in Shanghai from 2009-2015. Among 81 patients with increasing drug resistance, out of 390 patients enrolled, paired isolates from 59.3% (48/81) had different VNTR patterns indicating primary drug resistance. Our results have demonstrated that primary resistance due to exogenous reinfection is the major cause of drug resistance among treated TB patients in Shanghai; thus, strategies aimed at preventing and interrupting transmission are urgently needed to effectively reduce the epidemic of drug resistant tuberculosis.