To improve understanding of the factors influencing tuberculosis transmission and the role of pathogen variation, we sequenced all available specimens from patients diagnosed over 15 years in a whole district in Malawi. Mycobacterium tuberculosis lineages were assigned and transmission networks constructed, allowing ≤10 single nucleotide polymorphisms (SNPs) difference. We defined disease as due to recent infection if the network-determined source was within 5 years, and assessed transmissibility from forward transmissions resulting in disease. High-quality sequences were available for 1687 disease episodes (72% of all culture-positive episodes): 66% of patients linked to at least one other patient. The between-patient mutation rate was 0.26 SNPs/year (95% CI 0.21–0.31). We showed striking differences by lineage in the proportion of disease due to recent transmission and in transmissibility (highest for lineage-2 and lowest for lineage-1) that were not confounded by immigration, HIV status or drug resistance. Transmissions resulting in disease decreased markedly over time.DOI: http://dx.doi.org/10.7554/eLife.05166.001

Background. Recurrent tuberculosis is a major health burden and may be due to relapse with the original strain or reinfection with a new strain.Methods. In a population-based study in northern Malawi, patients with tuberculosis diagnosed from 1996 to 2010 were actively followed after the end of treatment. Whole-genome sequencing with approximately 100-fold coverage was performed on all available cultures. Results of IS6110 restriction fragment-length polymorphism analyses were available for cultures performed up to 2008.Results. Based on our data, a difference of ≤10 single-nucleotide polymorphisms (SNPs) was used to define relapse, and a difference of >100 SNPs was used to define reinfection. There was no evidence of mixed infections among those classified as reinfections. Of 1471 patients, 139 had laboratory-confirmed recurrences: 55 had relapse, and 20 had reinfection; for 64 type of recurrence was unclassified. Almost all relapses occurred in the first 2 years. Human immunodeficiency virus infection was associated with reinfection but not relapse. Relapses were associated with isoniazid resistance, treatment before 2007, and lineage-3 strains. We identified several gene variants associated with relapse. Lineage-2 (Beijing) was overrepresented and lineage-1 underrepresented among the reinfecting strains (P = .004).Conclusions. While some of the factors determining recurrence depend on the patient and their treatment, differences in the Mycobacterium tuberculosis genome appear to have a role in both relapse and reinfection.

BackgroundMixed, polyclonal Mycobacterium tuberculosis infection occurs in natural populations. Developing an effective method for detecting such cases is important in measuring the success of treatment and reconstruction of transmission between patients. Using whole genome sequence (WGS) data, we assess two methods for detecting mixed infection: (i) a combination of the number of heterozygous sites and the proportion of heterozygous sites to total SNPs, and (ii) Bayesian model-based clustering of allele frequencies from sequencing reads at heterozygous sites.ResultsIn silico and in vitro artificially mixed and known pure M. tuberculosis samples were analysed to determine the specificity and sensitivity of each method. We found that both approaches were effective in distinguishing between pure strains and mixed infection where there was relatively high (> 10%) proportion of a minor strain in the mixture. A large dataset of clinical isolates (n = 1963) from the Karonga Prevention Study in Northern Malawi was tested to examine correlations with patient characteristics and outcomes with mixed infection. The frequency of mixed infection in the population was found to be around 10%, with an association with year of diagnosis, but no association with age, sex, HIV status or previous tuberculosis.ConclusionsMixed Mycobacterium tuberculosis infection was identified in silico using whole genome sequence data. The methods presented here can be applied to population-wide analyses of tuberculosis to estimate the frequency of mixed infection, and to identify individual cases of mixed infections. These cases are important when considering the evolution and transmission of the disease, and in patient treatment.Electronic supplementary materialThe online version of this article (10.1186/s12864-018-4988-z) contains supplementary material, which is available to authorized users.

BackgroundThe proportion of tuberculosis attributable to transmission from close contacts is not well known. Comparison of the genome of strains from index patients and prior contacts allows transmission to be confirmed or excluded.MethodsIn Karonga District, Malawi, all tuberculosis patients are asked about prior contact with others with tuberculosis. All available strains from culture-positive patients were sequenced. Up to 10 single nucleotide polymorphisms between index patients and their prior contacts were allowed for confirmation, and ≥ 100 for exclusion. The population attributable fraction was estimated from the proportion of confirmed transmissions and the proportion of patients with contacts.ResultsFrom 1997–2010 there were 1907 new culture-confirmed tuberculosis patients, of whom 32% reported at least one family contact and an additional 11% had at least one other contact; 60% of contacts had smear-positive disease. Among case-contact pairs with sequences available, transmission was confirmed from 38% (62/163) smear-positive prior contacts and 0/17 smear-negative prior contacts. Confirmed transmission was more common in those related to the prior contact (42.4%, 56/132) than in non-relatives (19.4%, 6/31, p = 0.02), and in those with more intense contact, to younger index cases, and in more recent years. The proportion of tuberculosis attributable to known contacts was estimated to be 9.4% overall.ConclusionsIn this population known contacts only explained a small proportion of tuberculosis cases. Even those with a prior family contact with smear positive tuberculosis were more likely to have acquired their infection elsewhere.

Background-Patient retention in antiretroviral therapy (ART) programs remains a major challenge in sub-Saharan Africa. We examined whether and why retention in ART care has changed with increasing access.