Fine-grain population structure and transmission patterns of Mycobacterium tuberculosis in southern Mozambique, a high TB/HIV burden area

Abstract

Genomic studies of the Mycobacterium tuberculosis complex (MTBC) might shed light on the dynamics of its transmission, especially in high-burden settings, where recent outbreaks are embedded in the complex natural history of the disease. To this end, we conducted a 1 year prospective surveillance-based study in Mozambique. We applied whole-genome sequencing (WGS) to 295 positive cultures. We fully characterized MTBC isolates by phylogenetics and dating evaluation, and carried out a molecular epidemiology analysis to investigate further associations with pre-defined transmission risk factors. The majority of strains (49.5%, 136/275) belonged to lineage (L) 4; 57.8 % of them (159/275) were in genomic transmission clusters (cut-off 5 SNPs), and a strikingly high proportion (45.5%) shared an identical genotype (0 SNP pairwise distance). We found two 'likely endemic' clades, comprising 67 strains, belonging to L1.2, which dated back to the late 19th century and were associated with recent spread among people living with human immunodeficiency virus (PLHIV). We describe for the first time the population structure of MTBC in our region, a high tuberculosis (TB)/HIV burden area. Clustering analysis revealed an unforeseen pattern of spread and high rates of progression to active TB, suggesting weaknesses in TB control activities. The long-term presence of local strains in Mozambique, which were responsible for large transmission among HIV/TB-coinfected patients, calls into question the role of HIV in TB transmission.

Keywords: Mycobacterium tuberculosis; genomics; molecular epidemiology; transmission; tuberculosis.

Fig. 1.

Study flowchart. NTP: National Tuberculosis Programme; Xpert+: samples that tested positive by Xpert MTB/RIF; WGS: whole genome sequencing.

Fig. 2.

Phylogenetic tree, 275 strains. Lineages are represented in branch colours (pink for L1, blue for L2, purple for L3 and red for L4). Twenty-five BAPS groups are denoted by different colours in the outer ring of the phylogeny.

Fig. 3.

Proportion of strains linked by pairwise distance from 0 to 10 SNPs, using data from three different datasets. (a) Frequency of samples found at each pairwise distance. (b) Cumulative frequency (%) of isolates in transmission at pairwise distance from 0 to 10 SNPs. Colours: the blue line represents data from Mozambique (2013–2014), orange from Malawi (2009) and purple from Valencia (2015).

Fig. 4.

Phylogeographic approach. We reconstructed a phylogeny by combining isolates from Mozambique (MZ) and 8263 genomes representative of the MTBC global genetic diversity. This was used as input to infer the origin of BAPS ancestors by RASP (R package). Following defined criteria (see Methods), we classified two large endemic clades (E) and eight foreign clades (For: non-endemic). The rest of the BAPS groups could not be classified as endemic or not. MRCA: most recent common ancestor.

Fig. 5.

Endemic and non-endemic clades from L1 with BEAST dating results for the MRCA with origin in Mozambique. Pie charts represent the probability that the geographical origin of the ancestor was Mozambique (MZ). The 95 % HPD is drawn as orange intervals and refers to the ‘confidence interval’ for each ancestor estimated date. The posterior indicates the probability distribution over the parameter state space. This refers to the strength of the calculated temporal parameter. The red colour (posterior close to 1) reveals strong confidence on estimations performed for the date under the model of evolution applied. E: endemic; For: foreign (non-endemic); 95 % highest posterior density.