Abstract
BackgroundIn the context of advanced immunosuppression, M. tuberculosis is known to cause detectable mycobacteremia. However, little is known about the intra-patient mycobacterial microevolution and the direction of seeding between the sputum and blood compartments.MethodsFrom a diagnostic study of HIV-infected TB patients, 51 pairs of concurrent blood and sputum M. tuberculosis isolates from the same patient were available. In a previous analysis, we identified a subset with genotypic concordance, based on spoligotyping and 24 locus MIRU-VNTR. These paired isolates with identical genotypes were analyzed by whole genome sequencing and phylogenetic analysis.ResultsOf the 25 concordant pairs (49 % of the 51 paired isolates), 15 (60 %) remained viable for extraction of high quality DNA for whole genome sequencing. Two patient pairs were excluded due to poor quality sequence reads. The median CD4 cell count was 32 (IQR; 16–101)/mm3 and ten (77 %) patients were on ART. No drug resistance mutations were identified in any of the sequences analyzed. Three (23.1 %) of 13 patients had SNPs separating paired isolates from blood and sputum compartments, indicating evidence of microevolution.Using a phylogenetic approach to identify the ancestral compartment, in two (15 %) patients the blood isolate was ancestral to the sputum isolate, in one (8 %) it was the opposite, and ten (77 %) of the pairs were identical.ConclusionsAmong HIV-infected patients with poor cellular immunity, infection with multiple strains of M. tuberculosis was found in half of the patients. In those patients with identical strains, whole genome sequencing indicated that M. tuberculosis intra-patient microevolution does occur in a few patients, yet did not reveal a consistent direction of spread between sputum and blood. This suggests that these compartments are highly connected and potentially seed each other repeatedly.Electronic supplementary materialThe online version of this article (doi:10.1186/s12879-016-1737-2) contains supplementary material, which is available to authorized users.
Highlights
In the context of advanced immunosuppression, M. tuberculosis is known to cause detectable mycobacteremia
We considered a large cohort of human immunodeficiency virus (HIV)-positive patients who had concurrent pulmonary and blood M. tuberculosis strains and were categorized as identical, up to one spacer and/or locus difference, using the conventional methods of spoligotyping and Mycobacterial Interspersed Repetitive Units (MIRU)-Variable Number of Tandem Repeats (VNTR) 24 loci
Categorizing these isolates as identical by both spoligotyping and MIRU-VNTR 24 loci typing methods, yet with different single nucleotide polymorphisim (SNP), underscores the power of whole genome sequencing in ascertaining microevolution occurring outside the classical targeted genetic elements of M. tuberculosis compared to traditional molecular epidemiological methods [18, 19]
Summary
In the context of advanced immunosuppression, M. tuberculosis is known to cause detectable mycobacteremia. The recent advances in molecular analytical methods have increased our understanding of the possible heterogeneity of infection with Mycobacterium tuberculosis [1]. Several perspectives around this complexity in relation to HIV-infection have been documented [2]. The subtle genetic rearrangements caused by microevolution in IS6110 [7] are known to interrupt genes or modulate the expression of adjacent genes. These can affect interpretation of molecular epidemiological tests [8,9,10], whereas if this. It was suggested that such microevolution affected cavity formation, increased transmissibility of the emerging clonal variants [17]
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