Ssengooba Willy, de Jong Bouke C, Joloba Moses L, Cobelens Frank G, Meehan Conor J
Department of Medical Microbiology, College of Health Sciences Makerere University, Kampala, Uganda.
Unit of Mycobacteriology, Institute of Tropical Medicine, Antwerp, Belgium.
BMC Infect Dis. 2016 Aug 5;16:371. doi: 10.1186/s12879-016-1737-2.
In 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.
From 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.
Of 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)/mm(3) 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.
Among 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.
在深度免疫抑制的情况下，已知结核分枝杆菌会导致可检测到的分枝杆菌血症。然而，关于患者体内分枝杆菌的微观进化以及痰液和血液部分之间的播散方向，人们了解甚少。
从一项针对HIV感染的结核病患者的诊断研究中，获得了来自同一患者的51对同时采集的血液和痰液结核分枝杆菌分离株。在先前的分析中，我们基于间隔寡核苷酸分型（spoligotyping）和24个位点的MIRU-VNTR确定了一个基因分型一致的亚组。对这些基因型相同的配对分离株进行全基因组测序和系统发育分析。
在25对一致的配对（占51对配对分离株的49%）中，15对（60%）仍可存活以提取用于全基因组测序的高质量DNA。由于序列读取质量差，排除了2对患者。CD4细胞计数的中位数为32（四分位间距；16 - 101）/mm³，10名（77%）患者正在接受抗逆转录病毒治疗（ART）。在分析的任何序列中均未发现耐药突变。13名患者中有3名（23.1%）存在单核苷酸多态性（SNP），使血液和痰液部分的配对分离株出现差异，表明存在微观进化证据。使用系统发育方法确定祖先部分，在2名（15%）患者中，血液分离株是痰液分离株的祖先，在1名（8%）患者中情况相反，10对（77%）配对分离株相同。
在细胞免疫功能差的HIV感染患者中，一半患者发现感染了多种结核分枝杆菌菌株。在那些菌株相同的患者中，全基因组测序表明，少数患者体内确实发生了结核分枝杆菌的微观进化，但未揭示痰液和血液之间一致的传播方向。这表明这些部分高度连通，可能会反复相互播散。
