Macrophage susceptibility to infection by Ghanaian Mycobacterium tuberculosis complex lineages 4 and 5 varies with self-reported ethnicity

BackgroundThe epidemiology of Mycobacterium tuberculosis complex (MTBC) lineage 5 (L5) infections in Ghana revealed a significantly increased prevalence in Ewes compared to other self-reported ethnic groups. In that context, we sought to investigate the early phase of tuberculosis (TB) infection using ex vivo infection of macrophages derived from the blood of Ewe and Akan ethnic group volunteers with MTBC L4 and L5 strains.MethodsThe study participants consisted of 16 controls, among which self-reported Akan and Ewe ethnicity was equally represented, as well as 20 cured TB cases consisting of 11 Akans and 9 Ewes. Peripheral blood mononuclear cells were isolated from both healthy controls and cured TB cases. CD14+ monocytes were isolated and differentiated into monocyte-derived macrophages (MDMs) before infection with L4 or L5 endemic strains. The bacterial load was assessed after 2 hours (uptake) as well as 3 and 7 days post-infection.ResultsWe observed a higher capacity of MDMs from Ewes to phagocytose L4 strains (p < 0.001), translating into a higher bacillary load on day 7 (p < 0.001) compared to L5, despite the higher replication rate of L5 in Ewe MDMs (fold change: 1.4 vs. 1.2, p = 0.03) among the controls. On the contrary, within macrophages from Akans, we observed a significantly higher phagocytic uptake of L5 (p < 0.001) compared to L4, also translating into a higher load on day 7 (p = 0.04). However, the replication rate of L4 in Akan MDMs was higher than that of L5 (fold change: L4 = 1.2, L4 = 1.1, p = 0.04). Although there was no significant difference in the uptake of L4 and L5 among cured TB cases, there was a higher bacterial load of both L4 (p = 0.02) and L5 (p = 0.02) on day 7 in Ewe MDMs.ConclusionOur results suggest that host ethnicity (driven by host genetic diversity), MTBC genetic diversity, and individual TB infection history are all acting together to modulate the outcome of macrophage infections by MTBC

Bacterial load comparison of the three main lineages of Mycobacterium tuberculosis complex in West Africa

Funding: This work was funded by The Royal Society Africa Prize 2018 awarded to Dorothy Yeboah-Manu and supported by funding from Scottish Funding Council (SCF)-Global Challenges Research Fund (GCRF) to Prof Stephen Gillespie and Dr Wilber Sabiiti.Studies have shown an association between bacterial load and virulence; however, not much is known about the diversity in this phenotypic characteristic of Mycobacterium tuberculosis complex (MTBC). This study was therefore aimed to determine the differences in bacterial load of the three most prevalent MTBC genotypes (L4, L5 and L6) in West Africa at the time of diagnosis. A total of 170 paired fresh sputum samples were collected; one part in guanidinium thiocyanate (GTC) was used for RNA extraction and tuberculosis Molecular Bacterial Load Assay (TB-MBLA) and the other part without GTC was confirmed for TB positivity using Gene Xpert MTB/RIF, smear microscopy grading and culture on Löwenstein-Jensen media slants. The 170 sputum samples comprised of 155 new cases, 3 follow-up cases and 12 TB negative sputum samples. The time-to-culture-positivity (TTP) and degree of culture positivity (DCP) were recorded. All 122 isolates obtained were Spoligotyped for lineage (L) classification but spoligotypes were obtained from 120 isolates. Of the typed isolates, 70.0%, 10.8%, 10.8%, 4.2%, 2.5%, 0.8% and 0.8% were Lineage- 4, 5, 6, 2, 3, 1, and M. bovis respectively. Further analysis of the three most prevalent lineages showed significantly shorter TTP and higher DCP by L4 compared to L5 and L6 respectively: TTP 20.8, versus 26.5, and 28.2 days; p-value=0.005 and DCP 1.27, versus 0.81 and 0.29, p<0.001. Average TB-MBLA measured bacterial load of L4 was 3.82 Log10eCFU/mL which was not significantly different from 3.81 and 3.80 Log10eCFU/mL of L5 and L6 respectively, p=0.84. Degree of smear microscopy: L4=1.20, L5=1.20, L6=0.92 and Gene Xpert Cq values: L4=17.08, L5=18.37, L6=17.59; showed no significant difference between the lineages, p=0.72 and p=0.48 respectively. Retrospective analysis of a larger sample confirmed the difference in TTP, p<0.001. In conclusion, the observed shorter TTP and high DCP of L4 could signify high growth rate in culture that is independent of total bacterial load at diagnosis.Publisher PDFPeer reviewe