This, to the best of our knowledge, is the first report describing the species and strain diversity of M. tuberculosis complex isolates from TB patients in Rwanda. Characterization of prevailing M. tuberculosis strains focusing on different geographical levels is important for locating the origin, evolution and spread dynamics of particular M. tuberculosis clones, which is often difficult to be identified by traditional epidemiological investigations. In low-resource, high-disease burden settings, it is critical to identify circulating strains in order to understand the dynamics of spread of the causative agent. In Rwanda, there is no data about the species and strains of M. tuberculosis circulating in the country. This report, therefore, will provide baseline data for future country-wide molecular epidemiological studies to understand transmission dynamics of TB.
Regions of Difference (RD) analysis using 16S-rRNA, RD9 and TbD1 loci showed that all the strains investigated were characterized by presence of both 16S-rRNA and RD9 loci, and deletion in the TbD1 regions, a pattern confirming that they all were M. tuberculosis
strict sense. Most studies in the East African region have reported predominance of M. tuberculosis
], while most M. africanum
strains isolated to date are from West Africa [2
A majority (68.2%) of the spoligotypes obtained in this study belong to previously identified shared spoligotype international types (SITs). A significant proportion of the total isolates (48/151, 31.8%) belonged to SIT 52, while only 8/151 (5.3%) were SIT 135, a strain type commonly seen in Uganda. SIT 52 was found to be 7.6% (26/344) of isolates in a study in Central Uganda [18
] and 4.8% (6/125) of isolates from South Western Uganda [30
], while not a single strain of this type was seen in a collection of 130 isolates from Northern Tanzania [31
]. Generally, this genotype together with the related SIT 135 and SIT 128 are known to be the commonest strain types causing TB in Central African human host populations [4
The 151 isolates in the study show 53 different spoligopatterns, displaying a wide diversity of the spoligotypes in this collection. It is known that the structure of the TB populations is determined by geography, demography, and human migration. The large diversity of strains observed in this study may be attributed to increased transborder human movement in this region due to a large influx of former refugees from different neighboring countries in the last 15 years. Additionally, true orphan spoligotypes accounted for only 17% of all the spoligotypes in this study, this low percentage further supporting the hypothesis of increased recent human traffic in this setting, since countries with a history of isolation have been shown to have a large number of new spoligotypes, which is not the case in this scenario [32
Spoligotyping identified T2 to be the most predominant family of strains in Rwanda, accounting for up to 55.0% (83/151) of the total sample (Figures
and ). Results from a previous molecular study of recurrent TB in Rwanda by spoligotyping and mycobacterial interspersed repetitive unit variable number of tandem repeat (MIRU-VNTR) typing did not show species and strain types in the collection [16
] hence we cannot compare the two studies. Findings from the current study, however, are in agreement with the previous data from Uganda, in which two studies showed predominance of T2 family, the first having been conducted at the National referral hospital, Kampala, in which 67% of the isolates were T2 [33
] and the second a systematic community based study in Rubaga, one of the divisions of Kampala, which reported 70% isolates being of the T2 family [18
]. This result is in further agreement with those elsewhere reporting predominance of single genotypes in the respective populations across Africa [2
]. Collectively, these results depict a tendency for local genotypes that are well established to form a larger proportion of circulating strains compared to others as previously postulated [3
]. Since our sample collection may not reflect a national picture, a future national survey could genotype all isolates so as to give a clear situation of strain types as well as transmission pattern in this locale.
In Rwanda, the most recent national anti-tuberculosis drug resistance survey (2002–2005) on 616 new cases [14
] showed that 6.2% of the isolates were resistant to isoniazid, 3.9% to rifampicin and 3.9% were multi-drug resistant. In neighboring Uganda, a much earlier national survey (1996–1997) showed that of 586 patients, resistance to isoniazid was 6.7 %, that to rifampicin was 0.8% while MDR was 0.5% [36
]. The current study tested 46 new TB case, 94 retreatment cases and 11 cases with no known history. Overall MDR was 42.4%, a very high increase, most likely attributed to the high proportion of retreatment cases (94/115) in our study population as opposed to new TB cases in the previous studies.