Search

Background: Ethiopia is one of the high multidrug-resistant tuberculosis (MDR-TB) burden countries. However, phenotypic drug susceptibility testing can take several weeks due to the slow growth of Mycobacterium tuberculosis complex (MTBC) strains. In this study, we assessed the performance of a Sanger sequencing approach to predict resistance against five anti-tuberculosis drugs and the pattern of resistance mediating mutations., Methods: We enrolled 226 MTBC culture-positive MDR-TB suspects and collected sputum specimens and socio-demographic and TB related data from each suspect between June 2015 and December 2016 in Addis Ababa, Ethiopia. Phenotypic drug susceptibility testing (pDST) for rifampicin, isoniazid, pyrazinamide, ethambutol, and streptomycin using BACTEC MGIT 960 was compared with the results of a Sanger sequencing analysis of seven resistance determining regions in the genes rpoB, katG, fabG-inhA, pncA, embB, rpsL, and rrs., Result: DNA isolation for Sanger sequencing was successfully extracted from 92.5% (209/226) of the MTBC positive cultures, and the remaining 7.5% (17/226) strains were excluded from the final analysis. Based on pDST results, drug resistance proportions were as follows: isoniazid: 109/209 (52.2%), streptomycin: 93/209 (44.5%), rifampicin: 88/209 (42.1%), ethambutol: 74/209 (35.4%), and pyrazinamide: 69/209 (33.0%). Resistance against isoniazid was mainly mediated by the mutation katG S315T (97/209, 46.4%) and resistance against rifampicin by rpoB S531L (58/209, 27.8%). The dominating resistance-conferring mutations for ethambutol, streptomycin, and pyrazinamide affected codon 306 in embB (48/209, 21.1%), codon 88 in rpsL (43/209, 20.6%), and codon 65 in pncA (19/209, 9.1%), respectively. We observed a high agreement between phenotypic and genotypic DST, such as 89.9% (at 95% confidence interval [CI], 84.2%-95.8%) for isoniazid, 95.5% (95% CI, 91.2%-99.8%) for rifampicin, 98.6% (95% CI, 95.9-100%) for ethambutol, 91.3% (95% CI, 84.6-98.1%) for pyrazinamide and 57.0% (95% CI, 46.9%-67.1%) for streptomycin., Conclusion: We detected canonical mutations implicated in resistance to rifampicin, isoniazid, pyrazinamide, ethambutol, and streptomycin. High agreement with phenotypic DST results for all drugs renders Sanger sequencing promising to be performed as a complementary measure to routine phenotypic DST in Ethiopia. Sanger sequencing directly from sputum may accelerate accurate clinical decision-making in the future., Competing Interests: The authors declare that they have no competing interests.

Background: Previous studies suggest the burden of pulmonary tuberculosis (PTB) in Ethiopia may be greater in university students relative to the overall population. However, little is known about the transmission dynamics of PTB among students and members of the communities surrounding university campuses in Eastern Ethiopia., Methods: A cross sectional study was conducted in Eastern Ethiopia among prevalent culture-confirmed PTB cases from university students (n = 36) and community members diagnosed at one of four hospitals (n = 152) serving the surrounding area. Drug susceptibility testing (DST) was performed on Mycobacterium tuberculosis complex (MTBC) isolates using BD Bactec MGIT 960 and molecular genotyping was performed using spoligotyping and 24-loci MIRU-VNTR. MTBC strains with Identical genotyping patterns were assigned to molecular clusters as surrogate marker for recent transmission and further contact tracing was initiated among clustered patients., Results: Among all study participants, four MTBC lineages and 11 sub-lineages were identified, with Ethiopia&#95;3 (Euro-American lineage) being most common sub-lineage (29.4%) in both cohorts and associated with strain clustering (P = 0.016). We further identified 13 (8.1%) strains phylogenetically closely related to Ethiopia&#95;3 but with a distinct Spoligotyping pattern and designated as Ethiopia&#95;4. The clustering rate of MTBC strains was 52.9% for university students and 66.7% for community members with a Recent Transmission Index (RTI) of 17.6% and 48.4%, respectively. Female gender, urban residence, and new TB cases were significantly associated with strain clustering (P<0.05). Forty-eight (30%) of the study participants were resistant to one or more first line anti TB drugs, three patients were classified as multidrug resistant (MDR)., Conclusion: We found evidence for recent transmission of PTB among Ethiopian university students and the local community in Eastern Ethiopia, mainly linked to strains classified as Ethiopia&#95;3 sub lineage. Drug resistance didn't have a major impact on recent transmission but comprehensive molecular surveillance in combination with drug resistance profiling of MTBC strains is desirable to better characterize TB transmission dynamics in high risk congregate living environments such as university campuses and guide regional TB control programs., Competing Interests: The authors have declared that no competing interests exist.

Background: Recently, newly defined clades of Mycobacterium tuberculosis complex (MTBC) strains, namely Ethiopia 1-3 and Ethiopia H37Rv-like strains, and other clades associated with pulmonary TB (PTB) were identified in Ethiopia. In this study, we investigated whether these new strain types exhibit an increased ability to cause TB lymphadenitis (TBLN) and raised the question, if particular MTBC strains derived from TBLN patients in northern Ethiopia are genetically adapted to their local hosts and/or to the TBLN., Methods: Genotyping of 196 MTBC strains isolated from TBLN patients was performed by spoligotyping and 24-loci mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTR) typing. A statistical analysis was carried out to see possible associations between patient characteristics and phylogenetic MTBC strain classification., Results: Among 196 isolates, the majority of strains belonged to the Delhi/CAS (38.8%) lineage, followed by Ethiopia 1 (9.7%), Ethiopia 3 (8.7%), Ethiopia H37RV-like (8.2%), Ethiopia 2 and Haarlem (7.7% each), URAL (3.6%), Uganda l and LAM (2% each), S-type (1.5%), X-type (1%), and 0.5% isolates of TUR, EAI, and Beijing genotype, respectively. Overall, 15 strains (7.7%) could not be allocated to a previously described phylogenetic lineage. The distribution of MTBC lineages is similar to that found in studies of PTB samples. The cluster rate (35%) in this study is significantly lower (P = 0.035) compared to 45% in the study of PTB in northwestern Ethiopia., Conclusion: In the studied area, lymph node samples are dominated by Dehli/CAS genotype strains and strains of largely not yet defined clades based on MIRU-VNTR 24-loci nomenclature. We found no indication that strains of particular genotypes are specifically associated with TBLN. However, a detailed analysis of specific genetic variants of the locally contained Ethiopian clades by whole genome sequencing may reveal new insights into the host-pathogen co-evolution and specific features that are related to the local host immune system.

Background: Although Ethiopia ranks seventh among the world's 22 high-burden tuberculosis (TB) countries, little is known about strain diversity and transmission. In this study, we present the first in-depth analysis of the population structure and transmission dynamics of Mycobacterium tuberculosis strains from Northwest Ethiopia., Methods: In the present study, 244 M. tuberculosis isolates where analysed by mycobacterial interspersed repetitive unit - variable number tandem repeat 24-loci typing and spoligotyping methods to determine phylogenetic lineages and perform cluster analysis. Clusters of strains with identical genotyping patterns were considered as an indicator for the recent transmission., Results: Of 244 isolates, 59.0% were classified into nine previously described lineages: Dehli/CAS (38.9%), Haarlem (8.6%), Ural (3.3%), LAM (3.3%), TUR (2.0%), X-type (1.2%), S-type (0.8%), Beijing (0.4%) and Uganda II (0.4%). Interestingly, 31.6% of the strains were grouped into four new lineages and were named as Ethiopia&#95;3 (13.1%), Ethiopia&#95;1 (7.8%), Ethiopia&#95;H37Rv like (7.0%) and Ethiopia&#95;2 (3.7%) lineages. The remaining 9.4% of the isolates could not be assigned to the known or new lineages. Overall, 45.1% of the isolates were grouped in clusters, indicating a high rate of recent transmission., Conclusions: This study confirms a highly diverse M. tuberculosis population structure, the presence of new phylogenetic lineages and a predominance of the Dehli/CAS lineage in Northwest Ethiopia. The high rate of recent transmission indicates defects of the TB control program in Northwest Ethiopia. This emphasizes the importance of strengthening laboratory diagnosis of TB, intensified case finding and treatment of TB patients to interrupt the chain of transmission.