Profile and Frequency of Mutations Conferring Drug-Resistant Tuberculosis in the Central, Southeastern and Eastern Ethiopia.

Purpose: Advances in molecular tools that assess genes harboring drug resistance mutations have greatly improved the detection and treatment of drug-resistant tuberculosis (DR-TB). This study was conducted to determine the frequency and type of mutations that are responsible for resistance to rifampicin (RIF), isoniazid (INH), fluoroquinolones (FLQs) and second-line injectable drugs (SLIDs) in Mycobacterium tuberculosis (MTB) isolates obtained from culture-positive pulmonary tuberculosis (TB) patients in the central, southeastern and eastern Ethiopia. Patients and Methods: In total, 224 stored culture-positive MTB isolates from pulmonary TB patients referred to Adama and Harar regional TB laboratories between August 2018 and January 2019 were assessed for mutations conferring RIF, INH, FLQs and SLIDs resistance using GenoType®MTBDRplus (MTBDRplus) and GenoType®MTBDRsl (MTBDRsl). Results: RIF, INH, FLQs and SLIDs resistance-conferring mutations were identified in 88/224 (39.3%), 85/224 (38.0%), 7/77 (9.1%), and 3/77% (3.9%) of MTB isolates, respectively. Mutation codons rpoB S531L (59.1%) for RIF, katG S315T (96.5%) for INH, gyrA A90V (42.1%) for FLQs and WT1 rrs (100%) for SLIDs were observed in the majority of the isolates tested. Over a 10th of rpoB mutations detected in the current study were unknown. Conclusion: In this study, the most common mutations conferring drug resistance to RIF, INH, FLQs were identified. However, a significant proportion of RIF-resistant isolates manifested unknown rpoB mutations. Similarly, although few in number, all SLID-resistant isolates had unknown rrs mutations. To further elucidate the entire spectrum of mutations, tool such as whole-genome sequencing is imperative. Furthermore, the expansion of molecular drug susceptibility testing services is critical for tailoring patient treatment and preventing disease transmission.

Expanding molecular diagnostic capacity for COVID-19 in Ethiopia: operational implications, challenges and lessons learnt.

Efforts towards slowing down coronavirus (COVID-19) transmission and reducing mortality have focused on timely case detection, isolation and treatment. Availability of laboratory COVID-19 testing capacity using reverse-transcriptase polymerase chain reaction (RT-PCR) was essential for case detection. Hence, it was critical to establish and expand this capacity to test for COVID-19 in Ethiopia. To this end, using a three-phrased approach, potential public and private laboratories with RT-PCR technology were assessed, capacitated with trained human resource and equipped as required. These laboratories were verified to conduct COVID-19 testing with quality assurance checks regularly conducted. Within a 10-month period, COVID-19 testing laboratories increased from zero to 65 in all Regional States with the capacity to conduct 18,454 tests per day. The success of this rapid countrywide expansion of laboratory testing capacity for COVID-19 depended on some key operational implications: the strong laboratory coordination network within the country, the use of non-virologic laboratories, investment in capacity building, digitalization of the data for better information management and establishing quality assurance checks. A weak supply chain for laboratory reagents and consumables, differences in the brands of COVID-19 test kits, frequent breakdowns of the PCR machines and inadequate number of laboratory personnel following the adaption of a 24/7 work schedule were some of the challenges experienced during the process of laboratory expansion. Overall, we learn that multisectoral involvement of laboratories from non-health sectors, an effective supply chain system with an insight into the promotion of local production of laboratory supplies were critical during the laboratory expansion for COVID-19 testing. The consistent support from WHO and other implementing partners to Member States is needed in building the capacity of laboratories across different diagnostic capabilities in line with International Health Regulations. This will enable efficient adaptation to respond to future public health emergencies.

Factors affectinsg the utilization of Xpert MTB/RIF assay among TB clinic health workers in Addis Ababa.

INTRODUCTION: The diagnostic accuracy of Xpert MTB/RIF is well documented but underutilization is a major challenge in most high burden countries. This appears to be linked with insufficient knowledge of health professionals of using the tool. However, this has not been well studied. OBJECTIVE: Our objective was to assess the knowledge of health professionals on Xpert MTB/RIF assay and associated factors in detecting TB/TB drug resistance. METHODS: An institution based cross-sectional study was conducted from April 4 to June 5, 2015, in Addis Ababa that involved 209 healthcare providers working in TB clinics. Structured questionnaire through self-administered interview technique was used to collect the data. We asked them about Xpert on whether they are aware of its place in TB diagnosis, when and for whom it shall be used, its role in treatment monitoring, result interpretation and patient's registration that are diagnosed by Xpert MTB/RIF. We used binary logistic regression analysis to identify associated factors. Odds ratio with 95% CI was computed to assess the strength of the associations. RESULTS: Of the 209 participants interviewed, the majority 151 (72.2%) were nurses. More than a half of the respondents 114 (54.6%) had poor knowledge. Health professionals with age above 35 years (AOR = 6.253, 95% CI (1.1995, 19.604)) and those who read the Xpert guideline (AOR = 4.231, 95% CI (2.011, 8.900)) were more likely to have good knowledge on Xpert. CONCLUSION AND RECOMMENDATION: This study revealed that the overall magnitude of knowledge status was found to be low. Health workers above 35 years and those who read the guideline on Xpert had higher knowledge status on Xpert. Distribution of national guideline on Xpert and assigning experienced clinicians in TB DOTs clinics are recommended.

Performance evaluation of tuberculosis smear microscopists working at rechecking laboratories in Ethiopia.

BACKGROUND: Tuberculosis is an infectious disease caused by the bacillus Mycobacterium tuberculosis. According to the Ethiopian Federal Ministry of Health's 2013-2014 report, the tuberculosis case detection rate was 53.7%, which was below the target of 81% set for that year. OBJECTIVE: This study assessed the performance of tuberculosis smear microscopists at external quality assessment rechecking laboratories in Ethiopia. METHODS: A cross-sectional study was conducted at 81 laboratories from April to July 2015. Panel slides were prepared and validated at the National Tuberculosis Reference Laboratory. The validated panel slides were used to evaluate the performance of microscopists at these laboratories compared with readers from the reference laboratory. RESULTS: A total of 389 external quality assessment rechecking laboratory microscopists participated in the study, of which 268 (68.9%) worked at hospitals, 241 (62%) had more than five years of work experience, 201 (51.7%) held Bachelors degrees, and 319 (82%) reported tuberculosis smear microscopy training. Overall, 324 (83.3%) participants scored ≥ 80%. Sensitivity for detecting tuberculosis bacilli was 84.5% and specificity was 93.1%. The overall percent agreement between participants and reference readers was 87.1 (kappa=0.72). All 10 slides were correctly read (i.e., scored 100%) by 80 (20.6%) participants, 156 (40.1%) scored 90% - 95%, 88 (22.6%) scored 80% - 85% and 65 (16.7%) scored below 80%. There were 806 (20.7%) total errors, with 143 (3.7%) major and 663 (17%) minor errors. CONCLUSION: The overall performance of participants in reading the slides showed good agreement with the reference readers. Most errors were minor, and the ability to detect tuberculosis bacilli can be improved through building the capacity of professionals.

Performance evaluation of malaria microscopists working at rechecking laboratories in Ethiopia.

BACKGROUND: Microscopic diagnosis of Giemsa-stained thick and thin blood films has remained the standard laboratory method for diagnosing malaria. High quality performance of microscopists that examine blood slides in health facilities remains critically important. MATERIALS AND METHODS: A cross-sectional study was conducted to assess the performance of 107 malaria microscopists working at 23 malaria rechecking laboratories in Ethiopia. A set of 12 blood film slides was distributed to each microscopist. Data was collected and exported to SPSS version 20 for analysis. Chi-square, sensitivity, specificity, percent agreement, and kappa scores were calculated to assess performance in detecting and identification of Plasmodium species. RESULTS: The mean age of the participants was 30 ± 5 yrs and most of them (54; 50.5%) were working at regional reference laboratories. Overall, the sensitivity of participants in detecting and identifying malaria parasite species was 96.8% and 56.7%, respectively. The overall agreement on detection and identification of malaria species was 96.8% (Kappa = 0.9) and 64.8% (Kappa = 0.33), respectively. The least accurately identified malaria parasite species was P. malariae (3/107; 2.8%) followed by P. ovale (35/107; 32.7%). Participants working at hospital laboratories had the highest percentage (72.3 %, Kappa=0.51) of accurate species identification. Study participants that had participated in malaria microscopy and quality assurance trainings were significantly better at quantifying parasite densities (P<0.001). CONCLUSION: The accuracy of parasite identification and quantification differed strongly between participants and expert microscopists. Therefore, regular competency assessment and training for malaria microscopists should be mandatory to assure proper diagnosis and management of malaria in Ethiopia.

Molecular typing and drug sensitivity testing of Mycobacterium tuberculosis isolated by a community-based survey in Ethiopia.

BACKGROUND: The identification of circulating TB strains in the community and drug sensitivity patterns is essential for the tuberculosis control program. This study was undertaken to identify M. tuberculosis strains circulating in selected communities in Ethiopia as well as to evaluate the drug sensitivity pattern of these strains. METHOD: This study was a continuation of the Ethiopian National TB Prevalence Survey that was conducted between 2010 and 2011. Culture-positive isolates of M. tuberculosis from previous study were typed using region of difference (RD) 9-based polymerase chain reaction (PCR) and spoligotyping. Drug sensitivity testing was conducted using the indirect proportion method on Lowenstein-Jensen media. RESULT: All 92 isolates were confirmed as M. tuberculosis by RD9-based PCR and spoligotyping of 91 of these isolates leds to the identification of 41 spoligotype patterns. Spoligotype revealed higher diversity (45 %) and among this 65.8 % (27/41) were not previously reported. The strains were grouped into 14 clusters consisting of 2-15 isolates. The dominant strains were SIT53, SIT149 and SIT37 consisting of 15, 11, and 9 isolates, respectively. Our study reveals 70 % (64/91) clustered strains and only 39.1 % (25/64) occurred within the same Kebele. Further assignment of the strains to the lineages showed that 74.7 % (68/91) belonged to Euro-American lineage, 18.6 % (17/91) to East Africa Indian lineage and the remaining 6.5 % (6/91) belonged to Indo-oceanic lineage. Valid drug susceptibility test results were available for 90 of the 92 isolates. Mono-resistance was observed in 27.7 % (25/90) and poly-resistance in 5.5 % (5/90) of the isolates. Moreover, multi-drug resistance (MDR-TB) was detected in 4.4 % of the isolates whilst the rest (60/90) were susceptible to all drugs. The highest level of mono-resistance, 26.6 % (24/90), was observed for streptomycin with majority (91.1 %) of streptomycin mono-resistant strains belonging to the Euro-American lineage. CONCLUSION: In this study, the strains of M. tuberculosis circulating in selected sites of Ethiopia were identified along with the drug sensitivity patterns. Thus, these findings are useful for the TB Control Program of the country.

Evaluation of microscopic observation drug susceptibility assay for detection of multidrug-resistant Mycobacterium tuberculosis.

Early detection of multidrug-resistant Mycobacterium tuberculosis (MDR-TB) is of primary importance for both patient management and infection control. Optimal methods for identifying drug-resistant Mycobacterium tuberculosis in a timely and affordable way in resource-limited settings are not yet available. This study prospectively evaluated a low-technology but rapid drug susceptibility testing method, the microscopic observation drug susceptibility assay (MODS), in the concurrent detection of M. tuberculosis and its susceptibilities to isoniazid and rifampin (two drugs defining multidrug-resistant M. tuberculosis) directly from sputum specimens. Sputum samples were collected from 262 smear-positive TB patients in Addis Ababa, Ethiopia. To undertake MODS, 100 mul of decontaminated samples was inoculated into a 24-well plate containing 1 ml of 7H9 broth with and without appropriate drugs. The assay uses an inverted-light microscope to detect characteristic mycobacterial growth in liquid culture. Of 262 smear-positive patients, MODS detected 254 (96.9%) and culture in Löwenstein-Jensen medium detected 247 (94.3%) (P = 0.016). For the 247 cultures, the sensitivity, specificity, and accuracy of MODS for detecting MDR-TB were 92.0, 99.5, and 98.8%, respectively, using the method of proportion as a reference (concordance, 98.8%; kappa value, 0.932). Results for MODS were obtained in a median time of 9 days. MODS is an optimal alternative method for identifying MDR-TB in a timely and affordable way in resource-limited settings.