Molekylær epidemiologi blant pasienter med multiresistent tuberkulose i Tigray, Etiopia

Tuberculosis (TB), which is caused by closely related Mycobacterium tuberculosis complex (MTBC) species, is an ancient human disease that gravely affects millions of people every year. TB is a preventable and treatable infectious disease. The continuing emergence and spread of multidrug-resistant tuberculosis (MDR-TB ) threaten the global TB control efforts. TB is the first killer among infectious diseases worldwide. According to the WHO report, there were an estimated 10.0 million incident cases, 1.4 million TB deaths, with more than 95% of these deaths in developing countries in 2019. Ethiopia is among the three high TB, TB/HIV, and MDR-TB burden countries. In 2019 in Ethiopia, there were 157,000 new TB cases, 1,400 MDR/RR-TB cases and 23,800 death from TB. This thesis aimed to describe the molecular epidemiology of multidrugresistant Mycobacterium tuberculosis among pulmonary TB patients in Tigray Region, Ethiopia. Three hundred sputum samples were collected from six hospitals of the Tigray Region between July 2018 and August 2019. The 227 samples culture positive for MTBC were subjected to drug susceptibility test to 1st- and 2nd- line anti-TB drugs using line probe assay. Among the 227 positive cultures, 74 samples were sequenced using whole-genome sequencing (WGS). WGS analysis showed diversified Mycobacterium tuberculosis genotypes circulating in the region, with L4 as the predominant genotype. The overall proportion of MDR-TB was high. The high proportion of MDR-TB among new and previously treated patients is alarming and calls for an urgent intervention to improve patient management. The high proportion of MDR-TB among newly diagnosed cases and the high level of recent transmission index indicates an ongoing transmission, which suggests the need for an enhanced TB control program performance to interrupt transmission. The study highlighted the usefulness of mutations at rpoB, katG, embB, rpsL, pncA, ethA, gyrA and rrs genes as a molecular marker for the rapid detection of resistance to RIF, INH, ETB, SM, PZA, ETH, FLQs and injectable 2nd-line anti-TB drugs, respectively. Besides the canonical mutations, a significant number of disputed rpoB mutations were also reported. Overall, the regional TB control program should be strengthened to detect and provide appropriate early treatment and follow-up for drug-resistant TB (DR-TB) cases. Abiding by the five WHOrecommended priority actions for DR-TB management is necessary to reduce the current high MDR-TB burden in the study region. Periodic surveillance of drug-resistance conferring mutations, early diagnosis and treatment of TB, and scaling up of drug susceptibility testing facilities to prevent and control the transmission of DR-TB in the community is recommended.Tuberkulose (TB) forårsakes av nært beslektede arter innen Mycobacterium tuberculosiskomplekset (MTBC), og er en eldgammel, alvorlig sykdom hos mennesker som rammer millioner hvert år. TB er en smittsom sykdom som kan forebygges og behandles, men fremveksten og spredningen av multiresistent tuberkulose (MDR-TB) truer den globale bekjempelsesinnsatsen. TB er den smittsomme sykdommen som gir størst antall dødsfall i verden. WHO anslår at det i 2019 var anslagsvis 10,0 millioner tilfeller, 1,4 millioner TB dødsfall, med mer enn 95% av disse dødsfallene i utviklingsland. Etiopia er blant de tre land som er mest belastet, med høye tall av TB, TB / HIV og MDR-TB. I Etiopia var det i 2019 157.000 nye TB-tilfeller, 1400 MDR tilfeller og 23 800 dødsfall fra TB. Denne doktorgradens overordnede mål var å beskrive den molekylære epidemiologien til sjukdom forårsaket av multiresistent Mycobacterium tuberculosis blant lungepasienter i Tigray-regionen, Etiopia. Tre hundre sputumprøver ble samlet inn fra seks sykehus i Tigray-regionen mellom juli 2018 og august 2019. Av disse ble de 227 prøvene som var positive for MTBC, undersøkt med en følsomhetstest mot 1. og 2. linje anti-TB medisiner ved bruk av Line Probe Assay. Av de 227 positive kulturene ble 74 prøver sekvensert ved bruk av helgenomsekvensering (WGS). WGS-analysene viste at forskjellige MTBC-genotyper som sirkulerte i regionen, med L4 som den dominerende genotypen. Den høye andelen MDR-TB blant nye og tidligere behandlede pasienter er alarmerende og stiller store krav til en forbedret pasienthåndtering. Den høye andelen MDR-TB blant nylig diagnostiserte tilfeller og det høye nivået av nylig overførte infeksjoner indikerer en pågående overføring, og antyder behovet for en forbedret ytelse av TB-kontrollprogrammet for å avbryte overføringen. Studien fremhevet nytten av mutasjoner ved rpoB, katG, embB, rpsL, pncA, ethA, gyrA og rrs gener som en molekylær markør for rask påvisning av resistens mot RIF, INH, ETB, SM, PZA, ETH, FLQ og injiserbare andrelinje anti-TB medisiner. Samlet sett bør det regionale TB-kontrollprogrammet styrkes for å oppdage og gi tilpasset passende behandling og oppfølging av TB-tilfeller. Å identifisere TB-kontrollprogrammets begrensninger i regionen, og følge de fem WHO-anbefalte prioriterte tiltakene for styring av DRTB, er nødvendig for å redusere den nåværende høye MDR-TB-belastningen i studieområdet. Det anbefales periodisk overvåking av mutasjoner som gir antibiotikaresistens, tidlig diagnose og behandling av TB, og oppskalering av fasiliteter for testing av isolater for antibiotikafølsomhet for å forhindre og kontrollere overføring av DR-TB i samfunnet.Mekelle Universit

Molecular epidemiology of multidrug-resistant tuberculosis among pulmonary tuberculosis patients in Tigray Region, Ethiopia

Tuberculosis (TB), which is caused by closely related Mycobacterium tuberculosis complex (MTBC) species, is an ancient human disease that gravely affects millions of people every year. TB is a preventable and treatable infectious disease. The continuing emergence and spread of multidrug-resistant tuberculosis (MDR-TB ) threaten the global TB control efforts. TB is the first killer among infectious diseases worldwide. According to the WHO report, there were an estimated 10.0 million incident cases, 1.4 million TB deaths, with more than 95% of these deaths in developing countries in 2019. Ethiopia is among the three high TB, TB/HIV, and MDR-TB burden countries. In 2019 in Ethiopia, there were 157,000 new TB cases, 1,400 MDR/RR-TB cases and 23,800 death from TB. This thesis aimed to describe the molecular epidemiology of multidrugresistant Mycobacterium tuberculosis among pulmonary TB patients in Tigray Region, Ethiopia. Three hundred sputum samples were collected from six hospitals of the Tigray Region between July 2018 and August 2019. The 227 samples culture positive for MTBC were subjected to drug susceptibility test to 1st- and 2nd- line anti-TB drugs using line probe assay. Among the 227 positive cultures, 74 samples were sequenced using whole-genome sequencing (WGS). WGS analysis showed diversified Mycobacterium tuberculosis genotypes circulating in the region, with L4 as the predominant genotype. The overall proportion of MDR-TB was high. The high proportion of MDR-TB among new and previously treated patients is alarming and calls for an urgent intervention to improve patient management. The high proportion of MDR-TB among newly diagnosed cases and the high level of recent transmission index indicates an ongoing transmission, which suggests the need for an enhanced TB control program performance to interrupt transmission. The study highlighted the usefulness of mutations at rpoB, katG, embB, rpsL, pncA, ethA, gyrA and rrs genes as a molecular marker for the rapid detection of resistance to RIF, INH, ETB, SM, PZA, ETH, FLQs and injectable 2nd-line anti-TB drugs, respectively. Besides the canonical mutations, a significant number of disputed rpoB mutations were also reported. Overall, the regional TB control program should be strengthened to detect and provide appropriate early treatment and follow-up for drug-resistant TB (DR-TB) cases. Abiding by the five WHOrecommended priority actions for DR-TB management is necessary to reduce the current high MDR-TB burden in the study region. Periodic surveillance of drug-resistance conferring mutations, early diagnosis and treatment of TB, and scaling up of drug susceptibility testing facilities to prevent and control the transmission of DR-TB in the community is recommended

Characteristics of pulmonary multidrug-resistant tuberculosis patients in Tigray Region, Ethiopia: A cross-sectional study.

Background: Tuberculosis (TB) is among the top 10 causes of mortality and the first killer among infectious diseases worldwide. One of the factors fuelling the TB epidemic is the global rise of multidrug resistant TB (MDR-TB). The aim of this study was to determine the magnitude and factors associated with MDR-TB in the Tigray Region, Ethiopia. Method: This study employed a facility-based cross-sectional study design, which was conducted between July 2018 and August 2019. The inclusion criteria for the study participants were GeneXpert-positive who were not under treatment for TB, PTB patients' ≥15 years of age and who provided written informed consent. A total of 300 participants were enrolled in the study, with a structured questionnaire used to collect data on clinical, sociodemographic and behavioral factors. Sputum samples were collected and processed for acid-fast bacilli staining, culture and drug susceptibility testing. Drug susceptibility testing was performed using a line probe assay. Logistic regression was used to analyze associations between outcome and predictor variables. Results: The overall proportion of MDR-TB was 16.7% (11.6% and 32.7% for new and previously treated patients, respectively). Of the total MDR-TB isolates, 5.3% were pre-XDR-TB. The proportion of MDR-TB/HIV co-infection was 21.1%. A previous history of TB treatment AOR 3.75; 95% CI (0.7-2.24), cigarette smoking AOR 6.09; CI (1.65-2.50) and patients who had an intermittent fever (AOR = 2.54, 95% CI = 1.21-5.4) were strongly associated with MDR-TB development. Conclusions: The magnitude of MDR-TB observed among new and previously treated patients is very alarming, which calls for an urgent need for intervention. The high proportion of MDR-TB among newly diagnosed cases indicates ongoing transmission, which suggests the need for enhanced TB control program performance to interrupt transmission. The increased proportion of MDR-TB among previously treated cases indicates a need for better patient management to prevent the evolution of drug resistance. Assessing the TB control program performance gaps and an optimal implementation of the WHO recommended priority actions for the management of drug-resistant TB, is imperative to help reduce the current high MDR-TB burden in the study region

Whole Genome Sequencing of Drug Resistant and Drug Susceptible Mycobacterium tuberculosis Isolates From Tigray Region, Ethiopia

Background: Tuberculosis, mainly caused by Mycobacterium tuberculosis (Mtb), is an ancient human disease that gravely affects millions of people annually. We wanted to explore the genetic diversity and lineage-specific association of Mtb with drug resistance among pulmonary tuberculosis patients. Methods: Sputum samples were collected from pulmonary tuberculosis patients at six different healthcare institutions in Tigray, Ethiopia, between July 2018 and August 2019. DNA was extracted from 74 Mtb complex isolates for whole-genome sequencing (WGS). All genomes were typed and screened for mutations with known associations with antimicrobial resistance using in silico methods, and results were cross-verified with wet lab methods. Results: Lineage (L) 4 (55.8%) was predominant, followed by L3 (41.2%); L1 (1.5%) and L2 (1.5%) occurred rarely. The most frequently detected sublineage was CAS (38.2%), followed by Ural (29.4%), and Haarlem (11.8%). The recent transmission index (RTI) was relatively low. L4 and Ural strains were more resistant than the other strains to any anti-TB drug ( P < 0.05). The most frequent mutations to RIF, INH, EMB, SM, PZA, ETH, FLQs, and 2nd-line injectable drugs occurred at rpoB S450L, katG S315T, embB M306I/V, rps L K43R, pncA V139A, ethA M1R, gyr A D94G, and rrs A1401G, respectively. Disputed rpoB mutations were also shown in four (16%) of RIF-resistant isolates. Conclusion: Our WGS analysis revealed the presence of diverse Mtb genotypes. The presence of a significant proportion of disputed rpoB mutations highlighted the need to establish a WGS facility at the regional level to monitor drug-resistant mutations. This will help control the transmission of DR-TB and ultimately contribute to the attainment of 100% DST coverage for TB patients as per the End TB strategy

Frequency and patterns of first- and second-line drug resistance-conferring mutations in Mycobacterium tuberculosis isolated from pulmonary tuberculosis patients in a cross-sectional study in Tigray Region, Ethiopia

Objectives Tuberculosis (TB) is a preventable and treatable infectious disease, but the continuing emergence and spread of multidrug-resistant TB is threatening global TB control efforts. This study aimed to describe the frequency and patterns of drug resistance-conferring mutations of Mycobacterium tuberculosis (MTB) isolates detected from pulmonary TB patients in Tigray Region, Ethiopia. Methods A cross-sectional study design was employed to collect sputum samples from pulmonary TB patients between July 2018 to August 2019. Culture and identification tests were done at Tigray Health Research Institute (THRI). Mutations conferring rifampicin (RIF), isoniazid (INH) and fluoroquinolone (FQ) resistance were determined in 227 MTB isolates using GenoType MTBDRplus and GenoType MTBDRsl. Results Mutations conferring resistance to RIF, INH and FQs were detected in 40/227 (17.6%), 41/227 (18.1%) and 2/38 (5.3%) MTB isolates, respectively. The majority of mutations for RIF, INH and FQs occurred at codons rpoB S531L (70%), katG S315T (78%) and gyrA D94Y/N (100%), respectively. This study revealed a significant number of unknown mutations in the rpoB, katG and inhA genes. Conclusion High rates of mutations conferring resistance to RIF, INH and FQs were observed in this study. A large number of isolates showed unknown mutations, which require further DNA sequencing analysis. Periodic drug resistance surveillance and scaling-up of drug resistance testing facilities are imperative to prevent the transmission of drug-resistant TB in the community

Characteristics of pulmonary multidrug-resistant tuberculosis patients in Tigray Region, Ethiopia: A cross-sectional study.

BackgroundTuberculosis (TB) is among the top 10 causes of mortality and the first killer among infectious diseases worldwide. One of the factors fuelling the TB epidemic is the global rise of multidrug resistant TB (MDR-TB). The aim of this study was to determine the magnitude and factors associated with MDR-TB in the Tigray Region, Ethiopia.MethodThis study employed a facility-based cross-sectional study design, which was conducted between July 2018 and August 2019. The inclusion criteria for the study participants were GeneXpert-positive who were not under treatment for TB, PTB patients' ≥15 years of age and who provided written informed consent. A total of 300 participants were enrolled in the study, with a structured questionnaire used to collect data on clinical, sociodemographic and behavioral factors. Sputum samples were collected and processed for acid-fast bacilli staining, culture and drug susceptibility testing. Drug susceptibility testing was performed using a line probe assay. Logistic regression was used to analyze associations between outcome and predictor variables.ResultsThe overall proportion of MDR-TB was 16.7% (11.6% and 32.7% for new and previously treated patients, respectively). Of the total MDR-TB isolates, 5.3% were pre-XDR-TB. The proportion of MDR-TB/HIV co-infection was 21.1%. A previous history of TB treatment AOR 3.75; 95% CI (0.7-2.24), cigarette smoking AOR 6.09; CI (1.65-2.50) and patients who had an intermittent fever (AOR = 2.54, 95% CI = 1.21-5.4) were strongly associated with MDR-TB development.ConclusionsThe magnitude of MDR-TB observed among new and previously treated patients is very alarming, which calls for an urgent need for intervention. The high proportion of MDR-TB among newly diagnosed cases indicates ongoing transmission, which suggests the need for enhanced TB control program performance to interrupt transmission. The increased proportion of MDR-TB among previously treated cases indicates a need for better patient management to prevent the evolution of drug resistance. Assessing the TB control program performance gaps and an optimal implementation of the WHO recommended priority actions for the management of drug-resistant TB, is imperative to help reduce the current high MDR-TB burden in the study region