Mollalign Hilina, Alemayehu Dawit Hailu, Beyene Dereje, Melaku Kalkidan, Ayele Abaysew, Chala Dawit, Diriba Getu, Yenew Bazezew, Getahun Muluwork, Adnew Bethlehem, Moga Shewki, Collins Jeffrey Michael, Ghodousi Arash, Bobosha Kidist, Wassie Liya
Ethiopian Public Health Institute.
Armauer Hansen Research Institute.
Res Sq. 2024 Dec 17:rs.3.rs-5302564. doi: 10.21203/rs.3.rs-5302564/v1.
Globally, drug-resistant tuberculosis (DR-TB) is responsible for 13% of mortality attributable to antimicrobial resistance. In Ethiopia, extrapulmonary tuberculosis (EPTB) is a significant public health challenge, and drug resistance (DR) in EPTB is often overlooked. In a cross-sectional study conducted between August 2022 and October 2023, we aimed to explore the magnitude of phenotypic drug resistance and identify genetic mutations linked to resistance using 189 Mycobacterium tuberculosis (MTB) isolates cultured from extrapulmonary clinical specimens. Additionally, we assessed the agreement of the phenotypic and whole genome sequencing (WGS) based genotypic drug resistance detection. We performed phenotypic drug sensitivity testing (pDST) using liquid culture BD BACTECTM MGITTM 960 system and WGS using Illumina NextSeq500/550. The genomic data analysis pipelines MTBSeq and TBProfiler were used to predict drug resistance-conferring mutations. The agreement between the pDST and WGS was analyzed using SPSS version 29.0 software. Our result demonstrated phenotypic resistance to at least one anti-TB drug was detected in 16.9% (32/189) of the study participants. Isoniazid-resistant rifampicin-susceptible-TB (Hr-TB) and multi-drug-resistant TB (MDR-TB) phenotypes accounted for 2.6% (5/189) and 4.2% (8/189) respectively. Prevalence of MDR-TB was 2.4% (4/170) among newly diagnosed and 21.1% (4/19) among previously treated cases. WGS identified more (14/160, 8.75%) rifampicin-resistant genotypes (RR-TB) compared to pDST (8/189, 4.2%). We have identified a putative compensatory mutation for rifampicin (rpoBSer450Leu, rpoCAsp747Ala) for the first time from an EPTB clinical specimen in Ethiopia. Overall, there was a 3.75% rifampicin mono-resistant-TB(RMR-TB) genotype, which remains undetected using the conventional pDST and represented 42.9% (6/14) of the identified RR-TB genotypes. Mutations conferring rifampicin resistance-interim (rpoB.Ser450Ala) represented the majority (83.3%) of RMR-TB. Changes in ethA genes associated with ethionamide resistance were the most common resistance (n=7, 87.5%) in MDR-TB cases. There was a strong agreement between the pDST and WGS-TB Profiler pipeline to detect RR-TB (kappa=0.8) compared to the MTBSeq pipeline (k=0.58). In conclusion, MDR-TB, Hr-TB, and interim-RMR-TB are equally important public health challenges in the realm of EPTB in Ethiopia. The role of WGS is tremendous in detecting borderline/interim RMR-TB, which will help for tailored, personalized treatment strategies.
在全球范围内，耐多药结核病（DR-TB）占抗菌药物耐药所致死亡率的13%。在埃塞俄比亚，肺外结核病（EPTB）是一项重大的公共卫生挑战，而EPTB中的耐药性（DR）常常被忽视。在2022年8月至2023年10月期间开展的一项横断面研究中，我们旨在探讨表型耐药的程度，并利用从肺外临床标本中培养出的189株结核分枝杆菌（MTB）分离株确定与耐药相关的基因突变。此外，我们评估了基于表型和全基因组测序（WGS）的基因型耐药检测的一致性。我们使用液体培养BD BACTECTM MGITTM 960系统进行表型药物敏感性试验（pDST），并使用Illumina NextSeq500/550进行WGS。基因组数据分析流程MTBSeq和TBProfiler用于预测赋予耐药性的突变。使用SPSS 29.0软件分析pDST和WGS之间的一致性。我们的结果表明，在16.9%（32/189）的研究参与者中检测到对至少一种抗结核药物的表型耐药。耐异烟肼利福平敏感结核病（Hr-TB）和耐多药结核病（MDR-TB）表型分别占2.6%（5/189）和4.2%（8/189）。新诊断病例中MDR-TB的患病率为2.4%（4/170），既往治疗病例中为21.1%（4/19）。与pDST（8/189，4.2%）相比，WGS鉴定出更多（14/160，8.75%）的利福平耐药基因型（RR-TB）。我们首次从埃塞俄比亚的一份EPTB临床标本中鉴定出利福平的一个推定补偿性突变（rpoBSer450Leu，rpoCAsp747Ala）。总体而言，存在3.75%的利福平单耐药结核病（RMR-TB）基因型，使用传统pDST未检测到，且占已鉴定RR-TB基因型的42.9%（6/14）。赋予利福平耐药性临时突变（rpoB.Ser450Ala）占RMR-TB的大多数（83.3%）。与乙硫异烟胺耐药相关的ethA基因变化是MDR-TB病例中最常见的耐药情况（n=7，87.5%）。与MTBSeq流程（κ=0.58）相比，pDST与WGS-TB Profiler流程在检测RR-TB方面具有很强的一致性（κ=0.8）。总之，MDR-TB、Hr-TB和临时RMR-TB在埃塞俄比亚EPTB领域同样是重要的公共卫生挑战。WGS在检测临界/临时RMR-TB方面发挥着巨大作用，这将有助于制定针对性的个性化治疗策略。
