Rifampicin-resistant Mycobacterium Tuberculosis Research Articles

Diagnostic approaches and prevalence of Rifampicin resistant Mycobacterium tuberculosis in District Mardan

The disease tuberculosis (TB) caused by Mycobacterium Tuberculosis (MTB) is most common infectious disease in developing countries. The disease is fatal if not treated during the early stages of infection, thereby early and precise detection is a decisive step in curing the disease. The aim of this study was to analyze the prevalence of tuberculosis in patients reporting to Mardan Medical Complex (MMC), located in the district Mardan, KPK, Pakistan. The sputum of patients was analysed by Ziehl-Nilsen (ZN) staining technique followed by light microscopy called Acid-Fast Bacillus (AFB) staining. The sputum samples were collected from the patients and analysed by special PCR method called GeneXpert MTB/RIF assay, for genomic detection and resistance assay for rifampicin antibiotic were used, are the commonly used medicine for the treatment of MTB infection. Total 121 patients reported to MMC, represented 74 % patients from Mardan, 12% from Nowshera and 14% from Swabi. These patients were screened for the aim to evaluate the techniques for the detection of MTB. The light microscopy method confirmed 66 (55%) of the patients positive for MTB, whereas the same samples reported 78 (68%) patients positive for MTB through GeneXpert MTB/RIF assay, The Positive Predictive Value (PPV) and Negative Predictive Value (NPV) found for light microscopy were 99% and 78.1% respectively. The most used drug rifampicin was found ineffective in 9 patients (7%). Additionally, 83% of the patients when interviewed had a folk history of tuberculosis. Keywords: Microscopy, PCR, Mycobacterium tuberculosis, Rifampicin.

The Magnitude of MTB and Rifampicin Resistance MTB Using Xpert-MTB/RIF Assay Among Tuberculosis Suspected Patients in Gedeo Zone, Southern Ethiopia.

BackgroundTuberculosis (TB) remains a major global health problem causing death among millions of people each year. The new barrier that challenges the control of tuberculosis is the emerging and the increasing number of drug-resistant TB that becomes a world concern. This study aimed to determine the magnitude of rifampicin-resistant Mycobacterium tuberculosis (RR-MTB) among presumptive TB patients attending Dilla University Referral Hospital, Gedeo Zone, Ethiopia.MethodsA retrospective cross-sectional study was conducted at Dilla University Referral Hospital from January 2014 to December 2020. Sputum results were done using Xpert MTB/RIF assay and other necessary data were collected from the registration logbooks using a standardized data extraction format and analyzed using SPSS version 23 statistical software.ResultsA total of 17,745 presumptive TB patients were included, of which 62.2% were males. The overall prevalence of Mycobacterium tuberculosis (MTB) was 11.8%, of which 5.1% were confirmed to have RR-MTB. Extra-pulmonary TB was reported in 1.5% of the study participants. The highest prevalence of MTB and RR-MTB was recorded in 2017 with a prevalence of 20.1% and 8.5%, respectively. All age groups were significantly associated with a higher prevalence of MTB (p < 0.036). TB patients with a history of previous treatment and HIV positive were significantly associated with MTB (P < 0.021), while RR-MTB was only significantly associated with patients with a history of previous treatment (P < 0.018).ConclusionA high magnitude of MTB and RR-MTB was reported among TB patients with HIV and a history of previous treatment. Therefore, coordinated efforts should be applied to the improvement of treatment adherence of known TB cases, and appropriate control and prevention methods to reduce the emergence and increase of MTB and RR-MTB cases.

Transmission patterns of rifampicin resistant Mycobacterium tuberculosis complex strains in Cameroon: a genomic epidemiological study

BackgroundDetermining factors affecting the transmission of rifampicin (RR) and multidrug-resistant (MDR) Mycobacterium tuberculosis complex strains under standardized tuberculosis (TB) treatment is key to control TB and prevent the evolution of drug resistance.MethodsWe combined bacterial whole genome sequencing (WGS) and epidemiological investigations for 37% (n = 195) of all RR/MDR-TB patients in Cameroon (2012–2015) to identify factors associated with recent transmission.ResultsPatients infected with a strain resistant to high-dose isoniazid, and ethambutol had 7.4 (95% CI 2.6–21.4), and 2.4 (95% CI 1.2–4.8) times increased odds of being in a WGS-cluster, a surrogate for recent transmission. Furthermore, age between 30 and 50 was positively correlated with recent transmission (adjusted OR 3.8, 95% CI 1.3–11.4). We found high drug-resistance proportions against three drugs used in the short standardized MDR-TB regimen in Cameroon, i.e. high-dose isoniazid (77.4%), ethambutol (56.9%), and pyrazinamide (43.1%). Virtually all strains were susceptible to fluoroquinolones, kanamycin, and clofazimine, and treatment outcomes were mostly favourable (87.5%).ConclusionPre-existing resistance to high-dose isoniazid, and ethambutol is associated with recent transmission of RR/MDR strains in our study. A possible contributing factor for this observation is the absence of universal drug susceptibility testing in Cameroon, likely resulting in prolonged exposure of new RR/MDR-TB patients to sub-optimal or failing first-line drug regimens.

Characterization of rifampicin-resistant Mycobacterium tuberculosis in Khyber Pakhtunkhwa, Pakistan

Tuberculosis (TB), caused by Mycobacterium tuberculosis, is endemic in Pakistan. Resistance to both firstline rifampicin and isoniazid drugs (multidrug-resistant TB; MDR-TB) is hampering disease control. Rifampicin resistance is attributed to rpoB gene mutations, but rpoA and rpoC loci may also be involved. To characterise underlying rifampicin resistance mutations in the TB endemic province of Khyber Pakhtunkhwa, we sequenced 51 M. tuberculosis isolates collected between 2016 and 2019; predominantly, MDR-TB (n = 44; 86.3%) and lineage 3 (n = 30, 58.8%) strains. We found that known mutations in rpoB (e.g. S405L), katG (e.g. S315T), or inhA promoter loci explain the MDR-TB. There were 24 unique mutations in rpoA, rpoB, and rpoC genes, including four previously unreported. Five instances of within-host resistance diversity were observed, where two were a mixture of MDR-TB strains containing mutations in rpoB, katG, and the inhA promoter region, as well as compensatory mutations in rpoC. Heteroresistance was observed in two isolates with a single lineage. Such complexity may reflect the high transmission nature of the Khyber Pakhtunkhwa setting. Our study reinforces the need to apply sequencing approaches to capture the full-extent of MDR-TB genetic diversity, to understand transmission, and to inform TB control activities in the highly endemic setting of Pakistan.

Rifampicin-Resistant Disseminated Tuberculosis in an Immunocompetent Adolescent Male Presenting with Retropharyngeal Abscess and Spinal Involvement

Retropharyngeal abscess is a rare manifestation in spinal tuberculosis. Early clinical diagnosis followed by microbiological confirmation and effective treatment is crucial to avoid irreversible damage to the spine. Here, we report a case of disseminated tuberculosis in an immunocompetent adolescent male who presented with retropharyngeal abscess, multifocal involvement of the spine, and skin tuberculids. Xpert MTB/RIF assay in this patient facilitated early lifesaving treatment by detecting rifampicin-resistant Mycobacterium tuberculosis (MTB) in the clinical specimen.

Profiling and identification of novel rpoB mutations in rifampicin-resistant Mycobacterium tuberculosis clinical isolates from Pakistan

IntroductionRifampicin (RIF) is one of the most effective anti-tuberculosis first-line drugs prescribed along with isoniazid. However, the emergence of RIF resistance Mycobacterium tuberculosis (MTB) isolates is a major issue towards tuberculosis (TB) control program in high MDR TB-burdened countries including Pakistan. Molecular data behind phenotypic resistance is essential for better management of RIF resistance which has been linked with mutations in rpoB gene. Since molecular studies on RIF resistance is limited in Pakistan, the current study was aimed to investigate the molecular data of mutations in rpoB gene behind phenotypic RIF resistance isolates in Pakistan. MethodA total of 322 phenotypically RIF-resistant isolates were randomly selected from National TB Reference Laboratory, Pakistan for sequencing while 380 RIF resistance whole-genome sequencing (WGS) of Pakistani isolates (BioProject PRJEB25972), were also analyzed for rpoB mutations. ResultAmong the 702 RIF resistance samples, 675 (96.1%) isolates harbored mutations in rpoB in which 663 (94.4%) were detected within the Rifampicin Resistance Determining Region (RRDR) also known as a mutation hot spot region, including three novel. Among these mutations, 657 (97.3%) were substitutions including 603 (89.3%) single nucleotide polymorphism, 49 (7.25%) double and five (0.8%) triple. About 94.4% of Phenotypic RIF resistance strains, exhibited mutations in RRDR, which were also detectable by GeneXpert. ConclusionMutations in the RRDR region of rpoB is a major mechanism of RIF resistance in MTB circulating isolates in Pakistan. Molecular detection of drug resistance is a faster and better approach than phenotypic drug susceptibility testing to reduce the time for transmission of RIF resistance strains in population. Such insights will inform the deployment of anti-TB drug regimens and disease control tools and strategies in high burden settings, such as Pakistan.

Rifampicin resistance in Mycobacterium tuberculosis in Iran: a two-centre study

Multidrug-resistant tuberculosis remains a challenge. In this study, we investigated the incidence of rifampicin (RIF) resistance in Mycobacterium tuberculosis in a large number of pulmonary specimens.A two-center study in Tehran, the capital of Iran, was performed with 6624 pulmonary samples of patients with tuberculosis (TB) who were subjected to detection of RIF-resistant TB by GeneXpert MTB/RIF assay between May 2014 and July 2018. Conventional drug susceptibility testing was performed to confirm the results.Xpert MTB/RIF identified a total of 96 positives for M. tuberculosis, of which 5 (5.3%) samples were found to be RIF-resistant TB. All RIF-resistant and sensitive isolates detected by GeneXpert were phenotypically confirmed by drug susceptibility testing.These results indicated that the Xpert MTB/RIF test can be used as a rapid diagnostic method and can potentially decrease the morbidity associated with diagnostic delay and mistreatment.

Assessment of the GenoType MTBDRsl VER 2.0 compared to the phenotypic drug susceptibility testing and whole genome sequencing for the rapid detection of resistance to fluoroquinolone and second-line injectable drugs among rifampicin-resistant Mycobacterium tuberculosis isolates.

Molecular techniques have considerable advantages for rapid detection, a reduction of infectiousness, prevention of further resistance development and surveillance of drug-resistant TB. MTBDRsl VER 2.0 was used to detect resistance to second-line anti-tuberculosis drugs on 35 rifampicin-resistant M. tuberculosis (RR-MTB) isolates compared to the minimum inhibitory concentrations (MICs) and whole genome sequencing (WGS). The MTBDRsl VER 2.0 (Hain Life Science, Nehren, Germany) and WGS (San Diego, CA, USA) were performed for tracing mutations in resistant-related genes involved in resistance to fluoroquinolone (FLQ) and second-line injectable drugs. The broth microdilution method using 7H9 Middlebrook media supplemented with OADC was used to determine the MICs. The MTBDRsl VER 2.0 correctly detected 5/6 (83.3%) of FLQ-resistant strains. The MUT1 A1401G (seven strains) and MUT2 G1484T (one strain) mutations in rrs gene were detected in eight AMK/KAN/CAP-resistant strains. Four low-level KAN-resistant strains with the G-10A/C-12T (three strains) and eis C-14T (one strain) mutations in eis gene was diagnosed using MTBDRsl VER 2.0. Five errors were found in detecting resistance to kanamycin and capreomycin compared to the phenotypic drug susceptibility testing and WGS. Failling wild-type bands without improved mutant bands did not indicate a reliable resistance. WGS could efficiently resolve the discrepancies of the results. MTBDRsl showed better performance in detecting XDR strains than pre-XDR.

Prevalence of Mycobacterium tuberculosis (Rifampicin-Resistant MTB) and Associated Risk Actors Among Pulmonary Presumptive TB Patients in Eastern Amhara, Ethiopia: 2015-2019.

IntroductionTuberculosis (TB) is a major health problem, mainly in resource-limited settings. The aim of this study was to determine the prevalence of TB and rifampicin-resistant Mycobacterium tuberculosis (RR-MTB) among presumptive tuberculosis patients using Xpert MTB/RIF assay in Eastern Amhara, Ethiopia.MethodsA retrospective cross-sectional study was conducted among presumptive TB patients from three governmental hospitals in Amhara Regional State. Records of sputum sample results using Xpert MTB/RIF assay from January 2015 to December 2019 were investigated from registration books and analyzed using SPSS v.21.ResultsOf the total of 26,656 (24,116 adults and 2540 children) TB presumptive patients included in the study, more than half, 14,624 (54.9%), were males and the median age was 36.87 (interquartile: 25.46–50.85 years). The majority of participants were new cases, 20,273 (76.1%), and with unknown HIV status, 18,981 (71.2%), respectively. MTB prevalence was 11% (95% CI: 9.34–12.08%) in all age groups, and 7.6% (95% CI 6.52–9.04%) among children. Of the MTB confirmed cases, prevalence of RR-MTB was 245 (8.3%) in adults and 14 (7.2%) in children. MTB infection was higher in the age groups of 18–35 years [adjusted odds ratio (AOR) = 2.17; 95% CI: 1.86–2.54, p < 0.001], 36–53 years (AOR = 1.31; 95% CI 1.11–1.54, p < 0.001), those who were relapse cases (AOR = 1.97; 95% CI 1.69–2.27, p < 0.0010), and failure cases (AOR = 4.67; 95% CI 3.36–6.50, p < 0.001). However, the age groups of 54–71 years (AOR = 0.79; 95% CI 0.65–0.95, p = 0.01) and over 71 years (AOR = 0.48; 95% CI 0.35–0.68, p < 0.001) were associated with lower MTB infection. Resistance to rifampicin was higher in the relapsed (AOR = 2.10; 95% CI 1.40–3.03, p < 0.001) and failure cases (AOR = 3.50; 95% CI 1.9–6.61, p < 001).ConclusionPrevalence of MTB and RR-MTB low. TB infection was higher in adult age groups and those who had previous TB treatment history. Similarly, resistance to rifampicin was higher among the relapsed and failure patients. Appropriate measurements in monitoring of TB treatment could reduce TB and RR-MTB in the study area.

Estimating TB diagnostic costs incurred under the National Tuberculosis Elimination Programme: a costing study from Tamil Nadu, South India.

BackgroundThe National Tuberculosis Elimination Programme (NTEP) of India is aiming to eliminate TB by 2025. The programme has increased its services and resources to strengthen the accurate and early detection of TB. It is important to estimate the cost of TB diagnosis in India considering the advancement and implementation of new diagnostic tools under the NTEP. The objective of this study was to estimate the unit costs of providing TB diagnostic services at different levels of public health facilities with different algorithms implemented under the NTEP in Chennai, Tamil Nadu, South India.MethodsThis costing study was conducted from the perspective of the health system. This study used only secondary data and information that were available in the public domain. Data were collected with the approval of health authorities. The patient's diagnostic path from the point of registration until the final diagnosis was considered in the costing exercise. The unit costs of different diagnostic tools used in the NTEP implemented by Chennai Corporation were calculated.ResultsWe estimated the unit cost of the eight laboratory tests (Ziehl–Neelsen [ZN], fluorescence microscopy [FM], x-ray, digital x-ray, gene Xpert MTB/RIF (cartridge-based nucleic acid amplification test [NAAT] that identifies rifampicin resistant Mycobacterium Tuberculosis) Mycobacterium Tuberculosis/Rifampicin [MTB/RIF], mycobacteria growth indicator tube [MGIT], line probe assay [LPA] and Lowenstein Jensen [LJ] culture) for diagnosis of drug-sensitive and drug-resistant TB. The unit costs included fixed and variable costs for smear examination by ZN microscopy (₹ [Indian Rupee] 326 [USn}{}{}4.72], FM (₹104 [USn}{}{}1.5]), x-ray (₹218 [USn}{}{}3.15]), digital X-ray (₹281 [USn}{}{}4.07]), gene Xpert MTB/RIF (₹1137 [USn}{}{}16.47]), MGIT (₹7038 [USn}{}{}102]), LPA (₹6448 [USn}{}{}93.44]) and LJ culture (₹4850 [USn}{}{}70.28]). Out of 10 diagnostic algorithms used for TB diagnosis, algorithms using only smear microscopy had the lowest cost, followed by smear microscopy with x-ray for drug-sensitive TB (₹104 [USn}{}{}1.5] to ₹544 [USn}{}{}7.88]). Diagnostic algorithms for drug-resistant TB involving LPA and gene Xpert MTB/RIF were the most expensive.ConclusionsUnderstanding the various costs contributing to TB diagnosis in India provides crucial evidence for policymakers, programme managers and researchers to optimise programme spending and efficiently use resources.

Status of rpoB gene mutation associated with rifampicin-resistant Mycobacterium tuberculosis isolated in a rural setting in Nepal.

Mycobacterium tuberculosis ranks among the top 10 causes of deaths in Nepal despite the country having a long history of national tuberculosis prevention programmes that have proved very successful in the control of tuberculosis. Several cases of active or latent tuberculosis are still missing despite that the number of infected individuals is increasing each year. Microscopy has its own limitations and factors like low bacterial load, quality of sample, quality of smear, experience of microscopist etc. influence the overall sensitivity of the test. The implementation of a molecular technique-based rapid, point-of-care testing system offers higher sensitivity in the early diagnosis of tuberculosis. Cepheid GeneXpert is the most commonly used molecular technology in Nepal. It is a cartridge-based semi-quantitative, nested real-time PCR-based diagnostic system. It detects mutations in the beta-subunit of RNA polymerase (rpoB) gene that lead to rifampicin resistance (RR) in M. tuberculosis complex. The present study aims to increase our understanding of the epidemiology of mutations in the rpoB gene in tuberculosis-positive patients by using the Xpert MTB/RIF assay in a rural setting in Pyuthan Hospital, Nepal. Sputum from 2733 patients was tested for the diagnosis of tuberculosis using the Cepheid GeneXpert system between July 2018 and January 2020 at Pyuthan Hospital. Two hundred and ninety-seven of these samples (10.86 %) were positive for M. tuberculosis , of which 3.3 % (10/297) were rifampicin-resistant. Among rifampicin-resistant tuberculosis (RR-TB) patients, 50.0 % (5/10) showed mutations located in codons 529–533 (probe E) of the rpoB gene, followed by others. The GeneXpert system can be a convenient, highly sensitive, rapid and accurate tool for the diagnosis of tuberculosis, also identifying RR-TB and at the same time determining the molecular epidemiology of rifampin resistance-associated mutations in rural and/or resource-limited laboratory settings.

Xpert MTB/RIF combined with T-SPOT.TB in the diagnostic value of drug-resistant tuberculous pleurisy

Objective To investigate the diagnostic value of Xpert MTB/RIF combined with T-SPOT.TB in the drug-resistant tuberculous pleurisy. Methods Sixty-two patients with drug-resistant tuberculous pleurisy were selected as tuberculous pleurisy group, and 60 patients with other diseases and pleural effusion admitted to Hebei Province Chest Hospital from January 2014 to December 2019 were selected as the control. General clinical data were collected and analyzed, two groups of pleural effusion samples were collected for detection of Xpert MTB/RIF, T-SPOT.TB and tubercle bacillus culture and drug sensitivity, etc, tubercle bacillus culture of pleural effusion was used as the gold standard for the diagnosis of tuberculous pleurisy. Results The sensitivity of Xpert MTB/RIF, T-SPOT.TB and two parallel methods was 79.0%, 88.7%, 96.8% respectively, the specificity was 90.0%, 81.7%, 95.0% respectively, the diagnostic sensitivity, specificity were significantly higher than those of ADA and acid fast staining (all P <0.05). Taking the tuberclebacillus drug sensitivity as the gold standard, the sensitivity of Xpert MTB/RIF test for rifampicin resistance was 89.47%, and the consistency between the two methods was good ( Kappa =0.900). Conclusion Xpert MTB/RIF combined with T-SPOT.TB in the diagnosis of tuberculous pleurisy can improve the sensitivity and specificity, reduce the misdiagnosis rate, at the same time, Xpert MTB/RIF can quickly detect rifampicin resistance of Mycobacterium tuberculosis in pleural effusion, be conducive to the early diagnosis and treatment of drug-resistant tuberculous pleurisy. 摘要：目的 探讨结核分枝杆菌/利福平耐药基因检测(Xpert MTB/RIF)联合结核感染T细胞斑点检测(T-SPOT.TB)对耐药性结核性胸膜炎诊断及治疗的价值。 方法 选取2014年1月至2019年12月河北省胸科医院收治的62例结 核性胸膜炎患者为结核性胸膜炎组, 选择同期来河北省胸科医院院救治的非结核性胸膜炎患者60例为对照组。分析 比较两组一般资料, 留取两组胸腔积液样本进行Xpert MTB/RIF、T-SPOT.TB、结核菌培养+药敏等检测, 胸腔积液结核 菌培养阳性作为诊断结核性胸膜炎的金标准。 结果 Xpert MTB/RIF、T-SPOT.TB、两种方法并联检测结核性胸膜炎的 灵敏度分别为79.0%、88.7%、96.8%, 特异度分别为90.0%、81.7%、95.0%, 均明显髙于ADA及抗酸染色诊断的敏感度及 特异度, 差异有统计学意义( P 均<0.05)。以结核菌药敏检测为金标准, XpertMTB/RIF检测利福平耐药的敏感度为 89.47%, 两者一致性较好( Kappa =0.900)。 结论 Xpert MTB/RIF联合T-SPOT.TB检测提髙了结核性胸膜炎诊断的敏感 度和特异度, 降低误诊率, 同时XpertMTB/RIF可快速检测出胸水中耐利福平的结核分枝杆菌感染, 有利于耐药结核性 胸膜炎的早期诊断及治疗。

Preparing the healthcare workforce in South Africa for short-course rifampicin-resistant TB treatment: inter-professional training and task-sharing considerations

BackgroundTreatment for rifampicin-resistant Mycobacterium tuberculosis (RR-TB) is complex, however, shorter treatment, with newer antimicrobials are improving treatment outcomes. The South African National Department of Health (NDoH) recently accelerated the rollout of 9-month, all-oral, RR-TB short-course regimens. We sought to evaluate an inter-professional training program using pre-test and post-test performance of Professional Nurses (PNs), Advanced Practice Professional Nurses (APPNs) and Medical Officers (MOs) to inform: (a) training needs across cadres; (b) knowledge performance, by cadres; and (c) training differences in knowledge by nurse type.MethodsA 4-day didactic and case-based clinical decision support course for RR-TB regimens in South Africa (SA) was developed, reviewed and nationally accredited. Between February 2017 and July 2018, 12 training events were held. Clinicians who may initiate RR-TB treatment, specifically MOs and PN/APPNs with matched pre–post tests and demographic surveys were analyzed. Descriptive statistics are provided. Pre–post test evaluations included 25 evidence-based clinically related questions about RR-TB diagnosis, treatment, and care.ResultsParticipants (N = 842) participated in testing, and matched evaluations were received for 800 (95.0%) training participants. Demographic data were available for 793 (99.13%) participants, of whom 762 (96.1%) were MOs, or nurses, either PN or APPNs. Average correct response pre-test and post-test scores were 61.7% (range 7–24 correct responses) and 85.9% (range 12–25), respectively. Overall, 95.8% (730/762) of participants demonstrated improved knowledge. PNs improved on average 25% (6.22 points), whereas MOs improved 10% (2.89 points) with better mean test scores on both pre- and post-test (p < 0.000). APPNs performed the same as the MOs on post-test scores (p = NS).ConclusionsThe inter-professional training program in short-course RR-TB treatment improved knowledge for participants. MOs had significantly greater pre-test scores. Of the nurses, APPNs outperformed other PNs, and performed equally to MOs on post-test scores, suggesting this advanced cadre of nurses might be the most appropriate to initiate and monitor treatment in close collaboration with MOs. All cadres of nurse reported the need for additional clinical training and mentoring prior to managing such patients.

GeneXpert: A new tool for the rapid detection of rifampicin resistance in mycobacterium tuberculosis

Background: present study was planned to evaluate the diagnostic accuracy of GeneXpert assay for the detection of rifampicin resistance in mycobacterium tuberculosis using conventional drug susceptibility testing as gold standard. Methods and materials: This cross-sectional study was conducted at Jinnah Hospital, Lahore,/Allama Iqbal Medical College, Lahore, Pakistan, from January 2012 to December 2014, and comprised clinically and radiologically diagnosed tuberculosis suspected cases. Pulmonary and extra-pulmonary specimens were collected from strong tuberculosis suspects. All specimens were processed for Ziehl Neelsen staining, Lowenstein-Jensen culture and GeneXpert assay. All mycobacterium tuberculosis positive cases on Lowenstein-Jensen culture were further processed for drug susceptibility testing. Results: Of the 2,200 cases, 840(49.46%) were positive for mycobacterium tuberculosis on GeneXpert assay. Of them, 134(15.6%) cases showed rifampicin resistance on GeneXpert assay. The sensitivity, specificity, positive predictive value and negative predictive value of GeneXpert assay for rifampicin resistance were 127(98.3%), 704(99.1%), 127(94.7%) and 704(99.4%), respectively, by comparing the results with drug susceptibility testing. Conclusion: GeneXpert assay was an extremely helpful diagnostic tool for the detection of rifampicin resistance in tuberculosis suspects with fairly high sensitivity and specificity along with short turnout time

Bedaquiline in the treatment regimen of multidrugresistant tuberculosis (clinical observation)

Current trials are underway to design chemotherapy regimens that include bedaquiline to treat children with rifampicin and isoniazid resistant mycobacterium tuberculosis. This clinical observation is interesting from the viewpoint of experience of the potential effective and safe use of the new anti-tuberculosis drug bedaquiline (Sirturo) in children under 12 years of age, as the publications of clinical trials of the drug use in children is limited. The female patient was diagnosed with primary tuberculosis complex complicated by infusional pleurisy based on the epidemiological anamnesis (family contact with tuberculosis patients), immunodiagnostic tests – a positive ATR-test result (Diaskin-test), pathological changes in the lung tissue and intrathoracic lymph nodes found on the chest CT scans. The diagnosis was confirmed by detection of MBT in pleural effusion by PCR and liquid culture method (BACTEC MGIT-960), which are resistant to isoniazid and rifampicin (multidrug resistance, MDR). The girl was enrolled in a clinical trial and treated according to chemotherapy regimen IV including a new anti-tuberculosis drug, which she received for 6 months. There was a positive clinical and X-ray dynamics of the tuberculosis process with minimal residual changes throughout the entire course of chemotherapy followed by confirmed clinical cure of TB (after 24-month follow-up). Based on the example of this clinical observation, it can be concluded that bedaquiline is safe and showed good clinical efficacy in the chemotherapy regimen to treat MDR MBT in children under 12 years of age.

Antimycobacterial Activity and Gas Chromatography-Mass Spectrometry (GC-MS) Analysis of Henna (Lawsonia Inermis) Leaves extract.

Lawsonia inermis or (henna) is used as a cosmetic dye and medicinal plant for its strong antimycobacterial and other therapeutic effects. In this study, we investigated the phytochemical and antimycobacterial activity of L. inermis leaves methanol extract against Rifampicin sensitive and resistant mycobacterium tuberculosis ( M.tb ) strains. Fresh Sudanese henna leaves were collected from the University of Gezira campus garden, shade dried, ground and was extracted by methanol. Crude henna methanolic extract was then tested for antimycobacterial activity against eight M.tb strains sensitive and resistant to rifampicin, using drug susceptibility test. The phytoconstituents were identified by using gas chromatograph interfaced to a mass spectrometer (GC-MS). GC-MS analysis revealed the presence of seven phytochemical components, with prevailing of squalene (34.433%), Lawsone (20.185%) and Coumarin-4-carboxylic acid (23.328%). The crude methanolic extract of henna leaves demonstrated an inhibitory effect on all tested M.tb strains, with antimycobacterial activity being ≤ 50 mg/ml. Henna leaves crude methanolic extract is rich in phytoconstituents that may involve in its remarkable antimycobacterial activity against rifampicin sensitive and resistant M.tb strains.

A highly rifampicin resistant Mycobacterium tuberculosis strain emerging in Southern Brazil

Here we described phenotypical, molecular and epidemiological features of a highly rifampicin-resistant Mycobacterium tuberculosis strain emerging in Southern Brazil, that carries an uncommon insertion of 12 nucleotides at the codon 435 in the rpoB gene. Employing a whole-genome sequencing-based study on drug-resistant Mycobacterium tuberculosis strains, we identified this emergent strain in 16 (9.19%) from 174 rifampicin-resistant clinical strains, all of them belonging to LAM RD115 sublineage. Nine of these 16 strains were available to minimum inhibitory concentration determination and for all of them was found a high rifampicin-resistance level (≥to 32 mg/L). This high resistance level could be explained by structural changes into the RIF binding site of RNA polymerase caused by the insertions, and consequent low-affinity interaction with rifampicin complex confirmed through protein modeling and molecular docking simulations. Epidemiological investigation showed that most of the individuals (56.25%) infected by the studied strains were prison inmate individuals or that spent some time in prison. The phylogenomic approach revealed that strains carrying on insertion belonged to same genomic cluster, evidencing a communal transmission chain involving inmate individuals and community. We stress the importance of tuberculosis genomic surveillance and introduction of measures to interrupt Mycobacterium tuberculosis transmission chain in this region.

A Glutamine Insertion at Codon 432 of RpoB Confers Rifampicin Resistance in Mycobacterium tuberculosis.

Tuberculosis (TB) is an infectious respiratory disease caused by Mycobacterium tuberculosis and one of the top 10 causes of death worldwide. Treating TB is challenging; successful treatment requires a long course of multiple antibiotics. Rifampicin (RIF) is a first-line drug for treating TB, and the development of RIF-resistant M. tuberculosis makes treatment even more difficult. To determine the mechanism of RIF resistance in these strains, we searched for novel mutations by sequencing. Four isolates, CDC-1, CDC-2, CDC-3, and CDC-4, had high-level RIF resistance and unique mutations encoding RpoB G158R, RpoB V168A, RpoB S188P, and RpoB Q432insQ, respectively. To evaluate their correlation with RIF resistance, plasmids carrying rpoB genes encoding these mutant proteins were transfected into the H37Rv reference strain. The plasmid complementation of RpoB indicated that G158R, V168A, and S188P did not affect the MIC of RIF. However, the MIC of RIF was increased in H37Rv carrying RpoB Q432insQ. To confirm the correlation between RIF resistance and Q432insQ, we cloned an rpoB fragment carrying the insertion (encoding RpoB Q432insQ) into H37Rv by homologous recombination using a suicide vector. All replacement mutants expressing RpoB Q432insQ were resistant to RIF (MIC > 1 mg/L). These results indicate that RpoB Q432insQ causes RIF resistance in M. tuberculosis.

Mutations Associated with Rifampicin Resistance in Mycobacterium tuberculosis Isolates from Moroccan Patients: Systematic Review.

Background In recent years, the treatment of tuberculosis has been threatened by the increasing number of patients with drug resistance, especially rifampicin resistance, which is the most effective first-line antibiotic against Mycobacterium tuberculosis. Methods We performed a systematic review of the literature by searching the PubMed database for studies of rifampicin-resistant Mycobacterium tuberculosis (MTB) isolates from Moroccan patients, published between 2010 and 2020. The aim of this review was to quantify the frequency of the most common mutations associated with rifampicin resistance, to describe the frequency at which these mutations co-occur. Identified studies were critically appraised according to the Quality Assessment of Diagnostic Accuracy Studies-2 (QUADAS-2) tool. Results 6 studies met our inclusion criteria. Results show that 99.36% of MTB isolates had a single-point mutation, and the most commonly mutated codon of rpoB gene is 531 with 70.33% of phenotypically resistant strains. However, 10.38% of MTB strains phenotypically resistant to RIF did not exhibit any mutation in the rpoB gene. Conclusion Identification of a resistance-associated mutation to rifampicin can be a good marker of drug-resistant TB, but lack of a mutation in the target sequence must be interpreted with caution.

Rifampicin-Resistant Mycobacterium tuberculosis Among Patients with Presumptive Tuberculosis in Addis Ababa, Ethiopia.

BackgroundDrug-resistant tuberculosis remains a major public health threat complicating tuberculosis control programs globally. Data on rifampicin resistance (RR), which is a surrogate marker for multidrug resistance, are limited among Ethiopian tuberculosis patients. This study aimed to determine the magnitude of rifampicin-resistant Mycobacterium tuberculosis (RR-MTB) among presumptive tuberculosis patients attending St. Peter Tuberculosis Specialized Hospital, Addis Ababa, Ethiopia.Patients and MethodsA retrospective cross-sectional study was conducted at St. Peter Tuberculosis Specialized Hospital from January 2016 to December 2018. After checking completeness of the necessary information, data of tuberculosis-presumptive cases who underwent Gene Xpert® testing were collected from medical records using a data-extraction format prepared for this study purpose. Data were double entered and analyzed using SPSS version 20 statistical software.ResultsA total of 12,685 presumptive tuberculosis patients were included; of whom 54.5% were males and the mean age of the study participants was 40.3±18.7 years. Mycobacterium tuberculosis (MTB) was detected in 1714 participants (13.5%). Of these MTB cases, 169 cases (9.8%) were confirmed to have RR-MTB. Prevalence of MTB was relatively higher among males (15.1%, P=0.78); whereas RR-MTB was higher among females (10.3%, P=0.81). The incidence of MTB and RR-MTB was significantly associated with treatment history (P=0.042 and P=0.025), respectively. HIV infection has significantly associated with incidence of RR-MTB (P=0.032), but not with MTB (P˃0.05). Prevalence of MTB and RR-MTB had a declining trend through time, being 16.7% and 12.9%, 12.8% and 9.1%, and 12.2% and 7.9% in 2016, 2017 and 2018, respectively.ConclusionThis study showed a decreasing trend of both MTB and RR-MTB from 2016 to 2018 in an MTB, MDR-MTB, and TB/HIV co-infection high-burden setting, Addis Ababa, Ethiopia. Occurrence of MTB and RR-MTB was associated with treatment history. Therefore, improvement in treatment adherence of identified cases would be helpful to prevent emergence or re-emergence of MTB and RR-MTB cases.