Estimation of country-specific tuberculosis resistance antibiograms using pathogen genomics and machine learning.

BACKGROUND: Global tuberculosis (TB) drug resistance (DR) surveillance focuses on rifampicin. We examined the potential of public and surveillance Mycobacterium tuberculosis (Mtb) whole-genome sequencing (WGS) data, to generate expanded country-level resistance prevalence estimates (antibiograms) using in silico resistance prediction. METHODS: We curated and quality-controlled Mtb WGS data. We used a validated random forest model to predict phenotypic resistance to 12 drugs and bias-corrected for model performance, outbreak sampling and rifampicin resistance oversampling. Validation leveraged a national DR survey conducted in South Africa. RESULTS: Mtb isolates from 29 countries (n=19 149) met sequence quality criteria. Global marginal genotypic resistance among mono-resistant TB estimates overlapped with the South African DR survey, except for isoniazid, ethionamide and second-line injectables, which were underestimated (n=3134). Among multidrug resistant (MDR) TB (n=268), estimates overlapped for the fluoroquinolones but overestimated other drugs. Globally pooled mono-resistance to isoniazid was 10.9% (95% CI: 10.2-11.7%, n=14 012). Mono-levofloxacin resistance rates were highest in South Asia (Pakistan 3.4% (0.1-11%), n=111 and India 2.8% (0.08-9.4%), n=114). Given the recent interest in drugs enhancing ethionamide activity and their expected activity against isolates with resistance discordance between isoniazid and ethionamide, we measured this rate and found it to be high at 74.4% (IQR: 64.5-79.7%) of isoniazid-resistant isolates predicted to be ethionamide susceptible. The global susceptibility rate to pyrazinamide and levofloxacin among MDR was 15.1% (95% CI: 10.2-19.9%, n=3964). CONCLUSIONS: This is the first attempt at global Mtb antibiogram estimation. DR prevalence in Mtb can be reliably estimated using public WGS and phenotypic resistance prediction for key antibiotics, but public WGS data demonstrates oversampling of isolates with higher resistance levels than MDR. Nevertheless, our results raise concerns about the empiric use of short-course fluoroquinolone regimens for drug-susceptible TB in South Asia and indicate underutilisation of ethionamide in MDR treatment.

Genotypic and phenotypic comparison of drug resistance profiles of clinical multidrug-resistant Mycobacterium tuberculosis isolates using whole genome sequencing in Latvia.

BACKGROUND: Multidrug-resistant tuberculosis (MDR-TB) remains a major public health problem in many high tuberculosis (TB) burden countries. Phenotypic drug susceptibility testing (DST) take several weeks or months to result, but line probe assays and Xpert/Rif Ultra assay detect a limited number of resistance conferring gene mutations. Whole genome sequencing (WGS) is an advanced molecular testing method which theoretically can predict the resistance of M. tuberculosis (Mtb) isolates to all anti-TB agents through a single analysis. METHODS: Here, we aimed to identify the level of concordance between the phenotypic and WGS-based genotypic drug susceptibility (DS) patterns of MDR-TB isolates. Overall, data for 12 anti-TB medications were analyzed. RESULTS: In total, 63 MDR-TB Mtb isolates were included in the analysis, representing 27.4% of the total number of MDR-TB cases in Latvia in 2012-2014. Among them, five different sublineages were detected, and 2.2.1 (Beijing group) and 4.3.3 (Latin American-Mediterranean group) were the most abundant. There were 100% agreement between phenotypic and genotypic DS pattern for isoniazid, rifampicin, and linezolid. High concordance rate (> 90%) between phenotypic and genotypic DST results was detected for ofloxacin (93.7%), pyrazinamide (93.7%) and streptomycin (95.4%). Phenotypic and genotypic DS patterns were poorly correlated for ethionamide (agreement 56.4%), ethambutol (85.7%), amikacin (82.5%), capreomycin (81.0%), kanamycin (85.4%), and moxifloxacin (77.8%). For capreomycin, resistance conferring mutations were not identified in several phenotypically resistant isolates, and, in contrary, for ethionamide, ethambutol, amikacin, kanamycin, and moxifloxacin the resistance-related mutations were identified in several phenotypically sensitive isolates. CONCLUSIONS: WGS is a valuable tool for rapid genotypic DST for all anti-TB agents. For isoniazid and rifampicin phenotypic DST potentially can be replaced by genotypic DST based on 100% agreement between the tests. However, discrepant results for other anti-TB agents limit their prescription based solely on WGS data. For clinical decision, at the current level of knowledge, there is a need for combination of genotypic DST with modern, validated phenotypic DST methodologies for those medications which did not showed 100% agreement between the methods.

Variation in missed doses and reasons for discontinuation of anti-tuberculosis drugs during hospital treatment for drug-resistant tuberculosis in South Africa.

BACKGROUND: Updated World Health Organization (WHO) treatment guidelines prioritize all-oral drug-resistant tuberculosis (DR-TB) regimens. Several poorly tolerated drugs, such as amikacin and para-aminosalicylic acid (PAS), remain treatment options for DR-TB in WHO-recommended longer regimens as Group C drugs. Incomplete treatment with anti-TB drugs increases the risk of treatment failure, relapse, and death. We determined whether missed doses of individual anti-TB drugs, and reasons for their discontinuation, varied in closely monitored hospital settings prior to the 2020 WHO DR-TB treatment guideline updates. METHODS: We collected retrospective data on adult patients with microbiologically confirmed DR-TB between 2008 and 2015 who were selected for a study of acquired drug resistance in the Western Cape Province of South Africa. Medical records through mid-2017 were reviewed. Patients received directly observed treatment during hospitalization at specialized DR-TB hospitals. Incomplete treatment with individual anti-TB drugs, defined as the failure to take medication as prescribed, regardless of reason, was determined by comparing percent missed doses, stratified by HIV status and DR-TB regimen. We applied a generalized mixed effects model. RESULTS: Among 242 patients, 131 (54%) were male, 97 (40%) were living with HIV, 175 (72%) received second-line treatment prior to first hospitalization, and 191 (79%) died during the study period. At initial hospitalization, 134 (55%) patients had Mycobacterium tuberculosis with resistance to rifampicin and isoniazid (multidrug-resistant TB [MDR-TB]) without resistance to ofloxacin or amikacin, and 102 (42%) had resistance to ofloxacin and/or amikacin. Most patients (129 [53%]) had multiple hospitalizations and DST changes occurred in 146 (60%) by the end of their last hospital discharge. Incomplete treatment was significantly higher for amikacin (18%), capreomycin (18%), PAS (17%) and kanamycin (16%) than other DR-TB drugs (P<0.001), including ethionamide (8%), moxifloxacin (7%), terizidone (7%), ethambutol (7%), and pyrazinamide (6%). Among the most frequently prescribed drugs, second-line injectables had the highest rates of discontinuation for adverse events (range 0.56-1.02 events per year follow-up), while amikacin, PAS and ethionamide had the highest rates of discontinuation for patient refusal (range 0.51-0.68 events per year follow-up). Missed doses did not differ according to HIV status or anti-TB drug combinations. CONCLUSION: We found that incomplete treatment for second-line injectables and PAS during hospitalization was higher than for other anti-TB drugs. To maximize treatment success, interventions to improve person-centered care and mitigate adverse events may be necessary in cases when PAS or amikacin (2020 WHO recommended Group C drugs) are needed.

Pharmacokinetics of standard versus high-dose isoniazid for treatment of multidrug-resistant tuberculosis.

BACKGROUND: The WHO-endorsed shorter-course regimen for MDR-TB includes high-dose isoniazid. The pharmacokinetics of high-dose isoniazid within MDR-TB regimens has not been well described. OBJECTIVES: To characterize isoniazid pharmacokinetics at 5-15 mg/kg as monotherapy or as part of the MDR-TB treatment regimen. METHODS: We used non-linear mixed-effects modelling to evaluate the combined data from INHindsight, a 7 day early bactericidal activity study with isoniazid monotherapy, and PODRtb, an observational study of patients on MDR-TB treatment including terizidone, pyrazinamide, moxifloxacin, kanamycin, ethionamide and/or isoniazid. RESULTS: A total of 58 and 103 participants from the INHindsight and PODRtb studies, respectively, were included in the analysis. A two-compartment model with hepatic elimination best described the data. N-acetyltransferase 2 (NAT2) genotype caused multi-modal clearance, and saturable first-pass was observed beyond 10 mg/kg dosing. Saturable isoniazid kinetics predicted an increased exposure of approximately 50% beyond linearity at 20 mg/kg dosing. Participants treated with the MDR-TB regimen had a 65.6% lower AUC compared with participants on monotherapy. Ethionamide co-administration was associated with a 29% increase in isoniazid AUC. CONCLUSIONS: Markedly lower isoniazid exposures were observed in participants on combination MDR-TB treatment compared with monotherapy. Isoniazid displays saturable kinetics at doses >10 mg/kg. The safety implications of these phenomena remain unclear.

The small-molecule SMARt751 reverses Mycobacterium tuberculosis resistance to ethionamide in acute and chronic mouse models of tuberculosis.

The sensitivity of Mycobacterium tuberculosis, the pathogen that causes tuberculosis (TB), to antibiotic prodrugs is dependent on the efficacy of the activation process that transforms the prodrugs into their active antibacterial moieties. Various oxidases of M. tuberculosis have the potential to activate the prodrug ethionamide. Here, we used medicinal chemistry coupled with a phenotypic assay to select the N-acylated 4-phenylpiperidine compound series. The lead compound, SMARt751, interacted with the transcriptional regulator VirS of M. tuberculosis, which regulates the mymA operon encoding a monooxygenase that activates ethionamide. SMARt751 boosted the efficacy of ethionamide in vitro and in mouse models of acute and chronic TB. SMARt751 also restored full efficacy of ethionamide in mice infected with M. tuberculosis strains carrying mutations in the ethA gene, which cause ethionamide resistance in the clinic. SMARt751 was shown to be safe in tests conducted in vitro and in vivo. A model extrapolating animal pharmacokinetic and pharmacodynamic parameters to humans predicted that as little as 25 mg of SMARt751 daily would allow a fourfold reduction in the dose of ethionamide administered while retaining the same efficacy and reducing side effects.

Xpert MTB/XDR for detection of pulmonary tuberculosis and resistance to isoniazid, fluoroquinolones, ethionamide, and amikacin.

BACKGROUND: The World Health Organization (WHO) End TB Strategy stresses universal access to drug susceptibility testing (DST). DST determines whether Mycobacterium tuberculosis bacteria are susceptible or resistant to drugs. Xpert MTB/XDR is a rapid nucleic acid amplification test for detection of tuberculosis and drug resistance in one test suitable for use in peripheral and intermediate level laboratories. In specimens where tuberculosis is detected by Xpert MTB/XDR, Xpert MTB/XDR can also detect resistance to isoniazid, fluoroquinolones, ethionamide, and amikacin. OBJECTIVES: To assess the diagnostic accuracy of Xpert MTB/XDR for pulmonary tuberculosis in people with presumptive pulmonary tuberculosis (having signs and symptoms suggestive of tuberculosis, including cough, fever, weight loss, night sweats). To assess the diagnostic accuracy of Xpert MTB/XDR for resistance to isoniazid, fluoroquinolones, ethionamide, and amikacin in people with tuberculosis detected by Xpert MTB/XDR, irrespective of rifampicin resistance (whether or not rifampicin resistance status was known) and with known rifampicin resistance. SEARCH METHODS: We searched multiple databases to 23 September 2021. We limited searches to 2015 onwards as Xpert MTB/XDR was launched in 2020. SELECTION CRITERIA: Diagnostic accuracy studies using sputum in adults with presumptive or confirmed pulmonary tuberculosis. Reference standards were culture (pulmonary tuberculosis detection); phenotypic DST (pDST), genotypic DST (gDST),composite (pDST and gDST) (drug resistance detection). DATA COLLECTION AND ANALYSIS: Two review authors independently reviewed reports for eligibility and extracted data using a standardized form. For multicentre studies, we anticipated variability in the type and frequency of mutations associated with resistance to a given drug at the different centres and considered each centre as an independent study cohort for quality assessment and analysis. We assessed methodological quality with QUADAS-2, judging risk of bias separately for each target condition and reference standard. For pulmonary tuberculosis detection, owing to heterogeneity in participant characteristics and observed specificity estimates, we reported a range of sensitivity and specificity estimates and did not perform a meta-analysis. For drug resistance detection, we performed meta-analyses by reference standard using bivariate random-effects models. Using GRADE, we assessed certainty of evidence of Xpert MTB/XDR accuracy for detection of resistance to isoniazid and fluoroquinolones in people irrespective of rifampicin resistance and to ethionamide and amikacin in people with known rifampicin resistance, reflecting real-world situations. We used pDST, except for ethionamide resistance where we considered gDST a better reference standard. MAIN RESULTS: We included two multicentre studies from high multidrug-resistant/rifampicin-resistant tuberculosis burden countries, reporting on six independent study cohorts, involving 1228 participants for pulmonary tuberculosis detection and 1141 participants for drug resistance detection. The proportion of participants with rifampicin resistance in the two studies was 47.9% and 80.9%. For tuberculosis detection, we judged high risk of bias for patient selection owing to selective recruitment. For ethionamide resistance detection, we judged high risk of bias for the reference standard, both pDST and gDST, though we considered gDST a better reference standard. Pulmonary tuberculosis detection - Xpert MTB/XDR sensitivity range, 98.3% (96.1 to 99.5) to 98.9% (96.2 to 99.9) and specificity range, 22.5% (14.3 to 32.6) to 100.0% (86.3 to 100.0); median prevalence of pulmonary tuberculosis 91.3%, (interquartile range, 89.3% to 91.8%), (2 studies; 1 study reported on 2 cohorts, 1228 participants; very low-certainty evidence, sensitivity and specificity). Drug resistance detection People irrespective of rifampicin resistance - Isoniazid resistance: Xpert MTB/XDR summary sensitivity and specificity (95% confidence interval (CI)) were 94.2% (87.5 to 97.4) and 98.5% (92.6 to 99.7) against pDST, (6 cohorts, 1083 participants, moderate-certainty evidence, sensitivity and specificity). - Fluoroquinolone resistance: Xpert MTB/XDR summary sensitivity and specificity were 93.2% (88.1 to 96.2) and 98.0% (90.8 to 99.6) against pDST, (6 cohorts, 1021 participants; high-certainty evidence, sensitivity; moderate-certainty evidence, specificity). People with known rifampicin resistance - Ethionamide resistance: Xpert MTB/XDR summary sensitivity and specificity were 98.0% (74.2 to 99.9) and 99.7% (83.5 to 100.0) against gDST, (4 cohorts, 434 participants; very low-certainty evidence, sensitivity and specificity). - Amikacin resistance: Xpert MTB/XDR summary sensitivity and specificity were 86.1% (75.0 to 92.7) and 98.9% (93.0 to 99.8) against pDST, (4 cohorts, 490 participants; low-certainty evidence, sensitivity; high-certainty evidence, specificity). Of 1000 people with pulmonary tuberculosis, detected as tuberculosis by Xpert MTB/XDR: - where 50 have isoniazid resistance, 61 would have an Xpert MTB/XDR result indicating isoniazid resistance: of these, 14/61 (23%) would not have isoniazid resistance (FP); 939 (of 1000 people) would have a result indicating the absence of isoniazid resistance: of these, 3/939 (0%) would have isoniazid resistance (FN). - where 50 have fluoroquinolone resistance, 66 would have an Xpert MTB/XDR result indicating fluoroquinolone resistance: of these, 19/66 (29%) would not have fluoroquinolone resistance (FP); 934 would have a result indicating the absence of fluoroquinolone resistance: of these, 3/934 (0%) would have fluoroquinolone resistance (FN). - where 300 have ethionamide resistance, 296 would have an Xpert MTB/XDR result indicating ethionamide resistance: of these, 2/296 (1%) would not have ethionamide resistance (FP); 704 would have a result indicating the absence of ethionamide resistance: of these, 6/704 (1%) would have ethionamide resistance (FN). - where 135 have amikacin resistance, 126 would have an Xpert MTB/XDR result indicating amikacin resistance: of these, 10/126 (8%) would not have amikacin resistance (FP); 874 would have a result indicating the absence of amikacin resistance: of these, 19/874 (2%) would have amikacin resistance (FN). AUTHORS' CONCLUSIONS: Review findings suggest that, in people determined by Xpert MTB/XDR to be tuberculosis-positive, Xpert MTB/XDR provides accurate results for detection of isoniazid and fluoroquinolone resistance and can assist with selection of an optimised treatment regimen. Given that Xpert MTB/XDR targets a limited number of resistance variants in specific genes, the test may perform differently in different settings. Findings in this review should be interpreted with caution. Sensitivity for detection of ethionamide resistance was based only on Xpert MTB/XDR detection of mutations in the inhA promoter region, a known limitation. High risk of bias limits our confidence in Xpert MTB/XDR accuracy for pulmonary tuberculosis. Xpert MTB/XDR's impact will depend on its ability to detect tuberculosis (required for DST), prevalence of resistance to a given drug, health care infrastructure, and access to other tests.

Phosphatidylcholine (18:0/20:4), a potential biomarker to predict ethionamide-induced hepatic steatosis in rats.

Ethionamide (ETH), a second-line drug for multidrug-resistant tuberculosis, is known to cause hepatic steatosis in rats and humans. To investigate predictive biomarkers for ETH-induced steatosis, we performed lipidomics analysis using plasma and liver samples collected from rats treated orally with ETH at 30 and 100 mg/kg for 14 days. The ETH-treated rats developed hepatic steatosis with Oil Red O staining-positive vacuolation in the centrilobular hepatocytes accompanied by increased hepatic contents of triglycerides (TG) and decreased plasma TG and total cholesterol levels. A multivariate analysis for lipid profiles revealed differences in each of the 35 lipid species in the plasma and liver between the control and the ETH-treated rats. Of those lipids, phosphatidylcholine (PC) (18:0/20:4) decreased dose-dependently in both the plasma and liver. Moreover, serum TG-rich very low-density lipoprotein (VLDL) levels, especially the large particle fraction of VLDL composed of PC containing arachidonic acid (20:4) involved in hepatic secretion of TG, were decreased dose-dependently. In conclusion, the decreased PC (18:0/20:4) in the liver, possibly leading to suppression of hepatic TG secretion, was considered to be involved in the pathogenesis of the ETH-induced hepatic steatosis. Therefore, plasma PC (18:0/20:4) levels are proposed as mechanism-related biomarkers for ETH-induced hepatic steatosis.

Detection of isoniazid, fluoroquinolone, ethionamide, amikacin, kanamycin, and capreomycin resistance by the Xpert MTB/XDR assay: a cross-sectional multicentre diagnostic accuracy study.

BACKGROUND: The WHO End TB Strategy requires drug susceptibility testing and treatment of all people with tuberculosis, but second-line diagnostic testing with line-probe assays needs to be done in experienced laboratories with advanced infrastructure. Fewer than half of people with drug-resistant tuberculosis receive appropriate treatment. We assessed the diagnostic accuracy of the rapid Xpert MTB/XDR automated molecular assay (Cepheid, Sunnyvale, CA, USA) to overcome these limitations. METHODS: We did a prospective study involving individuals presenting with pulmonary tuberculosis symptoms and at least one risk factor for drug resistance in four sites in India (New Delhi and Mumbai), Moldova, and South Africa between July 31, 2019, and March 21, 2020. The Xpert MTB/XDR assay was used as a reflex test to detect resistance to isoniazid, fluoroquinolones, ethionamide, amikacin, kanamycin, and capreomycin in adults with positive results for Mycobacterium tuberculosis complex on Xpert MTB/RIF or Ultra (Cepheid). Diagnostic performance was assessed against a composite reference standard of phenotypic drug-susceptibility testing and whole-genome sequencing. This study is registered with ClinicalTrials.gov, number NCT03728725. FINDINGS: Of 710 participants, 611 (86%) had results from both Xpert MTB/XDR and the reference standard for any drug and were included in analysis. Sensitivity for Xpert MTB/XDR detection of resistance was 94% (460 of 488, 95% CI 92-96) for isoniazid, 94% (222 of 235, 90-96%) for fluoroquinolones, 54% (178 of 328, 50-61) for ethionamide, 73% (60 of 82, 62-81) for amikacin, 86% (181 of 210, 81-91) for kanamycin, and 61% (53 of 87, 49-70) for capreomycin. Specificity was 98-100% for all drugs. Performance was equivalent to that of line-probe assays. The non-determinate rate of Xpert MTB/XDR (ie, invalid M tuberculosis complex detection) was 2·96%. INTERPRETATION: The Xpert MTB/XDR assay showed high diagnostic accuracy and met WHO's minimum target product profile criteria for a next-generation drug susceptibility test. The assay has the potential to diagnose drug-resistant tuberculosis rapidly and accurately and enable optimum treatment. FUNDING: German Federal Ministry of Education and Research through KfW, Dutch Ministry of Foreign Affairs, and Australian Department of Foreign Affairs and Trade.

New ethionamide boosters and EthR2: structural and energetic analysis.

Ethionamide (ETH) is a high-profile drug for the treatment of patients with multidrug-resistant Mycobacterium tuberculosis and, in order to produce its inhibitory effects, it needs to be bioactivated by monooxygenase EthA. This process is under the control of the transcriptional repressors EthR and EthR2, so that their inhibition results in the boosting of ethionamide activation. Herein, through crystallographic data and computer simulations, we calculated the interaction binding energies of four inhibitors with improved in vitro potency, namely BDM76060 (PDB ID: 6HS1), BDM72201 (PDB ID: 6HRX), BDM76150 (PDB ID: 6HS2) and BDM72719 (PDB ID: 6HRY), in complexes with the transcriptional repressor EthR2, using density functional theory (DFT) within the molecular fractionation with conjugated caps (MFCC) approach. It was observed that these ligands share the same binding site within a 10.0 Å radius of the EthR2 protein; however, their structural particularities have a significant impact on the global energies of systems. The BDM72201 and BDM72719 components are weakly attached to the binding site, while BDM76060 and BDM76150 components produce stronger bonds, corroborating with experimental studies demonstrating that BDM76060 and BDM76150 are more successful in producing inhibitory effects. BDM76060 and BDM76150 have many functional groups that increase the contact surface with the protein and attract a more significant number of amino acid residues, being able to produce polarities that generate stronger interactions. In the current scenario of a growing number of cases of bacterial resistance, the obtained data can be used to guide clinical trials of these inhibitors and other inhibitors that act on the alternative EthR2 pathway, focusing on improving the activity of ethionamide, its effectiveness, a reduction in the treatment time and exposure to cytotoxic effects.

Effectiveness of Shorter Treatment Regimen in Multidrug-Resistant Tuberculosis Patients in Pakistan: A Multicenter Retrospective Record Review.

In Pakistan, the treatment of multidrug-resistant tuberculosis (MDR-TB) with a shorter treatment regimen (STR), that is, 4-6 months of amikacin, moxifloxacin (Mfx), ethionamide, clofazimine (Cfz), pyrazinamide (Z), ethambutol (E), and high-dose isoniazid, followed by 5 months of Mfx, Cfz, Z, and E, was initiated in 2018. However, there is a lack of information about its effectiveness in Pakistani healthcare settings. Therefore, this retrospective record review of MDR-TB patients treated with STR at eight treatment sites in Pakistan aimed to fill this gap. Data were analyzed using SPSS 23. Multivariate binary logistic regression (MVBLR) analysis was conducted to find factors associated with death and treatment failure, and lost to follow-up (LTFU). A P-value < 0.05 was considered statistically significant. Of 912 MDR-TB patients enrolled at the study sites, only 313 (34.3%) eligible patients were treated with STR and included in the current study. Of them, a total of 250 (79.9%) were cured, 12 (3.8%) completed treated, 31 (9.9%) died, 16 (5.1%) were LTFU, and four (1.3%) were declared as treatment failures. The overall treatment success rate was 83.7%. In MVBLR analysis, patients' age of 41-60 (odds ratio [OR] = 4.9, P-value = 0.020) and > 60 years (OR = 3.6, P-value = 0.035), being underweight (OR = 2.7, P-value = 0.042), and previous TB treatment (OR = 0.4, P-value = 0.042) had statistically significant association with death and treatment failure, whereas patients' age of > 60 years (OR = 5.4, P-value = 0.040) and previous TB treatment (OR = 0.2, P-value = 0.008) had statistically significant association with LTFU. The treatment success rate of STR was encouraging. However, to further improve the treatment outcomes, special attention should be paid to the patients with identified risk factors.

Whole-genome sequence analysis and comparisons between drug-resistance mutations and minimum inhibitory concentrations of Mycobacterium tuberculosis isolates causing M/XDR-TB.

Drug resistance (DR) remains a major challenge for tuberculosis (TB) control. Whole-genome sequencing (WGS) provides the highest genetic resolution for genotypic drug-susceptibility tests (DST). We compared DST profiles of 60 Mycobacterium tuberculosis isolates which were drug resistant according to agar proportion tests (one poly DR-TB, 34 multidrug-resistant TB and 25 extensively drug-resistant TB). We additionally performed minimum inhibitory concentration (MIC) tests using Sensititre MYCOTBI plates (MYCOTB) and a WGS-based DST. Agreement between WGS-based DST and MYCOTB was high for all drugs except ethambutol (65%) and ethionamide (62%). Isolates harboring the -15 c/t inhA promoter mutation had a significantly lower MIC for isoniazid than did isolates with the katG Ser315Thr mutation (p < 0.001). Similar patterns were seen for ethambutol (embB Gly406Asp vs. embB Met306Ile), streptomycin (gid Gly73Ala vs. rpsL Lys43Arg), moxifloxacin (gyrA Ala90Val vs. gyrA Asp94Gly) and rifabutin (rpoB Asp435Phe/Tyr/Val vs. rpoB Ser450Leu). For genotypic heteroresistance, isolates with lower proportion of mapped read tended to has lower MIC of anti-TB drugs than those with higher proportion. These results emphasize the high applicability of WGS for determination of DR-TB and the association of particular mutations with MIC levels.

Irreversible neuropathy in extremely-drug resistant tuberculosis: An unfortunate clinical conundrum.

Drug-resistant tuberculosis is an increasing healthcare challenge. Drug regimen building demands the use of different therapeutic groups, many of which harbor neurotoxicity as a side-effect, whether central or peripheral. Peripheral neuropathy is a major concern as it tends to be severe and usually irreversible. Anti-tubercular drugs that may contribute to peripheral neuropathy include INH, ethambutol, linezolid, cycloserine and para-amino salicylic acid. This potential adverse effect must be balanced against the intrinsically grave prognosis that drug resistant tuberculosis harbors. We present such a clinically challenging case of a 25 years-old female with extremely drug resistant tuberculosis whose treatment necessitated the use of several neurotoxic anti-tubercular drugs, leading to severe sensory peripheral neuropathy who did not improve despite the withdrawal of culprit drugs. She developed positive and negative sensory symptoms in both lower limbs. Nerve conduction studies were suggestive of sensory neuropathy affecting both lower limbs. Alternate causes of peripheral neuropathy including HIV, vasculitis, B12 deficiency and diabetes were ruled out. Despite drug withdrawal, the patient did not improve significantly. This case emphasizes the irreversibility of anti-tubercular therapy-induced peripheral neuropathy, demanding more rigorous clinical screening for the same while managing such patients.

Gitelman-like syndrome: A rare complication of using aminoglycosides in tuberculosis - A case report.

Aminoglycosides are known to cause electrolyte disturbances. Approximately 8-26% of patients who receive an aminoglycoside for several days develop mild renal impairment that is almost always reversible (Brunton et al., 2013). A 46 year old male with multi-drug-resistant pulmonary tuberculosis with resistance to kanamycin is being presented, who was on injectable Capreomycin, Levofloxacin, Ethionamide, Cycloserine, pyrazinamide, linezolid and clofazamine for a period of four months. He presented to us with generalised weakness and pain in the lower limb muscles. Investigation revealed hypokalemia, metabolic alkalosis, hypomagnesemia, hypocalceuria and hypocalcemia. This features mimic Gitelman's syndrome which is an autosomal recessive disorder affecting kidneys causing electrolyte disturbances. The drug was immediately withdrawn and electrolyte correction was given and the condition reversed gradually.

Bedaquilina para pacientes com tuberculose resistentes à rifampicina, multirresistentes e extensivamente resistentes a medicamentos / Bedaquiline for tuberculosis-resistant patients rifampicin, multi-resistant and extensively resistant to medicines

INTRODUÇÃO: A tuberculose (TB), conhecida anteriormente como tísica, é uma doença que pode ser causada por sete espécies do gênero do complexo Mycobacterium sendo a mais importante, do ponto de vista de saúde pública, a M. tuberculosis. Globalmente cerca de 10 milhões de pessoas tiveram TB no ano de 2018. No Brasil, em 2018, foram diagnosticados 72.788 casos novos de TB o que representa uma incidência de 34,8 casos por 100 mil habitantes. A TB pode ser classificada como pulmonar e extrapulmonar, sendo a primeira forma mais prevalente. Além disso, a TB pode ser classificada conforme a resistência à medicamentos, tais como: RR-TB, MDR-TB e XDR-TB. PERGUNTA DE PESQUISA: A bedaquilina (BDQ) associada ao tratamento padrão para pacientes adultos com RR-TB, MDR-TB ou XDR-TB, é mais eficaz, efetiva e segura comparado ao tratamento padrão utilizado pelo SUS (levofloxacino, moxifloxacino, amicacina, capreomicina, etionamida, terizidona, linezolida, clofazimina, pirazinamida, etambutol, isoniazida, rifampicina e paraminossalicílico) ou placebo? TECNOLOGIA: Bedaquilina (Sirturo®). EVIDÊNCIAS CIENTÍFICAS: A revisão sistematizada recuperou nove estudos (uma revisão sistemática [RS] com meta-análise em rede [network meta-analysis - NMA], um ensaio clínico randomizado [ECR] com dois relatos e sete estudos de coorte [seis retrospectivas e uma prospectiva]). A RS, com NMA, avaliou a BDQ em comparação aos medicamentos delamanida, metronidazol, moxifloxacino e levofloxacino. A RS avaliou os desfechos conversão de cultura do escarro e aceitabilidade, e não foram verificados resultados estatisticamente significantes. Os estudos de coorte avaliaram a BDQ em comparação aos mais diversos tratamentos disponíveis para RR-TB, MDR-TB e XDR-TB. As coortes avaliaram os seguintes desfechos: sobrevida sucesso no tratamento, tratamento completo, cura, conversão da cultura do escarro e mortalidade. Os resultados não foram estatisticamente significantes na meta-análise de modelo de efeitos randomizados para todos os desfechos avaliados, porém os resultados dos efeitos fixos demostraram resultados estatisticamente significantes favorecendo o tratamento com BDQ em comparação ao tratamento sem BDQ. Vale salientar que foram realizadas análises de subgrupos com o ECR, TMC207, que avaliou eficácia e segurança da BDQ associado ao tratamento padrão em comparação ao grupo placebo associado ao tratamento padrão em até 120 semanas para os desfechos de conversão da cultura do escarro, cura e segurança (mortalidade), porém não mudaram a direção dos resultados nas duas modelagem da meta-análise. AVALIAÇÃO ECONÔMICA (AE): Os tratamentos com BDQ comparado aos tratamentos do SUS mostraram-se dominados na avaliação de custo-efetividade, para o desfecho paciente curado. Assim, os tratamentos do SUS para RR-TB, MDR-TB e XDR-TB dominaram todos os tratamentos com BDQ, ou seja, todos os tratamentos com BDQ foram menos efetivos e mais caros que os tratamentos do SUS para obter a cura dos indivíduos com RR-TB, MDR-TB e XDR-TB. AVALIAÇÃO DE IMPACTO ORÇAMENTÁRIO (AIO): A AIO, para os pacientes com RR-TB, variou entre um custo incremental R$ 936 mil no caso base a uma economia de -R$ 1 milhão ao final do quinto ano no cenário alternativo; para MDR-TB variou entre uma economia de -R$44 mil no caso base a um gasto de R$ 110 mil ao final do quinto ano no cenário alternativo; e para XDR-TB variou entre um custo incremental de R$ 188 mil no caso base a R$ 4 mil no cenário alternativo ao final do quinto ano. MONITORAMENTO DO HORIZONTE TECNOLÓGICO (MHT): Cinco medicamentos foram detectados no MHT para pacientes com MDR-TB e XDR-TB (canamicina, cicloserina, sutezolida, pretomanide e protionamida). CONSIDERAÇÕES FINAIS: Há resultados conflitantes nas evidências encontradas no relatório. O ECR, analisado como de alto risco de viés (Risk of Bias 2.0) mostrou que a BDQ associada ao tratamento padrão é eficaz em comparação ao grupo de tratamento placebo associado ao tratamento padrão, porém com maior número de mortes e episódios de náusea em comparação ao grupo de tratamento sem a BDQ. Os resultados da RS, com NMA, de qualidade moderada, não demonstraram diferenças estatisticamente significantes entre as tecnologias avaliadas. Os resultados das meta-análises dos estudos de coorte de baixa qualidade metodológica (Newcastle-Ottawa Scale), em combinação com o ECR da BDQ, foram demonstrados em efeitos fixos e randomizados. Os desfechos sucesso no tratamento, tratamento completo, cura, conversão da cultura do escarro e mortalidade não foram estatisticamente significantes no modelo de efeito randomizados na meta-análise. No entanto, foram estatisticamente significantes no modelo de efeito fixos da metaanálise, e favoreceram o tratamento com BDQ em comparação aos pacientes não tratados sem BDQ. A AE demonstrou que os tratamentos com BDQ foram dominados em relação aos tratamentos disponibilizados no SUS sem BDQ, para o desfecho paciente tratado, sendo, portanto, mais custosos e menos efetivos. A AIO, para pacientes com RR-TB, variou entre R$ 936 mil no caso base a uma economia de -R$ 1 milhão no cenário alternativo ao final do quinto ano, para MDRTB variou entre uma economia de -R$44 mil no caso base a um custo de R$ 110 mil ao final do quinto ano no cenário alternativo e para XDR-TB variou entre um custo adicional de R$ 188 mil no caso base a um custo adicional de R$ 4 mil ao final do quinto ano no cenário alternativo. RECOMENDAÇÃO PRELIMINAR DA CONITEC: A Conitec, em sua 87ª reunião ordinária, realizada nos dias 03 e 04 de junho de 2020, deliberou que a matéria fosse disponibilizada em consulta pública com recomendação preliminar favorável à incorporação no SUS da bedaquilina para pacientes com tuberculose resistente à rifampicina (RR-TB), a tuberculose multirresistente (MDR-TB) e para tuberculose extensivamente resistente a medicamentos (XDR-TB), condicionada ao monitoramento e apresentação dos dados de vida real, efetividade e segurança, da utilização da bedaquilina pela população brasileira e conforme critérios estabelecidos em protocolo do Ministério da Saúde. CONSULTA PÚBLICA: A Consulta Pública nº 24/2020 foi realizada entre os dias 22/06/2020 a 13/07/2020. Foram recebidas 66 contribuições no total, das quais 19 (29%) foram pelo formulário para contribuições técnico-científicas e 47 (71%) pelo formulário para contribuições sobre experiência ou opinião de pacientes, familiares, amigos ou cuidadores de pacientes, profissionais de saúde ou pessoas interessadas no tema. Das 19 contribuições de cunho técnico-científico, 95% submeteram a contribuição com opinião concordando totalmente com a recomendação preliminar da comissão. Apenas uma contribuição discordou da recomendação preliminar da Conitec, mas foi uma contribuição equivocada e se tratava de outro tema de consulta pública, portanto, foi excluída da análise. Das 47 contribuições recebidas sobre experiência ou opinião, apenas 15 foram analisadas, pois 32 estavam em branco, se tratavam de outro tema ou foram preenchidas inadequadamente. As 15 contribuições remanescentes concordaram 100% com a decisão preliminar da comissão. Após a apreciação das contribuições encaminhadas na consulta pública nº 24/2020, o plenário da Conitec considerou que: I) Foi apresentado um novo preço de USD 340 da bedaquilina pela Johnson & Johnson, sendo proposto um desconto de 15% no preço utilizado no relatório de recomendação preliminar (USD 400); II) Foram enviadas novas estimativas de incidência para pacientes com tuberculose multirresistente, bem como evidência de possíveis limitações na análise de impacto orçamentário; III) A nova análise de impacto orçamentário, utilizando os novos parâmetros enviados na consulta pública, aponta para economia de recursos na população com tuberculose multirresistente e um custo incremental com tuberculose resistente à rifampicina e tuberculose extensivamente resistente no cenário sem taxa de difusão gradual da bedaquilina (100% no primeiro ano de incorporação). No entanto, ao adotarmos o cenário com taxa difusão gradual da bedaquilina, 30% no primeiro ano de incorporação a 70% no quinto ano, os resultados mudam e proporcionam economia de recursos para pacientes com tuberculose resistente à rifampicina e um custo incremental para pacientes com tuberculose multirresistente e tuberculose extensivamente resistente. RECOMENDAÇÃO FINAL DA CONITEC: Os membros da Conitec presentes na 89ª reunião ordinária, no dia 05 de agosto de 2020, deliberaram por unanimidade recomendar a incorporação da bedaquilina para pacientes com tuberculose resistentes à rifampicina, multirresistentes e extensivamente resistente a medicamentos, condicionado a apresentação de dados de vida real e conforme preconizado pelo Ministério da Saúde. Foi assinado o Registro de Deliberação nº 538/2020. DECISÃO: Incorporar a bedaquilina para pacientes com tuberculose resistentes à rifampicina, multirresistentes e extensivamente resistente a medicamentos, condicionado a apresentação de dados de vida real e conforme preconizado pelo Ministério da Saúde, no âmbito do Sistema Único de Saúde - SUS, conforme Portaria nº 36, publicada no Diário Oficial da União nº 168, seção 1, página 77, em 01 de setembro de 2020.

Ethionamide Population Pharmacokinetic Model and Target Attainment in Multidrug-Resistant Tuberculosis.

Ethionamide (ETA), an isonicotinic acid derivative, is part of the multidrug-resistant tuberculosis (MDR-TB) regimen. The current guidelines have deprioritized ETA because it is potentially less effective than other agents. Our aim was to develop a population pharmacokinetic (PK) model and simulate ETA dosing regimens in order to assess target attainment. This study included subjects from four different sites, including healthy volunteers and patients with MDR-TB. The TB centers included were two in the United States and one in Bangladesh. Patients who received ETA and had at least one drug concentration reported were included. The population PK model was developed, regimens with a total of 1,000 to 2,250 mg daily were simulated, and target attainment using published MICs and targets of 1.0-log kill and resistance suppression was assessed with the Pmetrics R package. We included 1,167 ethionamide concentrations from 94 subjects. The final population model was a one-compartment model with first-order elimination and absorption with a lag time. The mean (standard deviation [SD]) final population parameter estimates were as follows: absorption rate constant, 1.02 (1.11) h-1; elimination rate constant, 0.69 (0.46) h-1; volume of distribution, 104.16 (59.87) liters; lag time, 0.43 (0.32) h. A total daily dose of 1,500 mg or more was needed for ≥90% attainment of the 1.0-log kill target at a MIC of 1 mg/liter, and 2,250 mg/day led to 80% attainment of the resistance suppression target at a MIC of 0.5 mg/liter. In conclusion, we developed a population PK model and assessed target attainment for different ETA regimens. Patients may not be able to tolerate the doses needed to achieve the predefined targets supporting the current recommendations for ETA deprioritization.

Is hypothyroidism rare in multidrug resistance tuberculosis patients on treatment? A systematic review and meta-analysis.

BACKGROUND: Hypothyroidism is one of the adverse drug reactions that associated with Multidrug Resistant Tuberculosis (MDR-TB) medications. Extremely variable magnitude of hypothyroidism in MDR-TB patients has been reported from different parts of the world. However, there is no evidence that tried to estimate the pooled prevalence of hypothyroidism to confirm the rareness of hypothyroidism in MDR-TB patients on treatment. Therefore, we did a systematic review and meta-analysis to estimate the prevalence of hypothyroidism in MDR-TB patients on treatment, and to summarize the demographic and clinical characteristics of the patients. METHODS: We conducted a systematic review and meta-analysis on studies reported around the world on the prevalence of hypothyroidism in MDR-TB patients on treatment. We searched electronic databases: PubMed/Medline, EMBASE, CINAHL, Science Direct, Academic Search Complete and Google scholar for English language articles without limiting publication year. We also reviewed the bibliographies of relevant studies and conducted an electronic search for relevant conference abstracts. Eligible studies were cross-sectional and cohort studies that included at least five participants. We used a random-effects model to estimate the pooled prevalence of hypothyroidism. The registration number of this review study protocol is CRD42018109237. RESULTS: We included 30 studies and pooled data on a total of 6,241 MDR-TB patients. The crude prevalence of hypothyroidism was extremely heterogeneous. The pooled prevalence of hypothyroidism in MDR-TB patients on treatment was 17.0% (95% CI: 13.0-20.0). Ethionamide and para-aminosalicylic acid (PAS) were the most frequently reported drugs that associated with the occurrence of hypothyroidism. CONCLUSION: This review revealed that hypothyroidism is not a rare adverse drug reaction in MDR-TB patients on treatment. Ethionamide and PAS were the most frequently reported drugs that associated with the occurrence of hypothyroidism. Screening of hypothyroidism in MDR-TB patients on treatment is important while targeting patients on Ethionamide and PAS based treatment regimen.

Initial experience of bedaquiline implementation under the National TB Programme at NITRD, Delhi, India.

BACKGROUND: Bedaquiline (BDQ) was approved for treatment of drug resistant TB (DR-TB) under Conditional Access Programme (CAP) of Revised National Tuberculosis Control Programme (RNTCP) and was also implemented in the National Institute of TB and Respiratory Diseases (NITRD). We present early efficacy and safety of BDQ containing regimens for DR-TB. OBJECTIVE: To ascertain the early efficacy and safety of Bedaquline containing regimens in treatment of DR-TB. METHODS: BDQ containing regimens along with other drugs were designed as per WHO recommendations for DR-TB patients. They were followed up for sputum smear and culture conversion, adverse events during the treatment. RESULTS: A cohort of 290 DR-TB patients (Median age-29.77) were initiated on BDQ containing regimens. Of the available Sputum results, smear conversion was seen in 51% and 91% patients at the end of 1st week and 3rd month respectively. Similarly, 93% and 98% patients had culture conversion at the end of 3rd and 6th month respectively. 201 adverse events (AE) including 47 deaths were reported among 109 patients. QTc prolongation was seen in 29% patients but only 4 required discontinuation of BDQ. Lost to follow up of treatment was about 6%. CONCLUSION: Bedaquiline along with an optimized background regimen has shown early sputum conversion in larger number of difficult to treat patients having additional resistance of second line drugs along with INH and Rifampicin. The regimen is feasible in programmatic conditions and is relatively safe.

Potentially High Number of Ineffective Drugs with the Standard Shorter Course Regimen for Multidrug-Resistant Tuberculosis Treatment in Haiti.

Multidrug-resistant tuberculosis (MDR-TB) outcomes are poor partly because of the long treatment duration; the World Health Organization conditionally recommends a shorter course regimen to potentially improve treatment outcomes. Here, we describe the drug susceptibility patterns of a cohort of MDR-TB patients in Haiti and determine the number of likely effective drugs if they were treated with the recommended shorter course regimen. We retrospectively examined drug susceptibility patterns of adults initiating MDR-TB treatment between 2008 and 2015 at the Haitian Group for the Study of Kaposi's Sarcoma and Opportunistic Infections in Port-au-Prince, Haiti. First- and second-line drug susceptibility testing (DST) was analyzed and used to determine the number of presumed effective drugs. Of the 239 patients analyzed, 226 (95%), 183 (77%), 135 (57%), and 38 (16%) isolates were resistant to high-dose isoniazid, ethambutol, pyrazinamide, and ethionamide, respectively. Eight patients (3%) had resistance to either a fluoroquinolone or a second-line injectable and none had extensively resistant TB. Of the 239 patients, 132 (55%) would have fewer than five likely effective drugs in the intensive phase of the recommended shorter course regimen and 121 (51%) would have two or fewer likely effective drugs in the continuation phase. Because of the high rates of resistance to first-line TB medications, about 50% of MDR-TB patients would be left with only two effective drugs in the continuation phase of the recommended shorter course regimen, raising concerns about the effectiveness of this regimen in Haiti and the importance of using DST to guide treatment.

Improved Treatment Outcomes With Bedaquiline When Substituted for Second-line Injectable Agents in Multidrug-resistant Tuberculosis: A Retrospective Cohort Study.

BACKGROUND: Bedaquiline is used as a substitute for second-line injectable (SLI) intolerance in the treatment of multidrug-resistant (MDR) tuberculosis, but the efficacy and safety of this strategy is unknown. METHODS: In this retrospective cohort study adults receiving bedaquiline substitution for MDR tuberculosis therapy, plus a matched control group who did not receive bedaquiline, were identified from the electronic tuberculosis register in the Western Cape Province, South Africa. The primary outcome measure was the proportion of patients with death, loss to follow-up, or failure to achieve sustained culture conversion at 12 months of treatment. RESULTS: Data from 162 patients who received bedaquiline substitution and 168 controls were analyzed; 70.6% were infected with human immunodeficiency virus. Unfavorable outcomes occurred in 35 of 146 (23.9%) patients in the bedaquiline group versus 51 of 141 (36.2%) in the control group (relative risk, 0.66; 95% confidence interval, .46 -.95). The number of patients with culture reversion was lower in those receiving bedaquiline (1 patient; 0.8%) than in controls (12 patients; 10.3%; P = .001). Delayed initiation of bedaquiline was independently associated with failure to achieve sustained culture conversion (adjusted odds ratio for every 30-day delay, 1.5; 95% confidence interval, 1.1-1.9). Mortality rates were similar at 12 months (11 deaths in each group; P = .97). CONCLUSIONS: Substituting bedaquiline for SLIs in MDR tuberculosis treatment resulted in improved outcomes at 12 months compared with patients who continued taking SLIs, supporting the use of bedaquiline for MDR tuberculosis treatment in programmatic settings.

Integrated analysis of ethionamide resistance loci in Mycobacterium tuberculosis clinical isolates.

Tuberculosis patients taking second line drugs such as ethionamide (ETH) have often experienced previous treatment failure and usually have a complex history of disease and treatment that can span decades. Mutations in the ETH activating enzyme, EthA, confer resistance through undescribed mechanisms. To explore the impact of EthA mutations on ETH resistance, data from a total of 160 ETHR isolates was analysed. The most frequently mutated positions are within regions that display sequence conservation with the active site of OTEMO, another FAD-containing NADH-binding Baeyer-Villiger monooxygenase (BVMO), or with the sugar binding site of galectin-4N. Additionally, to look at a possible role of EthR on ETH resistance we purified an EthR mutant identified in a clinical isolate, F110L, and found it to bind the ethA-ethR intergenic region with higher affinity than the wild type regulator in gel shift assays. The ability of cyclic di-GMP to enhance DNA binding is maintained in the EthR mutant. To our knowledge, this is the first ETH resistance study that combines sequence and resistance data of clinical isolates with functional and structural information.