Search Results (21)

Oxazolidinones, novel synthetic antibacterials, inhibit protein biosynthesis and show potent activity against Gram-positive bacteria, including Mycobacterium tuberculosis (MTB). In this study, we aimed to compare the in vitro activity of linezolid (LZD) and four oxazolidinones, including tedizolid (TZD), contezolid (CZD), sutezolid (SZD), and delpazolid (DZD), against multidrug-resistant tuberculosis (MDR-TB) and pre-extensively drug-resistant tuberculosis (pre-XDR-TB) isolates from Hainan. We established their epidemiological cut-off values (ECOFFs) using ECOFFinder software and analyzed mutations in rrl (23S rRNA), rplC, rplD, mce3R, tsnR, Rv0545c, Rv0930, Rv3331, and Rv0890c genes to uncover potential mechanisms of oxazolidinone resistance. This study included 177 MTB isolates, comprising 67 MDR and 110 pre-XDR-TB isolates. Overall, SZD exhibited the strongest antibacterial activity against clinical MTB isolates, followed by TZD and LZD, with CZD and DZD showing equivalent but weaker activity (SZD_MIC50 = TZD_MIC50 < LZD_MIC50 < CZD_MIC50 = DZD_MIC50; SZD_MIC90 < TZD_MIC90 = LZD_MIC90 < CZD_MIC90 = DZD_MIC90). Significant differences in MIC distribution were observed for TZD (p < 0.0001), CZD (p < 0.01), SZD (p < 0.0001), and DZD (p < 0.0001) compared to LZD but not between MDR-TB and pre-XDR-TB isolates. We propose the following ECOFFs: SZD, 0.5 µg/mL; LZD, TZD, and CZD, 1.0 µg/mL; DZD, 2.0 µg/mL. No statistically significant differences in resistance rates were observed among these five drugs (p > 0.05). We found that eight MTB isolates (4.52% [8/177]) resisted these five oxazolidinones. Among these, only one isolate, M26, showed an amino acid substitution (Arg79His) in the protein encoded by the rplD gene, which conferred cross-resistance to TZD and CZD. Three distinct mutations were identified in the mce3R gene; notably, isolate P604 displayed two insertions that contributed to resistance against all five oxazolidinones. However, no significant correlation was observed between mutations in the rrl, rplC, rplD, mce3R, tsnR, Rv0545c, Rv0930, Rv3331, and Rv0890c genes with oxazolidinone resistance in the clinical MTB isolates tested. In summary, this study provides the first report on the resistance of MTB in Hainan to the five oxazolidinones (LZD, TZD, CZD, SZD, and DZD). In vitro susceptibility testing indicated that SZD exhibited the strongest antibacterial activity, followed by TZD and LZD, while CZD and DZD demonstrated comparable but weaker effectiveness. Mutations in rplD and mce3R were discovered, but further research is needed to clarify their role in conferring oxazolidinone resistance in MTB. Full article

Extensively drug-resistant tuberculosis (XDR-TB) is a public health concern as drug resistance is outpacing the drug development pipeline. Alternative immunotherapeutic approaches are needed. Peripheral blood mononuclear cells (PBMCs) were isolated from pre-XDR/XDR-TB (n = 25) patients and LTBI (n = 18) participants. Thereafter, monocytic-derived dendritic cells (mo-DCs) were co-cultured with M. tb antigens, with/without a maturation cocktail (interferon-γ, interferon-α, CD40L, IL-1β, and TLR3 and TLR7/8 agonists). Two peptide pools were evaluated: (i) an ECAT peptide pool (ESAT6, CFP10, Ag85B, and TB10.4 peptides) and (ii) a PE/PPE peptide pool. Sonicated lysate of the M. tb HN878 strain served as a control. Mo-DCs were assessed for DC maturation markers, Th1 cytokines, and the ability of the DC-primed PBMCs to restrict the growth of M. tb-infected monocyte-derived macrophages. In pre-XDR/XDR-TB, mo-DCs matured with M. tb antigens (ECAT or PE/PPE peptide pool, or HN878 lysate) + cocktail, compared to mo-DCs matured with M. tb antigens only, showed higher upregulation of co-stimulatory molecules and IL-12p70 (p < 0.001 for both comparisons). The matured mo-DCs had enhanced antigen-specific CD8⁺ T-cell responses to ESAT-6 (p = 0.05) and Ag85B (p = 0.03). Containment was higher with mo-DCs matured with the PE/PPE peptide pool cocktail versus mo-DCs matured with the PE/PPE peptide pool (p = 0.0002). Mo-DCs matured with the PE/PPE peptide pool + cocktail achieved better containment than the ECAT peptide pool + cocktail [50%, (IQR:39–75) versus 46%, (IQR:15–62); p = 0.02]. In patients with pre-XDR/XDR-TB, an effector response primed by mo-DCs matured with an ECAT or PE/PPE peptide pool + cocktail was capable of restricting the growth of M. tb in vitro. Full article

Background/Objectives: Tuberculosis (TB) remains a significant global health challenge, with drug-resistant tuberculosis (DR-TB) posing a greater threat due to difficulty in treatment. This study aimed to investigate the relationship between comorbidities and treatment outcomes in patients diagnosed with DR-TB in rural Eastern Cape using logistic regression. Methods: Data on patient characteristics, comorbidities, and treatment outcomes were extracted from the medical records and analyzed using Python version 3.8. and R version 4.1.1 software. A logistic regression model was used to determine the effects of selected variables on treatment outcomes of DR-TB cases. Results: Hearing loss and hypertension (HTN) were the most frequently observed comorbidities across various DR-TB cases, particularly rifampicin-resistant (RR), multidrug-resistant (MDR), and pre-extensively drug-resistant (pre-XDR-TB) cases. A hearing loss prevalence of 5.8% (26/445) was found among patients receiving treatment for TB, with the intensity of impairment ranging from mild to severe. Gender is significantly associated with the occurrence of HTN among these patients (p-value: 0.022). Comorbidities such as epilepsy, hearing loss, and HTN significantly impact treatment success, with higher risks of mortality and incomplete cure. Using logistic regression, obesity (OR = 3.0884; e = 1.1277; p = 0.0408) and HIV-positive status (OR = 0.4458; e = 0.8078; p = 0.0001) were highly likely and less likely associated with better treatment outcomes, respectively. The logistic regression model achieved an accuracy of 64.0%, a precision of 63.0%, and a recall of 95.0%, with an F-1 score of 76.0%. Conclusions: The findings underscore the importance of implementing integrated management strategies that address both DR-TB and its comorbidities, particularly in resource-limited settings where such patients are prevalent. Public health policies should incorporate strategies to provide nutritional assessments and interventions, particularly for individuals with low BMI. This could include food supplementation programs or partnerships with local food kitchens to ensure that patients have access to adequate nutrition during DR-TB treatment. Full article

The management of resistant tuberculosis (tb) can be extremely difficult, especially in case of novel unpredicted complications. In this report, we present a case of a 48-year-old patient with pre-extensively drug-resistant (XDR) tb who received a treatment regimen including pretomanid, bedaquiline, linezolid, cycloserine, and amikacin and died due to myocarditis. Acquired resistance to first- and second-line drugs developed due to previous poor adherence to medication. The clinical presentation of the patient, along with her initial ultrasonographical, electrocardiogram (ECG), and laboratory examinations, were typical for acute myocarditis; however, the patient was considered unstable, and further investigations, including magnetic resonance imaging (MRI), pericardiocentesis, and endomyocardial biopsy were not performed. To our knowledge, this is the first case of myocarditis in such a patient, the clinical features of which raised a high suspicion of drug induction that could be attributed to the treatment regimen that was administered. Clinicians who manage cases of drug-resistant tb should be aware of this newly reported, potentially lethal, adverse event. Full article

A previous study in Pará, Northern Brazil, described a strain of Mycobacterium tuberculosis with a unique genotype (SIT2517/T1) associated with multidrug-resistant tuberculosis (MDR-TB). To improve our understanding of MDR-TB transmission dynamics of these strains within this region, we performed phenotypic and genotypic drug susceptibility testing (pDST/gDST), 24-loci mycobacterial interspersed repetitive units (MIRU-VNTR) genotyping, whole-genome sequencing (WGS) and geo-epidemiology analysis. Of the 28 SIT2517/T1 isolates, 19 (67.9%) could be genotyped by 24-loci MIRU-VNTR and 15 by WGS. All belonged to sublineage 4.1.1.3, distinct from other representative Lineage 4 isolates identified in Brazil. The MDR phenotype determined by pDST was confirmed by gDST, the latter also demonstrating the presence of additional mutations conferring pre-extensively drug-resistance (pre-XDR). Discrepancies between gDST and pDST were observed for pyrazinamide and fluoroquinolones. Thirteen out of 15 isolates analyzed by WGS were clustered when applying a 12 single nucleotide polymorphisms (SNPs) cutoff. The SIT2517/T1 isolates were distributed across the metropolitan regions of Belém and Collares municipalities, showing no geographic clustering. WGS-transmission network analysis revealed a high likelihood of direct transmission and the formation of two closely linked transmission chains. This study highlights the need to implement TB genomic surveillance in the Brazilian Amazon region. Full article

The present study aimed to determine the genetic diversity of isolates of Mycobacterium tuberculosis (Mtb) from presumed drug-resistant tuberculosis patients from several states of Brazil. The isolates had been submitted to conventional drug susceptibility testing for first- and second-line drugs. Multidrug-resistant (MDR-TB) (54.8%) was the most frequent phenotypic resistance profile, in addition to an important high frequency of pre-extensive resistance (p-XDR-TB) (9.2%). Using whole-genome sequencing (WGS), we characterized 298 Mtb isolates from Brazil. Besides the analysis of genotype distribution and possible correlations between molecular and clinical data, we determined the performance of an in-house WGS pipeline with other online pipelines for Mtb lineages and drug resistance profile definitions. Sub-lineage 4.3 (52%) was the most frequent genotype, and the genomic approach revealed a p-XDR-TB level of 22.5%. We detected twenty novel mutations in three resistance genes, and six of these were observed in eight phenotypically resistant isolates. A cluster analysis of 170 isolates showed that 43.5% of the TB patients belonged to 24 genomic clusters, suggesting considerable ongoing transmission of DR-TB, including two interstate transmissions. The in-house WGS pipeline showed the best overall performance in drug resistance prediction, presenting the best accuracy values for five of the nine drugs tested. Significant associations were observed between suffering from fatal disease and genotypic p-XDR-TB (p = 0.03) and either phenotypic (p = 0.006) or genotypic (p = 0.0007) ethambutol resistance. The use of WGS analysis improved our understanding of the population structure of MTBC in Brazil and the genetic and clinical data correlations and demonstrated its utility for surveillance efforts regarding the spread of DR-TB, hopefully helping to avoid the emergence of even more resistant strains and to reduce TB incidence and mortality rates. Full article

The treatment of drug-resistant Mycobacterium tuberculosis relies on complex antibiotic therapy. Inadequate antibiotic exposure can lead to treatment failure, acquired drug resistance, and an increased risk of adverse events. Therapeutic drug monitoring (TDM) can be used to optimize the antibiotic exposure. Therefore, we aimed to develop a single-run multiplex assay using high-performance liquid chromatography–mass spectrometry (HPLC–MS) for TDM of patients with multidrug-resistant, pre-extensively drug-resistant and extensively drug-resistant tuberculosis. A target profile for sufficient performance, based on the intended clinical application, was established and the assay was developed accordingly. Antibiotics were analyzed on a zwitterionic hydrophilic interaction liquid chromatography column and a triple quadrupole mass spectrometer using stable isotope-labeled internal standards. The assay was sufficiently sensitive to monitor drug concentrations over five half-lives for rifampicin, rifabutin, levofloxacin, moxifloxacin, bedaquiline, linezolid, clofazimine, terizidone/cycloserine, ethambutol, delamanid, pyrazinamide, meropenem, prothionamide, and para-amino salicylic acid (PAS). Accuracy and precision were sufficient to support clinical decision making (≤±15% in clinical samples and ±20–25% in spiked samples, with 80% of future measured concentrations predicted to fall within ±40% of nominal concentrations). The method was applied in the TDM of two patients with complex drug-resistant tuberculosis. All relevant antibiotics from their regimens could be quantified and high-dose therapy was initiated, followed by microbiological conversion. In conclusion, we developed a multiplex assay that enables TDM of the relevant first- and second-line anti-tuberculosis medicines in a single run and was able to show its applicability in TDM of two drug-resistant tuberculosis patients. Full article

Kazakhstan ranks among the countries with the highest number of MDR-TB patients per 100,000 population worldwide. The successful transmission of local MDR strains of Mycobacterium tuberculosis (Mtb) poses a significant threat to disease control. In this study, we employed whole-genome sequencing to examine drug resistance, compensatory mutations, population structure, and transmission patterns in a sample of 24 clinical isolates of L2/Beijing Mtb collected in Astana, Kazakhstan between 2021 and 2022. The genotypic prediction of Mtb susceptibility to anti-TB agents was consistent with the phenotypic susceptibility, except for bedaquiline. An analysis of resistance-associated genes characterized most of the isolates as pre-extensively drug-resistant tuberculosis (pre-XDR-TB) (n = 15; 62.5%). The phylogenetic analysis grouped the isolates into four transmission clusters; the dominant cluster was assigned to the “aggressive” Central Asia outbreak (CAO) clade of L2/Beijing (n = 15; 62.5%). Thirteen mutations with putative compensatory effects were observed exclusively in Mtb isolates containing the rpoB S450L mutation. The putative compensatory mutations had a stabilizing effect on RpoABC protein stability and dynamics. The high prevalence of the CAO clade in the population structure of Mtb may explain the rapid spread of MDR-TB in Kazakhstan. Full article

The emergence of pre-extensively drug-resistant tuberculosis (pre-XDR-TB) is a threat to TB control programs in developing countries such as Zambia. Studies in Zambia have applied molecular techniques to understand drug-resistance-associated mutations, circulating lineages and transmission patterns of multi-drug-resistant (MDR) Mycobacterium tuberculosis. However, none has reported genotypes and mutations associated with pre-XDR TB. This study characterized 63 drug-resistant M. tuberculosis strains from the University Teaching Hospital between 2018 and 2019 using targeted gene sequencing and conveniently selected 50 strains for whole genome sequencing. Sixty strains had resistance mutations associated to MDR, one polyresistant, and two rifampicin resistant. Among MDR strains, seven percent (4/60) had mutations associated with pre-XDR-TB. While four, one and nine strains had mutations associated with ethionamide, para-amino-salicylic acid and streptomycin resistances, respectively. All 50 strains belonged to lineage 4 with the predominant sub-lineage 4.3.4.2.1 (38%). Three of four pre-XDR strains belonged to sub-lineage 4.3.4.2.1. Sub-lineage 4.3.4.2.1 strains were less clustered when compared to sub-lineages L4.9.1 and L4.3.4.1 based on single nucleotide polymorphism differences. The finding that resistances to second-line drugs have emerged among MDR-TB is a threat to TB control. Hence, the study recommends a strengthened routine drug susceptibility testing for second-line TB drugs to stop the progression of pre-XDR to XDR-TB and improve patient treatment outcomes. Full article

This study aimed to determine phenotypic and genotypic drug resistance patterns of Mycobacterium tuberculosis strains from children with tuberculosis (TB) in China and Russia, two high-burden countries for multi/extensively-drug resistant (MDR/XDR) TB. Whole-genome sequencing data of M. tuberculosis isolates from China (n = 137) and Russia (n = 60) were analyzed for phylogenetic markers and drug-resistance mutations, followed by comparison with phenotypic susceptibility data. The Beijing genotype was detected in 126 Chinese and 50 Russian isolates. The Euro-American lineage was detected in 10 Russian and 11 Chinese isolates. In the Russian collection, the Beijing genotype and Beijing B0/W148-cluster were dominated by MDR strains (68% and 94%, respectively). Ninety percent of B0/W148 strains were phenotypically pre-XDR. In the Chinese collection, neither of the Beijing sublineages was associated with MDR/pre-XDR status. MDR was mostly caused by low fitness cost mutations (rpoB S450L, katG S315T, rpsL K43R). Chinese rifampicin-resistant strains demonstrated a higher diversity of resistance mutations than Russian isolates (p = 0.003). The rifampicin and isoniazid resistance compensatory mutations were detected in some MDR strains, but they were not widespread. The molecular mechanisms of M. tuberculosis adaptation to anti-TB treatment are not unique to the pediatric strains, but they reflect the general situation with TB in Russia and China. Full article

Background: Kazakhstan has a high burden of multidrug-resistant tuberculosis in the Central Asian region. This study aimed to perform genomic characterization of Mycobacterium tuberculosis strains obtained from Kazakhstani patients with pre-extensively drug-resistant tuberculosis diagnosed in Kazakhstan. Methods: Whole-genome sequencing was performed on 10 pre-extensively drug-resistant M. tuberculosis strains from different regions of Kazakhstan. All strains had high-confidence resistance mutations according to the resistance grading system previously established by the World Health Organization. The genome analysis was performed using TB-Profiler, Mykrobe, CASTB, and ResFinder. Results: Valuable information for understanding the genetic diversity of tuberculosis in Kazakhstan can also be obtained from whole-genome sequencing. The results from the Phenotypic Drug Susceptibility Testing (DST) of bacterial strains were found to be consistent with the drug resistance information obtained from genomic data that characterized all isolates as pre-XDR. This information can help in developing targeted prevention and control strategies based on the local epidemiology of tuberculosis. Furthermore, the data obtained from whole-genome sequencing can help in tracing the transmission pathways of tuberculosis and facilitating early detection of outbreaks. Conclusions: The results from whole-genome sequencing of tuberculosis clinical samples in Kazakhstan provide important insights into the drug resistance patterns and genetic diversity of tuberculosis in the country. These results can contribute to the improvement of tuberculosis control and management programs in Kazakhstan. Full article

Without the proper information on pyrazinamide (PZA) susceptibility of Mycobacterium tuberculosis (MTB), PZA is inappropriately recommended for the treatment of both susceptible and multidrug-resistant tuberculosis (MDR-TB) in Nepal. This study aimed to collect information regarding PZA susceptibility in MTB isolates from Nepal by analyzing pncA and its upstream regulatory region (URR). A total of 211 MTB isolates were included in this study. Sequence analysis of pncA and its URR was performed to assess PZA resistance. First-line drug susceptibility testing, spoligotyping, and sequence analysis of rpoB, katG, the inhA regulatory region, gyrA, gyrB, and rrs were performed to assess their association with pncA mutation. Sequencing results reveal that 125 (59.2%) isolates harbored alterations in pncA and its URR. A total of 57 different mutation types (46 reported and 11 novel) were scattered throughout the whole length of the pncA gene. Eighty-seven isolates (41.2%) harbored mutations in pncA, causing PZA resistance in MTB. There was a more significant association of pncA alterations in MDR/pre-extensively drug-resistant (Pre-XDR) TB than in mono-resistant/pan-susceptible TB (p < 0.005). This first report on the increasing level of PZA resistance in DR-TB in Nepal highlights the importance of PZA susceptibility testing before DR-TB treatment. Full article

The present study aimed at analyzing the treatment outcomes and risk factors associated with fluoroquinolone drug resistance having mutations in the gyrA and gyrB genes. A total of 258 pulmonary tuberculosis samples with first-line drug-resistant (H, R, or HR) were subjected to GenoType MTBDRsl assay for the molecular detection of mutations. Among the 258 samples, 251 were drug-resistant tuberculosis and seven were sensitive to all first-line TB drugs. Out of 251 DR-TB cases, 42 cases were MDR TB, 200 were INH mono-resistant and nine cases were RIF mono-resistant tuberculosis. Out of 251 DR-TB cases performed with a MTBDRsl assay, 14 had Pre-XDR-FQ, one patient had pre-XDR-SLID, one had extensively drug-resistant tuberculosis (XDR-TB) and 235 cases were sensitive to both FQ and SLID drugs. The study group had a mean average of 42.7 ± 16.4 years. The overall successful treatment outcomes among the MDR, INH mono-resistant, and pre-XRD patients were 70.6%, 82.0%, and 51%, respectively. The percentage of risk for the unfavorable outcomes in the pre-XDR, INH -mono-resistant, and XDR cases were 113.84% increased risk with RR 2.14; 95% CI 0.7821–5.8468. The independent risk factor associated with the unfavorable outcomes to failure was 77.78% increased risk with RR 1.78; 95% CI 0.3375–9.3655. Logistic regression analysis revealed that the percentage relative risk among MDR-TB patients for gender, male (RR: 1.85), age ≥ 61 years (RR: 1.96), and diabetics (RR: 1.05) were 84.62%, 95.83%, and 4.76%, respectively. The independent risk factors associated with INH mono-resistant cases of age 16–60 (RR: 1.86), ≥61 year (RR: 1.18), and treated cases (RR: 5.06). This study presaged the significant risk of INH mono-resistant, pre-XDR, and MDR among males, young adults, diabetics, and patients with previous treatment failure. Timely identification of high-risk patients will give pronounced advantages to control drug resistance tuberculosis diseases. Full article

Tuberculosis (TB) remains a life-threatening infection, and it is well-known that effective TB treatment is associated with multiple drugs administered to infected patients on a daily basis. Terizidone (TZD) is an anti-TB drug used in the treatment of multi-drug resistant and extensively drug-resistant TB but presents with polyneuropathic adverse effects in some patients. To counteract these adverse effects, TZD is typically prescribed with pyridoxine (PDX), well known as Vitamin B₆. As part of a pre-formulation study investigating the potential to co-formulate these two compounds, it became necessary to have a simple and reliable reversed-phase high-performance liquid chromatography (RP-HPLC) method. Optimal, simultaneous separation and detection of TZD and PDX were obtained using an isocratic mobile phase setup, consisting of ultrapure water and acetonitrile (30:70% v/v), with 1 mL glacial acetic acid added to the mobile phase mixture. A Discovery^® C₁₈, 150 × 4.6 mm, 5 μm column maintained at ambient temperature was utilized, with a detection wavelength of 260 nm. The method was validated in terms of linearity, accuracy, precision, limit of detection (LOD), limit of quantification (LOQ), specificity, robustness, and solution stability. Validation proved this method to be acceptable and reliable for the simultaneous accurate detection and quantification of TZD and PDX. Full article

Tuberculosis causes more than 1.2 million deaths each year. Worldwide, it is the first cause of death by a single infectious agent. The emergence of drug-resistant strains has limited pharmacological treatment of the disease and today, new drugs are urgently needed. Semi-synthetic mulinanes have previously shown important activity against multidrug-resistant (MDR) Mycobacterium tuberculosis. In this investigation, a new set of semi-synthetic mulinanes were synthetized, characterized, and evaluated for their in vitro activity against three drug-resistant clinical isolates of M. tuberculosis: MDR, pre-extensively Drug-Resistant (pre-XDR), and extensively Drug-Resistant (XDR), and against the drug-susceptible laboratory reference strain H37Rv. Derivative 1a showed the best anti-TB activity (minimum inhibitory concentration [MIC] = 5.4 µM) against the susceptible strain and was twice as potent (MIC = 2.7 µM) on the MDR, pre-XDR, and XDR strains and also possessed a bactericidal effect. Derivative 1a was also tested for its anti-TB activity in mice infected with the MDR strain. In this case, 1a produced a significant reduction of pulmonary bacilli loads, six times lower than the control, when tested at 0.2536 mg/Kg. In addition, 1a demonstrated an adjuvant effect by shortening second-line chemotherapy. Finally, the selectivity index of >15.64 shown by 1a when tested on Vero cells makes this derivative an important candidate for future studies in the development of novel antitubercular agents. Full article