Multidrug-resistant Mycobacterium Tuberculosis Research Articles

Characteristic SNPs defining the major multidrug-resistant Mycobacterium tuberculosis clusters identified by EuSeqMyTB to support routine surveillance, EU/EEA, 2017 to 2019.

BackgroundThe EUSeqMyTB project, conducted in 2020, used whole genome sequencing (WGS) for surveillance of drug-resistant Mycobacterium tuberculosis in the European Union/European Economic Area (EU/EEA) and identified 56 internationally clustered multidrug-resistant (MDR) tuberculosis (TB) clones.AimWe aimed to define and establish a rapid and computationally simple screening method to identify probable members of the main cross-border MDR-TB clusters in WGS data to facilitate their identification and track their future spread.MethodsWe screened 34 of the larger cross-border clusters identified in the EuSeqMyTB pilot study (2017-19) for characteristic single nucleotide polymorphism (SNP) signatures that could identify and define members of each cluster. We also linked this analysis with published clusters identified in previous studies and identified more distant genetic relationships between some of the current clusters.ResultsA panel of 30 characteristic SNPs is presented that can be used as an initial (routine) screen for members of each cluster. For four of the clusters, no unique defining SNP could be identified; three of these are closely related (within approximately 20 SNPs) to one or more other clusters and likely represent a single established MDR-TB clade composed of multiple recent subclusters derived from the previously described ECDC0002 cluster.ConclusionThe identified SNP signatures can be integrated into routine pipelines and contribute to the more effective monitoring, rapid and widespread screening for TB. This SNP panel will also support accurate communication between laboratories about previously identified internationally transmitted MDR-TB genotypes.

Bacteriophage therapy for the treatment of Mycobacterium tuberculosis infections in humanized mice

The continuing emergence of new strains of antibiotic-resistant bacteria has renewed interest in phage therapy; however, there has been limited progress in applying phage therapy to multi-drug resistant Mycobacterium tuberculosis (Mtb) infections. In this study, we show that bacteriophage strains D29 and DS6A can efficiently lyse Mtb H37Rv in 7H10 agar plates. However, only phage DS6A efficiently kills H37Rv in liquid culture and in Mtb-infected human primary macrophages. We further show in subsequent experiments that, after the humanized mice were infected with aerosolized H37Rv, then treated with DS6A intravenously, the DS6A treated mice showed increased body weight and improved pulmonary function relative to control mice. Furthermore, DS6A reduces Mtb load in mouse organs with greater efficacy in the spleen. These results demonstrate the feasibility of developing phage therapy as an effective therapeutic against Mtb infection.

Investigating Phenotypic and Genotypic Resistance to Pyrazinamide Antibiotic in Multidrug-resistant (MDR) Mycobacterium Tuberculosis Isolates in Khuzestan Province from 2017 to 2022

Background and Objectives Pyrazinamide is one of the effective drugs for treating tuberculosis. Inclusion of this drug in the treatment regimen shortens the duration of tuberculosis treatment. The mutation in pncA gene causes loss of pyrazinamidase activity, the most important resistance mechanism in Mycobacterium tuberculosis isolates. Therefore, the purpose of this study is to investigate the phenotypic and genotypic resistance to pyrazinamide antibiotic in multidrug-resistant (MDR) Mycobacterium tuberculosis isolates in Khuzestan province between 2017 and 2022. Subjects and Methods This study was conducted on 40 isolates of Mycobacterium tuberculosis in Ahvaz TB Regional Laboratory. Drug sensitivity of the isolates was determined by the relative method in the antibiotic pyrazinamide in the isolates. Then, the frequency of pncA gene mutations related to pyrazinamide resistance in MDR isolates was determined using sequencing. Results According to the results of the drug sensitivity to rifampin and isoniazid antibiotics, there were 25 isolates, of which 16 strains were resistant to two drugs, namely isoniazid and rifampin (MDR-TB), and 9 were single-drug resistant isolates (8 isolates resistant to rifampin and 1 isolate resistant to Isoniazid). Of the 17 samples that had the pncA gene and were resistant to pyrazinamide, 13 had mutations in the pncA gene while 4 did not have any mutations. The most common mutation was a non-synonymous mutation in which the amino acid Val had changed to Phe. Conclusion According to the results of this study, there is a high frequency of resistance to pyrazinamide in MDR strains. Also, there is a high percentage of single-resistance mutations in the pncA gene, while there is lower prevalence of mutations in the panD and rpsA genes, which provide quick and accurate information about the sensitivity to pyrazinamide for MDR-TB and mono-resistant isolates. Therefore, the best method for detecting resistance to pyrazinamide is sequencing and whole pncA DNA sequence to confirm pyrazinamide resistance instead of the usual methods covering mutated hotspots.

Antibiotic Skeletal Diversification via Differential Enoylreductase Recruitment and Module Iteration in trans-Acyltransferase Polyketide Synthases.

Microorganisms are remarkable chemists capable of assembling complex molecular architectures that penetrate cells and bind biomolecular targets with exquisite selectivity. Consequently, microbial natural products have wide-ranging applications in medicine and agriculture. How the "blind watchmaker" of evolution creates skeletal diversity is a key question in natural products research. Comparative analysis of biosynthetic pathways to structurally related metabolites is an insightful approach to addressing this. Here, we report comparative biosynthetic investigations of gladiolin, a polyketide antibiotic from Burkholderia gladioli with promising activity against multidrug-resistant Mycobacterium tuberculosis, and etnangien, a structurally related antibiotic produced by Sorangium cellulosum. Although these metabolites have very similar macrolide cores, their C21 side chains differ significantly in both length and degree of saturation. Surprisingly, the trans-acyltransferase polyketide synthases (PKSs) that assemble these antibiotics are almost identical, raising intriguing questions about mechanisms underlying structural diversification in this important class of biosynthetic assembly line. In vitro reconstitution of key biosynthetic transformations using simplified substrate analogues, combined with gene deletion and complementation experiments, enabled us to elucidate the origin of all the structural differences in the C21 side chains of gladiolin and etnangien. The more saturated gladiolin side chain arises from a cis-acting enoylreductase (ER) domain in module 1 and in trans recruitment of a standalone ER to module 5 of the PKS. Remarkably, module 5 of the gladiolin PKS is intrinsically iterative in the absence of the standalone ER, accounting for the longer side chain in etnangien. These findings have important implications for biosynthetic engineering approaches to the creation of novel polyketide skeletons.

Whole Genome Sequence Dataset of Mycobacterium tuberculosis Strains from Patients of Campania Region

Tuberculosis (TB) is one of the deadliest infectious disorders in the world. To effectively TB manage, an essential step is to gain insight into the lineage of Mycobacterium tuberculosis (MTB) and the distribution of drug resistance. Although the Campania region is declared a cluster area for the infection, to contribute to the effort to understand TB evolution and transmission, still poorly known, we have generated a dataset of 159 genomes of MTB strains, from Campania region collected during 2018–2021, obtained from the analysis of whole genome sequence. The results show that the most frequent MTB lineage is the 4 according for 129 strains (81.11%). Regarding drug resistance, 139 strains (87.4%) were classified as multi susceptible, while the remaining 20 (12.58%) showed drug resistance. Among the drug-resistance strains, 8 were isoniazid-resistant MTB, 4 multidrug-resistant MTB, while only one was classified as pre-extensively drug-resistant MTB. This dataset expands the existing available knowledge on drug resistance and evolution of MTB, contributing to further TB-related genomics studies to improve the management of this disease.

Adverse drug events associated with linezolid administration: a real-world pharmacovigilance study from 2004 to 2023 using the FAERS database.

Introduction: Linezolid is an oxazolidinone antibiotic that is active against drug-resistant Gram-positive bacteria and multidrug-resistant Mycobacterium tuberculosis. Real-world studies on the safety of linezolid in large populations are lacking. This study aimed to determine the adverse events associated with linezolid in real-world settings by analyzing data from the US Food and Drug Administration (FDA) Adverse Event Reporting System (FAERS). Methods: We retrospectively extracted reports on adverse drug events (ADEs) from the FAERS database from the first quarter of 2004 to that of 2023. By using disproportionality analysis including reporting odds ratio (ROR), proportional reporting ratio (PRR), Bayesian Confidence Propagation Neural Network (BCPNN), along with the multi-item gamma Poisson shrinker (MGPS), we evaluated whether there was a significant association between linezolid and ADE. The time to onset of ADE was further analyzed in the general population and within each age, weight, reporting population, and weight subgroups. Results: A total of 11,176 reports of linezolid as the "primary suspected" drug and 263 significant adverse events of linezolid were identified, including some common adverse events such as thrombocytopenia (n = 1,139, ROR 21.98), anaemia (n = 704, ROR 7.39), and unexpected signals that were not listed on the drug label such as rhabdomyolysis (n = 90, ROR 4.33), and electrocardiogram QT prolonged (n = 73, ROR 4.07). Linezolid-induced adverse reactions involved 27 System Organ Class (SOC). Gender differences existed in ADE signals related to linezolid. The median onset time of all ADEs was 6 days, and most ADEs (n = 3,778) occurred within the first month of linezolid use but some may continue to occur even after a year of treatment (n = 46). Conclusion: This study reports the time to onset of adverse effects in detail at the levels of SOC and specific preferred term (PT). The results of our study provide valuable insights for optimizing the use of linezolid and reducing potential side effects, expected to facilitate the safe use of linezolid in clinical settings.

Transmission of drug-resistant Mycobacterium tuberculosis isolates between Finnish- and foreign-born cases, 2014–2021: A molecular epidemiological study

BackgroundData on the molecular epidemiology and transmission of drug-resistant Mycobacterium tuberculosis (MTB) in low-incidence settings with immigration from high-incidence settings is limited. MethodWe included 115 drug-resistant (DR) MTB isolates with whole-genome sequencing data isolated in Finland between 2014 and 2021. Potential transmission clusters were identified using a threshold of 12 single-nucleotide polymorphisms (SNPs). Highly related clusters were identified using a threshold of 5 SNPs. ResultOf the 115 DR MTB isolates, 31 (27.0%) isolates were from Finnish-born cases and 84 (73.0%) were from foreign-born cases. The proportion of multidrug-resistant (MDR) MTB isolates (30/84, 35.7%) from foreign-born cases was higher than that of MDR MTB isolates from Finnish-born cases (8/31, 25.8%). Lineage 2 (40/115, 34.8%) and lineage 4 (40/115, 34.8%) were the most prevalent lineages. A total of 25 (21.7%) isolates were classified into eight potential transmission clusters (≤12 SNPs). Furthermore, five highly related clusters (≤5 SNPs) were identified, including three DR MTB isolates from Finnish-born cases and 14 DR isolates from foreign-born cases. ConclusionThe risk of DR MTB transmission between Finnish- and foreign-born persons is not negligible. Further research on clustering analysis in drug-susceptible MTB is worth to inform tuberculosis management and control in low-incidence settings with increasing immigration.

Prevalence, Transmission and Genetic Diversity of Pyrazinamide Resistance Among Multidrug-Resistant Mycobacterium tuberculosis Isolates in Hunan, China.

China is a country with a burden of high rates of both TB and multidrug-resistant TB (MDR-TB). However, published data on pyrazinamide (PZA) resistance are still limited in Hunan province, China. This study investigated the prevalence, transmission, and genetic diversity of PZA resistance among multidrug-resistant Mycobacterium tuberculosis isolates in Hunan province. Drug susceptibility testing (DST) with the Bactec MGIT 960 PZA kit and pyrazinamidase (PZase) testing were conducted on all 298 MDR clinical isolates. Moreover, 24-locus MIRU-VNTR and DNA sequencing of pncA, rpsA, and panD genes were conducted on 180 PZA-resistant (PZA-R) isolates. The prevalence of PZA resistance among MDR-TB strains reached 60.4%. Newly diagnosed PZA-R TB patients and clustered isolates with identical pncA, rpsA, and panD mutations showed that transmission of PZA-R isolates played a significant role in the formation of PZA-R TB. Ninety-eight mutation patterns were observed in the pncA among 180 PZA-R isolates, and seventy-one (72.4%) were point mutations. Twenty-four of these mutations are new, including 2 base substitutions (V93G and T153S) and 22 nucleotide deletions or insertions. The W119C was found in PZA-S isolates, on the other hand, F94L and V155A mutations were found in both PZA resistant and susceptible isolates with positive PZase activity, indicating that they were not associated with PZA resistance. This is not entirely in line with the WHO catalogue. Ten novel rpsA mutations were found in 10 PZA-R isolates, which all combined with mutations in pncA. Thus, it is unpredictable whether these mutations in rpsA can impact PZA resistance. No panD mutation was found in all PZA-R isolates. DNA sequencing of pncA and PZase activity testing have great potential in predicting PZA resistance.

Transient comparison of techniques to counter multi-drug resistant bacteria: prime modules in curation of bacterial infections

Multidrug-resistant organisms are bacteria that are no longer controlled or killed by specific drugs. One of two methods causes bacteria multidrug resistance (MDR); first, these bacteria may disguise multiple cell genes coding for drug resistance to a single treatment on resistance (R) plasmids. Second, increased expression of genes coding for multidrug efflux pumps, which extrude many drugs, can cause MDR. Antibiotic resistance is a big issue since some bacteria may withstand almost all antibiotics. These bacteria can cause serious sickness, making them a public health threat. Methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus (VRE), Multidrug resistant Mycobacterium tuberculosis (TB), and CRE are gut bacteria that resist antibiotics. Antimicrobial resistance is rising worldwide, increasing clinical and community morbidity and mortality. Superbugs have made antibiotic resistance in some environmental niches even harder to control. This study introduces new medicinal plants, gene-editing methods, nanomaterials, and bacterial vaccines that will fight MDR bacteria in the future.

Drug-Resistant Characteristics, Genetic Diversity, and Transmission Dynamics of Multidrug-Resistant Mycobacterium tuberculosis in Jiangxi, China.

In this study, we aimed to determine the transmission pattern of multidrug-resistant tuberculosis (MDR-TB) isolates circulating in Jiangxi Province with whole-genome sequencing (WGS). In addition, we also sought to describe mutational resistome of MDR-TB isolates. A total of 115 MDR-TB isolates determined by the phenotypic proportion method of drug susceptibility testing between January 2018 and December 2022 from provincial drug surveillance (DRS) in Jiangxi were included in our analysis. The demographic data and treatment history were extracted from the National TB Registry System. WGS was used to analyze the genotypic characteristics of drug resistance and transmissions. About 62.6% of MDR-TB strains were isolated from cases that received previous anti-tuberculosis treatment. According to the WGS results, 96.5% were genotypic MDR-TB, and more than half of MDR-TB isolates tested were also resistant to streptomycin (59.1%), ethambutol (56.5%), and fluroquinolones (53.0%), while resistance to cycloserine and linezolid was lowest, only in two (1.7%) and one (0.9%) isolate, respectively. Ser450Leu in rpoB (57.9%), Ser315Thr in katG (74.1%), Met306Val in embB (40.0%), Lys43Arg in rpsL (75.0%), Ala90Val in gyrA (32.8%) were predominant mutant types among the rifampin-, isoniazid-, ethambutol-, streptomycin-, fluoroquinolones-resistant isolates, respectively. Lineage 2 (East Asian genotype) occurred at the highest frequency with 97 cases (84.3%), followed by lineage 4 (Euro-American genotype) with 18 cases (15.7%). Additionally, 5 clusters consisting of 10 isolates were identified in the present study, demonstrating a clustering rate of 8.7%. MDR/Rifampicin-Resistant (RR)-TB epidemic in this region is driven by lineage 2 clade that also show higher resistance to other anti-tuberculosis drugs. Lower cluster rates compared with a relatively higher proportion of new MDR-TB cases indicate that a considerable number of MDR-TB cases remain undiagnosed.

Identification of potential mutations associated with multidrug resistance among isolates of Mycobacterium tuberculosis in Malaysia by in silico screening

The emergence of multidrug resistance tuberculosis (MDR-TB) is caused by Mycobacterium tuberculosis (MTB) adaptation to survive in the presence of antibiotic, that were contributed by mutations in the MDR-associated genes. Previous research has indicated that the gene expression knockdown of fhaA leads to an accumulation of peptidoglycan (PG) precursors at the bacillary septum and poles, which suggest a possible deficiency in PG biosynthesis. Consequently, the cell wall becomes resistant to antibiotics, leading to multidrug resistance (MDR). In this study, bioinformatics analyses were performed on MDR-TB isolates from 24 clinical samples to search for novel mutations that contribute to antibiotic resistance. We found out a potential deletion of nucleotides encoding 6 amino acids in all 12 samples, particularly in fhaA gene (RV0020c). Our subsequent structural analysis shows that the deletion is at the position 243-248, causing conformational change of the native FhaA protein. We postulated that the deletion will potentially cause the loss of its binding affinity to MviN (precursor) and STPK (protein kinase), resulting in the inhibition and blockage of the peptidoglycan polymerization, causing MDR in MTB. In the future, experimental validation is necessitated to substantiate the association of these mutations with MDR.

Omadacycline pharmacokinetics/pharmacodynamics and efficacy against multidrug-resistant Mycobacterium tuberculosis in the hollow fiber system model.

Seventy-five years ago, first-generation tetracyclines demonstrated limited efficacy in the treatment of tuberculosis but were more toxic than efficacious. We performed a series of pharmacokinetic/pharmacodynamic (PK/PD) experiments with a potentially safer third-generation tetracycline, omadacycline, for the treatment of multidrug-resistant tuberculosis (MDR-TB). Mycobacterium tuberculosis (Mtb) H37Rv and an MDR-TB clinical strain (16D) were used in the minimum inhibitory concentration (MIC) and static concentration-response studies in test tubes, followed by a PK/PD study using the hollow fiber system model of TB (HFS-TB) that examined six human-like omadacycline doses. The inhibitory sigmoid maximal effect (Emax) model and Monte Carlo experiments (MCEs) were used for data analysis and clinical dose-finding, respectively. The omadacycline MIC for both Mtb H37Rv and MDR-TB clinical strain was 16 mg/L but dropped to 4 mg/L with daily drug supplementation to account for omadacycline degradation. The Mycobacteria Growth Indicator Tube MIC was 2 mg/L. In the test tubes, omadacycline killed 4.39 log10 CFU/mL in 7 days. On Day 28 of the HFS-TB study, the Emax was 4.64 log10 CFU/mL, while exposure mediating 50% of Emax (EC50) was an area under the concentration-time curve to MIC (AUC0-24/MIC) ratio of 22.86. This translates to PK/PD optimal exposure or EC80 as AUC0-24/MIC of 26.93. The target attainment probability of the 300-mg daily oral dose was 90% but fell at MIC ≧4 mg/L. Omadacycline demonstrated efficacy and potency against both drug-susceptible and MDR-TB. Further studies are needed to identify the omadacycline effect in combination therapy for the treatment of both drug-susceptible and MDR-TB.

The heme oxygenase-1 metalloporphyrin inhibitor stannsoporfin enhances the bactericidal activity of a novel regimen for multidrug-resistant tuberculosis in a murine model.

Multidrug-resistant (MDR) Mycobacterium tuberculosis (Mtb) poses significant challenges to global tuberculosis (TB) control efforts. Host-directed therapies (HDTs) offer a novel approach to TB treatment by enhancing immune-mediated clearance of Mtb. Prior preclinical studies found that the inhibition of heme oxygenase-1 (HO-1), an enzyme involved in heme metabolism, with tin-protoporphyrin IX (SnPP) significantly reduced mouse lung bacillary burden when co-administered with the first-line antitubercular regimen. Here, we evaluated the adjunctive HDT activity of a novel HO-1 inhibitor, stannsoporfin (SnMP), in combination with a novel MDR-TB regimen comprising a next-generation diarylquinoline, TBAJ-876 (S), pretomanid (Pa), and a new oxazolidinone, TBI-223 (O) (collectively, SPaO), in Mtb-infected BALB/c mice. After 4 weeks of treatment, SPaO + SnMP 5mg/kg reduced mean lung bacillary burden by an additional 0.69 log10 (P = 0.01) relative to SPaO alone. As early as 2 weeks post-treatment initiation, SnMP adjunctive therapy differentially altered the expression of pro-inflammatory cytokine genes and CD38, a marker of M1 macrophages. Next, we evaluated the sterilizing potential of SnMP adjunctive therapy in a mouse model of microbiological relapse. After 6 weeks of treatment, SPaO + SnMP 10mg/kg reduced lung bacterial burdens to 0.71 ± 0.23 log10 colony-forming units (CFUs), a 0.78 log-fold greater decrease in lung CFU compared to SpaO alone (P = 0.005). However, adjunctive SnMP did not reduce microbiological relapse rates after 5 or 6 weeks of treatment. SnMP was well tolerated and did not significantly alter gross or histological lung pathology. SnMP is a promising HDT candidate requiring further study in combination with regimens for drug-resistant TB.

Pulmonary tuberculosis and multidrug-resistant Mycobacterium tuberculosis in northwestern Ethiopia: a hospital-based cross-sectional study among presumptive pulmonary tuberculosis patients.

Border areas are important sites for disseminating Mycobacterium tuberculosis among individuals living in such areas. This study examined patients with suspected pulmonary tuberculosis (PTB) visiting the Abrihajira and Metema hospitals in northwest Ethiopia to investigate the prevalence of rifampicin-resistant Mycobacterium tuberculosis (RR-MTB), multidrug-resistant Mycobacterium tuberculosis (MDR-MTB), and risk factors related to Mycobacterium tuberculosis infection. A hospital-based cross-sectional study was conducted from February to August 2021 among 314 PTB presumptive patients. Xpert MTB/RIF and line probe assays (LPA) were used to process sputum samples. Data were imported into the Epi-Data 3.1 program and exported to Statistical Package for the Social Sciences (SPSS) version 20.0 (SPSS, Chicago, IL, United States) to conduct the analysis. A logistic regression analysis was used to investigate the relationship between the dependent and independent variables. A value of p of <0.05 denoted statistical significance. Of the total (314) PTB presumptive patients who participated in this study, 178 (56.69%) were men, and 165 (52.5%) were from 25 to 50 years of age with a median age of 35.00 (inter-quartile: 25-45 years). Among all patients, 12.7% had PTB by Gene Xpert and 7/314 (2.23%) were resistant to rifampicin. Among patients enrolled, 4/314 (1.27%) had MDR-MTB (resistant to RIF and INH) by LPA. Regarding the risk factors assessed, primary level of education, sputum production, night sweating, respiratory disorder, contact history of TB, history of MDR-MTB infection, history of alcohol use, and cigarette smoking showed statistical significance with the prevalence of PTB (p ≤ 0.05). This study observed a high prevalence of PTB, RR-MTB, and MDR-MTB compared with many other previous studies conducted in Ethiopia. Among the assessed risk factors that could be associated with the prevalence of PTB, eight were statistically significant. This prevalence, resistance, and statistically significant variables are the evidence to which more emphasis should be given to the country's border areas.

Anti-Tuberculosis Potential of OJT008 against Active and Multi-Drug-Resistant Mycobacterium Tuberculosis: In Silico and In Vitro Inhibition of Methionine Aminopeptidase.

Despite the recent progress in the diagnosis of tuberculosis (TB), the chemotherapeutic management of TB continues to be challenging. Mycobacterium tuberculosis (Mtb), the etiological agent of TB, is classified as the 13th leading cause of death globally. In addition, 450,000 people were reported to develop multi-drug-resistant TB globally. The current project focuses on targeting methionine aminopeptidase (MetAP), an essential protein for the viability of Mtb. MetAP is a metalloprotease that catalyzes the excision of the N-terminal methionine (NME) during protein synthesis, allowing the enzyme to be an auspicious target for the development of novel therapeutic agents for the treatment of TB. Mtb possesses two MetAP1 isoforms, MtMetAP1a and MtMetAP1c, which are vital for Mtb viability and, hence, a promising chemotherapeutic target for Mtb therapy. In this study, we cloned and overexpressed recombinant MtMetAP1c. We investigated the in vitro inhibitory effect of the novel MetAP inhibitor, OJT008, on the cobalt ion- and nickel ion-activated MtMetAP1c, and the mechanism of action was elucidated through an in silico approach. The compound's potency against replicating and multi-drug-resistant (MDR) Mtb strains was also investigated. The induction of the overexpressed recombinant MtMetAP1c was optimized at 8 h with a final concentration of 1 mM Isopropyl β-D-1-thiogalactopyranoside. The average yield from 1 L of Escherichia coli culture for MtMetAP1c was 4.65 mg. A preliminary MtMetAP1c metal dependency screen showed optimum activation with nickel and cobalt ions occurred at 100 µM. The half-maximal inhibitory concentration (IC50) values of OJT008 against MtMetAP1c activated with CoCl2 and NiCl2 were 11 µM and 40 µM, respectively. The in silico study showed OJT008 strongly binds to both metal-activated MtMetAP1c, as evidenced by strong molecular interactions and a higher binding score, thereby corroborating our result. This in silico study validated the pharmacophore's metal specificity. The potency of OJT008 against both active and MDR Mtb was <0.063 µg/mL. Our study reports OJT008 as an inhibitor of MtMetAP1c, which is potent at low micromolar concentrations against both active susceptible and MDR Mtb. These results suggest OJT008 is a potential lead compound for the development of novel small molecules for the therapeutic management of TB.

Association of Mutations in gyrA Gene with Resistance to Fluoroquinolones in Clinical Isolates of Multidrug-Resistant Mycobacterium tuberculosis

Association of Mutations in gyrA Gene with Resistance to Fluoroquinolones in Clinical Isolates of Multidrug-Resistant Mycobacterium tuberculosis

Computer-aided identification of Mycobacterium tuberculosis resuscitation-promoting factor B (RpfB) inhibitors from Gymnema sylvestre natural products

Tuberculosis (TB), an infectious disease caused by multi-drug resistant Mycobacterium tuberculosis (Mtb), has been a global health concern. Mtb affects over a third of the world’s population, causing two million deaths annually due to its dormancy and propensity to spread infection during this period. Resuscitation-promoting factor B (RpfB) plays a pivotal role in the growth of Mtb during dormant periods, making it a critical target for eliminating Mtb and curing TB. Gymnema sylvestre is a famous medicinal plant with several medicinal properties, including antimicrobial activity; however, the therapeutic potential of the various reported metabolites of this plant against Mtb has not yet been explored. The aim of this study was to explore the reported natural products of G. sylvestre against the RpfB of the Mtb. A total of 131 reported secondary metabolites of this plant were collected and virtually screened against the RpfB. We particularly targeted the Glu292 residue of RpfB as it is crucial for the catalysis of this protein. From our in-house library, 114 compounds showed a binding affinity higher than the standard drug. The binding stability of the top three lead compounds was further confirmed through MD simulation analysis. Drug likeness analyses indicated that the ten hits had zero violations of the Lipinski rule of five. In addition, analyses of pharmacokinetics, toxicity, and target prediction revealed that the top compounds are devoid of toxicity and do not affect human proteins. Additionally, they reflect multifaceted approach as anti-TB agents. Our selected hits not only exhibit molecular properties favoring physiological compatibility but also exhibit properties enhancing their potential efficacy as therapeutic candidates. The compounds investigated here are worthy of experimental validation for the discovery of novel treatments against TB. Further, this study also provides a promising avenue for research on the pharmacological potential of G. sylvestre.

1954. Dynamic PET facilitated Modeling and Novel Antibiotic Regimens for Tuberculous Meningitis due to Multidrug-Resistant Strains

1954. Dynamic PET facilitated Modeling and Novel Antibiotic Regimens for Tuberculous Meningitis due to Multidrug-Resistant Strains

Toxin-antitoxin Genes Expression in Multidrug-Resistant Mycobacterium tuberculosis Isolates under Drug Exposure.

Toxin-antitoxin systems (TAs) are highly conserved in Mycobacterium tuberculosis (Mtb). The TAs role in maintaining and disseminating drug resistance in bacterial populations has been indicated. So, we aimed to analyze the expression level of MazEF-related genes in drug-susceptible and multi drug-resistant (MDR) Mtb isolates under isoniazid (INH) and rifampin (RIF) stress. We obtained 23 Mtb isolates, including 18 MDR and 5 susceptible isolates, from the Ahvaz Regional TB Laboratory collection. The expression level of mazF3, mazF6, mazF9 toxin genes and mazE3, mazE6, mazE9 antitoxin genes in MDR and susceptible isolates was evaluated by quantitative real-time PCR (qRT-PCR) after exposure to RIF and INH. The mazF3, F6, and F9 toxin genes were overexpressed in at least two MDR isolates in the presence of RIF and INH, in contrast to mazE antitoxin genes. More MDR isolates were induced to overexpress mazF genes by RIF than INH (72.2% vs. 50%). Compared to the H37Rv strain, and susceptible isolates, the expression levels of mazF3,6 by RIF and mazF3,6,9 by INH were significantly upregulated in MDR isolates (p<0.05) but no remarkable difference was detected in the expression level of mazF9 genes by INH between these groups. In susceptible isolates, the expression levels of mazE3,6 by RIF and mazE3,6,9 by INH were induced and enhanced significantly compared with MDR isolates, but there was no difference between MDR and H37Rv strain. Based on the results, we propose that mazF expression under RIF/INH stress may be associated with drug resistance in Mtb in addition to mutations, and the mazE antitoxins may be related to enhanced susceptibility of Mtb to INH and RIF. Further experiments are needed to investigate the exact mechanism underlying the TA system role in drug resistance.

Characterization of Mutations in Genes Related to Rifampicin and Isoniazid Resistance in Multidrug-resistant Mycobacterium tuberculosis Strains from Hangzhou, China

Characterization of Mutations in Genes Related to Rifampicin and Isoniazid Resistance in Multidrug-resistant Mycobacterium tuberculosis Strains from Hangzhou, China