Multidrug-resistant Mycobacterium Tuberculosis Research Articles

High-throughput screening to discover inhibitors of the CarD\xb7RNA polymerase protein\u2013protein interaction in Mycobacterium tuberculosis

Multidrug-resistant Mycobacterium tuberculosis (MDR-TB) accounts for 3.7% of new cases of TB annually worldwide and is a major threat to global public health. Due to the prevalence of the MDR-TB and extensively drug resistant tuberculosis (XDR-TB) cases, there is an urgent need for new drugs with novel mechanisms of action. CarD, a global transcription regulator in MTB, binds RNAP and activates transcription by stabilizing the transcription initiation open-promoter complex (RPo). CarD is required for MTB viability and it has highly conserved homologues in many eubacteria. A fluorescence polarization (FP) assay which monitors the association of MTB RNAP, native rRNA promoter DNA and CarD has been developed. Overall, our objective is to identify and characterize small molecule inhibitors which block the CarD/RNAP interaction and to understand the mechanisms by which CarD interacts with the molecules. We expect that the development of a new and improved anti-TB compound with a novel mechanism of action will relieve the burden of resistance. This CarD FP assay is amenable to HTS and is an enabling tool for future novel therapeutic discovery.

Genetic diversity of multidrug-resistant Mycobacterium tuberculosis Beijing family isolates in Nepal

Genetic diversity of multidrug-resistant Mycobacterium tuberculosis Beijing family isolates in Nepal

Antimicrobial Peptide against Mycobacterium Tuberculosis That Activates Autophagy Is an Effective Treatment for Tuberculosis.

Mycobacterium tuberculosis (MTB) is the principal cause of human tuberculosis (TB), which is a serious health problem worldwide. The development of innovative therapeutic modalities to treat TB is mainly due to the emergence of multi drug resistant (MDR) TB. Autophagy is a cell-host defense process. Previous studies have reported that autophagy-activating agents eliminate intracellular MDR MTB. Thus, combining a direct antibiotic activity against circulating bacteria with autophagy activation to eliminate bacteria residing inside cells could treat MDR TB. We show that the synthetic peptide, IP-1 (KFLNRFWHWLQLKPGQPMY), induced autophagy in HEK293T cells and macrophages at a low dose (10 μM), while increasing the dose (50 μM) induced cell death; IP-1 induced the secretion of TNFα in macrophages and killed Mtb at a dose where macrophages are not killed by IP-1. Moreover, IP-1 showed significant therapeutic activity in a mice model of progressive pulmonary TB. In terms of the mechanism of action, IP-1 sequesters ATP in vitro and inside living cells. Thus, IP-1 is the first antimicrobial peptide that eliminates MDR MTB infection by combining four activities: reducing ATP levels, bactericidal activity, autophagy activation, and TNFα secretion.

Mucosal Therapy of Multi-Drug Resistant Tuberculosis With IgA and Interferon-γ.

New evidence has been emerging that antibodies can be protective in various experimental models of tuberculosis. Here, we report on protection against multidrug-resistant Mycobacterium tuberculosis (MDR-TB) infection using a combination of the human monoclonal IgA 2E9 antibody against the alpha-crystallin (Acr, HspX) antigen and mouse interferon-gamma in mice transgenic for the human IgA receptor, CD89. The effect of the combined mucosal IgA and IFN-γ; treatment was strongest (50-fold reduction) when therapy was applied at the time of infection, but a statistically significant reduction of lung bacterial load was observed even when the therapy was initiated once the infection had already been established. The protection involving enhanced phagocytosis and then neutrophil mediated killing of infected cells was IgA isotype mediated, because treatment with an IgG version of 2E9 antibody was not effective in human IgG receptor CD64 transgenic mice. The Acr antigen specificity of IgA antibodies for protection in humans has been indicated by their elevated serum levels in latent tuberculosis unlike the lack of IgA antibodies against the virulence-associated MPT64 antigen. Our results represent the first evidence for potential translation of mucosal immunotherapy for the management of MDR-TB.

Population structure of multidrug-resistant Mycobacterium tuberculosis clinical isolates in Colombia

Emergence of multidrug-resistant (MDR) Mycobacterium tuberculosis complex (MTBC) isolates is a major public health problem that threatens progress made in tuberculosis (TB) care and control worldwide. In Colombia, the prevalence of MDR tuberculosis (MDR-TB) has increased slowly but steadily since 2001. However, the population structure of the MDR-TB strains circulating in Colombia is sparsely known. In this work, 203 MDR isolates isolated in 2012–2013 were collected, and characterized by spoligotyping, followed by 24-loci MIRU-VNTR (data available for 190 isolates). The most prevalent genotypes corresponded to SIT42/LAM9 (12.81%), SIT62/H1 (10.34%), and SIT190/Beijing (10.34%). A fine analysis showed that although the MDR strains came from 29 of the 33 departments of Colombia, the distribution of these main lineages was not at random and depended on the city of isolation (p-value <0.000001). Both LAM and Beijing lineage strains were significantly associated with MDR-TB (p-value <0.0001): LAM lineage was associated with 2 patterns of MDR, namely combined resistance to INH + Rifampin (HR), and to SHRE (Streptomycin + INH + Rifampin + Ethambutol), while the Beijing lineage strains were essentially associated with MDR (SHRE). Interestingly, distribution of genotypic lineages in function of drug resistance information (e.g. pansusceptible vs. MDR) was different in our setting as compared to other countries in Latin America. However, MIRU-VNTR patterns were unique for all strains, an observation that did not support active transmission of circulating MDR clones.

In silico screening of active constituent of Couroupita guianensis against Mycobacterium

Background: A vital need to ascertain novel anti-tubercular agent which is the volatile global spreading of multidrug resistant Mycobacterium tuberculosis. Enoyl-acyl carrier protein reductase is one among such target. It is one of the key enzymes involved in the type II fatty acid biosynthesis pathway of M. tuberculosis. Objective: In this study, in silico evaluations were employed in screening of active constituent of Couroupita guianensis against Enoyl-acyl carrier protein reductase of Mycobacterium tuberculi. Materials and Methods: Totally 16 compounds namely Isatin, Indigo, Coup 2, Indirubin, Calotronaphthalene, Coup, Alpha Amyrin, Nerol, Betasitosterol, Campesterol, Eugenol, Tryptanthrin, Benzyl Alcohol, Betaamyrin and Farnesol were subjected to in silico screening. Glide software of Schrodinger was used to carry out the current work. Results: The compounds exhibit good docking score and few with hydrogen bond interaction. Isoniazid was used as the standard and validation was performed. The results have shown that derivatives were proved to be highly potent inhibitors against Mycobacterium tuberculosis enoyl acyl carrier protein reductase. Conclusion: Most of the compounds exhibit hydrophobic interaction. Then isatin and eugenol can be tested against Mycobacterium tuberculi. Keywords: Couroupita guianensis, Enoyl acyl carrier protein reductase, In silico screening, Mycobacterium tuberculi.

Prevalence of Drug-Resistant Mycobacterium tuberculosis in the Veterans Health Administration (VHA)

Background: In 2018, the CDC reported that isoniazid (INH)-resistant and multidrug-resistant Mycobacterium tuberculosis (MDR-TB, ie, resistant to at least INH and rifampin) represented 9.3% and 1.5% of TB cases, respectively, in the United States. Objective: We analyzed the prevalence of drug-resistant TB within the Veterans Health Administration (VHA) to determine factors associated with hospitalization. Method: Patients were identified using Council of State and Territorial Epidemiologists case definition for laboratory-confirmed TB by querying VHA data sources from January 1, 2010, to June 30, 2019. Susceptibility results were included for isolates from all body sites. Using a 2-proportion z test, we compared the following demographic factors for susceptible versus drug-resistant TB: age, gender, race/ethnicity, location of residence and birth. We also assessed the following clinical and hospitalization factors: pulmonary versus extrapulmonary disease, latent TB infection (LTBI) screening and treatment, length of stay (LOS), intensive care unit (ICU) stay, and death. Results: In total, 878 patients had lab-confirmed TB, and 812 (92%) had electronic drug-susceptibility results available. Of 812 patients, 49 (6%) showed anti-TB drug resistance (Fig. 1), which was less than that reported nationally by the CDC (Fig. 2). No patients had extensively drug-resistant TB. Only 18 of 49 patients (37%) with resistant TB had LTBI screening ≥3 months prior to diagnosis. Among 6 patients with LTBI, 3 (50%) received treatment. Patient state of residence was the only demographic factor associated with resistant TB. Arizona, Iowa, Massachusetts, Montana, Nevada, South Dakota, and Utah were significantly associated with drug-resistant TB cases (P &lt; .05); however, overall numbers of VHA TB cases in these states were low. Patients with resistant TB were more likely to have extrapulmonary TB (13 of 49, 26.5%) than patients with susceptible TB (111 of 763, 14.5%; P &lt; .05) (Table 1). Mortality at 30 and 90 days for resistant versus susceptible TB cases did not differ significantly: 6.1% versus 5.1% at 30 days (P = .75) and 10.2% versus 10.6% at 90 days (P = 0.92), respectively. The proportion of drug-resistant TB cases hospitalized did not differ from susceptible cases (59% vs 63%), respectively (P = .62) nor did the proportion with ICU stay (14.3% vs 12.3%), respectively (P = .40). Median LOS for drug-resistant TB cases and susceptible cases were similar: 5 days (range, 0–303 versus 4 days (range, 0–111), respectively. Conclusions: Rates of drug-resistant TB are lower in the VHA than in the general US population. However, improvement is needed in LTBI screening and treatment rates. Little has been published on drug resistance in extrapulmonary TB; however, our findings should alert clinicians to the possibility of resistance in these challenging infections.Funding: NoneDisclosures: None

Insertion and deletion evolution reflects antibiotics selection pressure in a Mycobacterium tuberculosis outbreak.

In genome evolution, genetic variants are the source of diversity, which natural selection acts upon. Treatment of human tuberculosis (TB) induces a strong selection pressure for the emergence of antibiotic resistance-conferring variants in the infecting Mycobacterium tuberculosis (MTB) strains. MTB evolution in response to treatment has been intensively studied and mainly attributed to point substitutions. However, the frequency and contribution of insertions and deletions (indels) to MTB genome evolution remains poorly understood. Here, we analyzed a multi-drug resistant MTB outbreak for the presence of high-quality indels and substitutions. We find that indels are significantly enriched in genes conferring antibiotic resistance. Furthermore, we show that indels are inherited during the outbreak and follow a molecular clock with an evolutionary rate of 5.37e-9 indels/site/year, which is 23 times lower than the substitution rate. Inherited indels may co-occur with substitutions in genes along related biological pathways; examples are iron storage and resistance to second-line antibiotics. This suggests that epistatic interactions between indels and substitutions affect antibiotic resistance and compensatory evolution in MTB.

Emergence of Specific gyrA Mutations Associated High-Level Fluoroquinolone-Resistant Mycobacterium tuberculosis among Multidrug-Resistant Tuberculosis Cases in North India

Aim: This study aims to determine the frequency and pattern of gyrA/B mutations in multidrug-resistant (MDR) Mycobacterium tuberculosis (MTB) strains and also to assess the association between different gyrA/B mutations with phenotypic resistance to moxifloxacin (MOX) at clinical breakpoint (CB) drug concentration. Method: A total of 106 clinical MTB isolates carrying gyrA/B mutations were included consecutively. Culture-based MOX susceptibility was tested at CB (1.0 μg/mL) followed by its correlation with gyrA/B mutations using Genotype MTBDRsl assay. The mutations associated with phenotypic resistance were further analyzed on a large dataset of 1,825 MDR tuberculosis (TB) patients. Result: D94G and A90V mutations within gyrA were significantly associated with resistance and susceptible phenotype (p < 0.001), respectively. Of 1,825 MDR patients, gyrA/B mutations were found in 58.8% cases, of which fluoroquinolone (FQ) resistance was concluded among 97.9%, 0.8%, and 1.3% patients due to mutation in gyrA, gyrB, and in both the genes, respectively. D94G alone (45.9%) followed by A90V (21.2%) mutations in gyrA gene was most frequent. Conclusion: Our study showed that MDR-TB has emerged in northern India with additional FQ resistance. Different selection pressure and transmission may result in prevailing accumulation of specific gyrA mutations causing high-level FQ resistance, therefore, current control measures need to be strengthened.

Using a Heat Diffusion Model to Detect Potential Drug Resistance Genes of Mycobacterium tuberculosis.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is one of the oldest known and most dangerous diseases. Although the spread of TB was controlled in the early 20th century using antibiotics and vaccines, TB has again become a threat because of increased drug resistance. There is still a lack of effective treatment regimens for a person who is already infected with multidrug-resistant Mtb (MDR-Mtb) or extensively drug-resistant Mtb (XDRMtb). In the past decades, many research groups have explored the drug resistance profiles of Mtb based on sequence data by GWAS, which identified some mutations that were significantly linked with drug resistance, and attempted to explain the resistance mechanisms. However, they mainly focused on several significant mutations in drug targets (e.g. rpoB, katG). Some genes which are potentially associated with drug resistance may be overlooked by the GWAS analysis. In this article, our motivation is to detect potential drug resistance genes of Mtb using a heat diffusion model. All sequencing data, which contained 127 samples of Mtb, i.e. 34 ethambutol-, 65 isoniazid-, 53 rifampicin- and 45 streptomycin-resistant strains. The raw sequence data were preprocessed using Trimmomatic software and aligned to the Mtb H37Rv reference genome using Bowtie2. From the resulting alignments, SAMtools and VarScan were used to filter sequences and call SNPs. The GWAS was performed by the PLINK package to obtain the significant SNPs, which were mapped to genes. The P-values of genes calculated by GWAS were transferred into a heat vector. The heat vector and the Mtb protein-protein interactions (PPI) derived from the STRING database were inputted into the heat diffusion model to obtain significant subnetworks by HotNet2. Finally, the most significant (P < 0.05) subnetworks associated with different phenotypes were obtained. To verify the change of binding energy between the drug and target before and after mutation, the method of molecular dynamics simulation was performed using the AMBER software. We identified significant subnetworks in rifampicin-resistant samples. Excitingly, we found rpoB and rpoC, which are drug targets of rifampicin. From the protein structure of rpoB, the mutation location was extremely close to the drug binding site, with a distance of only 3.97 Å. Molecular dynamics simulation revealed that the binding energy of rpoB and rifampicin decreased after D435V mutation. To a large extent, this mutation can influence the affinity of drug-target binding. In addition, topA and pyrG were reported to be linked with drug resistance, and might be new TB drug targets. Other genes that have not yet been reported are worth further study. Using a heat diffusion model in combination with GWAS results and protein-protein interactions, the significantly mutated subnetworks in rifampicin-resistant samples were found. The subnetwork not only contained the known targets of rifampicin (rpoB, rpoC), but also included topA and pyrG, which are potentially associated with drug resistance. Together, these results offer deeper insights into drug resistance of Mtb, and provides potential drug targets for finding new antituberculosis drugs.

Molecular typing of drug-resistant Mycobacterium tuberculosis strains from Turkey

ObjectivesAppropriate antibiotic therapy and prevention of cross-contamination are the most important subjects in tuberculosis (TB) control. The aim of this study was to investigate the major phylogenetic clades and transmission rate of multidrug-resistant (MDR) Mycobacterium tuberculosis isolates (n = 200) from patients with TB in Sivas and Konya Provinces of Turkey. MethodsThe phylogenetic relationship among the isolates was investigated by spoligotyping method. In addition, the 24-locus mycobacterial interspersed repetitive unit–variable-number tandem repeat (MIRU-VNTR) typing method was used to reveal cross-contamination. ResultsSpoligotyping revealed 13 different spoligotypes. A total of 188 strains (94.0%) were included in the cluster. The most prominent spoligofamily was the T family (43.0% of strains), followed by LAM (26.0%), H (8.0%), X and S (both 6.0%) and U (5.0%). Also, 12 strains (6.0%) belonged to the Beijing profile. MIRU-VNTR results showed 176 (88.0%) different genotypes among the isolates. In total, 24 strains (12.0%) were in the cluster. ConclusionsAccording to spoligotyping, there is a heterogeneous M. tuberculosis population in Turkey. MIRU-VNTR results showed that cross-contamination observed between MDR M. tuberculosis isolates in Turkey is controllable.

Multidrug-resistant Mycobacterium tuberculosis: a report of cosmopolitan microbial migration and an analysis of best management practices

BackgroundTuberculosis (TB) control is a primary global health priority but the goal to eliminate TB is being threatened by the increase in incidence of multidrug-resistant tuberculosis (MDR-TB). With this series of seven MDR-TB cases in migrant patients with identical Mycobacterium tuberculosis strains we aim to illustrate the challenges encountered during therapy and follow-up: language barriers, access to care for migrant patients, depression due to isolation, adverse reactions to the treatment, management of pediatric TB, further contact tracing. We also discuss best practices for the management of complex MDR-TB cases in settings with low overall TB incidence focusing on modern diagnostic assays and an individualized and an interdisciplinary therapeutic approach.MethodsWe describe a case series of seven consecutively diagnosed MDR-TB patients, six of them treated at our tertiary care hospital between May 2018 and March 2020. Epidemiologic data was gained by semi-structured patient interviews and reconstruction of the migration route. The origin of the cluster was confirmed by genotyping of the TB-strains.ResultsSix related patients were diagnosed with pulmonary MDR-TB between May and August 2018. All had a positive Interferon-Gamma-Release Assay (IGRA), in five patients sputum microscopy was positive for acid-fast bacilli (AFB). The genetic and phenotypical drug susceptibility test did not match with MDR-TB strains from an East-African origin. The index patient was identified through genetical fingerprinting. By changing the therapy to a modern MDR-TB regime and using an interdisciplinary and culture-sensitive approach, all patients improved clinically and radiologically.ConclusionHuman migration plays an important role for the global spread of MDR-TB in low incidence countries. Early case detection and adequate treatment are key to prevention of outbreaks. Especially language barriers and complex migration routes make genotyping of TB-strains a crucial tool to identify cases clusters, the potential index patient and transmission dynamics. We are fortunate enough to experience times in which new TB-antibiotics were made available and in which molecular assays revolutionized TB-diagnostics. We need to take advantage of that and develop personalized therapies for patients suffering from drug resistant TB.

Exploring the Potential Inhibition of Candidate Drug Molecules for Clinical Investigation Based on their Docking or Crystallographic Analyses against M. tuberculosis Enzyme Targets.

Tuberculosis (TB) is a devastating disease responsible for millions of humans' deaths worldwide. It is caused by a mycobacterial organism, the tubercle bacillus or Mycobacterium tuberculosis. Although TB can be treated, cured and can be prevented if patients take prescribed medicines, scientists have never come close to wiping it out due to a sharp rise in the incidence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) mycobacterium strains. Due to long regimen treatment and emergence of MDR and XDR-TB, it is urgent to re-engineer and reposition old drugs for developing new antimycobacterial entities with novel mechanisms of action to achieve effective TB control even against the resistant forms of TB. To combat the dreadful MDR and XDR-TB, potential targets are being extensively searched for the last couple of years for the design and discovery of active potential antitubercular chemotherapeutics. To explore the disease virulence, potential new tubercular target enzymes such as InhA, MmpL3, ATP synthase, DprE1, QcrB and MenA have been taken into consideration in the present study and the structure-based design of the corresponding target inhibitors which are under clinical investigation has been attempted to identify structural features for the discovery of new chemical entities (NCEs) having specificity towards MDR and XDR Mycobacterium tuberculosis (M. tuberculosis).

Pyrazinamide Resistance and Mutation Patterns Among Multidrug-Resistant Mycobacterium tuberculosis from Henan Province.

PurposeThis study was designed to identify the phenotypic and genotypic characteristics of pyrazinamide (PZA) resistance among multidrug-resistant Mycobacterium tuberculosis (MDR-TB) from Henan and to evaluate the efficacy of pncA, rpsA, and panD mutations in predicting PZA resistance.Materials and MethodsA total of 152 MDR strains were included in this study. The Bactec MGIT system was used to determine PZA susceptibility for all strains. The pncA, rpsA, and panD genes were sequenced to identify any mutations, and the sequences were then aligned with the sequence of standard strain H37Rv. Moreover, the correlations between PZA-resistant phenotypes and treatment outcomes were analysed.ResultsOf the152 strains, 105 had a PZA-resistant phenotype, and 102 harboured the pncA mutation. The PZA resistance rate was higher in the strains with resistance to all four first-line drugs and those that were pre-extensively drug-resistant (pre-XDR) and extensively drug-resistant (XDR). A total of 100 different pncA mutation patterns were identified, including 80 point mutations and 20 insertions/deletions, and 32 new pncA mutation patterns were detected. In this study, 13 strains had multiple mutations. Of the11 PZA-resistant strains without pncA mutations, two harboured the rpsA mutation, and one harboured the panD mutation. With PZA susceptibility results as the reference, single-gene pncA sequencing had sensitivity of 89.52% and specificity of 89.36%. With the combination of rpsA and panD, the sensitivity increased to 92.38%, and the specificity remained the same. No significant differences were observed in the sputum smear/culture conversion rate between PZA-resistant patients and PZA-sensitive patients. However, PZA resistance was related to the time to sputum smear/culture conversion (P = 0.018).ConclusionThe combination of pncA, rpsA, and panD was beneficial for the timely diagnosis of PZA resistance and could provide a laboratory basis for customizing treatment regimens for MDR-TB patients.

QUANTITATIVE ANALYSIS OF GYRA AND GYRB KNOWN MUTATIONS IN DRUG-RESISTANT MYCOBACTERIUM TUBERCULOSIS STRAINS TREATED WITH OFLOXACIN

Objective: The aim of the study is to measure the minimum inhibitory concentrations (MICs) of ofloxacin antibiotic from gyrA and gyrB mutations present in fluoroquinolones (FQs) resistant strains of Mycobacterium tuberculosis (MTB) and to further concentrate the potential association between gene mutations and phenotypic resistance based on their MICs.&#x0D; Methods: Different levels of ofloxacin MICs levels were detected in 31 archived multi drug-resistant MTB isolates showing gyrA mutations in codon at A90V, S91P, D94A, D94N/Y, D94 G, and D94H and two gyrB probes (N538D and E540V). The MIC determinations were made using the 1% proportion method.&#x0D; Results: Genotypic assay detected 32 mutations in the gyrA (n=29) and gyrB (n=3) genes among the 31 FQs resistant isolates. Most frequently seen in gyrA mutations at codon D94G (n=16; 50%), these mutations had a clearly elevated MIC level from 2 to 16 μg/ml, that was phenotypically resistant. The A90V mutation region consistently exhibited the lowest levels of ofloxacin resistance, with three out of eight (37.50%) of these isolates had a MIC of &lt;2 μg/ml. In addition, a further strain of S91P mutations for MIC was determined to be less than the critical concentration (CC). These low levels of resistance have been detected in a phenotypic manner which is noticeable in the study. Furthermore, fewer mutations in codons at D94A, D94N/Y were identified. Three wild-type absent isolates from gyrB QRDR were identified and the MIC of those strains for ofloxacin was lower than the critical cutoff value.&#x0D; Conclusion: Based on the results, it is shown that different resistance mutations were associated with different levels of MICs and the current concentration for MGIT will be lowered from 2 μg/ml to 1 μg/ml for the ofloxacin drug.

Differential DNA methylomes of clinical MDR, XDR and XXDR Mycobacterium tuberculosis isolates revealed by using single-molecule real-time sequencing

Post-replicative DNA methylation is essential for diverse biological processes in both eukaryotes and prokaryotes. Mycobacterium tuberculosis (M. tuberculosis), the causative agent of tuberculosis, remains one of the most formidable threats worldwide. Although DNA methylation of M. tuberculosis has been documented, little information is available for clinical drug-resistant M. tuberculosis. Single-molecule real-time (SMRT) sequencing was used to profile the core methylome of three clinical isolates, namely multidrug-resistant (MDR), extensively drug-resistant (XDR) and extremely drug-resistant (XXDR) strains. 3812, 6808 and 6041 DNA methylated sites were identified in MDR-MTB, XDR-MTB and XXDR-MTB genome, respectively. There are two types of methylated motifs, namely N6-methyladenine (m6A) and N4-methylcytosine (m4C). A novel widespread 6 mA methylation motif 5′-CACGCAG-3′ was found in XDR-MTB and XXDR-MTB. The methylated genes are involved in multiple cellular processes, especially metabolic enzymes engaged in glucose metabolism, fatty acid and TCA cycle. Many methylated genes are involved in mycobacterial virulence, antibiotic resistance and tolerance. This provided a comprehensive list of methylated genes in drug-resistant clinical isolates and the basis for further functional elucidation.

Artemisia annua and Artemisia afra extracts exhibit strong bactericidal activity against Mycobacterium tuberculosis

Ethnopharmacological relevanceEmergence of drug-resistant and multidrug-resistant Mycobacterium tuberculosis (Mtb) strains is a major barrier to tuberculosis (TB) eradication, as it leads to longer treatment regimens and in many cases treatment failure. Thus, there is an urgent need to explore new TB drugs and combinations, in order to shorten TB treatment and improve outcomes. Here, we evaluated the potential of two Asian and African traditional medicinal plants, Artemisia annua, a natural source of artemisinin (AN), and Artemisia afra, as sources of novel antitubercular agents. Aim of the studyOur goal was to measure the activity of A. annua and A. afra extracts against Mtb as potential natural and inexpensive therapies for TB treatment, or as sources of compounds that could be further developed into effective treatments. Materials and methodsThe minimum inhibitory concentrations (MICs) of A. annua and A. afra dichloromethane extracts were determined, and concentrations above the MICs were used to evaluate their ability to kill Mtb and Mycobacterium abscessus in vitro. ResultsPrevious studies showed that A. annua and A. afra inhibit Mtb growth. Here, we show for the first time that Artemisia extracts have a strong bactericidal activity against Mtb. The killing effect of A. annua was much stronger than equivalent concentrations of pure AN, suggesting that A. annua extracts kill Mtb through a combination of AN and additional compounds. A. afra, which produces very little AN, displayed bactericidal activity against Mtb that was substantial but weaker than that of A. annua. In addition, we measured the activity of Artemisia extracts against Mycobacterium abscessus. Interestingly, we observed that while A. annua is not bactericidal, it inhibits growth of M. abscessus, highlighting the potential of this plant in combinatory therapies to treat M. abscessus infections. ConclusionOur results indicate that Artemisia extracts have an enormous potential for treatment of TB and M. abscessus infections, and that these plants contain bactericidal compounds in addition to AN. Combination of extracts with existing antibiotics may not only improve treatment outcomes but also reduce the emergence of resistance to other drugs.

Anti-tuberculosis Property of Various Extracts of Lannea against multi-drug resistant Mycobacterium tuberculosis

Tuberculosis had been a stubborn disease that human beings are combating with from ages. It is a disease that affects the pulmonary system, basically the trachea and upper respiratory organs. It is caused by bacteria called Mycobacterium tuberculosis. Bacteria in the respiratory tract will cause infection and inflammation in the respiratory passages. It leads to narrowing and pain, which results in difficulty in breathing. There had been raising the confidence of the safety and potency of the herbs and medicinal plants. We have been using herbs to treat many diseases. The diseases that are caused primarily due to the viruses and bacteria are effectively treated using herbs in the traditional systems of medicine. One such plant is Lannea coramandelica which is proven to treat many diseases and is rich in polyphenols and flavonoids. The leaves of lannea were extracted using various solvents based on the polarity. It was then investigated for the anti-bacterial property against mycobacterium tuberculosis. All the extracts showed potency by inhibiting the bacteria, but Methanol and aqueous extracts showed a significantly better activity compared to the standard.

Molecular Mechanism of Multi-Drug Resistance in Mycobacterium tuberculosis

In this study, we demonstrate the clinical applicability of molecular assays for the differential identification of M. tuberculosis isolates by Double Repetitive Element-PCR (DRE-PCR), Duplex PCR (DPCR), Random Amplified Polymorphic DNA-PCR (RAPD-PCR) and IS6110 flanking PCR and for the detection of specific codon mutations in antibiotic-resistant genes, rpoB and katG in 55 MDR-TB and 25 drug-susceptible clinical isolates by Multiplex PCR assays. The MAS-PCR assay was identified as the most prevalent rpoB gene mutations at codon 531 (83.6%), followed by codon 526 (12.7%), and no mutation was found in codon 516. Among the 55 MDR-TB isolates, 49 (89%) isolates had S315T, 5 (9%) had S315N mutations. DRE-PCR and RAPD-PCR generated similar banding of cluster III strains and suggested that MDR-TB strain genotype C may be responsible for the transmission of TB infection among the study population. PCR based differential identification of mtp40 and rpoB DPCR procedures identified two NTM strains among the isolates studied. Genotypic method DRE-PCR was found highly reproducible, followed by RAPD-PCR and mtp40, and rpoB DPCR methods effectively-identified NTM infection in this region. The presence of S315T mutation in katG gene and S531L, H526Y mutations in rpoB gene in MDR-TB isolates proved resistant phenotype. The simplicity of the MAS-PCR assay permits its implementation for the detection of resistance to INH and RIF in clinical laboratories in regions where this mutation is predominant among MDR-TB strains.

Hybrid Design of Isonicotinic Acid Hydrazide Derivatives: Machine Learning Studies, Synthesis and Biological Evaluation of their Antituberculosis Activity.

Tuberculosis (TB) is an infection disease caused by Mycobacterium tuberculosis (Mtb) bacteria. One of the main causes of mortality from TB is the problem of Mtb resistance to known drugs. The goal of this work is to identify potent small molecule anti-TB agents by machine learning, synthesis and biological evaluation. The On-line Chemical Database and Modeling Environment (OCHEM) was used to build predictive machine learning models. Seven compounds were synthesized and tested in vitro for their antitubercular activity against H37Rv and resistant Mtb strains. A set of predictive models was built with OCHEM based on a set of previously synthesized isoniazid (INH) derivatives containing a thiazole core and tested against Mtb. The predictive ability of the models was tested by a 5-fold cross-validation, and resulted in balanced accuracies (BA) of 61-78% for the binary classifiers. Test set validation showed that the models could be instrumental in predicting anti- TB activity with a reasonable accuracy (with BA = 67-79 %) within the applicability domain. Seven designed compounds were synthesized and demonstrated activity against both the H37Rv and multidrugresistant (MDR) Mtb strains resistant to rifampicin and isoniazid. According to the acute toxicity evaluation in Daphnia magna neonates, six compounds were classified as moderately toxic (LD50 in the range of 10-100 mg/L) and one as practically harmless (LD50 in the range of 100-1000 mg/L). The newly identified compounds may represent a starting point for further development of therapies against Mtb. The developed models are available online at OCHEM http://ochem.eu/article/11 1066 and can be used to virtually screen for potential compounds with anti-TB activity.