Enhanced respiration prevents drug tolerance and drug resistance in Mycobacterium tuberculosis

Persistence, manifested as drug tolerance, represents a significant obstacle to global tuberculosis control. The bactericidal drugs isoniazid and rifampicin kill greater than 99% of exponentially growing Mycobacterium tuberculosis (Mtb) cells, but the remaining cells are persisters, cells with decreased metabolic rate, refractory to killing by these drugs, and able to generate drug-resistant mutants. We discovered that the combination of cysteine or other small thiols with either isoniazid or rifampicin prevents the formation of drug-tolerant and drug-resistant cells in Mtb cultures. This effect was concentration- and time-dependent, relying on increased oxygen consumption that triggered enhanced production of reactive oxygen species. In infected murine macrophages, the addition of N-acetylcysteine to isoniazid treatment potentiated the killing of Mtb. Furthermore, we demonstrate that the addition of small thiols to Mtb drug treatment shifted the menaquinol/menaquinone balance toward a reduced state that stimulates Mtb respiration and converts persister cells to metabolically active cells. This prevention of both persister cell formation and drug resistance leads ultimately to mycobacterial cell death. Strategies to enhance respiration and initiate oxidative damage should improve tuberculosis chemotherapies.

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Analysis of drug resistance and mutation profiles in Mycobacterium tuberculosis isolates in a surveillance site in Beijing, China

Journal of International Medical Research ◽

10.1177/0300060520984932 ◽

2021 ◽

Vol 49 (1) ◽

pp. 030006052098493

Author(s):

Jie Zhang ◽

Yixuan Ren ◽

Liping Pan ◽

Junli Yi ◽

Tong Guan ◽

...

Keyword(s):

Drug Resistance ◽

Mycobacterium Tuberculosis ◽

Drug Testing ◽

Multidrug Resistant ◽

High Rate ◽

Drug Resistant ◽

Surveillance Site ◽

Mdr Tb ◽

Previously Treated Patients ◽

Beijing China

Objective This study analyzed drug resistance and mutations profiles in Mycobacterium tuberculosis isolates in a surveillance site in Huairou District, Beijing, China. Methods The proportion method was used to assess drug resistance profiles for four first-line and seven second-line anti-tuberculosis (TB) drugs. Molecular line probe assays were used for the rapid detection of resistance to rifampicin (RIF) and isoniazid (INH). Results Among 235 strains of M. tuberculosis, 79 (33.6%) isolates were resistant to one or more drugs. The isolates included 18 monoresistant (7.7%), 19 polyresistant (8.1%), 28 RIF-resistant (11.9%), 24 multidrug-resistant (MDR) (10.2%), 7 pre-extensively drug-resistant (XDR, 3.0%), and 2 XDR strains (0.9%). A higher rate of MDR-TB was detected among previously treated patients than among patients with newly diagnosed TB (34.5% vs. 6.8%). The majority (62.5%) of RIF-resistant isolates exhibited a mutation at S531L in the DNA-dependent RNA polymerase gene. Meanwhile, 62.9% of INH-resistant isolates carried a mutation at S315T1 in the katG gene. Conclusion Our results confirmed the high rate of drug-resistant TB, especially MDR-TB, in Huairou District, Beijing, China. Therefore, detailed drug testing is crucial in the evaluation of MDR-TB treatment.

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embB Gene association with Ethambutol drug Resistance in Mycobacterium Tuberculosis Patients

10.53350/pjmhs2115123273 ◽

2021 ◽

Vol 15 (12) ◽

pp. 3273-3276

Author(s):

Sana Hafeez ◽

Haleema Sajid ◽

Farouk Qamar Malik ◽

Imran Ali Zaidi ◽

Sobia Niaz ◽

...

Keyword(s):

Drug Resistance ◽

Mycobacterium Tuberculosis ◽

Codon Position ◽

Mutational Analysis ◽

Transmission Rate ◽

Drug Resistant ◽

Tuberculosis Patients ◽

Two Samples ◽

Suspected Tuberculosis ◽

Embb Gene

Background: Tuberculosis (TB) is fatal and life threatening infectious disease. The transmission rate of tuberculosis is very high. Various drugs are used as treatment for TB. Recently it has been observed that one of the most important factor for fast TB spread is development of anti-TB drug resistant mycobacterium tuberculosis (MTB). Various combination of drugs like isoniazid (INH), rifampicin (RIF), Streptomycin(SM), pyrazinamide (PZA) or ethambutol (EMB) are in global use for TB treatment. Improper usage of these drugs makes the person prone to develop anti-TB drug resistant tuberculosis. Aim: To evaluate association of embB gene with ethambutol resistance in Mycobacterium Tuberculosis. Methods: 104 Specimens of sputum from suspected tuberculosis patients were processed for inoculation in Lowenstein J Medium after it has been decontaminated properly. Kit method by using QIAamp DNA Mini kit was utilized for extraction of DNA. Then region from base 6953 to 10249 of embB gene was amplified through PCR and then followed by sequencing with the aid of softwares blast2seq and ClustalW2. Three primer sets were utilized to amplify embB gene. Ethambutol (EMB) Resistant MTB specimens were processed to study mutation in embB gene. Results: Out of the total 104 sputum specimens, 14 samples were found to have ethambutol resistance. These 14 samples were then processed for mutational analysis. DNA sequence analysis of these 14 samples confirmed embB gene mutation in 10 samples. Mutational analysis revealed that 08 samples showed mutation at codon 306 and two samples showed mutation at 319 codon. The reported mutation Methionine →Isoleucine was seen in 07 samples with ATG codon replaced by ATA codon at codon position 306. One sample showed mutation as Methionine →Isoleucine with ATG codon replaced by ATC codon at codon position 306. Two samples showed mutation as Tyrosine →Serine with TAT codon replaced by TCT at 319 codon position in embB gene. Conclusion: This study concludes that mutation of certain genes particularly point mutation of embB gene at codon 306 and 319 is associated with drug resistance of ethambutol in ethambutol resistant mycobacterium tuberculosis patients. Keywords: Ethambutol, embB gene, Mycobacterium tuberculosis.

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Genotypes of Mycobacterium tuberculosis isolates circulating in Shaanxi Province, China

PLoS ONE ◽

10.1371/journal.pone.0242971 ◽

2020 ◽

Vol 15 (12) ◽

pp. e0242971

Author(s):

Yan Li ◽

Yu Pang ◽

Tianhua Zhang ◽

Xiaoping Xian ◽

Jian Yang ◽

...

Keyword(s):

Genetic Diversity ◽

Drug Resistance ◽

Mycobacterium Tuberculosis ◽

Multidrug Resistant ◽

Shaanxi Province ◽

Drug Resistant ◽

Beijing Family ◽

Tuberculosis Patients ◽

Smear Positive Pulmonary Tuberculosis ◽

Discriminatory Index

Objectives The prevalence of drug-resistant TB in Shaanxi Province is higher than other areas. This study was aimed to investigate the genetic diversity and epidemiology of Mycobacterium tuberculosis clinical strains in Shaanxi Province, China. Methods From January to December 2016, a total of 298 Mycobacterium tuberculosis clinical isolates from smear-positive pulmonary tuberculosis patients were genotyped by Mcspoligotyping and 15-locus VNTR. Results We found that the Beijing family strains was the most prominent family(81.54%, 243/298). Other family strains included T family(9.06%, 27/298), U family(0.67%, 2/298), LAM9 family(0.34%, 1/298) and Manu family(0.34%, 1/298). The rates of multidrug-resistant (MDR) M.Tuberculosis, age, type of case and education between Beijing and non-Beijing family strains were not statistically different, while the distribution in the three different regions among these was statistically significant. VNTR results showed that strains were classified into 280 genotypes, and 33 (11.07%) strains could be grouped into 14 clusters. 11 of the 15-VNTR loci were highly or moderately discriminative according to the Hunter-Gaston discriminatory index. Conclusions We concluded that the Beijing family genotype was the most prevalent genotype and 15-locus VNTR typing might be suitable for genotyping of M. tuberculosis in Shaanxi Province. There was less association between Beijing family genotypes and drug resistance in our study area.

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Evaluation of Pyrosequencing for Detecting Extensively Drug-Resistant Mycobacterium tuberculosis among Clinical Isolates from Four High-Burden Countries

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.03614-14 ◽

2014 ◽

Vol 59 (1) ◽

pp. 414-420 ◽

Cited By ~ 20

Author(s):

Kanchan Ajbani ◽

Shou-Yean Grace Lin ◽

Camilla Rodrigues ◽

Duylinh Nguyen ◽

Francine Arroyo ◽

...

Keyword(s):

Drug Resistance ◽

Mycobacterium Tuberculosis ◽

The Philippines ◽

Drug Susceptibility Testing ◽

Clinical Isolates ◽

Drug Resistant ◽

Second Line ◽

Additional Advantage ◽

Content Type ◽

Extensively Drug Resistant

ABSTRACTReliable molecular diagnostics, which detect specific mutations associated with drug resistance, are promising technologies for the rapid identification and monitoring of drug resistance inMycobacterium tuberculosisisolates. Pyrosequencing (PSQ) has the ability to detect mutations associated with first- and second-line anti-tuberculosis (TB) drugs, with the additional advantage of being rapidly adaptable for the identification of new mutations. The aim of this project was to evaluate the performance of PSQ in predicting phenotypic drug resistance in multidrug- and extensively drug-resistant tuberculosis (M/XDR-TB) clinical isolates from India, South Africa, Moldova, and the Philippines. A total of 187 archived isolates were run through a PSQ assay in order to identifyM. tuberculosis(via the IS6110marker), and to detect mutations associated with M/XDR-TB within small stretches of nucleotides in selected loci. The molecular targets includedkatG, theinhApromoter and theahpC-oxyRintergenic region for isoniazid (INH) resistance; therpoBcore region for rifampin (RIF) resistance;gyrAfor fluoroquinolone (FQ) resistance; andrrsfor amikacin (AMK), capreomycin (CAP), and kanamycin (KAN) resistance. PSQ data were compared to phenotypic mycobacterial growth indicator tube (MGIT) 960 drug susceptibility testing results for performance analysis. The PSQ assay illustrated good sensitivity for the detection of resistance to INH (94%), RIF (96%), FQ (93%), AMK (84%), CAP (88%), and KAN (68%). The specificities of the assay were 96% for INH, 100% for RIF, FQ, AMK, and KAN, and 97% for CAP. PSQ is a highly efficient diagnostic tool that reveals specific nucleotide changes associated with resistance to the first- and second-line anti-TB drug medications. This methodology has the potential to be linked to mutation-specific clinical interpretation algorithms for rapid treatment decisions.

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Role of embB Codon 306 Mutations in Mycobacterium tuberculosis Revisited: a Novel Association with Broad Drug Resistance and IS6110 Clustering Rather than Ethambutol Resistance

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.49.9.3794-3802.2005 ◽

2005 ◽

Vol 49 (9) ◽

pp. 3794-3802 ◽

Cited By ~ 79

Author(s):

Manzour Hernando Hazbón ◽

Miriam Bobadilla del Valle ◽

Marta Inírida Guerrero ◽

Mandira Varma-Basil ◽

Ingrid Filliol ◽

...

Keyword(s):

Drug Resistance ◽

Mycobacterium Tuberculosis ◽

Odds Ratio ◽

Univariate Analysis ◽

Drug Susceptibility ◽

Drug Resistant ◽

Development Of Resistance ◽

Resistance Patterns ◽

Novel Association

ABSTRACT Mutations at position 306 of embB (embB306) have been proposed as a marker for ethambutol resistance in Mycobacterium tuberculosis; however, recent reports of embB306 mutations in ethambutol-susceptible isolates caused us to question the biological role of this mutation. We tested 1,020 clinical M. tuberculosis isolates with different drug susceptibility patterns and of different geographical origins for associations between embB306 mutations, drug resistance patterns, and major genetic group. One hundred isolates (10%) contained a mutation in embB306; however, only 55 of these mutants were ethambutol resistant. Mutations in embB306 could not be uniquely associated with any particular type of drug resistance and were found in all three major genetic groups. A striking association was observed between these mutations and resistance to any drug (P < 0.001), and the association between embB306 mutations and resistance to increasing numbers of drugs was highly significant (P < 0.001 for trend). We examined the association between embB306 mutations and IS6110 clustering (as a proxy for transmission) among all drug-resistant isolates. Mutations in embB306 were significantly associated with clustering by univariate analysis (odds ratio, 2.44; P = 0.004). In a multivariate model that also included mutations in katG315, katG463, gyrA95, and kasA269, only mutations in embB306 (odds ratio, 2.14; P = 0.008) and katG315 (odds ratio, 1.99; P = 0.015) were found to be independently associated with clustering. In conclusion, embB306 mutations do not cause classical ethambutol resistance but may predispose M. tuberculosis isolates to the development of resistance to increasing numbers of antibiotics and may increase the ability of drug-resistant isolates to be transmitted between subjects.

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Spatiotemporal dynamics of drug‐resistant Mycobacterium tuberculosis : Contrasting trends and implications for tuberculosis control in EU high‐priority country

Transboundary and Emerging Diseases ◽

10.1111/tbed.13758 ◽

2020 ◽

Cited By ~ 1

Author(s):

Igor Mokrousov ◽

Anna Vyazovaya ◽

Klavdia Levina ◽

Alena Gerasimova ◽

Viacheslav Zhuravlev ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Spatiotemporal Dynamics ◽

Drug Resistant ◽

Tuberculosis Control

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Transient drug-tolerance and permanent drug-resistance rely on the trehalose-catalytic shift in Mycobacterium tuberculosis

Nature Communications ◽

10.1038/s41467-019-10975-7 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 11

Author(s):

Jae Jin Lee ◽

Sun-Kyung Lee ◽

Naomi Song ◽

Temitope O. Nathan ◽

Benjamin M. Swarts ◽

...

Keyword(s):

Drug Resistance ◽

Mycobacterium Tuberculosis ◽

Drug Tolerance

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Association between embB Codon 306 Mutations and Drug Resistance in Mycobacterium tuberculosis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01516-06 ◽

2007 ◽

Vol 51 (7) ◽

pp. 2618-2620 ◽

Cited By ~ 36

Author(s):

Xin Shen ◽

Guo-miao Shen ◽

Jie Wu ◽

Xiao-hong Gui ◽

Xia Li ◽

...

Keyword(s):

Drug Resistance ◽

Mycobacterium Tuberculosis ◽

Odds Ratio ◽

Rapid Detection ◽

Multidrug Resistant ◽

Drug Resistant ◽

Drug Resistant Tuberculosis ◽

Candidate Marker ◽

Resistant Tuberculosis

ABSTRACT embB306 mutants were detected in both ethambutol (EMB)-resistant and EMB-susceptible strains of Mycobacterium tuberculosis. Multidrug-resistant (MDR) strains had a higher proportion of embB306 mutants than non-MDR strains (odds ratio, 6.78; P < 0.001). The embB306 locus is a candidate marker for rapid detection of MDR and extremely drug resistant tuberculosis.

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Molecular Mechanism of DAPD/DXG against Zidovudine- and Lamivudine- Drug Resistant Mutants: A Molecular Modelling Approach

Antiviral Chemistry and Chemotherapy ◽

10.1177/095632020201300205 ◽

2002 ◽

Vol 13 (2) ◽

pp. 115-128 ◽

Cited By ~ 11

Author(s):

Youhoon Chong ◽

Katyna Borroto-Esoda ◽

Phillip A Furman ◽

Raymond F Schinazi ◽

Chung K Chu

Keyword(s):

Drug Resistance ◽

Amino Acid ◽

Antiviral Activity ◽

Molecular Mechanism ◽

Binding Affinity ◽

Molecular Modelling ◽

Enzyme Inhibitor ◽

Drug Resistant ◽

Amino Acid Residues ◽

Resistant Mutants

In order to understand molecular mechanism of antiviral drug resistance of HIV-1 reverse transcriptase (RT) as well as potent antiviral activity of 2,6-diaminopurine dioxolane (DAPD) [prodrug of (–)-β-D-dioxolane guanine (DXG)] against drug-resistant RTs, molecular modelling studies of three structurally distinct nucleoside RT inhibitor (NRTI)-triphosphates (TP) [zidovudine (AZT)-TP, lamivudine (3TC)-TP and DXG-TP] complexed with the wild-type (WT) and mutated RT were conducted. The computational analyses indicated that the antiviral activity and the calculated relative binding energy of the RT inhibitor triphosphates can be correlated, and the minimized structures gave information on the molecular mechanism of drug resistance conferred by mutations. The interactions between the NRTI-TP and adjacent amino acid residues (Lys65, Lys70, Arg72, Tyr115 and/or Gln151) played important roles in stabilizing the enzyme—inhibitor complex. Particularly, Arg72 was found to stabilize the dioxolane and oxathiolane sugar moiety through hydrogen bonding, which was responsible for favourable binding affinity of DXG-TP to AZT- as well as 3TC-resistant mutants. The conformational changes in these amino acid residues caused by mutation always affected the changes in the tertiary structures of enzyme-inhibitor complexes through either closing or opening the gap between the fingers and palm domains. The enzyme-inhibitor complexes with good binding affinity showed tight binding modes by closing the gap between the two domains, whereas weak inhibitors gave open and loose complexes.

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