The relationship between embb306 and embb406 mutations and ethambutol resistant in Mycobacterium tuberculosis isolated from patiens in west of Iran.

Background: Mutations in embB gene have been reported in ethambutol (EMB) resistance Mycobacterium tuberculosis (M. tuberculosis) isolates. The aim of this study was survey on embB 306 and 406 EMB resistant M. tuberculosis isolated from patients in West of Iran (2014-2015). Methods: Fifty strains of M. tuberculosis from patients with pulmonary tuberculosis (TB) were considered. Drug susceptibility using proportional method was performed. Polymerase chain reaction (PCR) -DNA sequencing was applied for mutation in embB 306 and 406 codon detection. Data were analyzed in SPSS 16 software using descriptive statistics and Fisher's exact test (p<0.05). Results: In this study 7 (14%) M. tuberculosis isolates were resistant to EMB. 6 (85.71%) and 1 (14.28%) resistant isolates had embB 306 and 304 codon mutations, respectively. Between embB306 mutations and resistance to EMB and MDR isolates had a significant relationship (p<0.001). Conclusion: The data indicated that embB 306 mutations have effect on EMB resistant. Detection of EMB resistant and these mutations prominent for antibiotic prescription.

Structural dynamics insights into the M306L, M306V, and D1024N mutations in Mycobacterium tuberculosis inducing resistance to ethambutol.

Resistance to anti-tuberculosis drugs, especially ethambutol (EMB), has been widely reported worldwide. EMB resistance is caused by mutations in the embB gene, which encodes the arabinosyl transferase enzyme. This study aimed to detect mutations in the embB gene of Mycobacterium tuberculosis from Papua and to evaluate their impact on the effectiveness of EMB. We analyzed 20 samples of M. tuberculosis culture that had undergone whole-genome sequencing, of which 19 samples were of sufficient quality for further bioinformatics analysis. Mutation analysis was performed using TBProfiler, which identified M306L, M306V, D1024N, and E378A mutations. In sample TB035, the M306L mutation was present along with E378A. The binding affinity of EMB to arabinosyl transferase was calculated using AutoDock Vina. The molecular docking results revealed that all mutants demonstrated an increased binding affinity to EMB compared to the native protein (-0.948 kcal/mol). The presence of the M306L mutation, when coexisting with E378A, resulted in a slight increase in binding affinity compared to the M306L mutation alone. The molecular dynamics simulation results indicated that the M306L, M306L + E378A, M306V, and E378A mutants decreased protein stability. Conversely, the D1024N mutant exhibited stability comparable to the native protein. In conclusion, this study suggests that the M306L, M306L + E378A, M306V, and E378A mutations may contribute to EMB resistance, while the D1024N mutation may be consistent with continued susceptibility to EMB.

Frequency of Codon 306 Mutations in embB Gene of Mycobacterium tuberculosis Resistant to Ethambutol: A Systematic Review and Meta-Analysis.

BACKGROUND: Ethambutol (EMB) resistance is a major concern in patients with tuberculosis (TB). The aim of this study was to determine the frequency rate of mutations in the embB306 gene of Mycobacterium tuberculosis (M. tuberculosis) resistant to EMB, based on a systematic review and meta-analysis. METHODS: Thirty-seven original articles (1997-2015) that have been published in valid databases were considered for this research. The articles were systematically reviewed for the prevalence and rate of mutations in embB306 in EMB-resistant M. tuberculosis. Data were analyzed using meta-analysis and random effects models (CI 95%, P < 0.10). RESULTS: With a 6,931 sample size in 37 original articles, the lowest rate was related to EMB resistance that was observed in 2014 with 0.05 (95% CI: 0.04-0.07) and the highest prevalence rate was 0.84 (95% CI: 0.68-1.01), observed in 1997. Lowest and highest prevalence rates of embB306 gene mutation in M. tuberculosis were 0.03 (95% CI: 0.01-0.07) in 2014 and 0.78 (95% CI: 0.71-1.84) in 2005, in the USA, respectively. CONCLUSIONS: The present study revealed the prevalence and association of mutations in the embB306 gene of M. tuberculosis with resistance to EMB. Detecting EMB-resistant M. tuberculosis can help in controlling and correcting the administration of drugs for patients with TB.

Rapid detection of ethambutol-resistant Mycobacterium tuberculosis in clinical specimens by real-time polymerase chain reaction hybridisation probe method.

Background: Early diagnosis of drug resistance (DR) to ethambutol (EMB) in tuberculosis (TB) remains a challenge. Simple and reliable method (s) are needed for rapid detection of DR Mycobacterium tuberculosis (MTB) in clinical specimens. Objectives: The aim of this study was to design fluorescence resonance energy transfer hybridisation probe-based real-time polymerase chain reaction (PCR) method for the early detection of EMB-resistant MTB direct from clinical sputa. Materials and Methods: Primers and probes were designed against 306 codon of embB gene which is commonly associated with EMB resistance. A comparative study was done between Lowenstein-Jenson (L-J) proportion and hybridisation probe-based real-time PCR method for susceptibility testing. DNA sequencing was used in nine representative isolates to validate the efficiency of real-time PCR method to detect emb306 mutation of MTB. Results: A total of 52 clinical sputum samples and corresponding culture isolates (from category II pulmonary TB cases) were included in this study. Out of 52 MTB isolates, 32 and 20 were resistant and susceptible to EMB, respectively, as determined by L-J proportion method. Real-time PCR showed 95% specificity, 75% sensitivity and 82.69% accuracy when compared with L-J proportion method. A 100% of concordance was observed by validating the real-time PCR results with DNA sequencing. Conclusions: Our real-time PCR hybridisation probe method promises for rapid detection of EMB-resistant MTB directly from clinical specimens. However, future studies and modifications of method by incorporating other potential loci along with targeted mutation (emb306) are still required to increase the sensitivity of method.

Mutations in embB gene associated with resistance to ethambutol in Mycobacterium tuberculosis strains isolated from TB patients in the west of Iran (2014-15).

INTRODUCTION: Mutations in embB gene, especially those in ethambutol resistance-determining region (ERDR), are known as "hot spots". These mutations have frequently been reported in EMB-resistant M. tuberculosis isolates, using the Sequence analysis as a precise and effective method. The aim of this study was to detect mutations in embB gene associated with resistance to ethambutol in Mycobacterium tuberculosis strains isolated from TB patients in the west of Iran (2014-15). MATERIAL AND METHODS: This study was performed on smear-positive patients of the west side of Iran, in the TB research center located in Kermanshah city, during 2014-15. Out of 1069 strains of Mycobacterium tuberculosis, 50 strains with pulmonary TB were selected and evaluated (22 men and 28 women; aged between 23 and 86; median age: 54.5years). RESULTS: Mutation in the embB gene was detected in all of the seven EMB-resistance isolates and five (42.71%) cases of them were MDR. The most frequent substantiation of amino acid occurred at the codon 306 in six (64.85%) of the EMB-resistant isolates. The Met306Val substitution resulting from an A→G transition was detected in three (42.85%) EMB-resistant isolates; and the Met306Ile substitution, due to a G→A transition was also detected in three (42.85%) resistant isolates. No mutations at the embB gene were detected in susceptible strains. CONCLUSION: Our results were similar to those that were regularly reported in earlier studies. The only mutations in the EMB-resistant isolates were found in embB 306 and 406 codons. Substantiation amino acids at codon 306 were the most frequent. The data indicated that embB 306 mutations are sufficient to induce ethambutol resistance, and detection of these mutations is advisable to be considered in the development of rapid molecular tests. Sequence analysis of the ERDR in the embB gene is adequate for rapid detection of EMB resistance, and mutations in the codon 306 are good predictive markers for resistance to EMB.