Cycloserine Resistance-related Gene In Clinical Isolated Strains Of Mycobaterium Tuberculosis

Objective:To understand the mutations of cycloserine resistance-related genes in clinical isolates of Mycobacterium tuberculosis and their correlation with drug-susceptible phenotypes,and provide a reference for the clinical application of cycloserine.Methods:Collect sputum specimens of tuberculosis patients who visited our hospital from 2017 to 2018 for acid-fast staining and culture of tuberculosis bacteria.The clinical isolates of Mycobacterium tuberculosis with drug sensitivity test results were extracted using CTAB method to extract the DNA of all clinical isolates.Whole-genome sequencing,comparing the sequencing results in the NCBI database with the standard strain H₃₇Rv sequence to understand the mutations of the cycA?alr?ald?and ddlA genes of the cycloserine resistance-related genes;anti-tuberculosis of all culture-positive mycobacteria First-line and second-line drug sensitivity tests to understand the phenotype sensitivity of isolated strains;Analyze and compare the relationship between mutations of cycloserine resistance-related genes and drug susceptibility phenotype,and understand the correlation between cycloserine resistance and rifampicin resistance.Results:In this study,123 clinical isolates of Mycobacterium tuberculosis were subjected to whole-genome sequencing.A total of 11 cycA,alr,ald,and ddlA genes of four drug-resistance-related genes of cycloserine were found to be non-synonymous mutations,as follows;A total of 5 mutation sites were detected in the cycA gene,including CGG278CTG?Arg-Leu?,TAC105TTC?Tyr-Phe?,GGG368AGG?Gly-Arg?,CGC547CAC?Arg-His?,GTG171ATG?Val-Met?,and CGG278CTG?Arg-Leu?mutation appeared in all strains,the mutation rate was 100%?123/123?;three mutation sites were detected in the alr gene,namely CAC297CGC?His-Arg?,ACC155CCC?Thr-Rro?,GTG1GGG?Val-Gly?,with a mutation rate of 2.4%?3/123?;two mutation sites were detected in the ald gene,namely GCC169ACC?Ala-Thr?and CTG149CGG?Leu-Arg?,with a mutation rate of 1.6%?2/123?;The ddlA gene ACA365TCA?Thr-Ser?mutation appeared in all tested strains with a mutation rate of100%?123/123?.Phenotypic drug sensitivity tests of 123 clinical isolates of Mycobacterium tuberculosis isolates showed that cycloserine was sensitive;71 were rifampicin-sensitive and 52 were drug-resistant.Conclusion:This study successfully obtained the genome-wide sequencing results of 123 cases of clinical isolates of M.tuberculosis,and found 11 mutations of 4 genes of cycloserine?cycA Arg278Leu,Tyr105Phe,Gly368Arg,Arg547His,and Val171Met;alr His297Arg,Thr155Rro,and Val1Gly;ald Ala169Thr And Leu149Arg;and ddlA Thr365Ser?have no obvious relationship with cycloserine phenotype resistance.In addition,this study shows that rifampicin phenotypic resistance is not significantly related to cycloserine resistance.This study suggests that the incidence of cycloserine resistance is low and has good clinical application value.