Application value of matrix-assisted laser desorption ionization time-of-flight mass spectrometry in the detection of drug resistance of Mycobacterium tuberculosis
Objective To explore the matrix-assisted laser desorption ionization time-of-flight mass spectrometry(MALDI-TOF-MS)platform for identifying Mycobacterium tuberculosis and detecting drug-resistant mutations and evaluate its application value.Methods MALDI-TOF-MS was used to detect three national reference strains,and the minimum detection limits for tuberculosis-specific identifying genes and drug-resistant mutations were determined.Respiratory tract clinical specimens of suspected tuberculosis patients were collected from January to September 2023 in the Tuberculosis Department of Kunming Third People's Hospital by non-random sampling method,including 120 spu-tum samples and 100 bronchoalveolar lavage fluid samples,with the content of 3-5 mL in each sample.Nucleic acid was extracted from all the samples,and the tuberculosis positive samples with Ct value≤32 were identified by qPCR kit for tuberculosis identification.The drug-resistant mutations of rifampicin(RIF),isoniazid(INH),ethambutol(EMB),moxifloxacin(MXF),streptomycin(SM)and pyrazinamide(PZA)were detected by MALDI-TOF-MS.The pheno-typic drug susceptibility test(pDST)and third-generation sequencing(nanopore platfom)were used to detect the TB-positive samples,and the results were compared with those of MALDI-TOF-MS to verify its accuracy.Results The lowest detection limit of MALDI-TOF-MS was 60-100 CFU/mL for MTB identification genes IS6110 and ext_RD9,and 100-1000 CFU/mL for drug-resistance mutations.A tolal of 132 tuberculosis positive samples with Ct value≤32 were screened by qPCR.Compared with pDST results,the sensitivity,specificity and kappa value of MALDI-TOF-MS for detection of drug-resistant mutations were 93.75%,100%and 0.96 for RIF,98.33%,90.28%and 0.88 for INH,83.33%,97.50%and 0.78 for EMB and 95.45%,100%and 0.97 for MXF,respectively.Compared with nanopore se-quencing,the sensitivity,specificity and kappa values of MALDI-TOF-MS for detection of drug-resistant mutations were 90.91%,100%and 0.94 for RIF,92.96%,100%and 0.92 for INH,92.86%,100%and 0.96 for EMB,95.45%,100%and 0.97 for MXF,95.74%,100%and 0.97 for SM,66.67%,100%and 0.80 for PZA,respectively.Conclusion MALDI-TOF-MS can be used as an effective method for rapid detection of Mycobacterium tuberculosis drug-resistant mutations because its results were highly consistent with those of pDST and nanopore sequencing.
Matrix-assisted laser desorption ionization time-of-flight mass spectrometryMycobacterium tuberculosisDrug resistancePhentypic drug susceptibility testNanopore sequencing
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