Application of matrix assisted laser desorption ionization time of flight mass spectrometry in the identification of <i>Mycobacterium tuberculosis</i>_West China Medical Journal

Authors：

DAI Zhongqiu , TANG Sishi , FAN Yujie , SONG Qifei , WANG Yuanfang , YUAN Yu , SHU Ling ,  XIE Yi

Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P. R. China;

Corresponding?author：

XIE Yi, Email: xie_yi_77@163.com

Keywords：

Matrix assisted laser desorption ionization time of flight mass spectrometry; Identification of mycobacteria; Mycobacterium tuberculosis complex; Non-tuberculous mycobacteria

DOI：

10.7507/1002-0179.201907080

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Objective To evaluate the accuracy and practicability of matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) in clinical isolates of mycobacteria.Methods We collected all tested strains, which were positive for Mycobacterium tuberculosis culture and positive for acid-fast staining, from West China Hospital of Sichuan University from 2014 to 2017, eliminating duplicate strains sent by the same patient at the same time. The traditional method was used with the P-nitrobenzoic acid (PNB)/ 2-Thiophenecarboxylic acid hydrazide (TCH) indicator to initially identify acid-resistant positive strains. Mycobacteria was identified by MALDI-TOF MS; the specificity and sensitivity of the MALDI-TOF MS was analyzed by duplex primer-polymerase chain reaction (Duplex-PCR) method and DNA sequencing method as the "gold standard" for the identification.Results A total of 237 anti-acid positive strains were collected; Mycobacterium tuberculosis complex (MTC) and non-tuberculous mycobacteria (NTM) were identified by mycobacterium double primer PCR, and NTM was identified by 16S rRNA gene sequencing. There were 218 cases of MTC and 19 cases of NTM. The results of preliminary identification using the traditional identification method of PNB/TCH indicator showed that there were 209 cases of MTC (with the sensitivity of 95.9%, specificity of 100.0%, positive predictive value of 100.0%, and negative predictive value of 67.9%) and 28 cases of NTM (with the sensitivity of 100.0%, specificity of 95.9%, positive predictive value of 67.9%, and negative predictive value of 100.0%). The results of MALDI-TOF MS method indicated that there were 199 cases of MTC (with the sensitivity of 91.3%, specificity of 100.0%, positive predictive value of 100.0%, and negative predictive value of 50.0%), 32 cases of NTM (with the sensitivity of 68.4%. specificity of 94.0%, positive predictive value of 40.6%, and negative predictive value of 97.1%), and 6 cases of others. There were 168 strains (84.4%) with the identification score>1.9 obtained by MALDI-TOF MS method.Conclusions MALDI-TOF MS is a better method for identifying mycobacteria, which has the same identification results as the traditional methods, and has low cost and is suitable for routine use in clinical microbiology laboratories.

Citation： DAI Zhongqiu, TANG Sishi, FAN Yujie, SONG Qifei, WANG Yuanfang, YUAN Yu, SHU Ling, XIE Yi. Application of matrix assisted laser desorption ionization time of flight mass spectrometry in the identification of Mycobacterium tuberculosis. West China Medical Journal, 2019, 34(8): 844-849. doi: 10.7507/1002-0179.201907080 Copy

1.	World Health Organization. Global tuberculosis report 2017. (2017-10-30)[2019-07-03]. https://www.who.int/tb/publications/global_report/gtbr2017_main_text.pdf.
2.	全國第五次結核病流行病學抽樣調查技術指導組, 全國第五次結核病流行病學抽樣調查辦公室. 2010年全國第五次結核病流行病學抽樣調查報告. 中國防癆雜志, 2012, 34(8): 485-508.
3.	李玉靜, 馬京華, 張彥坤, 等. 8種結核病實驗室檢測方法診斷價值的對比分析. 醫學動物防制, 2017, 33(1): 25-28, 32.
4.	Body BA, Beard MA, Slechta ES, et al. Evaluation of the vitek MS v3.0 matrix-assisted laser desorption ionization-time of flight mass spectrometry system for identification of mycobacterium and nocardia species. J Clin Microbiol, 2018, 56(6): pii: e00237-18.
5.	中國防癆協會. 結核病診斷細菌學檢驗規程. 中國防癆雜志, 1996, 18(2): 80-85.
6.	李道群, 夏雪山, 宋玉竹, 等. 結核分枝桿菌分子分型技術的研究和應用. 中國人獸共患病學報, 2016, 32(1): 76-82.
7.	Kim BJ, Hong SK, Lee KH, et al. Differential identification of Mycobacterium tuberculosis complex and nontuberculous mycobacteria by duplex PCR assay using the RNA polymerase gene (rpoB). J Clin Microbiol, 2004, 42(3): 1308-1312.
8.	Lee AS, Jelfs P, Sintchenko V, et al. Identification of non-tuberculous mycobacteria: utility of the GenoType Mycobacterium CM/AS assay compared with HPLC and 16S rRNA gene sequencing. J Med Microbiol, 2009, 58(Pt 7): 900-904.
9.	Petti CA, Bosshard PP, Brandt ME, et al. Interpretive criteria for identification of bacteria and fungi by DNA target sequencing; approved guideline. Clinical and Laboratory Standards Institute (CLSI), 2008.
10.	Keys CJ, Dare DJ, Sutton H, et al. Compilation of a MALDI-TOF Mass spectral database for the rapid screening and characterization of bacteria implicated in human infectious diseases. Infect Genet Evol, 2004, 4(3): 221-224.
11.	Mediavilla-Gradolph MC, De Toro-Peinado I, Bermúdez-Ruiz MP, et al. Use of MALDI-TOF MS for identification of nontuberculous mycobacterium species isolated from clinical specimens. BioMed Res Int, 2015, 2015: 854078.
12.	周昭彥, 胡必杰, 鮑容, 等. MALDI-TOF質譜快速鑒定非結核分枝桿菌方法的建立和評價. 中華檢驗醫學雜志, 2013, 36(7): 610-614.
13.	Huang TS, Lee CC, Tu HZ, et al. Rapid identification of mycobacteria from positive MGIT broths of primary cultures by MALDI-TOF mass spectrometry. PLoS ONE, 2018, 13(2): e0192291.
14.	劉國威, 白大鵬, 梅早仙. 非結核分枝桿菌肺病的診斷與治療研究進展. 中國城鄉企業衛生, 2016, 31(6): 18-21.
15.	劉國標, 羅春明, 鄧虹, 等. 抗結核治療對非結核分枝桿菌肺病的療效觀察. 現代醫院, 2017, 17(3): 354-356.
16.	Vijay GS Kumar, Tejashree A Urs, Rajani R Ranganath. MPT 64 antigen detection for rapid confirmation of M.tuberculosis isolates. BMC Res Notes, 2011, Vol.4(1).
17.	?aml? A, ?lki A. Comparison of MALDI-TOF MS, nucleic acid hybridization and the MPT64 immunochromatographic test for the identificationof M. Tuberculosis and non-tuberculosis Mycobacterium species. New Microbiol, 2016, 4(39): 259-263.
18.	王雷, 施旭東. 水溶性對硝基苯甲酸在結核分枝桿菌菌種鑒定中的應用. 臨床醫學工程, 2010, 17(11): 47-48.
19.	蔡靜, 張鑫, 魏黛玉, 等. 甘肅省53 株非結核分枝桿菌對9 種常用抗結核藥物的敏感性分析. 疾病監測, 2018, 33(6): 515-519.
20.	Carbonnelle E, Mesquita C, Bille E, et al. MALDI-TOF mass spectrometry tools for bacterial identification in clinical microbiology laboratory. Clin Biochem, 2010, 44(1): 104-109.
21.	Akyar I, ?avu?o?lu C, Aya? M, et al. Evaluation of the performance of MALDI-TOF MS and DNA sequence analysis in the identification of mycobacteria species. Turk J Med Sci, 2018, 48(6): 1351-1357.
22.	van Veen SQ, Claas EC, Kuijper EJ, et al. High-throughput identification of bacteria and yeast by matrix-assisted laser desorption ionization-time of flight mass spectrometry in conventional medical microbiology laboratories. J Clin Microbiol, 2010, 48(3): 900-907.
23.	王勝芬, 趙雁林, 黃海榮, 等. 結核分枝桿菌北京基因型菌株與耐藥表型的關系. 中國醫學科學院學報, 2009, 31(4): 427-431.
24.	劉棟賓, 向延根, 馬小華, 等. 結核分枝桿菌L型特征及臨床意義研究進展. 實用預防醫學, 2018, 25(9): 1150-1153, 封3.
25.	Genc GE, Demir M, Yaman G, et al. Evaluation of MALDI-TOF MS for identification of nontuberculous mycobacteria isolated from clinical specimens in mycobacteria growth indicator tube medium. New Microbiol, 2018, 41(3): 214-219.

1. World Health Organization. Global tuberculosis report 2017. (2017-10-30)[2019-07-03]. https://www.who.int/tb/publications/global_report/gtbr2017_main_text.pdf.
2. 全國第五次結核病流行病學抽樣調查技術指導組, 全國第五次結核病流行病學抽樣調查辦公室. 2010年全國第五次結核病流行病學抽樣調查報告. 中國防癆雜志, 2012, 34(8): 485-508.
3. 李玉靜, 馬京華, 張彥坤, 等. 8種結核病實驗室檢測方法診斷價值的對比分析. 醫學動物防制, 2017, 33(1): 25-28, 32.
4. Body BA, Beard MA, Slechta ES, et al. Evaluation of the vitek MS v3.0 matrix-assisted laser desorption ionization-time of flight mass spectrometry system for identification of mycobacterium and nocardia species. J Clin Microbiol, 2018, 56(6): pii: e00237-18.
5. 中國防癆協會. 結核病診斷細菌學檢驗規程. 中國防癆雜志, 1996, 18(2): 80-85.
6. 李道群, 夏雪山, 宋玉竹, 等. 結核分枝桿菌分子分型技術的研究和應用. 中國人獸共患病學報, 2016, 32(1): 76-82.
7. Kim BJ, Hong SK, Lee KH, et al. Differential identification of Mycobacterium tuberculosis complex and nontuberculous mycobacteria by duplex PCR assay using the RNA polymerase gene (rpoB). J Clin Microbiol, 2004, 42(3): 1308-1312.
8. Lee AS, Jelfs P, Sintchenko V, et al. Identification of non-tuberculous mycobacteria: utility of the GenoType Mycobacterium CM/AS assay compared with HPLC and 16S rRNA gene sequencing. J Med Microbiol, 2009, 58(Pt 7): 900-904.
9. Petti CA, Bosshard PP, Brandt ME, et al. Interpretive criteria for identification of bacteria and fungi by DNA target sequencing; approved guideline. Clinical and Laboratory Standards Institute (CLSI), 2008.
10. Keys CJ, Dare DJ, Sutton H, et al. Compilation of a MALDI-TOF Mass spectral database for the rapid screening and characterization of bacteria implicated in human infectious diseases. Infect Genet Evol, 2004, 4(3): 221-224.
11. Mediavilla-Gradolph MC, De Toro-Peinado I, Bermúdez-Ruiz MP, et al. Use of MALDI-TOF MS for identification of nontuberculous mycobacterium species isolated from clinical specimens. BioMed Res Int, 2015, 2015: 854078.
12. 周昭彥, 胡必杰, 鮑容, 等. MALDI-TOF質譜快速鑒定非結核分枝桿菌方法的建立和評價. 中華檢驗醫學雜志, 2013, 36(7): 610-614.
13. Huang TS, Lee CC, Tu HZ, et al. Rapid identification of mycobacteria from positive MGIT broths of primary cultures by MALDI-TOF mass spectrometry. PLoS ONE, 2018, 13(2): e0192291.
14. 劉國威, 白大鵬, 梅早仙. 非結核分枝桿菌肺病的診斷與治療研究進展. 中國城鄉企業衛生, 2016, 31(6): 18-21.
15. 劉國標, 羅春明, 鄧虹, 等. 抗結核治療對非結核分枝桿菌肺病的療效觀察. 現代醫院, 2017, 17(3): 354-356.
16. Vijay GS Kumar, Tejashree A Urs, Rajani R Ranganath. MPT 64 antigen detection for rapid confirmation of M.tuberculosis isolates. BMC Res Notes, 2011, Vol.4(1).
17. ?aml? A, ?lki A. Comparison of MALDI-TOF MS, nucleic acid hybridization and the MPT64 immunochromatographic test for the identificationof M. Tuberculosis and non-tuberculosis Mycobacterium species. New Microbiol, 2016, 4(39): 259-263.
18. 王雷, 施旭東. 水溶性對硝基苯甲酸在結核分枝桿菌菌種鑒定中的應用. 臨床醫學工程, 2010, 17(11): 47-48.
19. 蔡靜, 張鑫, 魏黛玉, 等. 甘肅省53 株非結核分枝桿菌對9 種常用抗結核藥物的敏感性分析. 疾病監測, 2018, 33(6): 515-519.
20. Carbonnelle E, Mesquita C, Bille E, et al. MALDI-TOF mass spectrometry tools for bacterial identification in clinical microbiology laboratory. Clin Biochem, 2010, 44(1): 104-109.
21. Akyar I, ?avu?o?lu C, Aya? M, et al. Evaluation of the performance of MALDI-TOF MS and DNA sequence analysis in the identification of mycobacteria species. Turk J Med Sci, 2018, 48(6): 1351-1357.
22. van Veen SQ, Claas EC, Kuijper EJ, et al. High-throughput identification of bacteria and yeast by matrix-assisted laser desorption ionization-time of flight mass spectrometry in conventional medical microbiology laboratories. J Clin Microbiol, 2010, 48(3): 900-907.
23. 王勝芬, 趙雁林, 黃海榮, 等. 結核分枝桿菌北京基因型菌株與耐藥表型的關系. 中國醫學科學院學報, 2009, 31(4): 427-431.
24. 劉棟賓, 向延根, 馬小華, 等. 結核分枝桿菌L型特征及臨床意義研究進展. 實用預防醫學, 2018, 25(9): 1150-1153, 封3.
25. Genc GE, Demir M, Yaman G, et al. Evaluation of MALDI-TOF MS for identification of nontuberculous mycobacteria isolated from clinical specimens in mycobacteria growth indicator tube medium. New Microbiol, 2018, 41(3): 214-219.

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Application of matrix assisted laser desorption ionization time of flight mass spectrometry in the identification of Mycobacterium tuberculosis

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