Targeting the rpoB Gene Using Nested PCR-Restriction Fragment Length Polymorphism for Identification of Nontuberculous Mycobacteria in Hospital Tap Water

Ji-Hyun Shin; Hae-Kyung Lee; Eun-Jin Cho; Jae-Yon Yu; Yeon-Ho Kang

doi:10.1007/s12275-008-0102-6

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Targeting the rpoB Gene Using Nested PCR-Restriction Fragment Length Polymorphism for Identification of Nontuberculous Mycobacteria in Hospital Tap Water: Ji-Hyun Shin , Hae-Kyung Lee , Eun-Jin Cho , Jae-Yon Yu , Yeon-Ho Kang; Journal of Microbiology 2008;46(6):608-614.
DOI: https://doi.org/10.1007/s12275-008-0102-6
Published online: December 24, 2008

Division of Bacterial Respiratory Infection, Center for Infectious Diseases, National Institute of Health, Centers for Disease Control and Prevention, Seoul 122-701, Republic of KoreaDivision of Bacterial Respiratory Infection, Center for Infectious Diseases, National Institute of Health, Centers for Disease Control and Prevention, Seoul 122-701, Republic of Korea

Corresponding author: Yeon-Ho Kang , Tel: 82-2-380-2140,

Received: 23 April 2008 • Accepted: 2 September 2008

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Abstract

Nontuberculous mycobacteria (NTM) are ubiquitous in the environment and can cause nosocomial infections in immunocompromised patients. Recently the presence of NTM in public drinking water and hospital water distribution systems has been reported. Their ability to form biofilms and their resistance to chlorine both contribute to their survival and colonization in water distribution systems. Here we analyzed thirty-two hospital tap water samples that were collected from different locations in three hospitals so as to evaluate the prevalence of NTM species. The water samples were concentrated by membrane filtration and then eluted with sterilized water following sonication. Two-step direct PCR targeting the rpoB gene, restriction fragment length polymorphism (RFLP) using the MspI restriction enzyme, and sequence analysis were performed for identification of NTM to the species level. The sequences of each PCR product were analyzed using BLASTN. Seven samples (7/32, 21.9%) were positive for NTM as determined by nested-PCR. The PCR-RFLP results indicated five different patterns among the seven positive PCR samples. The waterborn NTM were identified, including M. peregrinum, M. chelonae (2 cases), M. abscessus, M. gordonae (2 cases), and Mycobacterium sp. JLS. The direct two-step PCR-RFLP method targeting the rpoB gene was effective for the detection and the differentiation of NTM species from hospital tap water.

Keywords: nontuberculous mycobacteria;hospital tap water;rpoB gene;PCR-RFLP

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Targeting the rpoB Gene Using Nested PCR-Restriction Fragment Length Polymorphism for Identification of Nontuberculous Mycobacteria in Hospital Tap Water

J. Microbiol. 2008;46(6):608-614. Published online December 24, 2008

DOI: https://doi.org/10.1007/s12275-008-0102-6

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