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J Microbiol Methods. 2014 Jul;102:32-6. doi: 10.1016/j.mimet.2014.04.009. Epub 2014 Apr 26.

Single-nucleotide polymorphism PCR for the detection of Mycoplasma pneumoniae and determination of macrolide resistance in respiratory samples.

Author information

1
Department of Laboratory Medicine, Asan Medical Center and University of Ulsan College of Medicine, Seoul, Republic of Korea.
2
SolGent Co., Ltd., Daejeon, Republic of Korea.
3
Department of Pediatrics, Seoul National University Hospital, Seoul, Republic of Korea.
4
Department of Laboratory Medicine, Sanggye Paik Hospital, Inje University College of Medicine, Seoul, Republic of Korea.
5
Department of Pediatrics, Sanggye Paik Hospital, Inje University College of Medicine, Seoul, Republic of Korea.
6
Department of Laboratory Medicine and Hematologic Malignancy Branch, Research Institute and Hospital, National Cancer Center, Goyang-si, Republic of Korea.
7
Department of Infectious Diseases, Asan Medical Center and University of Ulsan College of Medicine, Seoul, Republic of Korea.
8
Department of Laboratory Medicine, Asan Medical Center and University of Ulsan College of Medicine, Seoul, Republic of Korea. Electronic address: sung@amc.seoul.kr.

Abstract

The aim of this study was to develop a single-nucleotide polymorphism (SNP) PCR assay to be performed directly on respiratory samples for the simultaneous detection of Mycoplasma pneumoniae and its 23S rRNA gene mutations, which are responsible for macrolide resistance. For multiplex SNP PCR, two outer primers for amplification of the 23S rRNA gene and two mutant-specific primers for the discrimination of single base changes were designed. A total of 73M. pneumoniae-positive samples and 100M. pneumoniae-negative samples were analyzed using this assay. By SNP PCR, we detected two mutations conferring high-level macrolide resistance in 22 samples (A2063G from 20 and A2064G from 2 samples); these results are identical to those produced by the 23S rRNA gene sequencing of M. pneumoniae-positive samples. Thus, this assay can be used as a practical method for the simultaneous detection of M. pneumoniae and mutations associated with macrolide resistance directly from respiratory samples.

KEYWORDS:

23S rRNA gene; Macrolide resistance; Mutation; Mycoplasma pneumoniae; Single-nucleotide polymorphism PCR

PMID:
24780151
DOI:
10.1016/j.mimet.2014.04.009
[Indexed for MEDLINE]

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