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Arch Microbiol. 2016 Aug;198(6):591-4. doi: 10.1007/s00203-016-1228-2. Epub 2016 Apr 25.

Targeting single-nucleotide polymorphisms in the 16S rRNA gene to detect and differentiate Legionella pneumophila and non-Legionella pneumophila species.

Zhan XY1,2,3, Hu CH1,2, Zhu QY4,5.

Author information

1
Guangzhou KingMed Center for Clinical Laboratory, No.10, Luoxuan 3 Road, Guangzhou International Bio-island, Guangzhou, 510300, China.
2
KingMed School of Laboratory Medicine, Guangzhou Medical University, Guangzhou, 510300, China.
3
The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, 510080, China.
4
Guangzhou KingMed Center for Clinical Laboratory, No.10, Luoxuan 3 Road, Guangzhou International Bio-island, Guangzhou, 510300, China. zqy@kingmed.com.cn.
5
KingMed School of Laboratory Medicine, Guangzhou Medical University, Guangzhou, 510300, China. zqy@kingmed.com.cn.

Abstract

A PCR-based method targeting single-nucleotide polymorphisms (SNPs) in the 16S rRNA gene was developed for differential identification of Legionella pneumophila and non-Legionella pneumophila. Based on the bioinformatics analysis for 176 Legionella 16S rRNA gene fragments of 56 different Legionella species, a set of SNPs, A(628)C(629) was found to be highly specific to L. pneumophila strains. A multiplex assay was designed that was able to distinguish sites with limited sequence heterogeneity between L. pneumophila and non-L. pneumophila in the targeted 16S rRNA gene. The assay amplified a 261-bp amplicon for Legionella spp. and a set of 203- and 97-bp amplicons only specific to L. pneumophila species. Among 49 ATCC strains and 284 Legionella isolates from environmental water and clinical samples, 100 % of L. pneumophila and non-L. pneumophila strains were correctly identified and differentiated by this assay. The assay presents a more rapid, sensitive and alternative method to the currently available PCR-sequencing detection and differentiation method.

KEYWORDS:

16S rRNA; Detection; Differentiation; Legionella; Legionella pneumophila; Single-nucleotide polymorphisms

PMID:
27112927
DOI:
10.1007/s00203-016-1228-2
[Indexed for MEDLINE]

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