Open AccessDevelopment of a 16 S –23 S r RNA intergenic spacer‐based quantitative PCR assay for improved detection and enumeration of L actococcus garvieae
Author(s) -
Thanh Hien Dang,
Park Hee Kuk,
Kim Wonyong,
Shin HyoungShik
Publication year - 2013
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/1574-6968.12038
Subject(s) - lactococcus lactis , biology , lactococcus , microbiology and biotechnology , genomic dna , polymerase chain reaction , 23s ribosomal rna , real time polymerase chain reaction , ribosomal rna , intergenic region , genome , bacteria , dna , genetics , gene , rna , lactic acid , ribosome
L actococcus garvieae is an important foodborne pathogen causing lactococcosis associated with hemorrhagic septicemia in fish worldwide. A real‐time quantitative polymerase chain reaction (q PCR ) protocol targeting the 16 S –23 S r RNA intergenic spacer ( ITS ) region was developed for the detection and enum‐eration of L . garvieae . The specificity was evaluated using genomic DNA s extracted from 66 cocci strains. Fourteen L . garvieae strains tested were positive, whereas 52 other strains including L actococcus lactis ssp. lactis , L actococcus lactis ssp. hordniae and L actococcus lactis ssp. cremoris did not show a specific signal. The minimal limit of detection was 2.63 fg of purified genomic DNA , equivalent to 1 genome of L . garvieae . The optimized protocol was applied for the survey of L . garvieae in naturally contaminated fish samples. Our results suggest that the q PCR protocol using ITS is a sensitive and efficient tool for the rapid detection and enumeration of L . garvieae in fish and fish‐containing foods.