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J Med Microbiol 55 (2006), 273-277; DOI: 10.1099/jmm.0.46027-0
© 2006 Society for General Microbiology
ISSN 0022-2615

Development of a routine laboratory direct detection system of staphylococcal enterotoxin genes

Akifumi Nakayama1,{dagger}, Akiko Okayama1, Misao Hashida1, Yasuzumi Yamamoto1, Hisakatsu Takebe1, Takashi Ohnaka2, Tomoyuki Tanaka2 and Shunsuke Imai1

1 Nara Prefectural Institute for Hygiene and Environment, 57-6 Ohmori-cho, Nara City, Nara 630-8131, Japan

2 Sakai City Institute of Public Health, 3-2-8 Kaichyo Higashi, Sakai City, Osaka 590-0953, Japan

Correspondence
Akifumi Nakayama
pyonchan{at}asint.jp

Received 28 January 2005
Accepted 21 October 2005

A novel direct detection system has been developed for eight staphylococcal enterotoxin (SE)-encoding genes (sea, seb, sec, sed, see, seg, seh and sei) in milk. Specific detection by real-time PCR was successful for all SE-encoding genes in the reference strains. Furthermore, a novel DNA-preparation method with good reproducibility [coefficients of variation 0·31, 0·99 and 1·21 % at 106, 104 and 102 c.f.u. (ml milk sample)–1, respectively] was developed to overcome PCR inhibition in the milk samples. The combination of this DNA-preparation method and real-time PCR resulted in high sensitivity [between 1·1x102 and 1·0x104 c.f.u. (ml milk sample)–1] and allowed the completion of the entire procedure within 4 h. Results of an evaluation of this method for the detection of SE-encoding genes using known outbreak milk samples produced results showing good correspondence with the reversed passive latex agglutination assay. In addition, this newly developed system can be applied to clinical samples such as faeces and vomit. Consequently, the system should be useful in the routine direct detection of SE-encoding genes in food-borne-poisoning samples.

{dagger}Present address: Clinical Laboratory, Nara Prefectural Hospital, 1-30-1 Hiramatsu, Nara City, Nara 631-0846, Japan.






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