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Detection of Shigella spp. in food with a nested PCR method – sensitivity and performance compared with a conventional culture method
Author(s) -
Lindqvist R.
Publication year - 1999
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1046/j.1365-2672.1999.00777.x
Subject(s) - shigella , nested polymerase chain reaction , sensitivity (control systems) , biology , microbiology and biotechnology , polymerase chain reaction , genetics , bacteria , gene , salmonella , engineering , electronic engineering
A nested PCR method was developed and its performance evaluated by detection of Shigella flexneri in food. The nested PCR amplifies sequences within an invasion‐associated locus ( ial ) on the invasion plasmid specific for Shigella and enteroinvasive Escherichia coli (EIEC). The nested PCR detected Sh. flexneri in lettuce inoculated with 2, 20 and 200 cfu g −1 after 1, 7 and 18 d of storage, respectively. In comparison, a culture method (NMKL no. 151) detected 10 5 cfu g −1 after 1 but not after 7 d of storage. The presence of inhibitors in blue cheese and shrimps reduced the sensitivity of the PCR assay. To overcome this inhibition, a sample preparation step based on buoyant density centrifugation was developed. This treatment resulted in a successful recovery of Sh. flexneri in lettuce, milk, shrimp and blue cheese inoculated with 10 cfu g −1 . The proposed method, which includes a combination of enrichment, buoyant density centrifugation and a nested PCR, can be completed in less than two working days.