Rapid detection of Salmonella in milk by combined immunomagnetic separation-polymerase chain reaction assay.
During the past few years, milk has presented a risk of Salmonella contamination; it has been implicated as the cause in several outbreaks of salmonellosis. Because conventional detection methods require 5 to 7 d for completion and involve several subcultivation stages followed by biochemical and serological tests, rapid and sensitive methods have been sought, mainly at the DNA level. Therefore, a study including milk samples was conducted to evaluate the performance of a combination of 2 techniques--immunomagnetic separation and polymerase chain reaction (PCR)--for the detection of Salmonella. The 16-, 14-, 12-, 10-, and 8-h nonselective pre-enrichment steps before immunomagnetic separation and the high-pure DNA preparation method before PCR were used in a combined assay. Milk samples, which were found to be Salmonella-negative by a reference method, were first inoculated with Salmonella Enteritidis. Next, the shortest pre-enrichment time that is required for detection of 1 or 10 cfu of Salmonella/mL by combined immunomagnetic separation-PCR assay was found by using 16-, 14-, 12-, 10-, and 8-h incubation periods. The detection limit using a 16-, 14-, or 12-h nonselective pre-enrichment was 1 to 10 cfu/mL. However, the sensitivity decreased to 10(1) and 10(2) cfu/mL, respectively, when 10- and 8-h pre-enrichments were used. This assay, in conjunction with a 12-h pre-enrichment, proved to be rapid (overall 16 h) and sensitive (1-10 cfu/mL) for the detection of Salmonella in milk samples and promising for routine use in the detection of Salmonella in milk.
Mercanoglu Taban
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Gazi University, Vocational School of Health Services, Golbasi, Ankara, Turkey. birce@gazi.edu.tr
Ben
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Aytac
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D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D005516:Food Microbiology; D005524:Food Technology / Q000379:methods*; D018189:Immunomagnetic Separation / Q000379:methods*; D008892:Milk / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D019779:Salmonella enterica / Q000302:isolation & purification*
D001426:Bacterial Proteins; C075518:invA protein, Bacteria