title: Quantification of Staphylococcus aureus in unpasteurised bovine and caprine milk by real-time PCR.
abstract: A sensitive and reproducible real-time PCR assay targeting the nuc gene of Staphylococcus aureus was applied for quantification of this microorganism in artificially and naturally contaminated raw milk samples. The S. aureus cell equivalents (SCEs) estimated by the real-time PCR method were two log scales higher than colony forming units (CFUs) estimated from a plate count method in artificially contaminated milk. The repeatability of the real-time PCR assay including the DNA isolation procedure was assessed by analysing the data derived from naturally contaminated samples. The relative standard deviation of the log-transformed data of four real-time PCR measurements including duplicate DNA isolations ranged between 11.3 and 1.0%. When analysing 80 bovine and 107 caprine naturally contaminated raw milk samples, the real-time PCR method yielded 19.3% more positive samples than the plate count method. With the exception of one sample, SCEs were always higher than CFUs. The difference between SCEs and CFUs was highly variable, and it was not possible to correlate real-time PCR-derived SCEs and CFUs. However, as each SCE detected by real-time PCR indicates a S. aureus cell, which is or has been present in the sample, this method offers the advantage of a retrospective analysis even of processed samples to aid food poisoning-related risk assessment.
authors.affiliation: Institute for Milk Hygiene, Milk Technology, and Food Science, University for Veterinary Medicine, Veterinärplatz 1, 1210 Vienna, Austria. ingeborg.hein@vu-wien.ac.at
authors.forename: Ingeborg
authors.initials: I
authors.lastname: Hein
authors.forename: Hannah J
authors.initials: HJ
authors.lastname: Jørgensen
authors.forename: Semir
authors.initials: S
authors.lastname: Loncarevic
authors.forename: Martin
authors.initials: M
authors.lastname: Wagner
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D002417:Cattle; D015169:Colony Count, Microbial / Q000379:methods*; D004269:DNA, Bacterial / Q000235:genetics; D004720:Endonucleases / Q000235:genetics; D006041:Goats; D008836:Micrococcal Nuclease / Q000235:genetics; D008892:Milk / Q000382:microbiology*; D009664:Norway; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D013211:Staphylococcus aureus / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; C074002:nuc protein, staphylococcus; D004720:Endonucleases; D008836:Micrococcal Nuclease