Detection of Staphylococcus aureus in milk by use of polymerase chain reaction analysis.
OBJECTIVE
To evaluate polymerase chain reaction (PCR) analysis for detection of Staphylococcus aureus (nuc gene) in fresh and formalin-preserved milk.


METHODS
Samples from 80 lactating sheep and 100 lactating dairy cows.


METHODS
4 lactating sheep were inoculated with S aureus by intramammary infusion. A set of primers specific for the nuc gene of S aureus was used to develop a PCR technique, and modification of the rapid boil method was used to isolate bacterial DNA. Milk was obtained from experimentally infected sheep before and after infusion with S aureus, and from the 100 cows and remaining 76 sheep. Samples were screened by bacteriologic culture and PCR. To validate the PCR assay, S aureus or other pathogens were added to distilled water and "normal" sheep milk samples, with and without formalin.


RESULTS
The PCR assay was 100% specific for S aureus when known negative and positive samples were tested. Sensitivity was 100% for samples with added S aureus or other pathogens. Sensitivity was lower for samples obtained from experimentally infected sheep, but increased from 53% to 90% with increased washing of target DNA.


CONCLUSIONS
The PCR technique based on the nuc gene is able to detect S aureus in sheep milk yields results faster than does traditional culturing, is highly specific, and is able to detect S aureus in formalin-fixed milk samples.


CONCLUSIONS
The assay is particularly suitable for analysis of samples shipped or stored without refrigeration. Although antibiotics in milk may inhibit growth in culture, they should not affect the results of the PCR assay.
Khan
M A
MA
Department of Veterinary Clinical Medicine, College of Veterinary Medicine, University of Illinois, Champaign-Urbana 61801, USA.
Kim
C H
CH
Kakoma
I
I
Morin
E
E
Hansen
R D
RD
Hurley
W L
WL
Tripathy
D N
DN
Baek
B K
BK
D000818:Animals; D002417:Cattle; D004269:DNA, Bacterial / Q000032:analysis; D005260:Female; D005511:Food Handling / Q000379:methods; D007774:Lactation; D008892:Milk / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D012756:Sheep; D013211:Staphylococcus aureus / Q000302:isolation & purification*
D004269:DNA, Bacterial