The Detection Of Staphylococcus Aureus In Food Using PCR Method

Food-borne diseases are mainly food security issues in our country which are caused by invasive organisms such as salmonella, staphylococcus aureus (SA) and Vibrio parahaemolyticus, the three common pathogenic bacteria. 50% of the causes-known food poisonings are caused by staphyloentero-toxin, which led the first place in bacterial food poisonings. The symptoms appears 1~6 hours after eating staphyloentero-toxin, which manifested as nausea, vomiting, upper abdominal pain and diarrhea similar to acute gastroenteritis and vomiting is most dominant. Staphylococcus aureus widely exists in the nature, human skin and mucous membrane; therefore it has a bigger chance to contaminate food. Common contamination source comes from Foodstuff producer and carriers without symptoms. Animal foodstuffs, milk and dairy products are easy to be contaminated to cause food poisoning. At present the detection of SA are still using conventional methods such as isolation, culture, biochemistry identification which takes a 4 to 7 days to report with a low sensitivity and specificity as well as tedious operations. It is reported that polymerase chain reaction (PCR) has been established to test SA in food at abroad; however no such reports have been found at home. We designed specific primers according to pathopoiesis of SA to establish a PCR method to quickly detect SA in food. It aimed to improve the sensitivity, shorten the detection time and provide a technique support for quick detection of food-borne diseases caused by SA. We tested 180 food samples from 9 areas in Jilin province by using national standard approach (GB4789.10-2003) and PCR method respectively. 45 samples were tested positively by using PCR method, counting for 25% of the whole tested samples, and the positive rates of fish and cooked meat ranked top, accounting for 40% respectively, pouched milk were all found negative. 42 samples were detected positively by national standard approach, accounting for 23.3%, among which unpacked milk were found with the highest positive rate 37%, while pouched milk and vegetables were detected negative. There is no significant difference between the positive frequencies tested by national standard approach and those tested by PCR method(?2=3.0,Pï¹¥0.05). The establishment of PCR method in current study depended on the primers designed by the thermostable nuclease nuc gene which is strictly conserved and unique to SA. And the target segment to be amplified was 422bp. When PCR method was established, the reaction conditions were optimized and the tests of sensitivity and specificity were also conducted. The results showed that the detection limit was 4.32g DNA of SA, which suggested a better sensitivity. Strains of Cerea bacillus and salmonella were used to test the specificity of PCR method. Target gene segments were found in the amplification products of positive control strains, while no target segments were found in the amplified products of other strains. This suggested a good specificity. And PCR could be done in 24 hours. Therefore, the PCR method established according to nuc gene had a better sensitivity and specificity, and it was quicker and more convenient than national standard approach. Therefore, it is more eligible to quick detection of food.The present study showed that the PCR amplification of unc gene may detect SA specifically without cross-reactions with other bacteria strains. PCR method to detect SA in food has a better specificity and sensitivity and greatly shortens the detection periodicity, which is superior to the conventional approach.