雞肉檢體中沙門氏桿菌快速檢驗方法之探討   全文下載

Rapid Detection of Salmonella spp. in Chicken Samples

本研究主要是探討聚合?鏈鎖反應（PCR）應用在沙門氏桿菌檢驗之可行性，首先以沙門氏桿菌純菌菌株，同時使用兩組primers: 337-L/338-R及TS11/TS4進行multiplex PCR最適條件之探討，結果PCR之最適thermal program為94℃ 1 min，60℃ 2 min，72℃ 2 min, 35 cycles；同時以此條件檢驗沙門氏桿菌標準菌株53株、食品檢體分離菌株32株及非沙門氏桿菌菌株14株，其準確率為100%；以序列稀釋之菌液測試此PCR條件之靈敏度，得知此條件最低可偵測10^4 CFU。在探討檢驗雞肉檢體之條件：將沙門氏桿菌菌液添加於未含此菌之雞肉等檢體中，經離心過濾後進行PCR，可偵測出10^4 CFU，共需4小時。此外，未添加菌液之市售雞肉等檢體，經Lactose broth增菌24小時後，進行離心過濾及PCR，前後共需28小時可偵測出雞肉檢體中之沙門氏桿菌，並經CNS檢驗方法確認結果相同，顯示本研究所建立之PCR方法可應用於雞肉檢體中沙門氏桿菌之檢驗。"

The feasibility of detecting Salmonella spp. using polymerase chain reaction (PCR) was studied. Two sets of primers: 337-L/338-R and TS11/TS4 were used together with pure strains of Salmonella spp. to find the optimum thermal conditions of multiplex PCR. The results showed that the optimum thermal program was 94℃, 1 min; 60℃, 2 min; 72℃, 2 min; 35 cycles. Under the same conditions, 53 standard strains, 32 isolated strains from foods and 14 non Salmonella strains were tested; the accuracy obtained was 100%. Series diluted cultures were used to test the sensitivity of PCR. The results showed that both sets of primers could detect 10^4 CFU of Salmonella spp. Spiked samples were used to find the optimum conditions for detecting Salmonella spp. directly in chicken samples. After ultrafiltration, 10^4 CFU of Salmonella spp. could be detected. It took four hours to complete PCR. Non-spiked samples were incubated in 24 hours with Lactose broth and ultra-filtrated. In this case, 28 hours were totally needed to obtain results. The samples were also checked using the Chinese National Standard (CNS) method. The results showed that the PCR method was applicable to the detection of Salmonella spp. directly in chicken samples.