應用特異性聚合酶鏈反應引子快速鑑定與雞相關樣品之腸炎沙門氏菌   全文下載

Specific PCR Primers for the Identification of Salmonella enterica Serovar Enteritidis in Chicken-Related Samples

本研究根據腸炎沙門氏菌（Salmonella enterica serovar Enteritidis）之sefb 基因，發展特異性聚合?鏈反應（PCR）引子SefB127L-SefB661R，快速鑑定與雞相關樣品中之腸炎沙門氏菌，此PCR 引子首先應用於85 株沙門氏菌及17 株非沙門氏菌檢測，結果顯示只有24 株腸炎沙門氏菌得到535 bp大小之PCR 產物。接著應用於40 個接入腸炎沙門氏菌之樣品，經過預培養，此PCR 引子對腸炎沙門氏菌其靈敏度為101 cells/g。進一步檢測285 種天然樣品，包括雞肉、雞蛋、雞舍、雞身及雞糞等之腸炎沙門氏菌。由結果顯示，此PCR 與傳統方法（BAM）皆於雞蛋及糞便中檢出腸炎沙門氏菌，其檢測率為1%。

In this study, a designed pair of PCR primers, SefB127L-SefB661R, based on the sefb gene (accession number L11009) sequences was used in polymerase chain reaction (PCR) for rapid evaluation of Salmonella enterica serovar Enteritidis in chickenrelated samples. The specificity of this method was checked with 85 Salmonella strains and 17 non-Salmonella strains. The results showed that only 24 isolates of S. enterica serovar Enteritidis exhibited 535 bp PCR product. The detection limit of this PCR method were evaluated using 40 spiked samples under enrichment protocols. The data revealed that microbial extract from as few as 101 target cells per gram of the sample culture was required for this assay. Before PCR amplification, pre-culture and the cell lysates, rather than the DNA extracts, were used directly for all tested samples. To verify the usefulness of this PCR process for S. enterica serovar Enteritidis examination, 285 endogenous samples including chicken meats, eggs and swabs of chicken-related samples and coop’s facilities were tested and compared with that obtained by conventional BAM (Bacteriological Analytical Manual) method. About 1% (three in 285) of the S. enterica serovar Enteritidis samples was contaminated, approximately the same as that obtained from BAM method.