Investigation of different methods in detecting Salmonella from porltry samples

• Main Authors: Cheng-Wen Wang, 王建文
• Other Authors: Ter-Hsin Chen
• Format: Others
• Language: zh-TW
• Published: 2014
• Online Access: http://ndltd.ncl.edu.tw/handle/40276817760224983356

碩士 === 國立中興大學 === 獸醫病理生物學研究所 === 102 === Salmonella spp. is one of the major foodborne pathogens in the world. When Salmonella infected poultry, it not only occurred the clinical symptoms, but also transmit to the product of poultry. The traditional culture method for detecting the samples with Salmonella which spend at least 5 days. It costs much time and the process is complex. Thus, the aim of my study is to find the best method in the field included the characters of rapid detection, high specificity and easy to operation. The samples are all take from the fields, and it will be tested by ISO6579:2002 to make sure samples without Salmonella spp. Then inoculate 4 standard Salmonella strains: Salmonella Enteritidis, Salmonella Typhimurium (Motile strains; MT), Salmonella Pullorum and Salmonella Gallinarum (Non-motile strains; NMT) to each sample. Using the traditional culture method as the gold standard, PCR, LAMP and Kit will be calculated the accuracy (AC), sensitivity (SE), specificity (SP), positive predictive value (PPV) and negative predictive value (NPV). Finally, calculate the kappa coefficient to compare the consistency of each method. The results show that when PCR detected MT, AC, SP, PPV and NPV has a significant for LAMP and Kit (p<0.05). Otherwise, when PCR detect the fecal sample with NMT, it’s unable to show positive of samples. Though the traditional culture method detected the NMT would decreases the sensitivity, LAMP and Kit don’t be effect by microorganism in fecal samples. It still shows high specificity and sensitivity for detection the nucleic acid of Salmonella. But using the boiling method to extract the nucleic acid in egg yolk, LAMP and Kit can’t show positive expectedly. It was inhibited by some material in egg yolk. The kappa coefficient at the part of MT, PCR/traditional culture method is 0.86, shows almost perfect coincide. In the part of NMT is 0.53, shows moderate coincide. LAMP and Kit also show almost perfect coincide of each other for kappa coefficient in MT and NMT. In summary, PCR is more consistency with traditional culture method than LAMP and Kit, but it costs more time than these two methods. When LAMP and kit detect the sample of egg yolk, it needs another way to extract the nucleic acid of Salmonella. These two methods that detect the fecal samples of NMT are better than PCR, because it would not be inhibited by the microorganism in fecal sample. In conclusion, PCR is suitable to monitor and detect the MT samples besides fecal of the field. LAMP and Kit is suitable to monitor and detect NMT samples in fecal sample of the field.