The Detection Of Salmonella In Eggs And The Comparative Study Of Pulsed-field Gel Electrophoresis Typing

Salmonella is an intestinal bacterium with a wild bacteria type range and a very complex antigenic structure. As wildely distributed, it is usually intaken by food ingestion, especially the eggs without completely cooked. As a result, the human body produces a self-limited diarrhea and leads to a systemic infection. As an important dietary raw material, there is a great deal of microbial contamination risks in the process of the storage, transportation and sales, especially Salmonella contamination risks. In recent years, an endless stream of reports about salmonellosis, as a serious threat to human health and bring huge economic losses to the country, has aroused widespread concern. So as a wide range of Salmonella, recently there have been some new to be found. Therefore, the main problem of Salmonella research is to establish a rapid and accurate detection method, and to classify the pathogen to prevent the introduction and spread of Salmonella.The study included the following three parts:The first part:This study discussed the microbial contamination of the commercially available fresh eggs of jiufeng chicken farm in 37℃, room temperature of 20℃ and refrigerated at 4℃, both eggshell surface and the contents. The result show that, on the surface of the shell, the colony total number reached 1.65 ×105 cfu in 4 d under the condition of 37℃ and reached 1.32 ×105 cfu in 4 d under the condition of 20℃. The colony total number reached 1.18×105 cfu in 21 d at 4℃. The total number of eggshell surface colony increased with the time. While inside the egg, and the colony total number reached 7.82×105 cfu/mL in 19 d at 37℃. Under the condition of 20℃, the colony total number reached 1.25×105 cfu/mL in 34 d. It spoiled slowly under 4℃ refrigerated conditio,and the colony total number reached 2.18×105 cfu/mL in 46 d. The total number of egg content colony increased with the time.There were 9 strains of Salmonella detected in 126 eggs, and the detection rate was 7.14%. There were 7 Salmonella enteritidis strains and 2 Salmonella typhimurium strains among them.The second part:The study then used the 9 Salmonella strians which was isolated from eggs and saved in the laboratoris, and designed the PCR primers according to the Salmonella pathogenicity island. Then discussed and optimized the conditions of the Salmonella PCR detection. In this study, we finally selected the primer SPI-3, as it was good in specificity. In the detection of 72 Salmonella strains,68 strains were positive, and the positive rate was 94.4%. The detectability of the PCR primer SPI-3 was 103 cfu/mL.The third part:The study then used the Salmonella strains to do PFGE typing. The result showed that the 66 Salmonella strains were cut into different size of 12～18 fragments by the Xbal restriction enzyme,40% in similarity. According to PFGE electrophoresis, in accordance with the above genetic similarity 90% is a PFGE type, the PFGE electrophoresis of 66 Salmonella strains were divided into 18 types. Through the comparison of Salmonella serotypes of PFGE map results, they had a relatively similar. There were 4 PFGE groups with the same typing of serotype. Exept Salmonella paratyphoid, all the PFGE group were the same as the serotype group.