| Streptococcus suis (SS) is a kind of common pathogens of swine, It’s also a zoonotic disease caused by a variety of pathogenic Streptococcus suis infection. The pathogen can cause meningitis, endocarditis, arthritis, pneumonia, septicemia and sudden death, and even infect humans and cause death. According to the cell surface properties of Streptococcus suis capsular polysaccharide. SS is divided into 35 serotypes (typel-34 and type 1/2). Among them, serotype 2 pathogenicity is the strongest and the most widely distributed, serotype 1,7 and 9 followed. Firstly, for three important virulence factors CPS2J, MRP and EF of Streptococcus suis serotype 2, the study established a triple-fluorescent quantitative PCR method. Then, in that of clinical specimens collected easily contaminated and are not easily separated from the typical colonies. The study had selected a selective medium for Streptococcus suis, which can improve the clinical efficiency of the separation and reduce the detection time. On the basis of the two experiments above, the study have investigated the situation of apparently healthy pigs carrying Streptococcus suis in parts of Shanghai.With the software Primer Express 3.0, three pairs of specific primers and fluorogenic-labedled probes were designed and synthesized in accordance with the target genes CPS2J, MRP and EF. The 5’-end of CPS2J, MRP, EF probes was labeled with reporter FAM, HEX and CY5, the 3’-end was all labeled with quencher BHQ1. By the reaction system and optimization of amplification conditions, the study set up a detection method for rapid detection of Streptococcus suis serotype 2 with the taqman probes and evaluated its sensitivity, specificity and reproducibility. It was found that the reaction system had good amplification efficiency, CPS2J, MRP and EF gene detection sensitivity was the lowest detected 90 copics/μL,40 copies/μL and 60 copies/μL. The specific test had no cross-reactivity with other pathogens. Variation coefficient in repetitive test is less than 3%. Amplification of the entire detection within 60 min to complete, the advantages of this method are fast, good sensitivity, reproducibility and specificity.The experiment also screened out a selective medium containing a combination of antimicrobial drugs for Streptococcus suis. Firstly, a preliminary selected tested on susceptibility papers, referenceed these drugs antimicrobial spectrum, chose four drugs which are not sensitive to the Streptococcus suis. Use three Streptococcus suis stains and seven clinical common bacteria contamination, measured the four drugs’s minimum inhibitory concentration. Based on the analysis of results above, I selected polymyxin B, aztreonam and furazolidone to combine into antibiotics. Their concentrations were 16 μg/mL,64 μg/mL, and 32μg/mL. The selective medium inhibitory rate closed to 100% for clinical contaminants which usually carried E. coli, Enterococcus faecalis, Staphylococcus aureus and Salmonella choleraesuis. The simulation results can be seen the number of contaminated samples isolated typical colonies significantly increased, so as to establish a method for clinical samples Streptococcus suis selective enrichment, greatly reducing the detection rate of Streptococcus suis.On the basis of these two parts of the experiment, the study also investigated the carrier case of Streptococcus suis in clinical healthy pigs, in some areas of Shanghai. From between December 2013 to April 2014, collect mandibular lymph nodesin two pig slaughterhouse in Shanghai Fengxian, after the selective enrichment broth cultured, used conventional PCR methods to test Streptococcus suis GDH genes and species characteristics CPS1J, CPS2J, CPS7J and CPS9J. From the 122 samples, the overall detection rate of Streptococcus suis was 27%(33/122), In the detection of 33 samples of S. suis,serotype 1 was 3.03%(1/33), Serotype 2 was 18.2%(6/33), serotype 7 was 6.06%(6/33) and the serotype 9 was 18.2%(6/33). Thus, the apparent healthy pigs carried Streptococcus suis widely, attention should be paid. |
PDF Full Text Request