| In this paper, the study was concentrated on Salmonella isolates from ducks in7regions of Sichuan. And the main research includes:①Salmonella was isolated and identified in serotypes and pathogenicity tests;②PCR method was established and applied to these isolates according to invA gene;③Drug sensitivity was applied;④Resistance genes were detected and analyzed. The results were showed as follows:1. Isolation, biomedical identification, serotyping and pathogenicity testing of Salmonella from ducks:48Salmonella strains were isolated and identified from seven regions in Sichuan, including14strains from Peng zhou,9strains from Mian yang,8strains from Qiong lai,6strains from De yang,5strains from Du jiangyan,3strains from Mei shan and3strains from Xin jing. The results revealed that48Salmonella isolates belong to6serotypes, including Salmonella Typhimurium (37.5%), Duck Salmonella (18.7%), Salmonella Enteritidis (14.6%), Salmonella Typhoidfever (14.6%), Salmonella Ⅱ (8.3%) and Salmonella Dublin (6.3%), which means there are different serotypes in different regions. It also showed that the pathogenicity of all isolates was very high in animal experiment.2. Based on invA gene, PCR for the detection of Salmonella isolates from ducks was developed and applied:a pair of primers (P1/P2) was designed according to the Salmonella invA gene sequence for amplification of Salmonella genome DNA. PCR product of608bp was obtained with these primers for Salmonella genome amplification but no products could be detected with these primers for genome amplification of other pathogens, including ATCC25922E.coli, Staphylococcus Aureus and Shigella; the sensitivity of the primers of P1/P2was5.1×102CFU/mL;48Salmonella-positive samples and2Salmonella-negative samples were detectd by stablished PCR method. And the results showed that the expected PCR fragment (608bp) could be obtained from the48Salmonella-positive samples but no PCR fragment could be obtained from the2 Salmonella-negative samples. After BLAST analysis, the similarity between PCR products and the target sequence was up to97%. These results demonstrated that the established invA PCR detection method was reliable, sensitive and could be used in clinical diagnosis and surveillance of infection.3. Drug susceptibility testing of Salmonella from ducks:drug sensitivity testing of48Salmonella isolates showed that all the isolates were resistant to36commonly used antibiotics. Antibiotic resistance rates of14antibiotics were more.than50%:penicillin G (100%), oxacillin (100%), rifampicin resistance (100%), Ampicillin (93.8%), Tetracycline (83.3%), Erythromycin (79.2%), Sulfamethoxazole compound (72.9%), Imipenem (66.7%), Amoxicillin (66.7%), Doxycycline (62.5%), Nalidixic acid (62.5%), Furazolidone (58.3%), Chloramphenicol (54.2%) and Trimethoprim (50.0%). Antibiotic resistance rates of2antibiotics were less than10%:Cefoperazone (6.3%) and Amikacin (8.3%). Antibiotic resistance rates of the remaining20antibiotics were between10%-50%. Salmonella isolates from Pengzhou were resistant to34antibiotics, while Salmonella isolates from Mian yang, De yang, Qiong lai and Du jiangyan were resistant to31antibiotics and Salmonella isolates from Xin jing were resistant to16antibiotics.All Salmonella strains had the resistant capacity to multi-drug resistant. Specifically,41strains (85.6%) were resistant to more than10kinds of antibiotics,29strains (60.3%) were resistant to more than15kinds of antibiotics,16strains (33.3%) were resistant to more than20kinds of antibiotics and2atrains (2.1%) resistant to more than30kinds of antibiotics; multiple-drug resistance was very severe among these Salmonella isolates, such as18.8%of Salmonella isolates resistance to15-16antimicrobial agents,10.4%of Salmonella isolates resistance to8-10antimicrobial agents,10.4%of Salmonella isolates resistance to11ntimicrobial agents; the study also showed that resistance and multiple-drug resistance of these strains were different in different regions.4. Resistance genes of48Salmonella isolates from ducks were measured by molecular epidemiological study:8paris of resistance gene primers were designed and6resistance genes were detected, including aph(3’)-Ila, aadAl, aadA2, tet(A), dphs and parC, but bla-TEM and tet(G) were not detected; the detection of drug resistance genes and resistant phenotypes were consistent; PCR products of6drug resistance genes were sequenced which showed that the similarity was more than97%. Distribution of resistant genes in7regions was different and it is prevalent that the one salmonella strain from ducks carries a variety of resistance genes. |
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