The Establishment Of PCR Method For Detection Of Salmonella Of Pet Turtles

Salmonella is a kind of Gram-negative bacillus and can be infected not only in animals, but also in human, causing diarrhea, abdominal cramp, vomiting, fever, sepsis, gastroenteritis and so on. There have been many serious cases about human reptile-associated salmonellosis in abroad .As more and more turtles are being as pets, some potential public health threats are existed. This study aimed to establish a PCR detection of salmonella in pet turtles.64 samples were collected in the Veterinary Teaching Hospital of China Agricultural University and pet markets from July, 2008 to August, 2009, which were all free of antibiotics. Isolated organisms were identified by biochemical tests recommended by WHO and ISO. Then Antibiotic susceptibility tests were taken by K-B method. The results showed that salmonella were found in 18 turtles with incidence of 28.1%. 5 turtles were privately owned with incidence of 11.3%, the other 13 turtles were from pet markets with incidence as much as 65%. Drug susceptibility test results showed that salmonella had resistance to many commonly used antibiotics, the rate even as much as above 70%. For example, ampicillin (74%), tetracycline (79%), sulfamethoxazole / trimethoprim (70%). The results also showed salmonella was very sensitive to ceftriaxone, amikacin and enrofloxacin cefalotin with rate as much as 99%. So it was considered these drugs can be first choices for clinical treatment.The study first established PCR detection for salmonella in pet turtles. Salmonella had specific invA gene, which were used to amplify the standard bacteria and 7 non-salmonella species by PCR. A 372bp DNA amplified fragment was found in salmonella strains, but none in the non-Salmonella strains. The detection limit of the PCR was 2.4×102cfu/ml with serial dilutions of bacterial culture and PCR products were confirmed as invA genes by sequencing and had high homology compared with the reference sequences, which indicated that the PCR detective method had high specificity and sensitivity. 230 bacterial strains from the 64 samples were tested and 135 bacterial strains were salmonella-positive samples from turtles, the positive rate of reptile-associated salmonellosis was 58.7%. This research has established a simple, sensitive and specific method for the detection of reptile-associated salmonellosis genus.