Molecular Typing Of Clostridium Difficile Surveyed From Neonatal Pigs In Sichuan District

Clostridium difficile is a Gram-positive,strictly anaerobic,spore-forming bacterium,and widely spread in worldwide.In piglets,it can cause neonatal enteritis,and it can lead to diarrhea piglets death.This study aims to investigate the molecular typing of C.difficile in Sichuan district.Firstly,it is proved that the evidence of diarrhea piglets were infected through C.difficile in Sichuan district.In this study,a total of 110 fresh stool samples from piglets 10 days old or less were tested for CD using swabs.The survey covered 6 pig farms(including intensive swine enterprises and individual farms)in Sichuan district.The farms were distributed over 6 counties(6 cities)of Sichuan district.The swabs were immediately placed in ice boxes after sampling,and rapidly transported to laboratory for isolation.A total of 110 fecal samples were collected from different pig farms and 20 C.difficile wild strains were isolated successfully.The isolate rate of C.difficile was approximately 18.18 %(20/110).5 of 6 pig farms tested positive for C.difficile(83.3 %).Secondly,it was found that there were 3 kinds of toxin genotypes in 20 C.difficile wild strains.A+B+CDT+(3): The 2 isolates C1 and C2 from Chongzhou City of Chendu City were of A+B+CDT+ type,and one isolate P1 from Pengshan District of Meishan City was of A+B+CDT+ type.A+B+CDT-(6): The 4 wild strains B3,B4,B5 and B19 from Linshui County of Guang'an City are all A+B+CDT-type,and the two wild strains LW3 and LW6 from Langzhong City of Nanchong City were all A+B+CDT-type.A-B-CDT-(11): The 10 wild strains Y1-Y10 from Jiang'an County of Yibin City are all A-B-CDT-type,and one isolate P2 from Pengshan District of Meishan City was of A-B-CDT-type.The proportion of toxigenic strains to total isolates is 45 %(9/20).The proportion of hypervirulent strains to total isolates is 15 %(3/20).The proportion of hypervirulent strains to toxigenic isolates is 33.3 %(3/9).Finally,it was found that there were 5 kinds of multilocus sequence types in 20 C.difficile wild strains.Within the isolate population of 20,1,3,2,4,and 10 were found to belong to multilocus sequence types ST3,-11,-35,-36,and-109,respectively.Details are as follows:A isolate P2 from Pengshan District of Meishan City was of ST3 type.The 2 isolates C1 and C2 from Chongzhou City of Chendu City were of ST11 type,and one isolate P1 fromPengshan District of Meishan City was of ST11 type.The two wild strains LW3 and LW6 from Langzhong City of Nanchong City were all ST35 type.The 4 wild strains B3,B4,B5 and B19 from Linshui County of Guang'an City are all ST36 type.The 10 wild strains Y1-Y10 from Jiang'an County of Yibin City are all ST109 type.We found ST35,-36 correspond to A+B+CDT-,ST11 corresponds to A+B+CDT+,and ST3,-109 correspond to A-B-CDT-.This study showed that the C.difficile subpopulation in Sichuan to be genetically highly variable.