Isolation And Identification Of Corynebacterium Pseudotuberculus From Guanzhong Dairy Goats And Detection Of Virulence Genes And Drug Resistance

In recent years,with the development of large-scale and intensive sheep farming,dairy goat breeding industry has developed rapidly in our country.Ovine pseudotuberculosis is caused by Corynebacterium pseudotuberculosis,which is found in almost all major sheep and goat production areas in the world.It has been reported in Inner Mongolia,Shaanxi,Gansu,Xinjiang,Yunnan,Shanghai,Zhejiang,Guangxi and Guangdong.Due to the chronic and subclinical nature of Corynebacterium pseudotuberculosis infection,it has brought significant economic losses to the goat industry.Fuping is an important breeding base in China.Guanzhong dairy goats farming has become an indispensable economic pillar industry for Fuping County's rural areas.In recent years,the ovine pseudotuberculosis in the flock of Fuping dairy goats has become widespread,which has seriously affected the healthy production of the goat breeding industry.In this study,the pus samples of ovine pseudotuberculosis suspected goats were investigated and collected in farms in different areas of Fuping,Shaanxi.The pathogens were isolated and identified by bacterial isolation,purification,Gram staining,biochemical experiments,and PCR identification.The virulence gene was detected by PCR method;The K-B method was used to analyze the resistance of pathogenic bacteria,and the drug resistance gene was further detected by PCR method.The results obtained are as follows:1.A total of fourteen suspected strains of Corynebacterium pseudotuberculosis were isolated from twenty-three pus samples of sick dairy goats collected in Fuping,Shaanxi.The isolated strain was white on agar medium containing 5% sheep blood,the colony edge was neat,easy to be pushed,dry and brittle.Gram staining is positive,microscopic examination is mostly short rod-shaped or globular,no spores,no capsule,difficult move,They are mostly scattered or fenced,and arranged in a "V" shape.The colony morphology and staining microscopic features of the isolates were similar with the pseudotuberculosis coryneform bacteria.Molecular biological test showed that the fourteen strains were more than 99% homologous to the 16 S rRNA nucleotide of the C.p reference strain in GenBank.2.The PCR analysis showed that virulence genes of isolated Corynebacterium pseudotuberculosis carried five major virulence factors,including PLD,Fag A,Fag B,Fag C,Fag D.3.The results of drug susceptibility test showed that the isolated strain was sensitive tovarious antibiotics such as Tetracycline,Penicillin,Clarithromycin,Rifampicin and Cefazolin,and resistant to Amikacin and Streptomycin.The drug resistance gene detection found that86%(12/14)of the isolated strains carry drug resistance genes that mediate aminoglycosides,and 100%(14/14)of the isolated strains carry drug resistance genes that mediates ?-lactam drugs.The resistance gene of rifampicin was not detected.