The Effect Of Amikacin Alone And In Combination With Other Three Antibiotics On The Mic Of Streptococcus Separated From Cattle With Mastitis

In veterinary clinic,dairy cow mastitis is the most common disease in large-scale dairy farms,and bovine Streptococcus is one of the major pathogens causing this disease.At present,there is no ideal vaccine for dairy cow mastitis in the world,so the antibiotics are the main measures to prevent and control mastitis.However,due to abuse of antibiotics in recent years,the antibiotic resistance of clinical isolated Streptococcus increased rapidly,which had brought serious challenges to the disease prevention and control.This study collected 63 samples from large-scale dairy farms in some areas of China from2016 to 2017,and 35 strains of mastitis Streptococcus were isolated and obtained by PCR detection of housekeeping gene sodA.The overall separation rate reached 55.56%.Using PCR to detect 7 virulence genes,the results showed that the scpB was the most prevalent genes,accounting for 68.57% of the Streptococcus strains,followed by cyl(48.57%),glnA(31.43%),hylB(22.86%),lmb(8.57%)and bac gene was not detected.The drug susceptibility and resistance of total 35 strains of isolates Streptococcus were determined by KB disk diffusion method.Up to 30 kinds of antibiotic sensitivity and resistance were detected,and using PCR to detect the expression of 10 drug resistance genes.The results showed that strains showed extreme resistance of Sulfamethoxazole.The resistance rate were85.72% respectively.In contrast,Oxytetracycline,Doxycycline,Tetracycline were relatively sensitive,and the sensitive rate were 31.43%,22.86% and 20%,respectively.The results of the detection of drug resistance genes showed that the detection rates of Aminoacyl aacC2(57.14%)and aacC4(42.86%)were the highest;the detection rates of Beta lactam blaTEM(48.57%),blaCTX-M(31.43%)and Quinolones GyrA(37.14%),ParC(37.14%)were similar;Macrolides ermB(20%)and mefA(17.14%)are relatively low;Aminoglycoside AphA3 and Macrolide ermA were not detected.In this study,the minimal inhibitory concentration(MIC)of Amikacin(AK),Ceftiofur(EFT),Erythromycin(E)and Marbofloxacin(MBF)alone was determined against pathogenic Streptococcusisolated from cattleby broth dilution method,the combined MIC by checkerboard method,and then calculated fractional inhibitory concentration(FIC),and judging the interaction after drug combination.The mutant prevention concentration(MPC)both alone and combined was determined by the method of agar dilution method,and then the value of selectivety index(SI)(MPC/MIC)would be acquired.Determine whether to narrow the resistance mutation selection window(MSW)and the best combination of drug use is selected.The results show,when used alone,the MICs of AK,EFT,E,MBF were 8-64,0.5-8,0.125-16,1-16?g/mL,respectively;After combined use,the FICs index of the combination were 0.625-1,0.3125-0.751,0.3125-0.75 indicating that the two drugs were synergistic or additive effect.The MPCs of AK alone against 7strains of Streptococcus mastitisisolateswere 128-512,64-512,16-256,64-512?g/mL,respectively;After combined use,SI decreased to 1-2,1-2,2-4.The results showed that AK could be reduced to MSW after combined use of AK and other drugs respectively,and the combined effect of AK and MBF was more obvious.This study detected the virulence genes and drug resistance of pathogenic Streptococcus isolated from cattle,and providing a theoretical basis for the development of new vaccine based on streptococcal virulence gene.In addition,by studying the drug susceptibility and resistance of clinical isolated streptococcus,this study offers data for improving the current status of drug resistance and reduce the production of drug-resistant mutant strains of pathogenic bovine streptococcus and for the prevention and treatment of mastitis.