Research On Fauna Investigation Of Main Pathogenic Bactcria And Genotyping Of Staphylococcus Aureus Caused Bovine Mastitis

Bovine mastitis is a frequently encountered disease of cow and causes very tremendous economical losses. Mainly caused by a variety of unspecified pathogens. It is very important for studying the species and distributing of the main pathogenic bacteria causing bovine mastitis in different regions in China,identifying the susceptivity to antibiotic of mail pathogens,and understanding the genotype of the different districts of our country and definite the connection between the typing and the pathogenicity of S.aureus. Research showed, staphylococcus aureus was main pathogens of bovine mastitis, which separation rate from 20% to80, spread widely and easy to generate the drug resistance. Therefore,it may provide the necessary technical data on prevention and cure bovine mastitis by researching the S.aureus.541 milk samples from cows suffered from clinical or subclinical mastitis were collected from cattle farm in Inner Mongolia,Gansu,Shanghai,Sichuan and Guizhou,and the main pathogens were identified and used for drug sensitive test. The results showed that 570 strains of main bacteria were identified from the milk samples by traditional methods,the average rate were 78.00%. There are 36 strains among them are Staphlococcus aureus (29.03 %),36 are Streptococcus agalactiae(29.03%), 24 are Streptococcus dysgalactiae(19.35%),16 are E.coli(12.90%),5 are Staphylococcus epidermidis(4.03%),3 are Streptococcus uberis(2.42%) in Inner Mongolia.66 strains among them are Staphlococcus aureus (44.59 %) ,29 are Staphylococcus epidermidis(19.59 % ),17 are Streptococcus agalactiae(11.49 % ),13 are Streptococcus dysgalactiae(8.78%),11 are Streptococcus uberis(7.43%),1 are E.coli(0.67%)in Shanghai. 20 strains among them are Staphlococcus aureus (33.33 %) , 13 are Streptococcus uberis(19.40 % ),7 are Staphylococcus epidermidis(10.45 % ), 6 are Streptococcus dysgalactiae(8.99%), 5 are E.coli(7.46%),3 are Streptococcus agalactiae(4.48%), in Sichuan. 38 strains among them are Staphlococcus aureus (43.18 %) , 15 are Streptococcus dysgalactiae(17.05 % ),12 are Staphylococcus epidermidis(13.64 % ),5 are Streptococcus uberis(5.68%),2 are E.coli(2.27%)in Guizhou. 50 strains among them are Staphylococcus epidermidis (34.97 % ), 27 are Staphlococcus aureus (18.88%), 17 are Streptococcus dysgalactiae(11.89%),11are E.coli(7.69%), 8 are Streptococcus uberis(5.59%),4 are Streptococcus agalactiae (2.80%) in Gansu. S. aureus was the advantage pathogenic bacteria in different regions of dairy cow mastitis.The results of drug sensitive test showed that the main pathogenic bacteria almost were produced different degree drug resistance to Penicillin,Tetracyclines and Glycopeptides,etc.,while Quinolones and Glycopeptides,etc.had a obviously bacteriostasis.These isolated 176 S.aureus strains and standard strain CVCC2246 undertake the RAPD typing use three universal primers 4M,OLP13 and AP7. Then use BioNumerics software to RAPD amplification products clustering analysis.Three primers for 177 strains of Staphylococcus aureus RAPD amplified bands were between 1 to 10, molecular size bands obtained between 250 ~ 4500bp. According to differences in genetic distance, 172 strains staphylococcus aureus was divided into 11 major groups of clustering by primer OLP13, 175 strains staphylococcus aureus was divided into 8 groups of clustering by primer 4M, 174 strains staphylococcus aureus was divided into 12 mainly gather groups by primer AP7.From the classification rateand the number of amplified bands, the size range of amplified bands of view, 4M was the best primer, OLP13 second, AP7 poor, but the genetic map and its discrimination between clustering coefficient shows OLP13 best. Coagulase gene polymorphism typing used for genotyping of the identified S.aureus respectively,the results showed that coagtdase gene typing revealed five types,and product size of about 900bp of the PCR 2 type was the dominant type, accounting for 33.33% isolates. The PCR 2 type was divided into two different subtypes and other bands no difference after digestion.Identifying the 176 staphylococcus aureus and 1 negative control strains basing on the PCR of 16S rRNA and 23S rRNA.A polymerase chain reaction (PCR) assay was developed for the detection of the Staphlococcus aureus of bovine mastitis. The results showed that PCR identification basing on 16S rRNA and 23S rRNA were 100% both in line with the traditional identification methods.