| Dairy cow mastitis is an udder inflammation caused by physical, chemical or microbial factors stimulation, and the major cause of this disease is a variety of pathogenic microbes'infection. It exists in the world widely and has very high incidence, which not only bring huge economic losses of dairy cow production, but also do harm to dairy industry, public health and food safety. The aim of this study was to analyze the characteristics of the main pathogenic bacteria causing dairy cow mastitis, and to provide a theorical foundation for epidemiological investigation, serological and drug resistance studies, clinical administration, cure and prevention. In this study, we isolated and identified the main pathogenic bacteria causing dairy cow mastitis from dairy farms in Shanghai area, and analyzed its genetic typing and antibiotics resistance, then discussed the effect of recombinant plasmid containing human lysozyme cDNA (pcDNAKLYZ) for cure and prevention of dairy cow mastitis, and evaluated the safety of this gene pharmaceutical to environment. This study includes the following four main elements:(1) Isolation and identification of main pathogenic bacteria causing dairy cow mastitis in Shanghai area. 107 strains of bacteria were isolated from the 126 milk samples of dairy cows with mastitis or subclinical mastitis in Shanghai area. Results showed that 74 strains among them were Staphylococcus aureus (69.16%), 27 were Streptococcus agalactiae (25.23%). In the all 126 milk samples, there were 40 samples infected by only one kind of bacterium. 32 were infected by Staphylococcus aureus, and 8 infected by streptococcus agalactiae; while the others were multi-infected, mainly including Staphylococcus aureus and streptococcus agalactiae.(2) Genetic typing the Staphylococcus aureus causing dairy cow mastitis. 74 Staphylococcus aureus isolated from mastitis milk were typed by ERIC-PCR, which could be amplified out 4~9 straps, revealeing 4 genotype. The strains showed identical or near genetic relationship occupy 79.7%. Isolates from the same regions belonged to the different gene type, and isolates from different regions belonged to the same gene type. This indicated that there may be a main strain of Staphylococcus aureus in a farm or different farms, or many strains of Staphylococcus aureus.(3) Drug sensitivity of main pathogenic bacteria causing dairy cow mastitis in Shanghai area. Drug sensitivity test showed that all pathogenic bacteria were resistant to penicillin, while cefradine, ciprofloxacin and gentamycin had a good inhibiting effect on these main pathogenic bacteria. Especially, almost all the bacteria were sensitive to cefradine. (4) Therapeutic effect and environmental release experiment on treating dairy cow mastitis using gene pharmaceutical in Shanghai area. To discuss the therapeutic effect and evaluate the safety of recombinant plasmid containing human lysozyme cDNA (pcDNAKLYZ), we compared the result of SMT, the total number of bacteria and main pathogenic bacteria in pre-treatment to those in post-treatment, and also detected recombinant plasmid from blood samples and milk samples, tested transformation bacteria from spit, urine, feces and environment after treating. Results showed that the cure rate was 90.91% of dry cow, while the effective power was 82.4% in lactation period. The total number of bacteria and pathogens decreased significantly and had gradually declining trend. Positive rate of dairy cow was 70.1 %( 5/7) detected by PCR in milk only 1 d after treating. The gene pharmaceutical was impossible to disperse into environment by secretion and feces, and diffuse parallelly in environment because of transformation of bacteria. These results indicated that pcDNAKLYZ was effective to cure and prevent dairy cow mastitis and safe to environment.
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