| Dairy cattle mastitis was one of the most common diseases and the most economically losses of dairy cattle diseases affecting dairy industries around the world. And dairy cattle mastitis has brought the latent harm to the human health. Nowadays, there were the large numbers of pathogenic bacteria causing dairy cattle mastitis. Not only had the much use of the antibiotics under the treatment made the dairy cattle produce the resistance to pathogenic bacteria but also the residue of the antibiotics had been increasingly affecting the food sanitation security and the human health, et al. Utilizing the vaccines to preventing and controlling dairy cattle mastitis was a kind of safe, economical and effective method. In this study, the primary pathogenic bacteria causing dairy cattle mastitis were isolated and identified. The triple inactived vaccine was developed. The immunity effectiveness of this vaccine was evaluated in the aspects of immunity adjuvants and immunity routes by using various experimental animals. This study was divided into four parts:Trial 1 Isolation and identification of the primary pathogenic bacteria of dairy cattle mastitis and the analysis of the pathogenicityIn this study, the 160 milk samples of clinical and subclinical mastitis from five dairy cattle farms in Jinan, Taian, and Linyi of Shandong and so on, were collected and measured. And the primary pathogenic bacteria were isolated and identified from all collected milk samples by the methods of the conventional biochemistry experiments and PCR identification. The results indicated that: there were 127 positive milk samples of the dairy cattle mastitis in 160 milk samples. The clinical mastitis was 36.22% and the subclinical mastitis was 63.78% in these 127 milk samples of the mastitis cases. The primary pathogenic bacteria Staphylococcus aureus, Escherichia coli and Streptococcus agalactiae were isolated from these milk samples. Staphylococcus aureus was sorted toαhemolysis,βhemolysis,α+βhemolysis and non-hemolysis by the different types of hemolysis; Escherichia coli was sorted to O2,O21,O76 and O88 by the different serotypes of O antigens; Streptococcus agalactiae was mainlyβhemolysis by the different types of hemolysis. The pathogenicity of the the primary pathogenic bacteria was analyzed to screen the typical strains of the stronger pathogenicity.Trial 2 The development of the triple inactived vaccine of dairy cattle mastitis and the evaluation of the experimental immunity effectivenessIn this study, the typical Sta.aureus(Staphylococcus aureus),E.coli(Escherichia coli)and Str.aga(Streptococcus agalactiae)were isolated from the milk samples of the dairy cattle mastitis, and a kind of the triple inactived vaccine was developed for preventing the dairy cattle mastitis. In order to evaluate the immunity effectiveness of this vaccine initially, the immunity trial and challenge trial in mice have been done. The antibody titers of the mice were measured by indirect ELISA. The results showed that, this vaccine exhibited better immunity properties. Compared with the control group, this vaccine significantly enhanced the antibody titers of the mice (P<0.01), and protected the mice from the virulent bacteria challenge, and the average protection index was 73.3%.Trial 3 The evaluation of the immunity effectiveness of the triple inactived vaccines of the various immunity adjuvantsIn this study, the more practical, security and effective adjuvant vaccine was screened by the comparisons of the immunity effectiveness of the various immunity adjuvant vaccines. The New Zealand rabbits and the healthy Holstein dairy cattle during the mid-lactation were included in this experiment. The immunity trials were done by the immunization of the oil-tween adjuvant vaccine and Al(OH)3 adjuvant vaccine to comparing the immunity effectiveness of the two kinds of adjuvant vaccines. The results showed that: the oil-tween adjuvant vaccine and the Al(OH)3 adjuvant vaccine could both improve the immunity of the organism and significantly improve (P<0.01) the specific antibody in serum. But the oil-tween adjuvant vaccine could better significantly improve (P<0.01) the specific antibody in serum than the Al(OH)3 adjuvant vaccine, maintain the higher specific antibody of the longer time and significantly reduced the SCC of the milk samples. Moreover, the stability of the oil-tween adjuvant vaccine was better, the price was cheap and this vaccine was easy to apply to the practice.Trial 4 The evaluation of the immunity effectiveness of the triple inactived vaccines by the various immunity routesA more effective and more convenient immunity route was found by the comparisons of the immunity effectiveness of Houhai point inoculation and conventional inoculation of the triple inactived vaccine of dairy cattle mastitis. The healthy Holstein dairy cattle during the mid-lactation were included in this experiment. By the comparisons of Houhai point inoculation, intramuscular inoculation and subcutaneous inoculation of this vaccine, the results showed that: firstly, the group of Houhai point inoculation could save the dosage of the vaccine, produce the firm immunity in advance, significantly improve (P<0.01) the specific antibody in serum and maintain the higher specific antibody of the longer time. Secondly, the group of Houhai point inoculation significantly improved the number of the white blood cells and the percents of lymphocytes and the functions of the immunocyte by the functions of acupuncture. And finally, the group of Houhai point inoculation significantly reduced the SCC of the milk samples (P<0.01). |
