Protective Effect Of CpG-ODN On Induced Goat Mastitis By Combination Of Two Bacterial Pathogens

This study chooses Suining White goats in early stages lactation as experimental animal,and investigates the effect of CpG-ODN on goat mastitis co-induced by Staphylococcus Aureus and Escherichia Coli.Furthermore,the experiment preliminary establish and optimize the two-dimensional electrophoresis(2-DE) patterns of mammary tissues.This research includes three series:Series1:Protection of CpG-ODN to Mammary Tissues of Goat Experimental Mastitis Co-induced by Two Bacterial PathogensSix healthy Suining White goats in their early lactation,weighing 30～40 kg were used in this study.The mammary right quarters of 6 goats were infused with CpG-ODN according to a dosage of 100μg·kg^-1BW on the 5^th day of postpartum.Left quarters were used as self controls and infused with sterile PBS.On the 8^th day of postpartum the same dosage of CpG-ODN or PBS were infused respectively for the second time.Then on the 9^th day of postpartum,the mammary quarter(both right and left) of 6 goats were infused with 1.5 mL of 2×10⁹ CFU·mL^-1 S.aureus and 2×10⁹ CFU·mL^-1 E.coli per mammary tissues respectively.Goats were slaughtered on the 72 h postinfection and the mammary tissuess were harvested.Clinical observation and histopathological examinations revealed that infusion with 3 mL of 2×10⁹ CFU·mL^-1 S.aureus and 2×10⁹ CFU·mL^-1 E.coli per mammary tissues could induced mastitis of goats.The results showed that infusion of CpG-ODN reduced the concentration of tumor necrosis factor-α(TNF-α)(P＜0.05),interleukin-1β(IL-1β)(P＞0.05),and interleukin-6(IL-6)(P＞0.05) and the activity of N-acetyl-β-D-glucosaminidase (NAGase)(P＜0.01),Myeloperoxidase(MPO)(P＜0.05).The result of the study showed that CpG-ODN inhibits release of inflammatory response that causes damage to mammary epithelial cells during mastitis infections in goats.The study demonstrated that the CpG-ODN protected against mastitis by S.aureus and E.coli infection in goat.Series2:Effect of CpG-ODN on the Cytokines and Enzymes in the Milk and Serum of Goat Experimental Mastitis Co-induced by Two Bacterial PathogensThe milk and serum samples were collected and analyzed.The result of the study showed that TNF-αpeaked at 24 h in PBS-treated group and postinfection.in CpG-ODN treated group,but the TNF-αin CpG-ODN treated group decreased by 10.38%(P＜0.05) compared with PBS-treated group at 24 h postinfection.CpG-ODN induced significant elevated IL-2 before infection compared with the PBS-treated group.IL-1βin milk peaked at 24 h in CpG-ODN treated group(P＜0.01) and 48 h in PBS-treated group,and that in CpG-ODN treated group decreased by 11.87%(P＜0.05) at 24 h and 28.72%(P＜0.01) at 48 h postinfection compared with PBS-treated group.In CpG-ODN treated group,NAGase and MPO activities increased with the duration of infectin and both peaked at 24 h, However in PBS-treated group the dynamic changes of NAGase,MPO activities were similar to those in CpG-ODN treated group.But the NAGase peak value was higher than those in latter and the MPO peak value was lower.In serum,TNF-αincreased and peaked at 24 h,at 8 h had significantly different compared with 0 h(P＜0.05);IL-1β,IL-2 increased and peaked at 24 h,48 h respectively,anytime compared with 0 h had no remarkable changes;the activities of MPO lowered at 24 h,compared with 0 h had significantly different(P＜0.01);the activities of NAGase also lowered at 48 h and compared with 0 h had no remarkable changes.The result of the study showed that CpG-ODN stimulated the production of IL-2,prompted the release of the TNF-αand IL-1βattenuated the destroy of inflammation-mediator to mammary tissues and shortened the inflammation course.The study confirmed that the CpG-ODN protected against mastitis by S.aureus and E.coli infection in goat.Series3:Effect of CpG-ODN on the Content of Lactoferrin in Milk and Two-dimensional Electrophoresis of Mammary Tissues of Goat Experimental MastitisTo measure the content of lactoferrin in milk of experiment using ELISA method. The results show,in CpG-ODN treated group,the content of lactoferrin increased and peaked at 16th hour(P＜0.01,compared to 0 th hour);in PBS treated group,the content of lactoferrin similar to it in CpG-ODN treated group,but it peaked at 24th hour(P＜0.01, compared to 0 th hour).CpG-ODN maybe protect against goat mastitis through nonspecific immunity.Furthermore,the experiment preliminary establish and optimize the two-dimensional electrophoresis(2-DE) patterns of mammary tissues.The results indicated that the method was efficient to mammary tissues for proteomic analysis.