鳗弧菌和溶藻弧菌二联疫苗对大菱鲆的免疫效果

Immune efficacy of a bivalent vaccine for turbot against Vibrio anguillarum and V.alginolyticus

将鳗弧菌(Vibrio anguillarum)和溶藻弧菌(V.alginolyticus)分别用0.3%福尔马林灭活,制备成鳗弧菌和溶藻弧菌2种单苗,并将2种菌等量配制成二联疫苗,单次腹腔注射免疫大菱鲆(Scophtalmus maximus)。通过检测免疫后替代途径补体活力、溶菌酶活力、抗体凝集效价的变化并分别进行攻毒实验比较疫苗的免疫效果。结果显示,3种疫苗对所测定的各免疫指标都有一定的促进作用,但二联疫苗后期效果低于2种单苗。二联疫苗对鳗弧菌和溶藻弧菌的免疫保护率分别为83.52%和83.24%,而鳗弧菌单苗对鳗弧菌攻毒、溶藻弧菌单苗对溶藻弧菌攻毒的免疫保护率分别为80.37%和74.89%。可以认为2株菌具有制备二联疫苗的可能性。

The most commonly encountered Vibrio species of pathogen in fish is Vibrio anguillarum, which is responsible for the majority of losses worldwide. V. alginolyticus also is common pathogen. The main objective of this work was to design a bivalent vaccine against these two bacteria and to study its effectiveness. The V. anguillarum vaccine, V. alginolyticus vaccine anda bivalent vaccine were prepared against these two pathogens from 0.3 % formalin-killed cells without any adjuvant. The final concentrated bivalent vaccine that was produced using equal volumes of each monovalent vaccine were at 2×10^9 CFU/ mL. Turbots （ Scophtalmus maximus）, body weight （31.2±1.9） g, were divided into four groups, each with 30 or 60 individuals. The fish were immunized with the vaccines by single i.p. injection in a volume of 0.2 mL. Control group received 0.2 mL of PBS. The fish were starved for 2 days prior to vaccination. The efficacy of vaccines was evaluated by the measurement of alternative complement activity （ACH50）, lysozyme activity, antibody titer and challenge of the two bacteria respectively. Plasma were collected per week from 15 turbots in each group and stored at - 20℃. ACH50 and lysozyme activity were measured following Yano and Ellis. Antibody titer was determined by use of a microtitration agglutination test. At last the fish were challenged by i.p. injection of the bacterial suspension and monitored for 14 days. Mortalities were recorded daily. To evaluate the significance of the difference between vaccined and unvaccined turbots, statistical analysis was performed using SPSS software （11.0）. The results indicated that the three vaccines enhanced the tested non-specific and specific immune functions. But the lysozyme activity and antibody titer of bivalent vaccine were lower than two monovalent vaccines in the last 2 - 3 weeks. ACH50 of bivalent vaccine became lower and lower from the beginning of experiment. In this work, the three immune parameters might be affected by two antigens administered contemporary. It might be the evidence of antigenic competition. The bivalent vaccine induced protection rates of 83.52 % and 83.24 % against V. anguillarum and V. alginolyticus respectively. The protection rates of V. anguillarum vaccine against Vibrio anguillarum and V. alginolyticus vaccine against V. alginolyticus were 80.37 % and 74.89%. Immunological cross-reaction may be the possible mechanism between V. anguillarum and V. alginolyticus. In conclusion, this new bivalent vaccine is effective in protecting turbots against V. anguillarum and V. alginolyticus. It is possible to design a bivalent vaccine.