| Duck tembusu virus disease is caused by the duck tembusu virus,which spread in egg-laying ducks extensively.The infected egg-laying ducks showed a severe drop in egg production.Which bring a huge financial loss to the culture industry of waterfowl in China.As there is no drug for the treatment,the preventive immunity of the disease is particularly important.The virus strains for this study,which was identified by Anhui Agricultural University from the sick ducks in 2010,which was named AH-F10 strains.The oil emulsion inactivated vaccine generated by the allantoic fluid proliferated tembusu virus had an important significance of preventing and controlling the duck tembusu disease in Anhui.The AH-F10 isolates was took as the candidate strain,three batches vaccine(1401,1402,1403 batches)were generated and evaluated by the different immune ways,repetitive immune and high dose immune.The evaluated method was as follows:(1)80healthy ducks in 60 day-age were randomly divided into eight groups,the ducks of group1,2 and 3 were immuned respectively by the vaccine of the 1401,1402,1403 batches by the way of intramuscular injection,and the ducks of group 4 was injected aseptic PBS as a control;the ducks of group 5,6 and 7 were immuned respectively by the vaccine of the1401,1402,1403 batches by the way of subcutaneous injection,and the ducks of group 8was injected aseptic PBS as a control,the immune dose was 0.5mL/single.(2)In the first immunization,40 healthy duck in 60 days-age were randomly divided into four groups,1401,1402,1403 batches of vaccine were immuned to the duck of group 1,2 and 3respectively by intramuscular injection,and the ducks of group 4 was injected aseptic PBS as a control,which with a immune dose of 0.5mL/single;the second immunization was14 days later after the first immunization with the same immunity method and immune dose;(3)40 healthy ducks in 60 days-age were randomly divided into four groups named 1-4,40egg-laying ducks in 120 day-age were randomly divided into four group named 5-8,the ducks of group1,2 and 3 were immuned respectively by the vaccine of the 1401,1402,1403 batches by the way of subcutaneous injection,the ducks of group 4 was injected aseptic PBS as a control,which with a immune dose of 1.0m L/single.The same immunity method and immune dose to the group 5,6,7 and 8.The results showed that,the groups of ducks immuned by 0.5mL dose are not found partly or systemic adverse reactions,and have a normal condition of mental state,gathering food and other activities.weight tests results showed there was no difference between the experimental group and the controlgroup;to immune laying duck with overdose,four groups of laying duck total egg production from 24/d fell to the lowest 4/d.After 4 days,egg production began to recover,and back to 24/day after 20 d.This results denmostrated that the duck tembusu virus AH-F10 inactivated vaccine possess a good safety in clinical application.For the study of the immunity and protection effect of duck tembusu virus(AH-F10)oil emulsion inactivated vaccine to ducks,30 uninfected and antibody-negative healthy ducks were divided equally into three groups,only the ducks of group 2 were immuned1401 batches of inactivated vaccines in the dose of 0.4mL,the ducks of group 1 and group3 were inoculated 0.4mL aseptic PBS,16 days later,the ducks of group 1 and group 2 were challenged with 0.4mL DTMUV(ELD50 = 10-2.67/0.2mL).The results showed that ducks of the group 2 does not appear partly or systemic adverse reactions after immunization,the serum antibody of DTMUV was detected by the method of indirect ELISA,the OD450 nm value ducks serum antibody of Group 2 increases to 0.37,positive rate was 50%(5/10)at14 days after the immunization.The results of challenge test with DTMUV showed that,the ducks of group 2 were not seen morbidity or mortality,vaccine protection ratio reached100%;the ducks of group 1 incidence reached 100%(10/10),mortality rate of 60%(6/10).The results of weight test showed that the average weight of the ducks of group 2 has a decrease of 7 g at 3 days after the challenge,then return to growth,and reach the average weight of group 1 and group 3.After challenge test,the ducks of group 1,group 2 and group 3 average daily gain were 14.8g,23.7g and 24.3g.Therefore,the results showed that the 1401 batches of oil emulsion inactivated vaccines has a good safety for the ducks,had no effects to the growth of ducks,and the inactivated vaccines can effectively stimulate the ducks to produce antibodies and protect the ducks,and provided a certain data to the further study of the duck tembusu virus oil emulsion inactivated vaccine.The ELISA detection method was established due to the E protein of duck tembusu virus AH-F10 strains with the optimal working concentration of of 2.978?g/well and the optimal antigen coating tempreture of 4 ? for 16 h,the optimal dilution degree of serum was 1:100,the optimal dilution degree of HRP-IgG was 1:500,the optimal incubation time of serum and HRP-IgG were both 40 minutes.The results showed that the established method of indirect ELISA was a high specific and sensitive method for the antibody detection and epidemiology investigation of duck tembusu virus. |
PDF Full Text Request