传染性法氏囊病毒VP2基因重组新城疫病毒疫苗的胚胎免疫研究
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摘要
鸡胚免疫接种能使雏鸡出壳时至出壳后几天内就具有特异性主动免疫力,该方法具有提早免疫、应激反应低、注射精确且一致、劳动力成本低和疫苗污染少等优点,一直是国内外学者的研究热点。
本研究选用表达传染性法氏囊病病毒(IBDV)超强毒株VP2基因重组新城疫病毒(NDV)Lasota疫苗株(rL-VP2),对其半数鸡胚感染量(EID_(50))、平均鸡胚致死时间(MDT)、脑内接种指数(ICPI)和静脉内致病指数(IVPI)等毒力指标进行了测定,结果表明重组病毒rL-VP2具有低致病性和高滴度鸡胚生长特性。
选择18胚龄的SPF鸡胚,按不同的接种剂量经羊膜腔途径实施胚胎免疫接种。出雏后对不同剂量组的孵化率进行统计,并分别于出雏后的第9 d、第14 d和第21 d对免疫鸡血清中抗IBDV抗体和NDV抗体水平进行测定,结果显示按104 EID_(50)/egg剂量进行免疫不会影响SPF鸡胚出雏率和健雏率,并且能够使免疫鸡产生较高的抗体水平,因此确定104 EID_(50)/egg为rL-VP2实施胚胎免疫的最佳免疫剂量。
在出雏后第4周,对10 4 EID_(50)/egg免疫组的雏鸡分别用IBDV强毒vvIBDV Gx株和NDV强毒F48E9株进行攻击,结果显示rL-VP2免疫鸡能对两种强毒的攻击产生100%的免疫保护,攻毒后对雏鸡的增重未见显著的影响(P>0.05),免疫鸡的法氏囊组织在攻毒后未见明显的病理变化。
本研究表明用重组病毒rL-VP2进行胚胎免疫具有较好的安全性和有效性,为同时预防IBD和ND的疫苗在胚胎免疫上的应用奠定了理论基础。
Chicken embryo immunization can generate immune response sooner than conventional approach, lead low stress response and reduce vaccine pollution, inject accurate and consistent, decrease labor costs.This method has been the research focus of scholars in the worldwide.
A recombinant Newcastle disease virus(rL-VP2) expressing the VP2 gene of vvIBDV Gx strain was used in the study.After determining the EID_(50), MDT, ICPI and IVPI of rL-VP2, the vaccine was proved that kept the low pathogenicity similar with LaSota vaccine strain and replicated with high-titer similar to that of parental NDV strain LaSota in chicken embryos.
The vaccine was administered through amnion cavity to 18-day-old specific-pathogen-free embryos with different doses. The percentage hatchability of each group was counted. To evaluate the efficacy of the recombinant vaccine against NDV and IBDV, the antibody titers were detected at 9th day, 14th day and 21st day post hatching. The results showed that the hatchability or 3rd week survivability did not decrease and the hatched chicks could obtain specific active immunity with higher antibody levels than other groups when the dose of 104 EID_(50)/egg was administered. According to the results, the optimum dose of rL-VP2 using in ovo was 104 EID_(50)/egg.
The group which administered with the dose of 104 EID_(50)/egg was challenged with a highly virulent NDV strain F48E9 and vvIBDV Gx strain at four weeks post-hatched. The result showed that 100% of chickens was protected from challenge, the weight of the chickens did not have significant difference(P>0.05) from the control group, and bursa of Fabricius did not present significant damage in this group.
These results are promising for the development of embryo safe efficacious IBDV and NDV vaccines for in ovo application.
本研究选用表达传染性法氏囊病病毒(IBDV)超强毒株VP2基因重组新城疫病毒(NDV)Lasota疫苗株(rL-VP2),对其半数鸡胚感染量(EID_(50))、平均鸡胚致死时间(MDT)、脑内接种指数(ICPI)和静脉内致病指数(IVPI)等毒力指标进行了测定,结果表明重组病毒rL-VP2具有低致病性和高滴度鸡胚生长特性。
选择18胚龄的SPF鸡胚,按不同的接种剂量经羊膜腔途径实施胚胎免疫接种。出雏后对不同剂量组的孵化率进行统计,并分别于出雏后的第9 d、第14 d和第21 d对免疫鸡血清中抗IBDV抗体和NDV抗体水平进行测定,结果显示按104 EID_(50)/egg剂量进行免疫不会影响SPF鸡胚出雏率和健雏率,并且能够使免疫鸡产生较高的抗体水平,因此确定104 EID_(50)/egg为rL-VP2实施胚胎免疫的最佳免疫剂量。
在出雏后第4周,对10 4 EID_(50)/egg免疫组的雏鸡分别用IBDV强毒vvIBDV Gx株和NDV强毒F48E9株进行攻击,结果显示rL-VP2免疫鸡能对两种强毒的攻击产生100%的免疫保护,攻毒后对雏鸡的增重未见显著的影响(P>0.05),免疫鸡的法氏囊组织在攻毒后未见明显的病理变化。
本研究表明用重组病毒rL-VP2进行胚胎免疫具有较好的安全性和有效性,为同时预防IBD和ND的疫苗在胚胎免疫上的应用奠定了理论基础。
Chicken embryo immunization can generate immune response sooner than conventional approach, lead low stress response and reduce vaccine pollution, inject accurate and consistent, decrease labor costs.This method has been the research focus of scholars in the worldwide.
A recombinant Newcastle disease virus(rL-VP2) expressing the VP2 gene of vvIBDV Gx strain was used in the study.After determining the EID_(50), MDT, ICPI and IVPI of rL-VP2, the vaccine was proved that kept the low pathogenicity similar with LaSota vaccine strain and replicated with high-titer similar to that of parental NDV strain LaSota in chicken embryos.
The vaccine was administered through amnion cavity to 18-day-old specific-pathogen-free embryos with different doses. The percentage hatchability of each group was counted. To evaluate the efficacy of the recombinant vaccine against NDV and IBDV, the antibody titers were detected at 9th day, 14th day and 21st day post hatching. The results showed that the hatchability or 3rd week survivability did not decrease and the hatched chicks could obtain specific active immunity with higher antibody levels than other groups when the dose of 104 EID_(50)/egg was administered. According to the results, the optimum dose of rL-VP2 using in ovo was 104 EID_(50)/egg.
The group which administered with the dose of 104 EID_(50)/egg was challenged with a highly virulent NDV strain F48E9 and vvIBDV Gx strain at four weeks post-hatched. The result showed that 100% of chickens was protected from challenge, the weight of the chickens did not have significant difference(P>0.05) from the control group, and bursa of Fabricius did not present significant damage in this group.
These results are promising for the development of embryo safe efficacious IBDV and NDV vaccines for in ovo application.
引文
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