运用实时荧光PCR法检测感染鸡种蛋中的REV

Detection of REV in Eggs of Infected Chickens by Real-time Fluorescent Quantitative PCR

关键词:  网状内皮组织增生症病毒  荧光定量PCR  血清  蛋清  鸡胚成纤维细胞

Key words:  REV  Real-time PCR  Serum  Egg White  CEF

摘要:

为研究通过种蛋检测的方法监测SPF鸡群REV感染情况，运用已建立的荧光定量PCR方法检测攻毒母鸡种蛋中REV，并与血清REV水平进行比较。人工感染12月龄母鸡，从种蛋中提取蛋清，接种正常鸡胚成纤维细胞后传代一次，之后石蜡密封孵化至9日龄制成鸡胚成纤维细胞（CEF），运用荧光定量PCR方法检测蛋清，并和血清的检测结果进行比较，结果显示：83份蛋清，3份REV阳性；46份CEF，2份REV阳性，均在攻毒后的11天、14天收集，阳性样品的蛋清和CEF呈对应关系，两种方法的检测结果一致。与血清检测结果比较：阳性鸡蛋均在血清病毒血症持续的时间内采集，表明通过种蛋REV检测能反映SPF鸡的急性感染状态。本研究为通过种蛋抗原检测从而监测SPF鸡REV感染奠定基础。

Abstract:

In order to study the application of REV method to detect REV infection in SPF chickens.To study the detection methods of REV in eggs, the REV of eggs in infected hens was detected by Real-time PCR method which already established, and compared with REV in serum.12 month-old hens were injected REV.The egg white was extracted from the eggs and was inoculated into chicken embryo fibroblast and passage once, then these eggs were sealed with paraffin wax, then were hached, CEF was made from 9 days embryos. Using the real-time PCR to detect these eggs white and CEF,then compared with the serum, the result showed that: REV in three egg white samples could be detected from 83 samples, and two CEF samples could be detected from 46 samples, the egg white and the CEF was correspondence which were collected in the eleventh and the fourteenth day after infectiong,the result of the two methods are consistent.The result of the serum detection showed that the REV could be detected from the third day after infection and reached the highest level on the ninth day, wasn’t tested from the eighteenth day. Positive eggs were collected during the time of the serum viremia, which indicated that the REV test can reflect the acute infection status of SPF chicken.The research provided a foundation for monitoring REV infection in SPF chickens by egg antigen test.day.The experiment proved that the collection time of positive eggs was consistent with the viral disease, which indicated that the REV test can reflect the acute infection status of SPF chicken.The research provided a foundation for monitoring the quality of SPF chicken using antigen detection of eggs.