Validation Of VAXXITEK® HVT+IBD Vaccine Intake In Chickens Using VHVT13-Specific QPCR By Processing Different Tissue Samples

Lemiere S., Bublot M., de Saint-Vis B., Le-Gros F.X., Dancer A., Bollart A. & Carlotti A. | 16th WVPAC Marrakesh | 2009


A specific vHVT13 qPCR assay using a beacon probe was developed and validated for monitoring of VAXXITEK HVT+IBD (Herpes Virus Turkey + Infectious Bursal Disease) vaccine intake in chickens. The target was a specific zone of the construct between the linker and the VP2 IBD virus gene (243 base pair-long). A first set of clinical studies involved conventional chickens reared in isolation units as well as field chickens vaccinated with VAXXITEK HVT+IBD vaccine at one day of age were sampled by taking two wing feathers per bird. Maximum vHVT13 expression was observed in feather tip samples around D28 post-vaccination. Validation of technique of sampling two feathers per bird was based on the specificity and limit of detection of the sample matrix. It is essential that the following conditions for storage and shipment are followed: the samples must be refrigerated or, if stored for more than one week, they must be frozen at around - 20 °C. Samples should be processed as early as possible. FTA® card transfer is a critical operation. Its success is linked, as for all tissue transfer onto a FTA card, to the amount of live tissue transferred. In a recent field experiment, transfer material was not sufficient for standard qPCR testing using the extraction kit validated for feather samples. In a second series of studies, conventional chickens reared in French field conditions vaccinated with VAXXITEK HVT+IBD vaccine at one day of age were sampled by taking in laboratory conditions whole blood and extracting buffy coat white blood cells after centrifugation, bursa of Fabricius and spleen for each bird of a group of 4-5 on D4, D8, D11, D15 then D18 of age. vHVT13 virus replication could be detected in spleen tissues, either fresh or transferred onto a FTA card (Figure 1) for shipments to other countries, from D11 of age. Fresh tissues from bursa (Figure 2) or buffy coats could be used for vHVT13 qPCR from D11-D15 of age as well.