HighlightsDeveloped, optimised and validated q-PCR to distinguish CVI988 and virulent MDV.Based on differential detection of single nucleotide polymorphism in pp38 gene.Specific, sensitive, efficient, reproducible assay with no false-positive results.Accurate differential quantification over biological range of virus levels.Replication of CVI988 and virulent MDV examined in chicken feathers and blood.

Infectious bursal disease virus (IBDV) exists in several different antigenic and pathogenic forms. The immune suppression caused by this virus in young chickens is not always associated with clinical signs of disease. The antigenic variant viruses originally described in the United States typically do not cause clinical signs of disease but can cause a marked immune suppression via the destruction of B lymphocytes. Using a reverse transcription-polymerase chain reaction (RT-PCR) assay we conducted a survey of asymptomatic broiler chicken flocks in Europe for IBDV. Restriction fragment length polymorphisms in the viral protein 2 (VP2) gene of four isolates from Spain and four isolates from France indicated they may be different from the classic and very virulent (vv) IBDV strains found throughout Europe. Nucleotide sequence and phylogenetic analysis of the hypervariable region of the VP2 gene indicated that all eight viruses were more similar to U.S. variant viruses than classic viruses. In two viruses, one from France and one from Spain, threonine was observed at amino acid position 222 and serine was found at position 254. These two substitution mutations are characteristic of Delaware variant viruses. In addition, all eight viruses had mutated amino acid position 318 from glycine to aspartic acid, another substitution mutation commonly found in U.S. variant viruses. Although importation restrictions prevented us from directly testing the antigenicity of these viruses, their nucleotide and predicted amino acid sequences suggest they could be antigenically distinctive compared to classic and vvIBDV commonly found in Europe. Confirmation of the presence of antigenic variant IBDV strains in Europe requires additional immunologic studies to elucidate the exact nature of the viral epitopes. Our data support the need for these immunologic studies.

The use of the complete DNA sequence for the Marek's disease virus (MDV) serotype 1 vaccine strain CVI988 Rispens in comparative genomic studies with virulent strains of MDV has revealed the presence of a number of insertions, deletions and single-nucleotide polymorphisms. In this study, we investigated a SNP in the H/ACA box of the viral RNA subunit of telomerase (vTR). We sequenced vTR from four different batches of CVI988 vaccine originating from a single commercial company. The A-to-G mutation defining the SNP in the H/ACA box of CVI988 vTR was present in only some of the batches. Thus, although this mutation affects CVI988 vTR function, it is not shared by all CVI988 isolates and may be a stochastic rather than causative event in CVI988 attenuation.