Increased virulence of Marek's disease virus type 1 vaccine strain CV1988 after adaptation to qt35 cells.

CVI988/Rispens strain of Marek's disease virus type I (MDV-1) is widely used as efficient vaccine to control Marek's disease (MD) in chicken flocks. Similarly to other live MD vaccine viruses it is propagated in freshly prepared chicken embryo fibroblasts (CEF). In this study, MDV-1 CVI988/Rispens strain was adapted to QT35 cells. The adapted virus, designated QT-CVI, exhibited similar cytopathic effect (CPE) in vitro to that of parental virus propagated in CEE In contrast, QT-CVI induced MD symptoms typical for mild MDV-1 strains after injection to birds. For identification of differentially expressed transcripts that might be involved in increased virulence of QT-CVI, we performed subtractive suppression hybridization (SSH). Subtracted PCR products mapped within MDV-1 BamHI-A and -H fragments and differential gene expression was also confirmed by Northern blot analysis with probes derived from these regions. To examine possible divergence at the virus genome level, PCR analysis was carried out. The BamHI-H fragment- specific 132 bp repeats were present at variable copy number, ranging from 2 to more than 30 copies in both CVI988/ Rispens and QT-CVI DNAs. PCR assays with primers mapping at the US/lRS junction identified CVI988/ Rispens-specific insertion of 116 bp in the region upstream of the ICP4 open reading frame (ORF). PCR analysis was positive also for DNA from non-infected QT35 cells and was consistent with the observation of Yamaguchi et al. (J Virol. 74, 10176-10186, 2000) who have found that QT35 cells carry a latent MDV-1 genome. It is likely, that adaptation of CVI988/Rispens to QT35 cells resulted in reactivation of an endogenous MDV-1 or at least in induction of expression of virulence-related transcripts that have consequently led to QT-CVI pathogenicity for chickens.