- Volume 89, Issue 10, 2008

BRCA1-associated protein 2 (BRAP2) is known to interact with the kinase suppressor of Ras 1 (KSR1), inhibiting the ERK signal transduction cascade. This study found that an Epstein–Barr virus (EBV) immediate-early protein, Rta, is a binding partner of BRAP2 in yeast and confirmed the binding in vitro by a glutathione S-transferase pull-down assay and in vivo by co-immunoprecipitation in 293(maxi-EBV) cells. Binding studies also showed that Rta and KSR1 interacted with the C-terminal 202 aa region in BRAP2. Additionally, Rta appeared to prevent the binding of KSR1 to BRAP2, activating the ERK signal transduction pathway and the transcription of an EBV immediate-early gene, BZLF1. Activation of the ERK signal transduction pathway by Rta may be critical for the maintenance of the lytic state of EBV.

We have characterized a novel, captured and fully functional viral interleukin (IL)-10 homologue (OvHVIL-10) from the gammaherpesvirus ovine herpesvirus 2. Unlike IL-10 homologues from other gammaherpesviruses, the OvHVIL-10 peptide sequence was highly divergent from that of the host species. The OvHVIL-10 gene is unique amongst virus captured genes in that it has precisely retained the original cellular exon structure, having five exons of similar sizes to the cellular counterparts. However, the sizes of the introns are dramatically reduced. The OvHVIL-10 protein was shown to be a non-glycosylated, secreted protein of M r 21 000 with a signal peptidase cleavage site between amino acids 26 and 27 of the nascent peptide. Functional assays showed that OvHVIL-10, in a similar way to ovine IL-10, stimulated mast cell proliferation and inhibited macrophage inflammatory chemokine production. This is the first example of a captured herpesvirus gene retaining the full cellular gene structure.

Fowlpox virus (FWPV) is found worldwide in poultry and wild birds. FWPV is a natural example of recombination between viruses, as reticuloendotheliosis virus (REV) fragments have been found in all poultry FWPVs and these are implicated in virulence alteration. We aimed to determine the commonality of this phenomenon and analysed FWPVs collected from 128 poultry flocks and birds over the last 10 years. Various fragments of both viruses were amplified and sequenced at the FWPV integration site, located between FWPV open reading frames 201 and 203. Seven isolates were found to contain no REV insertions, including fragments of the REV env, gag and 5′ REV-long terminal repeat (LTR). We demonstrate here for the first time, the existence of poultry FWPVs without REV inserts (two from chickens, one from turkey FWPV and four from wild birds). The REV inserts were heterogeneous in size. In addition to poultry and wild bird isolates, three FWPV vaccine virus strains were examined and found to contain only remnant REV-LTR and no REV envelope gene fragments.

Keratinocytes can be induced to produce cytokines by exogenous stimuli, such as UVB, and dysregulation of this production has been described in various skin diseases, including cancer. In this study, we compared the effect of UVB on the secretion of several cytokines involved in inflammation by human keratinocytes immortalized or not with human papillomavirus (HPV)16 or HPV38 at the mRNA and protein levels. We show that expression of the HPV E6/E7 oncoproteins influences not only the basal cytokine secretion profile of keratinocytes, but also its modulation upon UVB irradiation. In particular, UVB upregulates interleukin (IL)-6, IL-8 and transforming growth factor (TGF)-β in HPV-immortalized cells to a higher extent than in control keratinocytes. Moreover, expression of other pro-inflammatory molecules such as S100A8/9 and interferon (IFN)-κ was downregulated in HPV-immortalized cells. These data support the functional similarity between HPV16 and 38, and suggest an active role of these viruses in modulation of the inflammatory process.

Recent studies have suggested an association between human papillomaviruses (HPVs), particularly species 2 members of the genus Betapapillomavirus, and squamous cell carcinoma (SCC) of the skin. As most of these viruses are uncharacterized, molecular characterization and epidemiology are needed to advance our understanding of their significance in carcinogenesis. This study determined the complete genomes of four betapapillomaviruses of species 2 from skin lesions designated HPV-107, -110 and -111 and FA75[KI88-03], an isolate of an unpublished HPV type, and analysed their prevalence and viral loads in biopsies from SCC, actinic keratosis (AK), basal cell carcinoma, seborrhoeic keratosis and the healthy skin of 263 immunocompetent patients by HPV type-specific real-time PCR assays. Seventeen patients (6.5 %) harboured at least one of the four HPV types in their lesion, whereas seven patients (2.7 %) harboured one or more of the HPV types in healthy skin. Overall, the four viruses were more common in AK than in healthy skin (odds ratio 5.0, 95 % confidence interval 1.4–17.5), but the prevalence and viral loads were low. This characterization of HPV-107, -110 and -111 and FA75[KI88-03] expands the heterogeneity of members of species 2 of the genus Betapapillomavirus. However, as these types were found in only a few samples and in low amounts, a possible role in carcinogenesis remains elusive.

The healthy skin of two female domestic pigs (Sus scrofa domestica) was sampled with cotton-tipped swabs. Total genomic DNA was extracted from the samples and subjected to PCR with degenerate papillomavirus (PV)-specific primers. Similarity searches performed with blastn showed that partial E1 and L1 sequences of two novel PVs were amplified. Subsequently, the complete genomes of these Sus scrofa papillomaviruses (SsPVs) were amplified by long-template PCR, cloned and sequenced using a transposon insertion method. They contained the typical PV open reading frames (ORFs) E1, E2, E4, E6, L1 and L2, but the E7 ORF was absent in both viruses. Pairwise nucleotide sequence alignment of the L1 ORFs of the SsPVs showed 98.5 % similarity, classifying these viruses as SsPV type 1 ‘variants’ (SsPV-1a and -1b). Based on a concatenated alignment of the E1, E2, L1 and L2 ORFs of SsPV-1 variants a and b, and 81 other human and animal PV type species, a neighbour-joining phylogenetic tree was constructed. This phylogenetic analysis showed that the SsPV-1a and -1b variants did not cluster with the other PVs of artiodactyls (cloven-hoofed) host species, but clustered on the edge of the genus Alphapapillomavirus, very near to the root of this genus.

Porcine circovirus type 2 (PCV2) is divided into two genetic clusters designated PCV2a and PCV2b. The objectives of this study were to determine whether isolates from different clusters vary in virulence and to determine whether infection with PCV2a isolates induces protective immunity against subsequent infection with a recent PCV2b isolate. One-hundred and thirteen conventional specific-pathogen-free (SPF) pigs were assigned randomly to treatment groups and rooms: pigs inoculated with PCV2a cluster isolates (ISU-40895 or ISU-4838), pigs inoculated with PCV2b cluster isolates (NC-16845 or Can-17639) and uninoculated pigs. Necropsies were performed at 16 or 51 days post-inoculation (p.i.). There were no significant differences in PCV2-associated lymphoid lesions between PCV2a and PCV2b clusters; however, within the same cluster, significant differences were found between isolates: ISU-4838- and Can-17639-inoculated pigs had significantly (P<0.05) less severe lesions compared with ISU-40895- and NC-16845-inoculated pigs. To evaluate cross-protection, six pigs within each group were challenged at 35 days p.i. with an isolate from the heterologous cluster and were necropsied 51 days p.i. The severity of PCV2-associated lesions was reduced in pigs with prior exposure to an isolate from the heterologous cluster in comparison with singly inoculated pigs. Results indicate that the virulence of PCV2a and PCV2b isolates is not different in the conventional SPF pig model; however, the virulence of isolates within the same cluster differs. Increased virulence as reported to be associated with PCV2b isolates in the field was not observed under the conditions of this study. Moreover, cross-protection between PCV2a and PCV2b exists.

Baculovirus occlusion-derived virions (ODVs) contain a number of infectivity factors essential for the initiation of infection in larval midgut cells. Deletion of any of these factors neutralizes infectivity by the per os route. We have observed that P74 of the group I alphabaculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV) is N-terminally cleaved when a soluble form of the protein was incubated with insect midgut tissues under alkaline conditions and that cleavage was prevented by soybean trypsin inhibitor (SBTI). Presently, biological assays were carried out that suggest SBTI inhibits and trypsin enhances baculovirus per os infectivity. We developed a method to rescue per os infectivity of a P74 null virus involving co-transfection of viral DNA with a plasmid that transiently expresses p74. We used this plasmid rescue method to functionally characterize P74. A series of site-directed mutants were generated at the N terminus to evaluate if trypsin cleavage sites were necessary for function. Mutagenesis of R195, R196 and R199 compromised per os infectivity and rendered P74 resistant to midgut trypsin.