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Abstract: Sequence analysis of the murine gamma-herpesvirus 68 (γHV68) genome previously identified several open reading frames (ORFs) located at the left end of the viral genome that do not share homology with other known herpesvirus or cellular genes. Here, we show that one of these ORFs, M4, encodes a secreted glycoprotein that influences the establishment of splenic latency at early times post-infection. We generated a mutant virus containing a premature translation termination codon in the M4 ORF (M4.STOP), and demonstrated that this mutant virus replicates in vitro equivalent to wild type and marker rescue (M4.MR) viruses. M4.STOP was also capable of high-titer lytic replication in vivo, but at 16 days post-infection the establishment of latency in the spleen was significantly impaired. The defect in the establishment of splenic latency was apparent following either intranasal or intraperitoneal inoculation. In contrast, the M4.STOP mutant did not exhibit a defect in the establishment of latency in peritoneal cells. These results suggest that M4 mediates an extracellular host-pathogen interaction that impacts the establishment of latent infection in the spleen, but not the peritoneum. [Copyright &y& Elsevier]

Shope fibroma virus and myxoma virus encode proteins predicted to be Type II photolyases. These are enzymes that catalyze light-dependent repair of cyclobutane pyrimidine dimers (CPDs). When the Shope fibroma virus S127L gene was expressed in an Escherichia coli strain lacking functional CPD repair pathways, the expressed gene protected the bacteria from 70–75% of the ultraviolet (UV) light-induced cytotoxic DNA damage. This proportion suggests that Leporipoxvirus photolyases can only repair CPDs, which typically comprise ∼70% of the damage caused by short wavelength UV light. To test whether these enzymes can protect virus genomes from UV, we exposed virus suspensions to UV-C light followed by graded exposure to filtered visible light. Viruses encoding a deletion of the putative photolyase gene were unable to photoreactivate UV damage while this treatment again eliminated 70–90% of the lethal photoproducts in wild-type viruses. Western blotting detected photolyase protein in extracts prepared from purified virions and it can be deduced that the poxvirion interior must be fluid enough to permit diffusion of this ∼50-kDa DNA-binding protein to the sites where it catalyzes photoreactivation. Photolyase promoters are difficult to categorize using bioinformatics methods, as they do not obviously resemble any of the known poxvirus promoter motifs. By fusing the SFV promoter to DNA encoding a luciferase open reading frame, the photolyase promoter was found to exhibit very weak late promoter activity. These data show that the genomes of Leporipoxviruses, similar to that of fowlpox virus, encode catalytically active photolyases. Phylogenetic studies also confirmed the monophyletic origin of poxviruses and suggest an ancient origin for these genes and perhaps poxviruses. [Copyright &y& Elsevier]

Abstract: Cynomolgus macaques have been widely used as an animal model in preclinical biomedical research and are becoming more popular among HIV/SIV vaccine researchers. Here we report the isolation and characterization of a cytomegalovirus from cynomolgus macaques (CyCMV). CyCMV was isolated from a healthy captive-bred 4-year-old cynomolgus macaque of Filipino origin. The virus was identified by its characteristic growth properties in cell culture, ultrastructural morphology and sequence of viral DNA polymerase and glycoprotein B (gB). CyCMV gB shows 77% identity and 88% homology to rhesus cytomegalovirus (RhCMV) gB and 58% identity and 76% homology to human cytomegalovirus gB at the amino acid level. Phylogenetic analysis using known CMV gB protein sequences show that CyCMV is more closely related to RhCMV than to other primate CMVs. CyCMV down-regulates MHC class I expression on infected cells and we show that the colony-bred cynomolgus macaques have detectable CyCMV-specific humoral and cell-mediated immune responses. [Copyright &y& Elsevier]