Archives of Virology

The complete genome sequence of a novel mononegavirus, Lepeophtheirus salmonis negative-stranded RNA virus 1 (LsNSRV-1), obtained from a salmonid ectoparasite, Lepeophtheirus salmonis was determined. The viral genome contains five open reading frames encoding three unknown proteins (ORF I, II and III), a putative glycoprotein (G), and a large (L) protein. Phylogenetic analysis placed LsNSRV-1 in the recently established mononegaviral family Artoviridae. LsNSRV-1 showed a prevalence of around 97% and was detected in all L. salmonis developmental stages. Viral genomic and antigenomic RNA was localized to nerve tissue, connective tissue, epithelial cells of the gut, subepidermal tissue, exocrine and cement glands, as well as the testis, vas deferens and spermatophore sac of male L. salmonis and the ovaries and oocytes of females. Viral RNA was detected in both the cytoplasm and the nucleoli of infected cells, and putative nuclear export and localization signals were found within the ORF I, III and L proteins, suggesting nuclear replication of LsNSRV-1. RNA interference (RNAi) was induced twice during development by the introduction of a double-stranded RNA fragment of ORF I, resulting in a transient knockdown of viral RNA. A large variation in the knockdown level was seen in adult males and off springs of knockdown animals, whereas the RNA level was more stable in adult females. Together with the localization of viral RNA within the male spermatophore and female oocytes and the amplification of viral RNA in developing embryos, this suggests that LsNSRV-1 is transmitted both maternally and paternally. Small amounts of viral RNA were detected at the site where chalimi were attached to the skin of Atlantic salmon (Salmo salar). However, as the RNAi-mediated treatment did not result in LsNSRV-1-negative offspring and the virus failed to replicate in the tested fish cell cultures, it is difficult to investigate the influence of secreted LsNSRV-1 on the salmon immune response.

The chemical structure of human serum low density lipoproteins (LDL) involved in the inhibition of the haemagglutination by Sindbis virus was studied. Separation of lipoproteins into their protein and lipid components demonstrated that the inhibiting activity is almost completely due to the lipid moiety. The treatment of LDL and extracted lipoprotein lipids by phospholipase A2, C and D produced different effects and in particular a marked increase in the HI titre by the action of phospholipase A2. Moreover treatment with glycosidases significantly reduced the HI titre of LDL, thus suggesting the importance of carbohydrate moiety in the inhibiting lipoprotein molecule.

We describe the sequence for the complete genome of spinach latent virus (SpLV). Comparisons of this genome with that of the only other complete genome described for a species within the genus Ilarvirus (citrus leaf rugose virus – CiLRV) indicate that while there are marked differences between the RNA 3 of the two viruses, their respective RNAs 1 and 2 share many similarities. However, the putative 2a protein of SpLV contains a C2H2 type “zinc finger”-like motif located towards the carboxy terminal of the protein which is absent in CiLRV and has not been reported for other members of the family Bromoviridae. A second open reading frame (2b), located at a similar position to that described for the cucumoviruses, occurs in the RNA 2 of both SpLV and CiLRV. The putative coat protein of SpLV is similar to that of citrus variegation virus (CVV) and asparagus virus 2 (AV-2), both members of subgroup 2 of the ilarviruses. We have subsequently demonstrated a serological relationship between SpLV and other viruses in subgroup 2 and suggest that SpLV should be included in this subgroup rather than remain in a separate group (subgroup 6). However, while the putative movement protein of SpLV is remarkably similar to that of AV-2, it shows little relationship with the corresponding protein of CVV and the lack of similarity suggests that a recombination event may have occurred in the past. The relationship between the genera Alfamovirus and Ilarvirus is discussed in the light of the data for the genome of SpLV and recently published information for other members of the genus Ilarvirus.

A search for the cause of the inactivation of the transfectivity of the RNA from poliovirions, in the absence of a protective agent such as L-histidine, revealed that the inactivation is associated with trace contamination with copper and with an impurity or impurities in the phenol used to release the RNA from the poliovirions. Cu2+ and the impurity(ies) interactin vitro to produce a proximate inactivator or inactivators of the RNA. Phenol free or nearly free of active impurity can be prepared by steam distillation. Light is not required for formation or for action of the proximate inactivator. Addition of L-histidine to RNA undergoing inactivation promptly stops the inactivation, probably by taking copper away from the proximate inactivator.

We evaluated the usefulness of dual priming oligonucleotide (DPO)-based multiplex PCR, Seeplex HBV Lami-DR assay (Seegene Institute of Life Sciences, Seoul, Korea), to detect lamivudine-resistant HBV mutants in a comparison with the use of TRUGENE™ HBV genotyping and restriction fragment mass polymorphism (RFMP). Sera from 44 chronic hepatitis B patients were analyzed for the presence of mutations at codons 180 and 204 by performing DPO-based multiplex PCR, RFMP, and TRUGENE. The overall concordance rate among the three assays was 40.9% (18/44). Concordance rates between multiplex PCR and RFMP or multiplex PCR and TRUGENE were 61.4% (27/44) and 50.0% (22/44), respectively. In ten patients, multiplex PCR identified additional mutants not found using the other two methods. DPO-based multiplex PCR is a highly sensitive method to identify minor mutant populations and could be a practical tool in the monitoring of lamivudine resistance.

The role of the humoral immune response in clearance or prevention of canine distemper viral encephalitis of dogs infected with a virulent strain of canine distemper virus has been evaluated. Dogs that have demyelinating lesions, CDV proteins and infectious virus in their brains demonstrate an impaired humoral immune response. In dogs that recover from infection and contain no demyelinating lesions, viral proteins or infectious virus in the brain, antibodies to the internal proteins of CDV are observed early after infection. Later antibodies to primarily the H protein are detectable in sera of these dogs and the appearance of antibodies against the surface glycoprotein (H) correlates with the absence of lesions, CDV antigen and infectious virus in the brains of these dogs. Very late after infection immunoprecipitating antibody to all CDV antigens diminished rapidly so that at about ten weeks post infection antibodies that precipitate CDV antigens are barely detectable.