Biological and physical comparison of mink enteritis virus feline panleukopenia virus and canine parvovirus.
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The growth characteristics were studied in the attenuated strains of canine parvovirus CPVA-BN 80/82, mink enteritis virus MEVA-BN 63/82 and feline panleucopenia virus FPVA-BN 110/83 on the stable feline kidney cell line FE, and in the attenuated canine distemper virus CDV-F-BN 10/83 on chicken embryo cell cultures (KEB) and cultures of the stable cell line VERO. When the FE cultures were infected with different parvoviruses in cell suspension at MOI 2-4 TKID50 per cell, the first multiplication of the intracellular virus was recorded 20 hours p. i. In the canine parvovirus, the content of intracellular and extracellular virus continued increasing parallelly until the fourth day; then, from the fourth to the sixth day, the content of extracellular virus still increased whereas that of intracellular virus fell rapidly. In the case of the mink enteritis virus the release of the virus into the culture medium continued parallelly with the production of the cellular virus until the sixth day. In the case of the feline panleucopenia virus the values concerning free virus and virus bound to cells were lower, starting from the second day p. i. When KEB or VERO cultures were infected in cell suspension with the canine distemper virus at MOI about 0.004 per 1 cell, the replicated intracellular virus was first recorded in the KEB cultures five hours after infection but in the VERO cultures only 20 hours after infection, with a timely release of the virus into the culture medium in both kinds of tissue. In the KEB and VERO cultures the highest values of infection titres were recorded on the fourth day p. i., the course of virus multiplication on the cells being parallel with its release into the culture medium.
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Canine parvovirus-2 (CPV-2), Mink enteritis virus (MEV) and feline panleukopenia virus (FPV) were produced using identical cell culture and purification techniques. The distributions of the haemagglutinating activity of the three different parvoviruses in a CsCl gradient were similar with haemagglutinating peaks identified at 1.48-1.49, 1.42, 1.36 and 1.30-1.31 g cm-3. The number and distribution of the viral proteins and the equivalent protein molecular weights are similar for all three viruses in SDS-polyacrylamide gels (10%). Four viral proteins were identified and their molecular weights were determined: protein A (77 500-79 500), protein B (63 000-63 500), protein C (61 500-63 000) and protein D (50 000-55 000). The viral protein D although reported for some other parvoviruses has not previously been demonstrated in CPV-2, MEV or FPV.
A dual infection by feline panleukopenia virus (FPV) and feline calicivirus (FCV) in a 7 month-old cat is described. The animal developed a severe illness with depression, anorexia, fever, leucopoenia, nasal and ocular discharge and oral ulcers. Both FPV and FCV were isolated in cell cultures from a rectal swab and the presence of FCV was confimed by polymerase chain reaction. Antibodies to both the viruses were detected in the serum. The severity of the disease induced by the mixed viral infection highlights the need for intensifying FPV vaccination in cats.
Forty-two seronegative cats received an initial vaccination at 8 weeks of age and a booster vaccination at 12 weeks. All cats were kept in strict isolation for 3 years after the second vaccination and then were challenged with feline calicivirus (FCV) or sequentially challenged with feline rhinotracheitis virus (FRV) followed by feline panleukopenia virus (FPV). For each viral challenge, a separate group of 10 age-matched, nonvaccinated control cats was also challenged. Vaccinated cats showed a statistically significant reduction in virulent FRV-associated clinical signs (P = .015), 100% protection against oral ulcerations associated with FCV infection (P < .001), and 100% protection against disease associated with virulent FPV challenge (P < .005). These results demonstrated that the vaccine provided protection against virulent FRV, FCV, and FPV challenge in cats 8 weeks of age or older for a minimum of 3 years following second vaccination.
The feline parvovirus subgroup is comprised of viruses isolated from various carnivores, including the dog, cat, mink, raccoon, Arctic fox, and raccoon dog. Those viruses are > 98% identical in their DNA sequences and are very similar antigenically. We have shown that although canine parvovirus (CPV) replicates in numerous feline cell lines in vitro it does not infect cats after parenteral inoculation (U. Truyen and C. R. Parrish, (1992) J. Virol. 66, 5399-5408). Here we use recombination mapping to locate some viral determinants required for feline host range, and show that the ability to replicate in cats was determined by the right-hand 45% of the genome, most likely a function of the capsid protein gene. Efficient replication in the cat appeared to require feline panleukopenia virus sequences from both ends of the VP2 molecule, which contained differences of VP2 amino acid residues 80, 564, and 568. The difference at amino acid 80 was also associated with expression of an FPV-specific antigenic epitope. The differences which affected the feline host range were located in a region of the capsid structure where three VP2 molecules interact, and the mutations gave rise to changes in the conformation of loops of the three adjoining VP2 monomers. The mechanism(s) of the in vivo feline host range restriction were not defined, and we were unable to show in vitro inhibition of virus infectivity by feline serum components or erythrocytes.
Parvoviruses from mink (mink enteritis virus [MEV]), cats (feline panleukopenia virus [FPV]), raccoons (raccoon parvovirus [RPV]), and dogs (canine parvovirus [CPV]) were compared. Restriction enzyme analysis of the viral replicative-form DNA revealed no consistent differences between FPV and RPV isolates, but CPV and MEV isolates could be distinguished readily from other virus types. Feline panleukopenia virus, RPV, and MEV, but not CPV, replicated to high titers in mink. However, on the first passage, disease and microscopic lesions were observed only in mink inoculated with MEV. Feline panleukopenia virus and RPV isolates replicated in ferrets, but disease or microscopic lesions were not observed. Feline panleukopenia virus and RPV isolates could be passaged repeatedly in mink and ferrets. Virulence of FPV and RPV isolates was low compared with that of MEV, and only a single mink inoculated with FPV or with RPV developed clinical disease on the sixth passage of virus.
Feline panleukopenia virus (FPV) and canine parvovirus (CPV) are autonomous parvoviruses which infect cats or dogs, respectively. Both viruses cause an acute disease, with virus replicating for less than seven days before being cleared by the developing immune responses. The viruses have a broad tropism for mitotically active cells. In neonatal animals the viruses replicate in a large number of tissues, and FPV infection of the germinal epithelium of the cerebellum leads to cerebellar hypoplasia, while CPV may infect the hearts of neonatal pups, causing myocarditis. In older animals the virus replicates systemically, primarily in the primary and secondary lymphoid tissues, and also in the rapidly replicating cells of the small intestinal epithelial crypts. A transient panleukopenia or relative lymphopenia is often observed after FPV or CPV infection, respectively. Whether the reduction in cell numbers in vivo is due to virus replicating in and killing cells, or due to other indirect effects, is not known. However, FPV kills both erythroid and myeloid colony progenitors in in vitro bone marrow cultures, and it has been suggested that virus replication in the myeloid cells in vivo could lead to the reduced neutrophil levels seen after FPV infection of cats.
In an attempt to distinguish feline panleukopenia virus (FPLV) live vaccine strains from FPLV field isolates in Japan, we compared restriction fragment length polymorphisms (RFLP) of polymerase chain reaction (PCR)-amplified fragments of live FPLV vaccine strains with those of FPLV Japanese field isolates. On the basis of nucleotide sequence differences between PLI-IV, a live vaccine strain, and FPV-483, a recent field isolate, two restriction enzymes, Dra I and Afa I, were selected for PCR-RFLP analysis of nucleotide (nt) differences at nt 3695 and 4508, respectively. Three live vaccine strains including the PLI-IV strain could be distinguished from the Japanese field isolates by their PCR-RFLP patterns by Afa I, but one live vaccine strain was indistinguishable from the Japanese isolates when Dra I and Afa I were used. The Japanese field isolates were divided into two groups by the profile of PCR-RFLP patterns generated by Dra I and Afa I, suggesting that PCR-RFLP analysis using several enzymes provides a good genetic estimate of strain differentiation. No isolate that shows a Dra I-negative/Afa I-negative pattern has emerged in Japan, indicating the possibility that the live vaccine viruses with a Dra I-negative/Afa I-negative pattern, such as the PLI-Iv strain, are candidates for use as live FPLV vaccine strain in Japan where they can be genetically distinguished from field strains.
Canine parvovirus (CPV), feline panleukopenia virus (FPLV) and mink enteritis virus (MEV) were compared serologically, by determination of their host range in cell cultures, as well as by restriction enzyme analysis. Maps of the virus genomes were established using seven different restriction enzymes cutting at a total of 56 sites. MEV and FPLV gave maps which were identical except for one restriction site. The map of CPV is closely related to those of FPLV/MEV since their DNAs share about 80% of the restriction sites tested. However, CPV is clearly distinct from FPLV/MEV since either eight (German isolate) or nine (Belgian, Swiss and American isolates) restriction sites are different. The DNAs of six vaccine strains of FPLV and MEV were also analysed. They gave maps which closely resembled those of the respective wild-type strains. CPV and FPLV/MEV also differed with respect to antigenicity, as well as to host range in cell cultures.
It was recently reported that canine parvoviruses (CPV) had entered cat populations and induced disease in infected cats, while they had affected only dogs in the past. It is important to determine whether conventional feline panleukopenia virus (FPLV) vaccines protect against recent CPV infections. In this study, the cross-reactivity of virus-neutralising (VN) and haemagglutinin-inhibition (HI) antibodies in cats induced by FPLV and CPV s were examined. Lower cross-reactivities of VN and HI antibodies against each CPV strain were observed in cats experimentally inoculated with FPLV or vaccinated with an inactivated FPLV vaccine. In addition, we revealed the existence of a novel type of FPLV, which reacted weakly with antibodies induced by the conventional FPLV vaccine.
Approximately 80% of the genome of feline panleukopenia virus was cloned into pBR322. This DNA included the transcription unit for the major viral mRNA species. The nucleotide sequence of the cloned portion of the genome was determined. Comparison of the feline panleukopenia virus sequence with the sequences of the parvoviruses minute virus of mice and H-1 revealed considerable homology between the three viruses on both the nucleic acid and protein levels. Based on this homology, a model for the generation of the two size classes of viral structural proteins (VP1 and VP2') is proposed.
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The NS1 and VP1/VP2 genes of canine parvovirus and feline panleukopenia virus were amplified by the polymerase chain reaction (PCR). The restriction cleavage patterns of the amplified DNA fragments were compared among these parvoviruses including vaccine strains. Differences of the restriction site in the NS1 portions were observed between the vaccine strain and the wild type strain as well as between canine parvoviruses and feline panleukopenia viruses. The restriction patterns of feline panleukopenia viruses were distinct from those of canine parvoviruses, and showed differences between the vaccine strain and its wild type strain. This PCR-based restriction cleavage can be used for ecological study of such viruses.
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Plaque formation with representative strains of feline panleukopenia virus (FPV) has been obtained using a permanent line of feline kidney cells under agarose overlay. FPV-infected cells appear as white plaques after neutral red staining. Plaque size is determined by the extent of cell division in the infected monolayer. FPV assay by the plaque procedure is rapid and gives infectivity titres which exceed those determined by the common inclusion body and immunofluorescent assays of FPV by a factor of about 100 and 10, respectively. Moreover, the plaque assay offers an effective means for the quantification of neutralizing antibodies in feline sera as well as for the detection of heat-stable substances in bovine sera which strongly interfere with replication of the virus.
Four monoclonal antibodies (mAb) against a feline panleukopenia virus (FPLV) TU 1 strain, one of the host range variants of feline parvovirus (FPV), were produced and applied for antigenic analysis of FPLV, canine parvovirus (CPV) and mink enteritis virus (MEV). All mAbs were considered to be directed at epitopes on the virus capsid surface because they neutralized the infectivity and inhibited the hemagglutination (HA) of the homologous virus as well as other FPV strains. They were of the mouse IgG1 type. High antigenic homogeneity among FPLV strains was confirmed by HA-inhibition (HI) test with the mAbs and polyclonal immune sera against FPLV or CPV. But the TU 11 strain of FPLV was antigenically distinguished from the remaining 14 FPLV strains by both the HI test and the micro-neutralization test with one of the mAbs produced. MEV Abashiri strain was found to be antigenically indistinguishable from FPLV. Most of the CPV strains isolated after 1981 were considered to be antigenically different from earlier CPV isolates when some mAbs were applied in the serological tests, confirming the replacement of CPV by an antigenic variant in Japan. However, antigenically different CPVs were detected at the end of 1984 from unrelated epizootics occurred a month apart in the same area.