Canine Parvovirus Strains Research Articles

Rapid method utilizing the polymerase chain reaction for detection of canine parvovirus in feces of diarrheic dogs

By using primers based on the sequence of the VP2 gene of canine parvovirus (CPV), we established a rapid and specific assay for identification of the virus from fecal specimens based on the polymerase chain reaction (PCR). By use of a pair of primers, a specific 226-bp sequence was amplified by the PCR. All strains of CPV tested gave a specific amplification product by the PCR, while neither porcine parvovirus nor host cell did so. The PCR assay can detect fewer particles of CPV than the conventional methods, being able to detect CPV from fecal specimens in a rapid manner, provided that gel filtration of the samples through a spun column was done to remove inhibitory substances from the fecal specimens. These results suggest that the PCR assay can detect the presence of CPV in dogs early enough to prevent secondary infection by CPV in veterinary hospitals.

Detection by PCR of wild-type canine parvovirus which contaminates dog vaccines.

A method for detecting wild-type canine parvovirus (CPV) strains which contaminate vaccines for dogs has been developed by PCR. PCR primers which distinguish vaccine strains from the most common, recent strains of wild-type CPV in many countries, including Japan and the United States, were developed. This PCR is based on the differences in nucleotide sequences which determine the two antigenic types of this virus. CPV vaccine strains derived from antigenically old-type virus prevalent in former times were not detected by PCR with differential primers. Detection sensitivity of PCR was 100- to 10,000-fold higher than that of the culture method in Crandell feline kidney cells.

The emergence and evolution of canine parvovirus—an example of recent host range mutation

Canine parvovirus (CPV) emerged in 1978 an a new pathogen of dogs, which spread around the world and now appears endemic in the domesticated and wild dog populations in all countries. CPV is over 98% identical in DNA sequence to viruses which had been known for many years in cats, mink and raccoons, and genetic analysis has revealed that the differences in canine host range are determined by a small number of changes in the capsid protein gene. Comparison of the atomic structures of the CPV and FPV capsids shows that the changes affecting host range and virus-specific antigenic sites are exposed on the capsid surface in three different positions within a raised region at the threefold axis of symmetry, which is also the site of major antigenic determinants on the capsid. Three types of CPV have been defined by antigenic analysis with monoclonal antibodies. The original CPV strain (called CPV type-2) was only present in nature for a few years, and by 1981 it had been largely replaced in nature by a variant of CPV (CPV type 2a), which in turn replaced between 1984 and 1990 by a further variant (CPV type-2b). Those viruses differed by less than 0.2% of their genome sequences, but in each case the replacement apparently occurred on a global scale. The true ancestry of CPV is not clear, but the apparent emergence of the new types of CPV and its subsequent evolution suggest that this is a useful model for the emergence of new viruses with extended host ranges and their continuing adaptation.

Two Dominant Neutralizing Antigenic Determinants of Canine Parvovirus Are Found on the Threefold Spike of the Virus Capsid

The 25-nm diameter parvovirus capsid is assembled from 60 copies of a sequence common to the overlapping VP1 and VP2 proteins. Here we examine the epitope specificity's of 28 monoclonal antibodies (MAb) prepared against canine parvovirus (CPV), feline panleukopenia virus (FPV), and raccoon-dog parvovirus or blue (Arctic) fox parvovirus. Comparing the reactivity of those MAb with various MAb-selected escape mutants, or with natural variants of CPV or mink enteritis virus (MEV) which differ at known sequences, showed that the binding of 20 of those MAb was strongly affected by variations of two regions on the threefold spike of the CPV capsid. One region was adjacent to the tip of the threefold spike, and the second was around VP2 residue 300, on the shoulder of that structure. MAb recognizing both antigenic sites efficiently neutralized the virus infectivity and inhibited hemagglutination. Mutations leading to natural antigenic variation have also been observed in both those sites in naturally variant strains of CPV or MEV, suggesting that they are important antigenic structures on these parvoviruses. The bindings of several MAb were not affected by the mutations at those antigenic sites, indicating that they recognized other, and perhaps conserved, structures.

Construction of an infectious DNA clone of the Y1 strain of canine parvovirus and characterization of the virus derived from the clone.

We have cloned genome fragments of canine parvovirus strain Y1 from replicative-form DNA and double-stranded DNA synthesized from virion DNA in vitro, and constructed a recombinant plasmid containing a full-length Y1 genome (pCPVY 1). When this recombinant plasmid was transfected into cell cultures, an infectious virus could be recovered. To characterize this pCPVY 1-derived virus, its biological properties were compared with those of the parental strain. No difference was observed between them in antigen expression, viral DNA replication, hemagglutination ability, and virus multiplication, indicating that the virus derived from the infectious plasmid inherited the biological properties of the authentic Y1 strain. Therefore, this recombinant plasmid appears to be useful for reverse genetics of canine parvovirus.

Nucleotide sequence of feline panleukopenia virus: comparison with canine parvovirus identifies host-specific differences

The nucleotide sequence of feline panleukopenia virus (FPV) strain 193 was determined and compared with the sequence of canine parvovirus (CPV) strain N and partial sequences of FPV strain Carl and CPV strain b. Base differences were identified at 115 positions in these 5.1 kb genomes and predicted amino acid differences occurred at 40 positions. The two overlapping capsid protein genes contained almost twice as many base differences as the single non-structural protein gene (49 compared to 26) and about the same ratio was calculated for predicted amino acid differences (27 compared to 13). The 27 variant amino acids in the capsid proteins were clustered at three sites in the primary sequence, whereas 10 of the 13 variant amino acids in the non-structural protein occurred in the 130 C-terminal amino acids. The two FPV strains differed consistently from the two CPV strains at 31 bases: 12 base changes in the capsid protein genes resulted in six amino acid changes, six base changes in the non-structural protein gene resulted in three amino acid changes, and 13 base changes occurred in the non-coding sequence.

Comparison of feline parvovirus subspecific strains using monoclonal antibodies against a feline panleukopenia 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.

Canine parvovirus: strain difference in haemagglutination activity and antigenicity.

Canine parvovirus (CPV) strains were compared for haemagglutination (HA) activity and antigenicity and were divided into two types by HA activity. Strains Cp49 and 29-F showed temperature-dependent HA, like feline panleukopenia virus (FPLV) and mink enteritis virus (MEV), whereas strains Sp-80 and Y-1 showed temperature-independent HA with erythrocytes from eight species of animals. The results of a cross HA inhibition test using immunized rat sera suggested that of the two types of CPV those showing temperature-dependent HA were antigenically like FPLV and MEV whereas those showing temperature-independent HA were not. This antigenic difference between the two types was confirmed by a HA inhibition test with monoclonal antibodies. A chronological survey revealed that CPV isolates from earlier years have HA activity and antigenicity similar to those of FPLV and MEV, whereas current CPV isolates do not. There are some exceptional isolates from the transitional period which have similar antigenicity to FPLV and MEV but different HA activity. These results suggest that the haemagglutinin of CPV altered from one form resembling that of FPLV to a somewhat different structure during passage in dogs in nature.

New canine parvo virus strain 154 and use in vaccine effective in young animals in the presence of maternal antibodies

New canine parvo virus strain 154 and use in vaccine effective in young animals in the presence of maternal antibodies

Failure of vaccine or virulent strains of canine parvovirus to induce immunosuppressive effects on the immune system of the dog.

It is generally believed that canine parvovirus (CPV) is immunosuppressive and that immunosuppression plays an important role in the pathogenesis of canine parvovirus disease. However, there is little experimental evidence to support this assertion. The effects of virulent and vaccine strains of CPV on natural effector (NE) cell activity, antibody-dependent cell-mediated cytotoxicity (ADCC), chemiluminescence (CL) of neutrophils, lymphocyte response to mitogen, specific antibody response to CPV, and dermal reaction to phytohemagglutinin (PHA) were examined. Neither a virulent nor a vaccine strain of CPV significantly suppressed the immune system. The only potential effect noted was that dogs infected with the virulent strain of CPV developed a transient peripheral blood leukopenia. It was concluded that CPV does not cause immunosuppression and that immunosuppression does not play a significant role in the pathogenesis of canine parvovirus disease.

An improved hemagglutination test for study of canine parvovirus

Optimal conditions for hemagglutination (HA) by canine parvovirus (CPV) strains were investigated using several buffers. Porcine erythrocytes often agglutinated spontaneously in phosphate-buffered salt solution, isotonic saline solution or barbitone-complement-fixation buffer. Results were reproducible when borate-buffered saline (BBS) was used as the diluent for antigen, and “virus adjusting diluent” (VAD), containing 0.15 M NaCl and 0.3 M phosphate was used as the diluent for erythrocytes. Highest HA titers were obtained at pH 6.0 using BBS and VAD. Specific HA with CPV was observed not only at 4°C but at 37°C, and erythrocytes from horse, shrew mouse, hamster, cat, sheep and dog, as well as pig and African green monkey were agglutinated by CPV using the improved method.