Canine parvovirus (CPV), a member of the genus Parvovirus, family Parvoviridae, is a significant causative agent in Canine reproductive failure, causing serious economic losses in the pet industry. The major capsid protein, VP2 is the main target protein for neutralizing antibodies in CPV. When VP2 was expressed using baculovirus, it was produced abundantly and assembled into virus-like particles (VLPs) similar in size and morphology to the original virions. It was named as CPV-VLP. Additionally, p35 sequences of canine distemper virus (CDV) as T-helper epitope were fusion-expressed with each of N-term, C-term or both sides of CPV-VP2. Production of double antigenic recombinant protein and formation of VLPs were analyzed by SDS-PAGE and transmission electron microscopy, respectively.

A female wild raccoon dog was referred with a history of generalized seizure. Mild leukocytosis was noted on laboratory tests. Gross lesions included nasal hemorrhage, hemothorax, and hemorrhage in the urinary bladder with hematuria. Microscopically, interstitial and purulent bacterial pneumonia was observed in the lungs. In the cerebellum, characteristic eosinophilic intracytoplasmic and intranuclear inclusion bodies were found in Purkinje cells, and severe demyelination was observed in the cerebellar white matter. Canine distemper virus (CDV) infection was suspected and confirmed after detection of CDV nucleoprotein RNA in the cerebrum and the lungs by nested reverse transcription polymerase chain reaction (RT-PCR) and reverse transcription loop-mediated isothermal amplification (RT-LAMP). Based on the histopathological and molecular diagnostic findings, it was concluded that the raccoon dog was infected with CDV.

Kidney cells of canine embryos were separated into single cells using collagenase and dispase. Primary culture was conducted using these cells. To remove fibroblasts, these cells were treated with edetate disodium dihydrate (Na2EDDA), and pure epithelial cells were separated. Recombinant retrovirus particles that manifest teromerase were produced and inoculated into primary culture cells to produce immortalized canine cell strains (JNUCK-1 and JNUCK-2). To examine the characteristics of the produced cell strains, the growth curve, maximum cultured households, and expressed proteins (keratin) were identified. The JNUCK-1 and JNUCK-2 cell lines showed division ability until the 30th generation without growth retardation. JNUCK-1 and JNUCK-2 cell lines clearly expressed telomerase until the 25th generation. The canine distemper virus (CDV) was inoculated into the JNUCK-1 and JNUCK-2 cell lines, as well as in the Madin–Darby canine kidney (MDCK) cell line. The maximum titer of CDV from the JNUCK-1 cell strain was about 200 times higher than that from the MDCK cell strain. However, the JNUCK-2 cell strain produced a lower titer than the MDCK cell strain. We established a new canine kidney epithelial cell line (JNUCK-1) that could produce CDV with high titer.

Canine distemper virus (CDV) causes serious and often fatal disease in dogs. Currently, various cells or cell lines have been used to detect or produce CDV. In order to set up the conditions, we separated two different cell lines from Madin-Darby canine kidney (MDCK) and named as MDCK-F (fibroblast-like) and MDCK-E (epithelial-like) by Na2EDDA treatment. CDV seed virus was prepared using MDCK cells and inoculated into MDCK-F and MDCK-E including MDCK with various ranges of multiplicity of infection (MOI) to confirm the optimal amount of virus inoculation. The virus titer of TCID50/ml was calculated by inoculation of serially diluted virus into 96-well plate of MDCK cells. The titer and cytopathic effect (CPE) in MDCK-E were compared to those in MDCK-F. The titer of seed CDV was 1.24×106 TCID50/ml. Optimal MOI was about 0.1 for both MDCK-F and MDCK-E to obtain highest titers of 108 TCID50/ml and 5 × 108 TCID50/ml respectively. CPE in MDCK-E was shown 4 days after inoculation whether in MNCK-F 5 6 days after inoculation. We can obtain highest titer of 5 × 108 TCID50/ml with 0.1 MOI using MDCK-E. MDCK-E was more susceptible for CDV production than MDCK-F.