Genomic characteristics and preliminary study on in vitro infection properties of canine parvovirus type 2c derived from a canine anal swab sample

Objective This study aimed to determine the genomic characteristics and genotype of a CPV isolate (MDC-CPV) recovered from a vaccinated dog, and to investigate its in vitro infectivity and replication features in A72 and CRFK cell lines. Methods Clinical samples were subjected to nucleic acid detection for common canine pathogens and CPV antigen testing. Sequencing was performed using the Illumina NovaSeq PE150 platform. Following quality control, assembly, and annotation, the near-complete genome sequence of MDC-CPV was obtained. Homology searches were conducted using BLASTn. Genotyping was determined based on key amino acid residues in VP2, and phylogenetic analyses were performed using both the full-length VP2 coding sequence (CDS) and the whole-genome reference sequences. In vitro infection was subsequently validated in A72 and CRFK cells. Viral infection-associated characteristics were evaluated by observing cytopathic effects (CPE), and through Western blot, immunofluorescence, qPCR, TCID50 assay, and transmission electron microscopy. Results The genome length of MDC-CPV was 5,358 nucleotides, with no ambiguous bases identified. Genome annotation revealed that MDC-CPV contains typical CPV coding regions, including NS1, NS2, VP1, and VP2. The VP2 CDS was 1755 nucleotides in length, encoding 584 amino acids. Key VP2 residues for MDC-CPV were 297Ala/426Glu, along with a combination of other residues (87Leu, 101Thr, 300Gly, 305Tyr, 324Ile, 375Asp, 555Val, 564Ser, and 568Gly), supporting its classification as CPV-2c. Phylogenetic analysis placed MDC-CPV within the CPV-2c clade, showing close genetic relatedness to several CPV-2c reference strains from China and other regions in East Asia. In in vitro infection assays, CPE was observed in both A72 and CRFK cells inoculated with MDC-CPV. Furthermore, CPV-related protein expression and virus-like particles were detected. Quantitative PCR and TCID50 assays in CRFK cells demonstrated productive replication, with viral DNA levels and infectious virus titers increasing substantially from 48 h post-infection, reaching peak titers of approximately 5.8 log10 TCID50/1 mL at 96 h post-infection. Conclusion MDC-CPV is a CPV-2c strain isolated from a clinical sample of a vaccinated dog. Its genomic and VP2 genotypic characteristics are consistent with recently circulating CPV-2c strains. These findings provide a reference for molecular epidemiological surveillance of CPV-2c in the Sichuan region and for research on CPV infection in the context of vaccination.