A canine parvovirus recovered from a vaccinated five-month-old Poodle in Chengdu, Sichuan Province, has been sequenced and classified as the CPV-2c genotype, according to a study published in Frontiers in Veterinary Science. The isolate, designated MDC-CPV, was obtained from an anal swab collected on March 28, 2026, from a male Toy/Miniature Poodle that had received three doses of a canine tetravalent vaccine covering distemper virus, parainfluenza virus, adenovirus, and parvovirus between December 2025 and February 2026. The dog developed vomiting, depression, weight loss, and leukopenia, though no overt diarrhea was observed, and all antibody tests performed during the vaccination period had yielded positive results.
The research team, led by investigators from West China Hospital of Sichuan University, Southwest Minzu University, and Chengdu Zhonghuan Animal Hospital, screened the clinical sample for a panel of common canine pathogens using PCR or reverse transcription PCR. Only canine parvovirus type 2 produced an amplification band at the expected target size, while no definitive positive results were obtained for canine coronavirus, distemper virus, adenovirus types 1 and 2, parainfluenza virus, herpesvirus 1, Bordetella bronchiseptica, or pathogenic Leptospira species. A commercial colloidal gold rapid immunochromatographic strip also indicated CPV antigen positivity, providing the basis for subsequent whole-genome sequencing and in vitro infection experiments.
Sequencing was performed on the Illumina NovaSeq platform using a paired-end PE150 strategy, generating approximately 12.7 million raw reads corresponding to roughly 1.9 billion bases. After quality filtering, adapter trimming, and removal of rRNA, host-derived, and bacterial contaminating sequences, de novo assembly with SPAdes and SOAPdenovo produced a principal CPV-associated contig of 5,358 nucleotides with no ambiguous bases. The assembled genome showed an average sequencing depth of 52.94-fold, a median depth of 53-fold, and complete nucleotide coverage, with 98.99 percent of positions covered at least tenfold. Genome annotation confirmed the presence of the typical CPV coding regions, including the non-structural proteins NS1 and NS2 and the structural capsid proteins VP1 and VP2.
The closest BLASTn match for the assembled contig was canine parvovirus strain CPV-SH1516, isolated in Shanghai in 2017, sharing 99.812 percent nucleotide identity over a 4,795-base-pair alignment. A broader homology search against the NCBI database identified the top 30 closely related sequences as primarily CPV-2c or related canine parvovirus strains reported from China and other East Asian regions, with the highest nucleotide identity reaching approximately 99.81 percent. These results place MDC-CPV in close genomic relationship to recently circulating CPV-2c strains at the whole-genome level.
Genotyping based on key amino acid residues in the VP2 capsid protein supported classification as CPV-2c. The VP2 coding sequence spans 1,755 nucleotides and encodes a protein of 584 amino acids. Residue 426 of VP2, a site commonly used to distinguish the CPV-2a, CPV-2b, and CPV-2c antigenic variants, carries glutamic acid in MDC-CPV, the defining marker for CPV-2c. The virus also carries alanine at position 297 and a combination of additional residues including leucine at 87, threonine at 101, glycine at 300, tyrosine at 305, isoleucine at 324, aspartic acid at 375, valine at 555, serine at 564, and glycine at 568. Comparison with representative CPV-2, CPV-2a, and CPV-2b reference sequences confirmed that MDC-CPV shares the CPV-2c typing profile at these key positions.
Phylogenetic analysis using maximum-likelihood methods on the full-length VP2 coding sequence, with feline panleukopenia virus sequences as an outgroup, placed MDC-CPV firmly within the CPV-2c clade. The virus clustered most closely with sequence PZ569219.1 in both the whole-genome and VP2 trees, with strong branch support values of 75.6/90 and 84/97 for SH-aLRT and ultrafast bootstrap, respectively. In the NS1 tree, the nearest sequence was PZ529563.1, though this association lacked statistical support. Recombination analysis using seven independent algorithms on a panel of MDC-CPV and its 40 closest non-identical reference sequences found no well-supported recombination event, as no consistent breakpoint region was corroborated by at least three methods.
To evaluate infectivity outside the host, the researchers inoculated the processed sample and passaged viral supernatants into two cell lines commonly used for parvovirus work: A72 cells, derived from canine tumor tissue, and CRFK cells, derived from feline kidney. Compared with mock-infected controls, cells inoculated with MDC-CPV exhibited cytopathic effects including cell rounding, detachment, and monolayer destruction. Western blot analysis detected expression of CPV VP2-related proteins in infected cells but not in controls, and immunofluorescence assays confirmed the presence of CPV antigen-positive signals within infected cells. Transmission electron microscopy of supernatants from infected CRFK cells revealed virus-like particles within the size range typical of parvovirus-compatible particles.
Quantitative PCR targeting a conserved region of the viral NS gene showed that viral DNA levels remained relatively low during the first 24 hours post-infection but increased markedly from 48 hours onward, with a progressive rise at 72, 96, and 120 hours, while mock-infected controls remained at background levels throughout. Infectious virus titers, measured by TCID50 assay using the Reed–Muench method, followed a similar trajectory: titers were low during the first 24 hours, rose sharply to approximately 4.5 log10 TCID50/mL at 48 hours, and reached approximately 5.3 and 5.8 log10 TCID50/mL at 72 and 96 hours, respectively. The titer remained high at 120 hours with only a slight decrease from the 96-hour peak. All experiments were performed independently three times.
The authors note that the parallel increase in viral DNA and infectious titers demonstrates that MDC-CPV undergoes productive replication in CRFK cells and produces infectious progeny during the 120-hour observation period. They also observed that viral DNA levels continued to rise toward 120 hours even as infectious titer peaked near 96 hours, a divergence they attribute tentatively to continued accumulation of viral genomic DNA and noninfectious particles after infectious virion production plateaus, or to cytopathic damage and depletion of viable proliferating target cells at later stages. The authors emphasize that distinguishing among these possibilities would require additional experiments separately quantifying intracellular and extracellular viral DNA, infectious virions, and viable cell numbers.
The study’s authors caution that detection of a field CPV-2c strain in a vaccinated animal should not be interpreted as direct evidence of vaccine failure or viral immune escape. Neutralizing antibody titers at the time of disease were unavailable, and the CPV component of the administered vaccine was not sequenced, so the current data cannot establish whether antigenic differences between MDC-CPV and the vaccine strain contributed to infection. Protection against CPV is influenced by multiple factors including age at vaccination, maternal antibody interference, completion and timing of the schedule, individual immune responses, viral exposure dose, and antigenic differences between vaccine and field strains. The authors also note that because MDC-CPV derives from a single clinical case, the study cannot independently estimate the prevalence of CPV-2c in Chengdu or Sichuan, though the findings align with prior surveillance indicating CPV-2c is an important component of the currently circulating CPV population in the region.
Subject of Research: Genomic characterization and in vitro infectivity of a canine parvovirus type 2c isolate from a vaccinated dog in Sichuan, China
Article Title: Genomic characteristics and preliminary study on in vitro infection properties of canine parvovirus type 2c derived from a canine anal swab sample
Article References: Mou, D., Chen, Y., Wu, S., Huang, J., Guo, Y., & Bai, X. (2026). Genomic characteristics and preliminary study on in vitro infection properties of canine parvovirus type 2c derived from a canine anal swab sample. Frontiers in Veterinary Science, 13, Article 1938079. https://doi.org/10.3389/fvets.2026.1938079
Image Credits: AI Generated
DOI: 10.3389/fvets.2026.1938079
Keywords: canine parvovirus, CPV-2c, whole-genome sequencing, VP2 genotyping, phylogenetics, in vitro infection, CRFK cells, A72 cells, vaccinated dog, Sichuan, TCID50 assay, molecular surveillance
News Source: William Thompson. (October 30, 2026). Canine parvovirus 2c genome recovered from vaccinated dog in Sichuan. Scienmag.
