Spillover of Canine Parvovirus Type 2 to Pigs, South Dakota, USA, 2020

In 1978, canine parvovirus type 2 originated from spillover of a feline panleukopenia–like virus, causing a worldwide pandemic of enteritis and myocarditis among canids. In 2020, the virus was identified in pigs in South Dakota, USA, by PCR, sequencing, in situ hybridization, and serology. Genetic analysis suggests spillover from wildlife.

the species Carnivore protoparvovirus 1, which can cause severe disease in carnivores of many species (1)(2)(3). Besides CPV-2, which causes enteritis in dogs of all ages and myocarditis in puppies, the virus species includes feline panleukopenia virus, which causes severe enteritis and leukopenia in cats of all ages (4). In 1978, CPV-2 emerged and caused a worldwide pandemic after spillover from a feline panleukopenia virus-like virus in wildlife. Subsequent adaptation to canine hosts led to genetic and antigenic diversification into subtypes 2a, 2b, and 2c (5). Continued CPV host switching has been documented; spillover to wildlife (including skunks, raccoons, coyotes) has resulted in clinical disease and asymptomatic infection (2).
In October 2020, a dead pig was submitted to South Dakota State University (Brookings, SD, USA) for diagnostic testing. Histopathologic examination revealed mild to moderate enteritis, hepatitis, and visceral edema. Hemolytic Escherichia coli was isolated. No significant lung lesions were noted. Approximately 8 months later, we performed viral metagenomic sequencing on archived lung tissue for an unrelated research project and unexpectedly identified CPV-2. Using a 5′-nuclease PCR (Integrated DNA Technologies, https://www. idtdna.com), we confirmed that the sample was CPV-2 positive; cycle threshold (C t ) was 24.4. Sanger sequencing of overlapping amplicons confirmed the CPV-2 genome sequence determined by metagenomic sequencing. We submitted the strain SDS21601 sequence to GenBank (accession no. MZ666397).
We used a 5′-nuclease PCR to test 90 archived porcine lung samples submitted for respiratory disease diagnostic testing for CPV-2. Of the 90 samples, 9 (10%) were positive for CPV-2, including those with strain SDS21601, and C t values were 22.4-36.3. The samples were collected September-November 2020 from swine farms within 150 miles of Brookings. We sequenced the genome from a second strongly positive sample (C t 22.4) and submitted strain SDS21608 to GenBank (accession no. MZ666398). An amplicon from 4 of the remaining 7 samples positive by PCR was generated by PCR and confirmed as CPV-2 by Sanger sequencing. The 3 samples that failed to yield a CPV-2-specific amplicon had C t values >32. Sequence comparison showed 99.9% nt identity between SDS21601 and SDS21608. blastp (https://blast. ncbi.nlm.nih.gov) analysis of SDS21601 virus capsid protein (VP) 2 found 100% identity to CPV-2 from a coyote sampled in Montana in 2012. Analysis of the VP2 amino acid sequences identified an F212I mutation previously identified only from US wildlife, mainly coyotes.
We performed in situ hybridization on archived formalin-fixed paraffin-embedded tissues from SDS21608 by using a commercially available CPV-2 probe. CPV-2 nucleic acids were hybridized sporadically as intracytoplasmic punctate signals in few monocyte-macrophage lineage cells in the medullary and subcapsular sinuses of a bronchial lymph node (Figure). However, the primary anatomic site of CPV-2 infection and replication was not determined. In other examined tissues, we observed neither typical  virus-associated microscopic lesions as seen in carnivores nor obvious CPV-2 hybridization signals.
To further investigate the extent of CPV-2 circulation among swine, 8 months after collection of the CPV-2-positive lung tissue, we collected 20 serum samples from multiparous sows on the farm where strain SDS21601 originated. Of the 20 samples, 13 (65%) were positive for CPV-2-specific antibodies by hemagglutination inhibition (HI) assay; titers were 10-40 (Table). Nearly all serum samples (19 of 20) had antibody titers to porcine parvovirus 1 (PPV-1), ranging from 16 to 4,096. This result was expected given that pigs on the farm received commercial PPV-1 vaccine before farrowing. There was no correlation between CPV-2 and PPV-1 HI titers, indicating a lack of cross-reactivity between CPV-2 and PPV-1 antibodies in the HI assay.
To further investigate seroprevalence of CPV-2 in South Dakota, we randomly selected 25 sow serum samples from unrelated submissions collected at 5 farms (5 samples/farm) and analyzed them by HI for CPV-2. Of the 25 samples, 23 (92%) were positive for CPV-2; titers were 10-80. Together with the 10% positivity detected by quantitative PCR, these results suggest widespread CPV-2 infection of swine in South Dakota.
Members of Carnivore protoparvovirus 1 display >98% identity. Amino acid residue 300 of VP2 has been shown to be a critical determinant for the cross-species transfer of CPV-2 between carnivores of different species (6). Glycine 300 and tyrosine 305, observed in the VP2 of both swine CPV-2 strains (SDS21601 and SDS21608), are diagnostic of CPV-2 isolates from canids (7). The F212I mutation present in both swine CPV-2 strains, which was previously found only in wildlife, suggests a sylvatic origin. Of the species in which F212I has been identified, only coyotes are common in the agricultural areas of the upper US Midwest and are peridomestic. We hypothesize that the source of swine CPV-2 infection is CPV-2-positive coyote feces.
Our results demonstrate spillover of CPV-2 to swine. CPV-2 has been associated with severe enteritis in insectivorous Taiwanese pangolin (Manis pentadactyla pentadactyla), further demonstrating the propensity of CPV-2 to overcome host barriers (8). The ability of CPV-2 to cause disease in swine remains unknown; further surveillance is warranted because this spillover may threaten the health of swine herds.

RESEARCH LETTERS
To determine the extent of exposure to Zika virus (ZIKV) and chikungunya virus (CHIKV) in Jamaica, we collected serum from 584 pregnant women during 2017-2019. We found that 15.6% had antibodies against ZIKV and 83.6% against CHIKV. These results indicate potential recirculation of ZIKV but not CHIKV in the near future. 1 Preliminary results from this study were presented at the 2nd