Abstract. An outbreak of canine parvovirus type 2 infection caused by the Glu-426 mutant in 2 litters of pups is reported. The infected pups (n ϭ 6) were monitored daily for evidence of clinical signs and hematological changes and for the evaluation of viral shedding in the feces. The disease induced by the Glu-426 mutant was mild in all the infected pups. Vomiting and hemorrhagic diarrhea were not observed; however, the pups developed mucoid diarrhea (3.5 median days), depression (1.5 median days), and relative leukopenia and lymphopenia (2.5 median days). Fever and loss of appetite were observed only in 2 pups. Virus was detected in the feces for 4.5, 6.5, and 46 median days by hemagglutination, virus isolation on cell cultures, and real-time polymerase chain reaction (PCR), respectively. By real-time PCR, the highest viral DNA titers were detected in the feces of both litters at day 10, reaching median values of more than 10 10 DNA copies/mg of feces.
Two genotype-specific fluorogenic RT-PCR assays were developed for the detection and quantitation of canine coronavirus (CCoV) type I and type II RNA in the faeces of dogs with diarrhoea. Both the fluorogenic assays showed high specificity, sensitivity and reproducibility, allowing a precise quantitation of CCoV type I and type II RNA over a linear range of about eight orders of magnitude (from 10(1) to 10(8) copies of standard RNA). Comparison with genotype-specific gel-based RT-PCR assays revealed that the fluorogenic assays were more sensitive and more rapid than conventional amplifications, with a large increase in throughput. The genotype-specific fluorogenic assays were then used to detect and measure viral loads in the faecal samples collected from dogs naturally or experimentally infected with type I, type II, or both genotypes. Of 174 samples collected from naturally infected dogs, 77 were positive for CCoV type I and 46 for CCoV type II. Thirty-eight dogs were found to be infected naturally by both genotypes, with viral RNA titres generally higher for type I in comparison to type II. At the same time, dogs infected experimentally shed type I RNA with higher titres with respect to type II.
A mammalian orthoreovirus (MRV) strain was isolated from a pup with fatal diarrhea, which had a concurrent infection by canine parvovirus type 2. The reovirus isolate showed an atypical hemagglutination pattern and a retarded electrophoretic mobility of the S1 segment, which is characteristic of MRV type 3 (MRV-3). Assignment of the isolated virus to MRV-3 was confirmed by type-specific RT-PCR assays, targeting the S1 gene, and by subsequent sequence analysis of the PCR product. By phylogeny based on the S1 gene of several MRVs, the isolate fell into lineage E, along with the murine strain T3C9/61 and the bovine strains T3C18/61 and T3C31/59. Conversely, L1 sequences were found to segregate regardless of the viral type. A total of 110 fecal samples, 56 nasal and 31 ocular swabs from dogs with diarrhea or nasal/ocular discharge were tested by a nested-PCR assay specific for reoviruses, and no sample was found to contain MRV RNA, a finding that is apparently in contrast with the seroprevalence (25.77%) observed in dogs.
Funding informationFondazione CARIPLO -Misura a sostegno dello sviluppo di collaborazioni per l'identificazione di terapie e sistemi di diagnostica, protezione e analisi per contrastare l'emergenza Coronavirus e altre emergenze virali del future;
Coronavirus disease 2019 patients in earlier stages exhaled millions of severe acute respiratory syndrome coronavirus 2 per hour. Clin Infect Dis. 2020;ciaa1283.
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