Sequence analysis of divergent canine coronavirus strains present in a UK dog population (original) (raw)
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Journal of Clinical Microbiology, 2001
A nested PCR (nPCR) assay for the detection of canine coronavirus (CCV) in fecal samples is described. The target sequence for the assay was a 514-bp fragment within the spike (S) glycoprotein gene. The sensitivity of the assay is extremely high, detecting as little as 25 50% tissue culture infective doses per g of unprocessed feces. A clinical trial using dogs challenged orally with CCV SA4 and CCV NVSL was used to compare viral isolation and the nPCR assay as detection techniques over a 2-week period of infection. Virus isolation detected CCV shedding from day 4 to 9 postchallenge, while the nPCR assay detected CCV shedding from day 4 to 13 postchallenge. Cloning and sequencing of the nPCR assay product enabled investigation of the evolutionary relationships between strains within the S gene. The simple and rapid procedure described here makes this assay an ideal alternative technique to electron microscopy and viral isolation in cell culture for detection of CCV shedding in feces...
Journal of Virology, 2009
Four canine coronavirus type II (CCoV-II) strains were identified in the guts and internal organs of pups which had died of acute gastroenteritis. The CCoV-II strains were strictly related to porcine transmissible gastroenteritis virus (TGEV) in the N-terminal domain of the spike protein, whereas in the other parts of the genome, a higher genetic relatedness to recent CCoV-II isolates was observed. Experimental infection of dogs with a TGEV-like isolate induced mild gastroenteritis without any systemic involvement. By virus neutralization tests, antigenic differences between reference and TGEV-like CCoVs were found. Our data support the potential recombinant origin of the TGEV-like CCoVs.
Journal of clinical microbiology, 2014
Emerging canine coronavirus (CCoV) variants that are associated with systemic infections have been reported in the European Union; however, CCoV-associated disease in the United States is incompletely characterized. The purpose of this study was to correlate the clinicopathological findings and viral antigen distribution with the genotypic characteristics of CCoV in 11 puppies from nine premises in five states that were submitted for diagnostic investigation at Cornell University between 2008 and 2013. CCoV antigen was found in epithelial cells of small intestinal villi in all puppies and the colon in 2 of the 10 puppies where colon specimens were available. No evidence of systemic CCoV infection was found. Comparative sequence analyses of viral RNA extracted from intestinal tissues revealed CCoV-II genotype in 9 out of 11 puppies. Of the nine CCoV-IIs, five were subtyped as group IIa and one as IIb, while three CCoVs could not be subtyped. One of the CCoV-IIa variants was isolated ...
Isolation and sequence analysis of canine respiratory coronavirus
Virus Research, 2007
Canine respiratory coronavirus (CRCoV) has frequently been detected in respiratory samples from dogs by RT-PCR. In this report the first successful isolation of CRCoV from a dog with respiratory disease is described. The isolate CRCoV-4182 was cultured in HRT-18 cells but failed to replicate in a number of other cell lines. The nucleotide sequence of the 3-terminal portion of the CRCoV genome was determined including all open reading frames from the NS2 gene to the N gene. Comparison with other coronavirus sequences showed a high similarity to bovine coronavirus (BCoV). The region between the spike and the E gene was found to be the most variable and was used for phylogenetic analysis of several CRCoV strains. CRCoV-4182 showed a mutation within the non-structural protein region downstream of the S gene leading to the translation of an 8.8 kDa putative protein comprising a fusion of the equivalent of the BCoV 4.9 kDa protein to a truncated version of the BCoV 4.8 kDa protein. The culture of CRCoV will enable analysis of the expression and function of this and other CRCoV proteins as well as allowing the study of the role of CRCoV in the aetiology of canine infectious respiratory disease.
Canine Enteric Coronaviruses: Emerging Viral Pathogens with Distinct Recombinant Spike Proteins
Viruses, 2014
Canine enteric coronavirus (CCoV) is an alphacoronavirus infecting dogs that is closely related to enteric coronaviruses of cats and pigs. While CCoV has traditionally caused mild gastro-intestinal clinical signs, there are increasing reports of lethal CCoV infections in dogs, with evidence of both gastrointestinal and systemic viral dissemination. Consequently, CCoV is now considered to be an emerging infectious disease of dogs. In addition to the two known serotypes of CCoV, novel recombinant variants of CCoV have been found containing spike protein N-terminal domains (NTDs) that are closely related to those of feline and porcine strains. The increase in disease severity in dogs and the emergence of novel CCoVs can be attributed to the high level of recombination within the spike gene that can occur during infection by more than one CCoV type in the same host.
Genetic diversity of a canine coronavirus detected in pups with diarrhoea in Italy
Journal of Virological Methods, 2003
The sequence of the S gene of a field canine coronavirus (CCoV), strain Elmo/02, revealed low nucleotide (61%) and amino acid (54%) identity to reference CCoV strains. The highest correlation (77% nt and 81.7% aa) was found with feline coronavirus type I. A PCR assay for the S gene of strain Elmo/02 detected analogous CCoVs of different geographic origin, all which exhibited at least 92 Á/96% nucleotide identity to each other and to strain Elmo/02. The evident genetic divergence between the reference CCoV strains and the newly identified Elmo/02-like CCoVs strongly suggests that a novel genotype of CCoV is widespread in the dog population. #
PCR assay for the detection and the identification of atypical canine coronavirus in dogs
Journal of Virological Methods, 2002
Comparative sequence analysis of the PCR products of the M gene and fragments of the pol1a and pol1b genes of canine coronavirus (CCoV) have demonstrated that two separate clusters of CCoV are present in dogs. This note describes a PCR assay to identify atypical CCoV strains with nucleotide substitutions in the M gene. A total of 177 faecal samples from dogs CCoV positive previously with the PCR assay were analysed. Sixty-two of the 177 samples were amplified with the PCR described in the present study and were thus considered atypical CCoVs. The specificity of the PCR typing assay was confirmed by sequence analysis of the PCR products. #
Digital PCR Platform as a Tool to Determine the Canine Coronavirus (CCoV) Genome in Clinical Samples
Archives on Veterinary Science and Technology, 2018
Canine coronavirus (CCoV) is an important pathogen that affect dogs. Here we assessed a digital real-time based PCR (dPCR) for the detection of CCoV from infected AF-72 cells and directly from faeces from 100 symptomatic dogs. The results obtained from dPCR were compared to real-time TaqMan® based PCR assay (qPCR) and positive samples were submitted to phylogenetic analysis. Thus, dPCR had an equal sensitivity, 101 copies/μl of partial M CCoV gene, when compared to qPCR results with a good agreement in all analysis. These results indicated that the dPCR could be an alternative technique for the diagnosis of CCoV from clinical samples with advantage of simplicity and sensitivity.