CPV-2a Strains Research Articles

Isolation and sequence analysis of the complete NS1 and VP2 genes of canine parvovirus from domestic dogs in 2013 and 2014 in China

Canine parvovirus (CPV) can cause severe disease in animals and continuously generates new variant and recombinant strains in dogs that have a strong impact on sanitation. It is therefore necessary to investigate epidemic CPV strains to improve our understanding of CPV transmission and epidemic behavior. However, most studies have focused on the analysis of VP2, and therefore, information about recombination and relationships between strains is still lacking. Here, 14 strains of CPV were isolated from domestic dogs suspected of hosting CPV between 2013 and 2014 in China. The complete NS1 and VP2 genes were sequenced and analyzed. The results suggest that the new CPV-2a and new CPV-2b types are the prevalent strains in China. In addition to a few mutations (residues 19, 544, 545, 572 and 583 of NS1 and residues 267, 370, 377 and 440 of VP2) that were preserved during transmission, new mutations (residues 60, 630 of NS1, and residues 21, 310 of VP2) were found in the isolated strains. A phylogenetic tree based on VP2 sequences illustrated that the new CPV-2a and new CPV-2b strains from China form single clusters that are distinct from lineages from other countries. Moreover, recombination between the new CPV-2a and new CPV-2b types was also identified in the isolated strains. Due to differences in selection pressures or recombination, there were a small number of inconsistencies between the phylogenetic trees for VP2 and NS1, which indicated that phylogenetic relationships based on VP2 might not be representative of those based on NS1. The data indicated that mutations and recombination are constantly occurring along with the spread of CPV in China.

An insulated isothermal PCR method on a field-deployable device for rapid and sensitive detection of canine parvovirus type 2 at points of need

Canine parvovirus type 2 (CPV-2), including subtypes 2a, 2b and 2c, causes an acute enteric disease in both domestic and wild animals. Rapid and sensitive diagnosis aids effective disease management at points of need (PON). A commercially available, field-deployable and user-friendly system, designed with insulated isothermal PCR (iiPCR) technology, displays excellent sensitivity and specificity for nucleic acid detection. An iiPCR method was developed for on-site detection of all circulating CPV-2 strains. Limit of detection was determined using plasmid DNA. CPV-2a, 2b and 2c strains, a feline panleukopenia virus (FPV) strain, and nine canine pathogens were tested to evaluate assay specificity. Reaction sensitivity and performance were compared with an in-house real-time PCR using serial dilutions of a CPV-2b strain and 100 canine fecal clinical samples collected from 2010 to 2014, respectively. The 95% limit of detection of the iiPCR method was 13 copies of standard DNA and detection limits for CPV-2b DNA were equivalent for iiPCR and real-time PCR. The iiPCR reaction detected CPV-2a, 2b and 2c and FPV. Non-targeted pathogens were not detected. Test results of real-time PCR and iiPCR from 99 fecal samples agreed with each other, while one real-time PCR-positive sample tested negative by iiPCR. Therefore, excellent agreement (k=0.98) with sensitivity of 98.41% and specificity of 100% in detecting CPV-2 in feces was found between the two methods. In conclusion, the iiPCR system has potential to serve as a useful tool for rapid and accurate PON, molecular detection of CPV-2.

Phylogenetic and genome-wide deep-sequencing analyses of canine parvovirus reveal co-infection with field variants and emergence of a recent recombinant strain.

Canine parvovirus (CPV), a fast-evolving single-stranded DNA virus, comprises three antigenic variants (2a, 2b, and 2c) with different frequencies and genetic variability among countries. The contribution of co-infection and recombination to the genetic variability of CPV is far from being fully elucidated. Here we took advantage of a natural CPV population, recently formed by the convergence of divergent CPV-2c and CPV-2a strains, to study co-infection and recombination. Complete sequences of the viral coding region of CPV-2a and CPV-2c strains from 40 samples were generated and analyzed using phylogenetic tools. Two samples showed co-infection and were further analyzed by deep sequencing. The sequence profile of one of the samples revealed the presence of CPV-2c and CPV-2a strains that differed at 29 nucleotides. The other sample included a minor CPV-2a strain (13.3% of the viral population) and a major recombinant strain (86.7%). The recombinant strain arose from inter-genotypic recombination between CPV-2c and CPV-2a strains within the VP1/VP2 gene boundary. Our findings highlight the importance of deep-sequencing analysis to provide a better understanding of CPV molecular diversity.

Analysis of the full-length VP2 protein of canine parvoviruses circulating in Hungary

In recent years, the number of cases of disease caused by canine parvovirus 2 (CPV-2) in vaccinated dogs has increased. The aim of the present study was to identify CPV-2 strains present in Hungary. Forty-two out of 50 faecal specimens examined were positive, and 25 VP2 sequences were determined and analysed. Based on the current classification, the Hungarian viruses belong to New CPV-2a type, except two viruses that are recombinants of vaccine viruses and CPV-2a strains. The Tyr324Ile alteration was detected for the first time in Europe, and a "Hungarian-specific" substitution (Ala516Thr) was also identified in this study. The immunologically important parts of the currently spreading canine parvoviruses were examined and found to differ greatly from the vaccine strains that are widely used in Hungary.

Phylogenetic analysis of canine parvovirus partial VP2 gene in India

A total of 85 samples (58.0 %) were found to be positive for Canine parvovirus (CPV) by PCR assay (Hfor/Hrev primers) out of 158 suspected faecal samples of dogs collected from various states/union territories of India. Nine CPV isolates could be obtained in A-72 cell line. The sequencing of the partial VP2 gene of CPV identified the predominant CPV strain as CPV-2a (Ser297Ala) with one CPV-2b (Ser297Ala) and another CPV-2a variant strain (Ser297Gly). Several non-synonymous and synonymous mutations were also recorded in this study. The phylogenetic tree revealed that most of the CPV sequences from Tamil Nadu (Southern India) and Maharashtra (Western India) obtained during 2011 and few sequences from Northern India obtained during 2012 were grouped together along with CPV-2a (Ser297Ala) strains from China and India and followed the same evolution; although there was definitive indication of separate lineages too by few other sequences.

High local genetic diversity of canine parvovirus from Ecuador

Canine parvovirus (CPV) comprises three antigenic variants (2a, 2b, and 2c) that are distributed globally with different frequencies and levels of genetic variability. CPVs from central Ecuador were herein analyzed to characterize the strains and to provide new insights into local viral diversity, evolution, and pathogenicity. Variant prevalence was analyzed by PCR and partial sequencing for 53 CPV-positive samples collected during 2011 and 2012. The full-length VP2 gene was sequenced in 24 selected strains and a maximum-likelihood phylogenetic tree was constructed using both Ecuadorian and worldwide strains. Ecuadorian CPVs have a remarkable genetic diversity that includes the circulation of all three variants and the existence of different evolutionary groups or lineages. CPV-2c was the most prevalent variant (54.7%), confirming the spread of this variant in America. Ecuadorian CPV-2c strains clustered in two lineages, which represent the first evidence of polyphyletic CPV-2c circulating in South America. CPV-2a strains constituted 41.5% of the samples and clustered in a single lineage. The two detected CPV-2b strains (3.8%) were clearly polyphyletic and appeared related to Ecuadorian CPV-2a or foreign CPV-2b strains. Besides the substitution at residue 426 that is used to identify the variants, two amino acid changes occurred in Ecuadorian strains: Val139Iso and Thr440Ser. Ser(440) occurred in a biologically relevant domain of VP2 and is here described for the first time in CPV. The associations of Ecuadorian CPV-2c and CPV-2a with clinical symptoms indicate that dull mentation, hemorrhagic gastroenteritis and hypothermia occurred more frequently in infection with CPV-2c than with CPV-2a.

Phylodynamics analysis of canine parvovirus in Uruguay: evidence of two successive invasions by different variants

Canine parvovirus (CPV) comprises three antigenic variants (2a, 2b, and 2c) with different frequencies and genetic variability among countries. Current CPV populations are considered to be spatially structured with relatively little movement of viruses between geographical areas. Here we describe the evolution and population dynamics of CPV in Uruguay from 2006-2011 using full-length capsid viral protein 2 (VP2) sequences. CPV-2c was the predominant variant in Uruguay for 4 years (2006-2009). The estimated time to the most recent common ancestor suggested that the CPV-2c variant appeared in Uruguay around 2004-2005. Comparative phylogenetic analysis revealed that South American CPV-2c strains did not emerge de novo but may have a European origin. In 2010, a remarkable epidemiological change occurred as a consequence of the emergence of a novel CPV-2a strain in the previously homogeneous CPV-2c population. The frequency of the novel CPV-2a strain increased to 85 % in 2011, representing the first example of a CPV-2a strain replacing a predominant CPV-2c strain in a dog population. The CPV-2a strains detected in 2010-2011 were not phylogenetically related to any other strain collected on the American continent but were identical to Asiatic strains, suggesting that its emergence was a consequence of a migration event. Taken together, our findings suggest that in the last decade, Uruguay has experienced two successive invasions by CPV-2c and CPV-2a variants of European and Asiatic origins, respectively. These results support the hypothesis that CPV invasion events are not rare in certain geographic regions and indicate that some current strains may exhibit an unexpectedly high invasion and replacement capability.

Analysis of the VP2 protein gene of canine parvovirus strains from affected dogs in Japan

To clarify the evolution of canine parvovirus 2 (CPV-2) that has recently been epidemic in Japan, VP2 gene sequences at positions 3556–4166 were analyzed in 107 CPV-2 strains obtained from rectal swabs of diarrheic dogs from 2009 to 2011. CPV-2b (95 strains) was more frequently detected than CPV-2a (nine strains), while CPV-2c was not detected. Remaining three strains were identified as the original type CPV-2, which should be derived from vaccines. These findings are similar to the previous results involving Japanese strains, suggesting there has been no great change in the recent CPV-2 epidemic in Japan. This epidemic is the same as that in Taiwan. Furthermore, a 324-lle mutant, which has been reported in Korean and Chinese strains, was detected in 66.7% of CPV-2a strains.

Role of Multiple Hosts in the Cross-Species Transmission and Emergence of a Pandemic Parvovirus

Understanding the mechanisms of cross-species virus transmission is critical to anticipating emerging infectious diseases. Canine parvovirus type 2 (CPV-2) emerged as a variant of a feline parvovirus when it acquired mutations that allowed binding to the canine transferrin receptor type 1 (TfR). However, CPV-2 was soon replaced by a variant virus (CPV-2a) that differed in antigenicity and receptor binding. Here we show that the emergence of CPV involved an additional host range variant virus that has circulated undetected in raccoons for at least 24 years, with transfers to and from dogs. Raccoon virus capsids showed little binding to the canine TfR, showed little infection of canine cells, and had altered antigenic structures. Remarkably, in capsid protein (VP2) phylogenies, most raccoon viruses fell as evolutionary intermediates between the CPV-2 and CPV-2a strains, suggesting that passage through raccoons assisted in the evolution of CPV-2a. This highlights the potential role of alternative hosts in viral emergence.

Recent spreading of a divergent canine parvovirus type 2a (CPV-2a) strain in a CPV-2c homogenous population

Canine parvovirus type 2 (CPV-2), which causes acute hemorrhagic enteritis in dogs, is comprised of three antigenic variants (2a, 2b, and 2c) that are distributed worldwide with different frequencies. Variant prevalence was analyzed in 150 CPV-2-positive samples collected from the Uruguayan dog population in 2007–2010. Samples were analyzed with polymerase chain reaction, restriction fragment length polymorphism, and sequencing of the coding region for the largest and most variable loop of the VP2 capsid protein. CPV-2c was the only strain detected from 2007 to 2009. Uruguayan CPV-2c showed high homogeneity in both nucleotide and amino acid sequences, indicating a low level of genetic variability. In 2010, an unexpected epidemiological change occurred in Uruguay as a consequence of the appearance of a novel CPV-2a strain. This variant rapidly spread through the Uruguayan dog population and was detected in 20 of the 52 cases (38%) analyzed in 2010. CPV-2a sequences were identical in all field viruses analyzed, and in addition to the characteristic 426Asn residue, the sequences showed amino acid substitutions (267Tyr, 324Ile, and 440Ala) not observed in the Uruguayan CPV-2c. These data and the first detection in April 2010 suggest that the CPV-2a variant recently emerged in Uruguay and underwent clonal expansion. This observation is the first case in which a CPV-2a variant increased its frequency in a dog population where CPV-2c was prevalent. Our results emphasize the dynamic changes in CPV variants and highlight the importance of ongoing surveillance programs to provide a better understanding of virus epidemiology.

Misleading results of the MboII-based identification of type 2a canine parvovirus strains from Hungary reacting as type 2c strains

Type 2 canine parvovirus (CPV2) infection is one of the most frequent causes of death in the young, susceptible canine populations worldwide. Since its emergence in the 1970s, several variants have been described. In the present study the authors describe the genetic analysis of 24 Hungarian CPV2 strains collected from 2004 to 2008. Surprisingly, the genetic and phylogenetic investigations of all these strains revealed that all of them were type 2a CPVs. On the other hand, the genetic analysis provided substantial evidence to demonstrate that due to a seemingly constant point mutation present in most of the Hungarian CPV2a strains, a previously described MboII-based rapid identification of CPV2c strains unfortunately cannot be reliably used any more.

Characterisation of canine parvovirus strains isolated from cats with feline panleukopenia

Unlike the original canine parvovirus type 2 (CPV-2), CPV-2 variants have gained the ability to replicate in vivo in cats but there is limited information on the disease patterns induced by these variants in the feline host. During 2008, two distinct cases of parvoviral infection were diagnosed in our laboratories. A CPV-2a variant was identified in a 3-month-old Persian kitten displaying clinical sign of feline panleukopenia (FPL) (acute gastroenteritis and marked leukopenia) and oral ulcerations, that died eight days after the onset of the disease. Two pups living in the same pet shop as the cat were found to shed a CPV-2a strain genetically identical to the feline virus and were likely the source of infection. Also, non-fatal infection by a CPV-2c strain occurred in a 2.5-month-old European shorthair kitten displaying non-haemorrhagic diarrhoea and normal white blood cell counts. By sequence analysis of the major capsid protein (VP2) gene, the feline CPV-2c strain showed 100% identity to a recent canine type-2c isolate. Both kittens had been administered multivalent vaccines against common feline pathogens including FPL virus. Whether and to which extent the FPL vaccines can protect cats adequately from the antigenic variants of CPV-2 should be assessed.

Diagnosis of canine parvovirus by rapid immunomigration on a membrane

Rapid immunomigration on a membrane was applied to the diagnosis of canine parvovirus (CPV) in 128 samples of faeces containing four strains of parvovirus (two CPV-2a strains, itncluding one vaccine...