Feline Calicivirus Virulent Systemic Disease: Clinical Epidemiology, Analysis of Viral Isolates and In Vitro Efficacy of Novel Antivirals in Australian Outbreaks.

Feline calicivirus (FCV) causes upper respiratory tract disease (URTD) and sporadic outbreaks of virulent systemic disease (FCV-VSD). The basis for the increased pathogenicity of FCV-VSD viruses is incompletely understood, and antivirals for FCV-VSD have yet to be developed. We investigated the clinicoepidemiology and viral features of three FCV-VSD outbreaks in Australia and evaluated the in vitro efficacy of nitazoxanide (NTZ), 2'-C-methylcytidine (2CMC) and NITD-008 against FCV-VSD viruses. Overall mortality among 23 cases of FCV-VSD was 39%. Metagenomic sequencing identified five genetically distinct FCV lineages within the three outbreaks, all seemingly evolving in situ in Australia. Notably, no mutations that clearly distinguished FCV-URTD from FCV-VSD phenotypes were identified. One FCV-URTD strain likely originated from a recombination event. Analysis of seven amino-acid residues from the hypervariable E region of the capsid in the cultured viruses did not support the contention that properties of these residues can reliably differentiate between the two pathotypes. On plaque reduction assays, dose-response inhibition of FCV-VSD was obtained with all antivirals at low micromolar concentrations; NTZ EC50, 0.4-0.6 µM, TI = 21; 2CMC EC50, 2.7-5.3 µM, TI > 18; NITD-008, 0.5 to 0.9 µM, TI > 111. Investigation of these antivirals for the treatment of FCV-VSD is warranted.

Isolation and phylogenetic analysis of strains of feline calicivirus in Beijing, China.

Feline calicivirus (FCV) is a contagious cat pathogen that causes oral ulceration and/or upper respiratory disease. In this study, we collected 61 samples from a pet hospital in Beijing and used PCR or RT-PCR to detect FCV and feline herpesvirus 1 (FHV-1). Approximately 44.3% (27/61) of the samples were FCV positive, and 23.0% (14/61) were coinfected with FCV and FHV-1. FCV was isolated from 15 samples. One isolate was from a cat with virulent systemic disease (VSD) signs, and 14 isolates were from cats with stomatitis or upper respiratory diseases. The range of genome sequence identity among these isolates was 76.1-100.0%. Four of the isolates were considered to be of the same strain, with sequence identity ranging from 99.5 to 99.7%, and two isolates, BJ-280 and BJ-288, had completely identical sequences. The genomic sequence identity ranged from 76.0 to 88.5% between the 15 isolates and several reference strains, including the F4 and F9 vaccine strains. These results demonstrate that many FCV strains are co-circulating in Beijing. Due to the diversity of FCV in Beijing, it is necessary to monitor the current prevalence of the virus. This study provides more information for the development of effective measures to control FCV.

Temporally separated feline calicivirus isolates do not cluster phylogenetically and are similarly neutralised by high-titre vaccine strain FCV-F9 antisera in vitro.

OBJECTIVES: Feline calicivirus (FCV) is a highly variable and globally important feline pathogen for which vaccination has been the mainstay of control. Here, we test whether the continued use of FCV-F9, one of the most frequently used vaccine strains globally, is driving the emergence of vaccine-resistant viruses in the field. METHODS: This study made use of two representative panels of field isolates previously collected from cats visiting randomly selected veterinary practices across the UK as part of separate cross-sectional studies from 2001 and 2013/2014. Phylogenetic analysis and in vitro virus neutralisation tests were used to compare the genetic and antigenic relationships between these populations and FCV-F9. RESULTS: Phylogenetic analysis showed a typically radial distribution dominated by 52 distinct strains, with strains from both 2001 and 2013/2014 intermingled. The sequence for FCV-F9 appeared to be integral to this phylogeny and there were no significant differences in the genetic distances within each studied population (intra-population distances), or between them (inter-population distances), or between each population and FCV-F9. A 1 in 8 dilution neutralised 97% and 100% of the 2001 and 2013/14 isolates, respectively, and a 1 in 16 dilution neutralised 87% and 75% of isolates, respectively. There was no significant difference either in variance between the FCV-F9 neutralising titres for the two populations, or in the distribution of neutralisation titres across the two populations. CONCLUSIONS AND RELEVANCE: Although FCV is a highly variable virus, we found no evidence for a progressive divergence of field virus from vaccine strain FCV-F9, either phylogenetically or antigenically, with FCV-F9 antisera remaining broadly and equally cross-reactive to two geographically representative and temporally separated FCV populations. We suggest this may be because the immunodominant region of the FCV capsid responsible for neutralisation may have structural constraints preventing its longer term progressive antigenic evolution.

Feline calicivirus infection in cats with virulent systemic disease, Italy.

Feline calicivirus (FCV) is a contagious viral pathogen that usually causes a mild, self-limiting respiratory disease. More recently, highly virulent FCV strains have emerged and have been associated with severe systemic infection, referred to as virulent systemic disease (VSD). The objective of this study is to report VSD cases in Italian cats along with the molecular characterization of two detected FCV strains. Three client-owned cats showed clinical signs resembling to those described for VSD cases. The cats were subjected to molecular investigations for detection of FCV and other feline pathogens. Histopathology and immunohistochemistry were performed on internal organs of one cat; molecular characterization of two detected FCV strains was obtained through sequence and phylogenetic analyses. Putative VS-FCV strains were detected in all three cats, which were co-infected with feline panleukopenia virus. The cat submitted to histopathology and immunohistochemistry displayed severe histological changes and FCV antigens in internal organs. Two Italian FCV strains, for which amplification of ORF2 was successful, were strictly related and formed a unique phylogenetic cluster. These viruses did not show consistent changes in the amino acid sequences with respect to reference VS-FCVs. The results of our study confirm that VS-FCV strains are circulating in Italy and that VSD diagnosis is complicated since both genetic and clinical markers have not been identified so far.

Molecular characterization of feline calicivirus variants from multicat household and public animal shelter in Rio de Janeiro, Brazil

ABSTRACT The aim of this study was to perform the molecular characterization of conserved and variable regions of feline calicivirus capsid genome in order to investigate the molecular diversity of variants in Brazilian cat population. Twenty-six conjunctival samples from cats living in five public short-term animal shelters and three multicat life-long households were analyzed. Fifteen cats had conjunctivitis, three had oral ulceration, eight had respiratory signs (cough, sneeze and nasal discharge) and nine were asymptomatic. Feline calicivirus were isolated in CRFK cells and characterized by reverse transcription PCR target to both conserved and variable regions of open reading frame 2. The amplicons obtained were sequenced. A phylogenetic analysis along with most of the prototypes available in GenBank database and an amino acid analysis were performed. Phylogenetic analysis based on both conserved and variable region revealed two clusters with an aLTR value of 1.00 and 0.98 respectively and the variants from this study belong to feline calicivirus genogroup I. No association between geographical distribution and/or clinical signs and clustering in phylogenetic tree was observed. The variants circulating in public short-term animal shelter demonstrated a high variability because of the relatively rapid turnover of carrier cats constantly introduced of multiple viruses into this location over time.

Feline chronic gingivostomatitis with calicivirus infection: case report / Gengivostomatite crônica felina com infecção por calicivírus: relato de caso

Feline chronic gingivostomatitis (FCGS) is an oral inflammatory condition that frequently affects felines. Its etiology is not well defined, but several viral agents are thought to be involved. Several therapeutic protocols have been described, yet treatment response is often variable, and the therapeutic success is transient with an unpredictable duration. Therefore, the therapeutic strategy needs to be tailored for each patient. This work relates a case characterized by viral involvement in its etiopathogenesis providing an alternative to the most widely-used methods that so often frustrate both veterinary doctors and pet owners.(AU)

A gengivostomatite crônica felina (FCGS) é uma condição inflamatória oral que frequentemente afeta felinos. A sua etiologia não está bem definida, mas acredita-se que vários agentes virais possam estar envolvidos. Muitos protocolos terapêuticos têm sido descritos, no entanto, a resposta ao tratamento é frequentemente variável e o sucesso terapêutico é transitório com uma duração imprevisível. Portanto, a estratégia terapêutica precisa ser adaptada para cada paciente. O presente trabalho propõe a caracterização do envolvimento viral na etiopatogenia da doença como uma alternativa aos métodos mais amplamente utilizados, que muitas vezes frustram médicos veterinários e os donos de animais de estimação.(AU)

Molecular characterization of feline calicivirus variants from multicat household and public animal shelter in Rio de Janeiro, Brazil.

The aim of this study was to perform the molecular characterization of conserved and variable regions of feline calicivirus capsid genome in order to investigate the molecular diversity of variants in Brazilian cat population. Twenty-six conjunctival samples from cats living in five public short-term animal shelters and three multicat life-long households were analyzed. Fifteen cats had conjunctivitis, three had oral ulceration, eight had respiratory signs (cough, sneeze and nasal discharge) and nine were asymptomatic. Feline calicivirus were isolated in CRFK cells and characterized by reverse transcription PCR target to both conserved and variable regions of open reading frame 2. The amplicons obtained were sequenced. A phylogenetic analysis along with most of the prototypes available in GenBank database and an amino acid analysis were performed. Phylogenetic analysis based on both conserved and variable region revealed two clusters with an aLTR value of 1.00 and 0.98 respectively and the variants from this study belong to feline calicivirus genogroup I. No association between geographical distribution and/or clinical signs and clustering in phylogenetic tree was observed. The variants circulating in public short-term animal shelter demonstrated a high variability because of the relatively rapid turnover of carrier cats constantly introduced of multiple viruses into this location over time.

A multi-national European cross-sectional study of feline calicivirus epidemiology, diversity and vaccine cross-reactivity.

BACKGROUND: Feline calicivirus (FCV) is an important pathogen of cats for which vaccination is regularly practised. Long-term use of established vaccine antigens raises the theoretical possibility that field viruses could become resistant. This study aimed to assess the current ability of the FCV-F9 vaccine strain to neutralise a randomly collected contemporary panel of FCV field strains collected prospectively in six European countries. METHODS: Veterinary practices (64) were randomly selected from six countries (UK, Sweden, Netherlands, Germany, France and Italy). Oropharyngeal swabs were requested from 30 (UK) and 40 (other countries) cats attending each practice. Presence of FCV was determined by virus isolation, and risk factors for FCV shedding assessed by multivariable logistic regression. Phylogenetic analyses were used to describe the FCV population structure. In vitro virus neutralisation assays were performed to evaluate FCV-F9 cross-reactivity using plasma from four vaccinated cats. RESULTS: The overall prevalence of FCV was 9.2%. Risk factors positively associated with FCV shedding included multi-cat households, chronic gingivostomatitis, younger age, not being neutered, as well as residing in certain countries. Phylogenetic analysis showed extensive variability and no countrywide clusters. Despite being first isolated in the 1950s, FCV-F9 clustered with contemporary field isolates. Plasma raised to FCV-F9 neutralized 97% of tested isolates (titres 1:4 to 1:5792), with 26.5%, 35.7% and 50% of isolates being neutralized by 5, 10 and 20 antibody units respectively. CONCLUSIONS: This study represents the largest prospective analysis of FCV diversity and antigenic cross-reactivity at a European level. The scale and random nature of sampling used gives confidence that the FCV isolates used are broadly representative of FCVs that cats are exposed to in these countries. The in vitro neutralisation results suggest that antibodies raised to FCV-F9 remain broadly cross-reactive to contemporary FCV isolates across the European countries sampled.

Genetic diversity and phenotypic associations of feline caliciviruses from cats in Switzerland.

Feline calicivirus (FCV) is a common viral pathogen in domestic cats worldwide. The variable regions of the capsid (VP1) gene of FCV have one of the highest recorded rates of molecular evolution. Understanding the genetic diversity and phylogeny of FCV is a prerequisite to exploring the epidemiology and pathogenesis of this virus and to the development of efficacious vaccine strategies. In this study, we undertook a nationwide molecular characterization of FCV using for the first time nearly complete capsid (VP1) gene sequences. Sequences from 66 FCV samples were used to investigate the correlation between viral phylogeny and several traits, including geographic origin, signalment, husbandry, FCV vaccination and co-infections. Codon-based nucleotide alignment showed that individual nucleotides and their corresponding amino acid sites were either invariant or highly variable. Using a threshold of 20 % genetic distance in variable region E, FCV samples were grouped into 52 strains, 10 of which comprised two to three samples. Significant associations between FCV phylogeny and host characteristics were found, specifically the pedigree status of the cats, and two well-supported lineages were identified in which the current FCV strain definition was confounded. No correlation between viral genetic distances and geographic distances was evident. The greater resolution of the FCV phylogeny in this study compared to previous studies can be attributed to our use of more conserved regions of the capsid (VP1) gene; nonetheless, our results were still hampered by sequence saturation. The study highlights the need for whole-genome sequences for FCV phylogeny studies.

Humoral response to calicivirus in captive tigers given a dual-strain vaccine.

The current feline vaccine with a single strain of calicivirus has been used for captive tigers, yet it may not protect against virulent systemic calicivirus infections. A cross-institutional study investigated the humoral response to a new dual-strain, killed-virus calicivirus vaccine for nine captive tigers. The subspecies of these tigers were Amur (Panthera tigris altaica), Bengal (Panthera tigris tigris), and Malayan (Panthera tigris jacksoni). Serum neutralization titers for virulent feline calicivirus strain FCV-DD1 were higher following dual-strain vaccine administration. There were no reports of adverse vaccine reactions. Dual-strain vaccination may afford broadened cross-protection against different calicivirus strains and is desirable to reduce the risk of virulent systemic calicivirus disease in tigers.

Genetic characterization of feline calicivirus strains associated with varying disease manifestations during an outbreak season in Missouri (1995-1996).

Feline calicivirus (FCV) is a common cause of mild to severe upper respiratory tract disease (URTD) in cats. FCV strain 21223 was isolated from a kitten with severe pneumonia in a disease outbreak with unusually high mortality (35 %) that occurred in a Missouri feline colony in 1995-1996. Phylogenetic analysis of the genome sequence of strain 21223 indicated the emergence of a new FCV strain. Analysis of the full-length genome sequence of a closely related (99.5 % nucleotide identity) strain, 3786, obtained from an asymptomatic animal in the same colony four months later, showed the presence of seven amino acid substitutions, with six of them located in the VP1 capsid sequence encoded by ORF2. Comparative analysis of the E-region sequences (426-521 aa ORF2) presumably involved in virus-host cell receptor interactions did not identify amino acid substitutions unique to the virulent strain. We determined the complete genome sequences of four virus isolates that were collected in regional catteries in the months following the outbreak that were associated with different manifestations of the disease (URTD, chronic stomatitis, and gingivitis). We show that genetically distinct FCV strains were cocirculating in the area, and no apparent correlation could be made between overall sequence and observed disease.

Large-scale spatial and temporal genetic diversity of feline calicivirus.

Feline calicivirus (FCV) is an important pathogen of domestic cats and a frequently used model of human caliciviruses. Here we use an epidemiologically rigorous sampling framework to describe for the first time the phylodynamics of a calicivirus at regional and national scales. A large number of FCV strains cocirculated in the United Kingdom at the national and community levels, with no strain comprising more than 5% and 14% of these populations, respectively. The majority of strains exhibited a relatively restricted geographical range, with only two strains (one field virus and one vaccine virus) spreading further than 100 km. None of the field strains were identified outside the United Kingdom. Temporally, while some strains persisted locally for the majority of the study, others may have become locally extinct. Evolutionary analysis revealed a radial phylogeny with little bootstrap support for nodes above the strain level. In most cases, spatially and temporally diverse strains intermingled in the phylogeny. Together, these data suggest that current FCV evolution is not associated with selective competition among strains. Rather, the genetic and antigenic landscape in each geographical location is highly complex, with many strains cocirculating. These variants likely exist at the community level by a combination of de novo evolution and occasional gene flow from the wider national population. This complexity provides a benchmark, for the first time, against which vaccine cross-protection at both local and national levels can be judged.

Genetic and phylogenetic analyses of capsid protein gene in feline calicivirus isolates from Rio Grande do Sul in southern Brazil.

Feline calicivirus (FCV) is an important pathogen that affects domestic cats, inducing acute oral and upper respiratory tract clinical signs. The aim of this study was to analyze the variability of the capsid protein in different FCV isolates from southern Brazil. The sequencing analyses of thirteen Brazilian FCV samples, phylogenetic analyses and assessments of ten previously published sequences were conducted by examining the open reading frame 2 (ORF2, regions B-F). Comparisons of the predicted amino acid sequences of the ORF2 in Brazilian FCV isolates with those of the FCV-F9 strain indicated that the main differences are located within the regions C and hypervariable E (HVR_E). Epitopes that were mapped to the regions D, 5'HVR_E and conserved E also presented with some variability when compared to the strain F9. This is the first study describing sequence analyses and the phylogenetic relationships among FCV isolates from Brazil. The results presented here may expand upon current knowledge regarding aspects of FCV biology, epidemiology and genetic diversity and provide insights into improving the efficacies of current FCV vaccines.

Feline calicivirus VP2 is involved in the self-assembly of the capsid protein into virus-like particles.

Feline calicivirus (FCV) is considered the most common upper respiratory tract disease (URTD) associated pathogen in cats. We previously expressed FCV VP1 capsid protein in insect cells by baculovirus system and we observed that this protein self-assemble into virus-like particles (VLPs) different in size and lacking the typical cup-like depressions of caliciviruses. In the present study, VP1 and the small basic structural protein VP2 of FCV were individually expressed by baculovirus system. Coinfection of insect cells with both recombinant viruses resulted in VP1 and VP2 self-assembly to form depressions similar to native capsids in size and appearance, demonstrating that VP2 interacts with the VP1 protein in the formation of VLPs.

A dual-strain feline calicivirus vaccine stimulates broader cross-neutralization antibodies than a single-strain vaccine and lessens clinical signs in vaccinated cats when challenged with a homologous feline calicivirus strain associated with virulent systemic disease.

Feline calicivirus (FCV) causes an array of clinical disease in cats. Traditionally this disease has been associated with respiratory disease, limping, or chronic stomatitis. Within the last 10 years, virulent systemic feline calicivirus (VS-FCV) has been recognized which causes additional clinical signs and has a higher fatality rate. A dual-strain FCV vaccine containing a strain of FCV associated with traditional respiratory disease and a VS-FCV strain stimulates serum cross-neutralization antibodies when tested against field strains from Europe and VS-FCV strains from USA. Following challenge with a homologous VS-FCV strain, vaccinated cats had significantly reduced clinical signs.

Feline calicivirus strains isolated in Italy.

Feline Calicivirus (FCV) has been recognised as major oral and respiratory pathogen of cats. The high correlation among the field viruses and FCV-F9 serotype has represented the immunological bases for the employ of FCV-F9 serotype as a vaccine for calicivirosis in cats. The aim of this paper was to evaluate, by in vitro neutralization assays, the antigenic correlation among the vaccine F9 and FCV field strains isolated in Sicily (Italy) from cats showing clinical forms referable to calicivirus infection. The results confirm the low correlation between FCV-F9 strain and calicivirus strains spread in the feline population.

Comparison of the ability of feline calicivirus (FCV) vaccines to neutralise a panel of current UK FCV isolates.

Feline calicivirus (FCV) comprises a large number of strains which are related antigenically to varying degrees. The antigenic variability creates problems for choosing antigens to include in vaccines. Historically, these have been selected for use based on their cross-reactivity with a high proportion of field strains. However, it is important to determine the current level of cross-reactivity of vaccines and whether or not this may be decreasing owing to widespread vaccine use. In this in vitro study, we have compared the ability of antisera to two vaccine viruses (FCV strain F9 and FCV strain 255) to neutralise a panel of 40 recent UK field isolates. These 40 isolates were obtained by randomised, cross-sectional sampling of veterinary practices in different geographical regions of the UK so as to ensure they were representative of viruses circulating in the veterinary-visiting population of cats in the UK. Virus neutralisation assays showed that both vaccine strains are still broadly cross-reactive, with F9 antiserum neutralising 87.5% and 255 antiserum 75% of isolates tested with antiserum dilutions of 1 in 2 or greater. However, when antibody units were used, in order to take account of differences in homologous titres between antisera, fewer isolates were neutralised, with F9 antiserum showing a slightly higher proportion of isolates neutralised than 255. Multivariable analysis of the sample population of 1206 cats from which the 40 isolates were derived found that vaccinated cats were at a decreased risk of being positive for FCV, whereas cats from households with more than one cat, and cats with mouth ulcers were at increased risk. In addition as cats became older their risk of shedding FCV decreased.

Longitudinal molecular epidemiological analysis of feline calicivirus infection in an animal shelter: a model for investigating calicivirus transmission within high-density, high-turnover populations.

The control of outbreaks of calicivirus infection in high-density, high-throughput populations is a challenge to both human and veterinary medicine. In such populations, the prevalence of infection is, in part, dependent on the levels of biosecurity and how this affects virus transmission. Here we show how longitudinal analysis of feline calicivirus (FCV) infection in an animal rescue shelter can be used as a model to examine the dynamics of calicivirus transmission and evolution in such environments. FCV was isolated from 33 of 116 cats sampled over a 15-month period (overall prevalence, 28%). Sequence analysis of the immunodominant variable regions of the viral capsid gene identified 16 strains circulating in the shelter, with no single strain appearing to predominate. The majority of these strains were introduced into the shelter from the community and did not appear to be transmitted within the population. However, for three of these strains, putative transmission events within the shelter were identified. The rates of evolution within hypervariable regions of the FCV capsid gene in individual cats ranged from 0.05 to 1.4% per week, with the highest rates generally being found in animals that either acquired the virus while in the shelter or were undergoing acute infection. These data suggest that despite the high prevalence and presence of multiple strains of FCV within the shelter, the spread of such pathogens may be restricted by various control measures, including good hygiene and biosecurity.

Genogrouping of vaccine breakdown strains (VBS) of feline calicivirus in Japan.

Although prevention of feline calcivirus (FCV) infection by vaccination has been attempted, and isolation of FCV, development of the disease, and a few fatal cases in vaccinated cats have been reported. Fifteen FCV strains isolated from cats that had been vaccinated with commercially available FCV vaccines (F9, FCV-255, and FC-7) were genogrouped. Molecular analysis of viral genomes involved the construction of a phylogenetic tree of capsid genes using the NJ method. Cat anti-F9 serum and rabbit anti-FCV-255 serum were used for virus neutralization tests. Molecular phylogenetic analysis of the amino acid sequences of 15 virus isolates and those of the previously published and GenBank-deposited 9 global and 14 Japanese strains showed that 8 (53%) of the 15 virus isolates as well as the vaccine strains F9 and FCV-255 belonged to genogroup I (G(A)I), and 7 (47%) belonged to genogroup II (G(A)II). Of the 8 G(A)I strains, 2 were isolated from cats that had been vaccinated with an F9 strain live vaccine, 5 from cats vaccinated with an FCV-255-derived vaccine, and 1 from a cat vaccinated with an FC-7-derived vaccine. Of the 7 GAll strains, 5 were isolated from cats that had been vaccinated with the F9 strain live vaccine, 1 from a cat vaccinated with the FCV-255-derived vaccine, and 1 from a cat vaccinated with the FC-7-derived vaccine. These results indicate that more vaccine breakdown strains isolated from the cats vaccinated with the F9 strain-derived vaccine belong to G(A)II than to G(A)I, whereas more vaccine breakdown strains isolated from the cats vaccinated with the FCV-255 strain-derived vaccine belong to G(A)I than to G(A)II, and that when the FC-7 strain-derived vaccine is used, the vaccine breakdown strains belong almost equally to G(A)I and G(A)II. Thus, the genogroups of virus isolates varied with the vaccine strain used (p < 0.05). On the other hand, the neutralizing titres of feline anti-F9 serum and rabbit anti-FCV-255 serum against the 15 isolates were very low, showing no relationships between neutralizing antibody titres and genogroups. The DNA sequence identities between the virus isolates and the vaccine strains were low, at 70.6-82.9%, and no strains were found to have sequences derived from the vaccine strains. Alignment of amino acid sequences showed that the G(A)I or G(A)II virus isolates from the F9-vaccinated cats differed at position 428 of the 5' hypervariable region (HVR) of capsid region of the F9 strain, whereas those from the FCV-255-vaccinated cats differed at positions 438, 453, and 460 of the 5'HVR of capsid region E of the F9 strain. We speculate that these differences influence genogrouping. The amino acid changes within the F9 linear epitopes common to G(A)I and G(A)II were noted at positions 450, 451, 457 of 5'HVR of the capsid region E in the isolates from F9-derived vaccine-treated cats, and 449, 450, and 451 of 5'HVR of capsid region E in the isolates from FCV-255-derived vaccine-treated cats, suggesting that these amino acid changes are involved in escapes. These results suggest that alternate vaccination with the F9 and FCV-255 strains or the use of a polyvalent vaccine containing GAll strains serves to inhibit development.

Feline caliciviruses (FCVs) isolated from cats with virulent systemic disease possess in vitro phenotypes distinct from those of other FCV isolates.

During the past decade, several outbreaks of severe systemic disease associated with Feline calicivirus (FCV) have occurred in the USA and the UK. This new disease has caused high mortality in the affected animals and has been termed virulent systemic (VS)-FCV disease. Currently, there are no genetic or in vitro diagnostic methods to distinguish viruses isolated from cases of VS-FCV disease from other isolates. Here, five in vitro properties, as well as the capsid and proteinase-polymerase (pro-pol) sequences, of a set of FCV isolates that included seven isolates from five distinct VS-FCV outbreaks ('VS isolates') were investigated. Although all of the FCV isolates investigated had similar kinetics of growth under single-cycle conditions, VS isolates infected tissue-culture cells more efficiently under multiple-cycle growth conditions. Moreover, it was found that cells infected with VS isolates showed cytopathic effects earlier than cells infected with non-VS isolates, although no difference in relative ATP levels were noted at times when morphological changes were first seen. Both VS- and other (non-VS) isolates of FCV demonstrated similar temperature stabilities. Phylogenetic analyses and alignments of the capsid and pro-pol regions of the genome did not reveal any conserved changes that correlated with virulence, and the VS isolates did not segregate into a unique clade. These results suggest that VS isolates have arisen independently several times since first being described and can spread more efficiently in tissue culture than other isolates when infected at low multiplicity.