key: cord-0001248-yrju4spi authors: Henzel, Andréia; Brum, Mário Celso Sperotto; Lautert, Cláudia; Martins, Mathias; Lovato, Luciane Teresinha; Weiblen, Rudi title: Isolation and identification of feline calicivirus and feline herpesvirus in Southern Brazil date: 2012-06-01 journal: Braz J Microbiol DOI: 10.1590/s1517-83822012000200017 sha: 5b2c74eaf9dc747307ec2d91ba16771cd505e44c doc_id: 1248 cord_uid: yrju4spi Feline calicivirus (FCV) and feline herpesvirus type 1 (FHV-1) are the two primary causes of upper respiratory tract disease in cats. The aim of this study was to demonstrate the distribution of FCV and FHV-1 among the feline population of several counties in Rio Grande do Sul State, Brazil. To this end, conjunctival and nasal swabs were collected from 302 cats from different locations, including households, breeding catteries, veterinary clinics, animal hospitals and experimental research facilities. The samples were collected between July 2006 to June 2009. The virus isolation was performed in CRFK cells and, subsequently, the identification was confirmed by PCR. FCV, FHV-1, or both were isolated from 55 cats from 28 different locations. FCV alone was isolated from 52.7% (29/55) of the animals that tested positively, FHV-1 alone was isolated from 38.2% (21/55) of the animals that tested positively, and co-infection were detected in 9.1% (5/55) of the animals that tested positively. Virus detection was more prevalent in cats that were less than 1 year old, among animals that shared a living space with other cats, and females. FCV and FHV-1 were isolated from vaccinated cats. In addition, both viruses were isolated from cats that showed no signs of disease. The results suggest that a carrier state is common for both viruses in the evaluated population. A search for other causes of respiratory disease in that population is necessary; and further studies relating to the molecular characterization of viruses and vaccine efficacy are also necessary. Infectious respiratory disease is a major clinical problem in feline medicine. Such infections are primarily caused by either one or both of two viruses: feline calicivirus (FCV) and feline herpesvirus type 1 (FHV-1) (11, 25) . These viruses have *Corresponding Author. Mailing address: Setor de Virologia, DMVP/CCR/UFSM, Av. Roraima, n° 1000, Santa Maria, Rio Grande do Sul, Brazil, 97105-900.; Tel/Fax.: 5555 3220-8034.; E-mail: rudi@smail.ufsm.br FHV-1 is the agent of feline viral rhinotracheitis (FVR) (11, 8) . It is a DNA virus that is a member of the Varicellovirus genus in the Alphaherpesvirinae subfamily (11) . Only one serotype of this virus exists, and like other alphaherpesvirus, FHV-1 induces latency in nervous ganglions (11) . Thus, clinically recovered cats are carriers that undergo periodic episodes of virus reactivation, particularly after stress (11) . In both experimental and natural infections, symptoms include the following: depression, sneezing, inappetence, pyrexia and serous ocular and nasal discharges (11) . Cats of any age, sex or breed are susceptible, but a severe syndrome is usually restricted to kittens of up to six months of age (23) . It is estimated that more than 90% of the cats are seropositive to FHV; and that a minimum of eighty percent remains latently infected with 45% shedding the virus by all life long (18) . The nasal, oral and conjunctival vias are the natural routes of infection for FCV and FHV-1 (11, 25) . Transmission occurs mainly through direct contact between infected and susceptible cats; however, indirect transmission can also occur in the case of FCV, particularly within a cattery where secretions may contaminate cages, feeding and cleaning utensils or personnel (11, 25) . Reports from cats with URTD have revealed a prevalence ranging among 20-53% for FCV and 10-34% for FHV-1 (2, 12, 16) . In the general healthy cat population from several European countries, USA and Korea, the prevalence of FCV has varied from 15% to 31% (2, 16) whereas values ranging from less than 1% to 63% have been estimated for the prevalence of FHV-1 (2, 13, 15) . These viruses are still prevalent in the feline population despite the fact that vaccination against FCV and FHV-1 has been practiced since the 1970s (2, 29) . Vaccinations may have reduced the overall severity of disease; however, in some vaccinated individuals, disease may still occur (11, 25, 29) . Commercially available vaccines are generally safe and protect reasonably well against disease, although they do not prevent infection, the shedding of virus or even the development of the carrier state (9, 11, 20, 25) . Live attenuated and inactivated vaccines are available (11, 25) , and vaccine virus shedding after vaccination has been described experimentally only for a temperature sensitive FHV-1 vaccine (33) . In that case, vaccine virus has been shed for 25 days after vaccination (33) . There is little available information about FCV and FHV-Feline calicivirus and feline herpesvirus 1 in Brazil. Vaccination is performed in Veterinary Clinics and Hospitals with live attenuated and inactivated vaccines however the percentage of the population that is actually vaccinated is unknown. The isolation of FCV has been described once in the southern part of the country (32), followed by an experimental study regarding the pathogenicity of the virus (22) . However, to our knowledge, there has not yet been a described isolation of FHV-1 in Brazil. Regardless, evidence of the presence of both viruses has been obtained from serologic surveys performed in populations of wild (6, 27) and domestic felines (14) . The aim of the present study was to generate insights into the epidemiology of FCV and FHV-1 in the southern part of Brazil. Thus, conjunctival, nasal, oral and oropharyngeal swabs were collected from 302 cats in several counties in Rio Grande do Sul State, and the resulting isolation and identification of FCV and FHV-1 are described. The samples consisted of conjunctival, nasal and, Table 2) . The classification by gender revealed that the difference between the number of samples collected from male and female cats was only two, although the number of females that tested positively for virus isolation was more than two-fold higher the number of males that tested positively ( Table 2) . With regards to the origin of the cats, females were positive in 16 locations, males in 9 locations and both male and female in 3 locations (data not shown). The epidemiological conditions of feline calicivirus (FCV) and feline herpesvirus type 1 (FHV-1) are known among the feline population worldwide (11, 25) . Although it is generally assumed that these conditions are similar in Brazil, no major study regarding these viruses has been performed in that country, with the exception of several serological surveys (6, 14, 27) and an experimental study regarding pathogenicity (22) . The diseases of the respiratory tract of felines are an important and recurrent problem for veterinarians and cat owners globally, and FCV and FHV-1 have been described as one of the primary causes of these clinical manifestations (2, 8) . In this study, FCV, FHV-1 or both were isolated and identified from 55 cats with or without clinical signs from a total of 302 animals sampled in some cities of the Rio Grande do Sul State in the southern part of Brazil. In the present article, FCV was isolated more often, in terms of the overall results, but its frequency was different when the groups were analyzed separately ( Table 2) . FCV was the primary virus isolated in most of the groups examined, although FHV-1 was the virus most frequently isolated from vaccinated cats and was also isolated slightly more frequently than FCV in cats exhibiting clinical signs of disease. A greater prevalence of FCV in comparison to FHV-1 has been reported in cats with clinical manifestations of disease as well for clinically healthy cats (2, 16), although it has been shown that FHV-1 isolation is generally related to the presence of clinical signs, whereas FCV is not (12) . Furthermore, FHV-1 is the virus most commonly identified when respiratory clinical manifestations are observed (7, 13, 16, 34) ; in the present study, signs of respiratory disease were present in 24 out of the 25 sick cats from which FCV, FHV-1 or both were isolated. showing signs of oral lesions in this study (data not shown), which matches results from other studies that have associated oral ulcers more consistently with FCV infection (25, 26, 34) . The higher prevalence of FCV in comparison to FHV-1 isolated in the United Kingdom has been attributed to vaccines that began to be applied in the 1970s (12) . This vaccination likely contributed to the reduction of the number of cats in the Feline calicivirus and feline herpesvirus population excreting FHV-1 but did not significantly affect the number of cats excreting FCV (12) . The biology of the viruses is one characteristic that could contribute to this finding; FCV is an RNA virus with wide genetic and antigenic diversity, whereas FHV-1 is a stable DNA virus (25) . In this article, the number of vaccinated cats excreting FHV-1 was higher than that excreting FCV (Table 2 ). However, it should be noted that there was a unique situation in this study that could cause such results. Most of the isolated FHV-1 came from breeding catteries where an outbreak of respiratory disease was occurring. Thus, many of the FHV-1-isolated cats came from the same location; furthermore, most of the vaccinated cats used in this study came from the same cattery. In addition, FCV was the virus most frequently isolated from cats that did not show signs of disease ( Table 2) . As previously noted, FHV-1 is typically isolated when animals present clinic manifestations, whereas FCV is isolated when they do not (12) . Again, virus biology is the most probable explanation for these results because FCV carriers excrete the virus continuously, whereas FHV-1 carriers excrete the virus only when it is reactivated (11, 13, 25) . The percentages of isolation were roughly 60% for FCV and 27% for FHV-1 (Table 2) . A comparison with other data shows only that FCV is more commonly isolated than FHV-1 because the prevalence detected varied as much as 15 to 25% for FCV and 0.2 to 33.3% for FHV-1 in one study (12) to 25% for FCV and 0.6% for FHV-1 in another study (3) . However, no virus was isolated from approximately 64% of the samples coming from cats showing signs of respiratory disease (Table 1) . This finding could be attributed to problems with sampling and storage conditions or even cases in which cats were sampled late in the course of disease, as has been reported in other studies (2, 29) . Nonetheless, there are other causes for respiratory and ocular diseases in felines other than FCV and FHV-1, including agents such as fungi, bacteria and other viruses (10) . Chlamydophila felis is a bacteria routinely identified in cases of conjunctival disease in cats. Bordetella spp., which is associated with mild respiratory signs, is also commonly identified (2) . Thus, cases of disease in cats from which neither virus was isolated could have other etiologic causes. The age of the individuals was taken in account and when the results were analyzed separately for each virus, it was observed that FCV was primarily isolated from adult cats, whereas FHV-1 was primarily isolated from younger animals (results shown in Table 2 ). A higher prevalence of FCV in adults has been already described (34) . An average age of 38 months for cats was demonstrated positive for FCV, and 29.9 months for FHV-1-positive cats (34) . The results for FHV-1 also match those of previous studies, which isolated FHV-1 from 16.9% of cats between one and three months old, 8.7% from cats between 4 and 11 months old, and less than half the percentage from cats above 11 months of age (2) . As previously noted, the female/male ratio was comparable, whereas the viral isolation frequency was twice as frequent in females as in males (Table 1 and Results). The large amount of positive samples obtained from females was not an expected result. In quite a few studies performed in cat populations from numerous locations, no gender difference was observed (11, 25) . The castration status of the cats appears to play a larger role than the gender in the epidemiology because some researchers have shown a smaller number of positive spayed females and neutered males in comparison to non-castrated cats (2, 30) . The higher prevalence of the virus among noncastrated cats has been attributed to the behavior of these animals; the likelihood of exposure to virus of neutered/spayed animals may be reduced because social interactions are less likely to occur (30) . In the present study, it was not possible to compare the virus distribution among castrated cats because the data were not available. The viruses were more often isolated from cats that shared a habitat (Table 1) . Such results are similar to those demonstrated in surveys performed in European countries (2, Feline calicivirus and feline herpesvirus 13). The high prevalence of these viruses in cats that share a living space is generally attributed to the method of transmission, which requires close contact between infected and susceptible animals (11, 25) . Furthermore, another important characteristic of the Brazilian cat population is that most cats, even ones with owners, live outside. This means that cats are more exposed to infectious diseases that are transmitted by contact than the general population analyzed in surveys performed in the USA and European countries. Taken together, the data shown here provide insight into the epidemiology of FCV and FHV-1 among the cat population in counties in southern Brazil. One point of particular concern is the detection of cats without clinical signs that excrete the viruses. Critically, this occurred even among animals that were vaccinated. Such animals could be a source of infection, particularly for kittens. In addition, because FCV is an RNA virus that varies widely, the molecular characterization of isolates of this virus is underway in our lab, and the data compiled in this article will aid in further studies relating to epidemiology and vaccine efficacy. Both FCV and FHV are prevalent in Southern Brazil despite sporadic vaccination. Cats likely encounter these viruses at a very young age and this contributes to a complicated epidemiology for both these viruses. Of significance cats showing no clinical signs were found to be excreting virus as well as those that had previously been vaccinated against FCV and FHV. This highlights the need for continued research into these important diseases of domesticated cats. Feline respiratory tract disease in Louisiana A study of feline upper respiratory tract disease with reference to prevalence and risk factors for infection with feline calicivirus and feline herpesvirus Isolation of feline respiratory viruses from clinically healthy cats at UK cat shows Lethal outbreak of disease associated with feline calicivirus infection in cats Jaundice in a Siamese cat with in utero feline calicivirus infection First evidence of feline herpesvirus, calicivirus, parvovirus, and Ehrlichia exposure in Brazilian free-ranging felids Isolation and classification of feline picornavirus and herpesvirus in Denmark Feline viral respiratory disease: a review with particular reference to its epizootiology and control Efficacy of an inactivated feline calicivirus (FCV) vaccine against challenge with United Kingdom field strains and its interaction with the FCV carrier state Feline respiratory disease Update Feline herpesvirus Isolation of feline calicivirus and feline herpesvirus from domestic cats 1980 to 1989 Factors associated with upper respiratory tract disease caused by feline herpesvirus, feline calicivirus, Chlamydophila felis and Bordetella bronchiseptica in cats: experience from 218 European catteries Serum survey for antibodies to coronavirus, herpesvirus, calicivirus, and parvovirus in domestics cats from Rio Grande do Sul, Brazil Prevalence of feline herpesvirus 1, feline calicivirus and Chlamydophila felis in clinically normal cats at a Korean animal shelter Prevalence of feline calicivirus, feline leukaemia virus and antibodies to FIV in cats with chronic stomatitis Studies on the role of feline calicivirus in chronic stomatitis in cats Evaluation of serologic and viral detection methods for diagnosing feline herpesvirus-1 infection in cats with acute respiratory tract or chronic ocular disease Detection of feline calicivirus (FCV) from vaccinated cats and phylogenetic analysis of its capsid genes Interaction of a combined feline viral rhinotracheitis-feline calicivirus vaccine and the FVR carrier state An isolated epizootic of hemorrhagic-like fever in cats caused by a novel and highly virulent strain of feline calicivirus A standardized serum neutralization test for feline viral rhinotracheitis. II. The virus-serum system Feline picornavirus infection: the in vivo carrier state Feline calicivirus Acute and chronic faucitis of domestic cats. A feline calicivirus-induced disease Neutralizing antibodies against feline herpesvirus type 1 in captive wild felids of Brazil Differential sensitivity of culture and the polymerase chain reaction for detection of feline herpesvirus 1 in vaccinated and unvaccinated cats Detection of feline calicivirus, feline herpesvirus 1 and Chlamydia psittaci mucosal swabs by multiplex RT-PCR/PCR Feline respiratory viruses -their prevalence in clinically healthy cats Feline calicivirus carrier state. A study of the host/virus relationship Isolation of feline calicivirus from cats in Brazil Effect of a live attenuated intranasal vaccine on latency and shedding of feline herpesvirus 1 in domestic cats Feline herpesvirus 1 and feline calicivirus infections in a heterogeneous cat population of a rescue shelter All the content of the journal, except where otherwise noted, is licensed under a Creative Commons License