Seroprevalence of Toxoplasma gondii and Neospora caninum in feral cats (Felis silvestris catus) in Majorca, Balearic Islands, Spain

Cryptosporidium spp. are common intestinal protozoan parasites that infect a wide range of hosts, including humans and livestock, worldwide. The objective of this study was to determine the prevalence of Cryptosporidium spp. in dairy calves in Prince Edward Island, Canada, and the potential for transmission of this parasite between dairy calves and humans. Fecal samples were collected from 183 dairy calves from 11 farms in Prince Edward Island. The prevalence of Cryptosporidium spp. infections in these animals was determined by examining for the presence of oocysts in the fecal samples, using immunofluorescence microscopy. Molecular characterization was done using a nested-PCR protocol to amplify fragments of the Cryptosporidium heat-shock protein 70 gene, followed by DNA sequencing. Ten calves (6.2%), representing 4 out of 11 farms tested, were positive for Cryptosporidium spp. DNA sequence analysis on five PCR positive samples demonstrated that Cryptosporidium parvum was the only species present in the calves tested, suggesting that there is a potential risk of zoonotic transmission between dairy calves and humans in this region.     

Genetic characterization of 2 novel feline caliciviruses isolated from cats with idiopathic lower urinary tract disease

Feline caliciviruses (FCVs) are potential etiologic agents in feline idiopathic lower urinary tract disease (I-LUTD). By means of a modified virus isolation method, we examined urine obtained from 28 male and female cats with nonobstructive I-LUTD, 12 male cats with obstructive I-LUTD, and 18 clinically healthy male and female cats. All cats had been routinely vaccinated for FCV. Two FCVs were isolated; I (FCV-U1) from a female cat with nonobstructive I-LUTD, and another (FCV-U2) from a male cat with obstructive I-LUTD. To determine the genetic relationship of FCV-U1 and FCV-U2 to other FCVs. capsid protein gene RNA was reverse transcribed into cDNA, amplified, and sequenced. Multiple amino acid sequence alignments and phylogenetic trees were constructed for the entire capsid protein, hypervariable region E, and the more conserved (nonhypervariable) regions A, B, D, and F. When compared to 23 other FCV isolates with known biotypes, the overall amino acid sequence identity of the capsid protein of FCV-U1 and FCV-U2 ranged from 83 to 96%; identity of hypervariable regions C and E ranged from 58 to 85%. Phylogenetically, FCV-U1 clearly separated from other FCV strains in phenograms based on nonhypervariable regions. In contrast, FCV-U2 consistently segregated with the Urbana strain in all phenograms. Clustering of isolates by geographic origin was most apparent in phenograms based on nonhypervariable regions. No clustering of isolates by biotype was apparent in any phenograms. Our results indicate that FCV-UI and FCV-U2 are genetically distinct from other known vaccine and field strains of FCV.     

Detection of virulent feline herpesvirus-1 in the corneas of clinically normal cats

To evaluate the clinically normal feline cornea for the presence of virulent feline herpesvirus-1 (FHV-1), corneas from 31 cats (25 with normal eyes and six with active disease or corneal scarring) euthanased at a shelter were collected. Corneas from two specific pathogen-free cats were included as negative controls. Virus isolation (VI), fluorescent antibody (FA) staining and real-time polymerase chain reaction (rt-PCR) were performed on all samples. The presence or absence of dexamethasone in the media was evaluated for its effect on VI. VI was positive for FHV-1 in six corneas from five cats, all with clinically normal eyes. One cornea was positive for feline calicivirus (FCV) in addition to FHV-1, but only in media that included dexamethasone. Eight corneas were positive on rt-PCR for FHV-1, all from cats with clinically normal eyes. All positive VI samples were confirmed with FA staining. VI and rt-PCR were negative for FHV-1 and FCV in cats with active disease or corneal scarring. Data from this study indicate that virulent FHV-1 and FCV can be present in feline corneas that are clinically normal. Dexamethasone may enhance viral spread through a cell receptor mechanism.     

Molecular subtyping of feline immunodeficiency virus from domestic cats in Australia

OBJECTIVE: To determine the prevalent subtypes of feline immunodeficiency virus (FIV) present in the domestic cat population of Australia. METHOD: Blood samples were collected from 41 FIV antibody positive cats from four cities across Australia. Following DNA extraction, polymerase chain reaction (PCR) was performed to amplify the variable V3-V5 region of the envelope (env) gene. Genotypes were assessed by direct sequencing of PCR products and comparison with previously reported FIV sequences. Phylogenetic analysis allowed classification of the Australian sequences into the appropriate subtype. RESULTS: Of the 41 FIV samples, 40 were found to cluster with previously reported subtype A isolates, whilst the remaining sample grouped within subtype B. CONCLUSIONS: Subtype A was found to be the predominant FIV subtype present in Australia, although subtype B was also found. These results broaden our knowledge of the genetic diversity of FIV and the associated implications for preventative, diagnostic and therapeutic approaches.     

Evaluation of terminal deoxynucleotidyl transferase expression in bone marrow of clinically normal and feline leukemia virus-exposed cats

Bone marrow cells from 8 specific-pathogen-free and 11 feline leukemia virus-exposed cats were examined for the expression of the nuclear antigen terminal deoxynucleotidyl transferase (TdT). Using a standard indirect immunofluorescence technique, feline leukemia-exposed cats had increased expression of TdT in bone marrow aspirates (2.0% to 29.0%) when compared with that in bone marrow cells from specific-pathogen-free cats (2.5% to 6.0%). Seemingly, TdT can be used as an antigenic marker in leukemogenesis of FeLV-infected cats.     

Pattern of seroreactivity against feline foamy virus proteins in domestic cats from Germany

The prevalence of feline foamy virus (FFV, spumaretrovirinae) in naturally infected domestic cats ranges between 30 and 80% FFV positive animals depending on age, sex and geographical region analyzed. Two serotypes have been reported for FFV designated FUV7-like and F17/951-like. Serotype-specific neutralization has been shown to correlate with sequence divergence in the surface (SU) domain of the envelope protein (Env). We analyzed a serum collection of 262 domestic cat sera from Germany using a GST-capture ELISA setup screening for Gag and Bet specific antibodies and identified 39% FFV positive animals. Due to the heterogeneity of the serological samples, cut-offs for Gag and Bet reactivity had to be experimentally determined since application of calculated cut-off values yielded some false-positive results; the new cut-off values turned out to be also fully applicable to a previous study. Using the already established FUV7 ElpSU antigen and the newly cloned and produced F17/951 ElpSU antigen, both consisting of the corresponding ectodomains of the envelope leader protein (Elp) and SU protein, we aimed at the detection of Env-specific antibodies and discrimination between the two known FFV serotypes within the diagnostic FFV ELISA. We validated the ElpSU antigens using cat reference sera of known serotype and screened with this assay domestic cat sera from Germany. Use of the FUV7- and F17/951 ElpSU antigens in ELISA resulted in the detection of Env-specific antibodies in both cat reference sera and sera from domestic cats in Germany, but failed to allow serotyping at the same time.     

Genetic analysis of the RNA polymerase gene of caliciviruses from dogs and cats.

Caliciviruses that infect animals including humans cause a specific disease syndrome in their respective hosts. Feline calicivirus (FCV) is a major pathogen of respiratory disease of cats, and human caliciviru is a causative agent of diarrhea. It has been suggested, furthermore, that FCV and newly recognized canine calicivirus (CaCV) may also be possible causes of diarrhea in these animal species. In this study nucleotide sequence of the RNA polymerase gene of two caliciviruses of canine origin, namely CaCV strain No. 48 and FCV-like strain Sapporo/283, and a number of FCV strains of respiratory and enteric origins was examined. The length of sequenced region, from the 5'LKDEL motif through the 3'YGDD motif of the gene, was 555 bp for CaCV No. 48 strain and 552 bp for the other FCV strains including Sapporo/283 strain. In phylogenetic analysis, CaCV No. 48 strain grouped as a distinct branch sharing ancestral roots with San Miguel sea lion virus, and FCVs formed one compact group in which Sapporo/283 strain was included.     

Frequent detection of papillomavirus DNA in clinically normal skin of cats infected and noninfected with feline immunodeficiency virus

Feline cutaneous squamous cell carcinomas (SCCs) often contain felis domesticus papillomavirus type 2 (FdPV‐2) DNA. While this may suggest FdPV‐2 causes feline SCC development, the proportion of cats that are asymptomatically infected by this PV is unknown. Infection by feline immunodeficiency virus (FIV) is associated with high rates of cutaneous SCC development, possibly due to increased PV infection. This study examines the frequency of cutaneous asymptomatic FdPV‐2 infections in cats and compares the rate of FdPV‐2 infection in 22 FIV‐positive cats with that in 22 FIV‐negative cats. FdPV‐2 sequences were detected in 39% of skin swabs. One or both swabs contained FdPV‐2 DNA from 52% of the cats. FIV status, age or sex of the cat did not significantly influence FdPV‐2 infection. Cats that shared a household with a PV‐infected cat could remain uninfected suggesting infection depends more on host factors than exposure to the PV. These results indicate that asymptomatic FdPV‐2 infections are common in cats, but do not provide evidence that FdPV‐2 causes feline SCC development.     

The experimental transmission of various mycoplasmas of feline origin to domestic cats (Felis catus)

Mycoplasma felis and M. gatae recovered from domestic cats, M. arginini from a cheetah, an unidentified glucose-fermenting mycoplasma from a lion, and M. pulmonis from a rat were inoculated into specific pathogen free (SPF) kittens by the oral, ocular and nasal routes. During the 21-day experimental period a persistent infection was established in a proportion of the kittens challenged with M. felis, M. gatae and M. arginini. The mycoplasma isolated from the lion created a transitory infection in one kitten, but M. pulmonis could not be recovered even 24 hr after inoculation. None of the organisms investigated showed any pathological effect. The implications of this work in relation to the host-specificity and pathogenicity of the feline mycoplasmas are discussed. Résumé. On a inoculé à des chatons axéniques par les voies orales, oculaires et nasales, un Myco-plasmafelis et un M. gatae recueillis de chats domestiques, un M. arginini recueilli d'un guépard, un mycoplasme non identifié à glucose fermentant recueilli d'un lion et un M. pulmonis recueilli d'un rat. Durant la période expérimentale de 21 jours, on a établi une infection persistante dans une proportion des chatons soumis aux tests de Mycoplasma felis, de M. gatae et de M. agrinini. Le mycoplasma isolé recueilli du lion a créé une infection transitoire chez un chaton, mais le M. pulmonis n'a pu être recueilli, même 24 heures après l'inoculation. Aucun des organismes sous investigation n'a montré d'effets pathologiques. On a discuté des implications de ce travail en relation avec la spécifité d'hôtes et la pathogénicité de mycoplasmes félins. Zusammenfassung. Mycoplasma felis und M. gatae von Hauskatzen erlangt, M. arginini von einem Tschita, ein unidentifiziertes Glukose-fermentierendes Mycoplasma von einem Löwen, und M. pulmonis von einer Ratte wurden in keimfreie (SPF) junge Katzen geimpft, auf oralem, Augen- und Nasenweg. Während der 21 tägigen Versuchsperiode wurde eine anhaltende Infektion in den Katzen festgestellt, die dem M. felis, M. gatae und dem M. agrinini ausgesetzt waren. Das Mycoplasma von dem Löwen abgesondert, verursachte eine vorübergehende Infektion eines Kätzchens, aber M. pulmonis konnte selbst 24 Stunden nach der Impfung nicht gefunden werden. Keines der untersuchten Organismen zeigte irgendwelche pathologischen Effekte. Die Folgerungen dieser Arbeit im Verhältnis zu der Gast-Spezifität und Pathogenizität des Katzen-mycoplasmas sind diskutiert.     

Bacteroides species from the oral cavity and oral-associated diseases of cats

One hundred and sixty-seven strains of Bacteroides were isolated from 71 subcutaneous fight-wound abscesses of cats, 21 cases of feline pyothorax, normal gingival margins from 10 cats and 6 cases of feline gingivitis. Bacteroides species constituted (as a proportion of all anaerobic isolates examined) 44.5% from subcutaneous abscesses, 33.7% from pyothoraxes, 37.5% from normal gingiva and 27.7% from diseased gingiva. Bacteroides tectum comprised 43.7% or 73 of 167 strains, followed by the black- or brown-pigmented asaccharolytic feline species of B. gingivalis, B. salivosus and Group B, comprising 32.3% or 54 of 167 strains. B. heparinolyticus (some 10% or 17 of 167 strains) was the next most common species described. The remainder consisted of two strains of B. fragilis and 21 unspeciated strains. Bacteroides tectum was frequently isolated from subcutaneous abscesses (43.7%) and pyothoraxes (46.6%), and it constituted some 33% of anaerobic isolated from normal gingiva. Bacteroides heparinolyticus was more commonly encountered in purulent lesions (abscesses and pyothoraxes) than in the oral cavity.     

Phylogenetic analysis of feline panleukopenia virus (FPLV) strains in Korean cats

Sixteen Korean feline panleukopenia virus (FPLV) strains were compared with 48 non-Korean strains and two vaccine strains to conduct phylogenetic analysis of the FPLVs currently circulating among cats in Korea. Most of the residues that discriminate between FPLVs and canine parvoviruses (CPV-2, -2a, -2b, and -2c), including 80-Lys, 93-Lys, 103-Val, 323-Asp, 564-Asn, and 568-Ala, were conserved in the Korean FLPVs; however, exceptions were observed in two strains, namely K50/08 (80-Gln) and V142 (323-Asn). Phylogenetic analysis using the Bayesian inference and Neighbor-joining method showed that FPLVs were not segregated on a clear temporal or geographical basis. Three clusters (G1, G2, and G3) were formed by the VP2 nucleotide sequences analysed and Korean strains belonged to the G1 (n = 13) and G2 (n = 3) clusters. The ratio of non-synonymous to synonymous substitutions (dN/dS) revealed that purifying selection acts on the VP2 gene of Korean FPLVs.     

Isolation of Corynebacterium pseudotuberculosis from the cervical canal of clinically normal sows

Two isolates from the cervical canal of clinically normal sows were indistinguishable from Corynebacterium pseudotuberculosis in their biochemical properties and genetic homology by dna-dna hybridisation. Furthermore, three sheep inoculated with one of the isolates developed typical lesions of caseous lymphadenitis and gave antibody responses specifically to C pseudotuberculosis.     

Serological differentiation of FIV-infected cats from dual-subtype feline immunodeficiency virus vaccine (Fel-O-Vax FIV) inoculated cats

Feline immunodeficiency virus (FIV) vaccine, Fel-O-Vax FIV, was released for sale in the US in 2002. The antibodies of vaccinated cats interfere with serological assays by currently available FIV diagnostic kits. In this study, we investigated whether it is possible to distinguish serologically cats vaccinated with Fel-O-Vax FIV from cats experimentally or naturally infected with FIV. A total of 153 sera taken from 97 cats were used as serum samples. Enzyme linked immunosorbent assay (ELISA) was performed using whole FIV antigen and formalin treated whole FIV antigen, recombinant-gag (r-gag) antigen, and transmembrane (TM) peptide. Statistical analysis was performed using ELISA optical density (O.D.) values obtained with each antigen as variables. Except for the ELISA O.D. values obtained with r-gag antigen, a significant difference in ELISA O.D. values was observed between the vaccinated and the infected groups. However, it was not possible to distinguish both groups unequivocally. Using discriminant analysis, it was possible to distinguish the two groups with an accuracy of 97.1% with two discriminating variables (ELISA O.D. values obtained with formalin treated whole FIV antigen, and TM peptide), 97.8% with three discriminating variables (ELISA O.D. values obtained with whole FIV antigen, formalin treated whole FIV antigen, and TM peptide). Therefore, it was considered possible to distinguish cats vaccinated with Fel-O-Vax FIV from FIV-infected cats by ELISA using two types of antigens including formalin treated whole FIV antigen and TM peptide, or three types of antigens including formalin treated whole FIV antigen, TM peptide and whole FIV antigen.     

Therapeutic effects of recombinant feline interferon-omega on feline leukemia virus (FeLV)-infected and FeLV/feline immunodeficiency virus (FIV)-coinfected symptomatic cats

The clinical efficacy of a recombinant feline interferon, rFeIFN-omega, was evaluated for the treatment of cats presented with clinical signs associated with feline leukemia virus (FeLV) infection and FeLV/feline immunodeficiency virus (FIV) coinfection in the field. In this multicentric, double-blind, placebo-controlled trial, 81 cats meeting the inclusion criteria were randomly placed into 2 groups and treated subcutaneously with rFelFN-omega (1 million [M]U/kg per day) or placebo once daily for 5 consecutive days in 3 series (day 0, 14, 60). The cats were monitored for up to 1 year for clinical signs and mortality. During the initial 4-month period, interferon (IFN)-treated cats (n = 39) had significantly reduced clinical scores compared with placebo (n = 42), with all cats having received concomitant supportive therapies. Compared with the control, the IFN-treated group showed significantly lower rates of mortality: 39% versus 59% (1.7-fold higher risk of death for controls) at the 9-month time point and 47% versus 59% (1.4-fold higher risk of death for controls) at the 12-month time point. The IFN treatment was associated with minor but consistent improvement in abnormal hematologic parameters (red blood cell count, packed cell volume, and white blood cell count), apparently underlying the positive effects of IFN on clinical parameters. These data demonstrate that rFeIFN-omega initially has statistically significant therapeutic effects on clinical signs and later on survival of cats with clinical signs associated with FeLV infection and FeLV/FIV coinfection.     

Long-term analysis of feline calicivirus prevalence and viral shedding patterns in naturally infected colonies of domestic cats

Feline calicivirus (FCV) is a highly infectious respiratory pathogen of domestic cats. The prevalence of FCV in the general cat population is high, particularly in multi-cat households, largely because many clinically recovered cats remain persistently infected carriers. In order to assess how FCV circulates in such groups and to assess the contribution that each individual animal makes to the epidemiology of the disease, we have carried out the first detailed analysis of long-term shedding patterns of FCV in individual cats within naturally infected colonies. The prevalence of FCV in each of the groups on individual sampling occasions ranged from 0% to 91%, with averages for the individual colonies ranging from 6% to 75%. Within each of the colonies, one to three distinct strains of FCV were identified. Individual cats showed a spectrum of FCV shedding patterns over the sampling period which broadly grouped into three categories: those that shed virus relatively consistently, those that shed virus intermittently, and those that appeared never to shed virus. This is the first report identifying non-shedder cats that appear resistant to FCV infection over long periods of time, despite being continually exposed to virus. Such resistance appeared to be age related, which may have been immune-mediated, although by analogy with other caliciviruses, factors such as host genetic resistance may play a role. Given that a proportion of the population appears to be resistant to infection, clearly the cohort of cats that consistently shed virus are likely to provide an important mechanism whereby infection can be maintained in small populations.     

Experimental infection of domestic cats (Felis domesticus) with Cytauxzoon manul from Pallas' cats (Otocolobus manul)

In a random, blind study, six domestic cats were assigned to two treatment groups that received either sterile water or dexamethasone by subcutaneous injection prior to intravenous inoculation with Pallas' cat (Otocolobus manul) blood infected with Cytauxzoon manul. A seventh domestic cat served as a control and was inoculated only with sterile water. Cats were monitored for clinical signs consistent with cytauxzoonosis, and periodically screened for hemoparasitemia. All domestic cats (6/6) that received Pallas' cat blood infected with C. manul developed a low but detectible parasitemia by 9 days post-inoculation, yet remained clinically healthy. All domestic cats (7/7) were subsequently challenged with Cytauxzoon felis and developed clinical signs typical of cytauxzoonosis within 5 days post-challenge. Affected animals were euthanized and cytauxzoonosis was confirmed by histopathology. While inoculation of domestic cats with Pallas' cat blood infected with C. manul induced a parasitemia, it did not cause disease or provide protection against challenge with C. felis. Further studies are warranted to determine the potential for interspecies transmission and disease with C. manul.     

Nonsense mutation of feline beta-hexosaminidase beta-subunit (HEXB) gene causing Sandhoff disease in a family of Japanese domestic cats

G(M2) gangliosidoses are inherited metabolic disorders and are caused by severely reduced enzymatic activity of lysosomal beta-hexosaminidase. In the present study, the open reading frame (ORF) of the HEXB gene in a family of Japanese domestic cats with G(M2) gangliosidosis variant 0 (Sandhoff disease) was determined. Two types of abnormal cDNA clones were obtained from the liver of an affected cat tissue. One showed a single nucleotide substitution from C to T at nucleotide position 667 of the HEXB ORF. In the deduced amino acid sequence, the codon of arginine was altered to a stop codon. The genotyping, using PCR-primer introduced restriction analysis confirmed that Sandhoff disease in this family is associated with this nonsense mutation. Discovery of the nonsense mutation will permit the confirmation of the clinical diagnosis of Sandhoff disease in conjugation with the already established enzyme-based test.