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The aim of this study was to determine the prevalence and risk factors for Mycoplasma haemofelis (Mhf) and 'Candidatus Mycoplasma haemominutum' (Mhm) infections in domestic cats tested for feline immunodeficiency virus (FIV) and feline leukemia virus (FeLV) with a commercial enzyme-linked immunosorbent assay (ELISA) kit. Based on serological testing, cats were grouped as i) FIV-positive (n=25); ii) FeLV-positive (n=39); iii) FIV/FeLV-positive (n=8); and iv) FIV/FeLV-negative (n=77). Complete blood counts were followed by DNA extraction, species-specific polymerase chain reaction (16S rRNA gene) for Mhf and Mhm and Southern blotting for all animals. Mhf DNA was found in 4.0, 2.6, 12.5 and 7.8% of the cats from groups i, ii, iii and iv, respectively, while 32, 5.1, 50 and 5.2% of these animals had an Mhm infection. Cats with FIV (OR=4.25, P=0.009) and both FIV and FeLV (OR=7.56, P=0.014) were at greater risk of being hemoplasma infected than retroviral-negative cats, mainly due to Mhm infection (OR=8.59, P=0.001 and OR=18.25, P=0.001, respectively). Among pure-breed cats, FIV-positive status was associated with hemoplasma infection (OR 45.0, P=0.001).

A four-year-old, female neutered domestic shorthair cat had a history of chronic intermittent vomiting and lymphocytosis. B cell chronic lymphocytic leukaemia was diagnosed by flow cytometry, which revealed abnormally large numbers of mature B lymphocytes in the peripheral blood. The cat was treated conservatively with antiemetic drugs and remained stable without chemotherapy for over a year.

Fourteen cDNA clones encoding epitopes of proteins of Toxoplasma gondii feline enteroepithelial-stages parasites were isolated and expressed in Escherichia coli in an effort to determine the antigenecity of the parasites. Sequence analysis showed that four of the cDNA clones had a 930-bp open-reading frame encoding a product showing similarity to the 14-3-3 protein mRNA sequence.(1) Southern hybridization of DIG-labeled positive clone with T. gondii genomic DNA cleaved with EcoRI, BamHI and HindIII resulted in one or two bands in each case. In an immunofluorescence assay, polyclonal and monoclonal antibodies raised against the expressed protein showed strong reactivity with feline enteroepithelial-stages parasites and sporozoites. In a complementation assay in which a plasmid carrying the protein-coding region of the isolated cDNA was introduced into a Saccharomyces cerevisiae mutant, strain DS9-22, the expressed protein showed complementation of the function of the 14-3-3 protein in yeast transformants. These findings suggest that T. gondii parasites produce a protein showing partial homology with members of the 14-3-3 protein family and this protein is expressed in feline enteroepithelial-stages parasites.

The clonality analysis of the bone marrow cells was carried out by detecting the integrated proviruses of feline leukemia virus (FeLV) to understand the pathogenesis of FeLV-associated hematopoietic disorders in cats. Bone marrow cells from 4 cases with acute myeloid leukemia (AML), 9 cases with myelodysplastic syndromes (MDS), 2 cases with pure red cell aplasia (PRCA) and 3 healthy carriers infected with FeLV were subjected to Southern blot analyses using an exogenous FeLV probe. Clonal hematopoiesis was found in all the cases with AML and in 6 of the 9 cases with MDS, but not in the cases with both PRCA and healthy carriers infected with FeLV. In the 2 cases with MDS, it was thought that the same clones of the hematopoietic cells might proliferate before and after the progression of the disease irrespective of the changes of the hematological diagnoses by cytological examination. This study indicates that MDS in cats is a disease manifestation as a result of clonal proliferation of hematopoietic cells and can be recognized as a pre-leukemic state of AML.

A cosmid clone bank of Bordetella bronchiseptica genomic DNA was screened for the presence of type III secretion (TTS) genes using a probe derived from the TTS system genes of Ralstonia solanacearum. A 3.35kb PstI fragment, sub-cloned from a hybridising cosmid clone, was sequenced and found to contain a 97bp overlap with the previously reported B. bronchiseptica bscIJKLNO TTS gene cluster. DNA and predicted protein homology analysis suggests that a bscPQRST cluster lies immediately downstream of bscIJKLNO. A PCR amplification assay indicated that the bscT locus was present in 27 B. bronchiseptica animal isolates tested (100%). Dot-blot DNA hybridisation using probes for bscT and bscP confirmed the presence of these loci in six canine isolates associated with a variety of clinical signs. Although TTS has been implicated in the pathogenicity of B. bronchiseptica, it is likely that different clinical manifestations may be due to variations in gene expression or host factors, rather than the absence or presence of TTS genes.

Papillomaviruses (PVs) are highly species- and site-specific pathogens of stratified squamous epithelium. Although PV infections in the various Felidae are rarely reported, we identified productive infections in six cat species. PV-induced proliferative skin or mucous membrane lesions were confirmed by immunohistochemical screening for papillomavirus-specific capsid antigens. Seven monoclonal antibodies, each of which reacts with an immunodominant antigenic determinant of the bovine papillomavirus L1 gene product, revealed that feline PV capsid epitopes were conserved to various degrees. This battery of monoclonal antibodies established differential expression patterns among cutaneous and oral PVs of snow leopards and domestic cats, suggesting that they represent distinct viruses. Clinically, the lesions in all species and anatomic sites were locally extensive and frequently multiple. Histologically, the areas of epidermal hyperplasia were flat with a similarity to benign tumors induced by cutaneotropic, carcinogenic PVs in immunosuppressed human patients. Limited restriction endonuclease analyses of viral genomic DNA confirmed the variability among three viral genomes recovered from available frozen tissue. Because most previous PV isolates have been species specific, these studies suggest that at least eight different cat papillomaviruses infect the oral cavity (tentative designations: Asian lion, Panthera leo, P1PV; snow leopard, Panthera uncia, PuPV-1; bobcat, Felis rufus, FrPV; Florida panther, Felis concolor, FcPV; clouded leopard, Neofelis nebulosa, NnPV; and domestic cat, Felis domesticus, FdPV-2) or skin (domestic cat, F. domesticus, FdPV-1; and snow leopard, P. uncia, PuPV-2).

Borna disease virus (BDV) infection has been suggested to cause spontaneous neurological disease in cats referred to as staggering disease. However the evaluation of BDV infection in neurologically asymptomatic cats remained unclear. In the present study, BDV infected, asymptomatic cats in Tokyo were surveyed both by the presence of plasma antibodies against BDV-p24 and -p40 and by RNA detection in peripheral blood mononuclear cells. Seven of 32 domestic cats (21.9%) were serologically or genetically judged to be BDV-infected. Six cats were positive for anti-BDV antibody and two cats were positive for BDV RNA. Within the 2 RNA-positive cats, only one was positive for anti-BDV antibodies. Furthermore, the findings of anti-BDV-p40 and anti-BDV-p24 antibody-positive cats did not completely overlap. These results suggest that there are neurologically asymptomatic domestic cats infected with BDV present in the Tokyo area.

Digoxigenin labelled whole chromosomal DNA probes directed against three feline members of the genus Porphyromonas (P. gingivalis VPB 3492, P. circumdentaria NCTC 12469T and P. salivosa VPB 3313) were used to identify and quantify organisms in samples taken from the gingival margins of 40 domestic cats with different grades of periodontal disease. At the right upper canine tooth, the grade of periodontal disease ranged from 0 to 5 and the cfu of facultative/obligate anaerobes ranged from 5.5 x 10(4) to 2.0 x 10(6)). In 38 of the 40 cats, at least one of the three Porphyromonas species was isolated and regression analysis showed that the cfu of total Porphyromonas sp. was a highly significant indicator of the grade of periodontal disease (p < 0.001, R2 0.510). Feline P. gingivalis was isolated from 37 of the 40 cats and regression analysis showed that it was a highly significant predictor of the grade of periodontal disease (p < 0.001, R2 0.561). The cfu of P. salivosa was a significant predictor of the grade of periodontal disease (p < 0.001, R2 0.286) and regression analysis showed that there was a significant positive relationship between cfu of P. circumdentaria and grade of periodontal disease (p = 0.018, R2 0.116). The periodontal grades at the right upper third premolar tooth ranged from 0 to 6. The cfu of facultative/obligate anaerobes isolated ranged from 1.2 x 10(5) to 7.9 x 10(6), and regression analysis showed that cfu was a significant predictor of periodontal grade (p < 0.001, R2 0.378). The cfu of total Porphyromonas species ranged from 1.2 x 10(4) to 1.7 x 10(6) and regression analysis of the cfu against the grade of periodontal disease showed a highly significant association (p < 0.001, R2 0.633). The cfu of P. gingivalis ranged from 0 to 1.1 x 10(6) and regression analysis of the cfu of P. gingivalis against the grade of periodontal disease showed a highly significant association (p < 0.001, R2 0.439). The cfu of P. salivosa was a significant predictor of the grade of periodontal disease (p < 0.001, R2 0.479) and the same association was found between cfu of P. circumdentaria and grade of periodontal disease (p = 0.002, R2 0.204). This study has established Porphyromonas as anumerically significant and highly prevalent genus in feline periodontal disease.

Samples were examined from 15 subcutaneous fight wound abscesses from 15 cats. All abscesses were closed at the time of sampling and cats had received no prior treatment. Samples were processed within 20 min and quantitative assessment made of total facultative and obligately anaerobic flora isolated. Digoxigenin labelled whole chromosomal DNA probes directed against three feline members of the genus Porphyromonas (P. gingivalis VPB 3492, P. circumdentaria NCTC 12469T and P. salivosa VPB 3313) were used to identify members of this genus and quantification of these species was made from each cat using colony lifts and southern hybridisation from nitrocellulose membranes taken from replicate plates from each abscess sample. Twelve of the 15 abscesses yielded a variety of facultative and obligately anaerobic (FOA) bacterial species and members of the genus Porphyromounas were enumerated from each of these 12 abscesses. Of the 12 abscesses in which Porphyromonas species were detected, seven contained one species only (five contained only P. gingivalis and two contained only P. salivosa) three abscesses contained two species (both P. gingivalis and P. circumdentaria) and two abscesses contained all three species of Porphyromonas. These results show that members of the genus Porphyromonas are likely to be significant contributors to the purulent disease process in subcutaneous abscesses in cats.

The purpose of this study was to determine the temporal appearance of T. gondii in aqueous humor of cats orally inoculated with T. gondii using polymerase chain reaction (PCR) for the detection of the B1 gene. Serum and aqueous humor were collected from five SPF cats prior to oral inoculation with T. gondii and days 7, 14, 21, 28, 42, 84, 140, 147, 154, 161, 168, and 182 after inoculation. Cats were inoculated orally with T. gondii tissue cysts on day 0 and day 140. T. gondii-specific IgM and IgG were measured in serum and aqueous humor from the cats at each sample data. T. gondii B1 gene PCR was performed on all the aqueous humor samples and the amplified DNA was detected by Southern blotting. Chorioretinitis developed in three out of the five cats, but anterior uveitis was not detected. All cats developed T. gondii-specific IgG titers in serum, and had T. gondii-specific IgG C-values > 1 in both eyes at varying times during the study. T. gondii was detected by PCR and Southern blotting in aqueous humor in both eyes of all cats at times varying from days 14-84 after primary inoculation and days 14-42 after challenge inoculation.

The recent isolation of Helicobacter pylori from cats obtained from a commercial supplier has potentially important public health implications. The present study investigated whether H. pylori infection was common in stray cats. Twenty-five cats were examined for the presence of H. pylori by histological examination, culture and two polymerase chain reaction (PCR) assays. Histologically, the gastric biopsy specimens from all cats showed large spiral organisms typical of H. felis and not H. pylori. Samples from 23 cats yielded bacterial growth and two had no growth. Colonies grossly similar to H. pylori were tested for catalase, oxidase, urease and Gram's stain reactions. None was H. pylori. All samples tested as positive by the Helicobacter 16S rRNA genus-specific PCR assay and only six cats and a mouse stomach infected with H. heilmannii gave positive results with the adhesin subunit A (hpaA)-specific PCR assay, which is consistent with either H. pylori or H. heilmannii. The helicobacters identified in these samples by PCR were not cultivable and hence were probably H. heilmannii. H. pylori infection is uncommon in stray cats and owning pet cats should not be a threat to public health in relation to H. pylori infection.

A polymerase chain reaction (PCR) assay was developed and used to detect feline herpesvirus-1 (FHV-1) in conjunctival and oropharyngeal swabs, and in latently infected tissues (trigeminal ganglia, optic nerves, optic chiasma, olfactory bulbs and corneas) collected from 10 experimentally infected cats. There was good agreement between parallel tests of the swab specimens by PCR and virus isolation assay during the phase of acute, latent and recurrent disease episodes (kappa = 0.63, P < 0.001). The PCR reliably detected < or = 240 copies of FHV-1 template DNA, significantly improving upon previously published PCR assays for the agent.