Mitika Kuribayashi Hagiwara

Experimental infection of dogs with a Brazilian strain of Rickettsia rickettsii: clinical and laboratory findings

The bacterium Rickettsia rickettsii is the etiological agent of an acute, severe disease called Rocky Mountain spotted fever in the United States or Brazilian spotted fever (BSF) in Brazil. In addition to these two countries, the disease has also been reported to affect humans in Mexico, Costa Rica, Panama, Colombia and Argentina. Like humans, dogs are also susceptible to R. rickettsii infection. However, despite the wide distribution of R. rickettsii in the Western Hemisphere, reports of R. rickettsii-induced illness in dogs has been restricted to the United States. The present study evaluated the pathogenicity for dogs of a South American strain of R. rickettsii. Three groups of dogs were evaluated: group 1 (G1) was inoculated ip with R. rickettsii; group 2 (G2) was infested by R. rickettsii-infected ticks; and the control group (G3) was infested by uninfected ticks. During the study, no clinical abnormalities, Rickettsia DNA or R. rickettsii-reactive antibodies were detected in G3. In contrast, all G1 and G2 dogs developed signs of rickettsial infection, i.e., fever, lethargy, anorexia, ocular lesions, thrombocytopenia, anemia and detectable levels of Rickettsia DNA and R. rickettsii-reactive antibodies in their blood. Rickettsemia started 3-8 days after inoculation or tick infestation and lasted for 3-13 days. Our results indicate that a Brazilian strain of R. rickettsii is pathogenic for dogs, suggesting that canine clinical illness due to R. rickettsii has been unreported in Brazil and possibly in the other South American countries where BSF has been reported among humans.

Diagnóstico sorológico de erliquiose canina com antígeno brasileiro de Ehrlichia canis

The present study describes a successful isolation of Ehrlichia canis and its establishment in DH82 cells, followed by the development of an Indirect Fluorescent Antibodies Test (IFAT). Leukocytes collected from an experimentally infected dog with the Jaboticabal strain of E. canis were used to inoculate a DH82 cell monolayer. Two weeks later, the inoculated culture was checked for infectivity, every 5-6 days by both cytological staining and PCR, targeting a fragment of the dsb gene. The cell culture showed to be infected by Ehrlichia on day 27 by PCR and on day 28 by cytological staining. By the day 33, the infection rate reached 20% and on day 53, 60%. Currently, the isolate is established in DH82 cells, with several passages reaching 90-100% of infected cells, within 7 to 10 days post inoculation. After sequencing, the amplicon was identical to other E. canis corresponding sequences available in the GenBank. DH82 infected cells were used to standardize an IFAT for the diagnosis of canine ehrlichiosis.

Humoral immunity and reinfection resistance in dogs experimentally inoculated with Babesia canis and either treated or untreated with imidocarb dipropionate

It is proposed that the chronic asymptomatic carrier state produced by Babesia canis infection could make dogs more resistant against subsequent infections. This suggests that treatment with imidocarb dipropionate, which removes the organism, can make dogs more susceptible to reinfection in a short period of time. Ten male and female dogs of approximately 4-5 months of age were inoculated with B. canis. Half of them received treatment with imidocarb dipropionate (7 mg/kg) on days 15 and 27 post-infection and the other half were untreated. All the animals were examined using clinical and laboratory methods (CBC, platelet counts and serological study by indirect immunofluorescence test) for a 6-month period. Antibodies were first detected on day 7 post-injection and remained at high levels (1:2560) over the period in the non-treated group. This result was significantly different (P<0.001) from the treated group in which antibodies titers declined after day 34 post-infection. Six months later, after a homologous challenge infection only the dogs of treated group showed parasitaemia, thrombocytopenia and splenomegaly, which was significantly different (P<0.05) from the non-treated group. The sterilizing treatment with imidocarb dipropionate was effective in clearing the infection, but inhibited the maintenance of protective antibodies, making the animals more susceptible to reinfection.

Toxoplasma gondii infection in Brazilian domestic outpatient cats

The occurrence of Toxoplasma antibodies in domestic outpatient cats in the city of São Paulo was evaluated using the indirect immunofluorescence assay. A total of 248 blood samples obtained from male and female cats seen at the Veterinary Teaching Hospital at the University of São Paulo between February 1996 and January 1997 were tested. Of these, 17.7% were positive, with a 64 titer being detected in most animals. The frequence of Toxoplasma antibodies was significantly higher in older cats, those fed raw meat and those with free access to the outdoor environment. There was no significant difference in reactivity between males and females. We conclude that diet and free access to the outdoor environment were equally important as predisposing factors to the risk of Toxoplasma infection.

Borrelia burgdorferi antibodies in dogs from Cotia county, São Paulo State, Brazil

Dogs sera samples collected from Cotia County, São Paulo were tested using indirect immunoenzymatic test (ELISA) in order to study Lyme disease serology in dogs. ELISA method was standardized and G39/40 North American strain of Borrelia burgdorferi was used as antigen. Positive results were confirmed employing the Western blotting technique. Because of the possibility of cross-reactions, sera were also tested for different serological strains of Leptospira interrogans and L. biflexa using microscopic sera agglutination test. Twenty-three of 237 (9.7%) serum samples were positive in the ELISA; 20 of them (86.9%) were confirmed by the Western blotting, what suggests that Cotia may be a risk area for Lyme disease. Although 4 samples (1.7%) were positive for Lyme disease and leptospirosis, no correlation was found between the results (X(2) = 0.725; p = 0.394) what suggests absence of serological cross reactivity.

Estudo clínico da doença do trato urinário inferior em gatos domésticos de São Paulo

Fifty cats with signs of lower urinary tract disease (hematuria, dysuria, pollakiuria or urethral obstruction) were studied as far as clinical, laboratory and radiographic abnormalities were concerned. These animals were distributed in two groups, the first one comprised thirty-six male cats with urethral obstruction and the second, fourteen male and female cats presenting hematuria, dysuria and/or pollakiuria. Another group of twenty-five healthy cats, both sexes, different ages and breeds, fed exclusively with commercial dried food was included as control. Serum urea and creatinine measurements, urine analysis, urine culture and excretory urography were performed in all cats. The results showed that urinary tract infection was rarely seen. No significant difference was detected between castrated or intact animals. Among the sick cats the most consistent abnormalities found were: urinary alkaline pH, crystalluria and thickness of the urinary bladder wall. The difference in urine pH observed among healthy and sick animals fed with the same kind of meal should be stressed and deserves further studies.

Ocorrência de anticorpos antitoxoplasma em gatos infectados naturalmente pelo vírus da imunodeficiência dos felinos

Foram avaliadas, pela tecnica de imunofluorescencia indireta, a frequencia e a magnitude dos titulos de anticorpos antitoxoplasma (Toxoplasma gondii, Nicolle; Manceaux, 1909), em gatos infectados naturalmente pelo virus da imunodeficiencia dos felinos (VIF). Utilizaram-se 115 amostras de soro sanguineo de gatos negativos ao virus da leucemia felina que foram divididas em 3 grupos. Os 22 animais do grupo I eram positivos ao VIF. Os 58 animais que compuseram o grupo II eram doentes porem negativos na pesquisa de anticorpos anti-VIF e os 35 felinos do grupo III eram higidos e negativos ao VIF. Os resultados obtidos permitiram concluir que a frequencia de anticorpos antitoxoplasma foi maior no grupo I do que nos grupos II e III. A analise estatistica mostrou forte associacao entre a infeccao pelo VIF e a presenca de anticorpos antitoxoplasma. Nao se observou diferenca entre a magnitude dos titulos de anticorpos antitoxoplasma nos animais positivos e negativos ao VIF. Embora gatos que desenvolvam imunidade raramente eliminem oocistos, nao se sabe exatamente como esta imunidade pode influenciar a eliminacao de oocistos naqueles gatos infectados pelo VIF. Em face da alta frequencia de anticorpos antitoxoplasma observada nos animais positivos ao VIF, acredita-se que todos os gatos positivos a esse virus devam ser avaliados quanto a presenca de anticorpos antitoxoplasma.

Experimental infection and horizontal transmission of Bartonella henselae in domestic cats

In order to study B. henselae transmission among cats, five young cats were kept in confinement for two years, one of them being inoculated by SC route with B. henselae (10(5) UFC). Only occasional contact among cats occurred but the presence of fleas was observed in all animals throughout the period. Blood culture for isolation of bacteria, PCR-HSP and FTSZ (gender specific), and BH-PCR (species-specific), as well as indirect immunofluorescence method for anti-B. henselae antibodies were performed to confirm the infection of the inoculated cat as well as the other naive cats. Considering the inoculated animal, B. henselae was first isolated by blood culture two months after inoculation, bacteremia last for four months, the specific antibodies being detected by IFI during the entire period. All contacting animals presented with bacteremia 6 months after experimental inoculation but IFI did not detect seroconversion in these animals. All the isolates from these cats were characterized as Bartonella (HSP and FTSZ-PCR), henselae (BH-PCR). However, DNA of B. henselae could not be amplified directly from peripheral blood by the PCR protocols used. Isolation of bacteria by blood culture was the most efficient method to diagnose infection compared to PCR or IFI. The role of fleas in the epidemiology of B. henselae infection in cats is discussed.

Histopatologia e morfometria da bexiga de gatos com doença idiopática do trato urinário inferior (DITUI)

This study was carried out to evaluate cystitis of cats with idiopathic lower urinary tract disease but without signs of the disease at the time of the study. Histopathologic and morphometric analysis of the urinary bladder were done in 15 cats, eight with the disease but with no clinical signs and seven healthy cats. There was bladder mucosal and muscle layers thickening with intense mononuclear inflammatory infiltrate, subepithelial petechiae, hyperplasia and epithelial wrinkles among the cats with the lower urinary tract disease, these findings suggest chronic cystitis.

Avaliação da resposta imune humoral em cães jovens imunizados contra a cinomose com vacina de vírus atenuado

This study was carried out in order to investigate the kinetics of the immune response in puppies vaccinated to canine distemper, with attenuated distemper virus vaccine, by serum-neutralization test. Eleven healthy Beagle dogs, observed from birth to 30 months old, isolated far from other dogs, received monovalent living attenuated canine distemper vaccine at 75, 105 and 135 days old, and twelve months after the third dose of vaccine. Antibodies development were measured by serum-neutralization immediately before vaccination, at the 30th day after the first and the third dose administration, at the 90th day after the third dose administration, nine and twelve months after the annual dose. There was no canine distemper antibody detectable titer before primevaccination. The antibodies titer ranged from 4,047 to 4,880 neutralizing doses at the 30th day after the first dose administration and six months after annual dose, respectively. All dogs showed a similar response, although with variation, with antibody production of higher antibody titers than 2, which is considered the minimum protection titer. Twelve months after annual revaccination, the majority of the dogs showed high antibody titers, suggesting that annual revaccination could be unnecessary.