Genetic and molecular Omp25 analyses from worldwide Brucella canis strains: Possible mutational influences in protein function.

Brucella canis is responsible for canine brucellosis, a neglected zoonotic disease. The omp25 gene has been described as an important marker for Brucella intra-species differentiation, in addition to the ability to interact with the host immune system. Therefore, this study investigated the omp25 sequence from B. canis strains associated to a phylogenetic characterization and the unveiling of the molecular structure. In vitro analyses comprised DNA extraction, PCR, and sequencing of omp25 from 19 B. canis strains. Moreover, in silico analyses were performed at nucleotide level for phylogenetic characterization and evolutionary history of B. canis omp25 gene; and in amino acid level including modeling, dynamics, and epitope prediction of B. canis Omp25 protein. Here, we identified a new mutation, L109P, which diverges the worldwide omp25 sequences in two large branches. Interestingly, this mutation appears to have epidemiology importance, based on a geographical distribution of B. canis strains. Structural and molecular dynamics analyses of Omp25 revealed that Omp25L109P does not sustain its native ß-barrel. Likewise, the conformation of B-cell epitope on the mutated region was changed in Omp25L109P protein. Even without an evolutive marker, the new identified mutation appears to affect the basic function of B. canis Omp25 protein, which could indicate virulence adaptation for some B. canis strains in a context of geographical disposition.

First genome sequence of Chilean Brucella canis SCL strain provides insights on the epidemiology and virulence factors, explaining differences between geographical origins

BACKGROUND: Brucella canis is the etiological agent of canine brucellosis, a worldwide neglected zoonosis that constitutes one of the major infectious causes of infertility and reproductive failure in dogs. Although genomic information available for this pathogen has increased in recent years, here we report the first genome sequencing of a B. canis strain in Chile, and the differences in virulence genes with other B. canis strains. RESULTS: Genome assembly produced a total length of 3,289,216 bp, N50 of 95,163 and GC% of 57.27, organized in 54 contigs in chromosome I, and 21 contigs in chromosome II. The genome annotation identified a total of 1981 CDS, 3 rRNA and 36 tRNA in chromosome I, and 1113 CDS and 10 tRNA in chromosome II. There is little variation between the different strains and the SCL isolate. Phylogenetic analysis showed that the Chilean SCL strain is closely related to B. canis and B. suis strains. Small differences were found when compared to the Serbian isolate, but all strains shared the same recent common ancestor. Finally, changes in the sequence of some virulence factors showed that the SCL strain is similar to other South American B. canis strains. CONCLUSIONS: This work sequenced and characterized the complete genome of B. canis strain SCL, evidencing the complete presence of all the genes of the virB operon, and minor changes in outer membrane proteins and in the urease operon. Our data suggest that B. canis was introduced from North America and then spread throughout the South American continent.

Prevalence and Genomic Characterization of Brucella canis Strains Isolated from Kennels, Household, and Stray Dogs in Chile.

Canine brucellosis caused by Brucella canis is a zoonotic disease that causes reproductive alterations in dogs, such as infertility, abortion, and epididymitis. This pathogen is especially prevalent in South America, and due to the lack of official control programs and the growing trend of adopting dogs it constitutes a public health risk that must be addressed. The aim of this study was to determine the prevalence of B. canis infection in kennel, shelter, and household dogs and to characterize the genomic properties of circulating strains, including ure and virB operons and omp25/31 genes. Samples from 771 dogs were obtained, and the infection was detected by blood culture and/or serology in 7.0% of the animals. The complete ure and virB operons and the omp25/31 genes were detected. Interestingly, we found different single-nucleotide polymorphisms (SNPs) in some of the analyzed genes, which could mean a change in the fitness or virulence of these strains. This study provides further evidence about dogs as a source of B. canis strains that can infect people. This also highlights the need to implement official control programs, including the mandatory testing of dogs, especially stray dogs, before adoption.

Seroprevalence and Risk Factors of Brucellosis in Dogs in Enugu and Anambra States, Nigeria / Seroprevalencia y factores de riesgo de la brucelosis en perros de los Estados Enugu y Anambra, Nigeria

Abstract Seroprevalence of the antibodies of Brucella canis and Brucella abortus in dogs was assessed using a cross-sectional survey in Anambra and Enugu States, Nigeria. A total of 123 Companion dogs made up of 65 clinic dogs, 34 slaughter dogs and 24 household dogs were screened. For B. abortus antibody assay, the collected serum was used for Rose Bengal plate test (RBPT), Serum agglutination test (SAT) and Solid Phase Immunoassay technique with Immunocomb® Canine Brucellosis Antibody Test Kit was used. Out of the 123 dogs screened, none was positive for Brucella abortus antibodies while 34 (27.7%) of the dogs screened were positive for B. canis antibodies. There was a significant association (P<0.05) between infection and sex, the infection was significantly higher (P<0.05) in female than male dogs. Prevalence was significantly higher (P<0.05) in Exotic breeds than in mixed and local dog breeds. There was no association (P>0.05) between infection and antibody titre levels in the different categories of dogs. However, there was significant association (P<0.05) between the presence of Brucella canis antibodies and free roaming of dogs. This study provides the first serological evidence of B. canis infection in dogs in Enugu and Anambra States. This shows that B. canis is endemic in both states, underscoring the need for further studies. Female dogs, exotic breeds and freely roaming dogs are at a higher risk of Brucella infection in the study area; therefore, preventive and control measures are strongly recommended.

Resumen Se evaluó la seroprevalencia de los anticuerpos de Brucella canis y Brucella abortus en perros usando un sondeo transversal en los Estados Anambra and Enugu, Nigeria. Se examinó un total de 123 perros de compañía, de los cuales 65 eran perros de clínica, 34 perros de matadero y 24 perros caseros. Para el ensayo de anticuerpos de B. abortus, el suero muestreado se usó para la prueba de Rosa de Bengala (RBPT), prueba de aglutinación del suero (SAT) y se usó la técnica de inmunoensayo en fase sólida con el kit de prueba de anticuerpos para brucelosis canina Immunocomb®. De los 123 perros analizados, ninguno dio positivo para los anticuerpos de Brucella abortus mientras que 34 (27.7%) de los perros analizados dieron positivo para los anticuerpos de B. canis. Hubo una asociación significativa (P<0.05) entre infección y género; la infección fue significativamente más alta (P<0.05) en las hembras que en los machos. La prevalencia fue significativamente más alta (P<0.05) en las razas exóticas que en las razas cruzadas y las razas locales. No hubo ninguna relación (P>0.05) entre la infección y los niveles de titulación de anticuerpos en las diferentes categorías de perros. Sin embargo, hubo una relación significativa (P<0.05) entre la presencia de anticuerpos Brucella canis y los perros que andan libremente por doquier. Este estudio provee la primera evidencia serológica de infección con B. canis en perros de los Estados Enugu y Anambra. Esto muestra que la B. canis es endémica en ambos estados, enfatizando la necesidad de hacer más estudios. Las hembras, las razas exóticas y los animales que deambulan libremente se encuentran en el riesgo más alto de infección con Brucella en el área de estudio; por consiguiente, se recomienda enormemente tomar medidas preventivas y de control.

Prevalence of presentation of some zoonotic agents transmitted by canines and felines in Medellín, Colombia / Prevalencia de presentación de algunos agentes zoonóticos transmitidos por caninos y felinos en Medellín, Colombia

ABSTRACT Objective. To determine in Medellín, Colombia, the prevalence of zoonotic agents in canines and felines. Materials and methods. 1501 individuals were sampled for the analysis of zoonotic gastrointestinal parasites by direct coprology and flotation. 500 canine sera were examined by PARP-2ME and MAT for the diagnosis of Brucella canis and Leptospira sp, respectively. 500 feline sera were processed by IFI for the diagnosis of Toxoplasma gondii. The frequency for each zoonosis and the statistical significance for the different variables were established (p≤0.05; OR≥1; 95% CI). Results. 23.6% of canines and 16.3% of felines were positive for gastrointestinal parasites; Ancylostomids and D. caninum were the most prevalent; species, age, sex, sector, socioeconomic level and the month of sampling showed associations with gastrointestinal parasitism in pets. Canines showed a seroprevalence of 6.6% in B. canis and 8.4%, Leptospira sp; in felines 56.2% for T. gondii. All of the above associated with the commune, month of sampling, age and stratum. Conclusions. Pets located in different communes and socioeconomic strata with lower quality of life conditions represent a risk of zoonotic transmission.

RESUMEN Objetivo. Determinar la prevalencia de agentes zoonóticos en caninos y felinos en Medellín, Colombia. Materiales y métodos. Se muestrearon 1501 individuos para el análisis de parásitos gastrointestinales zoonóticos por medio de coprología directa y flotación. Se examinaron 500 sueros caninos por medio de PARP-2ME y MAT para el diagnóstico de Brucella canis y Leptospira sp, respectivamente. Se procesaron 500 sueros felinos por medio de IFI para el diagnóstico de Toxoplasma gondii. Se estableció la frecuencia para cada zoonosis y la significancia estadística para las diferentes variables (p≤0.05; OR≥1; IC 95%). Resultados. El 23.6% de los caninos y 16.3% de los felinos fueron positivos a parásitos gastrointestinales, siendo los Ancylostomideos y D. caninum los más prevalente, respectivamente; la especie, edad, sexo, sector, estrato socioeconómico y el mes de muestreo presentaron asociaciones con el parasitismo gastrointestinal en mascotas. En caninos se evidenció una seroprevalencia del 6.6% para B. canis y 8.4% para Leptospira sp; en felinos del 56.2% para T. gondii. Todas las anteriores asociadas con la zona de muestreo, mes, edad y estrato. Conclusiones. Las mascotas ubicadas en diferentes comunas y estratos socioeconómicos con condiciones de calidad de vida menores representan un riesgo de transmisión zoonótica.

Comparison of four polymerase chain reaction assays for the detection of Brucella spp. in clinical samples from dogs.

AIM: This study aimed to compare the sensitivity (S), specificity (Sp), and positive likelihood ratios (LR+) of four polymerase chain reaction (PCR) assays for the detection of Brucella spp. in dog's clinical samples. MATERIALS AND METHODS: A total of 595 samples of whole blood, urine, and genital fluids were evaluated between October 2014 and November 2016. To compare PCR assays, the gold standard was defined using a combination of different serological and microbiological test. Bacterial isolation from urine and blood cultures was carried out. Serological methods such as rapid slide agglutination test, indirect enzyme-linked immunosorbent assay, agar gel immunodiffusion test, and buffered plate antigen test were performed. Four genes were evaluated: (i) The gene coding for the BCSP31 protein, (ii) the ribosomal gene coding for the 16S-23S intergenic spacer region, (iii) the gene coding for porins omp2a/omp2b, and (iv) the gene coding for the insertion sequence IS711. RESULTS: The results obtained were as follows: (1) For the primers that amplify the gene coding for the BCSP31 protein: S: 45.64% (confidence interval [CI] 39.81-51.46), Sp: 95.62% (CI 93.13-98.12), and LR+: 10.43 (CI 6.04-18); (2) for the primers that amplify the ribosomal gene of the 16S-23S rDNA intergenic spacer region: S: 69.80% (CI 64.42-75.18), Sp: 95.62 % (CI 93.13-98.12), and LR+: 11.52 (CI 7.31-18.13); (3) for the primers that amplify the omp2a and omp2b genes: S: 39.26% (CI 33.55-44.97), Sp: 97.31% (CI 95.30-99.32), and LR+ 14.58 (CI 7.25-29.29); and (4) for the primers that amplify the insertion sequence IS711: S: 22.82% (CI 17.89 - 27.75), Sp: 99.66% (CI 98.84-100), and LR+ 67.77 (CI 9.47-484.89). CONCLUSION: We concluded that the gene coding for the 16S-23S rDNA intergenic spacer region was the one that best detected Brucella spp. in canine clinical samples.

Brucellosis outbreak in a commercial kennel: a case report / Surto de brucelose em canil comercial: relato de caso

Brucella canis is a facultative intracellular bacterium responsible for brucellosis in dogs, a chronic and zoonotic infectious disease endemic in Brazil, which promotes reproductive failure and involvement of the lymphoid and articular systems. The infection has been commonly described in breeding kennels, being difficult to treat and control. This study reports an outbreak of brucellosis in a commercial kennel in Guarulhos, SP, Brazil, comprised of 32 dogs (four males and 28 females) from five different breeds, where episodes of abortion had been happening since October 2015. Brucellosis was considered a potential cause of reproductive impairment only a year and a half after the first episode. Anamnesis, clinical examination and serological and bacteriological tests were conducted in all dogs. Of the 32 dogs, 28 had at least one clinical sign compatible with brucellosis, with increased lymph node volume and abortion being the most frequently observed signs. Twenty-six dogs had positive results in at least one of the laboratory tests used for the diagnosis, indicating the infections prevalence to be 81.25% in the kennel. Regarding microbiological diagnosis, B. canis was isolated in blood samples of 22 dogs, in vaginal swabs of nine females and in semen or preputial swabs of two males. Twenty-three dogs had positive results in the serological test.

A Brucella canis é uma bactéria intracelular facultativa responsável pela brucelose nos cães, uma doença infectocontagiosa de caráter crônico e zoonótico, endêmica no Brasil e associada a problemas reprodutivos e comprometimento dos sistemas linfoide e articular. A infecção é comumente descrita em canis de reprodutores, sendo de difícil tratamento e controle. Este trabalho relata um surto de brucelose em um canil comercial localizado em Guarulhos (São Paulo), composto por 32 cães (4 machos e 28 fêmeas) de 5 raças distintas, no qual episódios de abortamento foram relatados desde outubro de 2015. A suspeita de brucelose foi aventada apenas um ano e meio após a ocorrência do primeiro episódio de abortamento, sendo realizados anamnese, exames clínicos e diagnósticos sorológico e bacteriológico em todos os cães do plantel. Dos 32 cães, 28 apresentaram pelo menos um sinal clínico compatível com brucelose, sendo o aumento de linfonodos e o abortamento os sinais mais frequentemente observados. Vinte e seis animais apresentaram resultado positivo em pelo menos um dos testes laboratoriais usados para o diagnóstico, indicando prevalência de 81,25% na criação. No diagnóstico microbiológico, houve isolamento de B. canis em amostras de sangue de 22 cães, em amostras de swab vaginal de 9 fêmeas e em sêmen ou swab prepucial de 2 machos.

Brucellosis outbreak in a commercial kennel: a case report / Surto de brucelose em canil comercial: relato de caso

Brucella canis is a facultative intracellular bacterium responsible for brucellosis in dogs, a chronic and zoonotic infectious disease endemic in Brazil, which promotes reproductive failure and involvement of the lymphoid and articular systems. The infection has been commonly described in breeding kennels, being difficult to treat and control. This study reports an outbreak of brucellosis in a commercial kennel in Guarulhos, SP, Brazil, comprised of 32 dogs (four males and 28 females) from five different breeds, where episodes of abortion had been happening since October 2015. Brucellosis was considered a potential cause of reproductive impairment only a year and a half after the first episode. Anamnesis, clinical examination and serological and bacteriological tests were conducted in all dogs. Of the 32 dogs, 28 had at least one clinical sign compatible with brucellosis, with increased lymph node volume and abortion being the most frequently observed signs. Twenty-six dogs had positive results in at least one of the laboratory tests used for the diagnosis, indicating the infections prevalence to be 81.25% in the kennel. Regarding microbiological diagnosis, B. canis was isolated in blood samples of 22 dogs, in vaginal swabs of nine females and in semen or preputial swabs of two males. Twenty-three dogs had positive results in the serological test.(AU)

A Brucella canis é uma bactéria intracelular facultativa responsável pela brucelose nos cães, uma doença infectocontagiosa de caráter crônico e zoonótico, endêmica no Brasil e associada a problemas reprodutivos e comprometimento dos sistemas linfoide e articular. A infecção é comumente descrita em canis de reprodutores, sendo de difícil tratamento e controle. Este trabalho relata um surto de brucelose em um canil comercial localizado em Guarulhos (São Paulo), composto por 32 cães (4 machos e 28 fêmeas) de 5 raças distintas, no qual episódios de abortamento foram relatados desde outubro de 2015. A suspeita de brucelose foi aventada apenas um ano e meio após a ocorrência do primeiro episódio de abortamento, sendo realizados anamnese, exames clínicos e diagnósticos sorológico e bacteriológico em todos os cães do plantel. Dos 32 cães, 28 apresentaram pelo menos um sinal clínico compatível com brucelose, sendo o aumento de linfonodos e o abortamento os sinais mais frequentemente observados. Vinte e seis animais apresentaram resultado positivo em pelo menos um dos testes laboratoriais usados para o diagnóstico, indicando prevalência de 81,25% na criação. No diagnóstico microbiológico, houve isolamento de B. canis em amostras de sangue de 22 cães, em amostras de swab vaginal de 9 fêmeas e em sêmen ou swab prepucial de 2 machos.(AU)

Development of improved enzyme-based and lateral flow immunoassays for rapid and accurate serodiagnosis of canine brucellosis.

Brucellosis is a widespread zoonotic disease caused by Brucella spp. Brucella canis is the etiological agent of canine brucellosis, a disease that can lead to sterility in bitches and dogs causing important economic losses in breeding kennels. Early and accurate diagnosis of canine brucellosis is central to control the disease and lower the risk of transmission to humans. Here, we develop and validate enzyme and lateral flow immunoassays for improved serodiagnosis of canine brucellosis using as antigen the B. canis rough lipopolysaccharide (rLPS). The method used to obtain the rLPS allowed us to produce more homogeneous batches of the antigen that facilitated the standardization of the assays. To validate the assays, 284 serum samples obtained from naturally infected dogs and healthy animals were analyzed. For the B. canis-iELISA and B. canis-LFIA the diagnostic sensitivity was of 98.6%, and the specificity 99.5% and 100%, respectively. We propose the implementation of the B. canis-LFIA as a screening test in combination with the highly accurate laboratory g-iELISA. The B. canis-LFIA is a rapid, accurate and easy to use test, characteristics that make it ideal for the serological surveillance of canine brucellosis in the field or veterinary laboratories. Finally, a blind study including 1040 serum samples obtained from urban dogs showed a prevalence higher than 5% highlighting the need of new diagnostic tools for a more effective control of the disease in dogs and therefore to reduce the risk of transmission of this zoonotic pathogen to humans.

Application of a polymerase chain reaction test for the detection of Brucella canis from clinical samples of canines and humans / Aplicación de una prueba de reacción en cadena de la polimerasa para detección de Brucella canis a partir de muestras clínicas de caninos y humanos / Aplicação de um teste de reacção em cadeia da polimerase para detecção de Brucella canis a partir de amostras clínicas de canino e humano

Background: laboratory diagnosis of canine brucellosis includes serological and bacteriological tests; the blood culture is considered the gold standard, but it presents issues of sensitivity and delay in results. Therefore, the polymerase chain reaction (PCR) could be useful to detect low amounts of bacterial DNA from clinical samples and provide results within hours. Objective: to evaluate the sensitivity and specificity of PCR for the detection of Brucella canis in whole blood samples. Methods: blood samples from 499 dogs from kennels in two Colombian regions and 91 co-inhabiting humans were used. The 2-mercaptoethanol rapid slide agglutination test (2ME-RSAT) from serum and blood culture and PCR tests from whole blood were performed on all samples. Bayes theorem was used to establish the sensitivity and specificity of the PCR test compared with the other tests performed. Results: 9.9% of the evaluated co-inhabiting humans yielded positive serological results and 0% were positive by PCR or blood culture tests. 10.8% of dog samples were positive by blood culture, 19% were positive by PCR and 13% were positive by 2ME-RSAT. 7% of the samples were positive by all tests. Compared with blood culture, PCR had a sensitivity of 92.6% and a specificity of 90% for canine samples. Compared with 2ME-RSAT, it had a sensitivity of 77.4% and a specificity of 89.2%. When PCR and 2ME-RSAT results were compared with blood culture, a higher number of positive samples were retrieved than when results of only individual tests were applied. Conclusions: PCR is useful to detect B. canis in clinical samples; however, it is preferable to include the 2ME-RSAT test, as this improves the accuracy of the diagnosis. The PCR results are obtained within 24 to 48 hours and do not require the presence of whole bacterial cells to detect DNA.

Antecedentes: el diagnóstico de laboratorio de brucelosis canina incluye pruebas serológicas y bacteriológicas. El hemocultivo se considera el estándar de oro, pero tiene problemas de sensibilidad y tiempo de entrega de los resultados. Por eso, la reacción en cadena de la polimerasa (PCR) podría ser útil para detectar pequeñas cantidades de ADN bacteriano de muestras clínicas y proporcionar resultados en horas. Objetivo: evaluar la sensibilidad y especificidad de una PCR para la detección de Brucella canis, en muestras de sangre total de 499 perros de perreras y 91 humanos cohabitantes de dos regiones de Colombia. Métodos: se realizaron pruebas de aglutinación rápida en placa con 2-mercaptoetanol (2ME-PARP) a partir de suero, PCR y hemocultivo a partir de sangre total. El teorema de Bayes se utilizó para establecer la sensibilidad y especificidad de la prueba de PCR respecto a las demás. Resultados: 9,9% de los humanos cohabitantes evaluados resultaron positivos para la prueba serológica, y el 0% fue positivo para PCR o hemocultivo. 10,8% de las muestras de perros fueron positivas para hemocultivo, 19% fueron positivas para PCR y 13% eran 2ME-PARP positivo. 7% de las muestras fueron positivas para todos los ensayos. En las muestras caninas, la PCR tuvo una sensibilidad del 92,6% y una especificidad del 90% en comparación con el hemocultivo. En comparación con 2ME-PARP, tuvo una sensibilidad del 77,4% y una especificidad del 89,2%. Cuando se compararon los resultados de la PCR y 2ME-PARP con el hemocultivo, un mayor número de muestras positivas fueron obtenidas que usando los resultados de cada una. Conclusiones: la PCR es útil para detectar B.canis en muestras clínicas, sin embargo, es recomendable incluir la prueba de 2ME-PARP, lo que mejora la exactitud del diagnóstico, se obtienen los resultados en 24 ó 48 horas y no requieren la presencia de células bacterianas completas para detectar ADN.

Antecedentes: o diagnóstico laboratorial da brucelose canina inclui testes sorológicos e bacteriológicos. O padrão ouro é a cultura de sangue, mas tem problemas com a sensibilidade e o tempo de entrega dos resultados. Por conseguinte, a reação em cadeia da polimerase (PCR), pode ser útil para detectar pequenas quantidades de ADN bacteriano diretamente a partir de amostras clínicas e fornecem resultados em 24 horas. Objetivo: para avaliar a sensibilidade e a especificidade da PCR para a detecção de Brucella canis em amostras de sangue total de 499 caninos provenientes de canis e 91 seres humanos em contato com estes cães em duas regiões da Colômbia. Métodos: foram realizados testes de aglutinação rápida em placa com 2-mercaptoetanol (2ME-PARP) a partir de soro, PCR y hemocultura a partir de sangue total. O teorema de Bayes utilizou-se para estabelecer a sensibilidade e especificidade da PCR em relação aos outros. Resultados: 9,9% (9) dos humanos avaliados resultaram positivos na prova sorológica, 100% foram negativos por PCR e hemocultura. 10.8% (54) das amostras de cães foram positivas na hemocultura, 19% (95) foram positivas na PCR e 13% (65) foram positivas no teste 2ME-PARP. 7% (35) das amostras foram positivas para todos os testes. Nas amostras caninas, a PCR teve sensibilidade do 92.6% e uma especificidade de 90% em comparação com a hemocultura. Em comparação com 2ME-PARP, teve uma sensibilidade de 77.4% e uma especificidade de 82.2%. Quando foram comparados os resultados da PCR e 2-ME-PARP com a hemocultura, um maior número de amostras positivas foram obtidas que quando foram usados os resultados de cada uma. Conclusões: a PCR é útil para detectar B. canis em amostras clínicas, porém, é recomendável incluir o teste de 2ME-PARP, para melhorar a acurácia do diagnóstico. Os resultados obtém-se em 24-48 horas e não requerem a presença de células bacterianas completas para detectar ADN.

Inquérito sorológico e molecular da brucelose canina no município de Natal, Estado do Rio Grande do Norte / Serological and molecular survey of canine brucellosis in the county of Natal, Rio Grande do Norte state

O objetivo do presente trabalho foi determinar a ocorrência de anticorpos anti-Brucella rugosa e anti-Brucella lisa em cães do município de Natal, Estado do Rio Grande do Norte, Brasil, bem como identificar fatores de risco associados à positividade e realizar a detecção molecular em animais soropositivos. Foram utilizados soros sanguíneos de 416 cães atendidos em clínicas veterinárias durante o período de março a novembro de 2011. Para o diagnóstico sorológico da infecção por Brucella rugosa, foi empregada a prova de imunodifusão em gel de ágar (IDGA), utilizando antígeno de lipopolissacarídeos e proteínas de Brucella ovis, amostra Reo 198 e, para o diagnóstico da infecção por Brucella lisa, foi utilizado o teste do antígeno acidificado tamponado (AAT). De animais soropositivos, foram coletadas amostras de sangue com citrato de sódio para o diagnóstico pela reação em cadeia pela polimerase (PCR). A frequência de anticorpos anti-Brucella rugosa foi de 28,9% (120/416). Todos os animais foram negativos para anticorpos anti-Brucella lisa. Dentre 80 animais soropositivos, o DNA de Brucella spp. foi amplificado em três animais (3,8%). Não foram identificados fatores de risco associados à soropositividade. Conclui-se que a infecção por Brucella rugosa está presente no município de Natal, bem como se sugere o monitoramento sorológico de animais atendidos em clínicas visando à identificação de fontes de infecção.

The aim of this study was to determine the occurrence of anti-rough Brucella and anti-smooth Brucella antibodies in dogs from the county of Natal, Rio Grande do Norte state, Brazil, as well as to identify risk factors associated with positivity and to perform molecular detection of the agent in seropositive animals. Sera from 416 dogs attended in veterinary clinics during the period from March to November 2011 were used. For the serological diagnosis of rough Brucella the agar gel immunodiffusion (AGID) test, using antigen of lipopolysaccharides and proteins from Brucella ovis, strain Reo 198, was carried, and for smooth Brucella the buffered plate agglutination test (BPAT) was used. From seropositive animals, blood samples with sodium citrate were collected for the diagnosis by polymerase chain reaction (PCR). Frequency of anti-rough Brucella antibodies was 28.9% (120/416). All animals were negative for anti-smooth Brucella antibodies. Of the 80 seropositive animals Brucella spp. DNA was amplified in three (3.8%). Risk factors associated with the seropositivity were not identified. It was concluded that rough Brucella infection is present in the county of Natal, as well as it is suggested the serological monitoring of animals attended at clinics aiming the identification of sources of infection.

Fatores de risco e presença de anticorpos contra Brucella canis e amostras lisas de Brucella em cães do município de Araguaína, Tocantins, Brasil / Risk factors and presence of antibodies to Brucella canis and smooth Brucella in dogs from the municipality of Araguaína, Tocantins, Brazil

The aim of this survey was to verify the occurrence of anti-Leptospira spp. antibodies in captive animals in the Parque Zoobotânico Arruda Câmara, João Pessoa, Paraíba State, Northeastern Brazil. Blood samples were collected from 49 animals: 26 mammals of the species Sapajus libidinosus, Cebus flavius, Saimiri sciureu, Coendu sp., Pseudalopex vetulus, Leopardus pardalis, Leopardus tigrinus, Galactitis vitata, Eira barbara, Nasua nasua, Tayassu tajacu and Ratus norvegicus; 10 birds of the species Penelope jacucaca, Pavo cristatus, Anodorhynchus hyacinthinus, Ara chlorothpterus, Pionites leucogaster, Polyborus plancus, Geranoaetus melanoleucus and Urubitinga urubitinga; and 13 reptiles of the species Caiman latirostris, Paleosuchus trigonatus, Caiman crocodilus, Tupinabis merinae, Tupinambis teguixin, Boa constrictor, Corallus hortulanus, Python molurus, Bufocephala vanderhaegei, Geochelone denticulata and Geochelone carboraria. Sera were examined by the microscopic agglutination teste (MAT) using 24 serovars as antigens and cut-off point of 1:100. One ocelot (Leopardo pardalis) presented positive reaction for the Icterohaemorrhagiae serovar with titer of 100, however, it did not show any clinical sign of the infection. Sinantropic rodents are the main reservoirs of this serovar, which suggests the need of maintenance and continuous evaluation of rodent control programs.

Canine brucellosis is an infectious disease of worldwide distribution that can affect dogs, wild canids and man. It is caused by Brucella canis, but dogs can also be infected by smooth Brucella such as B. abortus and B. suis. Due to the increasing importance of dogs in our society, to the scarcity of information about canine brucellosis in the country and its zoonotic character, the aims of the present study were (i) to conduct a survey on the infection by B. canis and smooth Brucella in dogs from the municipality of Araguaína, Tocantins, Brazil, and (ii) to evaluate the risk factors associated with these infections. Sera from 241 dogs were analyzed by agar gel immunodiffusion (AGID) to detect B. canisantibodies, and Buffered Acidified Plate Antigen test (BAPA) and fluorescence polarization assay (FPA) to detect antibodies to smooth Brucella. From the 241 tested dogs, 132 reacted in the AGID and 128 reacted in the BAPA, but only two were positive in FPA. The seroprevalences of B. canis and smooth Brucella infections in dogs in Araguaína were 54.77% (95% CI: 48.25 to 61.17%) and 0.83% (95% CI: 0.10 to 2.97%), respectively. The analysis of risk factors showed associations between B. canis infection and vaccination against leptospirosis, and between B. canis infection and use of manufactured food. In conclusion, data from the present study showed a low prevalence of infection by s

Inquérito sorológico e molecular da brucelose canina no município de Natal, Estado do Rio Grande do Norte

The aim of this study was to determine the occurrence of anti-rough Brucella and anti-smooth Brucella antibodies in dogs from the county of Natal, Rio Grande do Norte state, Brazil, as well as to identify risk factors associated with positivity and to perform molecular detection of the agent in seropositive animals. Sera from 416 dogs attended in veterinary clinics during the period from March to November 2011 were used. For the serological diagnosis of rough Brucella the agar gel immunodiffusion (AGID) test, using antigen of lipopolysaccharides and proteins from Brucella ovis, strain Reo 198, was carried, and for smooth Brucella the buffered plate agglutination test (BPAT) was used. From seropositive animals, blood samples with sodium citrate were collected for the diagnosis by polymerase chain reaction (PCR). Frequency of anti-rough Brucella antibodies was 28.9% (120/416). All animals were negative for anti-smooth Brucella antibodies. Of the 80 seropositive animals Brucella spp. DNA was amplified in three (3.8%). Risk factors associated with the seropositivity were not identified. It was concluded that rough Brucella infection is present in the county of Natal, as well as it is suggested the serological monitoring of animals attended at clinics aiming the identification of sources of infection.

O objetivo do presente trabalho foi determinar a ocorrência de anticorpos anti-Brucella rugosa e anti-Brucella lisa em cães do município de Natal, Estado do Rio Grande do Norte, Brasil, bem como identificar fatores de risco associados à positividade e realizar a detecção molecular em animais soropositivos. Foram utilizados soros sanguíneos de 416 cães atendidos em clínicas veterinárias durante o período de março a novembro de 2011. Para o diagnóstico sorológico da infecção por Brucella rugosa, foi empregada a prova de imunodifusão em gel de ágar (IDGA), utilizando antígeno de lipopolissacarídeos e proteínas de Brucella ovis, amostra Reo 198 e, para o diagnóstico da infecção por Brucella lisa, foi utilizado o teste do antígeno acidificado tamponado (AAT). De animais soropositivos, foram coletadas amostras de sangue com citrato de sódio para o diagnóstico pela reação em cadeia pela polimerase (PCR). A frequência de anticorpos anti-Brucella rugosa foi de 28,9% (120/416). Todos os animais foram negativos para anticorpos anti-Brucella lisa. Dentre 80 animais soropositivos, o DNA de Brucella spp. foi amplificado em três animais (3,8%). Não foram identificados fatores de risco associados à soropositividade. Conclui-se que a infecção por Brucella rugosa está presente no município de Natal, bem como se sugere o monitoramento sorológico de animais atendidos em clínicas visando à identificação de fontes de infecção.

Inquérito sorológico e molecular da brucelose canina no município de Natal, Estado do Rio Grande do Norte

The aim of this study was to determine the occurrence of anti-rough Brucella and anti-smooth Brucella antibodies in dogs from the county of Natal, Rio Grande do Norte state, Brazil, as well as to identify risk factors associated with positivity and to perform molecular detection of the agent in seropositive animals. Sera from 416 dogs attended in veterinary clinics during the period from March to November 2011 were used. For the serological diagnosis of rough Brucella the agar gel immunodiffusion (AGID) test, using antigen of lipopolysaccharides and proteins from Brucella ovis, strain Reo 198, was carried, and for smooth Brucella the buffered plate agglutination test (BPAT) was used. From seropositive animals, blood samples with sodium citrate were collected for the diagnosis by polymerase chain reaction (PCR). Frequency of anti-rough Brucella antibodies was 28.9% (120/416). All animals were negative for anti-smooth Brucella antibodies. Of the 80 seropositive animals Brucella spp. DNA was amplified in three (3.8%). Risk factors associated with the seropositivity were not identified. It was concluded that rough Brucella infection is present in the county of Natal, as well as it is suggested the serological monitoring of animals attended at clinics aiming the identification of sources of infection.

O objetivo do presente trabalho foi determinar a ocorrência de anticorpos anti-Brucella rugosa e anti-Brucella lisa em cães do município de Natal, Estado do Rio Grande do Norte, Brasil, bem como identificar fatores de risco associados à positividade e realizar a detecção molecular em animais soropositivos. Foram utilizados soros sanguíneos de 416 cães atendidos em clínicas veterinárias durante o período de março a novembro de 2011. Para o diagnóstico sorológico da infecção por Brucella rugosa, foi empregada a prova de imunodifusão em gel de ágar (IDGA), utilizando antígeno de lipopolissacarídeos e proteínas de Brucella ovis, amostra Reo 198 e, para o diagnóstico da infecção por Brucella lisa, foi utilizado o teste do antígeno acidificado tamponado (AAT). De animais soropositivos, foram coletadas amostras de sangue com citrato de sódio para o diagnóstico pela reação em cadeia pela polimerase (PCR). A frequência de anticorpos anti-Brucella rugosa foi de 28,9% (120/416). Todos os animais foram negativos para anticorpos anti-Brucella lisa. Dentre 80 animais soropositivos, o DNA de Brucella spp. foi amplificado em três animais (3,8%). Não foram identificados fatores de risco associados à soropositividade. Conclui-se que a infecção por Brucella rugosa está presente no município de Natal, bem como se sugere o monitoramento sorológico de animais atendidos em clínicas visando à identificação de fontes de infecção.

Fatores de risco e presença de anticorpos contra Brucella canis e amostras lisas de Brucella em cães do município de Araguaína, Tocantins, Brasil / Risk factors and presence of antibodies to Brucella canis and smooth Brucella in dogs from the municipality of Araguaína, Tocantins, Brazil

The aim of this survey was to verify the occurrence of anti-Leptospira spp. antibodies in captive animals in the Parque Zoobotânico Arruda Câmara, João Pessoa, Paraíba State, Northeastern Brazil. Blood samples were collected from 49 animals: 26 mammals of the species Sapajus libidinosus, Cebus flavius, Saimiri sciureu, Coendu sp., Pseudalopex vetulus, Leopardus pardalis, Leopardus tigrinus, Galactitis vitata, Eira barbara, Nasua nasua, Tayassu tajacu and Ratus norvegicus; 10 birds of the species Penelope jacucaca, Pavo cristatus, Anodorhynchus hyacinthinus, Ara chlorothpterus, Pionites leucogaster, Polyborus plancus, Geranoaetus melanoleucus and Urubitinga urubitinga; and 13 reptiles of the species Caiman latirostris, Paleosuchus trigonatus, Caiman crocodilus, Tupinabis merinae, Tupinambis teguixin, Boa constrictor, Corallus hortulanus, Python molurus, Bufocephala vanderhaegei, Geochelone denticulata and Geochelone carboraria. Sera were examined by the microscopic agglutination teste (MAT) using 24 serovars as antigens and cut-off point of 1:100. One ocelot (Leopardo pardalis) presented positive reaction for the Icterohaemorrhagiae serovar with titer of 100, however, it did not show any clinical sign of the infection. Sinantropic rodents are the main reservoirs of this serovar, which suggests the need of maintenance and continuous evaluation of rodent control programs.

Canine brucellosis is an infectious disease of worldwide distribution that can affect dogs, wild canids and man. It is caused by Brucella canis, but dogs can also be infected by smooth Brucella such as B. abortus and B. suis. Due to the increasing importance of dogs in our society, to the scarcity of information about canine brucellosis in the country and its zoonotic character, the aims of the present study were (i) to conduct a survey on the infection by B. canis and smooth Brucella in dogs from the municipality of Araguaína, Tocantins, Brazil, and (ii) to evaluate the risk factors associated with these infections. Sera from 241 dogs were analyzed by agar gel immunodiffusion (AGID) to detect B. canisantibodies, and Buffered Acidified Plate Antigen test (BAPA) and fluorescence polarization assay (FPA) to detect antibodies to smooth Brucella. From the 241 tested dogs, 132 reacted in the AGID and 128 reacted in the BAPA, but only two were positive in FPA. The seroprevalences of B. canis and smooth Brucella infections in dogs in Araguaína were 54.77% (95% CI: 48.25 to 61.17%) and 0.83% (95% CI: 0.10 to 2.97%), respectively. The analysis of risk factors showed associations between B. canis infection and vaccination against leptospirosis, and between B. canis infection and use of manufactured food. In conclusion, data from the present study showed a low prevalence of infection by s

Brucella canis infection in dogs attended in veterinary clinics from patos, Paraíba state, Brazil

To determine the frequency of anti-Brucella canis antibodies in dogs attended in veterinary clinics from Patos, Paraíba State, Brazil, as well as to identify risk factors and to isolate and identify the agent, 193 dogs were used. Agar gel immunodiffusion test (AGID) was used to detect B. canis antibodies in sera. Isolation of B. canis was carried out in blood and bone marrow from seropositive animals. Six animals tested seropositive in AGID, resulting in a frequency of 3.11 percent. B. canis was isolated from bone marrow of one seropositive animal, with confirmation by PCR. Lack of cleaning of the dog's environment was identified as risk factor (odds ratio = 7.91). This is the first report of isolation of B. canis in dogs from the Northeast region of Brazil.

Brucella canis infection in dogs attended in veterinary clinics from patos, ParabaIBA state, Brazil.

To determine the frequency of anti-Brucella canis antibodies in dogs attended in veterinary clinics from Patos, Paraíba State, Brazil, as well as to identify risk factors and to isolate and identify the agent, 193 dogs were used. Agar gel immunodiffusion test (AGID) was used to detect B. canis antibodies in sera. Isolation of B. canis was carried out in blood and bone marrow from seropositive animals. Six animals tested seropositive in AGID, resulting in a frequency of 3.11%. B. canis was isolated from bone marrow of one seropositive animal, with confirmation by PCR. Lack of cleaning of the dog's environment was identified as risk factor (odds ratio = 7.91). This is the first report of isolation of B. canis in dogs from the Northeast region of Brazil.

Anticorpos anti-Brucella canis e anti-Brucella abortus em cães de Araguaína, Tocantins / Antibodies anti-Brucella canis and anti-Brucella abortus in dogs from Araguaína, Tocantins, Brazil

Os objetivos deste estudo foram determinar a soroprevalência da infecção por Brucella canis e Brucella abortus e avaliaros possíveis fatores de risco associados à infecção em cães no município de Araguaína, Tocantins. Soros de 374 cães,pertencentes à zona urbana do município de Araguaína-Tocantins, foram analisados pelas técnicas de imunodifusãoem ágar gel (IDGA), para pesquisa de anticorpos contra Brucella canis, e antígeno acidificado tamponado (AAT)e polarização fluorescente (FPA) para detecção de anticorpos contra Brucella abortus. Dos 374 soros testados parapresença de anticorpos contra B. abortus, 21 foram reagentes no AAT, entretanto todos foram negativos pela FPA. Àprova do IDGA 167 animais foram reagentes resultando em uma prevalência para B. canis de 44,53% (IC 95%; 39,43 a49,72). A avaliação de possíveis fatores de risco associados à soropositividade para B. canis não revelou a existência derelação entre a infecção e as variáveis individuais estudadas. Assim, o presente estudo permite concluir que não houveanimais infectados por B. abortus e que a infecção por B. canis está disseminada nos cães do município de Araguaína,Tocantins.(AU)

The aims of the present study were to determine the seroprevalence of infection by Brucella canis and Brucella abortusand to evaluate possible risk factors for infection in dogs from Araguaína, Tocantins, Brazil. Sera from 374 dogs, of theurban zones of the municipality, from both sexes, were submitted to the agar-gel immunodiffusion for Brucella canisantibodiesand to rose Bengal test (AAT) and fluorescence polarization assay (FPA) for Brucella abortus-antibodies.From the 374 tested dogs, 21 reacted in the AAT, but no one was positive in the FPA. The seroprevalence of B. canisinfection found in Araguaína, Tocantins, Brazil, was 44.53% (95% IC; 39.43 to 49.72). No association was found amongseropositivity for B. canis and the risk factors studied. Thus, data from the present study showed that there was noinfection by B. abortus among dogs in the sample and that infection by B. canis is widespread and at high prevalence in Araguaína, Tocantins, Brazil.(AU)

Brucella canis infection in dogs attended in veterinary clinics from patos, Paraíba state, Brazil

To determine the frequency of anti-Brucella canis antibodies in dogs attended in veterinary clinics from Patos, Paraíba State, Brazil, as well as to identify risk factors and to isolate and identify the agent, 193 dogs were used. Agar gel immunodiffusion test (AGID) was used to detect B. canis antibodies in sera. Isolation of B. canis was carried out in blood and bone marrow from seropositive animals. Six animals tested seropositive in AGID, resulting in a frequency of 3.11%. B. canis was isolated from bone marrow of one seropositive animal, with confirmation by PCR. Lack of cleaning of the dog's environment was identified as risk factor (odds ratio = 7.91). This is the first report of isolation of B. canis in dogs from the Northeast region of Brazil.

Anticorpos anti-Brucella canis e anti-Brucella abortus emcães de Araguaína, Tocantins / Antibodies anti-Brucella canis and anti-Brucella abortus in dogs from Araguaína, Tocantins,Brazil

Os objetivos deste estudo foram determinar a soroprevalência da infecção por Brucella canis e Brucella abortus e avaliaros possíveis fatores de risco associados à infecção em cães no município de Araguaína, Tocantins. Soros de 374 cães,pertencentes à zona urbana do município de Araguaína-Tocantins, foram analisados pelas técnicas de imunodifusãoem ágar gel (IDGA), para pesquisa de anticorpos contra Brucella canis, e antígeno acidificado tamponado (AAT)e polarização fluorescente (FPA) para detecção de anticorpos contra Brucella abortus. Dos 374 soros testados parapresença de anticorpos contra B. abortus, 21 foram reagentes no AAT, entretanto todos foram negativos pela FPA. Àprova do IDGA 167 animais foram reagentes resultando em uma prevalência para B. canis de 44,53% (IC 95%; 39,43 a49,72). A avaliação de possíveis fatores de risco associados à soropositividade para B. canis não revelou a existência derelação entre a infecção e as variáveis individuais estudadas. Assim, o presente estudo permite concluir que não houveanimais infectados por B. abortus e que a infecção por B. canis está disseminada nos cães do município de Araguaína,Tocantins.

The aims of the present study were to determine the seroprevalence of infection by Brucella canis and Brucella abortusand to evaluate possible risk factors for infection in dogs from Araguaína, Tocantins, Brazil. Sera from 374 dogs, of theurban zones of the municipality, from both sexes, were submitted to the agar-gel immunodiffusion for Brucella canisantibodiesand to rose Bengal test (AAT) and fluorescence polarization assay (FPA) for Brucella abortus-antibodies.From the 374 tested dogs, 21 reacted in the AAT, but no one was positive in the FPA. The seroprevalence of B. canisinfection found in Araguaína, Tocantins, Brazil, was 44.53% (95% IC; 39.43 to 49.72). No association was found amongseropositivity for B. canis and the risk factors studied. Thus, data from the present study showed that there was noinfection by B. abortus among dogs in the sample and that infection by B. canis is widespread and at high prevalence inAraguaína, Tocantins, Brazil.