Identification of enteric viruses circulating in a dog population with low vaccine coverage

ABSTRACT Although the use of vaccines has controlled enteric diseases in dogs in many developed countries, vaccine coverage is still under optimal situation in Brazil. There is a large population of nonimmunized dogs and few studies about the identification of the viruses associated with diarrhea. To address this situation, stool samples from 325 dogs were analyzed by polymerase chain reaction for the detection of common enteric viruses such as Canine adenovirus (CAdV), Canine coronavirus (CCoV), Canine distemper virus (CDV), Canine rotavirus (CRV) and Carnivorous protoparvovirus 1 (canine parvovirus 2; CPV-2). At least one of these species was detected in 56.6% (184/325) of the samples. The viruses detected most frequently in either diarrheic or nondiarrheic dog feces were CPV-2 (54.3% of the positive samples), CDV (45.1%) and CCoV (30.4%), followed by CRV (8.2%) and CAdV (4.9%). Only one agent was detected in the majority of the positive samples (63%), but co-infections were present in 37% of the positive samples and mainly included CDV and CPV-2. The data presented herein can improve the clinical knowledge in regions with low vaccine coverage and highlight the need to improve the methods used to control these infectious diseases in domestic dogs.

Immunohaematological and rheological parameters in canine visceral leishmaniasis / Parâmetros reológicos e imunohematológicos na leishmaniose visceral canina

Abstract The aim of this study was to find correlations between haematological and rheological parameters in canine visceral leishmaniasis (CLV). Enrolled dogs with CVL (n=31) had a confirmed diagnosis using the rapid test Dual Path Platform (TR DPP®) kit and Polymerase Chain Reaction (PCR) assay. A control group (n=31) comprised healthy dogs with negative results for the TR DPP® kit and PCR assay. Haematological parameters and total protein, albumin, globulin, and antibody were assessed. The rheological properties of the blood samples were also determined. Erythrocytes, haematocrit and platelet values of dogs in the CVL group were found to be lower than those of the control group. The total protein, globulin and IgG concentrations were higher in serum samples from the CVL group. The blood flow curve demonstrated increased shear rates in the CVL group. The viscosity of the blood from the infected animals was lower than in the healthy dogs. A positive correlation was found between erythrocyte numbers and blood viscosity. These data suggest that changes in the flow curve and viscosity of blood as well as the erythrocyte values may be a viable low-cost alternative for the monitoring of dogs with visceral leishmaniasis.

Resumo O objetivo deste estudo foi encontrar correlações entre parâmetros hematológicos e reológicos na leishmaniose visceral canina (LVC). Cães com LVC (n=31) tiveram diagnóstico confirmado de LVC pelo teste rápido imunocromatográfico (TR DPP®) e o ensaio de Reação em Cadeia da Polimerase (PCR). Cães saudáveis ​​ negativos para o TR DPP® e PCR foram alocados no grupo controle (n=31). Foram avaliados parâmetros hematológicos, proteínas totais, albumina, globulina, anticorpos e as propriedades reológicas do sangue. Os valores de eritrócitos, hematócrito e plaquetas de cães do grupo LVC foram menores do que os encontrados no grupo controle. As concentrações de proteína total, globulina e imunoglobulina G (IgG) foram maiores em amostras de soro do grupo LVC. A curva do fluxo sanguíneo apresentou maior taxa de cisalhamento no grupo LVC. A viscosidade sanguínea dos animais infectados foi menor do que a viscosidade do sangue dos animais do grupo controle. Uma correlação positiva foi encontrada entre o número de eritrócitos e a viscosidade do sangue. Esses dados sugerem que as mudanças na curva de fluxo e viscosidade do sangue, bem como os valores de eritrócitos, podem ser uma alternativa viável e de baixo custo para o monitoramento de cães com leishmaniose visceral.

Seroprevalence and spatial distribution dynamics of Yersinia pestis antibodies in dogs and cats from plague foci in the State of Ceará, Northeastern Brazil

Abstract INTRODUCTION: In Brazil, the plague is established in several foci located mainly in the northeastern part of the country, where it alternates between active and quiescent periods. These foci in the State of Ceará have high epidemiological importance. In addition to other plague detection activities, plague areas can be monitored through serological surveys of dogs and cats (domestic carnivores), which, following feeding on plague-infected rodents, can develop mild to severe forms of the disease and produce long-lasting antibodies. This study aimed to characterize the circulation dynamics and spatial distribution of Yersinia pestis antibodies in dogs and cats in plague foci areas of Ceará. METHODS: An ecological study was conducted to analyze the temporal series and spatial distribution of secondary data obtained from domestic carnivore serum surveillance in Ceará's plague areas from 1990 to 2014. RESULTS: Joinpoint analysis revealed that the overall trend was a reduction in antibody-positive animals. The mean proportion of antibody-positivity during the whole study period was 1.5% (3,023/203,311) for dogs, and 0.7% (426/61,135) for cats, with more than 4% antibody-positivity in dogs in 1997 and 2002. Antibody titers ranging from 1/16 to 1/64 were frequent. Despite fluctuations and a significant reduction, in recent years, there were antibody-positive animals annually throughout the study period, and the localities containing antibody-positive animals increased in number. CONCLUSION: Yersinia pestis is actively circulating in the study areas, posing a danger to the human population.

T lymphocytes and macrophages in the intestinal tissues of dogs infected with Leishmania infantum / Linfócitos T e macrófagos nos tecidos intestinais de cães infectados com Leishmania infantum

Abstract This study was about a semi-quantitative analysis of T lymphocytes (CD4+ and CD8+, FoxP3+ regulatory T cells), and macrophages in the gut wall of dogs naturally infected with Leishmania infantum. Thirteen dogs were divided into three groups: group 1 (G1, n=5), dogs with canine visceral leishmaniasis (CVL) and infected with L. infantum amastigotes in the intestine; group 2 (G2, n=5), dogs with CVL but without intestinal amastigotes; and group 3 (G3, n=3), uninfected dogs (control group). There was no significant difference (p ≥ 0.05) on CD4+ and Treg cell numbers among the groups, whereas the levels of CD8+ T cells and macrophages were significantly higher in dogs from G1 group than in G2 and G3 (p ≤ 0.05), especially in intestinal segments with high parasite burden. Parasite burden correlated positively with levels of CD8+ T cells and macrophages (p ≤ 0.05), but was inversely correlated to levels of CD4+ T lymphocytes and FoxP3+ Treg cells. In conclusion, in the intestine of dogs with CVL, the increase of CD8+ T cells and macrophages population associated with high parasite burdens, but no changes of CD4+ T cells and FoxP3+ Treg cells suggest a possible immunoregulation by the parasite not dependent on Treg cells.

Resumo Este estudo foi uma análise semi-quantitativa de linfócitos T (CD4+, CD8+ e regulatórios - Treg FoxP3+) e macrófagos na parede intestinal de cães naturalmente infectados com Leishmania infantum. Treze cães foram divididos em três grupos: grupo 1 (G1, n=5) continha cães com leishmaniose visceral canina (LVC) e com amastigotas intestinais; grupo 2 (G2, n=5) continha cães com LVC, mas sem amastigotas intestinais e o grupo 3 (G3, n=3) continha cães não infectados (grupo controle). Verificou-se que não houve diferença significativa (p ≤ 0.05) no número de células CD4+ e de Treg entre os grupos, mas o número de células T CD8+ e macrófagos foi significativamente superior nos cães do grupo G1 em relação ao G2 e ao G3 (p ≤ 0,05), especialmente nos segmentos intestinais com altas cargas parasitárias. As altas cargas parasitarias correlacionaram positivamente com os números de CD8+ e macrófagos (p ≤ 0,05), mas negativamente com as células CD4+ e Treg. Em conclusão, no intestino dos cães com LVC, o aumento das populações de células T CD8+ e de macrófagos associado a altas cargas parasitárias, mas nenhuma alteração de células T CD4+ e células Treg FoxP3+ sugerem uma possível imunorregulação pelo parasita não dependente de células Treg.

Immunogenic potential of Rhipicephalus (Boophilus) microplus aquaporin 1 against Rhipicephalus sanguineus in domestic dogs / Potencial imunogênico da aquaporina 1 de Rhipicephalus (Boophilus) microplus contra o carrapato Rhipicephalus sanguineus em cães domésticos

Abstract This study evaluated a recombinant aquaporin 1 protein of Rhipicephalus (Boophilus) microplus (RmAQP1) as antigen in a vaccine against R. sanguineus. Five dogs were immunized with RmAQP1 (10 µg) + adjuvant (Montanide) (G1), and five were inoculated with adjuvant only (G2), three times. Twenty-one days after the last immunization, animals of both groups were challenged with R. sanguineus larvae, nymphs and adults, and their biotic potential was compared. Blood samples were collected before each immunization and every 28 days after the last immunization for 10 weeks. Serum antibody titers (IgG) were assessed by ELISA. We observed that: engorgement period of adult females from G1 was 12% shorter than G2; larvae from G1 had 8.7% longer engorgement period than G2 and weighed 7.2% less; nymphs from G1 had 4.5% shorter engorgement period than G2 and weighed 3.6% less; although the antibody titers increased following the second immunization, they rapidly decreased after the third immunization. Results indicated low immunoprotection of RmAQP1 against adult R. sanguineus ticks, and possible efficacy on larvae and nymphs fed on immunized dogs. Further studies should be performed for a full evaluation of the immunoprotection of RmAQP1 against R. sanguineus infestations in dogs.

Resumo Este estudo avaliou a proteína recombinante (aquaporina) do carrapato Rhipicephalus (Boophilus) microplus como antígeno em vacina contra Rhipicephalus sanguineus. Cinco cães foram imunizados com RmAQP1 (10 µg) + adjuvante (G1) e cinco foram inoculados apenas com adjuvante (G2), três vezes. 21 dias após a última imunização todos os animais foram desafiados com larvas, ninfas e adultos de R. sanguineus, e potencial biótico dos carrapatos foi comparado. Amostras de sangue foram coletadas antes de cada imunização e a cada 28 dias após a última imunização, durante 10 semanas. Títulos de anticorpos dos soros dos cães foram avaliados por ELISA. Resultados: o período de ingurgitamento das fêmeas do G1 foi 12% mais curto que o período de ingurgitamento de G2; o período de ingurgitamento das larvas do G1 8,7% foi mais longo e o peso 7,2% menor que no caso de G2; o período de ingurgitamento das ninfas do G1 4,5% foi mais curto e peso 3,6% menor que no caso do G2; aumento dos títulos de anticorpos do G1 após a segunda imunização e declínio após a terceira imunização. Os resultados indicaram baixo potencial de imunoproteção de RmAQP1 contra R. sanguineus adultos, e possível eficácia contra larvas e ninfas, na dose testada. Sugere-se desenvolver novos estudos para melhor avaliação da eficácia de RmAQP1 contra R. sanguineus em cães.

Detección de coproantígenos de Echinococcus granulosus en canes de trabajadores de camales y comercializadores de vísceras en Lima metropolitana / Detection of stool antigens of Echinococcus granulosus in dogs belonging to slaughterhouse workers and offal merchants in Metropolitan Lima

RESUMEN Objetivo Demostrar la presencia de Echinoccocus granulosus en el hospedero definitivo en la ciudad de Lima, Perú, mediante la detección de antígenos del parásito en heces de canes pertenecientes a trabajadores y comercializadores de vísceras de centros de beneficio autorizados en Lima metropolitana. Métodos Se recolectaron muestras de heces de 58 canes, que fueron evaluadas utilizando la técnica coproELISA para detectar antígenos secretorio/excretorio de E. granulosus. Mediante una encuesta se obtuvo información sobre las prácticas de alimentación y el manejo de las mascotas. Resultados El 13,8% (8/58) de canes fue positivo a E. granulosus. En 27,8% (5/18) de los hogares se encontró al menos un animal positivo y se estimó que en las familias que tenían más de cuatro canes las posibilidades de encontrar al menos uno positivo eran mayores. En todos los hogares con al menos un can positivo sus mascotas se alimentaban con vísceras. El 94,4% (17) de los participantes no tenía conocimiento de las formas de contagio de la equinococosis. Conclusiones Los resultados muestran la presencia de hospederos definitivos en la zona urbana de Lima y subrayan la necesidad de aumentar la difusión de las prácticas para evitar la transmisión del parasito.

ABSTRACT Objective To demonstrate the presence of Echinoccocus granulosus in the definitive host in the city of Lima, Perú, by detecting parasite antigens in the stool of dogs belonging to offal handlers and merchants in authorized slaughterhouses in Metropolitan Lima. Methods Stool samples were collected from 58 dogs and examined using the coproELISA technique for the detection of secretory/excretory antigens of E. granulosus. A survey was conducted to obtain information on pet feeding and handling practices. Results Positivity to E. granulosus was detected in 13.8% (8/58) of the dogs. In 27.8% (5/18) of the homes, at least one animal showed positivity, and in families that had more than four dogs the chances of finding positivity in at least one dog were higher (P < 0.05). In all homes where at least one dog tested positive the pets were fed on offal. Of study participants, 94.4% (17) knew nothing about the routes of transmission of hydatid disease. Conclusions Results show the presence of definitive hosts in the urban area of Lima and underscore the need to more widely disseminate practices for the prevention of parasite transmission.

Occurrence of Neospora caninum and Toxoplasma gondii antibodies in dogs from rural properties surrounding a biological reserve, Espirito Santo, Brasil / Ocorrência de anticorpos anti-Neospora caninum e Toxoplasma gondii em cães de propriedades rurais do entorno de uma reserva biológica, Espírito Santo, Brazil

Abstract The aim of this study was to determine the presence of antibodies to Neospora caninum and Toxoplasma gondii in serum samples of 187 dogs from 30 rural properties surrounding Córrego do Veado Biological Reserve, Espírito Santo State, Brazil. The Reserve is one of the last lowland Atlantic Forest remnants of the region, surrounded by agriculture farms and cattle pastures. The presence of IgG antibodies was determined by the Indirect Fluorescent Antibody Test for T. gondii (cut-off 1:16) and N. caninum (cut-off 1:50). Positive samples were diluted 2-fold until the last positive dilution. Antibodies to T. gondii were found in 77 (47.05%) dogs and antibodies to N. caninum in 22 dogs (11.76%) and one only dog was positive for both infections. No association between T. gondii and N. caninum infection and sex was observed (p>0.05). Control measures to prevent those infections in dogs that living surrounding the reserve and that had contact with wild animals are important to avoid the introduction of N. caninum in wild animals. This was the first study of frequency of occurrence of T. gondii and N. caninum in dogs from Espírito Santo, Brazil.

Resumo O objetivo deste estudo foi determinar a presença de anticorpos contra Neospora caninum e Toxoplasma gondii em amostras de soro de 187 cães de 30 propriedades rurais do entorno da Reserva Biológica Córrego do Veado, Espírito Santo, Brasil. A Reserva é um dos últimos remanescentes de Mata Atlântica da região, cercada por fazendas com agricultura e pastagens. A presença de anticorpos IgG foi determinada pela reação de imunofluorescência indireta para T. gondii (ponto de corte de 1:16) e para N. caninum (ponto de corte de 1:50). As amostras positivas foram diluídas na base dois e tituladas. Anticorpos para T. gondii foram encontrados em 77 (47,05%) cães e para N. caninum em 22 (11,76%). Um cão foi positivo para ambas as infecções. Não houve associação entre a infecção pelo T. gondii e N. caninum e o sexo (p>0,05). Medidas de controle, para prevenir essas infecções em cães que vivem no entorno da reserva e com possibilidade de contato com animais selvagens, são importantes para evitar a introdução de N. caninum nos animais selvagens. Este foi o primeiro relato da frequência de anticorpos contra estes coccídios em cães do Espírito Santo, Brasil.

Urticaria papular y sus agentes causales en Colombia / Papular urticaria: A review of causal agents in Colombia

Resumen La urticaria papular es una enfermedad alérgica causada por la picadura de insectos, la cual predomina en el trópico. El objetivo de esta revisión fue profundizar en sus aspectos epidemiológicos e inmunológicos, particularmente con base en datos publicados en Latinoamérica. Se hizo una revisión no sistemática mediante la búsqueda electrónica de artículos sobre la epidemiología de la urticaria papular, las características entomológicas de los agentes causales y los mecanismos inmunológicos asociados. Según los diversos reportes de centros médicos de Latinoamérica la urticaria papular es frecuente; el único estudio de prevalencia publicado indica que afecta a una cuarta parte de los niños escolares de Bogotá. Hay información sobre la relación causal entre la exposición domiciliaria a la pulga, la pobreza y la urticaria papular en Bogotá, una ciudad representativa de las altitudes andinas. No hay estudios que indaguen directamente sobre los insectos causales en zonas cálidas, aunque se sospecha clínicamente de los mosquitos Aedes aegypti y Culex quinquefasciatus. En cuanto a su patogenia, se destaca la participación de mecanismos celulares que involucran las células colaboradoras Th2, lo cual explica que sea una condición de hipersensibilidad retardada. El papel de la inmunoglobulina E (IgE) en la urticaria papular no está tan claro. Se desconocen los antígenos derivados de los insectos que causan la enfermedad, aunque se plantea que existen moléculas comunes de reacción cruzada entre los insectos, tales como el alérgeno Cte f 2 en la pulga, y sus homólogos en los mosquitos. La urticaria papular es una condición frecuente en Latinoamérica que debe investigarse en profundidad. La caracterización inmunológica de los componentes moleculares que causan esta condición puede resolver interrogantes sobre su etiología y su patogenia.

Abstract Papular urticaria is a chronic allergic reaction induced by insect bites, which is common in the tropics. The objective of this review was to deepen on epidemiological and immunological aspects of this disease, focused on data published in Latin American countries. We conducted a non-systematic review of the literature through electronic search on the epidemiology of papular urticaria, the entomological characteristics of the causative agents and associated immunological mechanisms. Several reports from medical centers suggest that papular urticaria is common in Latin America. Only one epidemiological survey designed to estimate prevalence of papular urticaria has been published, reporting that about a quarter of children under six years of age is affected by this condition in Bogotá. There is evidence on the causal relationship among exposure to indoor fleas, poverty and papular urticaria in Bogotá, a representative city of the Andean altitudes. Information about causal insects in tropical warmer areas is scarce, although from clinical reports Aedes aegypti and Culex quienquefasciatus appear to be the most common. Th2 cellular-mediated mechanisms are involved in its pathogenesis, which explains its delayed hypersensitivity. The role of immunoglobulin E is not clear in this disease. Insect-derived antigens directly involved in papular urticaria etiology are unknown. However, it is possible that common molecules among causal insects mediate cross-reactive reactions, such as Cte f 2 allergen, found in cat fleas, and its counterparts in mosquitoes. Papular urticaria is a frequent disease in Latin America that should be further investigated. Immunological characterization of the molecular components that cause this condition may solve questions about its pathogenesis.

Risk factors associated with seropositivity for Leishmania spp. and Trypanosoma cruzi in dogs in the state of Paraiba, Brazil / Fatores de risco associados às soropositividades para Leishmania spp. e Trypanosoma cruzi em cães no Estado da Paraíba, Brasil

Abstract The aim of this survey was to determine the seropositivity and risk factors forLeishmania spp. and Trypanosoma cruzi in dogs in the State of Paraíba, Northeastern Brazil. A total of 1,043 dogs were tested, and the serological diagnoses of Chagas disease (CD) and canine visceral leishmaniasis (CVL) was performed by the indirect fluorescent antibody test (IFAT). Animals that tested seropositive for both diseases (by IFAT) were further subjected to ELISA. Of the 1,043 dogs 81 (7.8%; 95% CI = 6.1-9.4%) tested seropositive for Leishmania spp., while 83 were seropositive for T. cruzi (7.9%; 95% CI = 6.3-9.6%). Simultaneous serological reactions were detected in 49 animals (4.6%; 95% CI= 3.6-6.2%). Semi-domiciled housing (OR = 2.044), free housing (OR = 4.151), and soil (OR = 3.425) and soil/cement (OR = 3.065) environmental conditions were identified as risk factors for CVL seropositivity. The risk factors identified for CD seropositivity were semi-domiciled (OR = 2.353) or free housing (OR = 3.454), and contact with bovine (OR = 2.015). This study revealed the presence of dogs in the Paraíba State seropositive for CVL and CD, suggesting the need for revisiting and intensification of disease control measures through constant monitoring of the canine population.

Resumo O objetivo do presente trabalho foi determinar a soropositividade paraLeishmania spp. e Trypanosoma cruzi em cães do Estado da Paraíba, Nordeste do Brasil, bem como identificar fatores de risco. Foram utilizados 1.043 cães e, para o diagnóstico sorológico de doença de Chagas (DC) e leishmaniose visceral canina (LVC), foi utilizada a reação de imunofluorescência indireta (RIFI). Animais positivos para ambas as doenças (pela RIFI) foram submetidos ao ELISA. Dos 1.043 cães investigados, 81 foram soropositivos para Leishmania spp., resultando em prevalência de 7,8% (IC 95% = 6,1-9,4%) e, para T. cruzi, 83 (7,9%; IC 95% = 6,3-9,6%) animais foram soropositivos. Quarenta e nove animais (4,6%; IC 95% = 3,6-6,2%) apresentaram sororeatividade mista. Criação semidomiciliar (OR = 2,044), criação solta (OR = 4,151), ambiente de terra (OR = 3,425) e ambiente de terra/cimento (OR = 3,065) foram apontados como fatores de risco para LVC, e criação semidomiciliar (OR = 2,353), criação solta (OR = 3,454) e contato com bovinos (OR = 2,015) para DC. Conclui-se que LVC e DC estão presentes em cães do Estado da Paraíba, o que sugere revisão e intensificação das medidas de controle através do constante monitoramento da população canina.

Toxoplasma gondii and Neospora caninum in dogs from the state of Tocantins: serology and associated factors / Toxoplasma gondii e Neospora caninum em cães provenientes de Tocantins: sorologia e fatores associados

Abstract This study investigated occurrences of anti-Neospora caninum and anti-Toxoplasma gondii IgG antibodies by means of the enzyme-linked immunosorbent assay (ELISA) and indirect immunofluorescence assay (IFAT), along with risk factors associated with toxoplasmosis and neosporosis, in 204 dogs from urban and rural areas of the municipality of Araguaína, state of Tocantins, Brazil. One hundred and thirty samples (63.7%) were positive for T. gondii using ELISA: 57.1% and 70.7% in the urban and rural areas, respectively. The seropositivity frequency for T. gondii observed through IFAT was 57.4%, distributed between rural and urban areas as 62.6% and 52.4%, respectively. The factors associated with canine toxoplasmosis were age and breed (p<0.05). In relation to N. caninum, 88 samples (43.1%) were positive, according to ELISA, distributed as 42.9% in urban areas and 43.3% in rural areas. Anti - N. caninum antibodies were detected through IFAT in 62 dogs (30.4%), distributed as 31.3% and 29.5% between rural and urban areas, respectively. Age and breed were associated with neosporosis occurrence (p<0.05) by IFAT. This study provides the first detection of IgG antibodies for canine toxoplasmosis and neosporosis in the state of Tocantins and highlights the importance of dogs in the epidemiological chain of these diseases.

Resumo Este estudo investigou a ocorrência de anticorpos anti-Neospora caninum e anti-Toxoplasma gondii por ensaio imunoenzimático indireto (ELISA) e reação de imunofluorescência indireta (RIFI), assim como os fatores de risco associados à toxoplasmose e à neosporose em 204 cães provenientes de áreas urbana e rural do município de Araguaína, Estado de Tocantins, Brasil. Cento e trinta amostras (63,7%) foram positivas para T. gondii, destas 57,1% e 70,7% oriundas de áreas urbanas e rurais, respectivamente. Considerando-se o teste RIFI, a frequência de soropositividade para T. gondii foi de 57,4% com distribuição de 62,6% e 52,4% entre áreas rurais e urbanas, respectivamente. Fatores associados à toxoplasmose canina foram raça e idade, com soropositividade maior para animais mais velhos (p<0,05). Em relação à N. caninum, 88 (43,1%) amostras foram positivas, segundo ELISA, sendo distribuídas em 42,9% para área urbana e 43,3% para área rural. Por meio da RIFI, anticorpos anti-N. caninum foram detectados em 62 (30,4%) cães, sendo distribuídos em 31,3% e 29,5% entre áreas rurais e urbanas, respectivamente. Os fatores associados à ocorrência de neosporose, pela RIFI, foram idade e raça (p<0,05). Este estudo representa a primeira detecção de anticorpos IgG para toxoplasmose e neosporose canina no Estado de Tocantins e evidencia a importância de cães na cadeia epidemiológica dessas doenças.

Antibodies to Leishmania spp. in domestic felines / Anticorpos anti-Leishmania spp. em felinos domésticos

Abstract Leishmaniasis is a vector-borne zoonotic disease caused by protozoa in the genus Leishmania, typical of rural and peri-urban environments. The causative agent of American visceral leishmaniasis is Leishmania (Leishmania) infantum chagasi and the main insect vector in Brazil is Lutzomyia longipalpis. Dogs (Canis familiaris) are important in the transmission of the disease, as a reservoir closely related to humans and an infection source for phlebotomine vectors. Since 1990, an increasing number of feline leishmaniasis cases have been reported, suggesting that domestic cats (Felis catus) might be involved in the epidemiology of the disease. The present study analyzed the prevalence of anti-Leishmania spp. antibodies in naturally infected domestic cats from various neighborhoods in the municipality of Belém, Pará, Brazil, using the indirect immunofluorescence assay (IFA) and the direct agglutination test (DAT). Among the 443 samples tested, 18 (4.06%) presented positive reactions in the IFA. The observed titers were 40 IU in 4.97% of the samples and 80 IU in 0.90%. In the DAT test, positive results were found in 25 (5.64%) of the samples. The observed titers were also 40 IU (4.97%) and 80 IU (0.68%). The agreement rate between the two tests was considered low (Kappa coefficient = 0.10).

Resumo As leishmanioses são zoonoses vetoriais causadas por protozoários do gênero Leishmania, características de ambientes rurais e periurbanos. A leishmaniose visceral americana (LVA) é causada pela Leishmania (Leishmania) infantum chagasi, cujo principal vetor no Brasil é Lutzomyia longipalpis. O cão (Canis familiaris) possui papel ativo na transmissão da doença, pois é um reservatório muito próximo do humano e uma fonte de infecção para o flebotomíneo. O aumento do número de casos de leishmaniose felina, descritos na literatura a partir de 1990, sugere que gatos também podem atuar na epidemiologia dessa enfermidade. O presente estudo avaliou a prevalência de anticorpos anti-Leishmania spp. em gatos domésticos (Felis catus) de diferentes bairros do Município de Belém, Pará (PA), pela Reação de Imunofluorescência Indireta (RIFI) e pelo Teste de Aglutinação Direta (TAD), utilizando-se como ponto de corte o título de 40 UI. Entre os 443 gatos estudados, 18 (4,06%) apresentaram reação sorológica positiva na RIFI. Os títulos observados foram de 40 UI em 0,90%. No TAD, foi encontrada positividade em 25 (5,64%) animais. Os títulos observados foram de 40 UI, em 4,97% dos gatos, e 80 UI, em 0,68%. A concordância entre os testes foi considerada baixa (coeficiente Kappa: 0,10).

A study of experimental autoimmune encephalomyelitis in dogs as a disease model for canine necrotizing encephalitis

In the present study, the use of dogs with experimental autoimmune encephalomyelitis (EAE) as a disease model for necrotizing encephalitis (NE) was assessed. Twelve healthy dogs were included in this study. Canine forebrain tissues (8 g), including white and grey matter, were homogenized with 4 mL of phosphate-buffered saline for 5 min in an ice bath. The suspension was emulsified with the same volume of Freund's complete adjuvant containing 1 mg/mL of killed Mycobacterium tuberculosis H37Ra. Under sedation, each dog was injected subcutaneously with canine brain homogenate at four sites: two in the inguinal and two in the axillary regions. A second injection (booster) was administered to all the dogs using the same procedure 7 days after the first injection. Clinical assessment, magnetic resonance imaging, cerebrospinal fluid analyses, necropsies, and histopathological and immunohistochemical examinations were performed for the dogs with EAE. Out of the 12 animals, seven (58%) developed clinically manifest EAE at various times after immunization. Characteristics of canine EAE models were very similar to canine NE, suggesting that canine EAE can be a disease model for NE in dogs.

Prevalence of antibodies against Toxoplasma gondii and Neospora caninum in dogs

Toxoplasmosis and neosporosis are parasitic diseases of global importance. The present study had the objective to determine the influence of age, sex and breed in the prevalence of antibodies against both diseases in dogs from Brotas city, São Paulo State, Brazil. Blood samples of 342 dogs were collected, and the age, sex and breed of each animal were recorded. The serological diagnosis for toxoplasmosis and neosporosis were performed using the immunofluorescent antibody test (IFAT). The Fischer's test was used to calculate the association probability of the variables, with a = 5%. For toxoplasmosis the prevalence of antibodies was 26.9% (CI 95% 22.4-31.8%), and for neosporosis 4.97% (CI 95% 3.1-7.8%). The statistical analysis revealed a higher risk of infection for T. gondii in dogs with more than five years.

Detección de coproantígenos para el diagnóstico de echinococosis canina en la zona fronteriza de La Quiaca-Villazón / Coproantigens detection for the diagnosis of canine Echinococcosis in the border area of La Quiaca-Villazón

Las técnicas de inmunodiagnóstico basadas en la identificación de antígenos parasitarios en las heces secas de los perros han sido desarrolladas para la vigilancia de la echinococosis canina. En la región fronteriza de La Quiaca-Villazón, se encontraron las condiciones ambientales que favorecerían el ciclo del parásito, dada la presencia del hospedador definitivo (perro) y de hospedadores intermediarios (ovejas y cabras). La actividad más importante de la Puna es la cría de ovinos y camélidos; la faena se realiza en el campo y a manos del dueño de los ovinos, y no se aplican medidas preventivas de sanidad. El objetivo de este trabajo fue estimar la presencia de caninos parasitados por Echinococcus granulosus en esta región. Durante el año 2006 se recolectaron 168 muestras de materia fecal. En La Quiaca se tomaron muestras de las siguientes localidades: Barrios (área semirrural), Santa Catalina, Yavi Chico, El Portillo, Pumahuasi y Cara Cara (zonas rurales) y La Quiaca (área urbana). En Villazón se seleccionó el área urbana y el área semirrural de Ojo de Agua, Lampaya y Matancillas. Las muestras se analizaron por la prueba de copro-ELISA y copro-Western blot. Las localidades de San Francisco y Barrios tuvieron una prevalencia de 14,3 % y 6,7 %, respectivamente. En Villazón se encontró un 3,4 % de prevalencia en el área urbana. En Lampaya se encontró un 30 % de prevalencia. Estos resultados sugieren la necesidad de implementar estrategias para el control de la hidatidosis, tanto a nivel urbano como rural, para evitar el aumento y la dispersión de la echinococosis en la región.

Cystic Echinococcosis is a major public health issue. Immunodiagnostic techniques based on the identification of parasite antigens in dog dry faeces have been developed as alternatives for the surveillance of canine Echinococcosis. The environmental conditions favouring the parasite cycle were met in the border of La Quiaca-Villazón, given the presence of definitive (dog) and intermediate hosts (sheep and goats). The most important activity in La Puna is sheep and goat rearing; slaughtering is carried out almost exclusively in the field by sheep's owners, and preventive health measures do not apply. The aim of this study was to determine the presence of dogs parasitized by Echinococcus granulosus in this border region. A hundred and sixty eight (168) stool specimens were collected during 2006. La Quiaca samples were taken from the following selected areas: the semi-rural area of Barrios, the rural areas of Santa Catalina, Yavi Chico, El Portillo, Pumahuasi and Cara Cara and the urban area of La Quiaca; selected urban areas in Villazón and the semi-rural area of Ojo de Agua, Lampaya and Matancillas. The samples were analyzed by copro-ELISA -Western blot test. The cities of San Francisco and Barrios had a prevalence of 14.3% and 6.7%, respectively. A prevalence of 3.4% was observed in the urban area of Villazón, which indicates that dogs become infected in the rural areas and bring the risk into the city. Lampaya showed a prevalence of 30%. These findings suggest the need to implement strategies for the control of hydatidosis, both in urban and rural areas to avoid the increase and spread of Echinococcosis in the region.

THE SERO-CONVERSION AND EVALUATION OF RENAL ALTERATIONS IN DOGS INFECTED BY Leishmania (Infantum) chagasi / Soro-conversão e avaliação das alterações renais em cães infectados por Leishmania (Infantum) chagasi

This study investigated the sero-conversion period in which dogs from endemic areas test positive for visceral leishmaniasis (VL) as well as the early post-infection period in which renal alterations are observed. Dogs that were initially negative for Canine Visceral Leishmaniasis (CVL) were clinically evaluated every three months by serological, parasitological and biochemical tests until sero-conversion was confirmed, and six months later a subsequent evaluation was performed. Samples of kidney tissues were processed and stained with Hematoxylin and Eosin (H&E), Periodic Acid Schiff (PAS) and Masson’s trichrome stain and lesions were classified based on the WHO criteria. Of the 40 dogs that initially tested negative for VL, 25 (62.5%) exhibited positive serological tests during the study period. Of these 25 dogs, 15 (60%) tested positive within three months, five (20%) tested positive within six months and five (20%) tested positive within nine months. The dogs exhibited antibody titers between 1:40 and 1:80 and 72% of the dogs exhibited clinical symptoms. The Leishmania antigen was present in the kidneys of recently infected dogs. We found higher levels of total protein and globulin as well as lower levels of albumin in the infected dogs when compared to the control dogs. Additionally, infected dogs presented levels of urea and creatinine that were higher than those of the uninfected dogs. Glomerulonephritis was detected in some of the dogs examined in this study. These data suggest that in Teresina, the sero-conversion for VL occurs quickly and showed that the infected dogs presented abnormal serum proteins, as well as structural and functional alterations in the kidneys during the early post-infection period.

Este estudo investigou o período em que o cão torna-se positivo para leishmaniose visceral (VL) em área endêmica e as alterações renais no período recente pós-infecção. Cães negativos para VL foram avaliados clinicamente a cada três meses por testes sorológicos, parasitológicos e bioquímicos até a soro-conversão e seis meses após. Foram colhidos tecido renal de seis cães, submetidos a processamento de rotina e corados com HE, PAS e Masson e as lesões foram classificadas com base nos critérios da OMS. Dos 40 cães nagativos para VL, 25 (62,5%) apresentaram sorologia positiva durante o estudo. Desses, 15 (60%) tornaram-se positiva nos primeiros três meses, cinco (20%) tornaram-se positivas dentro de seis meses e cinco (20%) tornaram-se positivas dentro de nove meses. Os cães apresentavam títulos de anticorpos entre 1:40 e 1:80, e 72% mostraram sinais clínicos. Antígeno de Leishmania estava presente no rim. Foram encontrados níveis mais elevados de proteína total e globulina, e menores níveis de albumina em cães infectados quando comparados aos controles. Além disso, os cães infectados apresentaram níveis de uréia e creatinina maior do que os cães controles. Glomerulonefrite foi observada em cinco cães. Os resultados sugerem que em Teresina a soro-conversão para VL ocorre rapidamente e os cães apresentam proteínas séricas anormais e alterações na estrutura e função dos rins em um período recente pós-infecção.

Evaluación de la seroconversión como respuesta a la vacunación antirrábica en perros en el departamento del Valle del Cauca, Colombia, 2009 / Evaluation of the seroconversion as a response to rabies vaccination in dogs, Valle del Cauca, Colombia, 2009

Introducción. El departamento del Valle del Cauca ha estado libre de rabia canina por más de 20 años, aunque persisten focos de rabia silvestre que amenazan a humanos y sus mascotas; por ello, como medida preventiva. se realizan anualmente campañas de vacunación antirrábica canina. Objetivos. Medir el impacto de la vacunación en términos de seroconversión de anticuerpos neutralizadores y de porcentaje de perros con respuesta inmunitaria humoral adecuada, relacionando variables propias de estos animales y de las condiciones de vacunación. Discutir el significado epidemiológico de los resultados y sus implicaciones en salud pública. Materiales y métodos. Se obtuvo suero e información de 569 perros vacunados en los 42 municipios de Valle del Cauca. La inmunidad humoral se investigó por ELISA cuantitativa. La información se analizó con el programa Epi-Info 6.0. Resultados. El 9,1 % de los perros de la muestra fueron seronegativos y el 25,1 % no evidenciaron respuesta inmunitaria humoral adecuada a la vacunación. La concentración de anticuerpos disminuyó gradualmente desde la aplicación de la vacuna, y estuvo asociada a edad y calidad de las vacunas, aunque no estuvo asociada a sexo ni a raza. Conclusiones. Con el fin de aumentar los porcentajes de perros seropositivos y con respuesta inmunitaria humoral adecuada, se hacen las siguientes recomendaciones: 1) utilizar vacunas antirrábicas viables;2) aplicar dos dosis de vacuna durante los primeros seis meses de vida de los cachorros; 3) aplicar refuerzos de vacuna, por lo menos, una vez al año; 4) que las autoridades vigilen las actividades y los procesos programáticos relacionados con la vacunación antirrábica por particulares.

Introduction. The province of Valle del Cauca has been free of dog rabies for more than 20 years. However, sylvatic rabies foci remain which are threats to the health of the populace and its pets. Rabies vaccination campaigns are carried out annually in all 42 counties of the province. Objectives. The impact of dog vaccination was evaluated on the basis of humoral immunoresponse, population parameters and correlation with variables inherent to the vaccination process and logistics. Materials and methods. Sera and associated data were obtained from each of the 42 counties for a total sample of 569 rabies-vaccinated dogs. Rabies neutralizing antibodies were measured by quantitative ELISA. The data were analyzed with the statistical programs in Epi-Info 6.0. Results. Nearly 10% of dogs were seronegative (9.1%) and an additional 25.1% did not elicit an adequate humoral immune response to vaccination. Concentration of rabies neutralizing antibodies diminished gradually with the time after vaccination and was correlated with dog age and vaccine quality. No associations were noted between dog gender or breed. Conclusions. These data permit the following recommendations: (1) only viable, non expired rabies vaccines must be used to immunize animals, (2) two doses of rabies vaccine must be applied during the first six months of dog life, (3) booster immunizations must be administered every year, (4) practices and processes related to rabies vaccination in private institutions must inspected regularly by health authorities.

Use of elisa employing homologous and heterologous antigens for the detection of IgG and subclasses (IgG1 and IgG2) in the diagnosis of Canine visceral leishmaniasis / A utilização do ELISA empregando antígenos homólogos e heterólogos para a detecção de IgG e subclasses (IgG1 e IgG2) no diagnóstico de Leishmaniose visceral canina

Indirect immunofluorescence is the method recommended for the diagnosis of visceral leishmanisis in dogs, however, the accuracy of this technique is low and its use on a large scale is limited. Since ELISA does not present these limitations, this technique might be an option for the detection of IgG or specific IgG1 and IgG2 subclasses. Canine ehrlichiosis is an important differential diagnosis of American Visceral Leishmaniasis (AVL). The present study compared ELISA using Leishmania chagasi and Leishmania braziliensis antigen for the detection of anti-Leishmania IgG and subclasses in serum samples from 37 dogs naturally infected with L. chagasi (AVL) and in samples from four dogs co-infected with L. braziliensis and L. chagasi (CI). The occurrence of cross-reactivity was investigated in control serum samples of 17 healthy dogs (HC) and 35 infected with Ehrlichia canis (EC). The mean optical density obtained for the detection of IgG was significantly higher when L. chagasi antigen was used, and was also higher in subgroup VLs (symptomatic) compared to subgroup Vla (asymptomatic). The correlation between IgG and IgG1 was low. The present results suggest that IgG ELISA using homologous antigen yields the best results, permitting the diagnosis of asymptomatic L. chagasi infection and the discrimination between cases of AVL and ehrlichiosis in dogs.

A imunofluorescência indireta é o método recomendado para o diagnóstico de leishmaniose visceral em cães, entretanto, a acurácia dessa técnica é baixa e seu uso em grande escala é limitado. Uma vez que o ELISA não apresenta essas limitações, essa técnica poderia ser uma opção para a detecção de IgG ou subclasses IgG1 e IgG2 específicas. A ehrlichiose canina é um importante diagnóstico diferencial de Leishmaniose Visceral Americana (LVA). O presente estudo comparou o ELISA usando antígenos de Leishmania chagasi e Leishmania braziliensis para a detecção de IgG e subclasses anti-Leishmania em amostras de soro de 37 cães naturalmente infectados com L. chagasi (LVA) e em amostras de quatro cães co-infectados (CI). A ocorrência de reatividade cruzada foi investigada em amostras de soro controle de 17 animais saudáveis (HC) e 35 de infectados por Ehrlichia canis (EC). A média de densidade óptica obtida para a detecção de IgG foi significantemente maior quando o antígeno de L. chagasi foi usado e também mais elevada no subgrupo LVs (sintomático) quando comparado ao subgrupo LVa (assintomático). A correlação entre IgG e IgG1 foi baixa. O presente resultado sugere que ELISA IgG empregando antígeno homólogo, produz os melhores resultados, permitindo o diagnóstico de infecção assintomática por L. chagasi e a discriminação entre casos de LVA e ehrlichiose em cães.

An experimental protocol for the establishment of dogs with long-term cellular immune reactions to Leishmania antigens

Domestic dogs are considered to be the main reservoirs of zoonotic visceral leishmaniasis. In this work, we evaluated a protocol to induce Leishmania infantum/Leishmania chagasi-specific cellular and humoral immune responses in dogs, which consisted of two injections of Leishmania promastigote lysate followed by a subcutaneous inoculation of viable promastigotes. The primary objective was to establish a canine experimental model to provide positive controls for testing immune responses to Leishmania in laboratory conditions. After inoculation of viable promastigotes, specific proliferative responses of peripheral blood mononuclear cells (PBMCs) to either Leishmania lysate or recombinant proteins, the in vitro production of interferon-γ by antigen-stimulated PBMCs and a significant increase in circulating levels of anti-Leishmania antibodies were observed. The immunized dogs also displayed positive delayed-type hypersensitivity reactions to Leishmania crude antigens and to purified recombinant proteins. An important finding that supports the suitability of the dogs as positive controls is that they remained healthy for the entire observation period, i.e., more than seven years after infection. Following the Leishmania antigen lysate injections, the infection of dogs by the subcutaneous route appears to induce a sustained cellular immune response, leading to an asymptomatic infection. This provides a useful model for both the selection of immunogenic Leishmania antigens and for immunobiological studies on their possible immunoprotective activities.

Ehrlichia canis (Jaboticabal strain) induces the expression of TNF-α in leukocytes and splenocytes of experimentally infected dogs / Amostra Ehrlichia canis (Jaboticabal) induz a expressão de TNF-α em leucócitos e esplenócitos de cães experimentalmente infectados

Canine ehrlichiosis is caused by the bacterium Ehrlichia canis and is characterized by a systemic febrile disease of unknown pathogenesis. This study evaluated the expression of cytokines TNF-α, IL-10, IFN-γ, in splenic cells and blood leukocytes during the acute phase of ehrlichiosis and after treatment with doxycycline hyclate in dogs experimentally infected with the E. canis Jaboticabal strain. The study results showed a significant expression of TNF-α 18 days post-inoculation, reducing by approximately 70 percent after treatment. There was a unique peak of expression of IL-10 and IFN-γ 18 and 30 days post-inoculation, respectively. This study suggests that TNF-α plays a role in the pathogenesis of the acute phase of canine ehrlichiosis and that treatment with doxycycline hyclate reduces the systemic effects of this cytokine, possibly by reducing or eliminating parasitemia.

A erliquiose canina é causada pela bactéria Ehrlichia canis, que desencadeia no hospedeiro uma doença febril e sistêmica, de patogênese pouco conhecida. O presente estudo avaliou a expressão das citocinas TNF-α, IL-10, IFN-γ, em células esplênicas e em leucócitos sanguíneos, durante a fase aguda da erliquiose e após o tratamento com hiclato de doxiciclina, em cães experimentalmente infectados com a amostra E. canis Jaboticabal. Os resultados mostraram expressão significativa de TNF-α 18 dias após a inoculação, reduzindo aproximadante 70 por cento após o tratamento. Houve um único pico de expressão de IL-10 e de IFN-γ entre 18 e 30 dias após a inoculação, respectivamente. Este estudo sugere que o TNF-α participa da patogenia da fase aguda da erliquiose canina, e que o tratamento com hiclato de doxiciclina reduz os efeitos sistêmicos dessa citocina, possivelmente por reduzir ou eliminar a parasitemia.

Prevalencia de anticuerpos contra Leptospira en población urbana humana y canina del Departamento del Tolima / The prevalence of antibodies against Leptospira in urban human and canine populations from the Tolima Department

Objetivo Determinar la seroprevalencia de anticuerpos contra Leptospira sp. y los serovares dominantes, en población urbana humana y canina de tres municipios del departamento del Tolima, Colombia. Materiales y Métodos Se realizó un estudio epidemiológico trasversal con selección de sujetos por conveniencia en 62 barrios. Se obtuvieron muestras de 850 personas.y 850 caninos durante los meses de junio, julio y agosto de 2007, las cuales fueron procesadas utilizando la prueba de microaglutinación (MAT). En la prueba se incluyeron 5 serovares: Hardjo, Pomona, Grippotyphosa, Canicola, Icterohaemorrhagiae y Bratislava. La prueba se interpretó como positiva por la presencia de una aglutinación >50 por ciento de las leptospiras con uno o más serovares, en una dilución del suero >100 para las dos especies. Resultados Se encontró evidencia de infección en el 6 por ciento de la población humana y en el 21,4 por ciento de los caninos, con al menos uno de los serovares. La mayor reactividad fue para los serovares Pomona y Grippotyphosa para humanos y caninos. Se presentaron coaglutinaciones en el 13,7 por ciento de los sueros humanos y en el 4,4 por ciento de los caninos. Fue evidente la disminución de la frecuencia de aglutinación de los serovares Canicola e Icterohaemorrhagiae en los perros. Conclusiones Se observa un cambio en el patrón de presentación de la leptospirosis en poblaciones urbanas, siendo necesario fortalecer la vigilancia epidemiológica activa y pasiva, la implementación de pruebas rutinarias de diagnóstico y medidas de prevención específica en los reservorios animales.

Objective Determining Leptospira sp. antibody seroprevalence and prevalent serovars in human and canine populations from three municipalities in the Tolima department of Colombia. Materials and Methods This was a cross-sectional study (subjects being selected by convenience sampling in 62 districts). Samples were collected from 850 dogs and 850 people during June, July and August 2007; these were processed using a microscopic agglutination test (MAT) and five serovars: Hardjo, Pomona, Grippotyphosa, Canicola, Icterohaemorrhagiae and Bratislava. MAT was considered positive when 50 percent or more leptospira were agglutinated with one or more serovars in a 1:100 serum dilution in both species. Results At least one serovar showed evidence of infection in 6 percent of the people and 21.4 percent of canines. The most prevalent serovars were Pomona and Grippotyphosa in two populations. Co-agglutinations were present in 49 percent of human sera and 19.2 percent of canine samples. Decreased Canicola and Icterohaemorrhagiae serovar agglutination frequency was evident in dogs. Conclusions There was a change in leptospirosis pattern in urban populations. Active and passive surveillance must be strengthened, routine diagnostic tests implemented and preventative measures taken in specific animal reservoirs.