Prevalence of Bartonella species in shelter cats and their ectoparasites in southeastern Brazil / Prevalência de espécies de Bartonella em gatos de abrigos e seus ectoparasitas no Sudeste do Brasil

Abstract Feline Bartonella can be transmitted to humans through cat scratches or bites, and between cats, by the flea Ctenocephalides felis. The study was carried out in order to investigate the occurrence of Bartonella DNA in cats living in shelters and their ectoparasites and the relationship between the infection status of cats and ectoparasites they host. Bartonella DNA was detected in 47.8% of the cat blood samples, 18.3% of C. felis fleas, 13.3% of flea egg pools and 12.5% of lice pools. B. henselae and B. clarridgeiae DNA were detected in cat fleas, while B. henselae, B. clarridgeiae and B. koehlerae were found in blood samples from bacteremic cats. Cats infested by positive ectoparasites showed approximately twice the odds of being infected. Our results indicate that shelter cats have high prevalence of Bartonella species that are known to be human pathogens. This highlights the importance of controlling infestations by ectoparasites to avoid cat and human infection.

Resumo Algumas espécies de Bartonella têm os felinos como principais hospedeiros reservatórios. Tais patógenos são transmitidos ao homem por intermédio da arranhadura ou mordedura de gatos e entre os gatos, por meio da pulga Ctenocephalides felis. O objetivo deste estudo foi investigar a ocorrência de DNA de Bartonella spp. em gatos de abrigos e seus ectoparasitas e a relação entre o estado de infecção dos gatos e dos ectoparasitas albergados por estes. Material genético bacteriano foi detectado em 47,8% das amostras de sangue de gatos, 18,3% das pulgas C. felis, 13,3% dos "pools" de ovos de pulgas e 12,5% dos "pools" de piolhos. DNA de B. henselae e B. clarridgeiae foi detectado em pulgas, e B. henselae, B. clarridgeiae e B. koehlerae, em amostras de sangue de gatos. Gatos infestados por ectoparasitas que carreavam DNA de Bartonella spp. demonstraram aproximadamente o dobro de chance de estarem infectados. Esses resultados indicam que os gatos de abrigos têm alta prevalência de infecção por espécies de Bartonella, capazes de causar doenças no homem. E também destacam a importância do controle e prevenção da infestação por ectoparasitas, no intuito de prevenir a infecção em gatos e humanos.

Occurrence of Bartonella genotypes in bats and associated Streblidae flies from Maranhão state, northeastern Brazil / Ocorrência de genótipos de Bartonella em morcegos e moscas Streblidae no Estado do Maranhão, Nordeste do Brasil

Abstract Bartonella is a genus of emerging zoonotic bacteria that are mainly associated with mammalian erythrocytes and endothelial cells. Bats are natural reservoirs for a variety of important pathogens that impact human and animal health. Recent reports have highlighted the role of bats and bat flies in the maintenance of Bartonella. Here, we showed that none of the 29 bat DNA blood samples obtained from five bat species in São Luís Island, state of Maranhão, northeastern Brazil, were positive for Bartonella in qPCR assays targeting nuoG. On the other hand, three out of 15 DNA samples (20%) from flies in the family Streblidae were positive for Bartonella. The BLASTn results showed that the gltA and rpoB sequences shared identities ranging from 97.2% to 100%, with Bartonella sequences amplified from bats or bat flies from Costa Rica and Brazil. These findings were supported by phylogenetic analyses based on Bayesian inferences. The present study showed that Bartonella genotypes are present in bat flies, thus shedding some light on the distribution of bat fly-related Bartonella genotypes in South America.

Resumo Bartonella é um gênero de bactérias zoonóticas emergentes associadas principalmente a eritrócitos e células endoteliais de mamíferos. Morcegos são reservatórios naturais para uma variedade de patógenos importantes que afetam a saúde humana e animal. Além disso, estudos recentes destacaram o papel dos morcegos e de moscas associadas a morcegos na manutenção de Bartonella. No presente estudo, nenhuma das 29 amostras de DNA obtidas a partir do sangue de cinco espécies de morcegos amostrados na ilha de São Luís, estado do Maranhão, Nordeste do Brasil, foi positiva para Bartonella nos ensaios de qPCR direcionados ao gene nuoG. Por outro lado, três das 15 (20%) amostras de DNA de moscas da família Streblidae foram positivas para Bartonella. Os resultados do BLASTn mostraram que as sequências dos genes gltA e rpoB compartilharam identidade, variando de 97,2% a 100%, com as sequências de Bartonella amplificadas em morcegos ou moscas amostrados na Costa Rica ou Brasil. Tais resultados corroboraram as análises filogenéticas realizadas por Inferência Bayesiana. O presente estudo mostrou a ocorrência de Bartonella em moscas de morcegos, auxiliando a esclarecer a distribuição dos genótipos de Bartonella relacionadas a moscas Streblidae na América do Sul.

Aortic valve endocarditis due to Bartonella clarridgeiae in a dog in Brazil / Endocardite de válvula aórtica por Bartonella clarridgeiae em um cão no Brasil

Abstract We report the first documented case of endocarditis associated with Bartonella clarridgeiae in a dog in Latin America. Infective vegetative valvular aortic endocarditis was diagnosed in a 10-year-old male mixed breed dog. The dog presented grade V/VI systolic and diastolic murmur, hyperthermia, and progressive weight loss. Cardiomegaly and presence of diffuse alveolar pattern in the lung fields were observed in the thorax radiography evaluation. Irregular and hyperechogenic structures adhered to the aortic leaflets, causing obstruction of the left ventricular outflow tract and severe aortic insufficiency, were observed in the echocardiography evaluation. A vegetative, whitish, hardened structure measuring 1.0 cm in diameter was observed in aortic semilunar valve at necropsy. Based on a combination of pre-enrichment insect-based medium liquid culture, quantitative real-time and conventional PCR assays based on nuoG and gltA genes, respectively, followed by sequencing and phylogenetic inferences, B. clarridgeiae DNA was detected in the patient's aortic valve lesions. Clinical, echocardiographic, anatomopathologic and molecular features supported the diagnosis of severe aortic vegetative endocarditis possibly caused by B. clarridgeiae in a dog in Brazil.

Resumo Relatamos o primeiro caso documentado de endocardite associada à Bartonella clarridgeiae em um cão na América Latina. Endocardite aórtica valvar vegetativa infecciosa foi diagnosticada em um cão sem raça definida de 10 anos de idade. O cão apresentou sopro sistólico e diastólico de grau V / VI, hipertermia e perda progressiva de peso. Cardiomegalia e presença de padrão alveolar difuso nos campos pulmonares foram observados na avaliação radiográfica do tórax. Estruturas irregulares e hiperecogênicas aderidas aos folhetos aórticos, causando obstrução da via de saída do ventrículo esquerdo e insuficiência aórtica grave, foram observadas na avaliação ecocardiográfica. À necropsia, foi observada uma estrutura vegetativa, esbranquiçada e endurecida medindo 1,0 cm de diâmetro na válvula semilunar aórtica. Por meio de uma combinação de cultura líquida baseada em meio de pré-enriquecimento de inseto, ensaios de PCR quantitativa em tempo real e convencional baseados nos genes nuoG e gltA, respectivamente, seguidos de sequenciamento e inferências filogenéticas, DNA de B. clarridgeiae foi detectado no tecido valvular lesionado do paciente. O diagnóstico de endocardite vegetativa aórtica grave, possivelmente causado por B. clarridgeiae em um cão no Brasil, foi apoiado por características clínicas, ecocardiográficas, anatomopatológicas e moleculares.

Bartonella species pathogenic for humans infect pets, free-ranging wild mammals and their ectoparasites in the Caatinga biome, Northeastern Brazil: a serological and molecular study

ABSTRACT This study verified the occurrence of Bartonella spp. in dogs, cats, wild mammals and their ectoparasites in Petrolina and Lagoa Grande Counties, Pernambuco, located in a semi-arid region in Northeastern Brazil. Anti-Bartonella spp. antibodies were detected by indirect immunofluorescence assay (IFA) in 24.8% of dogs (27/109) and in 15% of cats (6/40). Bartonella sp. DNA was identified by PCR performed on DNA extracted from blood and ectoparasites using primers targeting Bartonella sp. gltA and ribC genes in 100% (9/9) of Pulex irritans from Cerdocyon thous, 57.4% (35/61) of P. irritans from dogs, 2.3% (1/43) of Ctenocephalides felis felis from dogs, 53.3% (24/45) of C. felis felis from cats, and 10% (1/10) of Polyplax spp. from Thrichomys apereoides. DNA sequencing identified Bartonella clarridgeiae and Bartonella henselae in C. felis felis from cats, Bartonella rochalimae in P. irritans from dog and C. thous, and Bartonella vinsoni berkhofii in P. irritans from dog.

MULTIPLEX SYBR® GREEN-REAL TIME PCR (qPCR) ASSAY FOR THE DETECTION AND DIFFERENTIATION OF Bartonella henselae AND Bartonella clarridgeiae IN CATS / PCR em Tempo Real (qPCR) multiplex utilizando SYBR® Green para a detecção e diferenciação de Bartonella henselae e Bartonella clarridgeiae em gatos

A novel SYBR® green-real time polymerase chain reaction (qPCR) was developed to detect two Bartonella species, B. henselae and B. clarridgeiae, directly from blood samples. The test was used in blood samples obtained from cats living in animal shelters in Southern Brazil. Results were compared with those obtained by conventional PCR targeting Bartonella spp. Among the 47 samples analyzed, eight were positive using the conventional PCR and 12 were positive using qPCR. Importantly, the new qPCR detected the presence of both B. henselae and B. clarridgeiae in two samples. The results show that the qPCR described here may be a reliable tool for the screening and differentiation of two important Bartonella species.

Um novo teste baseado na reação em cadeia da polimerase em tempo real (qPCR) com SYBR ® Green foi desenvolvido para detectar duas espécies de Bartonella, B. henselae e B. clarridgeiae, diretamente em amostras de sangue. Este teste foi utilizado em amostras de sangue obtidas de gatos que vivem em abrigos de animais do sul do Brasil. Os resultados foram comparados aos obtidos pelo PCR convencional utilizado para a detecção de Bartonella spp. Das 47 amostras analisadas, oito foram positivas no PCR convencional e 12 foram positivas para qPCR. A reação de qPCR, permitiu a detecção da presença simultânea de B. henselae e B. clarridgeiae em duas destas amostras. Os resultados mostram que a qPCR aqui descrita pode ser uma ferramenta confiável para a detecção e diferenciação de duas espécies importantes de Bartonella spp.

Molecular characterisation of Bartonella species in cats from São Luís, state of Maranhão, north-eastern Brazil

Bartonella species are fastidious bacteria that predominantly infect mammalian erythrocytes and endothelial cells and cause long-lasting bacteraemia in their reservoir hosts. Reports that describe the epidemiology of bartonellosis in Brazil are limited. This study aimed to detect and characterise Bartonella spp DNA from cat blood samples in São Luís, Maranhão, north-eastern Brazil. Among 200 cats tested for multiple genes, nine (4.5%) were positive for Bartonella spp: six cats for Bartonella henselae and three for Bartonella clarridgeiae. Based on the phylogenetic analysis of four genes, the B. henselae strain matched strains previously observed in Brazil and was positioned in the same clade as B. henselae isolates from the United States of America. Moreover, sequence alignment demonstrated that the B. clarridgeiae strain detected in the present study was the same as the one recently detected in cats from southern Brazil.

Detection of hemoplasma and Bartonella species and co-infection with retroviruses in cats subjected to a spaying/neutering program in Jaboticabal, SP, Brazil / Detecção de hemoplasmas e Bartonella sp. e co-infecção com retrovírus em gatos submetidos a um programa de castração/esterilização em Jaboticabal, SP, Brasil

Hemotrophic mycoplasmas and Bartonella species are important pathogens that circulate between cats and invertebrate hosts, occasionally causing diseases in humans. Nevertheless, there are few reports on occurrences of these agents in cats in Brazil. The present study aimed to detect the presence of hemoplasma and Bartonella DNA by means of PCR and sequencing. FIV antigens and anti-FeLV antibodies, were studied by using a commercial kit on blood and serum samples, respectively, among 46 cats that were sampled during a spaying/neutering campaign conducted in Jaboticabal, SP. Three (6.5%) cats were positive for hemoplasmas: two (4.3%) for 'Candidatus M. haemominutum' and one (2.2%) for both M. haemofelis and 'Candidatus M. turicensis'. One of the two 'Candidatus M. haemominutum'-infected cats was also positive for FeLV antigens and showed antibodies for FIV. Two cats (4.3%) were positive for B. henselae. One of them was also positive for FeLV antigens. Eight cats (17.4%) were positive for FeLV, and just one (2.2%) showed anti-FIV antibodies. Bartonella species and hemoplasmas associated with infection due to retroviruses can circulate among apparently healthy cats.

Micoplasmas hemotróficos e espécies de Bartonella são importantes patógenos que circulam entre gatos e hospedeiros invertebrados, causando ocasionalmente doenças no homem. Apesar disto, poucos são os estudos acerca da ocorrência destes agentes entre gatos no Brasil. O presente estudo objetivou detectar o DNA de hemoplasmas e Bartonella sp. pela PCR e sequenciamento. Antígeno de FIV e anticorpos anti-FeLV foram estudados utilizando um "kit" comercial, em amostras de sangue e soro, respectivamente, de 46 gatos amostrados em uma campanha de castração em Jaboticabal, SP. Três gatos (6,5%) foram positivos para hemoplasmas: dois (4,3%) para 'Candidatus M. haemominutum' e um (2,2%) para M. haemofelis and 'Candidatus M. turicensis'. Um dos gatos positivos para 'Candidatus M. haemominutum' mostrou-se também positivo na detecção de antígeno de FeLV e de anticorpos para FIV. Dois (4,3%) gatos mostraram-se positivos para B. henselae, sendo que um deles também se mostrou positivo para antígeno de FeLV. Oito gatos (17,4%) foram positivos para FeLV, e apenas um gato mostrou anticorpos anti-FIV. Bartonella sp. e hemoplasmas associados à infecção por retrovírus podem circular entre gatos aparentemente saudáveis.

Prevalence of Bartonella henselae and Bartonella clarridgeiae in cats in the south of Brazil: a molecular study

Bartonella spp are the causative agent of cat scratch disease in humans. Cats are the natural reservoir of these bacteria and may infect humans through scratches, bites or fleas. Blood samples from 47 cats aged up to 12 months were collected for this study. All animals were lodged in municipal animal shelters in the Vale do Sinos region, Rio Grande do Sul, Brazil. Bartonella spp were detected by genus-specific polymerase chain reaction (PCR) and when the PCR was positive, the species were determined by DNA sequencing. A Giemsa-stained blood smear was also examined for the presence of intraerythrocytic elements suggestive of Bartonella spp infection. Phylogenetic analysis was also performed for all positive samples. Using molecular detection methods, Bartonella spp were detected in 17.02 percent (8/47) of the samples. In seven out of eight samples confirmed to be positive for Bartonella spp, blood smear examination revealed the presence of intraerythrocytic elements suggestive of Bartonella spp. Phylogenetic analysis characterized positive samples as Bartonella henselae (5) or Bartonella clarridgeiae (3). To the best of our knowledge, this is the first molecular study demonstrating the presence of Bartonella spp in cats from the Southern Region of Brazil.

Bartonelose: análise molecular e sorológica em gatos do Rio de Janeiro Brasil / Bartonellosis: molecular and serological analysis in cats from Rio de Janeiro – Brazil

Bartonella henselae e mais recentemente B. quintana têm sido apontados como agentes causais de diversas moléstias emhumanos, entre as quais a doença da arranhadura do gato, endocardite, meningoencefalite e neuroretinite, podendo levarao óbito, principalmente os imunocomprometidos. O gato doméstico é considerado o principal animal envolvido na transmissãodestes patógenos. Constituiu-se objetivo deste estudo a avaliação da frequência de Bartonella spp. em gatos domésticosdomiciliados do município de Vassouras (RJ) comparando-se os achados na reação em cadeia pela polimerase (PCR) e nasorologia por imunofluorescência indireta (IFA). Amostras sanguíneas de 37 (100%) gatos de um abrigo da cidade deVassouras (RJ) foram analisadas, sendo 36 (97,3%) positivas na PCR para Bartonella spp. Das amostras PCR positivas,nove (25%) e 27 (75%) apresentaram, respectivamente, reatividade e ausência de reatividade ao IFA. Apenas uma (2,7%)amostra de sangue foi concomitantemente negativa na PCR e IFA para Bartonella spp. Este é o primeiro registro de infecçãopor Bartonella spp. em felinos domésticos no estado do Rio de Janeiro (Brasil) identificada por análise molecular e sorológica,o que nos permite concluir que este agente zoonótico está presente em alta frequência em gatos domésticos do municípiode Vassouras (RJ).

Bartonella henselae and B. quintana have been pointed as causal agents of many diseases in humans, and can lead to death, mainly immunodefficient people. Domestic cat is considered the unique animal in transmission of these pathogens. The purpose of this study was to evaluate the frequency of Bartonella spp. in domestic cats from Vassouras city (RJ) by polimerase chain reaction (PCR) and indirect immunofluorescence test assay (IFA) and compare the results. Blood samples from 37 (100%) domestic cats from a shelter of Vassouras city (RJ) were analyzed and 36 (97.3%) were considered positive by polymerase chain reaction (PCR). The indirect immunofluorescence test assay (IFA) revealed 9 (25.0%) and 27 (75.0%) of that PCR positive samples showed, respectively, reaction and absence of reaction to IFA. Only one sample (2.7%) was negative in PCR and IFA. This is the first communication of Bartonella spp. infection in domestic cats in Rio de Janeiro State (Brazil) identified by molecular and serological assays, thus it can be concluded that this zoonotic agent is present in high frequencies in domestic cats from Vassouras city (RJ).