Sarcocystis sp. shed by the common boa snake (Boa constrictor) in Brazil.

The genus Sarcocystis contains around 200 species and 25 of these infect snakes. Two Sarcocystis spp. shed by snakes have called special attention of the scientific community. S. nesbitti, which is shed by scrub pythons (Simalia amethistina), causes myopathy in humans that consume water or food contaminated with the parasite. Sporocysts of S. singaporensis, excreted by reticulated pythons (Malayopython reticulatus), is letal for rats and was successfully tested in the biological control of these rodents. A high biodiversity of snakes is found in Brazil, however, scarce information is available about Sarcocystis spp. in Brazilian snakes. Herein, we investigated Sarcocystis sp. in feces of the common boa (Boa constrictor) from Salvador, as it is widely distributed in Brazil and it is also bred in other countries. Feces of 65 boas were examined, and Sarcocystis sp. was found in 1/65 (1.53%) snakes. All snakes were alive, and for this reason, intestinal scrapping, which is the most sensitive method to detect the parasite, was not performed. Morphometric evaluation of sporocysts showed significant differences in their sizes. PCR and multilocus sequencing of four genetic markers (cox1, 18S, ITS1, and 28S) revealed that sporocysts corresponded to a new Sarcocystis species. Sequences of cox1 and 18S had identities of 100% and higher than 98%, respectively, with sequences obtained from the rodent Lagostomus maximus in Argentina. ITS1 and 28S sequences did not match with any known Sarcocystis sp. No ITS1 and 28S sequences were available for the Sarcocystis sp. found in the Argentinian L.maximus. Bioassay using the boa sporocysts was conducted in three mouse lineages and in Rattus norvegicus, but no parasitic stages were detected in these rodents. We concluded that the common boa is probably the definitive host of a new species of Sarcocystis sp. that has L. maximus or related rodents as intermediate hosts.

Sarcocystis neurona and related Sarcocystis spp. shed by opossums (Didelphis spp.) in South America.

Protozoan parasites of the genus Sarcocystis are obligatory heteroxenous cyst-forming coccidia that infect a wide variety of animals and encompass approximately 200 described species. At least four Sarcocystis spp. (S. falcatula, S. neurona, S. lindsayi and S. speeri) use opossums (Didelphis spp.) as definitive hosts, and two of them, S. neurona and S. falcatula, are known to cause disease in horses and birds, respectively. Opossums are restricted to the Americas, but their distribution in the Americas is heterogeneous. Five Didelphis spp. are distributed in South America (D. aurita, D. albiventris, D. marsupialis, D. imperfecta and D. pernigra) whereas just one opossum species (D. virginiana) is found in North America. Studies conducted in the last decades show that Sarcocystis spp., derived from South American Didelphis spp., have biological and genetic differences in relation to Sarcocystis spp. shed by the North American opossum D. virginiana. The aim of this review was to address the peculiar scenario of Sarcocystis species shed by South American opossums, with a special focus on diagnosis, epidemiology, and animal infections, as well as the genetic characteristics of these parasites.

Serologic reactivity of canine sera to Sarcocystis neurona and Sarcocystis cruzi antigens.

Sarcocystis neurona, a coccidian parasite shed by opossums (Didelphis spp.) in the Americas, is the major cause of equine protozoal myeloencephalitis (EPM) and induces disease in other domestic and wild animal species, including domestic dogs. Sarcocystis cruzi, despite its low pathogenicity for cattle (intermediate hosts), is worldwide distributed and uses mostly dogs as definitive hosts. The aims of this study were to test serological reactivities of dog sera to S. neurona and S. cruzi antigens, and to investigate potential serological cross-reactivity to these parasites. Sera from 353 Brazilian dogs were obtained from rural areas in the municipality of Ilhéus, Bahia, and examined by immunofluorescent antibody tests (IFAT). Antigens used in serological reactions consisted of S. neurona merozoites from a North American strain (SN138), and bradyzoites of S. cruzi obtained from Brazilian bovine hearts, with parasite species identity confirmed by PCR and sequencing of the 18S gene of the rDNA. Seropositivity to S. neurona and to S. cruzi were detected in 3.39% (12/353) and 4.81% (17/353) of the dogs, respectively. Ten canine sera reacted solely to S. neurona and 15 serum samples reacted only to S. cruzi. Two serum samples were simultaneously positive for both parasites. Sera from 14 dogs that tested positive by IFAT (9 for S. neurona and 3 for S. cruzi) and from two dogs that were negative by IFAT for the two parasites, were examined by Western blot using S. neurona as antigen; these sera reacted to a great number of protein bands, including antigens on the 16 and 30 KDa positions, which encompass immunodominant antigens for S. neurona in horses. Western blot did not show any specific pattern for S. neurona infection/exposure using canine sera. Dogs act as definitive hosts for several Sarcocystis spp. that infect farm animals, including horses, sheep, goats, water buffaloes and pigs, and for this reason, should contain antibodies to a broad repertoire of Sarcocystis spp. antigens. In conclusion, low percentages of dogs from rural areas of Ilhéus, Bahia, were reactive to both S. neurona and S. cruzi antigens. It is possible that other Sarcocystis species, besides S. neurona and S. cruzi, might have contributed for the seropositivity observed in this study. IFAT was more specific than Western blot to differentiate canine serological reactions to S. neurona and S. cruzi antigens.

Toxoplasmatinae Parasites in Bats from Bahia State, Brazil.

Toxoplasma gondii and Neospora caninum are widespread cyst-forming coccidian parasites of the subfamily Toxoplasmatinae that infect a wide range of wild and domestic animals. Whereas T. gondii is a zoonotic disease, N. caninum is restricted to nonhuman animals. Some chiropteran species can be infected by T. gondii and present fatal toxoplasmosis. In most cases, T. gondii -infected bats are believed to remain asymptomatic and to act as an infection source to other animals. It is not known whether N. caninum can infect bats. We determined infection rates of T. gondii and N. caninum in free-living bats in the state of Bahia, Brazil. Brain samples from 97 bats of seven species, captured in 2008-15, were analyzed by PCRs for T. gondii and N. caninum . Two of the 97 samples were positive for T. gondii DNA. None of the samples were positive for N. caninum DNA, suggesting that the bats were not susceptible to N. caninum infection or that its prevalence was very low.

Brazilian donkeys (Equus asinus) have a low exposure to Neospora spp.

Donkeys (Equus asinus) are closely related to horses and are known to be infected by several equine pathogens. Neospora caninum and Neospora hughesi are protozoan parasites that infect horses, but they were not confirmed in donkeys up to this date. The aim of this study was to evaluate the exposure of donkeys (Equus asinus) to Neospora spp. using tachyzoites of N. caninum as antigen and employing two common serologic methods, IFAT and immunoblot. Sera from 500 donkeys were obtained from 30 municipalities in Bahia state and tested by IFAT. Two of 500 sera were positive for Neospora spp. by IFAT with antibody titers of 100, and recognized a 37kDa antigen in immunoblot. Approximately 22% of the samples showed strong apical reactions and/or incomplete fluorescence, what may cause confusion in the interpretation of IFAT. We concluded that Neospora spp. are possibly of minor importance for Brazilian donkeys. Future studies are necessary to prove that Neospora spp. can naturally infect donkeys.

Brazilian donkeys (Equus asinus) have a low exposure to Neospora spp / Asininos (Equus asinus) brasileiros têm uma baixa exposição à Neospora spp

Donkeys (Equus asinus) are closely related to horses and are known to be infected by several equine pathogens. Neospora caninum and Neospora hughesi are protozoan parasites that infect horses, but they were not confirmed in donkeys up to this date. The aim of this study was to evaluate the exposure of donkeys (Equus asinus) to Neospora spp. using tachyzoites of N. caninum as antigen and employing two common serologic methods, IFAT and immunoblot. Sera from 500 donkeys were obtained from 30 municipalities in Bahia state and tested by IFAT. Two of 500 sera were positive for Neospora spp. by IFAT with antibody titers of 100, and recognized a 37kDa antigen in immunoblot. Approximately 22% of the samples showed strong apical reactions and/or incomplete fluorescence, what may cause confusion in the interpretation of IFAT. We concluded that Neospora spp. are possibly of minor importance for Brazilian donkeys. Future studies are necessary to prove that Neospora spp. can naturally infect donkeys.

Asininos (Equus asinus) são próximos filogeneticamente a equinos e podem ser infectados por vários patógenos de cavalos. Neospora caninum e Neospora hughesi são parasitos protozoários que infectam equinos, porém não foram confirmados em asininos até o momento. O objetivo deste estudo foi avaliar a exposição de asininos (Equus asinus) a Neospora spp., usando-se taquizoítos de N. caninum como antígeno e empregando-se duas técnicas sorológicas comuns para esta finalidade, reação de imunofluorescência indireta (RIFI) e immunoblot. Soros de 500 asininos, obtidos em 30 municípios no Estado da Bahia, foram testados por meio da RIFI. Dois dos 500 soros foram positivos para Neospora spp. pela RIFI com títulos de anticorpos de 100, e reconheceram um antígeno de 37kDa no immunoblot. Aproximadamente, 22% das amostras apresentaram fortes reações apicais e/ou fluorescência incompleta, o que pode causar confusão na interpretação da RIFI. Conclui-se que Neospora spp. são, possivelmente, de pouca importância para asininos brasileiros. Estudos posteriores são necessários para provar que Neospora spp. podem causar infecção natural em asininos.

Frequency of antibodies against Sarcocystis neurona and Neospora caninum in domestic cats in the state of Bahia, Brazil.

Sarcocystis neurona is the major agent of equine protozoal myeloencephalitis. It infects several mammalian species in the Americas, where the definitive hosts, marsupials of the genus Didelphis (D. virginiana and D. albiventris) are found. Domestic cats are one of the confirmed intermediate hosts of the parasite; however, antibodies against S. neurona had never before been demonstrated in Brazilian cats. The aim of this study was to determine whether cats in Bahia, Brazil, are exposed to the parasite. A total of 272 feline serum samples (134 from feral and 138 from house cats) were subjected to an indirect fluorescent antibody test using cultured merozoites of S. neurona as antigen. Positivity was detected in 4.0% (11/272) of the tested samples, with titers ranging from 25 to 800. The feline sera were also tested for antibodies against the protozoan Neospora caninum, with an observed antibody frequency of 2.9%. To the author's knowledge, this is the first study to report antibodies against S. neurona in Brazilian cats. We conclude that cats are exposed to the parasite in the region of this study. Further investigations are needed to confirm the role of cats in the transmission cycle of S. neurona in Brazil.

Frequency of antibodies against Sarcocystis neurona and Neospora caninum in domestic cats in the state of Bahia, Brazil / Frequência de anticorpos contra Sarcocystis neurona e Neospora caninum em gatos domésticos do Estado da Bahia, Brasil

Sarcocystis neurona is the major agent of equine protozoal myeloencephalitis. It infects several mammalian species in the Americas, where the definitive hosts, marsupials of the genus Didelphis (D. virginiana and D. albiventris) are found. Domestic cats are one of the confirmed intermediate hosts of the parasite; however, antibodies against S. neurona had never before been demonstrated in Brazilian cats. The aim of this study was to determine whether cats in Bahia, Brazil, are exposed to the parasite. A total of 272 feline serum samples (134 from feral and 138 from house cats) were subjected to an indirect fluorescent antibody test using cultured merozoites of S. neurona as antigen. Positivity was detected in 4.0% (11/272) of the tested samples, with titers ranging from 25 to 800. The feline sera were also tested for antibodies against the protozoan Neospora caninum, with an observed antibody frequency of 2.9%. To the author's knowledge, this is the first study to report antibodies against S. neurona in Brazilian cats. We conclude that cats are exposed to the parasite in the region of this study. Further investigations are needed to confirm the role of cats in the transmission cycle of S. neurona in Brazil.

Sarcocystis neurona é o principal agente da mieloencefalite protozoária equina. Esse parasito infecta várias espécies de mamíferos nas Américas, onde são encontrados os hospedeiros definitivos, os marsupiais do gênero Didelphis (D. virginiana and D. albiventris). O gato doméstico é um dos hospedeiros intermediários do parasito. Contudo, anticorpos contra S. neurona ainda não tinham sido demonstrados em gatos brasileiros. O objetivo deste trabalho foi determinar se gatos da Bahia, Brasil, são expostos ao parasito. Amostras séricas de 272 felinos (134 de gatos errantes e 138 de gatos domiciliados) foram testadas pelo teste de imunofluorescência indireta, utilizando-se como antígeno, merozoítos produzidos em cultura celular. Entre as amostras testadas, 4,0% (11/272) foram positivas com títulos entre 25 e 800. Os soros dos felinos foram também testados para anticorpos contra o protozoário Neospora caninum, cuja frequência de anticorpos foi de 2,9%. Esse é o primeiro relato de anticorpos contra S. neurona em gatos brasileiros. Conclui-se que os gatos da região estudada são expostos a S. neurona. Estudos futuros são necessários, a fim de se confirmar o papel dos gatos no ciclo de transmissão de S. neurona no Brasil.

Factors associated with infection by Neospora caninum in dogs in Brazil.

From August 2006 to 2008, 411 dogs in northeastern Brazil were evaluated for seropositivity to Neospora caninum. The dogs were clinically examined, and their owners were interviewed about the conditions in which the animals were maintained in order to assess the factors associated with infection by this parasite. A serum sample was taken from each dog for serological examination in an indirect fluorescent antibody test for N. caninum. The Yates' Chi-square test or Fisher's exact test was used to select the variables for the multivariate logistic regression model. Seropositivity was detected in 9.26% of dogs. The seropositivity rates of dogs from different environments were 2.6% (4/156) in urban areas, 13.1% (28/214) in peri-urban areas, and 14.6% (6/41) in rural areas. Factors associated with seropositivity for N. caninum were the following: contact with other dogs, access to food outside the home and residing in the peri-urban or rural environments (p<0.05). Results of this study confirm that dogs in urban, rural and peri-urban areas of northeastern Brazil are exposed to N. caninum. Control measures to prevent infection of dogs in the studied region should be focused primarily on preventing access to potential sources of infection, which include environments with other dogs, bovines, and other small intermediate hosts, such as birds and rodents.

Prevalence of ehrlichial infection among dogs and ticks in Northeastern Brazil / Prevalência da infecção por Ehrlichia em cães e carrapatos no Nordeste do Brasil

This study investigated the epidemiology of canine ehrlichiosis in Northeastern Brazil, focusing the identification of the Ehrlichia species and vectors involved. Samples were collected from 472 domestic dogs residing in the health districts of Cajazeiras and Itapuã of Salvador city. The average prevalence of antibodies reactive to E. canis by immunofluorescent antibody test (IFAT) (titer > 1:80) was 35.6 percent (168/472). Blood samples from the E. canis-seropositive animals were tested by nested PCR in order to identify the Ehrlichia species responsible for the infection. Among the seropositives, 58 (34.5 percent) were found to be PCR-positive for E. canis. Ticks were found in 32 dogs. Nested-PCR analysis showed that 21.9 percent (7/32) of the Rhipicephalus sanguineus were infected by E. canis. In both dogs and Rhipicephalus sanguineus, nested-PCR for E. ewingii and E. chaffeensis was negative, with no amplification of DNA fragment.

Este estudo objetivou pesquisar a epidemiologia da erliquiose canina no Nordeste do Brasil, com especial atenção na identificação da espécie de Ehrlichia envolvida nas infecções caninas e vetoriais detectadas. Para isso foram coletadas amostras de 472 cães domiciliados nos distritos sanitários de Cajazeiras e Itapuã. A prevalência de anticorpos anti-E. canis, pela imunofluorescência indireta (título > 1:80), em cães foi de 35,6 por cento (168/472). Os animais soropositivos foram analisados por uma nested-PCR para identificação da espécie de Ehrlichia responsável pela infecção. Dentre os positivos, 58 (34,5 por cento) cães foram PCR-positivos para E. canis. Foram coletados e classificados os carrapatos em 32 cães. A nested-PCR de Rhipicephalus sanguineus resultou em 21,9 por cento (7/32) de infecção por E. canis. A nested-PCR de amostras de sangue de cães e Rhipicephalus sanguineus para E. chaffeensis e E. ewingii foi negativa, não havendo amplificação de fragmento de DNA.

Investigation of Neospora caninum, Hammondia sp., and Toxoplasma gondii in tissues from slaughtered beef cattle in Bahia, Brazil.

Neospora caninum, Hammondia sp., and Toxoplasma gondii are parasites with morphological and genetic similarities. N. caninum and T. gondii are important abortive agents of cattle and sheep, respectively, and may infect numerous animal species. Hammondia sp. is not known to induce disease in animals, but may cause confusion in the identification of closely related coccidia. The aim of this study was to investigate infection rates caused by N. caninum, Hammondia sp., and T. gondii in beef cattle using a nested PCR for Toxoplasmatinae rDNA, followed by sequencing of the PCR products. Antibodies to N. caninum and T. gondii were also investigated in the tested animals. Brains and hearts were obtained from 100 beef cattle in a slaughterhouse in Bahia. Seven samples from brain tested positive for Toxoplasmatinae DNA. No positive reactions were found in heart tissues. After sequencing of the PCR products from all positive tissues, five sequences matched with N. caninum and two matched with T. gondii. Antibodies to N. caninum and T. gondii were found in 20% and 26% of the animals, respectively. The confirmation of N. caninum and the absence of Hammondia heydorni in the tested animals is suggestive that cattle are not efficient intermediate hosts of H. heydorni; however further studies need to be performed using a greater variety of tissues and a higher sample size. The detection of T. gondii DNA in bovine tissues reinforces the potential risk of transmission of this parasite to humans and other animals through the consumption of bovine meat.

Prevalence and risk factors associated to infection by Toxoplasma gondii in ovine in the State of Alagoas, Brazil.

The aim of this study was to evaluate the prevalence of anti-Toxoplasma gondii antibodies and to identify risk factors associated to the infection in the three meso-regions of the State of Alagoas, Brazil. A total count of 23 towns and 27 meat sheep farms were visited where blood samples were collected in order to perform the indirect immunofluorescence test to evaluate the antibodies presence. Questionnaires exploring the production system and nutritional, sanitary, and reproduction handling were handed out. The prevalence rate was 32.9% and the number of foci was 100%. In the multivariate statistical analysis, there was a significant association for the following variables: age (OR = 4.01; C.I. 2.03-7.94), size of the property (or the farm; OR = 0.48; C.I. 0.26-0.90), semi-intensive rearing system (OR = 3.17; C.I. 1.24-8.13), running water source (OR = 3.13; C.I.-1.66-5.87), and presence of cats (OR = 1.72; C.I. 1.08-2.75). It is concluded that sheep of the three meso-regions of the State of Alagoas are exposed to the infection caused by T. gondii with high prevalence. Control and prophylactic measures must be adopted seeking the improvement of the rearing system and the implantation of health promoting programs in cooperation with sheep farmers in order to elucidate the transmission means of this disease.

Occurrence of antibodies to Neospora caninum and Toxoplasma gondii in dogs from Pernambuco, Northeast Brazil.

A serological survey was carried out to assess the occurrence of anti-Neospora caninum antibodies in dogs from the State of Pernambuco. A total of 625 serum samples of dogs (289 from Paulista, 168 from Amaraji and 168 from Garanhuns) were tested by an immunofluorescence antibody assay for the detection of anti-N. caninum antibodies. A total of 177 (28.3%; IC 95%, 24.9-32.1) samples were positive. The seropositivity rates found in Paulista, Amaraji and Garanhuns were 26% (IC 95%, 21-31.4), 26.2% (IC 95%, 19.7-33.5) and 34.5% (IC 95%, 27.4-42.2), respectively. Of the 177 serum samples positive to anti-N. caninum antibodies, 170 were additionally tested for the presence of anti-Toxoplasma gondii antibodies and out of these 57.6% (IC 95%, 49.8-65.2) were positive. The results indicate that dogs from Amaraji, Paulista and Garanhuns are exposed to both N. caninum and T. gondii infections. The presence of dogs infected by N. caninum in Pernambuco represents a potential risk factor for the occurrence of outbreaks of abortion in cattle and small ruminants in this state. This study is the largest serological survey on the presence of anti-N. caninum antibodies in dogs carried out in Brazil and reports for the first time the exposure to N. caninum and T. gondii in dogs from Pernambuco.

Loss of infectivity of Neospora caninum oocysts maintained for a prolonged time.

The purpose of this study was to investigate whether sporulated Neospora caninum oocysts, which had been stored for 46 mo in a 2% sulfuric acid solution at 4 degrees C, remain morphologically viable and infective to gerbils (Meriones unguiculatus). Six gerbils were orally inoculated with doses of 400 or 1,200 oocysts. Two mo after inoculation, the animals did not show any clinical signs, had no histological lesions, and were seronegative for N. caninum at 1: 50 in an immunofluorescent antibody test. PCR using the brain from each gerbil did not reveal N. caninum specific DNA. We conclude that oocysts preserved for 46 mo are not infective, despite being morphologically intact.

Frequência de anticorpos anti-Neospora caninum em cães nos municípios de Salvador e Lauro de Freitas, Estado da Bahia - Brasil / Frequency of antibodies anti-Neospora caninum on dogs from Salvador and Lauro de Freitas, Bahia state - Brasil

A freqüência de anticorpos IgG anti-Neospora caninum foi estudada em 415 amostras séricas de cães domiciliados e errantes, procedentes dos municípios baianos de Salvador e Lauro de Freitas, utilizando-se a técnica de imunofluorescência indireta, com ponto de corte igual a 1:50. Anticorpos da classe IgG anti-N. caninum foram detectados em 13,3% (22/165) dos cães domiciliados e em 11,2% (28/250) dos errantes. As freqüências de soropositivos machos e fêmeas foram 8,0% (6/75) e 18,4% (14/76) nos cães domiciliados e 12,6% (17/135) e 9,6% (11/115) nos errantes, respectivamente. Não houve diferenças estatisticamente significativas entre sexo, idade, raça e a freqüência de soropositividade ao N. caninum dos cães domiciliados e errantes.

The frequency of anti-Neospora caninum IgG antibodies was studied in 415 serum samples from owned and stray dogs from Salvador and Lauro de Freitas counties. The Indirect immunofluorescence was performed using a cut-off of 1:50. Anti-N. caninum IgG antibodies were detected in 13.3% (22/165) owned dogs and in 11.2% (28/250) stray dogs. The frequencies of males and females seropositivity were 8.0% (6/75) and 18.4% (14/76) for owned dogs and 12.6 % (17/ 135) and 9.6% (11/115) in the stray dogs population, respectively. There were not a statistical significant difference between sex, age, breed and the dogs anti-N. caninum IgG antibodies incidence.

Freqüência de anticorpos IgG anti-Toxoplasma gondii em soros de cães errantes da cidade de Salvador-Bahia, Brasil / Frequency of IgG antibodies against-Toxoplasma gondii in sera of stray dogs in the city of Salvador-Bahia, Brazil

O Toxoplasma gondii é um coccídio intestinal intracelular obrigatório dos felídeos, de distribuição cosmopolita, descoberto em 1908 por Nicolle & Manceaux. O primeiro relato na espécie canina ocorreu em 1910, na Itália e, no Brasil em 1911. Objetivando-se avaliar a freqüência deste parasito, na população de cães errantes da cidade de Salvador-Ba, foram coletadas 225 amostras de sangue, de animais provenientes de 10 distritos sanitários. Os soros foram submetidos a reação de imunofluorescência indireta (RIFI) para detecção de anticorpos IgG anti-Toxoplasma gondii, utilizando-se a cepa AS28. Foram detectados 143 amostras positivas, representando uma freqüência de 63,55.00 por cento. As freqüências nos distritos sanitários foram as seguintes: Itapagipe 33,33 por cento; São Caetano/Valéria 46,15.00 por cento; Brotas 42,11 por cento; Barra/Rio Vermelho 64,28 por cento; Boca do Rio 80,00 por cento; Itapuã 65,38 por cento; Cabula/Beiru 80,64 por cento; Pau da Lima 73,91 por cento; Cajazeiras 64,70 por cento e Subúrbio Ferroviário 73,33 por cento. Os títulos encontrados variaram de 1:16 à 1:16384, sendo 1:16 (28,67 por cento), 1:64 (44,76 por cento), 1:256 (21,68 por cento), 1:1024 (4,20 por cento) e 1:16384 (0,70 por cento). Dos 123 machos e 102 fêmeas, 67,48 por cento e 58,82 por cento foram sororeagentes, respectivamente. Com relação à idade, dos 198 adultos e 27 jovens, 70,20 por cento e 14,80 por cento apresentaram-se soropositivos, respectivamente. As variáveis idade e distrito sanitário apresentaram associação estatisticamente significativa (p<0,05).