Soil contamination by Ancylostoma spp. and Toxocara spp. eggs in elementary school playgrounds in the extreme south of Brazil.

Millions of people worldwide, and especially schoolchildren, may be infected by geohelminths due to their exposure to a contaminated environment. The aim of this study was to evaluate soil contamination by Ancylostoma spp. and Toxocara spp. eggs in recreation areas at elementary schools in Pelotas, state of Rio Grande do Sul, Brazil. Sand samples were collected from 22 schools and were processed using the centrifugal flotation method. Helminth eggs with zoonotic potential were found in 12 out of the 22 schools (54.5%). Contamination by Ancylostoma spp. and Toxocara spp. was observed in 36.4% (8/22) and 27.3% (6/22) of the soil samples collected at these schools, respectively. These findings of eggs show that the school communities are exposed to risks of zoonotic transmission.

Soil contamination by Ancylostoma spp. and Toxocara spp. eggs in elementary school playgrounds in the extreme south of Brazil / Contaminação do solo de praças de recreação de escolas de ensino fundamental por ovos de Ancylostoma spp. e Toxocara spp. no extremo sul do Brasil

Abstract Millions of people worldwide, and especially schoolchildren, may be infected by geohelminths due to their exposure to a contaminated environment. The aim of this study was to evaluate soil contamination by Ancylostoma spp. and Toxocara spp. eggs in recreation areas at elementary schools in Pelotas, state of Rio Grande do Sul, Brazil. Sand samples were collected from 22 schools and were processed using the centrifugal flotation method. Helminth eggs with zoonotic potential were found in 12 out of the 22 schools (54.5%). Contamination by Ancylostoma spp. and Toxocara spp. was observed in 36.4% (8/22) and 27.3% (6/22) of the soil samples collected at these schools, respectively. These findings of eggs show that the school communities are exposed to risks of zoonotic transmission.

Resumo Milhões de pessoas podem ser acometidas por geohelmintos, especialmente crianças em idade escolar, devido a sua maior exposição a ambientes contaminados. O objetivo deste estudo foi avaliar a contaminação do solo por ovos de Ancylostoma spp. e Toxocara spp. em áreas de recreação de escolas de ensino fundamental da cidade de Pelotas, Rio Grande do Sul, Brasil. Foram colhidas amostras de areia de 22 escolas e processadas pelo método de centrífugo-flutuação. Em 54,5% (12/22) das escolas houve registro da presença de ovos de helmintos com potencial zoonótico. A contaminação por Ancylostoma spp. e Toxocara spp. foi observada em 36,4% (8/22) e 27,3% (6/22) das amostras de solo das escolas, respectivamente. Existe a presença de ovos de Ancylostoma spp. e Toxocara spp., havendo risco de transmissão de zoonoses à comunidade escolar.

Helminth eggs with zoonotic potential in the vicinity of public schools in southern Brazil.

The aim of this study was to evaluate environmental contamination by helminth eggs with zoonotic potential that were found in dog feces in the vicinity of elementary schools. Seventy-nine samples of dog feces were collected from 28 municipal schools located in five neighborhoods in Pelotas, Rio Grande do Sul (RS), Brazil. The samples were processed using the Willis-Mollay technique and analyzed using an optical microscope (40X), to identify any parasite eggs present. All neighborhoods were positive and 74.7% of the samples exhibited one or more helminth genera. The agent with the highest prevalence was Ancylostoma spp. (93.2%), followed by Trichuris spp. (18.6%), Toxocara spp. (11.9%) and Toxascaris (1.7%). These data show that there is a need for greater care towards controlling these helminths with zoonotic potential, including responsible pet ownership and daily activities to clean and collect dog feces in the vicinity of schools, because these are places where children play and study.

Helminth eggs with zoonotic potential in the vicinity of public schools in southern Brazil / Ovos de helmintos com potencial zoonótico nas proximidades de escolas públicas no extremo sul do Brasil

Abstract The aim of this study was to evaluate environmental contamination by helminth eggs with zoonotic potential that were found in dog feces in the vicinity of elementary schools. Seventy-nine samples of dog feces were collected from 28 municipal schools located in five neighborhoods in Pelotas, Rio Grande do Sul (RS), Brazil. The samples were processed using the Willis-Mollay technique and analyzed using an optical microscope (40X), to identify any parasite eggs present. All neighborhoods were positive and 74.7% of the samples exhibited one or more helminth genera. The agent with the highest prevalence was Ancylostoma spp. (93.2%), followed by Trichuris spp. (18.6%), Toxocara spp. (11.9%) and Toxascaris (1.7%). These data show that there is a need for greater care towards controlling these helminths with zoonotic potential, including responsible pet ownership and daily activities to clean and collect dog feces in the vicinity of schools, because these are places where children play and study.

Resumo O objetivo deste estudo foi avaliar a contaminação ambiental por ovos de helmintos com potencial zoonótico, diagnosticados em fezes caninas depositadas nas proximidades de escolas primárias. Setenta e nove amostras de fezes foram colhidas em 28 escolas municipais localizadas em cinco bairros da cidade de Pelotas, Rio Grande do Sul (RS), Brasil. As amostras foram processadas pela técnica de Willis-Mollay e analisadas em microscópio óptico (40X), para identificar quaisquer parasitos presentes. Todos os bairros foram positivos e 74,7% das amostras apresentaram um ou mais gêneros de helmintos. O agente com maior prevalência foi Ancylostoma spp. (93,2%), seguido por Trichuris spp. (18,6%), Toxocara spp. (11,9%) e Toxascaris (1,7%). Esses dados mostram que há necessidade de maior cuidado no controle desses helmintos com potencial zoonótico, incluindo a posse responsável dos animais e atividades diárias de limpeza e colheita de fezes de cães nas proximidades das escolas, porque esses são locais onde as crianças brincam e estudam.

Expressão heteróloga da EMA-2 (equi merozoite antigen) de Theileria equi em Pichia pastoris com potencial utilização em imunobiológicos / Heterologous expression of EMA-2 (equi merozoite antigen) of Theileria equi in Pichia pastoris with potential use in immunobiologics

A piroplasmose equina causada por Theileria equi acomete os equinos de forma endêmica no Brasil e em diversos outros países tropicais e subtropicais. Considerada uma das mais importantes doenças de equinos, causa danos à saúde animal e perdas econômicas. A proteína equi merozoite antigen (EMA-2) é uma das principais proteínas de superfície, expressa nos diversos estágios do ciclo do parasita, estimula resposta imune em animais infectados, tornando-se um possível candidato para utilização em diagnóstico. O gene EMA-2 foi clonado e expresso na levedura Pichia pastoris. A proteína EMA-2 recombinante (rEMA-2) foi caracterizada antigenicamente por Western Blot e por ELISA indireto, utilizando-se soro de equino positivo para theileriose. O resultado do ELISA demonstrou uma especificidade de 90,9% e sensibilidade de 83,3%, quando comparado ao padrão, sendo superior à imunofluorescência (80,6% de especificidade e 75,0% de sensibilidade), o que sugere que a rEMA-2 expressa em P. pastoris é um promissor antígeno para ser utilizado como ferramenta no imunodiagnóstico de theileriose equina.

Theileria equi, the causative agent of equine piroplasmosis, is endemic in Brazil and many other tropical and subtropical countries. It is considered one of the most important diseases of horses causing animal health problems and significant economic loss. The Protein equi merozoite antigen-2 (EMA-2) is a major surface protein that is expressed in different parasite cycle stages and induces immune response in infected animals, being a possible candidate to be used in diagnose. EMA-2 gene was cloned and expressed in the yeast Pichia pastoris, and the recombinant protein EMA-2 (rEMA-2) was characterized by Western Blot and indirect ELISA using equine positive sera. The ELISA results demonstrated a specificity of 90.9% and a sensitivity of 83.3% compared to the standard ELISA and being superior to immunofluorescence (80.6% of specificity and 75.0% of sensitivity) suggesting that the rEMA-2 expressed in P. pastoris is a promising antigen to be used as a tool in immunodiagnostic of theileriasis.

Comparison of three larval bioassays to evaluate susceptibility of Rhipicephalus (Boophilus) microplus to amitraz / Comparação de três bioensaios larvais para avaliar a suscetibilidade de Rhipicephalus (Boophilus) microplus ao amitraz

Field samples of Rhipicephalus (Boophilus) microplus from the state of Rio Grande do Sul, Brazil, were assessed using the following methods: larval packet test (LPT), larval immersion test (LIT) and syringe immersion test (SIT). The following parameters were determined for each population and for the Mozo susceptible reference strain: lethal concentration for 50% (LC50) with its 95% confidence interval (95% CI), regression line slope and resistance ratio (RR). Using the LPT, only one population was susceptible to amitraz, presenting a RR of 1.9. Using the same technique, the other populations presented RRs of between 92.9 and 3445.8 and were considered resistant. The LC50 of the Mozo strain calculated using the LPT, LIT and SIT was 2.9, 27.3, and 52.7 µg/mL, respectively. In general, a good fit to the probit statistical model was only achieved using the LPT. The results obtained in this study impair recommendations for using the LIT and SIT to diagnose amitraz resistance in R. (B.) microplus populations. Additional studies are required to improve the sensitivity of these tests in relation to the LPT.

Amostras de Rhipicephalus (Boophilus) microplus coletadas à campo no Rio Grande do Sul, Brasil, foram analisadas pelos seguintes métodos: teste do pacote de larvas (TPL), teste de imersão de larvas (TIL) e teste de imersão em seringas (TIS). Os seguintes parâmetros foram determinados para cada população e para a amostra referência suscetível Mozo: concentração letal para 50% (CL50) e seu intervalo de confiança de 95% (IC95%), inclinação da reta de regressão e os fatores de resistência (FR). Pelo TPL, apenas uma população foi sensível ao amitraz, com FR de 1,9. Utilizando a mesma técnica, as outras amostras apresentaram FR entre 92,9 e 3445,8 sendo consideradas resistentes. As CL50 da cepa Mozo calculadas por meio do TPL, TIL e TIS foram 2,9, 27,3 e 52,7 µg/mL, respectivamente. De forma geral, a adequação ao modelo estatístico de probitos só foi alcançada com o uso do TPL. Os resultados obtidos no presente estudo limitam a recomendação de uso do TIL e TIS para diagnóstico de resistência ao amitraz em populações de R. (B.) microplus. Estudos adicionais são necessários para aprimorar a sensibilidade destes testes em relação ao LPT.

Comparison of three larval bioassays to evaluate susceptibility of Rhipicephalus (Boophilus) microplus to amitraz.

Field samples of Rhipicephalus (Boophilus) microplus from the state of Rio Grande do Sul, Brazil, were assessed using the following methods: larval packet test (LPT), larval immersion test (LIT) and syringe immersion test (SIT). The following parameters were determined for each population and for the Mozo susceptible reference strain: lethal concentration for 50% (LC50) with its 95% confidence interval (95% CI), regression line slope and resistance ratio (RR). Using the LPT, only one population was susceptible to amitraz, presenting a RR of 1.9. Using the same technique, the other populations presented RRs of between 92.9 and 3445.8 and were considered resistant. The LC50 of the Mozo strain calculated using the LPT, LIT and SIT was 2.9, 27.3, and 52.7 µg/mL, respectively. In general, a good fit to the probit statistical model was only achieved using the LPT. The results obtained in this study impair recommendations for using the LIT and SIT to diagnose amitraz resistance in R. (B.) microplus populations. Additional studies are required to improve the sensitivity of these tests in relation to the LPT.

Cloning and expression of a truncated form of envelope glycoprotein D of Bovine herpesvirus type 5 in methylotrophic yeast Pichia pastoris.

Meningoencephalitis caused by Bovine herpesvirus type 5 (BoHV-5) is responsible for heavy economic losses in the cattle industry. As in other Alphaherpesviruses, the envelope glycoprotein IV (gD), which mediates penetration into host cells, is one of the major candidate antigens for a recombinant vaccine, since it induces a strong and persistent immune response. The DNA coding for a truncated form of BoHV-5 gD (tgD) has been cloned into the Pichia pastoris expression vector pPICZalphaB to allow protein secretion into the medium. After induction with methanol, a approximately 55kDa protein was obtained. Enzyme deglycosylation with Endo H showed a smaller size band in SDS-PGAE, with approximately 50kDa, suggesting that tgD has N-linked oligosaccharides and that it is not hyperglycosylated. The approximately 55kDa protein was recognized by several polyclonal antibodies, including polyclonal antibody anti-tgD and polyclonal antibodies of different animal species immunized with BoHV-5 and BoHV-1. This is the first report of BoHV-5 gD expression in yeast. It was shown that the recombinant truncated form of BoHV-5 gD has antigenic and immunogenic properties similar to the native BoHV-5 gD. Expression of tgD as a secreted protein allows simple and inexpensive purification methods that can be used for further studies to evaluate its immunogenicity in cattle.

Frequency of seropositive equines for Theileria equi in the Southern Rio Grande do Sul State, Brazil

Foi realizado um levantamento sobre theileriose equina em uma propriedade da região sul do estado do Rio Grande do Sul (RS). Amostras de sangue foram coletadas de 108 éguas de um haras localizado no municipio de Bagé, latitude 31°30' Sul e longitude 54Ú10' Oeste. A sorologia foi realizada com o uso da imunofluorescência indireta (IFAT) e utilizou-se a técnica de Nested Reacão em Cadeia Polimerase (nPCR). Do total de amostras examinadas, 22 por cento (24) foram positivas por IFAT e 15 por cento (16) por nPCR). Na análise por raca, 15.05 por cento (14) dos animais Puro Sangue Inglês (PSI) foram soro-positivos, e dos animais da raça Crioula (RC), 55 por cento (11) foram positivos por IFAT.

A study on equine theileriosis was carried out in the southern region of the Brazilian state of Rio Grande do Sul (RS). Blood samples were collected from 113 mares from an equine breeding farm located in the city ofBagé, latitude 31 "30'S and a longitude of 54 U10' W. The serological testing was carried out with the use of indirect fluorescence test (IFAT) and compared with Nested Polimerase Chain Reaction (nPCR). Among the sera collected from 118 horses, 25 were found positive to Theileria equi by the IFAT, while by nPCR 17 positive for T. equi was observed, corresponding to a frequency of22.1 percent> and 15.0 percent, respectively. The racial analysis showed 15.05 percent (14) thoroughbred and 55 percent (11) Crioulo breed horses to be positive by IFAT.

Presence of Gasterophilus (Leach, 1817) (Díptera: Oestridae) in horses in Rio Grande do Sul State, Brazil

With the purpose of verify the presence of larvae from the genus Gasterophilus in the Rio Grande do Sul State, a study was undertaken using 395 horses taken to slaughter in the city of Pelotas. The material analysis revealed that 126 animals (31,90 percent) presented infection, with 100 animals (25, infected by Gasterophilus nasalis, and 47 animals (11,90 percent) infected by G intestinalis. These results also represent the first report of the definite establishment of G intestinalis as a horse parasite in Brazil. The probable implications of G. intestinalis occurrence in Brazil are discussed.

Com o objetivo de verificar a presença de larvas do gênero Gasterophilus no Estado do Rio Grande do Sul, foi realizado um estudo com 395 equinos sacrificados em um abatedouro da cidade de Pelotas. O material analisado revelou que 126 animáis (31,90 por cento) apresentaram a infecção, sendo que 100 animáis (25,32 por cento) estavam infectados por Gasterophilus nasalis e 47 (11,90 por cento) por G intestinalis. Estes resultados também representan o primeiro relato definitivo de estabelecimento de G. intestinalis como parásito de equinos no Brasil. As implicações prováveis da presença de G. intestinalis no Brasil são discutidas.

Chronic fascioliasis in farmed and wild greater rheas (Rhea americana).

From 50 farmed Rhea americana slaughtered for human consumption, adult forms and eggs of Fasciola hepatica were found in 4. The other three livers were free of flukes but did show lesions caused by larval fluke migration. Histological lesions were similar to those caused by flukes in cattle and sheep. The rheas were from an endemic area of ruminant fascioliasis in Southern Brazil. F. hepatica eggs were also found in faecal samples of wild rheas from another endemic area in Southern Brazil. It is likely that the rheas play a role in the transmission of the disease to ruminants and could be jeopardizing the control of this parasitosis in endemic areas. From the best of our knowledge this is the first report of fascioliasis in R. americana.