Molecular Characterization Of Intestinal Protozoa In Two Poor Communities In The State Of São Paulo, Brazil.

Several species of protozoa cause acute or chronic gastroenteritis in humans, worldwide. The burden of disease is particularly high among children living in developing areas of the world, where transmission is favored by lower hygienic standards and scarce availability of safe water. However, asymptomatic infection and polyparasitism are also commonly observed in poor settings. Here, we investigated the prevalence of intestinal protozoa in two small fishing villages, Porto Said (PS) and Santa Maria da Serra (SM), situated along the river Tietê in the State of São Paolo, Brazil. The villages lack basic public infrastructure and services, such as roads, public water supply, electricity and public health services. Multiple fecal samples were collected from 88 individuals in PS and from 38 individuals in SM, who were asymptomatic at the time of sampling and had no recent history of diarrheal disease. To gain insights into potential transmission routes, 49 dog fecal samples (38 from PS and 11 from SM) and 28 river water samples were also collected. All samples were tested by microscopy and PCR was used to genotype Giardia duodenalis, Blastocystis sp., Dientamoeba fragilis and Cryptosporidium spp. By molecular methods, the most common human parasite was Blastocystis sp. (prevalence, 45% in PS and 71% in SM), followed by D. fragilis (13.6% in PS, and 18.4% in SM) and G. duodenalis (18.2% in PS and 7.9% in SM); Cryptosporidium spp. were not detected. Sequence analysis revealed large genetic variation among Blastocystis samples, with subtypes (STs) 1 and 3 being predominant, and with the notable absence of ST4. Among G. duodenalis samples, assemblages A and B were detected in humans, whereas assemblages A, C and D were found in dogs. Finally, all D. fragilis samples from humans were genotype 1. A single dog was found infected with Cryptosporidium canis. River water samples were negative for the investigated parasites. This study showed a high carriage of intestinal parasites in asymptomatic individuals from two poor Brazilian villages, and highlighted a large genetic variability of Blastocystis spp. and G. duodenalis.810

Molecular characterization of intestinal protozoa in two poor communities in the state of Sao Paulo, Brazil

Several species of protozoa cause acute or chronic gastroenteritis in humans, worldwide. The burden of disease is particularly high among children living in developing areas of the world, where transmission is favored by lower hygienic standards and scarce availability of safe water. However, asymptomatic infection and polyparasitism are also commonly observed in poor settings. Here, we investigated the prevalence of intestinal protozoa in two small fishing villages, Porto Said (PS) and Santa Maria da Serra (SM), situated along the river Tiete in the State of Sao Paolo, Brazil. The villages lack basic public infrastructure and services, such as roads, public water supply, electricity and public health services. Multiple fecal samples were collected from 88 individuals in PS and from 38 individuals in SM, who were asymptomatic at the time of sampling and had no recent history of diarrheal disease. To gain insights into potential transmission routes, 49 dog fecal samples (38 from PS and 11 from SM) and 28 river water samples were also collected. All samples were tested by microscopy and PCR was used to genotype Giardia duodenalis, Blastocystis sp., Dientamoeba fragilis and Cryptosporidium spp. By molecular methods, the most common human parasite was Blastocystis sp. (prevalence, 45% in PS and 71% in SM), followed by D. fragilis (13.6% in PS, and 18.4% in SM) and G. duodenalis (18.2% in PS and 7.9% in SM); Cryptosporidium spp. were not detected. Sequence analysis revealed large genetic variation among Blastocystis samples, with subtypes (STs) 1 and 3 being predominant, and with the notable absence of ST4. Among G. duodenalis samples, assemblages A and B were detected in humans, whereas assemblages A, C and D were found in dogs. Finally, all D. fragilis samples from humans were genotype 1. A single dog was found infected with Cryptosporidium canis. River water samples were negative for the investigated parasites. This study showed a high carriage of intestinal parasites in asymptomatic individuals from two poor Brazilian villages, and highlighted a large genetic variability of Blastocystis spp. and G. duodenalis8COORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPESFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESPsem informação2011/52100-

In vitro ANTIGIARDIAL ACTIVITY OF THE CYSTEINE PROTEASE INHIBITOR E-64

The quest for new antiparasitic alternatives has led researchers to base their studies on insights into biology, host-parasite interactions and pathogenesis. In this context, proteases and their inhibitors are focused, respectively, as druggable targets and new therapy alternatives. Herein, we proposed to evaluate the in vitro effect of the cysteine protease inhibitor E-64 on Giardia trophozoites growth, adherence and viability. Trophozoites (105) were exposed to E-64 at different final concentrations, for 24, 48 and 72 h at 37 °C. In the growth and adherence assays, the number of trophozoites was estimated microscopically in a haemocytometer, whereas cell viability was evaluated by a dye-reduction assay using MTT. The E-64 inhibitor showed effect on growth, adherence and viability of trophozoites, however, its better performance was detected in the 100 µM-treated cultures. Although metronidazole was more effective, the E-64 was shown to be able to inhibit growth, adherence and viability rates by ≥ 50%. These results reveal that E-64 can interfere in some crucial processes to the parasite survival and they open perspectives for future investigations in order to confirm the real antigiardial potential of the protease inhibitors.As cisteína-proteases estão entre os alvos mais promissores para o desenvolvimento de novos agentes terapêuticos, visto que participam de eventos fundamentais do ciclo de vida de muitos microorganismos, inclusive Giardia. Como a atividade das proteases pode ser controlada por inibidores específicos, essas substâncias têm sido avaliadas quanto ao potencial antiparasitário. Diante disso, o presente estudo teve por objetivo avaliar o efeito in vitro do inibidor de cisteína-proteases E-64 sobre o crescimento, a aderência e a viabilidade de trofozoítos de cepa de Giardia isolada em Botucatu. Nos ensaios de crescimento e aderência, o número de trofozoítos foi estimado microscopicamente em hemocitômetro, enquanto que a viabilidade celular foi avaliada pelo método do MTT. No presente estudo, embora o metronidazol tenha se apresentado bastante efetivo, o E-64 mostrou ser capaz de inibir o crescimento, a aderência e a viabilidade em taxas superiores a 50%, especialmente nos cultivos expostos à concentração de 100 µM. A despeito de preliminares, esses resultados demonstram que o inibidor E-64 pode interferir em processos primordiais para a sobrevivência do parasita, além do que, abrem novas perspectivas para investigações futuras a fim de se avaliar o real potencial giardicida dos inibidores de proteases

Atividade in vitro do inibidor de cisteína-proteases E-64 sobre trofozoítos de Giardia

As cisteína-proteases estão entre os alvos mais promissores para o desenvolvimento de novos agentes terapêuticos, visto que participam de eventos fundamentais do ciclo de vida de muitos microorganismos, inclusive Giardia. Como a atividade das proteases pode ser controlada por inibidores específicos, essas substâncias têm sido avaliadas quanto ao potencial antiparasitário. Diante disso, o presente estudo teve por objetivo avaliar o efeito in vitro do inibidor de cisteína-proteases E-64 sobre o crescimento, a aderência e a viabilidade de trofozoítos de cepa de Giardia isolada em Botucatu. Nos ensaios de crescimento e aderência, o número de trofozoítos foi estimado microscopicamente em hemocitômetro, enquanto que a viabilidade celular foi avaliada pelo método do MTT. No presente estudo, embora o metronidazol tenha se apresentado bastante efetivo, o E-64 mostrou ser capaz de inibir o crescimento, a aderência e a viabilidade em taxas superiores a 50%, especialmente nos cultivos expostos à concentração de 100 µM. A despeito de preliminares, esses resultados demonstram que o inibidor E-64 pode interferir em processos primordiais para a sobrevivência do parasita, além do que, abrem novas perspectivas para investigações futuras a fim de se avaliar o real potencial giardicida dos inibidores de proteases.The quest for new antiparasitic alternatives has led researchers to base their studies on insights into biology, host-parasite interactions and pathogenesis. In this context, proteases and their inhibitors are focused, respectively, as druggable targets and new therapy alternatives. Herein, we proposed to evaluate the in vitro effect of the cysteine protease inhibitor E-64 on Giardia trophozoites growth, adherence and viability. Trophozoites (105) were exposed to E-64 at different final concentrations, for 24, 48 and 72 h at 37 °C. In the growth and adherence assays, the number of trophozoites was estimated microscopically in a haemocytometer, whereas cell viability was evaluated by a dye-reduction assay using MTT. The E-64 inhibitor showed effect on growth, adherence and viability of trophozoites, however, its better performance was detected in the 100 µM-treated cultures. Although metronidazole was more effective, the E-64 was shown to be able to inhibit growth, adherence and viability rates by ≥ 50%. These results reveal that E-64 can interfere in some crucial processes to the parasite survival and they open perspectives for future investigations in order to confirm the real antigiardial potential of the protease inhibitors

S-100 dendritic cells in normal and Dermatobia hominis infested cattle skin

Foram investigadas as células dendríticas (CD) na pele normal de cinco bezerros das raças Nelore, cinco da raça Holandesa Preta e Branca e cinco animais mestiços por meio da imunomarcação pela proteína S-100. Os animais mestiços foram infestados experimentalmente com 100 larvas de primeiro estádio de Dermatobia hominis e deles foram colhidas biópsias de pele parasitada às 24, 48, 72 e 168 horas após a infestação. Biópsias de pele destes animais, colhidas antes da infestação, foram utilizadas como controle. A imunomarcação das CDs foi feita empregando-se anticorpos de coelhos antiproteína S-100 e a técnica da avidina-biotina-peroxidase. Além das CDs, melanócitos, nervos e células endoteliais apresentaram imunomarcação pela proteína S-100. As DCs foram observadas exclusivamente na derme superficial, próximas à camada basal, tanto nos animais infestados como nos não-infestados. Não se detectou diferença significativa no número de CDs que pudesse ser atribuída à raça dos animais. Nos animais parasitados por D. hominis, as CDs apresentavam-se mais intensamente coradas e com os prolongamentos mais espessos do que nos controles não-parasitados. Além disso, nos animais parasitados observou-se um decréscimo significativo no número de CDs a partir de 24 horas após a infestação

De ante acta vita

Nella concezione del processo penale romano più antico, come è noto, largo spazio avevano gli elementi emozionali volti a commuovere i giudici. Tra questi, i racconti della vita passata degli accusati. L'accusa e la difesa utilizzavano spregiudicatamente questo artifizio retorico. Nel periodo intermedio si registra la tendenza a costruire la vita ante acta come presunzione

Eficácia terapêutica e residual de seis formulações inseticidas sobre o parasitismo por larvas de Dermatobia hominis em bovinos

The immediate efficacy and residual effect of six commercially available parasiticides to control infestations by Dermatobia hominis were evaluated in a Nelore cattle herb. Fortyeight naturally infested females were divided into six groups of eight animals on the basis of the number of D. hominis nodules present at the day of treatment. The number of cutaneous nodules was determined by manual counting of larvae on both side of each animal. The counting was performed before treatment and weekly after the treatment in the follow six weeks. The animals were dosed with commercial products containing the following principles and concentrations: Trichlorfon (1 mL/4 kg), DDVP (15 mL/10 L), Cipermetrin/Piperonil Butoxide(10 mL/10 L), Ivermectin 1% (200 mcg/kg), Doramectin 1% (200 mcg/kg) e Doramectin 1% (50 mcg/kg). All the treatments produced a significant reduction of larval infestation (p<0.01) in all animal groups. The immediate and residual effectiveness showed no statistical difference (p<0.05) contrasting with the empirical perception by which the macrocyclic lactones provides a higher effectiveness compared to organophosphates and pyrethroids drugs. Data obtained in this work showed that there is no evidence of resistance of D. hominis to any of the drugs tested.Se evaluaran la eficacia inmediata y el efecto residual de seis formulaciones químicas disponibles comercialmente para el control de infestaciones de larvas de Dermatobia hominis. Cuarenta y ocho hembras de la raza Nelore, infestadas naturalmente, fueran divididas en seis grupos de ocho animales, basados en el número de larvas presentes en el día de tratamiento. La contaje del número de larvas se realizó en ambos lados de los animales antes del tratamiento y cada semana después del tratamiento hasta la sexta semana. Los animales fueran dosificados con los siguientes ingredientes activos y respectivas concentraciones: triclorfón (1 ml / 4 kg), DDVP (15 ml/10 L), Cipermetrina / butóxido de piperonilo (10 ml/10 L), 1% de ivermectina (200 mcg / kg) Doramectina 1% (200 mcg / kg) y doramectina 1% (50 mcg / kg). La análisis de los resultados reveló que todos los tratamientos produjeron una reducción significativa de la infestación (p<0,01), sin diferencia estadística entre los productos. Esta falta de diferencia entre los distintos tratamientos (p<0,05) contrastó con la percepción visual y empírica de que las lactonas macrocíclicas fueran más eficaces que los organofosfatos o piretroides. Los resultados obtenidos refutan la sospecha de resistencia a las sustancias ensayadas.A eficácia imediata e o poder residual de seis formulações químicas comercialmente disponíveis para o controle de infestações por larvas de Dermatobia hominis foram avaliados em bovinos da raça Nelore. Quarenta e oito fêmeas, naturalmente infestadas, foram divididas em seis grupos de oito animais com base no número de larvas presentes no dia do tratamento. A contagem do número de larvas foi feita em ambos os lados dos animais antes do tratamento e semanalmente após o tratamento até a sexta semana. Os animais foram tratados com produtos contendo os seguintes princípios ativos e respectivas concentrações: Triclorfon (1 ml/4 kg), DDVP (15 mL/10 L), Cipermetrina/Butóxido de Piperonila (10 mL/10 L), Ivermectina 1% (200 mcg/kg), Doramectina 1% (200 mcg/kg) e Doramectina 1% (50 mcg/kg). As análises dos resultados revelaram que todos os tratamentos produziram redução significativa das infestações (p<0,01), não havendo diferença de eficácia entre os produtos. A inexistência de diferença estatística entre a eficácia imediata e residual proporcionada pelos diferentes tratamentos (p<0,05) contrastou com a percepção visual empírica de que as lactonas macrocíclicas teriam apresentado eficácia maior que os produtos a base de organofosforados ou piretróides. Os resultados obtidos não sustentam a suspeita de resistência a nenhuma das substâncias testadas