RESUMO

Echinococcus granulosus sensu lato (s.l.) is a species complex with the potential to cause cystic echinococcosis (CE). Contact with the feces of domestic dogs (Canis familiaris) fed with raw viscera of intermediate livestock hosts is a risk factor for this infection in the southern region of Brazil. Although the region has been considered endemic to CE for many years, molecular data regarding the species of the complex causing CE in humans are scarce. This study aimed to perform a molecular analysis of the biological fluid from a human liver cyst to investigate the species responsible for CE. Genetic material obtained from the hydatid fluid of a hepatic cyst from a human with CE was subjected to PCR to amplify mitochondrial and nuclear DNA sequences. The phylogenetic analysis confirmed the human infection by Echinococcus canadensis G7 in the state of Paraná, Brazil. This is the first molecular record of E. canadensis G7 infecting a human in Brazil, and it is important to reiterate the risk of human CE caused by this species in South America, as reported by a previous study in Patagonia, Argentina. From the epidemiological point of view, this finding is of great relevance for the southern region of Brazil, since this parasite has previously only been detected in pigs in the state of Rio Grande do Sul, neighboring Paraná. The finding points to the importance of this identification in the molecular epidemiology of E. granulosus s.l., especially in South America.

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The coproparasitological examination of dogs (n=278) from two Brazilian biomes (Amazon [AZ] and Atlantic Forest [AF]) by centrifugal flotation demonstrated positivity values of 54.2% (AF) and 48.5% (AZ). The most prevalent parasites in AF were hookworms (81.0% - 47/58), Toxocara sp. (17.3% - 10/58) and Trichuris vulpis (12.1% - 7/58); while in AZ they were hookworms (86.7% - 72/83), Toxocara sp. (18.1% - 15/83), Dipylidium caninum (13.3% - 11/83) and T. vulpis (10.8% - 9/83). PCR was performed using the partial mitochondrial genes cytochrome c oxidase subunit 1 (pcox1) and NADH dehydrogenase 1 (pnad1) in 25 fecal samples positive for Toxocara sp. eggs and found one sample positive for pcox1 and six positives for pnad1. The sequencing of these samples was unsuccessful due to the difficulties inherent in copro-PCR+sequencing. The sequencing of 14 samples of T. canis adult helminths retrieved 11 sequences of 414 bp for pcox1 and nine sequences of 358 bp for pnad1. The phylogenetic trees of these sequences confirmed the species T. canis. Intraspecific genetic variation was only observed for pnad1. This is the second study involving molecular analysis of T. canis in dogs from Brazil and adds new information through the use of pnad1.

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Human polycystic echinococcosis is a parasitic infection caused by the larval stage of Echinococcus vogeli, which occurs in rural areas of Central and South America. Until now, little information on the genetic variability of E. vogeli is available. Here, 32 samples from human-excised E. vogeli cysts had a 396-bp sequence of the mitochondrial cytochrome oxidase I (COI) gene sequenced and compared to another 17 COI sequences representing nine Echinococcus species. A Bayesian COI tree revealed that all E. vogeli sequences formed a monophyletic and well-supported clade with an E. vogeli reference sequence. The occurrence of geographically restricted E. vogeli COI haplotypes suggests retention of ancestral polymorphisms with little migration in Acre, Brazil.

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Although sheep farming has grown in the state of Acre over the past four decades, little is known about occurrences of helminthiases in the herds of this region. The objective of the study was to assess the occurrences of non-intestinal helminthiasis among sheep slaughtered in Rio Branco. A total of 110 sheep livers were inspected from two slaughter batches (july 2014 and march 2015) in a slaughterhouse in Rio Branco. Livers with macroscopic lesions were photographed and were then subjected to histopathological analysis under an optical microscope. The macroscopic lesions showed small nodes with inflammatory characteristics and areas of fibrosis, which appeared to be calcified, thus suggesting a granulomatous reaction. Of the 110 evaluated livers, we noticed 110 nodules in total; these nodules have an average size of 0.5 cm. The histopathological analysis showed alterations to the architecture of the hepatic lobe, with multiple foci of necrosis and polymorphonuclear cells. Two samples revealed the presence of helminths from Nematode class and Capillaria sp. eggs identified by the typical morphology and morphometry. This seems to be the first report of Capillaria sp. in sheep livers in Brazil, and it serves as an important alert regarding animal health surveillance and control and regarding the Capillaria sp. zoonotic role in humans.

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RESUMO

Although sheep farming has grown in the state of Acre over the past four decades, little is known about occurrences of helminthiases in the herds of this region. The objective of the study was to assess the occurrences of non-intestinal helminthiasis among sheep slaughtered in Rio Branco. A total of 110 sheep livers were inspected from two slaughter batches (july 2014 and march 2015) in a slaughterhouse in Rio Branco. Livers with macroscopic lesions were photographed and were then subjected to histopathological analysis under an optical microscope. The macroscopic lesions showed small nodes with inflammatory characteristics and areas of fibrosis, which appeared to be calcified, thus suggesting a granulomatous reaction. Of the 110 evaluated livers, we noticed 110 nodules in total; these nodules have an average size of 0.5 cm. The histopathological analysis showed alterations to the architecture of the hepatic lobe, with multiple foci of necrosis and polymorphonuclear cells. Two samples revealed the presence of helminths from Nematode class and Capillaria sp. eggs identified by the typical morphology and morphometry. This seems to be the first report of Capillaria sp. in sheep livers in Brazil, and it serves as an important alert regarding animal health surveillance and control and regarding the Capillaria sp. zoonotic role in humans.(AU)

Embora a ovinocultura tenha despertado o interesse de criadouros no estado do Acre nas últimas quatro décadas, pouco se conhece sobre a ocorrência de helmintoses no plantel de ovinos dessa região. O objetivo do presente estudo foi avaliar a possibilidade de ocorrência de helmintíases não intestinais entre ovinos abatidos no município de Rio Branco. Foram inspecionados 110 fígados de ovinos em dois abates (julho de 2014 e março de 2015) em um abatedouro no município de Rio Branco. Fígados com lesões macroscópicas foram fotografados com posterior análise histopatológica por microscopia de luz. Nas lesões macroscópicas foram encontrados pequenos nódulos apresentando características inflamatórias com áreas de fibrose, aparentemente calcificadas, sugerindo uma reação granulomatosa. Dos 110 fígados avaliados, observou-se 110 nódulos no total; estes nódulos têm um tamanho médio de 0,5 cm. A análise histopatológica mostrou alterações na arquitetura do lóbulo hepático, com múltiplos focos de necrose, além da formação de abscessos hepáticos constituídos por polimorfonucleares. Duas amostras revelaram a presença de helmintos da Classe Nematoda e ovos de Capillaria sp. identificados pela morfologia típica e morfometria. Esse resultado parece ser o primeiro registro de Capillaria sp. em fígado de ovino no Brasil, o que é um importante alerta para a vigilância no controle sanitário animal e o seu papel zoonótico para humanos.(AU)

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Abstract Although sheep farming has grown in the state of Acre over the past four decades, little is known about occurrences of helminthiases in the herds of this region. The objective of the study was to assess the occurrences of non-intestinal helminthiasis among sheep slaughtered in Rio Branco. A total of 110 sheep livers were inspected from two slaughter batches (july 2014 and march 2015) in a slaughterhouse in Rio Branco. Livers with macroscopic lesions were photographed and were then subjected to histopathological analysis under an optical microscope. The macroscopic lesions showed small nodes with inflammatory characteristics and areas of fibrosis, which appeared to be calcified, thus suggesting a granulomatous reaction. Of the 110 evaluated livers, we noticed 110 nodules in total; these nodules have an average size of 0.5 cm. The histopathological analysis showed alterations to the architecture of the hepatic lobe, with multiple foci of necrosis and polymorphonuclear cells. Two samples revealed the presence of helminths from Nematode class and Capillaria sp. eggs identified by the typical morphology and morphometry. This seems to be the first report of Capillaria sp. in sheep livers in Brazil, and it serves as an important alert regarding animal health surveillance and control and regarding the Capillaria sp. zoonotic role in humans.

Resumo Embora a ovinocultura tenha despertado o interesse de criadouros no estado do Acre nas últimas quatro décadas, pouco se conhece sobre a ocorrência de helmintoses no plantel de ovinos dessa região. O objetivo do presente estudo foi avaliar a possibilidade de ocorrência de helmintíases não intestinais entre ovinos abatidos no município de Rio Branco. Foram inspecionados 110 fígados de ovinos em dois abates (julho de 2014 e março de 2015) em um abatedouro no município de Rio Branco. Fígados com lesões macroscópicas foram fotografados com posterior análise histopatológica por microscopia de luz. Nas lesões macroscópicas foram encontrados pequenos nódulos apresentando características inflamatórias com áreas de fibrose, aparentemente calcificadas, sugerindo uma reação granulomatosa. Dos 110 fígados avaliados, observou-se 110 nódulos no total; estes nódulos têm um tamanho médio de 0,5 cm. A análise histopatológica mostrou alterações na arquitetura do lóbulo hepático, com múltiplos focos de necrose, além da formação de abscessos hepáticos constituídos por polimorfonucleares. Duas amostras revelaram a presença de helmintos da Classe Nematoda e ovos de Capillaria sp. identificados pela morfologia típica e morfometria. Esse resultado parece ser o primeiro registro de Capillaria sp. em fígado de ovino no Brasil, o que é um importante alerta para a vigilância no controle sanitário animal e o seu papel zoonótico para humanos.

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Polycystic echinococcosis (PE) is caused by Echinococcus vogeli metacestodes (larval stage) in Neotropical countries. E. vogeli is trophically-transmitted between predators bush dogs (Speothos venaticus) and prey pacas (Cuniculus paca). In Brazil, reported PE cases are restricted to the Amazon biome. In this study, metacestodes from a paca hunted in Mato Grosso do Sul state (Cerrado biome) were identified morphological and histopathological techniques and further confirmed by molecular testing (sequencing of cytochrome C oxidase subunit I (cox1) gene) for the first time. Images of the whole liver showed superficial bubble-like hepatic masses. The parasitological analysis revealed large hooks (41.3 ±â€¯1.2 µm length/12.8 ±â€¯0.8 µm width) and small hooks (33.0 ±â€¯1.5 µm length/11.1 ±â€¯1.2 µm width), consistent with E. vogeli. Microscopically, the liver showed protoscoleces, a thick laminated layer, fibrosis, and inflammatory infiltrate in the adventitial layer. The DNA sequencing confirmed E. vogeli with 99% homology with sequences deposited in the GenBank. In addition, this finding greatly extends the geographic range of animal polycystic echinococcosis into the Cerrado. It is likely to occur in new biomes, where bush dogs and pacas share a given area in a trophic relationship.

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Fasciola hepatica is a trematode showing genetic variation among isolates from different regions of the world. The objective of this work was to characterize for the first time F. hepatica isolates circulating in different regions of Argentina. Twenty-two adult flukes were collected from naturally infected bovine livers in different areas from Argentina and used for DNA extraction. We carried out PCR amplification and sequence analysis of the ribosomal internal transcribed spacer 1 (ITS1), mitochondrial nicotinamide adenine dinucleotide dehydrogenase subunits 4 and 5 (nad4 and nad5) and mitochondrial cytochrome c oxidase subunit I (cox1) genes as genetic markers. Phylogenies were reconstructed using maximum parsimony algorithm. A total of 6 haplotypes were found for cox1, 4 haplotypes for nad4 and 3 haplotypes for nad5. The sequenced ITS1 fragment was identical in all samples. The analyzed cox1 gene fragment is the most variable marker and is recommended for future analyses. No geographic association was found in the Argentinean samples.

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BACKGROUND: Echinococcus granulosus (sensu lato) (s.l.) and Echinococcus vogeli are causative agents of chronic zoonotic diseases such as cystic and polycystic echinococcosis, respectively. In Brazil, polycystic echinococcosis has a restricted geographical distribution in the North Region, while cystic echinococcosis is observed in the South Region. Domestic dogs (Canis familiaris) fed with raw viscera represent a risk factor for E. granulosus (s.l.) infection in the South Region. Although this practice is frequent, it remains unclear whether domestic dogs are infected with E. vogeli in the state of Acre, located in the Amazon basin in the North Region of Brazil. The aim of this study was to investigate this gap in the polycystic echinococcosis epidemiology. METHODS: Sixty-five fecal samples were collected from the ground in five municipalities (Sena Madureira, n = 14; Rio Branco, n = 06; Bujari, n = 06; Xapuri, n = 30; and Epitaciolândia, n = 09) located in the state of Acre, northern Brazil. The samples were screened for parasites by copro-PCR using the cox1 gene associated with automated sequencing. RESULTS: Echinococcus vogeli was molecularly confirmed in a sample from Sena Madureira and E. granulosus (sensu stricto) (s.s.) (G1) in a sample from Rio Branco. CONCLUSIONS: These findings indicate that molecular assays are useful in typing Echinococcus taxa from fecal samples of dogs in northern Brazil. The present study is the first molecular record of E. vogeli in domestic dogs found in the state of Acre, reinforcing their role as a source of infection for humans. Because E. granulosus (s.s.) (G1) was detected for the first time in the North Region, from the epidemiological standpoint this finding is highly relevant, because it expands the known geographical distribution, which was previously restricted to the South Region of Brazil.

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Chemotherapy of food-borne trematodes relies on two drugs, praziquantel and tricabendazole, and there is growing interest in finding alternative therapies. Plant oil extracts have long been used in traditional Chinese medicine as sources of bioactive compounds with antiparasitic activity. Species of the genus Echinostoma are used as good models to test effective compounds against food-borne trematodes. This study evaluated the anthelmintic activity of crude artesunate extracts in vitro on newly excysted metacercariae of Echinostoma paraensei by light and scanning electron microscopy (SEM). The flukes were incubated with 1 µg/mL, 10 µg/mL, 25 µg/mL, 50 µg/mL and 100 µg/mL of artesunate for 4, 12, 24, 48 and 72 h. When the exposure time and concentration of artesunate increased, there were changes in motor activity, tegument damage and death. Blebs and swelling were the most common damages quantified on the tegument. The in vitro study reproduced results described for other immature flukes incubated with artemisinin derivatives. Excysted metacercariae of E. paraensei constitute a good model to study in vitro drug effects.

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