Mentha villosa Hubs., M. x piperita and their bioactives against gastrointestinal nematodes of ruminants and the potential as drug enhancers.

Parasitic diseases caused by gastrointestinal nematodes (GIN) are responsible for a major impact on ruminant welfare. Although the available anthelmintics have a safe margin of toxicity to the animals, their indiscriminate use has increased the selection of resistant parasite populations. In this scenario, essential oils (EO) stand out as a promising ecofriendly therapeutic alternative against GIN. The objective of this work was to determine the effect of the EO of Mentha villosa Hubs (MVEO) collected in 2017 and 2018, M. x piperita (MPEO) and their main components, carvone and limonene, against the third stage larvae (L3) of Haemonchus spp. and Trichostrongylus spp. The solutions, including in nanoemulsion preparations, were tested in a range of concentrations using the larval migration inhibition test (LMIT). The EO and carvone were also tested in combination with nitroxynil (NTX) to determine their effect as drug enhancers (additive or synergy). MVEO/2017, MVEO/2018, MPEO and carvone showed 70.6 (73.4 mg/mL), 86.3 (74.9 mL/mL), 95.5 (143.6 mg/mL), and 88.2 % (38.3 mg/mL) efficacy against L3, respectively. Carvone alone had approximately a 3-fold higher efficacy when compared to its concentration in each EO: 68.8 % in MVEO/2017 and 83.9 % in MVEO/2018. Limonene did not show any significant effect on inhibiting L3 migration. The combination of MPEO and NTX, and carvone and NTX showed a statistically significantly (P <  0.05) synergic and additive effect, respectively, when compared to the isolated treatment. The nanoemulsion of MVEO/2017 at 0.367 mg/mL, inhibited L3 migration by 83.1 %, demonstrating to be highly effective (concentration ratio of 1:0.004), when compared to the MVEO/2017 (70.6 % at 73.4 mg/mL) extraction. The in vitro data from the combination of MPEO or carvone plus NTX suggest that these products can be considered for in vivo experiments against the most important GIN of ruminants as drug enhancers, possibly reducing the final concentration of NTX`. The efficacy of carvone was higher (EC50 = 1.96 mg/mL) than its expected efficacy, based on its concentrations on both EO. Therefore, this component does not need the entire EO composition to exert its L3 motility action. The remarkable efficacy demonstrated by the MVEO/2017/nanoemulsion (EC50 = 0.10 mg/mL), supports its potential to be a candidate to the next-generation therapy to alleviate clinical parasite infections and combat GIN resistant populations.

Sheep polyclonal antibody to map Haemonchus contortus mimotopes using phage display library / Anticorpo policlonal de ovinos para mapear mimetopos de Haemonchus contortus usando a biblioteca de Phage display

Abstract The aim of this study was to evaluate phage display technology for mapping Haemonchus contortus mimotopes. We screened the PhD-7 Phage Display Peptide Library Kit with a sheep polyclonal antibody against H. contortus. After four rounds of selection, 50 phage peptide clones were selected by biopanning and sequenced. Two clones displaying peptide mimotopes of H. contortus proteins were chosen for sheep immunization: clone 6 - mimotope of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and clone 17 - mimotope of a disorganized muscle family member (Dim 1). Twelve sheep were allocated into 3 groups of 4 animals as follow: G1: control group; G2/GAPDH: immunized with clone 6; and G3/Dim1: immunized with clone 17. Four immunizations were performed at intervals of seven days (0, 7, 14, and 21 days). On day 28 post initial vaccination, all groups were orally challenged with 2500 H. contortus infective larvae. The mimotope peptides selected by phage display were recognized by IgG from sheep naturaly infected with H. contortus. The immunization protocol showed an increasein IgG anti-M13 phage titers, but no effect was observed in IgG-specific for the anti-mimotope peptides. This is the first report of successful use of a phage display library for the identification of mimotopes of H. contortus proteins.

Resumo O objetivo deste estudo foi avaliar a tecnologia de phage display no mapeamento de mimetopos de Haemonchus contortus. Anticorpo policlonal de ovinos anti-H. contortus foi usado para seleção a partir da biblioteca PhD-7 Phage Display Peptide Library Kit (New England BioLabs). Após quatro rodadas, 50 clones de fagos expressando peptídeos foram selecionados e sequenciados. Dois clones que exibiram mimetopos de H. contortus foram escolhidos para imunização de ovinos: clone 6 - mimetopo de gliceraldeído-3-fosfato desidrogenase (GAPDH) e clone 17 - mimetopo da família do músculo desorganizado (Dim 1). Doze ovinos foram alocados em 3 grupos de 4 animais, da seguinte forma: G1: grupo controle, G2/GAPDH: imunizado com o clone 6 e G3/Dim1: imunizado com o clone 17. Quatro imunizações foram realizadas (0, 7, 14 e 21 dias). No dia 28 após a primeira imunização, todos os grupos foram desafiados oralmente com 2500 larvas infectantes de H. contortus . Os peptídeos mimetopos selecionados foram reconhecidos por IgG de ovinos naturalmente infectados por H. contortus. O ensaio de imunização revelou um aument dos títulos de IgG anti-fago M13, mas não ocorreu aumento de IgG anti-peptídeos mimetopos. Este é o primeiro relato de uso bem sucedido da biblioteca de Phage display para a identificação de mimetopos de H. contortus.

Sheep polyclonal antibody to map Haemonchus contortus mimotopes using phage display library.

The aim of this study was to evaluate phage display technology for mapping Haemonchus contortus mimotopes. We screened the PhD-7 Phage Display Peptide Library Kit with a sheep polyclonal antibody against H. contortus. After four rounds of selection, 50 phage peptide clones were selected by biopanning and sequenced. Two clones displaying peptide mimotopes of H. contortus proteins were chosen for sheep immunization: clone 6 - mimotope of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and clone 17 - mimotope of a disorganized muscle family member (Dim 1). Twelve sheep were allocated into 3 groups of 4 animals as follow: G1: control group; G2/GAPDH: immunized with clone 6; and G3/Dim1: immunized with clone 17. Four immunizations were performed at intervals of seven days (0, 7, 14, and 21 days). On day 28 post initial vaccination, all groups were orally challenged with 2500 H. contortus infective larvae. The mimotope peptides selected by phage display were recognized by IgG from sheep naturaly infected with H. contortus. The immunization protocol showed an increasein IgG anti-M13 phage titers, but no effect was observed in IgG-specific for the anti-mimotope peptides. This is the first report of successful use of a phage display library for the identification of mimotopes of H. contortus proteins.

In vitro efficacy of Duddingtonia flagrans against nematodes of sheep based on in vivo calculations / Eficácia in vitro de Duddingtonia flagrans contra nematoides de ovinos com base em cálculo in vivo

Abstract Duddingtonia flagrans has been tested as an alternative parasite control, but data from in vitro experiments based on in vivo calculations describing nematophagous fungi predation in nematodes are restricted. The objective of this work was to determine the efficacy of D. flagrans against sheep nematode larvae in vitro using in vivo calculations. Fecal samples were introduced to fungi in different concentrations: 0.0/control; 0.05; 0.1; 0.2; 0.4; 0.8; 1.6; 3.2; and 6.4 g corresponding, respectively, to 583.000; 1.166.000; 2.332.000; 4.664.000; 9.328.000; 18.656.000; 37.312.000 and 74.624.000 chlamydospores/kg of body weight. The material was incubated for 14 days, before the larvae recovery (Assay 1). Assay 2 was carried out with the doses of 0.00625; 0.0125; and 0.025 g. The results showed a negative correlation between fungal concentrations and larval numbers for both assays. The fungus demonstrated an efficacy above 89% in both assays. Thus, we consider that the data from in vitro studies based on in vivo calculations may optimize the fungi quantities for field experiments.

Resumo Duddingtonia flagrans tem sido testado como uma alternativa no controle de parasitos, entretanto, trabalhos in vitro da predação de nematoides por fungos nematófagos correlacionados com cálculos baseados para testes in vivo são restritos. O objetivo deste trabalho foi determinar a eficácia in vitro de D. flagrans contra larvas de nematoides de ovinos tendo como base cálculos in vivo. Amostras fecais receberam a adição do fungo em diferentes concentrações: 0.0/controle; 0,05; 0,1; 0,2; 0,4; 0,8; 1,6; 3,2 e 6,4 gramas correspondendo, respectivamente, às seguintes dosagens: 583.000; 1.166.000; 2.332.000; 4.664.000; 9.328.000; 18.656.000; 37.312.000 e 74.624.000 clamidósporos/Kg de peso vivo animal. O material foi incubado por 14 dias, para recuperação das larvas (Ensaio 1). O Ensaio 2 foi realizado com concentrações de 0,00625; 0,0125 e 0,025 g. Foi observada correlação negativa entre a concentração fúngica e o número de larvas, nos dois ensaios. O fungo demonstrou eficácia acima de 89% em ambos os ensaios. A partir destes dados, acreditamos que ensaios in vitro baseados em cálculos in vivo podem aprimorar as dosagens para a realização de experimentos a campo.

Occurrence of gastrointestinal parasites in wild animals in State of Paraná, Brazil

ABSTRACT The objective of this study was to determine the prevalence and diversity of gastrointestinal parasites in fecal samples from wild birds and mammals from the State of Paraná. In total, 220 stool samples were sent to Parasitic Diseases Laboratory of the Federal University of Paraná during 13 months (Jan/2013-Jan/2014). A total of 52.7% (116/220) of the animals were positive for cysts, oocysts, eggs and/or trophozoites. In birds, the positivity rate was 37.9% (25/66) and mammals was 59.1% (91/154). Strongyloidea superfamily eggs were observed in 37.3% (82/220) of the samples, Eimeria spp. in 10% (22/220), and Trichuris spp. in 4.5% (10/220). The most frequent mammal species were llamas (Lama glama), and dromedaries (Camelus bactrianus) with infection rate of 70.1% (54/77) and 60.8% (14/23), respectively. In other hand, cockatiels (Nymphicus hollandicus) and ring necks (Psittacula krameri), were the most researched birds, with infection rate of 20% (40/50) and 100% (6/6), respectively. A high prevalence of gastrointestinal parasites was observed in most of wildlife animals. Further investigations should be conducted focusing on parasite control strategies and the conservation measurements for harmonizing the human-animal interaction on the long-term, reducing associated health risks.

Occurrence of gastrointestinal parasites in wild animals in State of Paraná, Brazil.

The objective of this study was to determine the prevalence and diversity of gastrointestinal parasites in fecal samples from wild birds and mammals from the State of Paraná. In total, 220 stool samples were sent to Parasitic Diseases Laboratory of the Federal University of Paraná during 13 months (Jan/2013-Jan/2014). A total of 52.7% (116/220) of the animals were positive for cysts, oocysts, eggs and/or trophozoites. In birds, the positivity rate was 37.9% (25/66) and mammals was 59.1% (91/154). Strongyloidea superfamily eggs were observed in 37.3% (82/220) of the samples, Eimeria spp. in 10% (22/220), and Trichuris spp. in 4.5% (10/220). The most frequent mammal species were llamas (Lama glama), and dromedaries (Camelus bactrianus) with infection rate of 70.1% (54/77) and 60.8% (14/23), respectively. In other hand, cockatiels (Nymphicus hollandicus) and ring necks (Psittacula krameri), were the most researched birds, with infection rate of 20% (40/50) and 100% (6/6), respectively. A high prevalence of gastrointestinal parasites was observed in most of wildlife animals. Further investigations should be conducted focusing on parasite control strategies and the conservation measurements for harmonizing the human-animal interaction on the long-term, reducing associated health risks.

In vitro efficacy of Duddingtonia flagrans against nematodes of sheep based on in vivo calculations.

Duddingtonia flagrans has been tested as an alternative parasite control, but data from in vitro experiments based on in vivo calculations describing nematophagous fungi predation in nematodes are restricted. The objective of this work was to determine the efficacy of D. flagrans against sheep nematode larvae in vitro using in vivo calculations. Fecal samples were introduced to fungi in different concentrations: 0.0/control; 0.05; 0.1; 0.2; 0.4; 0.8; 1.6; 3.2; and 6.4 g corresponding, respectively, to 583.000; 1.166.000; 2.332.000; 4.664.000; 9.328.000; 18.656.000; 37.312.000 and 74.624.000 chlamydospores/kg of body weight. The material was incubated for 14 days, before the larvae recovery (Assay 1). Assay 2 was carried out with the doses of 0.00625; 0.0125; and 0.025 g. The results showed a negative correlation between fungal concentrations and larval numbers for both assays. The fungus demonstrated an efficacy above 89% in both assays. Thus, we consider that the data from in vitro studies based on in vivo calculations may optimize the fungi quantities for field experiments.

Comparison of McMaster and Mini-FLOTAC fecal egg counting techniques in cattle and horses.

The aim of this study was to compare two fecal egg count (FEC) techniques; McMaster (McM) and Mini-FLOTAC (mF), for the detection of cattle and horse gastrointestinal nematode eggs, in different locations. Experiment 1: feces were collected from 16 cattle and FEC was performed individually, using mF with the sensitivity of 5 eggs per gram of feces (EPG) and McM with a sensitivity of 50 EPG at Empresa de Pesquisa Agropecuária de Minas Gerais - EPAMIG and the Laboratory of Parasitic Diseases of the University of Parana - LDP/UFPR. Experiment 2: Fecal samples from 30 horses were analyzed with mF (sensitivity of 5 EPG) and McM (sensitivity of 25 EPG) at the University of Mato Grosso do Sul - UFMS and LPD/UFPR. Experiment 3: feces were collected from 14 foals and FEC was performed using mF (sensitivity of 5 EPG); and McM (sensitivity of 25 EPG) only at the LPD/UFPR. For cattle, the average FEC of mF was 962 at LPD; and 1248 at EPAMIG; for McM it was 1393 at LPD and 1563 at EPAMIG. For horses, the FEC average using the mF was 650 at LPD and 469 at UFMS; and for McM it was 677 at LPD and 554 at UFMS. For foals, the average FEC for mF was 404 and 436 for McM. In all experiments, the standard deviation and the coefficient of variation values were significantly lower for mF. Therefore, it is recommended the use of the Mini-FLOTAC technique, which is a method with less variability and higher accuracy, particularly for animals with low FEC.

Gene silencing of Dim-1, a member of the disorganized muscle family, in Haemonchus contortus.

RNA interference has been widely used in parasites. In Haemonchus contortus, reproducible silencing has been reported; however, in this species an altered phenotype has been observed for only a few genes silenced by RNA interference. The aim of this work was to evaluate the effect of silencing Dim-1, a member of the disorganized muscle family on third stage larvae (L3) of H. contortus. Effective silencing of Dim-1 in L3 led to reduced L3 migration and slowed larval development from L3 to early L4. To our knowledge, this is the first report to demonstrate Dim-1 silencing affecting phenotypic characteristics of this parasite.

In vitro efficacy of Duddingtonia flagrans against nematodes of sheep based on in vivo calculations

Abstract Duddingtonia flagrans has been tested as an alternative parasite control, but data from in vitro experiments based on in vivo calculations describing nematophagous fungi predation in nematodes are restricted. The objective of this work was to determine the efficacy of D. flagrans against sheep nematode larvae in vitro using in vivo calculations. Fecal samples were introduced to fungi in different concentrations: 0.0/control; 0.05; 0.1; 0.2; 0.4; 0.8; 1.6; 3.2; and 6.4 g corresponding, respectively, to 583.000; 1.166.000; 2.332.000; 4.664.000; 9.328.000; 18.656.000; 37.312.000 and 74.624.000 chlamydospores/kg of body weight. The material was incubated for 14 days, before the larvae recovery (Assay 1). Assay 2 was carried out with the doses of 0.00625; 0.0125; and 0.025 g. The results showed a negative correlation between fungal concentrations and larval numbers for both assays. The fungus demonstrated an efficacy above 89% in both assays. Thus, we consider that the data from in vitro studies based on in vivo calculations may optimize the fungi quantities for field experiments.

Resumo Duddingtonia flagrans tem sido testado como uma alternativa no controle de parasitos, entretanto, trabalhos in vitro da predação de nematoides por fungos nematófagos correlacionados com cálculos baseados para testes in vivo são restritos. O objetivo deste trabalho foi determinar a eficácia in vitro de D. flagrans contra larvas de nematoides de ovinos tendo como base cálculos in vivo. Amostras fecais receberam a adição do fungo em diferentes concentrações: 0.0/controle; 0,05; 0,1; 0,2; 0,4; 0,8; 1,6; 3,2 e 6,4 gramas correspondendo, respectivamente, às seguintes dosagens: 583.000; 1.166.000; 2.332.000; 4.664.000; 9.328.000; 18.656.000; 37.312.000 e 74.624.000 clamidósporos/Kg de peso vivo animal. O material foi incubado por 14 dias, para recuperação das larvas (Ensaio 1). O Ensaio 2 foi realizado com concentrações de 0,00625; 0,0125 e 0,025 g. Foi observada correlação negativa entre a concentração fúngica e o número de larvas, nos dois ensaios. O fungo demonstrou eficácia acima de 89% em ambos os ensaios. A partir destes dados, acreditamos que ensaios in vitro baseados em cálculos in vivo podem aprimorar as dosagens para a realização de experimentos a campo.

Duddingtonia flagrans in the control of gastrointestinal nematodes of horses.

Horses can harbor a large amount of parasites that may cause serious clinical signs even death. The aim of this study was to evaluate the predatory activity of the fungus Duddingtonia flagrans against infective larvae (L3) of gastrointestinal nematodes of horses in fecal culture. The experimental design was completely randomized with three treated groups (G1, G2 and G3) and one control (CG), using eight animals/group. The treated animals received G1: 1.5 × 10(5); G2: 3 × 10(5) and G3: 6 × 10(5) chlamydospores of D. flagrans/kg body weight during 21 days. The fungi preparation was given at every other three-day interval. Faecal samples were collected during 30 days, on the same interval, to perform the fecal egg counts (EPG) and fecal culture for each horse. All groups demonstrated similar results for the EPG (P > 0.05) counts. D. flagrans significantly reduced (P < 0.05) the number of infective larvae after 72 h-interval between treatments. The G2 and G3 promoted higher results (P < 0.05) of L3 reduction compared to the CG. The biological control with the predacious fungi D. flagrans is still a promising free-living parasite regulator alternative to be use in livestock.

Efeito anticoccidiano de extrato hidroalcoólico de Artemisia annua em camas de aves contaminadas com Eimeria sp / Anticoccidial effect of Artemisia annua hydroalcoholic extract in poultry beds contaminated with Eimeria sp

O objetivo deste estudo foi avaliar a eficácia do extrato hidroalcoólico de Artemisia annua frente a oocistos de Eimeria sp. em camas contaminadas. O extrato foi produzido com 7 dias de percolação a 4°C, sendo posteriormente realizada a marcha fitoquímica; dosagem de fenóis totais, quantificação de artemisinina, ensaio antioxidante e teste de toxicidade. Para testar a atividade anticoccidiana, camas de aves compostas de cepilho de árvores foram contaminadas com 5000 oocistos. Foram formados quatro tratamentos, em triplicata, nos quais foram usadas diferentes concentrações, sendo G1: 12mg/mL, G2: 8mg/mL, G3: 4mg/mL e C-: água. Após a contaminação, foram aspergidos, 800 mL dos extratos nas diferentes concentrações sobre as camas e coletadas, em triplicatas, 10 cm2 de cada local, aleatoriamente, nos tempos: 0, 3, 6, 24, 48, e 72 horas após a aplicação. Nas análises fitoquímicas, foram evidenciados diversos compostos com propriedades antiparasitárias, como flavonoides e taninos. O fitoterápico continha 59,409±1,47μg/dL de artemisinina. O produto na concentração de 12mg.mL-1 apresentou eficácia entre 45,5 e 42,1%. Os resultados dos testes bioquímicos, juntamente com os encontrados no teste anticoccidiano, evidenciaram que o extrato produzido possui alto potencial para combater Eimeria sp.

The aim of this study was to determine the efficacy of hydroalcoholic extract of Artemisia annua against oocysts of Eimeria sp. in contaminated poultry beds. The extract was produced after 7 days of storage at 4°C, which was used to perform the phytochemical screening; the artemisinin measurement; the total phenolic; antioxidant testing and toxicity test. To test the anticoccidial activity, the birds space composed of shaver trees, were contaminated with 5000 oocysts. Four treatment were formed, in triplicate, were used in different concentrations as G1: 12mg/mL, G2:= 9mg/mL, G3: 6mg/mL, and C-: water. After contamination 800 mL of the herbal at different concentrations were sprayed on the bed and collected, in triplicate, 10 cm2 each site, randomly, at times: 0, 3, 6, 24, 48, and 72 hours after application. In phytochemical analysis, were shown compounds with antiparasitic properties, such as flavonoids and tannins. The herbal contained 59.409±1.47mg/dL artemisinin. The product at a concentration of 12mg.mL-1 showed efficacy from 44.25 to 40.71%. The results of biochemical tests, with the in vitro test showed that the extract has produced high potential for combating Eimeria sp.

Efeito anticoccidiano de extrato hidroalcoólico de Artemisia annua em camas de aves contaminadas com Eimeria sp / Anticoccidial effect of Artemisia annua hydroalcoholic extract in poultry beds contaminated with Eimeria sp

O objetivo deste estudo foi avaliar a eficácia do extrato hidroalcoólico de Artemisia annua frente a oocistos de Eimeria sp. em camas contaminadas. O extrato foi produzido com 7 dias de percolação a 4°C, sendo posteriormente realizada a marcha fitoquímica; dosagem de fenóis totais, quantificação de artemisinina, ensaio antioxidante e teste de toxicidade. Para testar a atividade anticoccidiana, camas de aves compostas de cepilho de árvores foram contaminadas com 5000 oocistos. Foram formados quatro tratamentos, em triplicata, nos quais foram usadas diferentes concentrações, sendo G1: 12mg/mL, G2: 8mg/mL, G3: 4mg/mL e C-: água. Após a contaminação, foram aspergidos, 800 mL dos extratos nas diferentes concentrações sobre as camas e coletadas, em triplicatas, 10 cm2 de cada local, aleatoriamente, nos tempos: 0, 3, 6, 24, 48, e 72 horas após a aplicação. Nas análises fitoquímicas, foram evidenciados diversos compostos com propriedades antiparasitárias, como flavonoides e taninos. O fitoterápico continha 59,409±1,47μg/dL de artemisinina. O produto na concentração de 12mg.mL-1 apresentou eficácia entre 45,5 e 42,1%. Os resultados dos testes bioquímicos, juntamente com os encontrados no teste anticoccidiano, evidenciaram que o extrato produzido possui alto potencial para combater Eimeria sp...

The aim of this study was to determine the efficacy of hydroalcoholic extract of Artemisia annua against oocysts of Eimeria sp. in contaminated poultry beds. The extract was produced after 7 days of storage at 4°C, which was used to perform the phytochemical screening; the artemisinin measurement; the total phenolic; antioxidant testing and toxicity test. To test the anticoccidial activity, the birds space composed of shaver trees, were contaminated with 5000 oocysts. Four treatment were formed, in triplicate, were used in different concentrations as G1: 12mg/mL, G2:= 9mg/mL, G3: 6mg/mL, and C-: water. After contamination 800 mL of the herbal at different concentrations were sprayed on the bed and collected, in triplicate, 10 cm2 each site, randomly, at times: 0, 3, 6, 24, 48, and 72 hours after application. In phytochemical analysis, were shown compounds with antiparasitic properties, such as flavonoids and tannins. The herbal contained 59.409±1.47mg/dL artemisinin. The product at a concentration of 12mg.mL-1 showed efficacy from 44.25 to 40.71%. The results of biochemical tests, with the in vitro test showed that the extract has produced high potential for combating Eimeria sp...

Método FAMACHA para detectar anemia clínica causada por Haemonchus contortus em cordeiros lactentes e ovelhas em lactação / FAMACHA method for detecting clinical anemia caused by Haemonchus contortus in suckling lambs and lactating ewes

O controle das endoparasitoses gastrintestinais em pequenos ruminantes tem sido tradicionalmente realizado por meio do uso indiscriminado dos anti-helmínticos, porém essa prática tem resultado em grande pressão de seleção de parasitas resistentes. Métodos seletivos (ex: FAMACHA/FMC e contagem de ovos nas fezes/OPG) e que apresentem boa sensibilidade para identificar os animais que necessitam receber o tratamento químico são importantes alternativas aos métodos tradicionais. O objetivo desse trabalho foi determinar a sensibilidade e a especificidade do método FMC, usado para detectar anemia causada por Haemonchus contortus em ovinos da raça Suffolk, em duas categorias de alta suscetibilidade; cordeiros lactentes e ovelhas durante a fase de lactação...

The control of gastrointestinal parasites in small ruminants has traditionally involved the indiscriminate use of anthelmintic drugs that resulted in a high parasite selection pressure for anthelmintic resistance. Selective methods (ex.: FAMACHA/FMC and faecal egg count/EPG), which have good sensitivity to identify animals that need drug treatment, are important alternative methods for parasite control. The objective of this study was to determine the sensitivity and specificity of the FMC method to detect clinical anemia caused by Haemonchus contortus in two highly susceptible categories of Suffolk sheep; suckling lambs and lactating ewes...

Evaluation of resistance in a selected field strain of Haemonchus contortus to ivermectin and moxidectin using the Larval Migration on Agar Test / Avaliação da resistência em um isolado de campo selecionado de Haemonchus contortus à ivermectina e moxidectina usando o Teste de Migração de Larvas em Ágar

Haemonchus contortus is one of the most common and economically significant causes of disease in small ruminants worldwide, and the control programs of parasitic nematodes - including H. contortus - rely mostly on the use of anthelmintic drugs. The consequence of the use of this, as the sole sanitary strategy to avoid parasite infections, was the reduction of the efficacy of all chemotherapeutic products with a heavy selection for resistance. The widespread of anthelmintic resistance and the difficulty of its early diagnosis has been a major concern for the sustainable parasite management on farms. The objective of this research was to determine and compare the ivermectin (IVM) and moxidectin (MOX) effect in a selected field strain of H. contortus with a known resistance status, using the in vitro larval migration on agar test (LMAT). Third stage larvae of the selected isolate were obtained from faecal cultures of experimentally infected sheep and incubated in eleven increasing diluted concentrations of IVM and MOX (6, 12, 24, 48, 96, 192, 384, 768, 1536, 3072 and 6144µg/mL). The dose-response sigmoidal curves were obtained using the R² value of >0.90 and the lethal concentration (LC50) dose for the tested anthelmintic drugs using a four-parameter logistic model. The LC50 value for MOX was significantly lower than IVM (1.253µg/mL and 91.06µg/mL), identifying the H. contortus isolate as considerably less susceptible to IVM compared to MOX. Furthermore, the LMAT showed a high consistency (p<0.0001) and provided to be a useful diagnostic tool for monitoring the resistance status of IVM and MOX in H. contortus field isolate, as well as it may be used for official routine drug monitoring programs under the Ministry of Agriculture (MAPA) guidance.

Haemonchus contortus é uma das causas mais comuns e economicamente significativas de doença em produções de pequenos ruminantes em todo o mundo, e os programas de controle de parasitas nematoides - incluindo H. contortus - baseiam-se principalmente no uso de drogas anti-helmínticas. A consequência da utilização desses compostos, como sendo a única estratégia sanitária para evitar infecções por parasitas, tem sido a redução da eficácia de todos os produtos quimioterápicos, selecionando fortemente para resistência. O desenvolvimento generalizado da resistência anti-helmíntica e a dificuldade de seu diagnóstico precoce têm sido uma grande preocupação para o manejo sustentável de parasitas no campo. O objetivo desta pesquisa foi determinar e comparar o efeito da ivermectina (IVM) e da moxidectina (MOX) em um isolado de campo selecionado de H. contortus com um estado de resistência conhecido, utilizando o teste in vitro de migração de larvas em ágar (LMTA). Larvas de terceiro estágio de um isolado de H. contortus selecionado foram obtidas a partir de culturas de fezes de ovinos infectados experimentalmente e incubadas em onze concentrações diluídas crescentes de IVM e MOX (6, 12, 24, 48, 96, 192, 384, 768, 1536, 3072 e 6144µg/mL). As curvas sigmoides de dose-resposta foram obtidas utilizando o valor de R² >0,90 e a dose de concentração letal (CL50) para as drogas anti-helmínticas testadas, utilizando um modelo logístico de quatro parâmetros. O valor de CL50 para MOX foi significativamente menor do que para IVM (1,253µg/mL e 91,06µg/mL), identificando o isolado de H. contortus como consideravelmente menos suscetível à IVM em comparação à MOX. Além disso, o LMTA mostrou uma alta consistência (p<0,0001) e pode ser uma ferramenta útil de diagnóstico para monitorar o status de resistência de IVM e MOX em isolado de campo de H. contortus, assim como ser utilizado de forma oficial e em rotina para programas de monitoramento das drogas sob a demanda do Ministério da Agricultura (MAPA).

Partial selective treatment of Rhipicephalus microplus and breed resistance variation in beef cows in Rio Grande do Sul, Brazil.

Rhipicephalus microplus infestation causes heavy losses in cattle. The majority of farmers control the cattle tick by using long-acting drugs throughout the year, which evidently increases selection pressure for drug resistance. Partial selective treatment (PST) may be used to directly reduce selection pressure by treating only the portion of the herd that is most infested. The objective of this work was to determine the effect of PST in two herds of beef cows and the effect of breed tolerance. The Santiago (n=306 animals) and the São Francisco de Assis (n=204 animals) herds were composed of up to eight pure breeds of Bos taurus and Bos indicus and their crosses and were evaluated during the first and second (8 months) tick generations. Ticks larger than 4.5mm were counted on one side of the animal, and the animals were treated when numbers exceeded 20 ticks per host. On both farms, the tick-tolerant breeds with a high proportion of B. indicus (Braford, Brangus, and Nellore) consistently carried fewer ticks than Charolais. The economical evaluation showed an average profit margin in the use of PST of 674.25% and 1394.5% on the Santiago and São Francisco de Assis farms, respectively, in comparison to all-herd treatment. These results indicate that PST is a reliable tool for health management and may be used as a standard protocol for tick control with significant economic benefits on farms. In addition, the maintenance of the highest possible tolerant/resilient breeds is also an essential factor that should be considered in today's more sustainable animal production systems.

Acaricide activity of leaves extracts of Sambucus australis Schltdl. (Caprifoliaceae) at 2 percent on engorged females of Rhipicephalus (Boophilus) microplus / Atividade acaricida de extratos de folhas de Sambucus australis Schltdl (Caprifoliaceae) a 2 por cento sobre teleóginas de Rhipicephalus (Boophilus) microplus

Parasite infections caused by Rhipicephalus (Boophilus) microplus are responsible for the most of economic losses in producing-cattle countries in tropical and subtropical areas. Indiscriminate uses of chemical acaricides have contributed with the appearance of tick resistance to many drugs available in the marketplace, and it is a serious problem in the tick control. Flowers of Sambucus australis (South America), called "sabugueiro-from-Brazil", are used on infusions or decoctions forms in the folk medicine with diuretic, antipyretic, anti-inflammatory, and laxative purpose, also employed for treating respiratory diseases in human. The main goal of this study was to evaluate in vitro potential of S. australis leaves extracts as an acaricide agent. Ethanol extract at 70 percent has been dehydrated and a fraction was suspended in 70 percent ethanol or in distillated water at final concentration of 2 percent (0.2mg-1). Using the immersion test of engorged females the efficiency results were obtained in 34 percent and 66 percent with the leaves extract diluted in water and 70 percent ethanol, respectively. This study is the first report on acaricidal activity of S. australis against cattle tick. Further studies to determine the active metabolites in different stages of S. australis could aid to identify suitable extracts to be tested in the R. (B.) microplus control.

A infestação por Rhipicephalus (Boophilus) microplus é responsável por grandes perdas econômicas em países produtores de bovinos nas áreas tropicais e subtropicais. Usos indiscriminados de acaricidas químicos têm contribuído para o aparecimento da resistência dos ixodídeos a múltiplas drogas, representando um sério problema no controle de carrapatos. As flores da planta Sambucus australis, conhecidas pelo nome popular de sabugueiro do Brasil, são usadas, na forma de infusão ou emplastros, na medicina popular como diurético, antipirético, antiinflamatório e laxativo; também empregado no tratamento de doenças respiratórias em humanos. O objetivo principal deste trabalho foi avaliar o potencial in vitro de extratos de folhas de S. australis, como agente carrapaticida. O extrato etanólico a 70 por cento foi dessecado e retomado parte em etanol a 70 por cento ou em água destilada, numa concentração final de 2 por cento (0,2mg-1). No biocarrapaticidograma obteve-se resultados de eficácia entre 34 por cento e 66 por cento para o extrato de folhas diluído em água e etanol a 70 por cento, respectivamente. Esse estudo é o primeiro relato de ação carrapaticida de S. australis. Estudos adicionais para determinar os metabólitos ativos, em diferentes estágios de S. australis, fornecerão subsídios para identificar os extratos apropriados a serem testados no controle do R. (B.) microplus.

Atividade acaricida in vitro de Glechon spathulata Benth. sobre teleóginas de Rhipicephalus (Boophilus) microplus / Acaricide activity in vitro of Glechon spathulata Benth. On engorged females of Rhipcephalus (Boophilus) microplus

O objetivo deste estudo foi avaliar a atividade acaricida in vitro de extratos vegetais de Glechon spathulata (manjeroninha do campo) sobre teleóginas de Rhipicephalus (Boophilus) microplus. Fêmeas ingurgitadas desse carrapato foram coletadas de bovinos mestiços de uma propriedade do município de São Pedro do Sul, Rio Grande do Sul, Brasil. A experimentação in vitro consistiu na realização de testes de imersão de fêmeas ingurgitadas. O extrato bruto seco e a fração clorofórmica utilizados nos testes in vitro foram preparados na concentração de 2 por cento e diluídos em água e etanol, sendo todos os tratamentos realizados em triplicata. O extrato bruto seco de G. spathulata retomado em etanol a 70 por cento apresentou uma eficácia média de 99 por cento, enquanto que quando o mesmo extrato foi diluído em água, demonstrou eficácia média de 72 por cento. Observou-se que no controle etanol a 70 por cento houve diferença na eclodibilidade dos ovos, porém não interferiu no peso da postura. A fração clorofórmica de G. spathulata não apresentou eficácia no ensaio efetuado. Os resultados obtidos nesse estudo demonstraram que o extrato hidroetanólico a 70 por cento de G. spathulata apresenta eficácia acaricida in vitro. Este estudo é o primeiro relato de ação acaricida de G. spathulata.

The aim of this study was to evaluate the in vitro acaricide efficiency of Glechon spathulata extracts on engorged females of Rhipcephalus (Boophilus) microplus. Engorged ticks were collected from crossbred cattle from one farm in São Pedro do Sul municipality, Rio Grande do Sul, Brazil. in vitro experiments consisted on the immersion tests of engorged females in solutions of G. spathulata, where treatments were made at triplicate with fractions and extracts at 2 percent concentration, diluted in water and ethanol. Crude dry extract of G. spathulata suspended in 70 percent ethanol presented an efficiency average of 99 percent, while the same extract diluted in water demonstrated efficiency average of 72 percent. It was observed in 70 percent ethanol control difference in egg eclodibility; however there was no interference in egg weight. The chloroform fraction of G. spathulata showed no efficacy in the test. The results obtained suggest that the hydroethanolic extract at 70 percent of G. spathulata has significant acaricide activity in vitro. This research is the first communication of acaricidal effect of. G. spathulata.

Ação acaricida in vitro de Tropaeolum majus sob teleóginas de Rhipicephalus (Boophilus) microplus / in vitro acaricidal effect of Tropaeolum majus on the engorged female of Rhipicephalus (Boophilus) microplus

O objetivo desse estudo foi o de avaliar a eficácia acaricida in vitro do extrato aquoso e etanólico a 70 por cento da Tropaeolum majus em teleóginas do Rhipicephalus (Boophilus) microplus. Extrato das partes aéreas (caule e folhas) e de flores foram testados na concentração final de 2 por cento. A coleta de teleóginas de Rhipicephalus (Boophilus) microplus foi realizada em propriedades do município de São Francisco de Assis e São Pedro do Sul, Rio Grande do Sul, Brasil. O extrato das partes aéreas em etanol a 70 por cento alcançou eficácia de 99,1 por cento. Esse estudo é o primeiro relato de ação carrapaticida de Tropaeolum majus.

The aim of this study was to evaluate the in vitro acaricide effect of aqueous and ethanolic extracts of Tropaeolum majus on Rhipicephalus (Boophilus) microplus engorged females. Extracts from the aerial parts (stems and leaves) and from the flowers were tested at a final concentration of 2 percent. The Rhipicephalus (Boophilus) microplus engorged females were collected in farms in two municipalities (São Francisco de Assis and São Pedro do Sul), in Rio Grande do Sul state, Brazil. An efficacy of 99,1 percent was found using extract of aerial parts in ethanol at 70 percent. This is the first report of acaricidal effect of Tropaeolum majus.