Effect of the Combination of Synthetic Anthelmintics with Carvacryl Acetate in Emulsions with and without a Sodium Alginate Matrix on Haemonchus contortus.

The present study aimed to evaluate the effect of nanoemulsions using combined synthetic anthelmintics, thiabendazole (TBZ), levamisole (LEV), and ivermectin (IVM), with carvacryl acetate (CA) against Haemonchus contortus, and also tested the presence and absence of alginate (ALG). The anthelmintic effect of the CA/TBZ nanoemulsion was evaluated in the egg hatch test (EHT). The effects of CA/IVM and CA/LEV nanoemulsions were evaluated in the larval development test (LDT). The emulsions CA/TBZ/ALG and CA/TBZ showed a multimodal profile, with most particles on the nanometric scale. The encapsulation efficiency in CA/TBZ/ALG was 80.25%, and that in CA/LEV/ALG was 89.73%. In the EHT, CA/TBZ and CA/TBZ/ALG showed mean combination indices (CIs) of 0.55 and 0.36, respectively, demonstrating synergism in both. In LDT, CA/IVM had an average CI of 0.75, and CA/LEV and CA/LEV/ALG showed CI values of 0.4 and 0.93, respectively. It was concluded that CA/TBZ showed a synergistic interaction, and CA/TBZ/ALG showed an enhanced effect. In addition, the matrix brought stability to the product, encouraging its improvement to obtain higher efficacy.

In vitro anthelmintic activity of Lippia alba essential oil chemotypes against Haemonchus contortus.

The aim of the present study was to evaluate the effects of the essential oils of Lippia alba chemotypes carvone and citral on H. contortus. Chemical characterization was performed by means of gas chromatography coupled to mass spectrometry (GC/MS). The anthelmintic effects of the essential oils were assessed through the egg hatch test (EHT) and the adult worm motility test (AWMT) using a multidrug-resistant H. contortus Kokstad isolate. Confocal laser scanning microscopy (CLSM) of eggs and adults of H. contortus and scanning electron microscopy (SEM) of adults were performed after treatment with oils for qualitative observations of their effects. The carvone chemotype of L. alba (LaCV) presented 70% carvone, and the citral chemotype of L. alba (LaCT) presented 29.4% geranial and 20.4% neral, respectively. In the EHT, the EC50 values of LaCV and LaCT were 0.2 and 0.3 mg/mL, respectively. In AWMT, after 12 h of exposure to 2 mg/mL LaCV and 2 mg/mL LaCT, 100% of adult nematodes were immobile. CLSM showed changes in larval motility inside the egg caused by LaCV, while LaCT promoted changes in larval formation. In adults exposed to both chemotypes, alterations in the anterior portion of the oesophagus were observed. In SEM, morphological changes were observed in the buccal capsule and in the medial portion of H. contortus adults. It is concluded that the two essential oils of L. alba, the chemotypes carvone and citral, caused morphological changes and inhibited the hatching of eggs and the motility of adult H. contortus nematodes.

In vitro anthelmintic activity of an R-carvone nanoemulsions towards multiresistant Haemonchus contortus.

This work aimed to evaluate the in vitro anthelmintic effect of carvone nanoemulsions on Haemonchus contortus. Three R-carvone nanoemulsions were prepared: uncoated R-carvone nanoemulsions homogenized in a sonicator (UNAlg-son) and homogenized in an ultrahomogenizer (UNAlg-ultra) and sodium alginate-coated R-carvone (CNAlg-ultra). The physicochemical characterizations of the nanoemulsions were carried out. The anthelmintic activity was evaluated using egg hatch test (EHT), larval development test (LDT) and adult worm motility test (AWMT). Changes in cuticle induced in adult H. contortus were evaluated by scanning electron microscopy (SEM). The results were subjected to analysis of variance and compared using the Tukey test (P < 0.05). The effective concentration to inhibit 50% (EC50) of egg hatching and larval development was calculated. The particle sizes were 281.1 nm (UNAlg-son), 152.7 nm (UNAlg-ultra) and 557.8 nm (CNAlg-ultra), and the zeta potentials were −15 mV (UNAlg-son), −10.8 mV (UNAlg-ultra) and −24.2 mV (CNAlg-ultra). The encapsulation efficiency was 99.84 ± 0.01%. SEM of the nanoemulsions showed an increase in size. In EHT, the EC50 values of UNAlg-son, UNAlg-ultra and CNAlg-ultra were 0.19, 0.02 and 0.17 mg mL−1, respectively. In LDT, they were 0.29, 0.31 and 0.95 mg mL−1 for UNAlg-son, UNAlg-ultra and CNAlg-ultra, respectively. The adult motility inhibition was 100% after 12 h of exposure to UNAlg-ultra and CNAlg-ultra, while for UNAlg-son, it was 79.16%. SEM showed changes in the buccal capsule and cuticular damage. It was concluded that R-carvone nanoemulsions showed antiparasitic action demonstrating promise for the control of infections caused by gastrointestinal nematodes in small ruminants.

Phenotypic and genotypic approaches for detection of anthelmintic resistant sheep gastrointestinal nematodes from Brazilian northeast.

The aim of this study was to characterize the anthelmintic resistance (AR) of a sheep gastrointestinal nematode population, named Caucaia, from northeastern Brazil. Phenotypic tests performed were: egg hatch (EHT), larval development (LDT) and fecal egg count reduction (FECRT). Benzimidazoles (BZs) genotypic evaluation was by frequency of single nucleotide polymorphisms (SNPs) F200Y, F167Y and E198A, and for levamisole (LEV), by frequency of resistance alleles of Hco-acr-8 gene. The primers were designed specifically for Haemonchus contortus. Effective concentrations 50% (EC50) for BZs (EHT), and for macrocyclic lactones (MLs) and LEV (LDT) were 1.02 µg/mL, 1.81 ng/mL and 0.04 µg/mL, respectively. Resistance ratios for MLs and LEV were 0.91 and 3.07, respectively. FECRT efficacies of BZs, MLs, monepantel (MPTL) and LEV were 52.4; 87.0; 94.5 and 99.6%, respectively. qPCR for BZs demonstrated resistance allele frequencies of 0%, 26.24% and 69.08% for SNPs E198A, F200Y and F167Y, respectively. For LEV, 54.37% of resistance alleles were found. There was agreement between EHT, FECRT and qPCR for BZs, and agreement between LDT and qPCR for LEV. Thus, based on higher sensitivity of qPCR, and phenotypic evaluation, the Caucaia population was considered resistant to BZs, MLs, LEV and suspect for MPTL.

Ação inseticida in vitro do óleo essencial de Lippia gracilisSchauer sobre Damalinia Bovicola ovis (Schrank, 1781) / In vitro action of Lippia gracilis Schauer essential oil on Damalinia Bovicola ovis (Schrank, 1781)

O objetivo deste trabalho é avaliar a eficácia inseticida in vitro do óleo essencial de Lippia gracilis sobre Bovicolaovis. Os espécimes (n = 900) de B.ovis foram coletados de ovinos naturalmente infestados, distribuídos aleatoriamente em grupos, e submetidos aos diferentes tratamentos: (G1) 1.000 a 15,6 mg/ml óleo essencial de Lippia gracilis; (G2) Tween 80 a 3% (controle negativo), e: (G3) Amitraz 1,25 g/mL (controle positivo). Realizou-se 4 réplicas (25 piolhos por réplica) para cada tratamento. A viabilidade dos piolhos foi avaliada nos tempos de 1, 3, 6, 24, 48 e 72 h. Os dados foram expressos em eficácia média ± desvio padrão (SPSS versão 23.0) e diferenças estatísticas dos grupos experimentais foram obtidas por Kruskal-Wallis e Friedman. Para obtenção da CI50e CI90 foi realizada Regressão de Probit, com nível de significância de p< 0,05. O óleo essencial de L. gracilis nas concentrações de 100 a 62,5 mg/mL apresentaram eficácia de 100%, 1 h pós-exposição. Os valores de CI50 e CI90 foram 18,1 mg/mL e 44,9 mg/mL, respectivamente. Dessa forma, concluímos que o óleo essencial de L. gracilis apresentou eficácia sobre B.ovis in vitro.

The objective of this work is to evaluate the in vitro insecticidal efficacy of the essential oil of Lippia gracilis against Bovicolaovis. The specimens (n = 900) of B.ovis were collected from naturally infested sheep, randomly distributed in groups, and subjected to different treatments: (G1) 1,000 to 15.6 mg/ml essential oil of Lippia gracilis; (G2) 3% Tween 80 (negative control), and: (G3) Amitraz 1.25 g/mL (positive control). 4 replicates (25 lice per replica) were performed for each treatment. Louse viability was assessed at 1, 3, 6, 24, 48 and 72 h. The data were expressed as mean efficacy ± standard deviation (SPSS version 23.0) and statistical differences in the experimental groups were obtained by Kruskal-Wallis and Friedman. Probit regression was performed to obtain the IC50 and CI90, with a significance level of p <0.05. The essential oil of L. gracilis at concentrations of 100 to 62.5 mg/mL showed 100% efficacy, 1 h post-exposure. The IC50 and CI90 values were 18.1 mg/mL and 44.9 mg/mL, respectively. Thus, we conclude that L. gracilis essential oil is effective on B. ovis.

Ação inseticida in vitro do óleo essencial de Lippia gracilis Schauer sobre Damalinia Bovicola ovis (Schrank, 1781) / In vitro action of Lippia gracilis Schauer essential oil on Damalinia Bovicola ovis (Schrank, 1781)

O objetivo deste trabalho é avaliar a eficácia inseticida in vitro do óleo essencial de Lippia gracilis sobre Bovicolaovis. Os espécimes (n = 900) de B.ovis foram coletados de ovinos naturalmente infestados, distribuídos aleatoriamente em grupos, e submetidos aos diferentes tratamentos: (G1) 1.000 a 15,6 mg/ml óleo essencial de Lippia gracilis; (G2) Tween 80 a 3% (controle negativo), e: (G3) Amitraz 1,25 g/mL (controle positivo). Realizou-se 4 réplicas (25 piolhos por réplica) para cada tratamento. A viabilidade dos piolhos foi avaliada nos tempos de 1, 3, 6, 24, 48 e 72 h. Os dados foram expressos em eficácia média ± desvio padrão (SPSS versão 23.0) e diferenças estatísticas dos grupos experimentais foram obtidas por Kruskal-Wallis e Friedman. Para obtenção da CI50e CI90 foi realizada Regressão de Probit, com nível de significância de p< 0,05. O óleo essencial de L. gracilis nas concentrações de 100 a 62,5 mg/mL apresentaram eficácia de 100%, 1 h pós-exposição. Os valores de CI50 e CI90 foram 18,1 mg/mL e 44,9 mg/mL, respectivamente. Dessa forma, concluímos que o óleo essencial de L. gracilis apresentou eficácia sobre B.ovis in vitro.

The objective of this work is to evaluate the in vitro insecticidal efficacy of the essential oil of Lippia gracilis against Bovicolaovis. The specimens (n = 900) of B.ovis were collected from naturally infested sheep, randomly distributed in groups, and subjected to different treatments: (G1) 1,000 to 15.6 mg/ml essential oil of Lippia gracilis; (G2) 3% Tween 80 (negative control), and: (G3) Amitraz 1.25 g/mL (positive control). 4 replicates (25 lice per replica) were performed for each treatment. Louse viability was assessed at 1, 3, 6, 24, 48 and 72 h. The data were expressed as mean efficacy ± standard deviation (SPSS version 23.0) and statistical differences in the experimental groups were obtained by Kruskal-Wallis and Friedman. Probit regression was performed to obtain the IC50 and CI90, with a significance level of p <0.05. The essential oil of L. gracilis at concentrations of 100 to 62.5 mg/mL showed 100% efficacy, 1 h post-exposure. The IC50 and CI90 values were 18.1 mg/mL and 44.9 mg/mL, respectively. Thus, we conclude that L. gracilis essential oil is effective on B. ovis.

Anthelmintic activity of nanoencapsulated carvacryl acetate against gastrointestinal nematodes of sheep and its toxicity in rodents.

The objective of this study was to evaluate the efficacy of carvacryl acetate (CVA) and nanoencapsulated CVA (nCVA) on gastrointestinal nematodes of sheep. The CVA was nanoencapsulated with chitosan/gum arabic and the efficacy of nanoencapsulation (EE), yield, zeta potential, nanoparticle morphology and release kinetics at pH 3 and 8 were analyzed. Acute and subchronic toxicity were evaluated in rodents and reduction of egg counts in the faeces (FECRT) of sheep. The sheep were divided into four groups (n = 10): G1, 250 mg/kg CVA; G2, 250 mg/kg nCVA; G3, polymer matrix and G4: 2.5 mg/kg monepantel. EE and nCVA yield were 65% and 57%, respectively. The morphology of the nanoparticles was spherical, size (810.6±286.7 nm), zeta potential in pH 3.2 (+18.3 mV) and the 50% release of CVA at pHs 3 and 8 occurred at 200 and 10 h, respectively. nCVA showed LD50 of 2,609 mg/kg. CVA, nCVA and monepantel reduced the number of eggs per gram of faeces (epg) by 57.7%, 51.1% and 97.7%, respectively. The epg of sheep treated with CVA and nCVA did not differ from the negative control (P>0.05). Nanoencapsulation reduced the toxicity of CVA; however, nCVA and CVA presented similar results in the FECRT.

Anthelmintic activity of nanoencapsulated carvacryl acetate against gastrointestinal nematodes of sheep and its toxicity in rodents / Atividade anti-helmíntica do acetato de carvacrila nanoencapsulado sobre nematoides gastrintestinais de ovinos e toxicidade em roedores

Abstract The objective of this study was to evaluate the efficacy of carvacryl acetate (CVA) and nanoencapsulated CVA (nCVA) on gastrointestinal nematodes of sheep. The CVA was nanoencapsulated with chitosan/gum arabic and the efficacy of nanoencapsulation (EE), yield, zeta potential, nanoparticle morphology and release kinetics at pH 3 and 8 were analyzed. Acute and subchronic toxicity were evaluated in rodents and reduction of egg counts in the faeces (FECRT) of sheep. The sheep were divided into four groups (n = 10): G1, 250 mg/kg CVA; G2, 250 mg/kg nCVA; G3, polymer matrix and G4: 2.5 mg/kg monepantel. EE and nCVA yield were 65% and 57%, respectively. The morphology of the nanoparticles was spherical, size (810.6±286.7 nm), zeta potential in pH 3.2 (+18.3 mV) and the 50% release of CVA at pHs 3 and 8 occurred at 200 and 10 h, respectively. nCVA showed LD50 of 2,609 mg/kg. CVA, nCVA and monepantel reduced the number of eggs per gram of faeces (epg) by 57.7%, 51.1% and 97.7%, respectively. The epg of sheep treated with CVA and nCVA did not differ from the negative control (P>0.05). Nanoencapsulation reduced the toxicity of CVA; however, nCVA and CVA presented similar results in the FECRT.

Resumo O objetivo deste trabalho foi avaliar a eficácia do acetato de carvacrila (ACV) e do ACV nanoencapsulado (nACV) sobre nematóides gastrintestinais de ovinos. O ACV foi nanoencapsulado com quitosana/goma arábica e foi analisada a eficácia de nanoencapsulamento (EE), o rendimento, potencial zeta, morfologia das nanopartículas e cinética de liberação em pH 3 e 8. Foram avaliadas as toxicidades aguda e subcrônica em roedores e a redução da contagem de ovos nas fezes (RCOF) de ovinos. Os ovinos foram divididos em quatro grupos (n = 10): G1, 250 mg/kg ACV; G2, 250 mg/kg de nACV; G3, matriz polimérica e G4: 2,5 mg/kg de monepantel. A EE e o rendimento de nACV foram de 65% e 57%, respectivamente. A morfologia das nanopartículas foi esférica, tamanho (810,6±286,7 nm), potencial zeta no pH 3,2 (+18,3 mV) e a liberação de 50% de CVA nos pHs 3 e 8 ocorreu às 200 e 10 h, respectivamente. nACV apresentou DL50 de 2.609 mg/kg. ACV, nACV e o monepantel reduziram a contagem de ovos por grama de fezes (opg) em 57,7%, 51,1% e 97,7%, respectivamente. A contagem de opg de ovelhas tratadas com ACV e nCVA não diferiu do controle negativo (P>0,05). O nanoencapsulamento reduziu a toxicidade do AVC; no entanto, nACV e ACV apresentaram resultados semelhantes na RCOF.

Anthelmintic activity of Eucalyptus citriodora essential oil and its major component, citronellal, on sheep gastrointestinal nematodes.

This study aimed to evaluate the anthelmintic activity of Eucalyptus citriodora essential oil and citronellal on sheep gastrointestinal nematodes. Essential oil composition was determined by gas chromatography mass spectrometry. The substances were evaluated in vitro using adult worm motility test (AWMT) and transmission electron microscopy (TEM). The acute toxicity test in mice and the fecal egg count reduction test (FECRT) in sheep were performed. Citronellal was confirmed as the essential oil major constituent (63.9%). According to the AWMT, 2 mg/mL of essential oil and citronellal completely inhibited Haemonchus contortus motility at 6 h post exposure. H. contortus exposed to essential oil and citronellal exhibited internal ultrastructural modifications. The lethal dose 50 values in mice were 5,000 and 2,609 mg/kg for essential oil and citronellal, respectively. E. citriodora essential oil reduced sheep epg at 14 days post treatment by 69.5% (P<0.05). No significant differences were observed in epg between the citronellal and negative control groups (P>0.05). The interaction between citronellal and other constituents in the essential oil may be relevant for its in vivo anthelmintic activity. Thus, E. citriodora essential oil and citronellal pharmacokinetic studies may help elucidate the anthelmintic activity of these compounds.

Anthelmintic activity of Eucalyptus citriodora essential oil and its major component, citronellal, on sheep gastrointestinal nematodes / Atividade anti-helmíntica do óleo essencial de Eucalyptus citriodora e seu componente majoritário, citronelal, sobre nematoides gastrintestinais de ovinos

Abstract This study aimed to evaluate the anthelmintic activity of Eucalyptus citriodora essential oil and citronellal on sheep gastrointestinal nematodes. Essential oil composition was determined by gas chromatography mass spectrometry. The substances were evaluated in vitro using adult worm motility test (AWMT) and transmission electron microscopy (TEM). The acute toxicity test in mice and the fecal egg count reduction test (FECRT) in sheep were performed. Citronellal was confirmed as the essential oil major constituent (63.9%). According to the AWMT, 2 mg/mL of essential oil and citronellal completely inhibited Haemonchus contortus motility at 6 h post exposure. H. contortus exposed to essential oil and citronellal exhibited internal ultrastructural modifications. The lethal dose 50 values in mice were 5,000 and 2,609 mg/kg for essential oil and citronellal, respectively. E. citriodora essential oil reduced sheep epg at 14 days post treatment by 69.5% (P<0.05). No significant differences were observed in epg between the citronellal and negative control groups (P>0.05). The interaction between citronellal and other constituents in the essential oil may be relevant for its in vivo anthelmintic activity. Thus, E. citriodora essential oil and citronellal pharmacokinetic studies may help elucidate the anthelmintic activity of these compounds.

Resumo Este trabalho objetivou avaliar a atividade anti-helmíntica do óleo essencial de Eucalyptus citriodora e citronelal sobre nematoides gastrintestinais de ovinos. A composição do óleo essencial foi determinada por cromatografia gasosa acoplada à espectrometria de massas. As substâncias foram avaliadas in vitro utilizando-se teste de motilidade de vermes adultos (AWMT) e microscopia eletrônica de transmissão (TEM). Teste de toxicidade aguda em camundongos e teste de redução da contagem de ovos fecais (FECRT) em ovinos foram realizados. Citronelal foi confirmado como componente majoritário do óleo essencial (63,9%). No AWMT, 2 mg/mL de óleo essencial e citronelal inibiram completamente a motilidade de H. contortus 6 h pós-exposição. H. contortus expostos ao óleo essencial e citronelal exibiram modificações ultraestruturais internas. Os valores da dose letal 50 em camundongos foram 5.000 e 2.609 mg/kg para óleo essencial e citronelal, respectivamente. Óleo essencial de E. citriodora reduziu opg de ovinos 14 dias pós-tratamento em 69,5% (P<0,05). Não houve diferença significativa de opg entre grupo controle negativo e citronelal (P>0,05). A interação entre citronelal e outros constituintes do óleo essencial pode ser relevante na atividade anti-helmíntica in vivo. Portanto, avaliação farmacocinética do óleo essencial de E. citriodora e citronelal pode auxiliar a elucidar a atividade anti-helmíntica desses compostos.

Anthelmintic effect of Cymbopogon citratus essential oil and its nanoemulsion on sheep gastrointestinal nematodes.

The anthelmintic resistance stimulated the search for strategies for controlling gastrointestinal nematodes, including the use of free essential oils or its nanoemulsion. This study evaluated the anthelmintic efficacy of Cymbopogon citratus essential oil (CcEO) and C. citratus essential oil nanoemulsion (CcEOn). Pysicochemical analyses were performed. The in vitro effect was determined using the egg hatch test (EHT) on Haemonchus contortus and in vivo effect was evaluated in sheep infected with gastrointestinal nematodes. The animals were treated with CcEO (500 mg/kg) or CcEOn (450 mg/kg) for the fecal egg count (FEC) and the determination of worm burden. The main component of CcEO was citral. The CcEO content in the nanoemulsion was 20% (v/v), and the mean particle size was 248 nm. In EHT, CcEO and CcEOn (1.25 mg/mL) inhibited larval hatching by 98.4 and 97.1%, respectively. Three animals treated with CcEO died whereas in the group treated with CcEOn one animal died. The FEC and total worm burden of the treated groups did not differ from the negative control (p>0.05). The CcEOn showed efficacy only on H. contortus (p<0.05). In conclusion, nanoencapsulation reduced toxicity and increased efficacy on H. contortus.

Anthelmintic effect of Cymbopogon citratus essential oil and its nanoemulsion on sheep gastrointestinal nematodes / Efeito anti-helmíntico do óleo essencial de Cymbopogon citratus e sua nanoemulsão sobre nematoides gastrintestinais de ovinos

Abstract The anthelmintic resistance stimulated the search for strategies for controlling gastrointestinal nematodes, including the use of free essential oils or its nanoemulsion. This study evaluated the anthelmintic efficacy of Cymbopogon citratus essential oil (CcEO) and C. citratus essential oil nanoemulsion (CcEOn). Pysicochemical analyses were performed. The in vitro effect was determined using the egg hatch test (EHT) on Haemonchus contortus and in vivo effect was evaluated in sheep infected with gastrointestinal nematodes. The animals were treated with CcEO (500 mg/kg) or CcEOn (450 mg/kg) for the fecal egg count (FEC) and the determination of worm burden. The main component of CcEO was citral. The CcEO content in the nanoemulsion was 20% (v/v), and the mean particle size was 248 nm. In EHT, CcEO and CcEOn (1.25 mg/mL) inhibited larval hatching by 98.4 and 97.1%, respectively. Three animals treated with CcEO died whereas in the group treated with CcEOn one animal died. The FEC and total worm burden of the treated groups did not differ from the negative control (p>0.05). The CcEOn showed efficacy only on H. contortus (p<0.05). In conclusion, nanoencapsulation reduced toxicity and increased efficacy on H. contortus.

Resumo A resistência anti-helmíntica estimulou a busca por estratégias de controle de nematoides gastrintestinais, incluindo óleos essenciais livres ou em nanoemulsão. Este estudo avaliou a eficácia anti-helmíntica do óleo essencial de Cymbopogon citratus (OECc) e da nanoemulsão do óleo essencial de C. citratus (nOECc). Análises físico-químicas foram realizadas. O efeito in vitro foi determinado no teste de eclosão de ovos (TEO) sobre Haemonchus contortus e o efeito in vivo foi avaliado em ovinos infectados com nematoides gastrintestinais. Os animais foram tratados com OECc (500 mg/kg) ou nOECc (450 mg/kg) para determinação do número de ovos por grama de fezes (OPG) e carga parasitária. O principal constituinte do OECc foi citral. O conteúdo de OECc na nanoemulsão foi 20% e o tamanho médio de partícula foi 248 nm. No TEO, OECc e nOECc (1,25 mg/mL) inibiram 98,4 e 97,1% da eclosão larvar, respectivamente. Três animais tratados com o OECc morreram, enquanto um animal do grupo tratado com a nOECc morreu. O OPG e a carga parasitária total dos grupos tratados não diferiram do controle negativo (p>0.05). A nOECc apresentou eficácia somente sobre H. contortus (p<0.05). Como conclusão, o nanoencapsulamento reduziu a toxicidade e aumentou a eficácia sobre H. contortus.

Haemonchus contortus ß-tubulin isotype 1 gene F200Y and F167Y SNPs are both selected by ivermectin and oxfendazole treatments with differing impacts on anthelmintic resistance.

Parasitism by Haemonchus contortus is one of the main limiting factors in small ruminant production in tropical areas. Benzimidazoles (BZ) and macrocyclic lactones (ML) are the most used anthelmintic classes in gastrointestinal nematodes control. There is considerable scientific evidence of a possible relation between the anthelmintic resistance to BZ and ML. This study aimed to characterize the dynamics of anthelmintic resistance in an H. contortus susceptible isolate under selection pressure for BZ and ML alone or in combination and the role of isotype 1 ß-tubulin gene SNPs in these situations. A total of 12 Somali sheep were infected with 5000 third stage larvae of H. contortus Inbred-Susceptible Edinburgh (ISE) isolate. Once infection was established, animals were distributed in three groups (n=4), each treated with crescent doses of oxfendazole (OXF), ivermectin (IVM) and oxfendazole plus ivermectin (IVMOXF). An additional control group with untreated animals was maintained during the entire experiment. After each treatment, eggs were collected and real-time PCR was performed to identify single nucleotide polymorphisms (SNPs) F167Y, F200Y and E198A, in addition to egg hatch test (EHT) for BZ and larval development test (LDT) for ivermectin resistance. All treatments led to increased resistance allelic frequencies at SNPs F200Y and F167Y (p <0.05). In vitro results showed increased phenotypic resistance against both anthelmintic classes in groups IVM and IVMOXF while group OXF only developed resistance against BZ. Finally, we provide evidence that while isotype 1 ß-tubulin gene SNPs may have some involvement with ML resistance, the presence of these ß-tubulin SNPs alone are not sufficient to develop ML resistance.

Anthelmintic effect of thymol and thymol acetate on sheep gastrointestinal nematodes and their toxicity in mice.

Thymol is a monoterpene and acetylation form of this compound can reduce the toxicity and enhance its biological effects. The objective of this study was to evaluate the effect of thymol and thymol acetate (TA) on egg, larva and adult Haemonchus contortus and the cuticular changes, acute toxicity in mice and the efficacy on sheep gastrointestinal nematodes. In vitro tests results were analyzed by analysis of variance (ANOVA) and followed by comparison with Tukey test or Bonferroni. The efficacy of in vivo test was calculated by the BootStreet program. In the egg hatch test (EHT), thymol (0.5 mg/mL) and TA (4 mg/mL) inhibited larval hatching by 98% and 67.1%, respectively. Thymol and TA (8 mg/mL) inhibited 100% of larval development. Thymol and TA (800 µg/mL) reduced the motility of adult worms, by 100% and 83.4%, respectively. Thymol caused cuticular changes in adult worm teguments. In the acute toxicity test, the LD50 of thymol and TA were 1,350.9 mg/kg and 4,144.4 mg/kg, respectively. Thymol and TA reduced sheep egg count per gram of faeces (epg) by 59.8% and 76.2%, respectively. In in vitro tests thymol presented better anthelmintic activity than TA. However TA was less toxic and in in vivo test efficacy was similar.

Anthelmintic effect of thymol and thymol acetate on sheep gastrointestinal nematodes and their toxicity in mice / Efeito anti-helmíntico do timol e acetato de timila sobre nematoides gastrintestinais de ovinos e toxicidade em camundongos

Abstract Thymol is a monoterpene and acetylation form of this compound can reduce the toxicity and enhance its biological effects. The objective of this study was to evaluate the effect of thymol and thymol acetate (TA) on egg, larva and adult Haemonchus contortus and the cuticular changes, acute toxicity in mice and the efficacy on sheep gastrointestinal nematodes. In vitro tests results were analyzed by analysis of variance (ANOVA) and followed by comparison with Tukey test or Bonferroni. The efficacy of in vivo test was calculated by the BootStreet program. In the egg hatch test (EHT), thymol (0.5 mg/mL) and TA (4 mg/mL) inhibited larval hatching by 98% and 67.1%, respectively. Thymol and TA (8 mg/mL) inhibited 100% of larval development. Thymol and TA (800 µg/mL) reduced the motility of adult worms, by 100% and 83.4%, respectively. Thymol caused cuticular changes in adult worm teguments. In the acute toxicity test, the LD50 of thymol and TA were 1,350.9 mg/kg and 4,144.4 mg/kg, respectively. Thymol and TA reduced sheep egg count per gram of faeces (epg) by 59.8% and 76.2%, respectively. In in vitro tests thymol presented better anthelmintic activity than TA. However TA was less toxic and in in vivo test efficacy was similar.

Resumo Timol é um monoterpeno e a acetilação deste composto pode reduzir a toxicidade e potencializar os seus efeitos biológicos. O objetivo deste trabalho foi avaliar o efeito do timol e acetato de timolila (AT) sobre ovos, larvas e adultos de Haemonchus contortus e suas alterações cuticulares, toxicidade aguda em camundongos e a eficácia sobre nematoides gastrintestinais de ovinos. Os resultados dos testes in vitro foram analisados por análise de variância (ANOVA) e comparados pelo testes de Tukey ou Bonferroni. A eficácia do teste de redução da contagem de ovos nas fezes (RCOF) foi calculada pelo programa BootStreet. No teste de inibição da eclosão de ovos (TEO), timol (0,5 mg/mL) e AT (4 mg/mL) inibiram a eclosão das larvas em 98% e 67,1%, respectivamente. Timol e AT (8 mg/mL) inibiram 100% do desenvolvimento larval. Timol e AT (800 μg/mL) reduziram a motilidade dos nematoides adultos, em 100% e 83,4%, respectivamente. O timol provocou alterações cuticulares nos nematoides adultos. No teste de toxicidade aguda, a DL50 do timol e AT foi de 1.350,9 mg/kg e 4.144,4 mg/kg, respectivamente. Timol e AT reduziram a contagem de ovos por gramas de fezes (OPG) dos ovinos em 59,8% e 76,2%, respectivamente. Nos testes in vitro timol apresentou atividade melhor anti-helmíntica do que AT. Entretanto, AT foi menos tóxico do que o timol e no teste in vivo apresentaram eficácia semelhante.

Monitoramento do ciclo estral de fêmeas equinas por meio de citologia vaginal, ultrassonografia e dosagem hormonal / Monitoring equine estrous cycle through vaginal cytology, ultrasonography and hormone dosage / Monitoreo del ciclo estral de hembras equinas por medio de citología vaginal, ultrasonido y dosis hormonal

O ciclo estral é a sincronia entre eventos comportamentais, anatômicos e endócrinos que resultam na ovulação, e pode ser dividido em fase lútea e fase folicular. A identificação das fases do ciclo estral da égua pode ser acompanhada por meio de várias biotécnicas reprodutivas como, por exemplo, citologia vaginal classificando os diversos tipos de células do epitélio vaginal, ultrassonografia para avaliação da dinâmica folicular e a dosagem hormonal no plasma sanguíneo ou das fezes do animal para caracterizar a fase do ciclo estral que o animal possivelmente se encontra. Sendo assim, o objetivo desta revisão foi discutir a fisiologia do ciclo estral de éguas, a fim de favorecer novas pesquisas relacionadas à biotécnicas reprodutivas para equinos.

The estrous cycle is the synchrony of behavioral, anatomical and endocrine events that result in ovulation. It can be divided into luteal and follicular phases. The identification of the stages in the estrous cycle of mares can be accomplished through several reproductive biotechnologies, such as, for instance, vaginal cytology classifying the different types of cells in the vaginal epithelium, follicular dynamics evaluated through ultrasound examination, and hormone dosage in blood plasma or feces from the animal to characterize the likely estrous cycle phase. Thus, the objective of this review is to discuss the physiology of the estrous cycle in mares in order to encourage new research related to reproductive biotechnologies for horses.

El ciclo estral es la sincronía entre eventos de comportamiento, anatómicos y endocrinos que resultan en la ovulación, y se puede dividir en fase lútea y fase folicular. La identificación de las etapas del ciclo estral de la yegua puede ser acompañada a través de varias biotécnicas reproductivas como, por ejemplo, citología vaginal, clasificando los diferentes tipos de células del epitelio vaginal, ultrasonido para evaluación de la dinámica folicular y la dosificación hormonal en el plasma sanguíneo o de las heces del animal para caracterizar la fase del ciclo estral que el animal posiblemente se encuentra. Así, el objetivo de esta revisión ha sido examinar la fisiología del ciclo estral de yeguas, con el fin de fomentar nuevas investigaciones relacionadas con las biotecnologías reproductivas para equinos.

Leishmaniose tegumentar americana canina na zona rural do município de São Miguel, Rio Grande do Norte* / Leishmaniasis Tegumentary American dog in rural area of the county of São Miguel, Rio Grande do Norte Francisco Rodrigues de Freitas Araújo,** Weibson Paz Pinheiro Andre,*** Paulo Fernando Cisneiros da Costa Reis,****

Leishmaniose Tegumentar Americana (LTA) constitui um problema de saúde pública emergente no Rio Grande do Norte, com relatos de casos humanos principalmente na Região Serrana do Alto Oeste do estado, uma vez que o cão tem um importante papel na manutenção da doença neste ambiente. O presente estudo teve por objetivo analisar prevalência da LTA canina em quatro localidades localizadas na Zona Rural do município de São Miguel, RN. Foram realizados exames clínicos e diagnóstico sorológico através das técnicas de ELISA e RIFI em 100 cães de diferentes sexo, idade e categorias companhia e de caça. Todas as localidades apresentaram cães soropositivos com resultados de 21% de positividade para a técnica de ELISA e 10% para a de RIFI. Maiores prevalências foram observadas em cães machos, assintomáticos, na faixa etária de dois anos de idade e de caça. O presente estudo confirma a importância epidemiológica como reservatório da infecção e poderá auxiliar nas medidas de prevenção e controle da doença no município. .

American Cutaneous Leishmaniasis ( ACL) is an emerging public health problem in Rio Grande do Norte , with reports of human cases mainly in the mountainous region of the Upper Midwest state . Once the dog has an important role in maintaining the disease in this environment the present study aimed to evaluate the prevalence of canine ACL at four sites located in the Rural Area of São Miguel , RN . Clinical and serological diagnosis through ELISA and IFA in 100 dogs of different sex, age and company categories and hunting were performed . All towns had seropositive dogs with results of 21 % positivity for ELISA and 10 % for the IFA. Highest prevalence rates were observed in males , healthy dogs aged two years old and hunting . The present study confirms the epidemiological importance as a reservoir of infection and may assist in the prevention and control of disease in the city.

Leishmaniose tegumentar americana canina na zona rural do município de São Miguel, Rio Grande do Norte / Leishmaniasis Tegumentary American dog in rural area of the county of São Miguel, Rio Grande do Norte

Leishmaniose Tegumentar Americana (LTA) constitui um problema de saúde pública emergente no Rio Grande do Norte, com relatos de casos humanos principalmente na Região Serrana do Alto Oeste do Estado. Uma vez que o cão tem um importante papel na manutenção da doença neste ambiente. O presente estudo teve por objetivo analisar prevalência da LTA canina em quatro localidades localizadas na Zona Rural do município de São Miguel, RN. Foram realizados exames clínicos e diagnóstico sorológico através das técnicas de ELISA e RIFI em 100 cães de diferentes sexo, idade e categorias companhia e de caça. Todas as localidades apresentaram cães soropositivos com resultados de 21% de positividade para a técnica de ELISA e 10% para a de RIFI. Maiores prevalências foram observadas em cães machos, assintomáticos, na faixa etária de dois anos de idade e de caça. O presente estudo confirma a importância epidemiológica como reservatório da infecção e poderá auxiliar nas medidas de prevenção e controle da doença no município.

American Cutaneous Leishmaniasis ( ACL) is an emerging public health problem in Rio Grande do Norte , with reports of humancases mainly in the mountainous region of the Upper Midwest state . Once the dog has an important role in maintaining the diseasein this environment the present study aimed to evaluate the prevalence of canine ACL at four sites located in the Rural Area of SãoMiguel , RN . Clinical and serological diagnosis through ELISA and IFA in 100 dogs of different sex, age and company categoriesand hunting were performed . All towns had seropositive dogs with results of 21 % positivity for ELISA and 10 % for the IFA.Highest prevalence rates were observed in males , healthy dogs aged two years old and hunting . The present study confirms theepidemiological importance as a reservoir of infection and may assist in the prevention and control of disease in the city.

Monitoramento do ciclo estral de fêmeas equinas por meio de citologia vaginal, ultrassonografia e dosagem hormonal / Monitoring equine estrous cycle through vaginal cytology, ultrasonography and hormone dosage / Monitoreo del ciclo estral de hembras equinas por medio de citología vaginal, ultrasonido y dosis hormonal

O ciclo estral é a sincronia entre eventos comportamentais, anatômicos e endócrinos que resultam na ovulação, e pode ser dividido em fase lútea e fase folicular. A identificação das fases do ciclo estral da égua pode ser acompanhada por meio de várias biotécnicas reprodutivas como, por exemplo, citologia vaginal classificando os diversos tipos de células do epitélio vaginal, ultrassonografia para avaliação da dinâmica folicular e a dosagem hormonal no plasma sanguíneo ou das fezes do animal para caracterizar a fase do ciclo estral que o animal possivelmente se encontra. Sendo assim, o objetivo desta revisão foi discutir a fisiologia do ciclo estral de éguas, a fim de favorecer novas pesquisas relacionadas à biotécnicas reprodutivas para equinos...

The estrous cycle is the synchrony of behavioral, anatomical and endocrine events that result in ovulation. It can be divided into luteal and follicular phases. The identification of the stages in the estrous cycle of mares can be accomplished through several reproductive biotechnologies, such as, for instance, vaginal cytology classifying the different types of cells in the vaginal epithelium, follicular dynamics evaluated through ultrasound examination, and hormone dosage in blood plasma or feces from the animal to characterize the likely estrous cycle phase. Thus, the objective of this review is to discuss the physiology of the estrous cycle in mares in order to encourage new research related to reproductive biotechnologies for horses...

El ciclo estral es la sincronía entre eventos de comportamiento, anatómicos y endocrinos que resultan en la ovulación, y se puede dividir en fase lútea y fase folicular. La identificación de las etapas del ciclo estral de la yegua puede ser acompañada a través de varias biotécnicas reproductivas como, por ejemplo, citología vaginal, clasificando los diferentes tipos de células del epitelio vaginal, ultrasonido para evaluación de la dinámica folicular y la dosificación hormonal en el plasma sanguíneo o de las heces del animal para caracterizar la fase del ciclo estral que el animal posiblemente se encuentra. Así, el objetivo de esta revisión ha sido examinar la fisiología del ciclo estral de yeguas, con el fin de fomentar nuevas investigaciones relacionadas con las biotecnologías reproductivas para equinos...