Monitoring environmental conditions on the speed of development and larval migration of gastrointestinal nematodes in Urochloa decumbens in northeastern Brazil.

The study aimed to evaluate the speed of development of gastrointestinal nematodes (GIN) eggs to infective larvae (L3) and its migration under effect of meteorological variables: temperature, rainfall, relative humidity, solar radiation to Urochloa (=Brachiaria) decumbens grass pasture during rainy season, from June to September 2019, and dry season, October to March 2020, in Recôncavo baiano region of Bahia state, Brazil. Monthly, fecal samples obtained from goats with recurrent GIN infection were deposited in six plots of one square meter. In +7, +14 and + 21-days post-deposition, lower and upper strata grass (0-15 and > 15 cm, respectively) and remaining feces were collected and submitted to Baermann's technique modified to perform larval count and identification. Meteorological data were obtained from a local weather station database. The log-transformed larval count results were analyzed regarding the collection day effect, stratum effect, comparing the means by Tukey's test (p < 0.05). Multivariate regression analysis and correlation of meteorological variables with larval counts was performed. In the rainy season, the largest proportion of recovered L3 was concentrated in +7-days post-deposition in the months of June and July, both in the remaining feces and herbage samples, while in August development took place more slowly, after +14-days post-deposition. During the dry season, L3 development only occurred after the first collection in January and February. Lower strata had higher proportion of recovery than in upper strata. Negative correlation was found for solar radiation. Greater rainfall in the rainy season compared to dry season was a favorable condition for high contamination of grass samples. However, reduced vertical migration to upper strata may have been influenced by low temperatures during the rainy season. Regarding the genera present in the fecal samples used in the experiment were found Haemonchus sp., Trichostrongylus sp. and Oesophagostomum sp. Thus, it is possible to conclude that in the region of the study, during the rainy season, there is a rapid larval development and the L3 are able to migrate more effectively to the grass. On the other hand, in the dry season, larval development tends to be slower, with less migration to the grass. Solar radiation can be useful for predicting months with the highest risk of infection. Therefore, prophylactic measures should be employed in goat herds during the rainy season whilst natural reduced pasture contamination in the dry season may favor animal maintenance for a longer time.

Combination of entomopathogenic nematodes with acaricides or essential oil of Lippia triplinervis against Rhipicephalus microplus (Acari: Ixodidae).

This study aimed to evaluate the combination effect of Heterorhabditis bacteriophora HP88 and H. indica LPP1, with the acaricides deltamethrin, amitraz and chlorfenvinphos, and the essential oil (EO) of Lippia triplinervis, against engorged females of Rhipicephalus microplus. In order to verify the effect of acaricides and EO, the adult immersion test was used, and in the groups treated only with entomopathogenic nematodes (EPNs), 150 infective juveniles were used per female. In the treatments with nematodes in combination with the acaricides or EO, the females were immersed in the solutions (acaricide or EO) and then transferred to Petri dishes for application of the nematodes. The treatment with acaricides resulted in a control percentage lower than 70%, except in the group treated with chlorfenvinphos in the second experiment (84.3%). The control percentage was 73% for L. triplinervis EO, and greater than 90% in all the groups treated with nematodes. For treatments with EPNs combined with the acaricides or EO, the efficacy was greater than 95% (except for deltamethrin + HP88), and reached 100% in the treatment with LPP1 + amitraz. It can be concluded that the EPNs at the concentrations tested were compatible with the acaricides deltamethrin, amitraz and chlorfenvinphos, and with the EO of L. triplinervis. These combinations enhance the effect of these control agents.

In vitro efficacy of two commercial products of Metarhizium anisopliae s.l. for controlling the cattle tick Rhipicephalus microplus.

The effects of two different products - Metarril® SP Organic (dry conidia) and Metarril® SC Organic (emulsifiable concentrated conidia in vegetable oil) - on eggs, larvae and Rhipicephalus microplus engorged females were here explored. Three concentrations (108, 107, and 106 conidia mL-1) for both products were prepared in water + 0.1% Tween® 80 (v/v); afterward, bioassays were carried out for all R. microplus stages by immersion in suspensions (Metarril® SP) or formulations (Metarril® SC). Metarril® SP suspensions showed low efficacy and did not affect biological parameters of treated engorged females; for eggs and larvae, only slight decreases in hatchability and larvae population were observed. Despite a delay in germination, Metarril® SC presented better results; for females, reductions in Egg Mass Weight (EMW) and Egg Production Index (EPI) were reported. On eggs, 108 conidia mL-1 increased Incubation Period (IP), shortened Hatching Period (HP) and decreased hatchability by up to 61%; for larvae, 107 and 108 conidia mL-1 reached 99.6 and 100% larval mortality respectively, 10 days after fungal exposure. Thus, further studies involving the use of oil-based formulations for ticks such as Metarril® SC need to be performed, especially to control the most susceptible stages (eggs and larvae).

In vitro efficacy of two commercial products of Metarhizium anisopliae s.l. for controlling the cattle tick Rhipicephalus microplus / Eficácia in vitro de dois produtos comerciais de Metarhizium anisopliae s.l. no controle do carrapato bovino Rhipicephalus microplus

Abstract The effects of two different products - Metarril® SP Organic (dry conidia) and Metarril® SC Organic (emulsifiable concentrated conidia in vegetable oil) - on eggs, larvae and Rhipicephalus microplus engorged females were here explored. Three concentrations (108, 107, and 106 conidia mL-1) for both products were prepared in water + 0.1% Tween® 80 (v/v); afterward, bioassays were carried out for all R. microplus stages by immersion in suspensions (Metarril® SP) or formulations (Metarril® SC). Metarril® SP suspensions showed low efficacy and did not affect biological parameters of treated engorged females; for eggs and larvae, only slight decreases in hatchability and larvae population were observed. Despite a delay in germination, Metarril® SC presented better results; for females, reductions in Egg Mass Weight (EMW) and Egg Production Index (EPI) were reported. On eggs, 108 conidia mL-1 increased Incubation Period (IP), shortened Hatching Period (HP) and decreased hatchability by up to 61%; for larvae, 107 and 108 conidia mL-1 reached 99.6 and 100% larval mortality respectively, 10 days after fungal exposure. Thus, further studies involving the use of oil-based formulations for ticks such as Metarril® SC need to be performed, especially to control the most susceptible stages (eggs and larvae).

Resumo No presente trabalho, os efeitos de dois diferentes produtos foram avaliados - Metarril® SP Organic (conídios secos) e Metarril® SC Organic (conídios concentrados em óleo vegetal) - para ovos, larvas e fêmeas ingurgitadas de Rhipicephalus microplus. Três concentrações (108, 107 e 106 conídios mL-1) para cada produto foram preparadas em água + Tween® 80 0,1% (v/v); os bioensaios foram realizados para todos os estágios de R. microplus por imersão nas suspensões (Metarril® SP) ou formulações (Metarril® SC). Metarril® SP não afetou os parâmetros biológicos das fêmeas, demonstrando assim baixa eficácia; para ovos e larvas, foram observadas discretas diminuições na eclodibilidade e na população de larvas. Apesar de um atraso na germinação, Metarril® SC apresentou melhores resultados; para as fêmeas, foram detectadas reduções no Peso da Massa de Ovos (PMO) e no Índice de Produção de Ovos (IPO). Para os ovos, a concentração de 108 conídios mL-1 aumentou o Período de Incubação (PI), reduziu o Período de Eclosão (PE) e também o da eclodibilidade em até 61%; para larvas, 107 e 108 conídios mL-1 atingiram 99,6 e 100% de mortalidade larval, respectivamente, 10 dias após a exposição fúngica. Com isso, estudos adicionais que envolvem o uso de formulações à base de óleo para carrapatos, como Metarril® SC, precisam ser realizados, especialmente para controlar os estágios mais suscetíveis (ovos e larvas).

Unveiling the oxidative metabolism of Rhipicephalus microplus (Acari: Ixodidae) experimentally exposed to entomopathogenic fungi.

Rhipicephalus microplus is an important tick in tropical regions due to the high economic losses caused by its parasitism. Metarhizium anisopliae and Beauveria bassiana are well-known entomopathogenic fungi that can afflict R. microplus ticks. The development of new targets and strategies to control this parasite can be driven by studies of this tick's physiology. Recently, it was reported that when exposed to adverse physiological conditions, ticks can activate fermentative pathways, indicating transition from aerobic to anaerobic metabolism. Nevertheless, the precise mechanism by which entomopathogenic fungi influence R. microplus metabolism has not been clarified, limiting understanding of the tick-fungus association. Thus, the present study aimed to evaluate the effect of infection of ticks by M. anisopliae and B. bassiana on the amount of selected carboxylic acids present in the hemolymph, enabling increased understanding of changes previously reported. The results showed preservation in the concentrations of oxalic, lactic, and pyruvic acids in the hemolymph 24 and 48 h after dropping from cattle; while there were variations in the concentration of these carboxylic acids after infection of female ticks to M. anisopliae and B. bassiana. Significant increases were observed in the concentration of oxalic and lactic acids and significant reduction of pyruvic acid for both observation times (24 and 48 h) after infection by entomopathogenic fungi. These results indicate that B. bassiana and M. anisopliae infection alters the basal metabolism of R. microplus females, resulting in the activation of fermentative pathways.

Lab-on-a-chip and SDS-PAGE analysis of hemolymph protein profile from Rhipicephalus microplus (Acari: Ixodidae) infected with entomopathogenic nematode and fungus.

In the present study, lab-on-a-chip electrophoresis (LoaC) was suggested as an alternative method to the conventional polyacrylamide gel electrophoresis under denaturing conditions (SDS-PAGE) to analyze raw cell-free tick hemolymph. Rhipicephalus microplus females were exposed to the entomopathogenic fungus Metarhizium anisopliae senso latu IBCB 116 strain and/or to the entomopathogenic nematode Heterorhabditis indica LPP1 strain. Hemolymph from not exposed or exposed ticks was collected 16 and 24 h after exposure and analyze by SDS-PAGE or LoaC. SDS-PAGE yielded 15 bands and LoaC electrophoresis 17 bands. Despite the differences in the number of bands, when the hemolymph protein profiles of exposed or unexposed ticks were compared in the same method, no suppressing or additional bands were detected among the treatments regardless the method (i.e., SDS-PAGE or chip electrophoresis using the Protein 230 Kit®). The potential of LoaC electrophoresis to detect protein bands from tick hemolymph was considered more efficient in comparison to the detection obtained using the traditional SDS-PAGE method, especially when it comes to protein subunits heavier than 100 KDa. LoaC electrophoresis provided a very good reproducibility, and is much faster than the conventional SDS-PAGE method, which requires several hours for one analysis. Despite both methods can be used to analyze tick hemolymph composition, LoaC was considered more suitable for cell-free hemolymph protein separation and detection. LoaC hemolymph band percent data reported changes in key proteins (i.e., HeLp and vitellogenin) exceptionally important for tick embryogenesis. This study reported, for the first time, tick hemolymph protein profile using LoaC.

Compatibilidade de Heterorhabditis amazonensis (Rhabditida: Heterorhabditidae) isolado RSC-5 com diferentes carrapaticidas utilizados no controle de Rhipicephalus microplus (Acari: Ixodidae) / Compatibility of Heterorhabditis amazonensis (Rhabditida: Heterorhabditidae) strain RSC-5 with different acaricides used for the control of Rhipicephalus microplus (Acari: Ixodidae)

O estudo teve como objetivo avaliar a viabilidade de Heterorhabditis amazonensis isolado RSC-5 após exposição a diferentes carrapaticidas utilizados no controle de Rhipicephalus microplus. Foram constituídos seis tratamentos, cada um composto por um produto, sendo cada grupo com 75.000 nematoides (NEPs) em suspensão de 20 mL de solução de diferentes carrapaticidas, em concentração comercial. O controle foi formado por 75.000 NEPs e 20 mL de água destilada, e todos os grupos foram mantidos em câmara climatizada a 25°C. A avaliação do percentual de sobrevivência e infectividade em lagartas Galleria mellonella foi realizada 24 e 72 horas após o início do experimento. A mortalidade de lagartas no teste de infectividade foi analisada após 72 e 120 horas. Com 24 horas de exposição, o percentual de sobrevivência de H. amazonensis RSC-5 não foi significativamente reduzido (p > 0,05) somente na exposição ao princípio ativo deltametrina. O mesmo foi observado no período de 72 horas em relação à associação clorpirifós + cipermetrina + butóxido de piperonila + citronelal. Não foi constatada sobrevivência de nenhum juvenil infectivo nos grupos expostos à associação clorfenvinfós + diclorvós. A exposição ao clorfenvinfós resultou em percentual de sobrevivência inferior a 50% após 72 horas. O potencial de infectar lagartas de G. mellonella foi reduzido apenas no grupo tratado com o princípio ativo clorfenvinfós. Dessa forma, é possível concluir que o princípio ativo clorfenvinfós e a associação clorfenvinfós + diclorvós não foram compatíveis com H. Amazonenses RSC-5, causando redução no percentual de sobrevivência e infectividade dos juvenis desse nematoide. Os outros produtos foram compatíveis, não causando redução na infectividade do isolado testado.(AU)

The aim of this study was to assess the viability of Heterorhabditis amazonensis strain RSC-5 after exposure to different acaricides used for Rhipicephalus microplus control. Six treatment groups were formed, one for each product. Each group was composed of 75,000 nematodes in a 20 mL solution of different acaricides, at commercial concentration. The control group was formed by the same number of nematodes in 20 mL of distilled water. All the groups were kept in a climate-controlled chamber at 25°C. The percentage of survival and infectivity in Galleria mellonella caterpillars were determined 24 and 72 hours after the beginning of the experiment. The mortality of the caterpillars in the infectivity test was assessed 72 and 120 hours. After 24 hours of exposure, only the active ingredient deltamethrin did not significantly reduce the survival percentage of H. amazonensis RSC-5 (p > 0.05). The same was observed after 72 hours of exposure to the combination of chlorpyriphos + cypermethrin + piperonyl butoxide + citronellal. There was no survival of infective juveniles in the groups exposed to the combination of chlorphenvinphos + dichlorvos. The exposure to chlorphenvinphos for 72 hours resulted in 50% of mortality. The potential to infect G. mellonella caterpillars was only reduced in the group treated with the active ingredient chlorphenvinphos. Chlorphenvinphos and the combination of chlorphenvinphos + dichlorvos were not compatible with H. amazonensis RSC-5, causing a reduction in the survival and infectivity of juveniles of this nematode, while the other products were compatible, causing no reduction in the infectivity of this isolate.(AU)

Effects of destruxin A on Rhipicephalus (Boophilus) microplus ticks (Acari: Ixodidae).

This study evaluated the effects of destruxin A on Rhipicephalus (Boophilus) microplus females, since this toxin is one of the likely causes of high mortality induced by the entomopathogenic fungus Metarhizium anisopliae in arthropods. Ticks were immersed or inoculated with different concentrations of destruxin A. Despite the doses applied, there were no deaths or significant alterations in oviposition between the groups treated with destruxin A and the control groups. No other external effect caused by destruxin, such as tetanic paralysis, was observed in these engorged female ticks after the treatment.

Effects of destruxin A on Rhipicephalus (Boophilus) microplus ticks (Acari: Ixodidae) / Efeitos da destruxina A no carrapato Rhipicephalus (Boophilus) microplus (Acari: Ixodidae)

This study evaluated the effects of destruxin A on Rhipicephalus (Boophilus) microplus females, since this toxin is one of the likely causes of high mortality induced by the entomopathogenic fungus Metarhizium anisopliae in arthropods. Ticks were immersed or inoculated with different concentrations of destruxin A. Despite the doses applied, there were no deaths or significant alterations in oviposition between the groups treated with destruxin A and the control groups. No other external effect caused by destruxin, such as tetanic paralysis, was observed in these engorged female ticks after the treatment.

Este estudo avaliou os efeitos da destruxina A em fêmeas de Rhipicephalus (Boophilus) microplus, uma vez que essa toxina é uma das prováveis causas da alta mortalidade induzida pelo fungo entomopatogênico Metarhizium anisopliae em artrópodes. Os carrapatos foram imersos ou inoculados com diferentes concentrações de destruxina A. Apesar das doses aplicadas, não houve mortes ou alterações significativas de postura entre os grupos tratados com destruxinas A e os grupos controle. Nenhum outro efeito externo provocado pela destruxina A, tal como paralisia tetânica, foi observado nas fêmeas ingurgitadas de carrapato após o tratamento.

Potential synergistic effect of Melia azedarach fruit extract and Beauveria bassiana in the control of Rhipicephalus (Boophilus) microplus (Acari: Ixodidae) in cattle infestations.

The use of a concentrate emulsion of Melia azedarach green fruits and a suspension of the fungus Beauveria bassiana was evaluated in the control of Rhipicephalus microplus on artificially infested cattle. The evaluation was conducted following the protocol established by the Brazilian Agriculture Ministry. Five groups of 4 or 5 animals were allocated to one of the following treatments: emulsion concentrate of M. azedarach at 0.25% (T AZED 0.25%), emulsion concentrate of M. azedarach at 0.5% (T AZED 0.5%), B. bassiana at 2.4 × 10(8) conidia (T BASS), association of the concentrate of M. azedarach at 0.25% with B. bassiana at 2.4 × 10(8) conidia (T AZED 0.25%+BASS), and control (untreated). The association of the two compounds provided better results than any one isolated treatment, indicating compatibility or perhaps a synergy between M. azedarach and B. bassiana. This treatment resulted in fewer engorged females (129 ± 70) than in the control group (233 ± 82), showing high performance against all developmental stages of the tick. Results revealed an apparent synergistic effect of M. azedarach and B. bassiana in the control of R. microplus that should be further investigated.