New Field Technique to Evaluate the Action of the Fungus Metarhizium Anisopliae on Rhipicephalus (boophilus) Microplus Tick

The tick Rhipicephalus microplus parasitizes cattle and transmits infectious agents, thus generating losses estimated at US$ 3.24 billion per year. Although chemical treatment is the main means of control, there are records of populations resistant to these products in several regions of Brazil. The losses associated with production animals and resistance to acaricides have given rise to searching for alternative control strategies. The entomopathogenic fungus Metarhizium anisopliae is a promising biological control agent. The objective of this work was to evaluate the action of the fungus M. anisopliae for controlling R. microplus, using a novel semi-field test protocol. The experiment began with application of a solution of M. anisopliae (isolate IBCB 425) at a concentration of 2 x 1013 conidia/hectare, to the soil. The broth containing the fungus was made up of water and mineral oil in the proportions of 19:1. The same broth was applied without addition of fungus in the control group. Females of R. microplus were placed on the soil, wrapped in voile fabric, in both plots. The groups were left in the pasture for 15 days, after which they were collected and kept in a BOD chamber for the end of incubation and hatching. Identical deposition was repeated three times every 14 days for both groups in the same areas. Engorged females of both groups were evaluated in relation to fungal infection, in histological sections that were then stained with hematoxylin and eosin (HE) and by means of in situ hybridization with oligonucleotides labeled with biotin. The results showed that the effectiveness of the fungus continued even beyond 30 days after application. This suggested that the colonization of the pasture by the fungus was stable. HE staining and in situ hybridization showed, respectively, changes to the reproductive system of the females and confirmation of infection by the species M. anisopliae. Based on these results, it is possible to demonstrate the action of this entomopathogen on the tick and suggested that the field markers technique can be used and replicated to evaluate the action of the fungus M. anisoplie in females of R. microplus.