Simone Quinelato

Susceptibility of different populations of ticks to entomopathogenic fungi

This study aimed to evaluate the in vitro effect of the entomopathogenic fungi Metarhizium anisopliae sensu lato (s.l.) and Beauveria bassiana sensu lato (s.l.) on two distinct populations of Rhipicephalus microplus, from two different experimental farms. Bioassays were performed with engorged females, eggs and larvae. Fungal infection was evaluated based on biological parameters of treated engorged females, percentage of hatch from treated eggs, and percentage of mortality and mean lethal time (LT90) of treated larvae. When the treatments were compared between the two populations, there were significant differences in the following parameters: pre-oviposition period, hatching period and egg production index. Moreover, the results showed that B. bassiana s.l., isolate Bb 986, was more virulent than M. anisopliae s.l., isolate 959, for engorged females, showing a control percentage of 49%. In the bioassay with eggs, the hatching percentage ranged from 3.1% to 49.5% in one population and from 3.4% to 42.7% in the other, with no significant difference between the two populations. In the bioassay with unfed larvae, the mortality percentage ranged from 91.8% to 98.7% in one population and from 71.0% to 94.0% in the other. The LT(90) varied from 19.52 to 27.51 days in one of the populations and 22.89 to 37.31 days in the other. These results suggest that populations of R. microplus show distinct variation in their susceptibility to B. bassiana s.l. and M. anisopliae s.l.

Effect of oil-based formulations of acaripathogenic fungi to control Rhipicephalus microplus ticks under laboratory conditions

The formulations of acaripathogenic fungi to control ticks have been widely studied. The present study evaluated the efficacy of oil-based formulations of Metarhizium anisopliae sensu lato (s.l.), isolate Ma 959, and Beauveria bassiana, isolate Bb 986, on different Rhipicephalus microplus stages, comparing the efficacy between aqueous suspensions and 10, 15 and 20% mineral oil formulations. Twelve groups were formed: one aqueous control group; three mineral oil control groups, at 10, 15 or 20%; two aqueous fungal suspensions of M. anisopliae s.l. or B. bassiana; and three formulations of M. anisopliae (s.l.) or B. bassiana containing 10, 15, and 20% mineral oil. To prepare aqueous suspensions and oily formulations, fungal isolates were cultivated on rice grains in polypropylene bags. The conidial suspensions and formulations had a concentration of 10(8)conidia/mL. Bioassays were repeated twice. After treatment, the following biological parameters of engorged females were evaluated: hatching percentage, egg production index, nutritional index, and percentage of tick control. The following parameters were evaluated in the bioassays with eggs: period of incubation, period of hatch, and hatching percentage. Mortality was evaluated in bioassays with larvae. M. anisopliae s.l. and B. bassiana oil-based formulations were more effective than aqueous suspensions against R. microplus eggs, larvae and engorged females, however, there was no significant difference between the three oil concentrations used. M. anisopliae s.l. and B. bassiana formulated in mineral oil reached 93.69% and 21.67% efficacy, respectively, while M. anisopliae s.l. and B. bassiana aqueous suspensions attained 18.70% and 1.72% efficacy, respectively. M. anisopliae s.l. oil-based formulations caused significant effects in all biological parameters of engorged females while B. bassiana oil-based formulations modified significantly the nutritional index only. Eggs treated with M. anisopliae s.l. and B. bassiana oil-based formulations showed hatching rates that decreased 102.5 and 3.65 times, respectively. In the bioassay with larvae, M. anisopliae s.l. oil-based formulations caused nearly 100% mortality five days after treatment, while larva treated with B. bassiana oil-based formulations reached 100% mortality at day 20 after treatment. Larva from oil-based control groups showed mortality at day 15 after treatment, which indicated a possible toxic effect of the oil for this R. microplus stage. The results showed that the fungal mineral oil formulations tested were more effective than the aqueous suspension. Oil-based formulations at 10, 15 and 20% enhanced the activity of M. anisopliae s.l. Ma 959, and B. bassiana Bb 986, isolates against R. microplus eggs, larvae, and engorged females tick. Mineral oil was effective as an adjuvant in formulations of M. anisopliae s.l., Ma 959, and B. bassiana, Bb 986, for the control of R. microplus under laboratory conditions.

The ecology of horse cyathostomin infective larvae (Nematoda-Cyathostominae) in tropical southeast Brazil

Experimental studies about the recovery, survival and migration to pasture of cyathostomin infective larvae (L(3)) from fresh feces depositions were conducted from February 2005 to March 2007 in a tropical region of southeast Brazil. Grass and feces were collected weekly at 8 a.m., 1 and 5 p.m. and processed by the Baermann technique. Multivariate analysis (principal components method) showed the influence of time and environmental variables on the number of infective larvae recovered from the feces and pasture. In the rainy period (October-March), more infective larvae were recovered on the feces and grass apex. In contrast, in the dry period (April-September), the recovery was higher only on the grass base, as well as the L(3) survival on feces and grass. More larvae were recovered at 8 a.m., except from the grass apex, where the highest recovery was at 1 p.m. Few studies investigating the seasonal transmission of equine cyathostomin have been conducted in South American tropical climates. These results demonstrate that in tropical conditions L(3) are available on feces and pasture throughout the year. Knowledge of climatic influences on the development and survival of L(3) is crucial to designing integrated parasite control programs that provide effective protection while slowing the development of anthelmintic resistance.

Virulence potential of Metarhizium anisopliae s.l. isolates on Rhipicephalus (Boophilus) microplus larvae.

The use of entomopathogenic fungi to control arthropods has been reported worldwide for decades. The aim of the present study was to evaluate the virulence of 30 Metarhizium anisopliae s.l. Brazilian isolates from different geographical regions, hosts or substrates on the larvae of Rhipicephalus (Boophilus) microplus ticks under in vitro conditions to the selection of virulent isolates in order to be further used in biological control programs. The current study confirmed the lethal action of M. anisopliae s.l. isolates on R. (B.) microplus larvae with different mortality levels, usually directly proportional to the conidia concentration. No relationship was found between the origin of the isolate and its virulence potential or between the virulence potential and conidia production. Three isolates (CG 37, CG 384 and IBCB 481) caused a high percentage of larval mortality, reaching LC(50) at 10(6) conidia ml(-1), thus requiring a lower conidia concentration to cause an approximately 100% larval mortality. The results of this study suggest that these three isolates are the most promising for use in programs aimed at microbial control in the field.

Commercial formulation of Metarhizium anisopliae for the control of Rhipicephalus microplus in a pen study

The present study evaluated, for the first time, the effect of the commercial formulation Metarril(®) SP Organic of Metarhizium anisopliae plus 10% mineral oil to control Rhipicephalus microplus in a pen study. Three groups were formed with six animals each: the first group was exposed to Metarril(®) plus 10% mineral oil and 1% Tween 80; the second group was exposed to sterile distilled water, mineral oil and Tween 80 (oil control group); and the third group received no treatment (control group). The fungal formulation contained 1 × 10(8)conidiaml(-1). Each animal was sprayed with 3L of formulation. Fallen ticks were counted daily and a sample of 20 engorged females per day was incubated for assessment of biological parameters. Throughout the study period, Metarril(®) oil-based formulation showed an efficacy ranging from 19.20% to 67.39% in comparison with the control group; and from 8.18% to 61.38% in comparison with the oil control group. The average efficacy of Metarril(®) oil-based formulation was 47.74% and 40.89% in comparison with control and oil control groups, respectively. Changes in the biological parameters of engorged R. microplus females were observed in the first three days after treatment, with a significant reduction in hatching percentage and egg production index. We concluded that Metarril(®) SP Organic plus 10% mineral oil was efficient against R. microplus in pen studies. However, further in vivo studies are required to increase the efficacy and to establish a protocol for the use of this product in the field against the cattle tick.

Seasonal dynamics of cyathostomin (Nematoda – Cyathostominae) infective larvae in Brachiaria humidicola grass in tropical southeast Brazil

The ecology of cyathostomin larvae was evaluated in different seasons, from July 2007 to June 2008, in the municipality of Seropédica, Rio de Janeiro state, southeastern Brazil. Samples of feces and grass were collected every 15 days at 8 AM and 5 PM and the infective larvae were recovered by the Baermann technique. Leaves of the grass Brachiaria humidicola were cut to 20 cm, which is the length containing most of the larvae. The highest number of larvae was recorded at 8 AM the winter (8300 L(3)kg(-1)dm) and spring (5300 L(3)kg(-1)dm). These results demonstrate that climate conditions can affect the recovery of larvae and that rain and temperature contributed to the migration and survival of the larvae, which were available throughout the year in the study area.

In vitro pathogenicity of different Metarhizium anisopliae s.l. isolates in oil formulations against Rhipicephalus microplus

ABSTRACT The present study aimed to evaluate the in vitro pathogenicity of the Metarhizium anisopliae s.l. CG 112, CG 347, CG 32, CG 148 and CG 629 isolates formulated in vegetable or mineral oil against Rhipicephalus microplus. The bioassays were performed with R. microplus engorged females. The ticks were immersed for three minutes in oil-based formulations containing 10% oil (mineral or vegetable). The effects of the different fungal isolates were evaluated analyzing the females’ reproductive parameters (the total weight of the egg mass, hatching percentage, egg production index and nutritional index). The present study showed that all of the tested isolates and both oil formulations (vegetable or mineral oil) changed the biological parameters of the R. microplus females. However, the mineral oil formulation was more effective than the vegetable oil formulation, as the former showed a higher tick control percentage. It was concluded that there was variation in the virulence among the different M. anisopliae s.l. isolates and between the different types of oil. Moreover, that either of the isolates GC 148 and GC 629 formulated in oil confers the good potential for controlling R. microplus engorged females.