Following this procedure, five groups were formed, three with five animals each and two with four. Each group was maintained in a separate 3 m × 6 m stall that
had been built entirely of cement and was partially covered. The animals were fed corn silage supplemented with protein concentrates and received water “ad libitum. One of the following treatments was allocated to each of the five groups: 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 × 108 conidia (T BASS), association of the concentrate of M. azedarach Z-VAD-FMK concentration at 0.25% with B. bassiana at 2.4 × 108 conidia (T AZED 0.25% + BASS), and the control (untreated). Each animal was sprayed with
5 L of water or the testing solution, using a 20 L costal bomb, and each mixture was prepared with tap water, at the time of use. From days −3 to +20, the female ticks measuring between 4.5 and 8.0 mm and attached to the right side of each bovine were collected and counted. This number was doubled to provide an estimate of the total burden for each animal. Per day, from the total female ticks of each group, the 20 largest were selected, collectively weighed and incubated (27 °C and RH ≥ 80%). After BYL719 ic50 20 days of incubation, the eggs selected and weighed to obtain the index of conversion in eggs (ICO) = [(weight of eggs/weight of the females) × 100]. The eggs were incubated and after 25 days evaluated to determine the hatchability. The index of effectiveness of the treatment was calculated in accordance with the following formulae (Holdsworth et al., 2006): Daily percentage control = 100 − daily percentage tick survival (DPTS): DPTS=Ticks counted in treated groupNumber of ticks expected in treated group if left untreated (ADEQ)×100 ADEQ=Total pre-treatment count in treated groupTotal pre-treatment count in control group×Daily control count The total burden, log
(x + 1), the index of effectiveness, and the reproductive parameters were subjected to an analysis of variance (ANOVA) and differences among means were determined by Tukey’s all pairwise comparison (P < 0.05). Comparisons were made between the three following periods (in days): 1st to 6th, 7th to 13th, and 14th to 20th, which corresponded to the modal periods of larvae, nymphs, and adults, respectively. The T AZED 0.25% + BASS either treatment, which had the two compounds, produced better results in the control of R. microplus than any isolated treatment, indicating a compatibility or perhaps a synergy between M. azedarach and B. bassiana. Fewer engorged females were observed at all intervals as compared to the control group, indicating greater activity against all stages of the tick. On the other hand, the highest concentration of M. azedarach (T AZED 0.5%) worked mainly against adult and larval ticks, producing lower counts in the first and last intervals than in the control group.