Strategic control of the cattle tick Rhipicephlaus microplus applied to rotational and silvopastoral grazing systems in subtropical areas.

This work evaluated if strategic control based on no more than three or four annual treatments is useful to control Rhipicephalus microplus infestations on cattle when it is applied to intensive rotational grazing and silvopastoral systems with high stocking rates in subtropical areas. In the intensive rotational grazing system, three annual treatments with chemical acaricides were applied on cattle in two different schemes: between spring and early summer and from late winter and late spring. Strategic control based on three treatments with chemical acaricides from late winter to late spring plus an additional fourth treatment in summer was tested in the silvopastoral system. In the intensive rotational grazing systems, the control schemes allow to reach a significant reduction in the tick load on cattle considering a time interval from spring to autumn. However, the efficacy levels were not high enough in some specific moments, namely, the tick counts of summer and autumn (there were not significant differences between treated and control groups). The scheme evaluated in the silvopastoral grazing system yielded better results than those tested for the intensive rotational system, because significant differences in tick load between treated and control groups were observed in all post-treatment counts and when the analysis was performed for the whole study period. However, values of efficacy in the count-by-count comparison were disparate, ranging from 64.1 to 99.7. Although the efficacy values obtained in the silvopastoral system were better than those of the rotational grazing systems, the total tick load on treated cattle in autumn was not low enough (mean abundance values 25.14 and 38.14). Ticks were more evenly distributed among hosts in late summer and autumn than in spring or early summer, where few hosts carry most of the ticks. Some management strategies as intensive rotational systems or silvopastoral structures can lead to a more efficient forage use, but they imply greater tick challenge than in extensive grazing systems. In these situations, the schemes of strategic control bases on three or four annual treatments should be complemented with additional tactical treatments in late summer or autumn.

A comparison of two in vitro bioassays to detect resistance of the cattle tick Rhipicephalus microplus to fipronil.

The aim of this work is to compare the sensitivity of two in vitro bioassays to detect resistant to fipronil in Argentinean populations of the cattle tick Rhipicephalus microplus. Two different larval bioassays prepared with technical grade (97%) fipronil were compared: larval immersion test (LIT) and larval packet test (LPT). Seven strains from different provinces were treated with both assays. Colonia Tabay, Colonia Benítez, Intiyaco and Quimili strains were considered resistant in both LIT and LPT bioassays. The 95% confidence intervals (IC95) for lethal concentration 50% (LC50) did not overlap with the susceptible reference strain (SRS) and all the values of RR50 obtained were higher than 2. Garabato and Federal strains were considered as susceptible for both techniques because the IC95 for the LC50 overlapped with those of the SRS and the RR50 values were lesser than 2. An ambiguous situation occurs with Reconquista strain. This strain was considered as susceptible with LPT and with incipient resistant after LIT trial. The analysis of the results indicates that both LIT and LPT trials have enough sensibility to differentiate resistant and susceptible strains, but LIT was more sensitive than LPT when the resistance is incipient.

Migration of Rhipicephalus microplus ticks among cattle.

The aim of this work was to quantitatively analyse the magnitude of the migration of Rhipicephalus microplus ticks among cattle under field conditions, in groups of bovines with different stocking rates. The role of this phenomenon in the epidemiology of Anaplasma marginale has been discussed. Cattle naturally infested with R. microplus (donors) and cattle non-infested with R. microplus (recipients) were clustered for 13 days into two groups with low and high stocking rates (G1 and G2 respectively). The initial prevalence of infestation (infested cattle / total cattle) was 20% in both groups. Adult migratory ticks from donor to recipient cattle were recorded by examination of the recipient cattle on days 2, 6, 9, and 13. The tick infestation rate, calculated using the Kaplan-Meier survival test, was higher in G2 (p = 0.05). The cumulative incidence on day 13 was 25% in G1 and 65% in G2, with no significant differences. The results demonstrate that migration of adults of R. microplus under field conditions occurs, in accordance with previous studies, and that its magnitude is associated with the stocking rate. These results highlight the relevance of R. microplus in the epidemiology of A. marginale through its role as a vector in the intrastadial transmission of this pathogen of cattle.

Resistance of the cattle tick Rhipicephalus (Boophilus) microplus to fluazuron in Argentina.

The aim of this work is to report the presence of resistance to fluazuron in a population of Rhipicephalus microplus in Argentina. The evidence was obtained from field and in vitro trials. In the field trial, cattle infested with ticks was treated with two commercial formulations of fluazuron. The in vitro trial (adult immersion test, AIT) was performed by using technical grade fluazuron. In the field trial, there were no significant differences between the treated and control groups between days 2 and 34 post-treatment. The only exceptions (treated group I in day 14 post-treatment, treated group II in days 23 and 29 post-treatment) had a significantly lower tick load than the untreated group, but the efficacy was not higher than 70%. Viable engorged females were collected on both groups of treated bovines in all counts, and the production of viable larvae was not precluded with the application of the two commercial formulations of fluazuron evaluated in this study. The results obtained with the in vitro assay (AIT) also indicate that the R. microplus population tested in this work has a higher level of resistance to fluazuron than another susceptible field strain. The integrated analysis of the field and in vitro trials clearly reveals the emergence of resistance to fluazuron in a R. microplus population from Argentina. This diagnosis of resistance does not imply that the fluazuron has lost its functionality at a regional scale, but it highlights the need to establish control strategies that minimize the use of this drug in order to preserve its functionality as an acaricide.

Alternative applications of the strategic control against the cattle tick Rhipicephalus (Boophilus) microplus in a subtropical area.

Although different evaluations on the efficacy of the strategic control against Rhipicephalus microplus have been performed, the effects of successive applications of these schemes on the abundance of cattle ticks have not been evaluated. The aim of this work was to analyse the long-term effect of strategic applications of chemical acaricides on the R. microplus infestation in cattle in a subtropical area. These schemes are based on the application of three annual treatments between late winter and late spring. Additionally, a trial to evaluate the efficacy of the strategic control by deferring the first treatment from late winter to spring and the third treatment from late spring to summer was also carried out. The efficacy of the strategic control applied on 3 consecutive years was significant. The tick infestation in the treated group always remained at low levels, because mean number of ticks was almost never higher than 20. Regarding the trial where the third application of acaricide was deferred from spring to summer, and the first one from late winter to spring, the differences between treated and control group were significant in all post-treatment counts. The results of this study add evidence that support the sustainability of the strategic control in subtropical areas where the population dynamics of R. microplus is characterized by a well-marked seasonal pattern. Three relevant aspects were determined: (i) the feasibility and efficacy of successive applications of the strategic control in consecutive years; (ii) the time window to start the sequence of treatments is from late winter to mid-spring; (iii) it is achievable deferring the last treatment from late spring to summer if the tick infestation levels on cattle are low enough to allow it.

A Trypanosoma species detected in Rhipicephalus (Boophilus) microplus ticks from Argentina.

Specimens of a Trypanosoma sp. were found in a haemolymph sample of Rhipicephalus microplus from Argentina. Polymerase chain reaction (PCR) was done targeting the SSU rRNA gene of Trypanosoma spp. and a fragment of 2300 base pairs (bp) was amplified, subsequently a phylogenetic analysis was conducted, based on an alignment of 905 bp, containing the sequence of the Argentina isolate and sequences of different Trypanosoma species retrieved from GenBank. Phylogenetic analysis revealed that this trypanosome is not related to Trypanosoma theileri as was previously thought, instead the strain of Trypanosoma detected in this study can be provisionally determined as belonging to the recently described organism Trypanosoma rhipicephalis. Furthermore, phylogenetic analysis performed in this work revealed that T. rhipicephalis belongs to a novel clade of tick-related trypanosomes, most with limited genetic data, for which essential aspects of both the vertebrate and invertebrate life cycles are lacking. The lack of basic information restricts the inferences that can be done from the present finding and, in addition, points out a clear knowledge gap in the biology of this group of trypanosomes.

In vitro diagnosis of resistance of the cattle tick Rhipicephalus (Boophilus) microplus to fipronil in Argentina.

The aim of this work was to adapt and apply the in vitro larval bioassay packet test (LPT) to diagnose resistance of Rhipicephalus microplus ticks to fipronil in Argentina. Ten serial dilutions of fipronil were included in the LPT bioassay. Seven populations of R. microplus from six Argentinean Provinces were evaluated. Samples from Colonia Benítez, Quimilí and Intiyaco were unambiguously classified in the category 'resistant'. In all these cases the 95% confidence intervals did not overlap with that of the susceptible reference strain and the slope values were lower than the slope value obtained for the susceptible reference strain. The resistance ratio (RR50) values of the populations from Colonia Benítez I and II, Quimilí and Intiyaco were 2.23, 3.49, 6.84 and 3.39, respectively. Populations from El Colorado I and II, Acambuco, Vera and Feliciano were classified as 'susceptible' because the 95% confidence intervals overlapped with that of the susceptible reference strain and the RR50 values never exceeded 1.32. The results of these LPT bioassays indicate the presence of fipronil resistant R. microplus populations in Argentina.

Resistance of the cattle tick Rhipicephalus (Boophilus) microplus to ivermectin in Argentina.

Resistance to ivermectin in populations of the cattle tick Rhipicephalus microplus in Argentina was diagnosed in this work. The in vitro larval immersion test (LIT) was used to determine quantitatively the levels of resistance to ivermectin in different populations of R. microplus. Additionally, field trials to control natural infestations of R. microplus on cattle with a commercial formulation of ivermectin 3.15% were carried and jointly analyzed with the in vitro assays. The phenotypic response of the populations analyzed was not uniform. Five of them were classified as susceptible, four populations as resistant, and one in the category "incipient resistance". Regarding the field trials, the therapeutic efficacy in a population classified with LIT as susceptible achieved values higher than 94% two weeks after treatment, and no reproductively viable females were observed after the second day post-treatment. Conversely, the values of efficacy percentage in a population (named as "San Martín") classified with LIT in the category "incipient resistance" never exceeded the 70.8%, and engorged females were collected in practically all counts. The population "San Martín" was classified in the category "incipient resistant" with LIT analysis, but the field trial unambiguously shows that this tick population is resistant. The comparison of the results obtained with LIT in vitro assays and through field trials shows that biased estimations of resistance levels may occur when resistance ratios (RR) values are ≤2, and additional field efficacy trials could be needed to know with precision the status of the tick populations evaluated.

Relationship between pharmacokinetics of ivermectin (3.15%) and its efficacy to control the infestation with the tick Rhipicephalus (Boophilus) microplus in cattle.

This study aimed to determine the relationship between the variation in plasma concentration of ivermectin 3.15% over time and its efficacy against the cattle tick Rhipicephalus (Boophilus) microplus. In addition, a trial was conducted to infer if the application of successive treatments with ivermectin 3.15% could affect its accumulation in cattle. A noticeable variation of ivermectin plasma concentration was observed among the treated heifers. However, these differences did not have a significant effect onthe therapeutic efficacy of the treatment at the end of the trial. No significant differences were observed in the levels of tick infestations between heifers of the treated group; moreover, no significant correlation was detected between the plasma AUC0-21 of ivermectin 3.15% and the cumulative number of ticks of each heifer. Levels of therapeutic efficacy higher than 80% were observed only from day 7 post-treatment, when levels of ivermectin concentration were higher than 8 ng/ml. The lowest values of therapeutic efficacy were observed during the first and the second days post-treatment, when plasma concentrations of ivermectin 3.15% were lower than 8 ng/ml. Viable engorged females were collected from the heifers belonging to the treated group from days 1-5 post-treatment. There was a significant accumulation of the drug after the second dose of ivermectin 3.15%. Ivermectin concentrations in fat biopsies were 366 ng/g (51 days after the first treatment), 275 ng/g (51 days after the second treatment) and 15 ng/g (64 days after the second treatment). These results suggest that applications of successive treatments with ivermectin 3.15% might increase its accumulation in cattle tissues, extending the withdrawal period indicated for the commercial formulation.