**3. Molecular basis of pesticide resistance**

In general, acaricide resistance can be the result of increased metabolic detoxication or target site modification [4]. Metabolic detoxication is frequently the result of increased enzymatic activity by isozymes encoded by multigene families such as cytochrome P450, glutathione-S- transferase, and carboxylesterase [16, 17]; all these enzymes hydrolyze or sequester different kinds of pesticides. Exposure to a pesticide can exert enough pressure to select an enzymatic system or a specific isozyme within each family. Esterase isozyme overexpression is generally accepted as a mechanism involved in OP resistance. However, in the Coatzacoalcos laboratory strain of *R*. *microplus*, designated as such to reflect the name of the village in Mexico where the original tick population sample was obtained, metabolic detoxication has been identified by its efficient esterase activity resulting from enzyme overexpression as a resistance mechanism for permethrin that belongs to the pyrethroid chemical class of pesticides. This strain has been toxicologically characterized using the larval packet test (LPT) [18], which helped to elucidate the esterase-based mechanism of resistance to permethrin. The *R*. *microplus* Coatzacoalcos strain exhibits a significant enhanced capacity to hydrolyze permethrin as well as an increased esterase activity. This suggests an esterase based metabolic mechanism as a main component of permethrin resistance [19]. The esterase gene responsible for permethrin resistance was identified and named *CzEST9*. It is also known that the overexpression mechanism of this isozyme is the result of *CzEST9* duplication in the Coatzacoalcos strain that leads to metabolic detoxication through the overexpression of esterase 9 activity in *R. microplus* [20]. The sequence of *CzEST9* gene has been determined and the recombinant product yielded a 62.8 kDa protein [19]. Since the Coatzacoalcos strain does not include the *Kdr* variation in the sodium channel gene found in other Mexican strains of *R*. *microplus*, it is suggested that there are two inde‐ pendent mechanisms of acaricide resistance to pyrethroids. However, common mechanisms of acaricide resistance to pyrethroids in Mexico apparently involve the presence of sequence variation in the sodium channel gene [21].

The sodium channel is the known target site of pyrethroids. Sequence variation in the sodium channel prevents pyrethroids from attaching to the target site due to an alteration in the sodium channel stereochemical structure. For this reason, the process is described as target site modification mechanism, or *Kdr*-type resistance (Knock down resistance). This is one of the mechanisms of pesticide resistance in insects that is better understood.

Two important allele variants occurring in the sodium channel gene associated with pyreth‐ roid resistance in the cattle tick *R. microplus* are the variation occurring in domain III segment 6 (III-S6) [22] and the variation occurring at the bridge joining segments 4–5 in domain II (II-S4-5) [23]. The former is a Phe-Ile substitution produced by a nucleotide variation at domain III-S6 that was first reported in Mexican tick strains. Its role and contribution to pyrethroid resistance has been confirmed [21]. The other is a Le-Ile substitution found thus far only in Australian tick strains; this variation is very similar to a variation found in the crop insect pest *Bemicia tabaci* [23].

Findings on the diversity of allele variants occurring in the sodium channel gene associated with pyrethroid resistance in the cattle tick *R. microplus* have been confirmed by experiments based on differences in melting temperatures (Tm) of sodium channel allele specific gene fragments obtained with single larvae DNA from México and Australia. These experiments revealed that substitution III-S6 (Phe-Ile) only occurs in Mexican tick populations whereas substitution II-S4-5 (Le-Ile) only occurs in Australian tick strains [24]. The information available suggests that there are at least two different and independent mechanisms involved based on the different amino acid substitutions (Phe-Ile and Le-Ile) residing in different positions of the sodium channel protein sequence.
