Submitted to: Journal of Medical Entomology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 5/3/2002
Publication Date: 7/31/2002
Citation: GUERRERO, F., LI, A.Y., HERNANDEZ, R. MOLECULAR DIAGNOSIS OF PYRETHROID RESISTANCE IN MEXICAN STRAINS OF BOOPHILUS MICROPLUS (ACARI: IXODIDAE). JOURNAL OF MEDICAL ENTOMOLOGY. 2002. v. 39. p. 770-776. Interpretive Summary: We have utilized two rapid diagnostic polymerase chain reaction (PCR) assays to study individual larvae from two pyrethroid resistant strains of Boophilus microplus. One strain, designated SF, appears to be resistant due to insensitivity of the target site, the sodium channel, to the effects of pyrethroids. The second strain, designated Cz, appears to be resistant due to the presence of an enzyme capable of degrading pyrethroids. Groups of larvae from each of the two strains were treated with various doses of permethrin and larvae which survived the treatment were separated from those which were killed. DNA was isolated from individual larvae from both the surviving and killed ticks of both strains and tested for the presence or absence of two mutations which were suspected of being involved in the resistance mechanism. One mutation is in a gene which codes for a sodium channel protein which regulates the flow of sodium ions into the cell. A similar sodium channel gene mutation has been shown to be responsible for pyrethroid resistance in a number of insect species. The second mutation is in an esterase, CzEst9, a type of protein capable of binding and degrading various types of pesticides. The sodium channel gene mutation preferentially associated with permethrin exposure survivors while the esterase mutation was found in slightly elevated numbers in surviving larvae. Thus, the sodium channel gene mutation likely plays a significant role in permethrin resistance while the mutation in the permethrin- degrading esterase plays a lesser role. The reason the Cz strain is resistant to permethrin appears more likely due to the Cz strain
Technical Abstract: PCR diagnostic assays were utilized to identify possible resistance- associated roles of two amino acid substitutions found in pyrethroid resistance-associated genes of Boophilus microplus. Individual larvae from the San Felipe (SF) target site resistant strain and the Coatzoacoalcos (Cz) metabolic resistant strain were separated into resistant and susceptible groups by larval packet bioassays and analyzed by PCR. A Phe to Ile amino acid mutation in the sodium channel gene S6 transmembrane segment of domain III was found to have a close association with survival of acaricide treatments containing as high as 30% permethrin. As the permethrin dose was increased, an increase was seen in the proportion of surviving larvae which possessed two mutated sodium channel alleles. An Asp to Asn amino acid substitution, originally found in high allele frequency in alleles of the CzEst9 esterase of the Cz strain, appeared to provide some resistance to permethrin, although the presence of the mutation did not associate with resistance in the dose response fashion seen with the sodium channel amino acid mutation. Resistance provided by CzEst9 might be more dependent on concentration of CzEst9 more so than the presence of a mutated allele.