RESUMO
Dendroctonus-bark beetles are natural components and key ecological agents of coniferous forests. They spend most of their lives under the bark, where they are exposed to highly toxic terpenes present in the oleoresin. Cytochrome P450 (CYP) is a multigene family involved in the detoxification of these compounds. It has been demonstrated that CYP6DE and CYP6DJ subfamilies hydroxylate monoterpenes, whose derivatives can act as pheromone synergist compounds or be pheromones themselves in these insects. Given the diversity and functional role of CYPs, we investigated whether these cytochromes have retained their function throughout the evolution of these insects. To test this hypothesis, we performed a Bayesian phylogenetic analysis to determine phylogenetic subgroups of cytochromes in these subfamilies. Subgroups were mapped and reconciled with the Dendroctonus phylogeny. Molecular docking analyses were performed with the cytochromes of each subgroup and enantiomers of α-pinene and ß-pinene, (+)-3-carene, ß-myrcene and R-(+)-limonene. In addition, functional divergence analysis was performed to identify critical amino acid sites that influence changes in catalytic site conformation and/or protein folding. Three and two phylogenetic subgroups were recovered for the CYP6DE and CYP6DJ subfamilies, respectively. Mapping and reconciliation analysis showed different gain and loss patterns for cytochromes of each subgroup. Functional predictions indicated that the cytochromes analyzed are able to hydroxylate all monoterpenes; however, they showed preferential affinities to different monoterpenes. Functional divergence analyses indicated that the CYP6DE subfamily has experimented type I and II divergence, whereas the CYP6DJ subfamily has evolved under strong functional constraints. Results suggest cytochromes of the CYP6DE subfamily evolve to reinforce their detoxifying capacity hydroxylating mainly α- and ß-pinene to (+) and (-)-trans-verbenol, being the negative enantiomer used as a pheromone by several Dendroctonus species; whereas cytochromes of the CYP6DJ subfamily appear to retain their original function related to the detoxification of these compounds.
RESUMO
The western pine beetle (WPB), Dendroctonus brevicomis LeConte, is a major mortality agent of pines in North America. A total of 706 adults of WPB from 81 geographical sites were analyzed with traditional and geometric morphometric methods to evaluate the variation of discrete and quantitative morphological characters with particular attention to the antenna, spermatheca, and seminal rod. Principal coordinates and canonical variate analyses supported three geographical groups in WPB: (1) West, from British Columbia to southern California along the Pacific coast, Idaho, and Montana; (2) East-SMOC, including Nevada, Utah, Colorado, Arizona, New Mexico, Texas, Chihuahua, and Durango; and (3) SMOR, including Coahuila, Nuevo Leon, and Tamaulipas. The pubescence length on the elytral declivity was a robust character for separating West specimens from the other groups. Additionally, the genitalia shape both female and male in dorsal view was a reliable character for discriminating among groups. Based on these results, which agree with genetic and chemical ecology evidence, we herein reinstate Dendroctonus barberi Hopkins (East-SMOC group) and remove it from synonymy with D. brevicomis (West group). Differences in the spermatheca and seminal rod shape of SMOR specimens suggest that these populations might be a different species from D. barberi and D. brevicomis.
