Abstract
Four cytochrome P450 cDNAs, CYP6AA7, CYP9J40, CYP9J34, and CYP9M10, were isolated from mosquitoes, Culex quinquefasciatus. The P450 gene expression and induction by permethrin were compared for three different mosquito populations bearing different resistance phenotypes, ranging from susceptible (S-Lab), through intermediate (HAmCqG0, the field parental population) to highly resistant (HAmCqG8, the 8th generation of permethrin selected offspring of HAmCqG0). A strong correlation was found for P450 gene expression with the levels of resistance and following permethrin selection at the larval stage of mosquitoes, with the highest expression levels identified in HAmCqG8, suggesting the importance of CYP6AA7, CYP9J40, CYP9J34, and CYP9M10 in the permethrin resistance of larva mosquitoes. Only CYP6AA7 showed a significant overexpression in HAmCqG8 adult mosquitoes. Other P450 genes had similar expression levels among the mosquito populations tested, suggesting different P450 genes may be involved in the response to insecticide pressure in different developmental stages. The expression of CYP6AA7, CYP9J34, and CYP9M10 was further induced by permethrin in resistant mosquitoes. Taken together, these results indicate that multiple P450 genes are up-regulated in insecticide resistant mosquitoes through both constitutive overexpression and induction mechanisms, thus increasing the overall expression levels of P450 genes.
Highlights
Cytochrome P450s have long been of particular interest because they are critical for the detoxification and/or activation of xenobiotics such as drugs, pesticides, plant toxins, chemical carcinogens and mutagens
A significant characteristic of insect P450s associated with the enhanced metabolic detoxification of insecticides is the increase in the levels of P450 proteins and P450 activity that results from constitutive overexpression of P450 genes in insecticide resistant insects, which has been implicated in the development of resistance to insecticides [3,6,7,8,9,10,11] and tolerance to plant toxins [12,13]
Results of the pilot experiment, in which the induction of P450s in both resistant HAmCq mosquito populations showed a clear concentration (LC50)- and time (24 h)dependent response. Based on these preliminary results, two different permethrin treatment experiments were conducted: 1),1000 late 3rd instar larvae of each of the three Culex mosquito strains were treated with permethrin at their respective LC50 concentrations (0.007ppm, 0.07ppm, and 20ppm for the S-Lab, HAmCqG0, and HAmCqG8 strains, respectively) and the expression of the P450 genes were examined 12, 24, 48, and 72h after the permethrin treatment; and 2) mosquito strains were treated with their corresponding LC10, LC50 and LC90 concentrations of permethrin (Table 1) and the surviving mosquitoes were collected for RNA extraction 24 h after permethrin challenge
Summary
Cytochrome P450s have long been of particular interest because they are critical for the detoxification and/or activation of xenobiotics such as drugs, pesticides, plant toxins, chemical carcinogens and mutagens. A significant characteristic of insect P450s associated with the enhanced metabolic detoxification of insecticides is the increase in the levels of P450 proteins and P450 activity that results from constitutive overexpression of P450 genes in insecticide resistant insects, which has been implicated in the development of resistance to insecticides [3,6,7,8,9,10,11] and tolerance to plant toxins [12,13] Another feature of some insect P450 genes is that their expression can be induced by both exogenous and endogenous compounds [3], a phenomenon known as induction
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