Liverpool, United Kingdom
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Khajehali J.,Ghent University | van Leeuwen T.,Ghent University | Grispou M.,Greek National Agricultural Research Foundation | Morou E.,Agricultural University of Athens | And 7 more authors.
Pest Management Science | Year: 2010

Backround: In Tetranychus urticae Koch, acetylcholinesterase insensitivity is often involved in organophosphate (OP) and carbamate (CARB) resistance. By combining toxicological, biochemical and molecular data from three reference laboratory and three OP selected strains (OP strains), the AChE1 mutations associated with resistance in T. urticae were characterised. Results: The resistance ratios of the OP strains varied from 9 to 43 for pirimiphos-methyl, from 78 to 586 for chlorpyrifos, from 8 to 333 for methomyl and from 137 to 4164 for dimethoate. The insecticide concentration needed to inhibit 50% of the AChE1 activity was, in the OP strains, at least 2.7, 55, 58 and 31 times higher for the OP pirimiphos-methyl, chlorpyrifos oxon, paraoxon and omethoate respectively, and 87 times higher for the CARB carbaryl. By comparing the AChE1 sequence, four amino acid substitutions were detected in the OP strains: (1) F331W (Torpedo numbering) in all the three OP strains; (2) T280A found in the three OP strains but not in all clones; (3) G328A, found in two OP strains; (4) A201S found in only one OP strain. Conclusions: Four AChE1 mutations were found in resistant strains of T. urticae, and three of them, F331W, G328A and A201S, are possibly involved in resistance to OP and CARB insecticides. Among them, F331W is probably the most important and the most common in T. urticae. It can be easily detected by the diagnostic PCR-RLFP assay developed in this study. © 2009 Society of Chemical Industry.


Dowd A.J.,Vector Research | Morou E.,Vector Research | Steven A.,Vector Research | Ismail H.M.,Vector Research | And 5 more authors.
International Journal of Environmental Analytical Chemistry | Year: 2010

Recombinant glutathione-S-transferases (GSTs) can be used as analytical tools for the development of simple insecticide quantification assays. This assay explores the ability of pyrethroids to promote inhibition of the GST-catalysing 1-chloro-2,4-dinitrobenzene (CDNB)/glutathione (GSH) conjugation reaction. The sensing scheme is based on the pH change occurring in a weak buffer system by the GST reaction, which is measured using a spectrophotometer and the dye indicator bromothymol blue (616 nm). Practical use depends on the recognition affinity of the GST for insecticides, inhibition kinetics, enzyme stability and compatibility with the detection assay. In this study we compared the recombinant GSTs AgGSTD1-6 and AdGSTD1-1 from the mosquito vectors Anopheles gambiae and Anopheles dirus, respectively, with high affinity for pyrethroids, for their suitability for detecting pyrethroids in vector disease control programmes. The results showed that AgGSTD1-6 was the most suitable enzyme with the best structural stability at higher temperatures (Tm57°C) and pH optima in the alkaline range (pH 7.7). Using the AgGSTD1-6, we subsequently developed a pH- change colourimetric assay for detecting pyrethroids. Linear calibration curves were obtained for deltamethrin (R20.99) with useful concentration ranges of 0-50 μgmL-1. The effect of temperature in the range 25-40°C on the pyrethroid quantification assay was negligible. The assay was validated with extracts from insecticide sprayed surfaces and found to be reproducible and reliable compared with the standard reverse-phase high performance liquid chromatography (Rp-HPLC) method. The potential of the assay for monitoring insecticide residues in the frame of insecticide based malaria control interventions is discussed. © 2010 Taylor & Francis.

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