Development of permethrin resistance in Culex quinquefasciatus Say in Kuala Lumpur,Malaysia

The resistance status towards permethrin among the laboratory strain, the permethrin-selected strain and four field strains of Culex quinquefasciatus collected in Kuala Lumpur, Malaysia was determined using three standard laboratory methods: WHO larval bioassay, WHO adult bioassay and biochemical microplate assay. Cx. quinquefasciatus permethrin-selected strain larvae were the least susceptible to permethrin with a resistance ratio of 47.28-folds, whereas all field strain larvae of the same species were tolerant to permethrin with resistance ratios of more than 3-folds. In contrast, in adult stage, the permethrin exposed permethrin-selected strain (resistance ratio = 1.27) was found to be more susceptible to permethrin than all permethrin-exposed field strains (resistance ratios = 2.23–2.48). Complete mortalities for all strains of Cx. quinquefasciatus adults proved the effectiveness of the synergist; piperonyl butoxide (PBO). For the biochemical microplate assay, the reduction of the mean optical density of elevated oxidase activity of three field strains upon exposure to PBO confirmed the association between oxidase activity and permethrin tolerance. On the other hand, irregular patterns of the mean optical density of elevated oxidase activity in the laboratory strain, permethrin-selected strain and Jalan Fletcher strain illustrated the gene variation within these mosquito colonies as well as the involvement of other enzyme activities in the permethrin resistance occurred.     

Splice variant specific increase in Ca2+/calmodulin‐dependent protein kinase 1‐gamma mRNA expression in response to acute pyrethroid exposure

In mammals, pyrethroids are neurotoxicants that interfere with ion channel function in excitable neuronal membranes. Previous work demonstrated increases in the expression of Ca²⁺/calmodulin‐dependent protein kinase 1‐gamma (Camk1g) mRNA following acute deltamethrin and permethrin exposure. In the rat, this gene is expressed as two distinct splice variants, Camk1g1 and Camk1g2. The present study tests the hypothesis that changes in Camk1g mRNA expression in the rat following acute pyrethroid exposure are due to a specific increase in the Camk1g1 splice variant and not the Camk1g2 splice variant. Long‐Evans rats were acutely exposed to permethrin, deltamethrin, or corn oil vehicle. Frontal cortex was collected at 6 h postdosing. In addition, rats were exposed to permethrin (100 mg/kg) or deltamethrin (3 mg/kg), and frontal cortex was collected at 1, 3, 6, 9, 12, or 24 h along with time‐matched vehicle controls. Expression of Camk1g1 and Camk1g2 mRNA was measured by quantitative real‐time RT‐PCR and quantified using the 2^{‐Δ Δ C}T method. Dose‐dependent increases in Camk1g1 mRNA expression were observed for both pyrethroids at 6 h. In addition, a dose‐dependent increase in Camk1g2 was observed at 6 h although it was very small in magnitude. The increases in Camk1g1 expression for deltamethrin and permethrin peak between 3 and 6 h postexposure and returns to control levels by 9 h. There was no increase in CAMK1G1 protein as measured with Western blots. The present data demonstrate that pyrethroid‐induced changes in Camk1g are driven mainly by increased expression of the Camk1g1 splice variant. © 2010 Wiley Periodicals, Inc. J Biochem Mol Toxicol 24:174–186, 2010; Published online in Wiley InterScience ( www.interscience.wiley.com ). DOI 10.1002/jbt.20324     

Simultaneous determination of pyridostigmine bromide, N,N-diethyl-m-toluamide, permethrin, and their metabolites in rat plasma and urine by high-performance liquid chromatography

A rapid and simple method was developed for the separation and quantification of the anti nerve agent drug pyridostignmine bromide (PB; 3-dimethylaminocarbonyloxy-N-methyl pyridinium bromide) its metabolite N-methyl-3-hydroxypyridinium bromide, the insect repellent DEET (N,N-diethyl-m-toluamide), its metabolites m-toluamide and m-toluic acid, the insecticide permethrin (3-(2,2-dichloro-ethenyl)-2,2-dimethylcyclopropanecarboxylic acid(3-phenoxyphenyl)methylester), and two of its metabolites m-phenoxybenzyl alcohol, and m-phenoxybenzoic acid in rat plasma and urine. The method is based on using C₁₈ Sep-Pak^® cartridges for solid-phase extraction (SPE) and high-performance liquid chromatography (HPLC) with reversed-phase C₁₈ column, and gradient UV detection ranging between 208 and 230 nm. The compounds were separated using gradient of 1 to 99% acetonitrile in water (pH 3.20) at a flow-rate ranging between 0.5 and 1.7 ml/min in a period of 17 min. The retention times ranged from 5.7 to 14.5 min. The limits of detection were ranged between 20 and 100 ng/ml, while limits of quantitation were 150–200 ng/ml. Average percentage recovery of five spiked plasma samples were 51.4±10.6, 71.1±11.0, 82.3±6.7, 60.4±11.8, 63.6±10.1, 69.3±8.5, 68.3±12.0, 82.6±8.1, and from urine 55.9±9.8, 60.3±7.4, 77.9±9.1, 61.7±13.5, 68.6±8.9, 62.0±9.5, 72.9±9.1, and 72.1±8.0, for pyridostigmine bromide, DEET, permethrin, N-methyl-3-hydroxypyridinium bromide, m-toluamide, m-toluic acid, m-phenoxybenzyl alcohol and m-phenoxybenzoic acid, respectively. The relationship between peak areas and concentration was linear over the range between 100 and 5000 ng/ml. This method was applied to analyze the above chemicals and metabolites following their administration in rats.     

Evaluation of permethrin-impregnated mosquito-nets against mosquitoes in China

In Yishan County, Guangxi Province, China, where malaria and Brugian filariasis are transmitted by vectors belonging to the Anopheles hyrcanus Pallas group, a study was conducted from June to November 1986 to evaluate the effectiveness of permethrin-impregnated mosquito bednets to reduce man/vector contact. In three experimental villages a total of 246 mosquito nets were treated with permethrin EC at a dosage of 0.5 g/m2. Control bioassays (1 h exposure) of treated nets gave up to 100% mortality of Armigeres subalbatus (Coquillett), Anopheles sinensis Wiedemann and Culex tritaeniorhynchus Giles for up to 19 weeks. During 5 months evaluation in the villages, permethrin-impregnated mosquito-nets reduced the number of mosquitoes found inside nets by 99.3% for Culex quinquefasciatus Say, by 94.4% for Anopheles sinensis Wiedemann and by 75% for Anopheles lesteri anthropophagus Xu & Feng. This measure could thus be considered an appropriate means of controlling vectors in villages.     

Olive leaf extract modulates permethrin induced genetic and oxidative damage in rats

Permethrin is a common synthetic chemical, widely used as an insecticide in agriculture and other domestic applications. The previous reports indicated that permethrin is a highly toxic synthetic pyrethroid pesticide to human and environmental health. Therefore, the present experiment was undertaken to determine the effectiveness of olive leaf extract in modulating the permethrin induced genotoxic and oxidative damage in rats. The animals used were broadly divided into four (A, B, C and D) experimental groups. Group A rats served as control animals and received distilled water intraperitoneally (n = 5). Groups B and C rats received intraperitoneal injections of permethrin (60 mg kg⁻¹ b.w) and olive leaf extract (500 mg kg⁻¹ b.w), respectively. Group D rats received permethrin (60 mg kg⁻¹ b.w) plus olive leaf extract (500 mg kg⁻¹ b.w). Rats were orally administered their respective feed daily for 21 days. At the end of the experiment rats were anesthetized and serum and bone marrow cell samples were obtained. Genotoxic damage was assessed by micronucleus and chromosomal aberration assays. Total antioxidant capacity and total oxidant status were also measured in serum samples to assess oxidative status. Treatment of Group B with permethrin resulted in genotoxic damage and increased total oxidant status levels. Permethrin treatment also significantly decreased (P < 0.05) total antioxidant capacity level when compared to Group A rats. Group C rats showed significant increases (P < 0.05) in total antioxidant capacity level and no alterations in cytogenetic parameters. Moreover, simultaneous treatments with olive leaf extract significantly modulated the toxic effects of permethrin in Group D rats. It can be concluded that olive leaf extract has beneficial influences and could be able to antagonize permethrin toxicity. As a result, this investigation clearly revealed the protective role of olive leaf extract against the genetic and oxidative damage by permethrin in vivo for the first time.     

Determination of LC50 of permethrin acaricide in semi-engorged females of the tick Rhipicephalus sanguineus (Latreille, 1806) (Acari: Ixodidae)

Currently, the most efficient and widely used method of tick control still is the treatment with acaricides, especially permethrin (active ingredient of the Advantage^® Max3, Bayer), a pyrethroid with neurotoxic action. Due to the wide use of this acaricide in the control of the tick Rhipicephalus sanguineus, this study carried out laboratorial procedures to determine the LC₅₀ (lethal concentration fifty) of permethrin in semi-engorged females of R. sanguineus. Based on the result of 14 dilutions of permethrin in distilled water and later Probit analysis, the LC₅₀ of permethrin for R. sanguineus was 2062 ppm (1549-2675 ppm). This work can be used as a protocol with other chemicals, to determinate the LC₅₀, basic procedure for studies of control, resistance and behavior of ticks treated with acaricides, especially the brown dog tick R. sanguineus. Also, the knowledge of the LC₅₀ provides information on the potency of chemicals, the sensitivity of Arthropods to them and even estimates on pest control.     

Reduction in swimming performance in juvenile rainbow trout (Oncorhynchus mykiss) following sublethal exposure to pyrethroid insecticides

While the lethal toxicity of pyrethroid insecticides to fish is well documented, their sublethal physio-behavioral effects remain poorly characterized. Known pyrethroid-associated changes to insect neuromuscular function may translate into similar effects in fish, thereby altering swimming ability and affecting foraging, predator avoidance, and migration. Three experiments were conducted using critical (U_crit) and burst (U_max) swimming speeds to assess the sublethal effects of the pyrethroids permethrin and deltamethrin in juvenile rainbow trout (Oncorhynchus mykiss). Fish were exposed to deltamethrin (100, 200, or 300 ng/L) or permethrin (1, 2, or 3 μg/L) in water for 4 d, and assessed for swimming performance. Deltamethrin (200 and 300 ng/L) reduced U_crit, but not U_max, while both swim performance measurements were unaffected by permethrin. Subsequent experiments used only U_crit to assess deltamethrin exposure. In a time course experiment, deltamethrin (300 ng/L) reduced U_crit after 1 and 4 d of exposure, but after 7 d of exposure U_crit was fully recovered. Finally, deltamethrin (1, 2, or 3 μg/L) reduced U_crit after 1 h bath exposures similar to recommended protocols for deltamethrin based sea-lice treatment in aquaculture. The real-world implications of the revealed pyrethroid-associated swimming ability reductions in salmon may be important in areas close to aquaculture facilities.     

The role of the Aedes aegypti Epsilon glutathione transferases in conferring resistance to DDT and pyrethroid insecticides

The Epsilon glutathione transferase (GST) class in the dengue vector Aedes aegypti consists of eight sequentially arranged genes spanning 53,645 bp on super contig 1.291, which maps to chromosome 2. One Epsilon GST, GSTE2, has previously been implicated in conferring resistance to DDT. The amino acid sequence of GSTE2 in an insecticide susceptible and a DDT resistant strain differs at five residues two of which occur in the putative DDT binding site. Characterization of the respective recombinant enzymes revealed that both variants have comparable DDT dehydrochlorinase activity although the isoform from the resistant strain has higher affinity for the insecticide. GSTe2 and two additional Epsilon GST genes, GSTe5 and GSTe7, are expressed at elevated levels in the resistant population and the recombinant homodimer GSTE5-5 also exhibits low levels of DDT dehydrochlorinase activity. Partial silencing of either GSTe7 or GSTe2 by RNA interference resulted in an increased susceptibility to the pyrethroid, deltamethrin suggesting that these GST enzymes may also play a role in resistance to pyrethroid insecticides.     

Permethrin Alters Adipogenesis in 3T3‐L1 Adipocytes and Causes Insulin Resistance in C2C12 Myotubes

Pyrethroids are a class of insecticides structurally derived from the naturally occurring insecticides called pyrethrins. Along with emerging evidence that exposure to insecticides is linked to altered weight gain and glucose homeostasis, exposure to pyrethroids has been linked to altered blood glucose levels in humans. Thus, the purpose of this study was to determine the role of permethrin on lipid and glucose metabolisms. Permethrin was treated to 3T3‐L1 adipocytes and C2C12 myoblasts to determine its role in lipid and glucose metabolisms, respectively. Permethrin treatment resulted in increased expression of key markers of adipogenesis and lipogenesis in adipocytes. Permethrin significantly reduced insulin‐stimulated glucose uptake in myotubes. This is the first report on the role of permethrin in altered lipid metabolism in adipocytes and impaired glucose homeostasis in myotubes. These results may help elucidate fundamental underlying mechanisms between insecticide exposure, particularly permethrin, and potential risk of developing obesity and its comorbidities.