Biomedical subjects
Publications and source records attributed to L McCarroll.
Treatment of head louse infection is primarily through topical insecticides. However, there is growing evidence of resistance. A representative population sample was tested using biochemical and molecular methods; it was shown that, in Wales, treatments containing pyrethroids are likely to be less effective in controlling head louse infection than those containing organophosphates.
Resistance to permethrin in an East African population of the major malaria vector, Anopheles gambiae is multifactorial. A mutated sodium channel allele and enhanced insecticide metabolism contribute to the resistance phenotype. We used microsatellite markers to scan the genome for quantitative trait loci (QTL) associated with permethrin resistance. Two major and one minor QTL were identified. The first QTL, rtp1, colocalizes with the sodium channel gene on chromosome 2L thus further supporting the importance of mutations in this gene in conferring permethrin resistance. The second two loci are located on the third chromosome and one of these, rtp2, flanks a large cluster of cytochrome P450 genes. Further detailed mapping of these regions will help elucidate the molecular mechanisms of metabolic resistance to insecticides.
Samples of the dengue vector mosquito Aedes aegypti (L.) (Diptera: Culicidae) were collected from 13 localities between 1995 and 1998. Two laboratory strains, Bora (French Polynesia) and AEAE, were both susceptible to DDT and permethrin; all other strains, except Larentuka (Indonesia) and Bouaké (Ivory Coast), contained individual fourth-instar larvae resistant to permethrin. Ten strains were subjected to a range of biochemical assays. Many strains had elevated carboxylesterase activity compared to the Bora strain; this was particularly high in the Indonesian strains Salatiga and Semarang, and in the Guyane strain (Cayenne). Monooxygenase levels were increased in the Salatiga and Paea (Polynesia) strains, and reduced in the two Thai strains (Mae Kaza, Mae Kud) and the Larentuka strain. Glutathione S-transferase activity was elevated in the Guyane strain. All other enzyme profiles were similar to the susceptible strain. The presence of both DDT and pyrethroid resistance in the Semarang, Belem (Brazil) and Long Hoa (Vietnam) strains suggested the presence of a knock-down resistant (kdr)-type resistance mechanism. Part of the S6 hydrophobic segment of domain II of the voltage-gated sodium channel gene was obtained by RT-PCR and sequenced from several insects from all 13 field strains. Four novel mutations were identified. Three strains contained identical amino acid substitutions at two positions, two strains shared a different substitution, and one strain was homozygous for a fourth alteration. The leucine to phenylalanine substitution that confers nerve insensitivity to pyrethroids in a range of other resistant insects was absent. Direct neurophysiological assays on individual larvae from three strains with these mutations demonstrated reduced nerve sensitivity to permethrin or lambda cyhalothrin inhibition compared to the susceptible strains.
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Recombinant glutathione S-transferase (agGST1-6) from the malaria vector mosquito Anopheles gambiae Giles (Diptera: Culicidae) was expressed in Escherichia coli using a pET3a vector system. The expressed enzyme was biochemically active with reduced glutathione (GSH) and 1-chloro-2,4-dinitrobenzene (CDNB). Activity of agGST1-6 with GSH and CDNB was inhibited to different degrees by both alpha-cyano and non-alpha-cyano pyrethroid insecticides. This inhibition was used to develop an assay for quantification of pyrethroids. Standard curves of insecticide concentration against percentage of enzyme inhibition or volume of iodine solution were established by spectrophotometry and iodine volumetric titration, respectively, for permethrin and deltamethrin. These assays allowed estimation of pyrethroid concentrations both spectrophotometrically and visually. For the residue assay of each insecticide, a cut-off point of 50% of the initial pyrethroid impregnation concentration was used, which should differentiate between biologically active and inactive treated bednets. The cross-reactivity of the primary permethrin photodegradants (3-phenoxyalcohol and 3-phenoxybenzoic acid) with the recombinant agGST1-6 was assayed in the same system. No agGST1-6 inhibition by the insecticide metabolites was observed, suggesting that the system is unaffected by primary permethrin metabolites and will accurately measure insecticide parent compound concentrations. The estimated pyrethroid insecticide concentrations, given spectrophotometrically and by iodine titration assay, were comparable to those obtained by direct HPLC quantification of residues extracted from bednets. Hence, it should be relatively easy to adapt this method to produce a test kit for residue quantification in the field.
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The evolution and spread of insecticide resistance is an important factor in human disease prevention and crop protection. The mosquito Culex quinquefasciatus is the main vector of the disease filariasis and a member of a species complex which is a common biting nuisance worldwide. The common insecticide resistance mechanism in this species involves germline amplification of the esterases estalpha21 and estbeta21. This amplification has arisen once and rapidly spread worldwide. Less common and more variable resistance phenotypes involve coamplification of estalpha3 and estbeta1, or individual amplification of a single estbeta1, different alleles of the same estalpha and estbeta gene loci. Estalpha21 and estbeta21 are on the same large fragment of amplified DNA (amplicon) 2.7 kb apart. We have now shown that this amplicon contains another full-length gene immediately 5' of estalpha21 which codes for a molybdenum-containing hydroxylase, with highest homology to aldehyde oxidase (AO) from other organisms. The full-length putative AO gene is not present on the estalpha3/estbeta1 or estbeta1 amplicons, but multiple truncated 5' ends of this gene are present around the presumed estalpha3/estbeta1 amplicon breakpoint. Polymerase chain reaction (PCR) analysis of insecticide-susceptible genomic DNA demonstrated that a different allele of the putative AO gene in its non-amplified form is immediately 5' of estalpha. The 'AO' gene on the estalpha21/estbeta21 amplicon is expressed and resistant insects have greater AO activity. This AO activity is sensitive to inhibition by an aldehyde-containing herbicide and pesticide. This enzyme may confer a selective advantage to these insects in the presence of insecticide, as AO in mammals is believed to be important in the detoxification process of several environmental pollutants.
The inflammatory cytokines tumor necrosis factor-alpha (TNF alpha) and interleukin-1 alpha (IL-1 alpha) have angiogenic properties but generally inhibit cultured endothelial cell (EC) proliferation. Investigations into the growth-regulatory effects of these two agents on a variety of cultured EC types showed that they exert mitogenic, anti-proliferative or cytotoxic effects depending upon cell type and cytokine combinations. The anti-proliferative effect was distinct from cytotoxicity. Nitric oxide (NO) release from EC, examined as a potential mechanism underlying some of these effects, did not appear to mediate the anti-proliferative effects of these cytokines. However, NO also seemed to have a bimodal effect on EC proliferation depending upon whether the NO was endogenous or exogenous. These data underscore the diversity in cytokine and NO effects on cultured EC which, if reproducible in vivo, may be partly responsible for the variable and sometimes contradictory results obtained with regards to the role of inflammatory cytokines and NO on angiogenesis.
Insect cells are relatively cheap to maintain and are capable of producing accurately translated and correctly processed heterologous proteins. Recent research has focused on the development of improved expression vectors for continuous, high-level production of foreign proteins, including a number of membrane-targeted receptors, in Drosophila and lepidopteran insect cells. Mosquito cells have also been employed for studies on the control of vector-borne diseases, such as malaria.