Selective and Irreversible Inhibitors of Aphid Acetylcholinesterases: Steps Toward Human-Safe Insecticides

Aphids, among the most destructive insects to world agriculture, are mainly controlled by organophosphate insecticides that disable the catalytic serine residue of acetylcholinesterase (AChE). Because these agents also affect vertebrate AChEs, they are toxic to non-target species including humans and birds. We previously reported that a cysteine residue (Cys), found at the AChE active site in aphids and other insects but not mammals, might serve as a target for insect-selective pesticides. However, aphids have two different AChEs (termed AP and AO), and only AP-AChE carries the unique Cys. The absence of the active-site Cys in AO-AChE might raise concerns about the utility of targeting that residue. Herein we report the development of a methanethiosulfonate-containing small molecule that, at 6.0 µM, irreversibly inhibits 99% of all AChE activity extracted from the greenbug aphid (Schizaphis graminum) without any measurable inhibition of the human AChE. Reactivation studies using β-mercaptoethanol confirm that the irreversible inhibition resulted from the conjugation of the inhibitor to the unique Cys. These results suggest that AO-AChE does not contribute significantly to the overall AChE activity in aphids, thus offering new insight into the relative functional importance of the two insect AChEs. More importantly, by demonstrating that the Cys-targeting inhibitor can abolish AChE activity in aphids, we can conclude that the unique Cys may be a viable target for species-selective agents to control aphids without causing human toxicity and resistance problems

Selective and Irreversible Inhibitors of Mosquito Acetylcholinesterases for Controlling Malaria and Other Mosquito-Borne Diseases

New insecticides are urgently needed because resistance to current insecticides allows resurgence of disease-transmitting mosquitoes while concerns for human toxicity from current compounds are growing. We previously reported the finding of a free cysteine (Cys) residue at the entrance of the active site of acetylcholinesterase (AChE) in some insects but not in mammals, birds, and fish. These insects have two AChE genes (AP and AO), and only AP-AChE carries the Cys residue. Most of these insects are disease vectors such as the African malaria mosquito (Anopheles gambiae sensu stricto) or crop pests such as aphids. Recently we reported a Cys-targeting small molecule that irreversibly inhibited all AChE activity extracted from aphids while an identical exposure caused no effect on the human AChE. Full inhibition of AChE in aphids indicates that AP-AChE contributes most of the enzymatic activity and suggests that the Cys residue might serve as a target for developing better aphicides. It is therefore worth investigating whether the Cys-targeting strategy is applicable to mosquitocides. Herein, we report that, under conditions that spare the human AChE, a methanethiosulfonate-containing molecule at 6 µM irreversibly inhibited 95% of the AChE activity extracted from An. gambiae s. str. and >80% of the activity from the yellow fever mosquito (Aedes aegypti L.) or the northern house mosquito (Culex pipiens L.) that is a vector of St. Louis encephalitis. This type of inhibition is fast (∼30 min) and due to conjugation of the inhibitor to the active-site Cys of mosquito AP-AChE, according to our observed reactivation of the methanethiosulfonate-inhibited AChE by 2-mercaptoethanol. We also note that our sulfhydryl agents partially and irreversibly inhibited the human AChE after prolonged exposure (>4 hr). This slow inhibition is due to partial enzyme denaturation by the inhibitor and/or micelles of the inhibitor, according to our studies using atomic force microscopy, circular dichroism spectroscopy, X-ray crystallography, time-resolved fluorescence spectroscopy, and liquid chromatography triple quadrupole mass spectrometry. These results support our view that the mosquito-specific Cys is a viable target for developing new mosquitocides to control disease vectors and to alleviate resistance problems with reduced toxicity toward non-target species

Palaeosymbiosis Revealed by Genomic Fossils of Wolbachia in a Strongyloidean Nematode

Wolbachia are common endosymbionts of terrestrial arthropods, and are also found in nematodes: the animal-parasitic filaria, and the plant-parasite Radopholus similis. Lateral transfer of Wolbachia DNA to the host genome is common. We generated a draft genome sequence for the strongyloidean nematode parasite Dictyocaulus viviparus, the cattle lungworm. In the assembly, we identified nearly 1 Mb of sequence with similarity to Wolbachia. The fragments were unlikely to derive from a live Wolbachia infection: most were short, and the genes were disabled through inactivating mutations. Many fragments were co-assembled with definitively nematode-derived sequence. We found limited evidence of expression of the Wolbachia-derived genes. The D. viviparus Wolbachia genes were most similar to filarial strains and strains from the host promiscuous clade F. We conclude that D. viviparus was infected by Wolbachia in the past, and that clade F-like symbionts may have been the source of filarial Wolbachia infections

What socio-demographic factors influence poverty and financial health care access among disabled people in Flanders : a cross-sectional study

BACKGROUND: Current literature shows that people with a disability have a lower income than people without a disability. Disabled people often experience difficulties with health care access. The objective of this study is to assess the current financial situation and poverty rate amongst disabled people in Flanders. Furthermore we wanted to analyze factors that contribute to the risk of poverty and problems with financial health care access in adult people with a disability in Flanders. METHODS: An online and paper survey were constructed and made available through two large organizations for people with different types of disability in Flanders. Descriptive statistics and logistic regression analysis were performed. RESULTS: In this convenience sample, 20.9% of the 889 respondents live under the poverty threshold. Important contributing factors to the risk of poverty are having children (OR 3.43, 95% CI 2.10-5.59) and a low level of dependence (OR 16.40, 95% CI 6.21-43.28). 25.2% of the respondents did not access health care because of financial shortcomings. A low level of dependence is one important contributing factor (OR 3.16, 95% CI 1.41-6.98) to limited financial health care access. CONCLUSION: This research confirms that disability is associated with a higher risk of poverty and impaired financial health care access