Comparative capability to detoxify vegetable allelochemicals by larval mosquitoes.

In order to confirm the phytotoxicological basis for the ecological specialization of larval culicine fauna among different subalpine mosquito breeding sites, we compared the capability of six different Aedes larval taxa or populations of different ecological origin to detoxify dietary leaf litter originating from the environmental vegetation. Detoxification experiments were performed through in vitro digestion of a toxic leaf litter fraction using larval extracts as the enzymatic sources. Comparison of toxicological and detoxifying properties among the different larval samples indicates an association between their tolerance to leaf litter toxicants and their detoxification capability, which vary according to ecological origin. The fact that the detoxifying factor within the larval extracts appears to be a protein-like compound with a molecular weight bigger than 30 kDa suggests the possible involvement of detoxifying enzymes in larval tolerance to leaf litter toxicants. This is congruent with previous biochemical data that suggests the involvement of cytochrome P450 monooxygenase and esterase activities in the detoxification process.

Factors influencing the toxicity of xenobiotics against larval mosquitoes.

In order to examine the factors influencing xenobiotic toxicity against larval mosquitoes, the larvicidal performances of two conventional insecticides (temephos and Bacillus thuringiensis var. israelensis: Bti) and a new potential phyto-insecticide (decomposed leaf litter) were compared under different conditions against three detritivorous larval mosquito types. Bioassays performed under standard conditions indicated differential tolerance levels according to the xenobiotic and the larval type. Bioassays performed under different conditions of xenobiotic dose and geometry of the water column indicated differential effects of those parameters on mortality rates. This allowed us to distinguish the performances of temephos versus those of Bti and leaf litter. These toxicological performances were examined as indicators for analysis of xenobiotic bioavailability for mosquito larvae in environmental water, and also for their comparative interest in field mosquito control.

Hot extraction and characterization of a ligninlike fraction involved in larvicidal effects of decomposed leaf litter against mosquito.

Hot water-extraction was performed on decomposed leaf litter in order to solubilize the toxic fraction involved in the dietary interaction against mosquito larvae in subalpine breeding sites. The toxic fraction was partially extracted by water with an optimum temperature of 60 degrees C and recovered in an insoluble form. Phytochemical characterization was achieved through differential enzymatic hydrolyses, using the laccase mediator delignifying system, and aluminum chloride chelation monitored by standard bioassays; comparative spectrophotometric analyses in ultraviolet light after solubilization in acetyl bromide; and comparative reversed-phase high-performance liquid chromatography of the phenolic aldehydes after alkaline nitrobenzene oxidation. The results suggested the involvement of ligninlike compounds in the toxicity of the isolated fraction. Toxicity of this fraction appeared far stronger than that of the crude leaf litter. The involvement of this ligninlike fraction in the dietary toxicity of leaf litter against larval mosquito was then investigated.

Dietary toxicity of decomposed arborescent leaf litter against larval mosquito: involvement of a lignin-polypeptidic complex.

To characterize the toxic compounds involved in the dietary toxicity of decomposed arborescent leaf litter against larval mosquito, a toxic fraction was extracted from crude leaf litter by using hot water. Preliminary characterization of this fraction, called the insoluble fraction (IF) because it progressively precipitates after extraction, has suggested the involvement of lignin-like compounds in the toxicity. Further analyzes are currently being performed by using additional phytotoxicity-based methods. The involvement of lignin-like compounds in the toxicity was indicated by both the comparative effects of different enzymatic oxidative treatments and reversed-phase high-performance liquid chromatography analysis of the phenolic aldehydes and acids obtained after alkaline nitrobenzene oxidation. However, these lignin-like compounds may not be involved alone in the toxicity, as no specific feature of those components was associated with the toxicity. Among the possible compounds associated with lignin-like compounds in the toxicity, peptidic compounds were suggested by comparative determination of the C/N ratio and then revealed by denaturation experiments, use of specific binding protein molecules, and thin-layer chromatography analysis. A possible role of these peptidic compounds associated with lignin-like compounds in the dietary toxicity of the leaf litter against the larval mosquito midgut is discussed.

Taste sensitivity of detritivorous mosquito larvae to decomposed leaf litter.

Dietary leaf litter chemistry is known to play an important ecotoxicological role in the plant-mosquito interaction in subalpine flooded areas surrounded by vegetation because of differential larvicidal effects of insoluble polyphenols formed during the leaf decaying process. This dietary interaction was investigated through comparative evaluation of the role of toxic/nontoxic leaf litter in both larval foraging and feeding behavior, by using different samples of decomposed alder leaf litter and larval Aedes aegypri as experimental references. Track analysis showed significant differences in larval foraging behavior in the absence or presence of leaf litter. Comparative alimentary preference investigations and further track analysis suggested that larvae are unable to detect leaf litter toxicity. These characteristics of the larval behavioral feeding pattern suggested that: (1) decomposed leaf litter may be involved as an important attractive food source in the habitat selection and evolutionary history of culicids, and (2) preingestive behavioral mechanisms appear to be minimally involved in the differential larval dietary adaptation to toxic leaf litter. These results may have interesting consequences for culicid biological control.

Larvicidal properties of decomposed leaf litter in the subalpine mosquito breeding sites.

The larvicidal properties of the dietary leaf litter originating from the vegetation surrounding the subalpine mosquito breeding sites were investigated by using 10-month decomposed alder leaf litter against different field collections of culicine taxa of various ecological origin (Aedes cantans, Aedes caspius, Aedes cataphylla, Aedes detritus, Aedes punctor, Aedes pullatus, Aedes rusticus, Anopheles claviger, Culex hortensis, Culex pipiens, Culiseta morsitans). Larvae originating from sites with polyphenol-poor vegetation appeared more sensitive to ingested leaf litter than those originating from sites with polyphenol-rich vegetation. Within a given taxon (e.g., A. rusticus, A. cataphylla, C. hortensis), the overall levels of cytochrome P450 monooxygenase and esterase activities appeared higher in larvae able to feed on leaf litter than in pupae and adults unable to feed on leaf litter. This suggests the involvement of these enzymes in the detoxification mechanisms responsible for larval tolerance to polyphenols of the dietary leaf litter. Such a tolerance of the larval stage thus appears as fundamental in the ecotoxicological adaptation of mosquito taxa to the polyphenolic profiles of the riparian vegetation.

Premolt calcium secretion in midgut posterior caeca of the crustacean Orchestia: Ultrastructure of the epithelium.

Both structural and functional changes are observed within the posterior caeca (PC) of Orchestia during the molt cycle. During the intermolt period, there are two segments which are structurally different: a distal segment lined by type I epithelial cells and a proximal segment lined by type II cells. During molting, the PC cells are active in calcium turnover. Calcium is secreted and stored as calcareous concretions in the caecal lumen during the preexuvial period; then during the postexuvial period it is reabsorbed to mineralize the new cuticle. During the preexuvial period, cellular type III differentiates along the whole length of the PC in poster-anterior sequence and functions in ionic calcium secretion, from the basal part to the cellular apex. During the postexuvial period, this cellular type turns into cellular type IV engaged in calcium reabsorption from successive generations of spherites, from the cellular apex to the basal part. The role played by the caecal epithelium during both formation and reabsorption of the concretions was investigated by experiments in which caeca were transplanted to host pericardial cavities or were blocked by causing an abdominal hernia. The main structural characteristic features of cellular type III are as follows: an extracellular network of channels extends from basal to apical ends; microvilli are long and often apically dilated; multivacuolar complexes are localized in extracellular channels and within dilated tips of microvilli before secretion into caecum lumen; bundles of microtubules are oriented in parallel around the luminal orifices of the extracellular network; ribosomes are abundant in cytoplasm. Cellular type III develops progressively from the distal end of the caecum to the proximal one as the preexuvial period advances and concretions form in the caecum lumen.