Acute toxicity and esterase response to carbaryl exposure in two different populations of amphipods Hyalella curvispina.

During the last years, a carbaryl insecticide was extensively applied in the valley of Río Negro and Neuquén, North Patagonia Argentina, to manage codling moths (Cydia pomonella), the main pest of pear and apple trees. In this study carbaryl susceptibility and B-esterase activity from both insecticide-exposed and non-exposed field populations of amphipods Hyalella curvispina were studied. Two subpopulations, one susceptible to carbaryl (LC50=213±7.5µg/L carbaryl) and one resistant to it (LC50=14,663±2379µg/L carbaryl), were found in the agricultural area selected in this study. Both populations were, in turn, more resistant to carbaryl than the population from a pristine area (LC50=11.31±2.27µg/L carbaryl). The in vivo 48h-IC50 values for cholinesterase (ChE) were close to the corresponding 48h-LC50 values as determined for the non-exposed population (IC50=7.16±0.86µg/L carbaryl) and for the susceptible subpopulation from the insecticide-exposed site (IC50=193±99µg/L carbaryl). Carbaryl exposure of the amphipods from the agricultural area mentioned above produced a significant decrease of carboxylesterase (CabE) activity, at a sublethal concentration (10µg/L) that was not able to significantly inhibit ChE, thereby showing a protective role of CabE and its usefulness as early biomarker. However, at lethal concentrations the inhibition of ChE activity was higher than that of CabE. On the other hand, CabE of amphipods from the pristine site was less sensitive to carbaryl than ChE, suggesting a different participation of CabE in ChE protection in the susceptible population of H. curvispina. Pulse exposure to carbaryl for 2h caused a significant inhibition of ChE in amphipods from both populations, with a fast recovery as expected for a carbamate insecticide. In conclusion, we proved that amphipods from the said agricultural area have developed resistance to carbaryl and showed the presence of two subpopulations with a different response to the insecticide. Moreover, these results reinforce the use of ChE together with CabE inhibition as indicators of carbamate exposure in H. curvispina.

Acute toxicity and biochemical effects of azinphos methyl in the amphipod Hyalella curvispina.

We evaluated the acute toxicity and biochemical effects of the organophosphorus pesticide azinphos methyl (AM) in the amphipod Hyalella curvispina that inhabits ponds and irrigation channels of an intensive fruit-producing region in Rio Negro and Neuquén valley, North Patagonia, Argentina. The analysis by nonlinear regression of data from the 96 h-acute toxicity tests indicated the coexistence of two subpopulations of H. curvispina with different susceptibilities to AM. The 96 h-LC50 for the resistant subpopulation (166 ± 56 µg/L) was 216-fold higher than the 96h-LC50 value for the susceptible one (0.77 ± 1.33 µg/L).The two subpopulations could not be distinguished based on the biochemical measurements in control amphipods. Cholinesterase activity was significantly inhibited in AM-exposed amphipods in a concentration-dependent manner. The IC50 value obtained after 96 h of exposure (2.18 ± 1.95 µg/L) was significantly lower than the 48 h-IC50 value (29.6 ± 17.4 µg/L). Carboxylesterase activity was significantly inhibited after 48 h of exposure to 12.5 and 62.5 µg/L AM (inhibition, 51%). This enzyme was thus able to protect cholinesterase from inhibition at 12.5 µg/L AM. Reduced glutathione and catalase showed a significant increase after 24 h of exposure as an adaptive response to AM, whereas glutathione S-transferase activity was not significantly modified. The analysis of species sensitivity distribution showed that both subpopulations of H. curvispina were more tolerant to AM than most amphipod species, and that the susceptible subpopulation was more sensitive to AM than the other local aquatic species analyzed. The maximum concentration of AM in drainage water within the fruit-producing area reported by other studies would affect most of the amphipod species (99%) and also a 44% of local aquatic ones. The results obtained in this study point out the usefulness of including amphipods like H. curvispina in ecotoxicity studies and monitoring programs to perform pesticide risk assessments.

Evolution of insecticide resistance in non-target black flies (Diptera: Simuliidae) from Argentina.

Black flies, a non-target species of the insecticides used in fruit production, represent a severe medical and veterinary problem. Large increases in the level of resistance to the pyrethroids fenvalerate (more than 355-fold) and deltamethrin (162-fold) and a small increase in resistance to the organophosphate azinphos methyl (2-fold) were observed between 1996-2008 in black fly larvae under insecticide pressure. Eventually, no change or a slight variation in insecticide resistance was followed by a subsequent increase in resistance. The evolution of pesticide resistance in a field population is a complex and stepwise process that is influenced by several factors, the most significant of which is the insecticide selection pressure, such as the dose and frequency of application. The variation in insecticide susceptibility within a black fly population in the productive area may be related to changes in fruit-pest control. The frequency of individuals with esterase activities higher than the maximum value determined in the susceptible population increased consistently over the sampling period. However, the insecticide resistance was not attributed to glutathione S-transferase activity. In conclusion, esterase activity in black flies from the productive area is one mechanism underlying the high levels of resistance to pyrethroids, which have been recently used infrequently. These enzymes may be reselected by currently used pesticides and enhance the resistance to these insecticides.

Evolution of insecticide resistance in non-target black flies (Diptera: Simuliidae) from Argentina

Black flies, a non-target species of the insecticides used in fruit production, represent a severe medical and veterinary problem. Large increases in the level of resistance to the pyrethroids fenvalerate (more than 355-fold) and deltamethrin (162-fold) and a small increase in resistance to the organophosphate azinphos methyl (2-fold) were observed between 1996-2008 in black fly larvae under insecticide pressure. Eventually, no change or a slight variation in insecticide resistance was followed by a subsequent increase in resistance. The evolution of pesticide resistance in a field population is a complex and stepwise process that is influenced by several factors, the most significant of which is the insecticide selection pressure, such as the dose and frequency of application. The variation in insecticide susceptibility within a black fly population in the productive area may be related to changes in fruit-pest control. The frequency of individuals with esterase activities higher than the maximum value determined in the susceptible population increased consistently over the sampling period. However, the insecticide resistance was not attributed to glutathione S-transferase activity. In conclusion, esterase activity in black flies from the productive area is one mechanism underlying the high levels of resistance to pyrethroids, which have been recently used infrequently. These enzymes may be reselected by currently used pesticides and enhance the resistance to these insecticides.

Biomarkers of effect in toads and frogs.

Amphibians are good bioindicators of environmental pollution due to their susceptibility to chemicals during their freshwater cycles. The effects of environmental pollution, together with changes in human activity and climate, have contributed to the reduction in the amphibian population over recent decades. However, toxicological research on amphibians has been rather scarce compared with that on other vertebrates. In this article we review the biochemical alterations underlying xenobiotic action and/or the detoxifying responses described for anuran species, with the aim of establishing possible biomarkers of effect. During the embryonic development of anurans, morphological and behavioural alterations are the effects most frequently cited in connection with chemical exposures. However, such biomarkers have a low sensitivity and are unspecific compared with biochemical alterations. Some primary pesticide targets, in particular cholinesterases for organophosphates and carbamates, have been evaluated. Esterases change seasonally and with the stage of development, and their sensitivity to anticholinesterase agents varies between species. Thus their use as biomarkers in anurans must be carefully analysed. Enzymes and endogenous compounds related to oxidative metabolism may also be used as biomarkers of effect. Glutathione pool, glutathione-S-transferases and metallothioneins respond in different ways to pesticides and heavy metals in anuran embryos and tadpoles. Mixed-function oxidases, in turn, are less developed in amphibians, and show a reduced induction in response to pesticide exposures. Endogenous polyamine levels are also proposed as good age-related biomarkers of damage. Finally, molecular biomarkers related to receptor binding, signal transduction and genetic response have gained increasing relevance, as they have been implicated in the fertilisation process and the earliest events in anuran development. The identification of transcription factors associated with the exposure of amphibians to xenobiotics as well as other alterations in hormone signalling appears highly promising. However, these techniques are likely to complement other methods. In conclusion, the use of several biomarkers with multiple endpoints is needed to link exposure to response and to provide better predictive tools for the environmental protection of endangered anuran species.