Lidia Gauna

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.

Thiols and polyamines in the potentiation of malathion toxicity in larval stages of the toad Bufo arenarum.

Treatment with exogenous spermidine enhanced acute malathion toxicity during larval development of the toad Bufo arenarum Hensel. The polyamine was rapidly incorporated in the larvae with a subsequent metabolization to putrescine and spermine, which were excreted to the media. Endogenous polyamine levels were not changed by either spermidine or malathion treatments. However, 0.5-mM spermidine modified malathion uptake and bioavailability increasing the concentration of the xenobiotic in the larvae. The amount of reduced thiols was decreased by both compounds, but the depletion was insufficient to induce cytotoxicity. The oxidative degradation of polyamines competes for the pool of reduced glutathione used in the conjugation of malathion in the larvae, thus leading to the reported potentiation of toxicity. Our results suggest that exposure to thiols-depleting agents may induce alteration of organophosphate degradation in amphibian larvae.

Early biochemical changes produced by malathion on toad embryos

The exposure of newly fertilized amphibian embryos to malathion (44 mg/L) produces 67% mortality on the fifth day of exposure. Clinical signs of intoxication are evident on the fourth day. A variety of biochemical parameters, such as enzyme activity, lipid and protein content, was examined in control and malathion-treated embryos. The activity of acetyl, butyrylcholinesterase and aliesterase are quickly inhibited. An interference with cellular protein synthesis and redistribution is possible. A slight increase in total phospholipids is evident after 24 hr of exposure.

Effect of exogenously applied polyamines on malathion toxicity in the toad Bufo arenarum Hensel

The polyamines putrescine, spermidine, and spermine, are able to potentiate the toxicity of malathion in Bufo arenarum Hensel toad larvae. This action is synergistic and maximal with spermidine, which elevated up to 13-fold the mortality produced by this organophosphorus compound. Spermidine increased the degree of inhibition of acetylcholinesterase activity elicited by malathion, and impaired the recovery of this activity at the end of the treatment. Spermidine had no effect on the enzyme when applied alone. Toxic effects were also observed with the polyamines themselves when applied at concentrations similar to the intracellular levels described for rapid-growing organisms.

Comparative toxicity of parathion in early embryos and larvae of the toad, Bufo arenarum hensel

Received: 31 March 1993/Accepted: 24 October 1993 Parathion (O,O-diethyl O-p-nitrophenyl phosphorothioate) is a widely used agricultural insecticide highly toxic to mammals; however, very little is known about the detoxication capacity in some natural enemies of insects such as amphibians (Harriet al 1979; Llamas et al. 1985; Caballero de Castro et al 1991; Gauna et al. 1991). The biota may be stressed by discharges of pesticides, and it is therefore essential to know its potential effect on non-target organisms before irreversible change occurs. Some monitoring techniques use amphibian larvae as an useful indicator for aquatic contamination (Dumpert and Zeitz 1984; Beiswenger 1988; Williams et al. 1989). An ideal insecticide should be efficacious against pest species, but relatively safe for non-target organisms. To develop selective insecticides it is important to understand the differences and similarities of the defense systems between insects and the beneficial fauna such as predators and parasitoids. Thus, this report deals with a comparative study of parathion susceptibility on early embryonic and larval stages of the Argentinian toad

Mechanisms of resistance to DDT and pyrethroids in Patagonian populations of Simulium blackflies.

Mixed populations of the pest blackflies Simulium bonaerense Coscarón & Wygodzinsky, S. wolffhuegeli (Enderlein) and S. nigristrigatum Wygodzinsky & Coscarón (Diptera: Simuliidae) are highly resistant to DDT and pyrethroids in the Neuquén Valley, a fruit‐growing area of northern Patagonia, Argentina. As these insecticides have not been used for blackfly control, resistance is attributed to exposure to agricultural insecticides. Pre‐treatment with the synergist piperonyl butoxide (PBO) reduced both DDT and fenvalerate resistance, indicating that resistance was partly due to monooxygenase inhibition. Pre‐treatment with the synergist tribufos to inhibit esterases slightly increased fenvalerate toxicity in the resistant population. Even so, biochemical studies indicated almost three‐fold higher esterase activity in the resistant population, compared to the susceptible. Starch gel electrophoresis confirmed higher frequency and staining intensity of esterase electromorphs in the resistant population. Incomplete synergism against metabolic resistance indicates additional involvement of a non‐metabolic resistance mechanism, such as target site insensitivity, assumed to be kdr‐like in this case. Glutathione S‐transferase activities were low and inconsistent, indicating no role in Simulium resistance. Knowing these spectra of insecticide activity and resistance mechanisms facilitates the choice of more effective products for Simulium control and permits better coordination with agrochemical operations.

Effects of dieldrin treatment on physiological and biochemical aspects of the toad embryonic development

Dieldrin is a cylclodiene insecticide highly persistent in nature due to its chemical stability. The exposure of toad embryos to Dieldrin induces hyperactivity in the swimming larvae and inhibition of cholinesterases. However, the inhibition of these enzymes during early development is not life threatening. The present report provides a physiological and biochemical study of the noxious effect of Dieldrin on the toad embryonic development.

MODEL STUDY OF FACTORS INFLUENCING STEADY STATE CLEARANCE FOR LIPOPHILIC TOXICANTS IN AQUATIC MICROCOSMS

We have studied the distribution and bioelimination of the organophosphorus pesticide parathion in a native microcosm consisting of water, sediment, bivalves and aquatic plants. A common apparent clearance constant for biotic and abiotic components was suggested from the analysis of parathion accumulation and degradation. In this work we developed a global model explaining the toxicokinetics of a lipophilic compound and particurlarly the common steady state degradation in an aquatic microcosm, using a set of linear differential equations. We simulated the distribution and degradation of the compound in the microcosm, and fitted single-compartment equation models to data, estimating the apparent sorption and elimination constants. We verified the existence of a common apparent degradation constant for all the compartments. We infer from the mathematical expressions and corroborate from the simulated data that the apparent degradation constant is equal to the sum of the metabolization rates at each biotic compartment multiplied by the compound mass ratios established at steady state between the biotic and the abiotic compartments. Product kinetics simulation showed that steady state might also be achieved in the different compartments, with the same apparent constant as that obtained for toxicant clearance. As a practical result, the total radioactivity in water would serve to calculate the global clearance constant in a simple experimental way if a radiotracer is used. Physical and chemical degradation and chemical loss due to volatilization and CO2 diffusion were analyzed in the microcosm model. The assessments of the cases where these factors affect the clearance process as well as the implications emerged are discussed.