Acute Toxic Effects of the Herbicide Formulation Focus® Ultra on Embryos and Larvae of the Moroccan Painted Frog, Discoglossus scovazzi

Abstract

For regulatory and scientific purposes, there is a need to understand the sensitivity of a wider variety of wild species of amphibians and the sensitivities within their life stages to chemicals of widespread use such as herbicides. We investigated the acute toxic effects of the herbicide formulation Focus Ultra [with the active ingredient (a.i.) cycloxydim plus solvent naphtha and sodium dioctylsulphosuccinate as added substances] on embryos and early stage larvae of the Moroccan painted frog (Discoglossus scovazzi). Different clinical signs (twitching, convulsion, and narcosis) occurred at 40 and 80 mg/L in embryos (4 and 8 mg a.i./L) and narcotic effects (total immobilization or irregular escape responses) at 10, 15, and 20 mg/L in larvae (1, 1.5, and 2 mg a.i./L). Growth inhibition (total length), starting at 20 mg/L in embryos and 2.5 mg/L in larvae (2 and 0.25 mg a.i./L, respectively) was understood as sign of toxicity (retardation) and not as sign of teratogenicity. However, the connection to teratogenesis remained unclear though total length reduction occurred at concentrations <20 % of the 96-h LC50 value and at a minimum concentration that inhibits growth of only 17 % of the 96-h LC50 value. Starting at 20 mg/L, mortality in embryos significantly increased and at 15 mg/L in early larvae (2 and 1.5 mg a.i./L, respectively). Mortality of larvae was enhanced during the first 24 h of exposure to 15 and 20 mg/L (1.5 and 2 mg a.i./L). Morphology of the embryos remained unobtrusive. In contrary, axial malformations significantly increased in the early larvae starting at 10 mg/L (1 mg a.i./L), a concentration free of lethal effects. In all considered end points, larvae were significantly more sensitive than embryos, probably because of developmental and physiological properties or different exposure and bioavailability of the compound. Focus Ultra induced comparable lethal and immobilization effects in D. scovazzi as it does to standard test organisms in pesticide approval. However, to validate the apparent safety in the field, which is based on calculated surface water concentrations of the a.i., more data on real contamination levels is necessary (e.g., peak concentrations, concentrations of added substances). Furthermore, sufficient buffer strips between the farmland and amphibian ponds must be considered, and the effects of the substance on terrestrial life stages have not been assessed yet.

Population and life‐stage–specific effects of two herbicide formulations on the aquatic development of European common frogs (Rana temporaria)

“…For both test substances, the added vehicles are mainly responsible for adverse effects . This is also discussed for narcotic effects observed for Focus Ultra in other studies and seems be true as well for Roundup UltraMax in the present study.…”

“…Growth inhibition due to Focus Ultra exposure in the early R. temporaria larvae of the present study started at comparable low test concentrations as in a previous study [17]. In the present study, growth inhibition was significantly increased in the lowest Roundup UltraMax test concentrations (Figure 4).…”

“…Focus Ultra significantly increased axial malformations in anuran larvae in a previous study [17]. This kind of malformation was also observed in X. laevis larvae, but did not reach the level of significance [16].…”

“…Calculated concentrations for Roundup UltraMax were 0 mg a.i./L, 2.25 mg a.i./L, 4.5 mg a.i./L, 9 mg a.i./L, 11.25 mg a.i./L, and 13.5 mg a. i./L; and for Focus Ultra, concentrations were 0 mg a.i./L, 0.5 mg a.i./L, 1 mg a.i./L, 1.5 mg a.i./L, 2 mg a.i./L, and 3 mg a. i./L. Because no range-finding was possible with R. temporaria individuals, the concentrations were chosen based on rangefinding tests with a discoglossid species that also develops suspension feeding larvae [17]. The same water parameters already measured at the breeding sites were taken.…”

“…Causal relationships between increasing pesticide use and amphibian population declines remain unclear due to the current lack of a broad database flanked by appropriate knowledge of the toxic potential of pesticides at low concentrations in the field . Most adverse effects on amphibians have been observed in laboratory studies and in studies using mesocosms . Field data show that pesticide use can affect amphibians at least at the individual level .…”