The criteria applied to the acute ECOTOX dataset for measured concentrations to be reported potentially resulted in fewer acute comparisons than may have been expected for an analysis of this type. However, the reduced uncertainty associated with this refined acute dataset is likely to outweigh the potential benefits of a larger dataset, which may include unreliable studies. Because Aldrich  did not require studies to be based on measured concentrations, some amphibian data based on nominal concentrations were included.
In this analysis, amphibians were between 10-fold and 100-fold more sensitive than fish in comparisons for aluminum chloride (25-fold), 2,4 dichloroaniline (12-fold), malathion (34-fold), and pentachlorophenol-sodium salt (12-fold). In all of these cases, application of the standard EU assessment factor of 100 to the regulatory fish LC50 value would have covered the amphibian LC50 value, and these results are therefore not discussed further.
The only two comparisons of acute LC50 values that suggested greater than 100-fold sensitivity for amphibians were those for p-nonylphenol (2,111-fold) and dimethoate (7,300-fold). The acute amphibian data for p-nonylphenol were taken from a paper by Bridges et al. , in which southern leopard frog (Rana sphenocephala) tadpoles were exposed to measured concentrations of p-nonylphenol for 96 h. They reported a 96-h LC50 of 0.34 µg/L, which is much lower than the corresponding fish geometric mean value from ECOTOX of 718 µg/L (560–920 µg/L), which is based on two 96-h LC50 values reported by Ernst et al. . However, along with data on amphibian sensitivity to p-nonylphenol, Bridges et al.  also cite a rainbow trout 96-h LC50 value for p-nonylphenol of 0.19 µg/L and a fathead minnow 96-h LC50 value for p-nonylphenol of 0.27 µg/L from U.S. EPA . They also cite a boreal toad (Bufo boreas) tadpole 96-h LC50 for p-nonylphenol of 0.12 µg/L . These additional data suggest that there is in fact little difference in the acute toxicity of p–nonylphenol to amphibians and fish. This is further supported by the chronic data comparison for p–nonylphenol, which shows that fish are at least 10 times more sensitive than amphibians (see Table 1).
In our analysis and in the analysis by Aldrich , the acute amphibian LC50 for dimethoate was taken from a paper by Khangarot et al. , in which Rana hexadactyla tadpoles were exposed to formulated dimethoate (Rogor 30 EC). The reported 96-h LC50 is 7.82 ppb Rogor 30 EC (nominal concentration), which would correspond to 2.4 µg/L dimethoate active substance, although Khangarot et al.  are not clear on the expression of endpoints in units of formulation or active substance. However, other data sources suggest that amphibians and fish are similarly sensitive to dimethoate. For example, the Environment Agency of England and Wales  collated acute and chronic data for dimethoate, and the value reported by Khangarot et al.  is again identified as the most sensitive amphibian result (however, the study is considered not reliable, because of missing analytical values and insufficient description of the test methodology). For fish, the Environment Agency report also identified a low 96-h LC50 value for mullet exposed to dimethoate of 2.3 µg/L from a paper by Aboul-Eta and Khalil , which was also considered not reliable, because of missing chemical analysis. Furthermore, the chronic data pair for dimethoate shows a higher sensitivity of fish (see Table 1). Therefore, the weight of evidence suggests that fish and amphibians are similarly sensitive, rather than amphibians being significantly more sensitive than fish.
After additional scrutiny of the two instances of greater than 100-fold amphibian sensitivity initially identified in this analysis, in general, amphibians are of comparable acute sensitivity to fish in laboratory toxicity studies and, on average, slightly less sensitive than fish (Fig. 1). This finding was conspicuous despite the stringent data selection criteria applied in the present study, which allowed any amphibian species to be included in the analysis, whereas fish data were limited to only two species (O. mykiss and P. promelas) which potentially excluded more sensitive fish species, as demonstrated in the case of dimethoate.
Other authors have found that amphibians are, in general, either equally or less sensitive than fish to acute chemical exposures [3-13, 29]. For example, Hoke and Ankley  found that LC50 values from the frog embryo teratogenesis assay-Xenopus (FETAX) were not the most sensitive result when compared with other acute toxicity endpoints from traditional aquatic test species (including fish) for Cd, Cu, Se, Hg, Zn, ammonia, aniline, pentachlorophenol, atrazine, malathion, and parathion. The only exception was aluminum, for which FETAX was more sensitive, but only by a factor of approximately 2. Bridges et al.  report comparative 96-h LC50 results for southern leopard frog tadpoles (Rana sphenocephala), boreal toad tadpoles (Bufo boreas), bluegill sunfish (Lepomis macrochirus), fathead minnow, and rainbow trout for five chemicals (p-nonylphenol, carbaryl, Cu, pentachlorophenol, and permethrin). The most sensitive tadpole and fish results were within a factor of 2 for all five chemicals.
Therefore, the overall picture from this analysis and other reviews is that fish are generally more acutely sensitive than amphibians, and the standard EU risk assessment factor of 100 accounts for potential species sensitivity differences between fish and amphibians.
Further details of the studies with potentially unreliable NOECs that suggest that amphibians are more sensitive than fish are provided in the following sections.
The azoxystrobin amphibian NOEC of 10 µg/L was taken from a study by Johannsson et al.  in which common frogs (Rana temporaria) were exposed to 1 or 10 µg/L azoxystrobin. No effects were observed at the highest test concentration of 10 µg/L. This unbounded NOEC cannot be regarded as a true NOEC and is therefore an unreliable basis for comparison with the fish NOEC of 147 µg/L taken from the OPP database. Equally, the amphibian NOEC for acetochlor, which was reported by Helbing et al. , is based on exposure of Rana catesbeiana tadpoles to either 1 or 10 nM acetochlor. At 10 nM (equivalent to 2.7 µg/L, based on a molecular weight of 269.77 g/mol), no effects on apical endpoints (forelimb emergence, mouth development, or tail regression) were seen. This again is an unbounded NOEC and an unreliable basis for comparison with the fish NOEC of 130 µg/L taken from the OPP database. In both of these instances, the true amphibian NOEC will likely be higher than the reported NOEC initially used for the comparisons, and this therefore does not provide evidence of greater amphibian sensitivity to these chemicals, despite the ratios calculated in this analysis.
The amphibian NOEC of 59 µg/L, for effects on development of X. laevis exposed to ammonium perchlorate, is based on a paper by Goleman et al. . This study employed a very large gap between exposure concentrations with a corresponding LOEC of 14,000 µg/L. The large spacing factor between the NOEC and LOEC means that this result is insufficient for a reliable comparison of amphibian and fish sensitivity to ammonium perchlorate, as the true NOEC could span more than two orders of magnitude between the reported NOEC and LOEC values. Furthermore, this study was conducted in FETAX medium, which may have contributed to an artificially high sensitivity (see later discussion on sodium perchlorate). The corresponding fish NOEC, based on a study by Crane et al. , is reported as 1,000 µg/L, which is between the NOEC and LOEC for amphibians as reported by Goleman et al. . In this case, the maximum acceptable toxicant concentration ratio is 3.48 (compared with the NOEC ratio of 16.9), indicating that the observed sensitivity difference is at least partly determined by experimental design and may in fact be much smaller.
After removing the above data, amphibian NOECs were more than 10 times more sensitive than corresponding fish NOECs for three substances: carbaryl (42-fold), dexamethasone (11-fold), and sodium perchlorate (218-fold).
The amphibian NOEC for carbaryl is based on results from a paper by Rohr et al. . Streamside salamander (Ambystoma barbouri) embryos were exposed to carbaryl at 0.5, 5, or 50 µg/L. Survival was reduced at 50 µg/L by approximately 20%, which is why an NOEC for survival of 5 µg/L is reported. The corresponding fish NOEC of 210 µg/L stems from a fathead minnow early life-stage test taken from the OPP database. The maximum acceptable toxicant concentration ratio is 10.6 (compared with the NOEC ratio of 42), indicating that the observed sensitivity difference is at least partly determined by experimental design and may in fact be much smaller. The U.S. EPA Office of Prevention, Pesticides, and Toxic Substances reviewed the relative acute and chronic toxicity of carbaryl to amphibian and fish species, including the studies already discussed, as part of a Pesticide Effects Determination of the risks of carbaryl use to the federally listed endangered Barton Springs salamander (Eurycea sosorum ). This assessment concluded that none of the amphibian toxicity data available in the open literature was sufficiently robust to use for quantitative risk assessment. They also report that the available evidence suggests that amphibians are less sensitive to carbaryl than the most sensitive fish species for which data were available (i.e., Atlantic salmon, Salmo salar, with an NOEC of 6.8 µg/L).
The amphibian NOEC for the corticosteroid drug dexamethasone is derived from a study by Lorenz et al. , who exposed X. laevis tadpoles (stage 51) to 10, 100, or 500 nM dexamethasone for 21 d. They reported effects on hindlimb growth at 100 nM, leading to identification of a NOEC of 10 nM from their study, which is 3.9 µg/L (based on a molecular weight of 392.5 g/mol). The corresponding fish value is a growth NOEC of 42.7 µg/L (the LOEC is 424 µg/L) from a 29-d fish early life-stage test with fathead minnow . The NOEC ratio is 11, with a similar maximum acceptable toxicant concentration ratio, as both studies employed a (relatively large) spacing factor of 10. Both of these studies seem to have been performed and reported adequately, so amphibians may be more sensitive than fish to dexamethasone. Lorenz et al.  suggest that the effects that they measured in Xenopus are likely to have resulted from complex mechanisms involved in the modulatory actions of corticosteroids on amphibian metamorphosis and thyroid hormones, via prolactin synthesis. This biochemical pathway does not occur in fish species, which could explain the apparent difference in sensitivity.
The amphibian NOEC for sodium perchlorate is derived from a study by Brausch et al. , who investigated the relative sensitivity of X. laevis to perchlorate (as sodium perchlorate) in natural stream water and synthetic (FETAX) test media. Embryos (Nieuwkoop-Faber Stage 11) of X. laevis were exposed in the laboratory to measured concentrations of perchlorate in either FETAX medium (33.5 and 70.4 µg/L) or natural stream water (29.3 and 98.8 µg/L) for 66 d. The NOEC and LOEC values of 33.5 µg/L and 70.4 µg/L were derived for developmental effects on metamorphosis and hindlimb length, respectively, in FETAX media. However, no statistically significant effects were observed in X. laevis exposed to perchlorate in natural stream water. In a separate experiment, Brausch et al.  report that exposure of juvenile (5-d-old) S. multiplicata to perchlorate in natural stream water at measured concentrations of 50.1, 107.1, and 1,038 µg/L for 42 d resulted in no statistically significant effects on either metamorphosis or survival. The authors conclude that natural surface water mitigates the anti-metamorphic effect of perchlorate in X. laevis and is likely to contribute to the lack of effects observed in S. multiplicata. The precise mechanism by which perchlorate effects are modified by test media has not been established, but it may be that natural surface waters contain sufficient iodide or some other water quality characteristic that mitigates the anti-metamorphic effects in amphibians. Similar contrasting observations on perchlorate toxicity (high in FETAX medium, low or absent in natural water) have been reported by Carr et al. . Goleman and Carr  report a NOEC for sodium perchlorate for X. laevis exposed in FETAX medium of 22.6 µg/L, which is similar to the NOEC in FETAX medium reported by Brausch et al. . The Goleman and Carr study was not selected as a key NOEC for the present study, because the spacing factor between test concentrations was greater than 100. However, it and other studies  provide supporting evidence for the NOEC reported by Brausch et al.  in FETAX medium. Perchlorate is a potent inhibitor of the uptake of iodide into the thyroid, reducing the formation of thyroid hormones that control amphibian metamorphosis [91, 108]. The specific mechanism of perchlorate toxicity may explain the relative sensitivity of amphibians to this substance, although the environmental relevance of laboratory studies on perchlorate using FETAX medium is uncertain. The observed high sensitivity of Xenopus to perchlorate in FETAX media could be a potential artifact of the medium. Garber  also reported on deficiencies of the FETAX medium, which hinders tadpole metamorphosis. Because fish are usually tested in natural well water or dechlorinated tap water, such artifacts would not occur under chronic testing conditions for fish. Finally, after 40 weeks of exposure to measured concentrations of perchlorate of up to 1,500 µg/L in natural water, X. tropicalis did not show any effects on apical endpoints. Hence the NOEC was at least 1,500 µg/L . Thus, in comparable media no difference is found between fish and amphibians in their chronic sensitivity to perchlorate.
Several of the chronic studies with amphibians that are evaluated here have been conducted to study the effects of chemicals on the hypothalamic–pituitary–thyroid axis; that is, they specifically investigate the influence on amphibian development of an endocrine mode of action. To characterize such effects, the amphibian metamorphosis assay with X. laevis (Amphibian Metamorphosis Assay, OECD test guideline 231) was developed. For other endocrine modes of action, such as (anti-)estrogenic or (anti-)androgenic mechanisms, regulatory concerns for aquatic vertebrates are usually addressed through testing with fish. Nevertheless, the present analysis of chronic data shows that even for chemicals that are known to influence the hypothalamic–pituitary–thyroid axis in amphibians, such as ethylenethiourea and propylthiouracil, fish apical endpoints are comparably sensitive (the only possible exception being the drug dexamethasone). Hence, fish data are appropriate to cover the chronic risks of such substances to amphibians. The potential to study specific thyroid-mediated effects in fish, including an assessment of thyroid histology and sensitivity considerations, was explored for propylthiouracil by Schmidt and Braunbeck , sodium perchlorate by Mukhi and Patino , and ammonium perchlorate by Crane et al. . This demonstrates the potential to use fish higher-tier toxicity tests to address the risk of substances active on the hypothalamic–pituitary–thyroid endocrine axis.
Kerby et al.  found that amphibians were highly acutely sensitive to the three phenolic chemicals that they analyzed (triclosan and two unidentified others). In our analysis of chronic data, amphibians were not more sensitive than fish to the phenolic substances 17-β–estradiol, 4-tert-octylphenol, 17-α-ethinylestradiol, bisphenol A, p-nonylphenol, nonylphenol, and triclosan. Amphibians were slightly more sensitive to pentachlorophenol. However, in this instance a standard assessment factor of 10 would be sufficient to cover the observed difference in sensitivity. Our analysis therefore does not suggest a need for additional amphibian testing for phenolic substances.
Therefore, the overall picture from this analysis and other reviews is that amphibians are not generally more chronically sensitive than fish and the standard EU risk assessment factor of 10 accounts for potential species sensitivity differences between fish and amphibians.