How glyphosate and its associated acidity affect early development in zebrafish (Danio rerio)

Abstract

Background Glyphosate is among the most extensively used pesticides worldwide. Following the ongoing highly controversial debate on this compound, its potential impact on non-target organisms is a fundamental scientific issue. In its pure compound form, glyphosate is known for its acidic properties. Methods We exposed zebrafish (Danio rerio) embryos to concentrations between 10 μM and 10 mM glyphosate in an unbuffered aqueous medium, as well as at pH 7, for 96 hours post fertilization (hpf). Furthermore, we investigated the effects of aqueous media in the range of pH 3 to 8, in comparison with 1 mM glyphosate treatment at the respective pH levels. Additionally, we exposed zebrafish to 7-deoxy-sedoheptulose (7dSh), another substance that interferes with the shikimate pathway by a mechanism analogous to that of glyphosate, at a concentration of one mM. The observed endpoints included mortality, the hatching rate, developmental delays at 24 hpf, the heart rate at 48 hpf and the malformation rate at 96 hpf. LC10/50, EC10 and, if reasonable, EC50 values were determined for unbuffered glyphosate. Results The results revealed high mortalities in all treatments associated with low pH, including high concentrations of unbuffered glyphosate (>500 μM), low pH controls and glyphosate treatments with pH < 3.4. Sublethal endpoints like developmental delays and malformations occurred mainly at higher concentrations of unbuffered glyphosate. In contrast, effects on the hatching rate became particularly prominent in treatments at pH 7, showing that glyphosate significantly accelerates hatching compared with the control and 7dSh, even at the lowest tested concentration. Glyphosate also affected the heart rate, resulting in alterations both at pH 7 and, even more pronounced, in the unbuffered system. In higher concentrations, glyphosate tended to accelerate the heart rate in zebrafish embryos, again, when not masked by the decelerating influence of its low pH. At pH > 4, no mortality occurred, neither in the control nor in glyphosate treatments. At 1 mM, 7dSh did not induce any mortality, developmental delays or malformations; only slightly accelerated hatching and a decelerated heart rate were observed. Our results demonstrate that lethal impacts in zebrafish embryos can be attributed mainly to low pH, but we could also show a pH-independent effect of glyphosate on the development of zebrafish embryos on a sublethal level.

Recommendations for Improving Methods and Models for Aquatic Hazard Assessment of Ionizable Organic Chemicals

“…Another recent study on pharmaceuticals in the zebrafish embryo assay showed higher toxicity with higher fraction of neutral species (Alsop and Wilson ). We did not include a study on the pH‐dependent toxicity of glyphosate toward zebrafish embryos in Supplemental Data, Table S1, because the zebrafish embryo assay was performed unbuffered and pH was <4, most likely leading to pH effects rather than effects of glyphosate because glyphosate did not cause any mortality up to 10 mM when buffered to pH 7 and the negative control was almost equally toxic at low pH values (Schweizer et al ).…”

Effects of parental exposure to glyphosate-based herbicides on embryonic development and oxidative status: a long-term experiment in a bird model

“…Glyphosate and commercial products (e.g. RoundUp) made with glyphosate have been repeatedly reported to cause embryo mortality and deformations in fish (zebrafish 10uM to 1 mM pure glyphosate 9 , 10 mg/L RoundUp or pure glyphosate 10 ), and aquatic amphibians (Xenopus 0.3-1.3 mg/L RoundUp 11 , 500 pg/egg pure glyphosate 12 ). In contrast, mammal and bird embryos and fetuses are exposed to glyphosate residues only via maternal transfer of the chemicals, which may result in malformations, altered sex ratios, and low sperm quality in rodent models (doses: 500 mg/kg RoundUp 13 , 5 g/L pure glyphosate 14 , 50-450 mg/kg RoundUp 15 ).…”

Impacts of a glyphosate-based herbicide on the gut microbiome of three earthworm species (Alma millsoni, Eudrilus eugeniae and Libyodrilus violaceus): A pilot study

“…In field studies, the half-life of glyphosate can range between a few days to around 6 months [ 50 ]. In soils, glyphosate competes with phosphorus and thus its absorption depends on mineral composition [ 51 ]. All these environmental factors will impact GBH availability to earthworms in agricultural settings.…”