Combined effects of glyphosate and chemical hypoxia in zebrafish: a new toxicological point of view

Glyphosate (Gly), a systemic and non-selective post-emergence herbicide used worldwide, has 3 emerged as a pollutant. However, its toxic effects are debated by regulatory authorities. In 4 addition, in the aquatic environment, often the presence of pollutants is associated with a 5 hypoxia condition that could change their toxicological effects. We used zebrafish embryos to 6 evaluate the toxic effects of Gly and its mechanisms in a hypoxic condition chemically induced 7 by cobalt chloride (CoCl2). We found that Gly induced toxicity in a time and concentration8 dependent manner. The toxicity of Gly was determined at 96 hours post fertilization as a lethal 9 concentration (LC), and LC10, LC20, and LC50 values were 85.7, 97, and 122.9 mg/L, 10 respectively. When Gly was combined with CoCl2 the toxicological endpoints were lower than 11 values referred to the Gly alone indicating the worse effects of chemical hypoxia on Gly 12 toxicity. Histological observations were performed at 25, 50, 75, and 100 mg/L for Gly both 13 alone and in combination with 10 mM CoCl2. Fisher’s exact test showed significant differences 14 in the presence of hepatic and gut inflammation at 75 and 100 mg/L of Gly both alone and in 15 combination with CoCl2. To deeply investigate the effects of hypoxia on Gly toxicity we 16 decided to test the lowest dose of Gly, 50 mg/L, alone or in combination with CoCl2 10 mM on 17 liver glycogen storage and oxidative stress. Again the results obtained indicate the worse effects 18 of chemical hypoxia on Gly toxicity. Thus Gly toxicity could be reconsidered in light of the 19 damage it causes to the liver and intestines and its effect in combination with factors that induce 20 chemical hypoxia.