Glyphosate-Induced Specific and Widespread Perturbations in the Metabolome of Soil Pseudomonas Species

Abstract

Previous studies have reported adverse effects of glyphosate on crop-beneficial soil bacterial species, including several soil Pseudomonas species. Of particular interest is the elucidation of the metabolic consequences of glyphosate toxicity in these species. Here we investigated the growth and metabolic responses of soil Pseudomonas species grown on succinate, a common root exudate, and glyphosate at different concentrations. We conducted our experiments with one agricultural soil isolate, P. fluorescens RA12, and three model species, P. putida KT2440, P. putida S12, and P. protegens Pf-5. Our results demonstrated both species-and strain-dependent growth responses to glyphosate. Following exposure to a range of glyphosate concentrations (up to 5 mM), the growth rate of both P. protegens Pf-5 and P. fluorescens RA12 remained unchanged whereas the two P. putida strains exhibited from 0 to 100% growth inhibition. We employed a 13 C-assisted metabolomics approach using liquid chromatography-mass spectrometry to monitor disruptions in metabolic homeostasis and fluxes. Profiling of the whole-cell metabolome captured deviations in metabolite levels involved in the tricarboxylic acid cycle, ribonucleotide biosynthesis, and protein biosynthesis. Altered metabolite levels specifically in the biosynthetic pathway of aromatic amino acids (AAs), the target of toxicity for glyphosate in plants, implied the same toxicity target in the soil bacterium. Kinetic flux experiments with 13 C-labeled succinate revealed that biosynthetic fluxes of the aromatic AAs were not inhibited in P. fluorescens Pf-5 in the presence of low and high glyphosate doses but these fluxes were inhibited by up to 60% in P. putida KT2440, even at sub-lethal glyphosate exposure. Notably, the greatest inhibition was found for the aromatic AA tryptophan, an important precursor to secondary metabolites. When the growth medium was supplemented with aromatic AAs, P. putida S12 exposed to a lethal dose of glyphosate completely recovered in terms of both growth rate and selected Aristilde et al.Metabolomics of Glyphosate Effects on Soil Bacteria metabolite levels. Collectively, our findings led us to conclude that the glyphosateinduced specific disruption of de novo biosynthesis of aromatic AAs accompanied by widespread metabolic disruptions was responsible for dose-dependent adverse effects of glyphosate on sensitive soil Pseudomonas species.

The soil microbial hydrolytic activity, content of nitrogen and organic carbon were enhanced by organic farming management using cover crops and composts in potato cultivation

“…Our results are in accordance with previous findings that show the inhibition effect of pesticide residues on the soil biota and activity of microorganisms (Makaw et al 1979;Angelini et al 2013). Based on the literature, glyphosate affects adversely the freeliving bacteria populations by reducing the amount of soil bacteria as well as the ratios of various bacteria groups and thereby decreasing the soil fertility (Newman et al 2016;Aristilde et al 2017).…”

The Introduction of Thousands of Tonnes of Glyphosate in the food Chain—An Evaluation of Glyphosate Tolerant Soybeans

“…A recent meta-analysis showed a significant link between exposure to glyphosate and Non-Hodgkin lymphoma [32]. The focus on the putative carcinogenicity of glyphosate has been to the neglect of evidence of adverse effects of various types and end-points, e.g., in soil bacteria [33], and direct and indirect (transgenerational) effects on male sperm quantity and quality in rats [34,35]. In our opinion, the needed re-evaluation of glyphosate toxicity should not be delayed by, or narrowly focused on, the question of whether or not glyphosate should be categorized as a carcinogen [36][37][38].…”

Re-registration Challenges of Glyphosate in the European Union

“…A recent study provides a Europe-wide assessment of the dispersal of glyphosate and AMPA in EU agricultural topsoils, being present in 45% of the topsoils collected, originating from 11 countries and six crop systems, with a maximum concentration of 2 mg/kg, as well as their potential spreading by wind and water erosion , further affected by small-scale sediment transport in water erosion (Bento et al, 2018), persisting under low bacterial activity in limited aerobic conditions or non-neutral pH (la Cecilia and Maggi, 2018), and adversely affecting soil microbial and nematodal diversity (Dennis et al, 2018). From the soil glyphosate can be translocated by plant roots; and it can affect non-target plants near agricultural ditches (Saunders and Pezeshki, 2015), and affect soil rhizosphere-associated bacterial communities (Newman et al, 2016) and soil Pseudomonas species (Aristilde et al, 2017). Toxicity of Roundup R to the soil filamentous fungus Aspergillus nidulans was reported with a median lethal dose (LD 50 ) corresponding to glyphosate concentrations of 90-112 mg/l, ∼100-fold below agricultural application levels (Nicolas et al, 2016).…”