Glyphosate, but not its metabolite AMPA, alters the honeybee gut microbiota

Abstract

The honeybee (Apis mellifera) has to cope with multiple environmental stressors, especially pesticides. Among those, the herbicide glyphosate and its main metabolite, the aminomethylphosphonic acid (AMPA), are among the most abundant and ubiquitous contaminant in the environment. Through the foraging and storing of contaminated resources, honeybees are exposed to these xenobiotics. As ingested glyphosate and AMPA are directly in contact with the honeybee gut microbiota, we used quantitative PCR to test whether they could induce significant changes in the relative abundance of the major gut bacterial taxa. Glyphosate induced a strong decrease in Snodgrassella alvi, a partial decrease of a Gilliamella apicola and an increase in Lactobacillus spp. abundances. In vitro, glyphosate reduced the growth of S. alvi and G. apicola but not Lactobacillus kunkeei. Although being no bee killer, we confirmed that glyphosate can have sublethal effects on the honeybee microbiota. To test whether such imbalanced microbiota could favor pathogen development, honeybees were exposed to glyphosate and to spores of the intestinal parasite Nosema ceranae. Glyphosate did not significantly enhance the effect of the parasite infection. Concerning AMPA, while it could reduce the growth of G. apicola in vitro, it did not induce any significant change in the honeybee microbiota, suggesting that glyphosate is the active component modifying the gut communities.

Fig 1. Abundances of the main bacterial taxa of the honeybee microbiota after chronic exposure to glyphosate and/or AMPA, relative to untreated controls.

Summer and winter honeybees were chronically exposed to 1.5 mM or 7.5 mM of glyphosate (GLY) and/or AMPA in their feeding sugar syrup. Some bees were also infected with spores of N. ceranae (Ncer). Control bees (Ct) were not exposed to any stressor. 15 days after treatment initiation, the DNA from guts was extracted and the 16S rDNA of the major taxa of the microbiota was quantified by QPCR and normalized to the total bacterial 16S content. The data are represented as the log2 of the ratio of the abundance in treated condition relative to untreated controls. Data were gathered from five independent colony-replicates. Bars represent the 95% confidence intervals. Stars and hash marks indicate significant differences (paired t-test, p<0.05 and p<0.005 respectively). When a difference was significant, a similar effect was observed in all the five colony replicates. nd: not determined.

Fig 2. Growth of bacterial strains isolated from the honeybee gut in the presence of glyphosate and AMPA.

Strains were inoculated in HIB or MRS medium in the presence of increasing concentrations of glyphosate (GLY) or AMPA. After 48h of growth under anaerobic conditions, the optical density of the cultures was measured at 600 nm. The results are presented as the mean ratio between treated culture and control culture without xenobiotic that was used as a calibrator. Data were gathered from six independent replicates. Bars represent the 95% confidence intervals. A significant difference (Student t-test or Mann-Whitney test) is indicated by a star (p<0.05) or a hash mark (p<0.001).