Daphnia stress response to environmental concentrations of chloramphenicol-multi-omics approach

Abstract

Commonly used medicines, when discarded or improperly disposed of, are known to contaminate freshwater ecosystems. Pharmaceuticals can be toxic and mutagenic, and can modify freshwater organisms, even at environmentally relevant concentrations. Chloramphenicol (CAP) is an antibiotic banned in Europe. However, it is still found in surface waters around the world. The aim of this study was to evaluate the impact of chloramphenicol contamination in freshwater on the model organism Daphnia magna. Specific life history parameters, proteome, and host-associated microbiome of four D. magna clones were analyzed during a three-generation exposure to CAP at environmental concentrations (32 ng L^-1). In the first generation, no statistically significant CAP effect at the individual level was detected. After three generations, exposed animals were smaller at first reproduction and on average produced fewer offspring. The differences in D. magna's life history after CAP treatment were in accordance with proteome changes. D. magna's response to CAP presence indicates the high stress that the tested organisms are under, e.g., male production, upregulation of ubiquitin-conjugating enzyme E2 and calcium-binding protein, and downregulation of glutathione transferase. The CAP-exposed D. magna proteome profile confirms that CAP, being reactive oxygen species (ROS)-inducing compounds, contributes to structural changes in mitochondria. Microbiome analysis showed a significant difference in the Shannon index between control and CAP-exposed animals, the latter having a more diverse microbiome. Multilevel analyses, together with long exposure in the laboratory imitating conditions in a polluted environment, allow us to obtain a more complete picture of the impact of CAP on D. magna.

Keywords: Antibiotics; Life history; Microbiome; Proteome.

Fig. 1

Experimental setup

Fig. 2

Daphnia magna (all clones) life history parameters. Boxplot showing the distribution of life history parameters across generation and treatment. The central line within each box represents the median, the edges of the box represent the first and third quartiles (Q1 and Q3), and the whiskers extend to the minimum and maximum values. Outliers are plotted as individual points beyond the whiskers as white circles. Star indicates significant differences among treatments (repeated-measure ANOVA, post hoc Tukey HSD)

Fig. 3

Gender percentage (females – white bar; males – grey bar) of Daphnia magna (B, D, K, and P clone) cultured without pharmaceutical (empty bars), in the presence of CAP (striped bars) after three generations

Fig. 4

Daphnia microbiome analysis. A star symbol indicates a p-value less than 0.05. Bacterial taxa with a mean relative abundance of less than 0.1% in all samples were filtered out