Metabolomics revealed more deleterious toxicity induced by the combined exposure of ammonia and nitrite on Ruditapes philippinarum compared to single exposure.

NH3-N and NO2-N always co-exist in the aquatic environment, but there is not a clear opinion on their joint toxicities to the molluscs. Presently, clams Ruditapes philippinarum were challenged by environmental concentrations of NH3-N and NO2-N, singly or in combination, and analyzed by metabolomics approaches, enzyme assays and transmission electron microscope (TEM) observation. Results showed that some same KEGG pathways with different enriched-metabolites were detected in the three exposed groups within one day, and completely different profiles of metabolites were found in the rest of the exposure period. The combined exposure induced heavier and more lasting toxicities to the clams compared with their single exposure. ACP activity and the number of secondary lysosomes were significantly increased after the combined exposure. The present study shed light on the joint-toxicity mechanism of NH3-N and NO2-N, and provided fundamental data for the toxicity research on inorganic nitrogen.

Comparative analysis of microRNA expression profiles in clam Ruditapes philippinarum after ammonia nitrogen exposure.

Ammonia nitrogen is a long-lasting pollutant along the Chinese coast. In our previous studies, the clam Ruditapes philippinarum exhibited several toxic responses to environmental concentrations of ammonia nitrogen. To elucidate the underlying mechanism of ammonia nitrogen toxicity in clams at the post-transcriptional level, microRNA (miRNA) expression profiles were investigated by high-throughput sequencing after the clams were exposed to 0.1 mg/L ammonia nitrogen for 30 days. A total of 238 miRNAs were identified, including 49 conserved miRNAs and 189 novel miRNAs. After comparative analysis, six miRNAs were significantly expressed after 1 day of exposure, with three up-regulated and three down-regulated miRNAs. In addition, 35 miRNAs were significantly expressed after 30 days of exposure, of which 16 were up-regulated and 19 were down-regulated. Furthermore, the target genes of each differentially expressed miRNA were predicted, followed by Gene Ontology (GO) category and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. The target genes were predicted to be involved in the immune response, protein processing and transport, DNA damage repair, cellular communication, neural signaling, redox homeostasis, lipid metabolism, and biotransformation. A biological phagocytosis assay proved the speculation that ammonia nitrogen regulated the immunity of clams with the aid of a novel miRNA (novel_29). These findings support further research on miRNA levels in R. philippinarum exposed to ammonia nitrogen.