Transcriptomic and enzymatic analyses revealed synergistic toxicity of ammonia and nitrite on Ruditapes philipparum

A prior study revealed that combined exposure to ammonia nitrogen (NH₃-N) and nitrite nitrogen (NO₂-N) led to significant changes in metabolites and increased levels of phagocytosis-related enzymes in Ruditapes philipparum clams, compared to individual exposures. However, the underlying mechanism remains unknown. In this study, R. philipparum were exposed to single or combined NH₃-N and NO₂-N for 7 days, and differentially expressed genes (DEGs) were identified through comparative transcriptome analysis, followed by GO and KEGG analyses. The results indicated that both exposure treatment and duration influenced the outcomes of the GO and KEGG analyses. The combined exposure group exhibited higher toxicity, as shown by the early activation of pathways related to mitochondrial damage, inflammation, CYP450 detoxification, and immune response. Additionally, the combined exposure significantly induced more severe toxicity pathways, which would worsen the clams' condition by disrupting carbohydrate and protein metabolism, energy production, detoxification, and causing cell disintegration and death. Mitochondrial damage was identified as a critical factor in combined toxicity, supported by increased enzyme activities related to carbohydrate metabolism (α-KGDH and SDH) and redox parameters (SOD, CAT, GR, GSH, GPX, and MDA). In summary, this study reveals a synergistic toxicity of NH₃-N and NO₂-N at environmental pollution concentrations on R. philipparum and highlights significant threats to marine bivalve aquaculture.