Polystyrene nanobeads enhance polycylic aromatic hydrocarbon genotoxicity in an in vitro fish model

Microscopic plastic particles – microplastics – are a ubiquitous contaminant in aquatic environments, which potentially exist at the nanoscale. They bind hydrophobic chemicals, such as polycyclic aromatic hydrocarbons (PAHs), altering their environmental fate, as well as interactions with biota. Using rainbow trout intestine (RTgutGC) and gill (RTgill-W1) cell lines, the present study investigates the effects of polystyrene micro-(MB) (4 µm) and nanobeads (NB) (0.22 µm) on the cyto- and genotoxicity of two PAHs, benzo[a]pyrene (BaP) and 3-nitrobenzanthrone (3-NBA) over 48 hrs. Using the Alamar Blue bioassay to assess cytotoxicity, PAHs (0.1, 1 and 10 µM) decreased cell viability by 10-20% at 10 µM in both cell lines, whereas MBs and NBs (1,000 – 10,000 particles / ml) were non-toxic. Genotoxicity was assessed using the Comet assay and a measure of oxidative DNA damage via the formamidopyrimidine DNA-glycosylase (FPG)-modified Comet assay. In the gill cell line only minor increases (1-4%) in DNA damage were seen in all treatments. The gut cell line DNA damage increased to 8% for BaP and 15% for 3-NBA, with a greater induction of oxidative DNA damage. The most dramatic result was in the presence of both 0.22 µm NBs and 10µM 3-NBA, with oxidative DNA damage reaching 69%. This was not observed for the 4µm MBs. Results show the presence of plastic NBs enhances genotoxicity of 3-NBA, causing a significant increase in DNA damage in the intestine cell line. Dietary exposure to this mixture of contaminants may be of greater concern than respiratory pathways in aquatic species.