Histochemical alteration, protein nitration, and antioxidant expression in oysters as early warning indicators for...

Microplastics are widely distributed in aquatic environments and have been detected in numerous organisms. Marine bivalves are among the most affected aquatic organisms by microplastics due to their filter-feeding behavior. In this study, 60 oysters were collected from two sites along the Texas Gulf Coast. Fourier transform infrared (FTIR) spectroscopy and Nile red staining were used to detect microplastics in oyster (Crassostrea virginica) tissues. We compared field and laboratory samples collected after a two-week depuration period to identify differences in the shape and size of microplastics in oyster tissues. Results from the field sites showed a large number of microplastics, mostly polyethylene (PE) fibers (~3-4 µm in size), in the gills and digestive glands. Conversely, laboratory samples showed a notable decrease in microplastic size (~1-2 µm), reinforcing the argument for cleaner waters. We performed immunohistochemical (IHC) analysis to assess the effects of PE fibers on the expression of 3-nitrotyrosine protein (NTP, a biomarker of protein nitration), superoxide dismutase (SOD), and catalase (CAT) in oyster tissues. IHC results revealed higher expression (P < 0.05) of NTP, SOD, and CAT in the tissues of field oysters compared to laboratory oysters, indicating that microplastics induce nitrative stress, which in turn elevates antioxidant enzyme activity in oyster tissues. Using Periodic-acid Schiff staining, we observed less mucus secretion in the tissues of field oysters than in those of laboratory oysters. This occurrence strengthens the argument that microplastics are causing heightened stress and leaving oysters vulnerable in the transport system and in immune function. Biochemical analysis showed significant (P < 0.05) differences in the pH of extrapallial (EF) fluid between field and laboratory oysters, but there were no differences in EF glucose levels. Overall, these findings suggest that PE microplastics cause histochemical and biochemical alterations, increased protein nitration, and increased antioxidant expression, which may impair physiological functions in oysters.