This study aimed to determine the microplastic prevalence in eastern oysters (C. virginica) in three sites in the Chesapeake Bay in Virginia and optimize the digestion methods. The digestion results illustrate that the lowest recovery rate and digestion recovery were related to enzymatic, enzymatic + hydrogen peroxide (H₂O₂), and HCl 5% treatments, while the highest digestion recovery and recovery rate were observed in H₂O₂ and basic (KOH) treatments. Nitric acid digestion resulted in satisfying digestion recovery (100%), while no blue polyethylene microplastics were observed due to the poor recovery rate. In addition, nitric acid altered the color, changed the Raman spectrum intensity, and melted polypropylene (PP) and polyethylene terephthalate (PET). In order to determine the number of microplastics, 144 oysters with an approximately similar size and weight from three sites, including the James River, York River, and Eastern Shore, were evaluated. Fragments were the most abundant microplastics among the different microplastics, followed by fibers and beads, in the three sites. A significantly higher number of fragments were found in the James River, probably due to the greater amount of human activities. The number of microplastics per gram of oyster tissue was higher in the James River, with 7 MPs/g tissue, than in the York River and Eastern Shore, with 6.7 and 5.6 MPs/g tissue.