Since 1978, the Lake Ontario preyfish community survey has provided information on the status and trends of the benthic preyfish community related to Fish Community Objectives that includes understanding preyfish population dynamics and community diversity. Beginning in 2015, the benthic preyfish survey expanded from US-only to incorporate lake-wide sampling sites which increased the survey’s spatial coverage, and resumed sampling in eastern embayments (Black River, Chaumont, Guffin, and Henderson Bays) that were historically sampled during a September bottom trawl survey to index yellow perch from 1978 to 2007. In 2021, the collaborative benthic preyfish survey completed 195 bottom trawl tows across main lake and embayments at depths from 5 to 226 m. New embayment sites at Bay of Quinte, Sodus, and Little Sodus Bay were added to the survey in 2021 to compare fish communities across nearshore sites. In total, the 2021 survey sampled 109,178 fish from 35 species. Round goby (Neogobius melanostomus) was the most numerically abundant species comprising 44% of the total catch, followed by deepwater sculpin (Myoxocephalus thompsonii), and alewife (Alosa pseudoharengus) at 17% and 11%, respectively. Deepwater sculpin accounted for most (406 kg) of the fish biomass sampled during the 2021 survey (total=1,995 kg), followed by round goby (257 kg), and common carp (252 kg). Slimy sculpin (Cottus cognatus) biomass was higher in 2021 than in 2020, when spatial coverage was reduced. Deepwater sculpin biomass remained high in 2021 and similar to observations since 2019. White perch biomass (Morone americana) in Black River Bay has increased compared to observations from historical surveys. Yellow perch (Perca flavescens) accounted for most of the benthic preyfish biomass across the embayments surveyed in 2021 except for the Bay of Quinte and Black River Bay, where white perch accounted for a greater proportion of the fish community biomass.