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Natural wildfire regimes are important for ecosystem succession but can have negative ecological effects depending on fire characteristics. A portion of a granite rock barrens landscape that extends along the eastern shoreline of Georgian Bay, Lake Huron to eastern Ontario, Canada, burned in 2018 during a wildfire that affected >11,000 ha. This landscape is a biodiversity hotspot providing habitat for many species at risk where freshwater turtles nest in soil deposits in cracks and crevices in the bedrock dominated by moss (Polytrichum spp.) and lichen (Cladonia spp.) cover. To assess the initial effect of wildfire on freshwater turtle nesting habitat, we measured soil depths and estimated moss, lichen, and vascular plant cover at 2 morphology types (crevice, flat) in burned and unburned areas of the landscape. The probability that burned flat plots supported soil was near zero; the burned flat plots had 98% less soil volume compared to unburned flat plots. Although crevices were more resistant to soil loss, burned crevices still had a 15% lower probability of having soil and 35% less soil volume compared to unburned crevice plots. We estimated nest site availability by calculating the number of locations with shallow (5–10 cm), intermediate (10–20 cm), and deep (>20 cm) soils required for a small (5 cm × 5 cm) or medium (10 cm × 10 cm) nest chamber. Overall, the burned open rock barrens had 71–73% fewer sites with suitable soil depth and volume for a nest chamber of either size. Furthermore, burned plots had almost no lichen and moss cover but were dominated by bare soil, forbs, and jack pine (Pinus banksiana) seedlings. Although the loss of tree cover in previously forested areas may increase nest site availability for freshwater turtles in newly open areas, we suggest that organic soil combustion and soil erosion may require restoration activities in the post‐fire landscape to support successful nesting of at‐risk turtles. © 2020 The Wildlife Society. After an 11,000‐ha fire in a rock barrens landscape, the availability of nesting habitat for freshwater turtles was lower at burned compared to unburned sites, and the suitability of remaining potential nest sites was likely negatively affected through alterations in soil properties. Extensive organic soil combustion and soil erosion, leading to loss and alteration of nesting habitat, may require restoration activities in post‐fire landscapes to support successful nesting of at‐risk turtles. [ABSTRACT FROM AUTHOR]