Several unconformities have been previously recognized in the Late Ordovician and early Silurian strata of southern Ontario. We examined the Georgian Bay, Queenston, Whirlpool, Manitoulin, Power Glen, Cabot Head formations and associated unconformities. Detailed sequence stratigraphic and chemostratigraphic analysis of Late Ordovician and early Silurian outcrops between Niagara, New York and Manitoulin Island, Ontario reveals new insights on the timing of the erosional unconformities and the Ordovician–Silurian boundary. We recognize three significant lowstand unconformities in this interval, which are referred to as the Cherokee, S1B and S2 unconformities. Additional small-scale surfaces are present but do not reflect any major change in sea level or pause in sedimentation. Using δ13Ccarb chemostratigraphy, biostratigraphy and sequence stratigraphy we tentatively correlate units and the mentioned unconformities from southern Ontario to other eastern North American sections of comparable age, showing how glacioeustasy had a widespread effect on the deposition and removal of strata in far-field, subtropical basins. The Cherokee Unconformity appears to be a composite erosion surface found across eastern North America that formed during the two or more episodes of glacioeustatic sea level fall in the early to middle Hirnantian. The overlying S1B and S2 unconformities can also be found across eastern North America and appear to be the result of glacioeustatic sea level fall occurring during the early Silurian. Additionally, these new insights on the timing of these erosional unconformities help better constrain the placement of the Ordovician/Silurian Boundary in Ontario.