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Approximately 600 km of high-resolution seismic reflection data were collected to investigate the late-Quaternary stratigraphic development of Lake Titicaca. The focus of this report is on two seismic sequence boundaries, which are interpreted as erosional surfaces formed at times of low lake level. The younger erosional surface occurs as much as 90 m below the present lake level and up to 8 m below the present sediment–water interface. This erosional surface is interpreted to be coeval with a well-documented early- to mid-Holocene lowstand, dated between ~8,000 and 3,600 cal yr BP. An earlier and previously unknown erosional surface occurs at a sub-bottom depth of approximately 30 m, and as much as 240 m below the present lake level, which implies a major late-Pleistocene lowstand of Lake Titicaca. By extrapolation of sedimentation rates from the upper ~14 m of sediment, we estimate the age of this older lowstand at >90,000 cal yr BP. Both lowstands of Lake Titicaca indicated by the seismic data are likely to have been a response to climatic change in the region.