I conducted a study of population subdivision and genetic diversity among populations of the giant river turtle Podocnemis expansa using microsatellite DNA markers. Turtles were sampled from four nesting beaches distributed within 100 km of one another along the Middle Caquetá River in Colombian Amazonia. I detected significant genetic differentiation within the Caquetá basin with Weir and Cockerham's Fst estimator, θ, but not the rho-st estimator of Slatkin's Rst. I also detected differences in allele and genotype frequencies using probability and exact tests. Although significant, population subdivision appears subtle enough to pass undetected by Rst with the sample sizes available. This is presumably due to the relatively higher variance of Rst. The differentiation found among beaches within the Caquetá River is surprising given that individuals are able to migrate more than 400 km between nesting seasons, and females come to nest at the Caquetá River from common tributaries. Potential explanations of the subdivision found are the segregation of females into nesting family groups (including natal homing) or of males into mating family groups. The genetic subdivision has some geographic component such that the three beaches closest genetically are also the closest in distance along the river. I detected strong differentiation between Colombian and Brazilian populations using Rst which I discuss in light of the available information about the biology of P. expansa. I also discuss the implications of these results to conservation issues of this endangered species.