Middle Eocene-Oligocene calcareous nannoplankton have been analyzed quantitatively for eight DSDP and ODP sites ranging from the equatorial zone to 65°S latitude in the South Atlantic Ocean. There is a general decrease in calcareous nannoplankton species diversity from the middle Eocene through the Oligocene, and species diversity also decreases towards the higher latitudes. Similar to findings of other studies, the drop of the species diversity of calcareous nannoplankton across the Eocene/Oligocene boundary is not drastic. Delineation of latitudinal distribution patterns of individual species or groups and R-mode cluser analysis permit the classification of warm-water taxa (Coccolithus formosus, discoasters, helicosphaerids, and sphenoliths), temperate-water taxa (Coccoliths pelagicus, Cyclicargoliths floridanus group, Reticulofenestra bisecta, Reticulofenestra samodurovii/Reticulofenestra umbilica), and cool-water taxa (chiasmoliths, Isthmolithus recurvus, and Reticulofenestra daviesii). The abundance of warm-water taxa plummets whereas the abundance of cool-water taxa sharply increases across the Eocene/Oligocene boundary at the middle- and high-latitude sites, reflecting a profound cooling event in these areas. Polar ordination and similarity analyses reveal that steep latitudinal biogeographic gradients had been established at least by the middle Eocene. The gradient greatly increases near the Eocene/Oligocene boundary, indicating more severe and permanent cooling at the high latitudes. The biogeographic gradient data of this study conflict with the widely accepted inference from the oxygen isotopic data that the thermal gradients between mid latitudes and high latitudes are low or nearly flat for the Paleogene oceans. Lower surface water salinities in the high latitudes may have lowered the δ18O values of the planktonic microfossils, but apparently did not have significant effect on latitudinal distribution patterns of calcareous nannoplankton, which offers an independent means for estimating latitudinal thermal gradients. © 1990.