The aim of this study is to identify significant biotic regions (groups of areas with similar biotas) and biotic elements (groups of taxa with similar distributions) for the marsupial fauna in a part of northern South America using physiographical areas as Operational Geographical Units (OGUs). We considered Venezuela a good model to elucidate this issue because of its high diversity in landscapes and the relatively vast amount of information available on the geographical distribution of marsupial species. Based on the presence-absence of 33 species in 15 physiographical sub-regions (OGUs) we identified Operational Biogeographical Units (OBUs) and chorotypes using a quantitative analysis that tested statistical significance of the resulting groups. Altitudinal and/or climatic trends in the OBUs and chorotypes were studied using a redundancy analysis. The classification method revealed four OBUs. Strong biotic boundaries separated: i) the xerophytic zone of the Continental coast (OBU I); ii) the sub-regions north of the Orinoco River (OBU III and IV); and those south to the river (OBU II). Eleven chorotypes were identified, four of which included a single species with a restricted geographic distribution. As for the other chorotypes, three main common distribution patterns have been inferred: i) species from the Llanos and/or distributed south of the Orinoco River; ii) species exclusively from the Andes; and iii) species that either occur exclusively north of the Orinoco River or that show a wide distribution throughout Venezuela. Mean altitude, evapotranspiration and precipitation of the driest month, and temperature range allowed us to characterize environmentally most of the OBUs and chorotypes obtained.