As rainfall in South-Central Chile has decreased in recent decades, local communities and industries have developed an understandable concern about their threatened water supply. Reconstructing streamflows from tree-ring data has been recognized as a useful paleoclimatic tool in providing long-term perspectives on the temporal characteristics of hydroclimate systems. Multi-century long streamflow reconstructions can be compared to relatively short instrumental observations in order to analyze the frequency of low and high water availability through time. In this work, we have developed a Biobío River streamflow reconstruction to explore the long-term hydroclimate variability at the confluence of the Mediterranean-subtropical and the Temperate-humid climate zones, two regions represented by previous reconstructions of the Maule and Puelo Rivers, respectively. In a suite of analyses, the Biobío River reconstruction proves to be more similar to the Puelo River than the Maule River, despite its closer geographic proximity to the latter. This finding corroborates other studies with instrumental data that identify 37.5°S as a latitudinal confluence of two climate zones. The analyzed rivers are affected by climate forcings on interannual and interdecadal time-scales, Tropical (El Niño Southern Oscillation) and Antarctic (Southern Annular Mode; SAM). Longer cycles found, around 80-years, are well correlated only with SAM variation, which explains most of the variance in the Biobío and Puelo rivers. This cycle also has been attributed to orbital forcing by other authors. All three rivers showed an increase in the frequency of extreme high and low flow events in the twentieth century. The most extreme dry and wet years in the instrumental record (1943–2000) were not the most extreme of the past 400-years reconstructed for the three rivers (1600–2000), yet both instrumental record years did rank in the five most extreme of the streamflow reconstructions as a whole. These findings suggest a high level of natural variability in the hydro-climatic conditions of the region, where extremes characterized the twentieth century. This information is particularly useful when evaluating and improving a wide variety of water management models that apply to water resources that are sensitive to agricultural and hydropower industries.