We examined mitochondrial control-region haplotype diversity and allelic frequency distributions for three polymorphic microsatellite loci in 541 coastalOncorhynchus mykisscollected from six habitats associated with different levels of human activity and ocean access in southern California. Extensive urbanization, climatic unpredictability, and the accelerated rate of decline in anadromous fish suggested a probable loss of genetic diversity in this area due to habitat fragmentation, geographic isolation, and population bottlenecks. Unexpectedly high levels of genetic diversity were found in southern California populations ofO. mykiss. Haplotype diversity (HS) was highest in anadromous fish (HS= 0.74) and lowest in the Whale Rock Hatchery trout (HS= 0.32). The proportion of variation attributable to population differentiation among habitat groups (GST) was 10%. Haplotype frequencies showed a close relationship between anadromous steelhead and resident rainbow trout from closed habitats (DST= 0.03). Combined microsatellite allelic diversity (at lociOmy77,Omy207, andSsa289) was highest in rainbow trout from closed habitats (88%), and lowest in Whale Rock Hatchery fish (29%). Greatest microsatellite distance (δμ = 17.1) was between anadromous steelhead and reservoir rainbow trout, and closest identity (δμ, = 1.8) was among rainbow trout from closed habitats, hatchery rainbow trout, and reservoir rainbow trout. Analysis of genetic distance measures for both molecular markers showed that considerations of life history patterns and freshwater habitats that retain ocean access remain important factors in the preservation of the unique genetic diversity found in southern California coastalO. mykiss.