Southern California remains an important focus of West Nile virus (WNV) activity, with persistently elevated incidence after invasion by the virus in 2003 and subsequent amplification to epidemic levels in 2004. Eco-epidemiological studies of vectors-hosts-pathogen interactions are of paramount importance for better understanding of the transmission dynamics of WNV and other emerging mosquito-borne arboviruses. We investigated vector-host interactions and host-feeding patterns of 531 blood-engorged mosquitoes in four competent mosquito vectors by using a polymerase chain reaction (PCR) method targeting mitochondrial DNA to identify vertebrate hosts of blood-fed mosquitoes. Diagnostic testing by cell culture, real-time reverse transcriptase-PCR, and immunoassays were used to examine WNV infection in blood-fed mosquitoes, mosquito pools, dead birds, and mammals. Prevalence of WNV antibodies among wild birds was estimated by using a blocking enzyme-linked immunosorbent assay. Analyses of engorged Culex quinquefasciatus revealed that this mosquito species acquired 88.4% of the blood meals from avian and 11.6% from mammalian hosts, including humans. Similarly, Culex tarsalis fed 82% on birds and 18% on mammals. Culex erythrothorax fed on both birds (59%) and mammals (41%). In contrast, Culex stigmatosoma acquired all blood meals from avian hosts. House finches and a few other mostly passeriform birds served as the main hosts for the blood-seeking mosquitoes. Evidence of WNV infection was detected in mosquito pools, wild birds, dead birds, and mammals, including human fatalities during the study period. Our results emphasize the important role of house finches and several other passeriform birds in the maintenance and amplification of WNV in southern California, with Cx. quinquefasciatus acting as both the principal enzootic and "bridge vector" responsible for the spillover of WNV to humans. Other mosquito species, such as Cx. tarsalis and Cx. stigmatosoma, are important but less widely distributed, and also contribute to spatial and temporal transmission of WNV in southern California.