To characterize the etiologic agent (WA1) of the first reported case of babesiosis acquired in Washington State. Case report, and serologic, molecular, and epizootiologic studies. South-central Washington State. A 41-year-old immunocompetent man with an intact spleen who developed a moderately severe case of babesiosis. Serum specimens from the patient were assayed by indirect immunofluorescent antibody (IFA) testing for reactivity with seven Babesia species and with WA1, which was propagated in hamsters inoculated with his blood. A Babesia-specific, ribosomal-DNA (rDNA) probe was hybridized to Southern blots of restriction-endonuclease-digested preparations of DNA from WA1, Babesia microti, and Babesia gibsoni. Serum specimens from 83 family members and neighbors were assayed for IFA reactivity with WA1 and B. microti. Small mammals and ticks were examined for Babesia infection. The patient's serum had very strong IFA reactivity with WA1, strong reactivity with B. gibsoni (which infects dogs), but only weak reactivity with B. microti. DNA hybridization patterns with the rDNA probe clearly differentiated WA1 from B. gibsoni and B. microti. Four of the patient's neighbors had IFA titers to WA1 of 256. The tick vector and animal reservoir of WA1 have not yet been identified, despite trapping 83 mammals and collecting 235 ticks. WA1 is morphologically indistinguishable but antigenically and genotypically distinct from B. microti. Some patients elsewhere who were assumed to have been infected with B. microti may have been infected with WA1. Improved serodiagnostic and molecular techniques are needed for characterizing Babesia species and elucidating the epidemiology of babesiosis, an emergent zoonosis.