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Dusky-footed wood rats (Neotoma fuscipes Baird) and two species of Peromyscus mice (P. maniculatus Wagner and P. truei Shufeldt) were collected over a 16-month period from three sites in Sonoma County, California. Blood was collected from 93 wood rats and 177 mice and serum or plasma was tested for seroreactivity with Ehrlichia phagocytophila sensu lato (also known as the human granulocytic ehrlichiosis agent). Thirty-five (37.6%) wood rats and 15 (8.5%) mice were seropositive. Positive Neotoma serology by site ranged from 9.4% to 62.1%. Polymerase chain reaction (PCR) testing for the Ehrlichia groESL heat shock operon was performed on all the seropositive and selected seronegative wood rats; 24 (68.6%) seropositive animals were PCR positive. Two seroconversions and no seroreversions were detected among 18 of the seropositive wood rats that were recaptured and tested multiple times (range = 2-6). Fourteen (77.8%) of the 18 were also PCR positive with six of these positive at every testing point (range = 2-6). One wood rat remained serologically and PCR positive in six specimens collected over a 14-month period. One male of 84 questing adult Ixodes pacificus Cooley & Kohls collected was PCR-positive for E. phagocytophila. Borrelia burgdorferi, the agent of Lyme disease, was cultured from ear punch biopsies from six of seven E. phagocytophila seropositive and one of four seronegative wood rats.