Western burrowing owls (Athene cunicularia hypugaea) are small, ground-dwelling owls ofwestern North America that frequent prairie dog (Cynomys spp.) towns and other grasslands.As they rely on rodent prey and occupy burrows once or concurrently inhabited by fossorialmammals, the owls often harbor fleas. We examined the potential role of fleas found onburrowing owls in plague dynamics by evaluating prevalence of Yersinia pestis in fleas and inowl blood. During 2012-2013 fleas and blood were collected from burrowing owls in portionsof five states with endemic plague: Idaho, Oregon, Washington, Colorado, and South Dakota.Fleas were enumerated, taxonomically identified, pooled by nest and assayed for Y. pestis usingculturing and molecular (PCR) approaches. Owl blood underwent serological analysis for plagueantibodies and nested PCR for detection of Y. pestis. Of >4750 fleas collected from owls, Pulexirritans, a known plague vector in portions of its range, comprised more than 99.4%. However,diagnostic tests for Y. pestis of flea pools (culturing and PCR) and owl blood (PCR and serology)were negative. Thus, despite that fleas were prevalent on burrowing owls, and the potentialfor a relationship with burrowing owls as a phoretic host of infected fleas exists, we found noevidence of Y. pestis in sampled fleas or in owls that harbored them. We suggest that studiessimilar to those reported here during plague epizootics will be especially useful for confirmingthese results.