Island foxes (Urocyon littoralis) are endemic to six of the eight California Channel Islands (USA). The island fox is classified as a threatened species by the State of California, and recently three of the six subspecies have experienced abrupt population declines. As part of a continuing effort to determine the cause of the declines, we tested island fox serum samples collected in 1988 (n = 176) and 1997–98 (n = 156) over the entire geographic range of the species for seroprevalence of canine heartworm (Dirofilaria immitis) antigen. Using a commercially available enzyme-linked immunosorbent assay (PetChek®, Idexx Laboratories, Westbrook, Maine, USA) we detected heartworm antigen in four of the six populations of island foxes. On San Miguel and Santa Rosa Islands, seroprevalence in adult foxes was >85% (n = 62) in 1988 and increased to 100% (n = 24) in 1997–98. On Santa Cruz Island, seroprevalence in adult foxes decreased from 83% (n = 30) to 58% (n = 26), whereas on San Nicolas Island, seroprevalence increased from 25% (n = 32) to 77% (n = 30) during the same period. All of the pups assayed (n = 33) were seronegative. The seroprevalences of heartworm reported herein for the four populations of island foxes are the highest yet reported for a fox species. However, additional demographic data reported elsewhere suggests that heartworm has not been a major factor in the recent declines of island fox populations.