Polymerase Chain Reaction-Based Method for the Detection of Canine Heartworm (Filarioidea: Onchocercidae) in Mosquitoes (Diptera: Culicidae) and Vertebrate Hosts

A polymerase chain reaction (PCR)-based test was developed to aid in identification of Dirofilaria sp. and for use in surveys for infected vectors of Dirofilaria immitis (Leidy). A set of PCR primers was designed based on the DNA sequence of a D. immitis surface antigen gene. The predicted product was a 378 base-pair DNA fragment. The target fragment was amplified from free D. immitis larvae (both L3 and microfilariae), individual infected mosquitoes, pools of 30 mosquitoes (including a single infected mosquito), infected mosquitoes stored desiccated at room temperature for 7 mo, and whole blood from a dog infected with D. immitis. These primers did not amplify a homologous fragment from a mermithid or from 4 other species of filarioid nematodes (including 1 other Dirofilaria species). Third-stage larvae from field-collected mosquitoes also were tested. Field-collected L3, identified tentatively as D. immitis, were confirmed by PCR. There was no amplification using the PCR test for L3 identified tentatively as Dirofilaria sp. (possibly D. tenuis Chandler). These data provide a strong indication of the specificity of these primers. The potential utility of this technique for detecting the presence of D. immitis in field populations of mosquitoes is discussed.