Species compositions of Culicoides paraensis (Goeldi) (Diptera: Ceratopogonidae), the major vector of Oropouche virus to humans in Central and South American urban cycles, and Culicoides insinuatus Ortiz & Leon differed along a northeast-to-southwest transect across Iquitos, Department of Loreto, Peru. The relative distributions of the species were consistent with patterns of human outbreaks along the Amazon River. We resumed collection of biting midges between May 2000 and January 2004 at three sites previously sampled (1996–1997) to determine whether the known vector was expanding its range relative to the earlier survey. C. paraensis did not replace C. insinuatus across the region surveyed. Instead, C. insinuatus dominated the more southern sites and significantly increased its relative proportion at all three sites. Apparently, microhabitat differences and not range expansion by C. paraensis were responsible for differences in species compositions across the sample sites.