The study reported here evaluated the distribution, relative abundance, and malaria transmission potential of Anopheles mosquitoes at 30 sites representing different ecological strata in western Kenya. Seasonal variation in anopheline densities and transmission potential, as expressed by Entomological inoculation rates (EIR), was investigated. Of the 6491 indoor resting anopheline mosquitoes collected at the 30 sites, 91.3 % (n = 5926) were An. gambiae s.l. and 8.7 % (n = 565) were An. funestus with an average house density of 6.58 and 0.63, respectively. Analysis of the data indicated significant variation in mosquito densities between study sites, species and season. High densities of both An. gambiae and An. funestus were recorded in the northern and northeastern parts of the district, while generally low densities were recorded in the south. Anopheles gambiae s.s. and An. arabiensis comprised 60.3 % (n = 3573) and 39.7 % (n = 2352) of the total number of An. gambiae s.l. mosquitoes collected, respectively. The composition of the An. gambiae s.l. sibling species showed temporal and spatial variation. Entomologic inoculation rates were estimated at 1.55 and 0.12 infective bites per person per month for An. gambiae s.l. and An. funestus, respectively. This study reveals considerable seasonal and site-specific variation in vector distribution, composition and transmission potential. Application of control interventions must therefore consider seasonal variations since the vectorial system changes quite rapidly over a short period of time.