We examined latitudinal patterns of generic richness in stream insects across the Mackenzie River system in northern Canada. We focused on low- to mid-order tributaries flowing into the river and spanning ∼11° of latitude. Physical-habitat characteristics and water chemistry were examined as factors affecting diversity both at the local and regional scale. No overall latitudinal trend was found in richness, but trends were apparent at the order level. Local generic richness of Ephemeroptera and Plecoptera declined toward the north, Trichoptera generic richness was lowest in the middle of the latitudinal range, and Diptera generic richness increased with latitude. Most orders of aquatic insects followed the expected decline in richness with increasing latitude, but the family Chironomidae was an exception, probably because of their ability to tolerate subarctic environmental conditions and because of the relative absence of potential predation and competition from other invertebrates. Streams were characterized along principal component axes corresponding to stream size and terrain, substrate, water source, and turbidity. Most groups were slightly more diverse in larger streams. Ephemeroptera and Plecoptera had higher richness on coarse substrates, and Trichoptera had lower richness in streams in the Norman Range that possessed unique hydrology and water-chemistry characteristics because of the presence of taliks. Chironomid richness was positively correlated with latitude and stream size. Latitudinal gradients in regional diversity largely followed the trends in local richness, but all taxa had small regional numbers of genera near the middle of the latitudinal range, and a decline in overall aquatic insect richness between the south and north was more apparent at the regional scale. We conclude that regional environmental factors control the maximum richness that can be expected in a locality at a given latitude, whereas local environmental factors control the number of taxa present at a site. Spatial gradients in benthic insect richness are likely to affect the applicability of biomonitoring metrics that use groups whose richness varies with latitude.