Although considerable information exists on habitat use by stream salmonids, only a small portion has quantitatively examined diurnal and nocturnal habitat variation. We examined diel variation in habitat use by age-0 and age-1+ brook trout (Salvelinus fontinalis) during summer and autumn in a headwater stream in northern Pennsylvania. Habitat variables measured included cover, depth, substrate, and velocity. The most pronounced diel variation occurred in the use of cover during both seasons. Both age-0 brook trout and age-1+ trout were associated with less cover at night. Age-0 brook trout occupied swifter water during the day than at night during both seasons, but the difference was not significant. Increased cover, depth, and substrate size governed the habitat of age-1+ brook trout. Our findings support the need for a better understanding of diel differences in habitat use of stream salmonids when considering habitat enhancement and protection.