Sediment bacteria are ubiquitous and important to the cycling of organic matter and nutrients in streams. We experimentally investigated the effect of sediment grain size on bacterial communities by using artificial substrata of different sizes incubated in a northeast Ohio (USA) stream. We examined abundances of specific bacterial taxa with fluorescent in situ hybridization. In general, taxon abundances were highest on 5.0-mm particles and lowest on 0.1-mm particles, and all taxa responded similarly to particle size. Few differences in abundance were found between substrata incubated in a pool or riffle in the stream or between substrata with or without organic matter amendment. Particle surface area and particle packing (sediment permeability) both affected bacterial abundance in our experiments. However, our results suggest that other factors, such as chemistry of interstitial spaces and susceptibility to predation, also contribute to differences in abundance among substrata of different particle sizes.