Abstract: Little is known about the microbial community structure associated with soil aggregation in microaggregates (0.25–0.05 mm) that are held within macroaggregates (>0.25 mm). We investigated the effects of irrigation, tillage, and cropping system on aggregate distribution and the community structure of the predominant culturable bacteria in microaggregates held within macroaggregates from 2005 to 2008 at a site in western North Dakota. Managements were malt barley (Hordeum vulgare L.) under no-till and conventional till and an established no-tilled Conservation Reserve Program (CRP) plantation on Lihen loam (sandy, mixed, frigid, Entic Haplustolls) under irrigated and dryland systems. We predicted that microaggregates held within macroaggregates of irrigated no-till barley and CRP management have a higher proportion of the predominant culturable bacterial species that have the capability to aggregate soil than in irrigated and nonirrigated conventional till barley managements. Fatty acid profiling and DNA sequencing were used to identify bacterial isolates and a soil sedimentation assay to determine their soil-aggregating ability. The proportion of soil aggregating Gram-negative bacteria, dominated by pseudomonads and Stenotrophomonas maltophilia, was higher under irrigated no-till barley and CRP compared with the other managements. Species abundance and diversity of the soil-aggregating bacterial isolates were the highest in irrigated no-till barley and CRP and the lowest in irrigated conventional till barley. We demonstrated that, under irrigated management with high residue input, microaggregates held within macroaggregates represent a distinct environment that selects for specific soil-binding microbial communities that may contribute to soil structural stability at the 0- to 5-cm depth.