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Soybean roots are colonized and nodulated by multiple strains of compatible nitrogen-fixing rhizobia primarily belonging to the Genus Bradyrhizobium. Motility towards the root and attachment to root hairs are key determinants of competitive colonization and subsequent nodulation. Bacterial surface properties and motility are known to vary with chemical composition of the culture medium, and root adhesion and nodulation occur in a soil environment rather than laboratory medium. We asked whether the nodulation-promoting factors motility, surface hydrophobicity and surface adhesion of Bradyrhizobium are affected by growth in a soil nutrient environment. B. diazoefficiens USDA 110, 126, 3384, and B. elkanii USDA 26 were grown in mineral salt medium with peptone, yeast extract and arabinose (PSY), and in a soil extracted soluble organic matter (SESOM) medium. Surface hydrophobicity was determined by partitioning into hydrocarbon, motility by transition through soft agar, and surface-exposed saccharides by lectin profiling, followed by biofilm formation and soybean root adhesion capacity of populations. SESOM-grown populations were generally less motile and more hydrophobic. They bound fewer lectins than PSY-grown populations, indicating a simpler surface saccharide profile. SESOM populations of USDA 110 did not form detectable biofilm, but showed increased binding to soy roots. Our results indicate that growth in a soil environment impacts surface properties, motility, and subsequent soy root adhesion propensity. Hence, evaluation of Bradyrhizobium for nodulation efficiency should be performed using soil from the specific field where the soybeans are to be planted, rather than laboratory culture media.