Colonization of glass substrata by populations of three or four bacterial species over periods of 4 weeks or more was investigated using recirculating, model laboratory systems. Numbers of a coryneform,Aeromonas hydrophila, Pseudomonas fluorescens, andXanthomonas maltophiliaon surfaces and in the liquid phase were monitored to determine whether any species inhibited or facilitated the colonization by another organism. Each system contained 800 ml of culture in a peptone/yeast extract/artificial lakewater medium, pulse‐fed every 3 days, and recirculated at 12 ml·min−1. Numbers of each species in the liquid phase and in biofilms that had been removed from the surfaces and dispersed were determined by viable counts on differential media. Biofilm coverage was evaluated by computer‐enhanced microscopy. The coryneform quickly colonized the surfaces, and there was little change in suspended or attached numbers over the experimental period. Colonization byA. hydrophilaincreased in the presence ofP. fluorescens. X. maltophiliawas a highly adhesive strain and promoted microcolony formation on the surface. The influence of nutrient concentration on colonization was determined by testing media at 10−2, 10−1, and 10−1dilutions of the standard concentration. The relative proportions of species both in the liquid and on surfaces altered with different nutrient concentrations, and with increase in nutrient concentration, there was a greater increase in total numbers of bacteria in the liquid (ranging from 7·7 × 106.ml−1to 1·7 × 1010.ml−1) than on the surfaces (ranging from 6·3 × 105.cm−2to 1·2 × 101.cm2). The results indicated that the ability of bacteria to colonize surfaces is to a large extent related to their ability to colonize the liquid phase. However, when a species has strong adhesive characteristics, this can result in relatively greater colonization of surfaces. In addition, synergistic interspecies interactions may allow a poor colonizer to become established.