Aims: To determine the incidence of multiple antibiotic-resistant strains of the emergent human pathogens Legionella pneumophila, Pseudomonas aeruginosa and mesophilic Aeromonas species among those isolated from water reservoirs and industrial cooling systems. Methods and Results: Water from four natural water reservoirs and four industrial cooling towers was sampled for 1 year period. The total heterotrophs, mesophilic Aeromonas, Pseudomonas spp. and Legionella spp. counts were performed as recommended by standard procedures, and the sensitivity of the isolates to 27 antibiotics was tested. A total of 117 Aeromonas, 60 P. aeruginosa and 15 L. pneumophila strains were isolated and identified by means of biochemical tests and DNA probes. 46.4% of Aeromonas, and 100% of P. aeruginosa isolates presented multiple resistance. Legionella pneumophila strains were generally sensitive to the drugs used. Conclusions: Antibiotic-resistant pathogenic bacteria belonging to P. aeruginosa and mesophilic Aeromonas species are common in natural aquatic environments. Thus, the risk of waterborne diseases owing to domestic and industrial uses of freshwater should be re-examined from the increase of bacterial resistance point of view. Significance and Impact of the Study: These data confirm the emergence of bacteria resistant to antibiotics in aquatic environments.