The increase of nutrients in water bodies, in particular nitrogen (N) and phosphorus (P) due to the recent expansion of agricultural and other human activities is accelerating environmental degradation of these water bodies, elevating the risk of eutrophication and reducing biodiversity. To evaluate the ecological effects of the influx of nutrients in an oligotrophic and stoichiometrically imbalanced environment, we performed a replicated in situ mesocosm experiment. We analyzed the effects of a N- and P-enrichment on the bacterial interspecific interactions in an experiment conducted in the Cuatro Cienegas Basin (CCB) in Mexico. This is a desert ecosystem comprised of several aquatic systems with a large number of microbial endemic species. The abundance of key nutrients in this basin exhibits strong stoichiometric imbalance (high N:P ratios), suggesting that species diversity is maintained mostly by competition for resources. We focused on the biofilm formation and antibiotic resistance of 960 strains of cultivated bacteria in two habitats, water and sediment, before and after 3 weeks of fertilization. The water habitat was dominated by Pseudomonas, while Halomonas dominated the sediment. Strong antibiotic resistance was found among the isolates at time zero in the nutrient-poor bacterial communities, but resistance declined in the bacteria isolated in the nutrient-rich environments, suggesting that in the nutrient-poor original environment, negative inter-specific interactions were important, while in the nutrient-rich environments, competitive interactions are not so important. In water, a significant increase in the percentage of biofilm-forming strains was observed for all treatments involving nutrient addition.