Adaptation to a specific niche theoretically constrains a population's ability to subsequently diversify into other niches. We tested this theory using the bacterium Pseudomonas fluorescens, which diversifies into niche specialists when propagated in laboratory microcosms. Numerically dominant genotypes were allowed to diversify in isolation. As predicted, populations increased in fitness through time but showed a greatly decreased ability to diversify. Subsequent experiments demonstrated that niche generalists and reductions in intrinsic evolvability were not responsible for our data. These results show that niche specialization may come with a cost of reduced potential to diversify.