The population size of diazotrophic bacteria naturally associated with the maize rhizosphere can be affected by biotic and environmental factors. In this work we have evaluated the effect of two genotypes of maize, with and without nitrogen fertilization, on the population dynamics and distribution of diazotrophic bacteria associated with maize plants over different plant ontogenic stages. The study was carried out in a field experiment with and without nitrogen fertilization, using two maize cultivars (Santa Helena 8447 and Santa Rosa 3063) previously selected from 32 maize cultivars for the lowest and highest response to nitrogen fertilization, respectively. Microbiological and molecular approaches were used to characterize the diazotrophic bacterial population structure. Bacterial population was assessed by the most probable number using semi-solid N-free media, and by DNA isolation from soil and plant tissue followed by amplification of nifH gene fragments using nested PCR, and by RFLP analysis using the restriction endonucleases TaqI and HaeIII. The dynamics of the diazotrophic bacterial population were affected by the ontogenic stage of the maize plants, but not by the cultivar type. Roots were the preferred site of colonization, independent of cultivar type and growth stage. During the first stage of maize growth, addition of nitrogen fertilizer negatively affected the diazotrophic bacterial population.