A combination of cultivation-based methods with a molecular biological approach was used to investigate whether isolated bacteria with identical 16S rRNA gene sequences can represent distinct eco – and – genotypes. A set of 8 strains of Pseudomonas putida and Actinetobacter calcoaceticus were isolated from mushroom casing compost community by conventional plating or by using a liquid most probable number (MPN) dilution series. All the 8 strains showed genetic diversity but each strain utilized a specific combination of 31 carbon substrates, and the niche overlap indices were low, suggesting that each strain occupied a different ecological niche. Our results concluded that the extent of physiological diversity masked by identical partial 16S rRNA sequences is much large and that this so-called micro diversity has ecological relevance.