Aims: The aim of this study was to identify the prevalence of environmentally adapted enterococci strains by analysing biochemical fingerprinting (BF) data of 3952 enterococci isolates collected over 5 years from the six catchments in Southeast Queensland, Australia. Methods and Results: A BF method was used to type 3952 enterococci isolates from six catchments. The environmental isolates were compared with a large existing BF library comprised of 5803 enterococci isolates from 10 host groups. Environmental isolates belonged to 801 biochemical phenotypes (BPTs), of which, an average of 29.2% was specific to each catchment. When compared with the BF library, an average of 79.5% BPTs from each catchment was identical to those in the library (i.e. host-origin BPTs). The remaining 20.5% was regarded as non-host origin BPTs, as they were not in the library and constituted only 5.3% of the total isolates tested for each catchment. Conclusions: Our data suggest that less than 5% of studied environmental strains was not identical to those in the library and seemed to be of environmental origin. From a microbial source tracking context, such low level of environmentally adapted strains can have a minimal impact on the performance of the library-based methods if a large number of isolates were tested from both the host groups and environmental waters. Significance and Impact of the Study: These data shed light on the importance of the size and representativeness of library-based source-tracking methods and their implications for the identification of faecal pollution in environmental waters.