Streptococcus castoreus is a rarely encountered beta-haemolytic group A Streptococcus with high tropism for the beaver as host. Based on 27 field isolates under study, evidence strongly suggests that S. castoreus behaves as an opportunistic pathogen in beavers. Although it belongs to the resident mucosal microbiota, this Streptococcus species is associated with purulent lesions in diseased animals. With few exceptions, isolates proved to be highly similar in a panel of phenotypic (including biochemistry, resistance pattern, MALDI-TOF mass spectrometry and Fourier transform-infrared spectroscopy) and classic molecular (16S rRNA and sodA gene) analyses, and thus did not show any specific pattern according to host species or spatio-temporal origin. Conversely, S. castoreus isolates were differentiated into a multitude of pulsed-field gel electrophoresis 'pulsotypes' that did not seem to reflect true epidemiologic lineages. In contrast, single reactions of genomic fingerprinting using BOX-, (GTG)5- and RAPD-PCRs revealed at least subclusters with respect to host species, geographic origin or year, and confirmed the co-colonization of individuals with more than one isolate. In addition to isolates from free-ranging Eurasian beavers (Castor fiber), this study includes S. castoreus from captive North American beavers (Castor canadensis) for the first time.