Isolates were retrieved from drinking water and from animal and human sources associated with a waterborne outbreak of giardiasis. This is the first report of water-source and epidemic-associated Giardia isolates being adapted to in vitro propagation. Outbreak-associated, non-out-break-associated, and reference isolates were characterized using isoenzyme electrophoresis and pulsed-field gel electrophoresis (PFGE). All outbreak-associated and 2 other isolates were in one of eight zymodemes. The chromosomal complement of the outbreak-associated isolates was relatively homogeneous; this PFGE karyotype was distinguishable from other karyotypes. Overall results of both characterization methods were similar, although PFGE appears to be a more discriminating biotyping technique. Banding patterns of the outbreak-associated Giardia isolates remained the same even though the parasite passed through different hosts during the outbreak. Heterogeneity of isolates was also demonstrated for the first time within a single community not associated with the outbreak.