The first five viral haemorrhagic septicaemia virus (VHSV) isolates found in Spain were examined for in vitro growth characteristics, neutralisation by trout antiserum and immunisation challenge of trout fingerlings by water immersion. The viruses had come from different host species in various geographical locations in different years. Three reference VHSV serotypes (F1, F2 and 23.75) were also included in the study. There appeared to be little relationship between the in vivo delay of mortality or the protection of the immunised trout after challenge and the in vitro characteristics studied. In contrast to the in vitro results, the in vivo delay of mortality suggested a closer relationship of the Spanish VHSV isolates to the F2 serotype than to the F1 or the 23.75 serotypes. If the final protection figures are analysed, however, there could be three or four groups of viruses.