Standardized challenge viruses are essential for the evaluation of Marek's disease (MD) vaccines with many MD challenge preparations consisting of lymphocytes or whole blood from infected birds. Virus present in these preparations is difficult to quantify by tissue culture assays and, therefore, the infectious bird dose and long-term storage viability cannot be assured. We report on the properties of two low-passage virulent Australian MD viruses, the Woodlands No. 1 strain and strain MPF 57. Both strains were isolated in chicken embryo kidney cultures and adapted to grow in chicken embryo fibroblast cultures for a maximum of 14 passages. Both strains could be readily assayed in tissue culture and produced titres of 10(3) to 10(4) 50% tissue culture infectious doses per ml (TCID(50)). Birds inoculated at three different doses were observed over 10 weeks, and tissues examined for gross and histological lesions, bursarbody weight ratios and the presence of viraemia. Tissue culture-grown preparations of both strains were only slightly less virulent than the original lymphocyte challenge material and produced similar pathological responses and around 80% death or gross lesions. From bursarbody weight ratios strain MPF 57 appeared to be more virulent than the Woodlands No. 1 strain.