Transgenic tobacco (Nicotiana tabacum) plants of (i) cv. Samsun NN containing the cauliflower mosaic virus 35S constitutive promoter linked to a defective replicase (DR) gene derived from cucumber mosaic virus (CMV) subgroup I isolate Fny, and (ii) cv. Xanthi containing the CaMV 35S promoter linked to the coat protein (CP) gene of CMV subgroup I isolate C were tested for resistance to various Australian isolates of CMV. The tobacco plants were challenged with 3 CMV subgroup I isolates (BNRR, BMR and B6) using sap inoculation. When used to challenge nontransgenic tobacco plants with 5 subgroup II CMV isolates from lupins (LY, LCH, LAcc, LGu and LD), this inoculation method did not result in systemic infection so graft inoculation was used instead to challenge transgenic plants with these 5 isolates. When plants of the line with the DR gene were challenged with the 3 subgroup I isolates, extreme resistance was revealed as none showed symptoms and CMV was not detectable by ELISA. When the same 3 isolates were inoculated to the 3 lines with the CP gene, resistance was characterised by fewer plants becoming virus infected, delayed systemic movement and, in the plants that were infected, partial remission of symptoms plus somewhat decreased virus concentration. Challenge of transgenic plants with DR or CP with the 5 subgroup II isolates resulted in fewer plants becoming infected. Actual numbers of plants infected varied with line and subgroup II isolate and the DR gene was as effective as the CP gene at decreasing infection. With subgroup II isolate LY, infection was associated with remission of symptoms and with the other 4 isolates with delayed systemic movement. Thus the DR gene approach was more effective than the CP approach in obtaining extreme resistance against Australian subgroup I isolates of CMV. These results suggest that introducing a similar DR gene construct made from a subgroup II isolate from lupins into commercial lupin cultivars may be a suitable strategy for obtaining extreme resistance to subgroup II isolates from lupins.