A current goal of malaria vaccine research is the development of vaccines that will cross-protect against multiple strains of malaria. Inthe present study, the breadth of cross-reactivity induced by a 30K multivalent DNA vaccine has been evaluated in susceptible A/J mice(H-2a) against infection with the Plasmodium chabaudi adami DK strain and a virulent parasite subspecies, Plasmodium chabaudi chabaudi AS. Immunized A/J mice were significantly protected against infection with both P. c. adami DK (31'40% reduction in cumulativeparasitemia) and P. c. chabaudi AS parasites, where a 30'39% reduction in cumulative parasitemia as well as enhanced survival wasobserved. The 30K vaccine-induced specific IFN-c production by splenocytes in response to native antigens from both P. c. chabaudi AS and P. c. adami DK. Specific antibodies reacting with surface antigens expressed on P. c. adami DS and P. c. chabaudi AS infected red blood cells, and with opsonizing properties, were detected. These results suggest that multivalent vaccines encoding conserved antigenscan feasibly induce immune cross-reactivity that span Plasmodium strains and subspecies and can protect hosts of distinct major histocompatibility complex haplotypes.