Importantly, this vaccine also induced partial cross-species protection against challenge with P. berghei parasites. Sterile protective immunity was also demonstrated with blood-stage vaccines containing plasmepsin-4-deficient P. berghei parasites although the mechanisms of protective immunity were not determined [34]. Many studies have shown that powerful CD8+ T-cell responses are associated with protection induced by vaccination with whole attenuated sporozoites compared with subunit vaccines [35, 36]. While the latter were HSP mutation considered to have more potential, clinical trials have been disappointing. For example, in the latest trial of RTS,s/AS01E,

an efficacy of only 16·8% was observed over the 4-year

follow-up period [37]. By contrast, a recent trial with irradiation-attenuated sporozoites was largely successful, although six doses were required to induce protection [38]. Whole-parasite vaccines have consistently conferred the best immunity, through the development of both strong CD4+ T and CD8+ T-cell responses [35, 39]. The limited success of SAR245409 cell line clinical trials with subunit blood-stage antigens and the polymorphic nature of the candidate vaccine antigens MSP-1, MSP-2 and AMA-1 pose major problems for vaccine development [40]. Moreover, in some clinical trials with MSP-1, MSP-2 and RESA, reduction in parasitaemia was parasite strain specific [41]. Our findings of strong protective immunity in mice vaccinated with whole-parasite vaccines or with semi-purified soluble antigens suggest that mixtures of antigens would induce a strong T-cell response against many antigens and provide the most efficient protective immune responses against infection. This Quinapyramine observation has more recently been substantiated. [42]. Immunization of human volunteers with a small

number of blood-stage parasites followed by drug cure gave protection that was associated with CD4+ and CD8+ T-cell proliferation, IFN-γ and nitric oxide synthase activity in peripheral blood mononuclear cells [43]. The success of this trial led to more experimental studies in mice to determine the correlates of protective immunity. In the most recent studies from Michael Good’s laboratory, immunization with chemically attenuated parasites, or with very low doses of killed blood-stage parasites together with the adjuvant CpG-ODN, gave cross-strain protection in mice through the development of a strong CD4+ T-cell-dependent IFN-γ and nitric oxide response [44, 45]. Although these findings are encouraging and suggest that a similar approach might be considered for human use, vaccines composed of whole blood-stage parasites face major safety concerns.