Transgenic rodent Plasmodium berghei parasites as tools for assessment of functional immunogenicity and optimization of human malaria vaccines.

Functional assessments of immune responses against stagespecific antigens of Plasmodium falciparum, the agent of the deadliest form of human malaria, are limited mainly to in vitro assays because P. falciparum cannot infect small animals, such as rodents. Hence, antibodies from individuals who have been naturally infected or vaccinated against malaria during clinical trials have been evaluated by enzyme-linked immunosorbent assays and, more functionally, by inhibition of liver-stage development assays (ILSDA), growth inhibition assays (GIA), or membrane feeding assays (MFA). Although the results obtained by these methods have been informative, these techniques remain in vitro assays that may not provide full explanations of the functional relevance and effectiveness of vaccine-elicited immune responses. Transgenic murine parasites expressing human malaria antigens have been developed to facilitate the optimization of the immunogenicity of vaccines and employed to directly address fundamental questions about the biological role of antibodies in vivo. ASSESSMENT OF VACCINE-INDUCED IMMUNITY A tremendous amount of effort has been exerted in the past two decades in identifying, characterizing, and testing various stage-specific malaria antigens as potential vaccine candidates. As a result, a number of candidate vaccines have undergone phase I and phase II clinical trials, with promising results. Some of the antigens (30) that have been well-characterized include circumsporozoite protein (CS) and thrombospondinrelated anonymous protein, expressed in sporozoites; merozoite surface protein 1 (MSP-1), apical membrane antigen 1, and erythrocyte binding antigen 175, expressed on asexual blood stages; and P25, P48/45, and P230, expressed on the sexual stages of the parasites. Though much is now known about the mechanisms of immunity and immune responses to some of these candidate vaccines, the main challenge that has been encountered in evaluating the functional in vivo efficacy of vaccine-induced immune responses is the lack of suitable small-animal models. By and large, most of the available assays are only in vitro surrogates, such as ILSDA (8) and hepatic invasion assays (17, 21, 31) for assessing neutralizing antisporozoite immunity, GIA (22, 24, 25, 32, 33) for assessing functional immune responses to asexual parasite stages, and MFA (3, 14, 18) for measuring immune responses against surface antigens present on gametocytes, gametes, or ookinete stages of the parasite. Nonhuman primates such as Aotus and Saimiri monkeys can be infected with adapted human malaria parasites, enabling the assessment of functional immune responses against malaria antigens in vivo; however, these animal models are not widely accessible, and the cost of maintaining primates is a limiting factor (38). The availability of suitable smallanimal models for the in vivo assessment of vaccine-induced functional immune responses may play a significant role in the development and functional assessment of vaccines against human malaria. High-efficiency transfection protocols (11) have enabled the transfer of genes from human malaria parasites into rodent malaria parasites with relative ease, and here, we discuss and review the potential and feasibility of using such transgenic parasites (Table 1) in assessing antibody responses to various human malaria parasite stage-specific target antigens.