Quantitative Bioluminescent Imaging of Pre-Erythrocytic Malaria Parasite Infection Using Luciferase-Expressing Plasmodium yoelii

Abstract

The liver stages of Plasmodium parasites are important targets for the development of anti-malarial vaccine candidates and chemoprophylaxis approaches that aim to prevent clinical infection. Analyzing the impact of interventions on liver stages in the murine malaria model system Plasmodium yoelii has been cumbersome and requires terminal procedures. In vivo imaging of bioluminescent parasites has previously been shown to be an effective and non-invasive alternative to monitoring liver stage burden in the Plasmodium berghei model. Here we report the generation and characterization of a transgenic P. yoelii parasite expressing the reporter protein luciferase throughout the parasite life cycle. In vivo bioluminescent imaging of these parasites allows for quantitative analysis of P. yoelii liver stage burden and parasite development, which is comparable to quantitative RT-PCR analysis of liver infection. Using this system, we show that both BALB/cJ and C57BL/6 mice show comparable susceptibility to P. yoelii infection with sporozoites and that bioluminescent imaging can be used to monitor protective efficacy of attenuated parasite immunizations. Thus, this rapid, simple and noninvasive method for monitoring P. yoelii infection in the liver provides an efficient system to screen and evaluate the effects of anti-malarial interventions in vivo and in real-time.

Artesunate versus Chloroquine Infection–Treatment–Vaccination Defines Stage-Specific Immune Responses Associated with Prolonged Sterile Protection against Both Pre-erythrocytic and Erythrocytic Plasmodium yoelii Infection

“…1). These cells were cocultured with primary hepatocytes infected with luciferase-expressing P. yoelii spz (24), and parasite development was quantified as previously described (29). We observed that addition of 5 3 10 5 liver CD8 + T cells from CQ-or AS-ITVtreated mice significantly inhibited parasite development (81.2 and 78.5%, respectively) as compared with those from mockimmunized, untreated mice (Fig.…”

“…using an RNeasy kit (Qiagen) following the manufacturer's instructions. cDNA generation and quantitative PCR (qPCR) were performed as previously described (24). The P. yoelii 18S rRNA signal was normalized to that of mouse GAPDH.…”

Immunization of Mice with Live-Attenuated Late Liver Stage-Arresting Plasmodium yoelii Parasites Generates Protective Antibody Responses to Preerythrocytic Stages of Malaria

“…Luciferase activity was measured 42 to 44 h after mosquito bite or i.v. challenge of mice with PyGFP-luc as previously described (34).…”

An Expanding Toolkit for Preclinical Pre-Erythrocytic Malaria Vaccine Development: Bridging Traditional Mouse Malaria Models and Human Trials

“…These approaches are mutually exclusive in the same animal, though, as qPCR analysis requires animal euthanasia and liver extraction prior to blood-stage infection -resulting in a doubling of animal numbers if both liver burden and time to patency are to be measured. Recently, measurement of liver-stage burden without animal sacrifice has become possible through live bioluminescent imaging techniques and the generation of parasites expressing the firefly luciferase gene [34,[66][67][68][69][70]. This approach strikes a balance between sensitivity of measuring liver-stage burden while keeping the animal alive to measure parasite progression to bloodstage patency, thus reducing animal use and also enabling recurrent challenge for investigation of duration of protection and immunological memory.…”