Safety and Reproducibility of a Clinical Trial System Using Induced Blood Stage Plasmodium vivax Infection and Its Potential as a Model to Evaluate Malaria Transmission

Abstract

Background: Interventions to interrupt transmission of malaria from humans to mosquitoes represent an appealing approach to assist malaria elimination. A limitation has been the lack of systems to test the efficacy of such interventions before proceeding to efficacy trials in the field. We have previously demonstrated the feasibility of induced blood stage malaria (IBSM) infection with Plasmodium vivax. In this study, we report further validation of the IBSM model, and its evaluation for assessment of transmission of P. vivax to Anopheles stephensi mosquitoes.

Methods: Six healthy subjects (three cohorts, n = 2 per cohort) were infected with P. vivax by inoculation with parasitized erythrocytes. Parasite growth was monitored by quantitative PCR, and gametocytemia by quantitative reverse transcriptase PCR (qRT-PCR) for the mRNA pvs25. Parasite multiplication rate (PMR) and size of inoculum were calculated by linear regression. Mosquito transmission studies were undertaken by direct and membrane feeding assays over 3 days prior to commencement of antimalarial treatment, and midguts of blood fed mosquitoes dissected and checked for presence of oocysts after 7-9 days.

Results: The clinical course and parasitemia were consistent across cohorts, with all subjects developing mild to moderate symptoms of malaria. No serious adverse events were reported. Asymptomatic elevated liver function tests were detected in four of six subjects; these resolved without treatment. Direct feeding of mosquitoes was well tolerated. The estimated PMR was 9.9 fold per cycle. Low prevalence of mosquito infection was observed (1.8%; n = 32/1801) from both direct (4.5%; n = 20/411) and membrane (0.9%; n = 12/1360) feeds.

Conclusion: The P. vivax IBSM model proved safe and reliable. The clinical course and PMR were reproducible when compared with the previous study using this model. The IBSM model presented in this report shows promise as a system to test transmission-blocking interventions. Further work is required to validate transmission and increase its prevalence.

Trial registration: Anzctr.org.au ACTRN12613001008718.

Conflict of interest statement

I have read the journal's policy and the authors of this manuscript have the following competing interests: PG and SE are employed by Q-Pharm Pty Ltd; CO and KI are employed by PATH MVI.

Figures

Fig 1. Study flow diagram.

Fig 2. Most frequent adverse events (AEs) reported in the study due to all causes.

AE are schematized according to their causality and severity. Fever was the most common AE reported (n = 22). The majority of AEs reported were mild. Only 14 severe AEs were reported and all were attributed to malaria. Abbreviations: ALT: alanine amino transferase; AST: aspartate aminotransferase.

Fig 3. Clinical adverse events attributed to malaria infection by severity.

The majority of clinical AEs were mild with only 11 moderate and 5 severe clinical AEs attributable to malaria infection.

Fig 4. Laboratory parameters recorded as adverse events.

All clinically significantly abnormal laboratory parameters are represented according to their severity. All were attributed to malaria infection. There were no significantly abnormal laboratory parameters recorded as AEs attributable to other causes. Of the 25 laboratory parameters that were recorded as AEs, 15 were abnormalities of LFTs, i.e. either an elevation of ALT or AST. The majority of laboratory AEs were moderate in severity with 2 mild and 9 severe. Abbreviations: LDH: lactate dehydrogenase; AST: aspartate aminotransferase; ALT: alanine amino transferase.

Fig 5. Liver function tests.

Levels of (A) ALT (alanine aminotransferase), (B) AST (aspartate aminotransferase) and (C) total bilirubin versus study day for each of the six subjects. The horizontal dotted line indicates normal range. Abbreviations: ULN: upper limit of normal; LLN: lower limit of normal).

Fig 6. Parasitemia and gametocytemia pre and post antimalarial treatment.

(A) Parasitemia as determined by qPCR of the 18S rDNA target. (B) Estimate of gametocytemia as determined by pvs25 qRT-PCR. Closed lines or “pre” depicts subjects from first parasitemia detection to treatment with artemether/lumefantrine on Day 14 as per protocol. Dashed lines or “post” represents clearance of the parasitemia following treatment. Day represents study day with inoculation occurring on Day 0.

Fig 7. Structures observed on mosquito midguts.

(A) and (B) Oocyts observed on midguts of mosquitoes fed with P. vivax infected blood. (C) and (D) Other ovoid structures observed on midguts of mosquitoes fed with P. vivax infected blood. (E) and (F) Midguts of mosquitoes fed with non-infected blood (negative control). No structures were present on the mosquito midgut represented in panel E. Panel F illustrates ovoid structures observed on the mosquito midguts of some negative controls. Magnification: 20x in panels A, C, D and F; 10x in panel B; 4x in panel E.