Oulton, T. | bioRxiv Science

bioRxiv2023

Repeated Plasmodium vivax blood stage infection provides sterile protection against homologous challenge in non-human primates

The malaria parasite Plasmodium vivax remains a major global public health challenge, causing major morbidity across tropical and subtropical regions. Several candidate vaccines are in preclinical and clinical trials, however no vaccine against P. vivax malaria is approved for use in humans. Here we assessed whether P. vivax strain-transcendent immunity can be achieved by repeated infection in Aotus monkeys. For this purpose, we repeatedly infected six animals with blood stages of the P. vivax Salvador 1 (SAL-1) strain until sterile immune, and then challenged with the AMRU-1 strain. Sterile immunity was achieved in 4/4 Aotus monkeys after two homologous infections with the SAL-1 strain, while partial protection against a heterologous AMRU-1 challenge (i.e., delay to infection and reduction in peak parasitemia compared to control) was achieved in 3/3 monkeys. IgG levels based on P. vivax lysate ELISA and protein microarray increased with repeated infections and correlated with the level of homologous protection. Analysis of parasite transcriptional profiles across inoculation levels provided no evidence of major antigenic switching upon homologous or heterologous challenge. In contrast, we observed significant transcriptional differences in the P. vivax core gene repertoire between SAL-1 and AMRU-1. Together with the strain-specific genetic diversity between SAL-1 and AMRU-1 these data suggest that the partial protection upon heterologous challenge is due to molecular differences between strains (at genome and transcriptome level) rather than immune evasion by antigenic switching. Our study demonstrates that sterile immunity against P. vivax can be achieved by repeated homologous blood stage infection in Aotus monkeys, thus providing a benchmark to test the efficacy of candidate blood stage P. vivax malaria vaccines. Author summaryPlasmodium vivax is the most widespread human malaria parasite. Elimination efforts are complicated due to the peculiar biology of P. vivax including dormant liver forms, cryptic reservoirs in bone marrow and spleen and a large asymptomatic infectious reservoir in affected populations. Currently there is no vaccine against malaria caused by P. vivax. Here we induce sterile immunity by repeated P. vivax infection with the SAL-1 strain in non-human primates. In contrast, heterologous challenge with the AMRU-1 strain only provided partial protection. Antibody levels against a crude antigen and a protein microarray correlated with the level of homologous protection. Parasite transcriptional profiles across inoculation levels failed to show major antigenic switching across SAL-1 infections or upon heterologous challenge, instead suggesting other mechanisms of immune evasion. Our study demonstrates that sterile immunity against P. vivax can be achieved by repeated blood stage infection in Aotus monkeys, thus providing a benchmark to test the efficacy of candidate blood stage P. vivax malaria vaccines.

microbiology↗

bioRxiv2020

Plasmodium falciparum serology: A comparison of two protein production methods for analysis of antibody responses by protein microarray

The evaluation of protein antigens as putative serologic biomarkers of infection has increasingly shifted to high-throughput, multiplex approaches such as the protein microarray. In vitro transcription/translation (IVTT) systems - a similarly high-throughput protein expression method - are already widely utilised in the production of protein microarrays, though purified recombinant proteins derived from more traditional whole cell based expression systems also play an important role in biomarker characterisation. Here we have performed a side-by-side comparison of antigen-matched protein targets from an IVTT and purified recombinant system, on the same protein microarray. The magnitude and range of antibody responses to purified recombinants was found to be greater than that of IVTT proteins, and responses between targets from different expression systems did not clearly correlate. However, responses between amino acid sequence-matched targets from each expression system were more closely correlated. Despite the lack of a clearly defined relationship between antigen-matched targets produced in each expression system, our data indicate that protein microarrays produced using either method can be used confidently, in a context dependent manner, though care should be taken when comparing data derived from contrasting approaches. Statement of significance of the studyProtein microarray technology is increasingly being realised as a powerful tool in disease biomarker identification. Protein-based, serologic arrays are already well utilised in the characterisation of antibody responses to the malarial Plasmodium spp., drastically improving throughput and efficiency compared to more typical experimental assays. Such approaches have commonly made use of in vitro transcription/translation (IVTT) protein expression systems, though other protein expression methods are regularly employed. In this study we have directly compared antibody responses in a malaria endemic population, to matched protein antigens derived from both an IVTT and purified bacterial recombinant system in the same protein microarray. We demonstrated that the magnitude of measured antibody response to matched protein antigens tends to be greater to purified recombinants, rather than IVTT products. Further, our analysis showed highly variable levels of correlation of response between antigen-matched targets derived from each expression system, although relationships in antibody response were stronger between proteins with overlapping amino acid sequences. This study highlights the importance of considering the strengths and weaknesses of each expression system in the context of a protein microarray according to experimental hypotheses, and illustrates the need for attention when comparing data generated by these different methodologies.

immunology↗