Identification of novelPlasmodium vivaxproteins associated with protection against clinical malaria (preprint)

As progress towards malaria elimination continues, the challenge posed by the parasite species Plasmodium vivax has become more evident. In many regions co-endemic for P. vivax and Plasmodium falciparum , as transmission has declined the proportion of cases due to P. vivax has increased. Novel tools that directly target P. vivax are thus warranted for accelerated elimination. There is currently no advanced vaccine for P. vivax and only a limited number of potential candidates in the pipeline. In this study we aimed to identify promising P. vivax proteins that could be used as part of a subunit vaccination approach. We screened 342 P. vivax protein constructs for their ability to induce IgG antibody responses associated with protection from clinical disease in a cohort of children from Papua New Guinea. This approach has previously been used to successfully identify novel candidates. We were able to confirm previous results from our laboratory identifying the proteins reticulocyte binding protein 2b and StAR-related lipid transfer protein, as well as at least four novel candidates with similar levels of predicted protective efficacy. Assessment of these P. vivax proteins in further studies to confirm their potential and identify functional mechanisms of protection against clinical disease are warranted.

A comparison of non-magnetic and magnetic beads for measuring IgG antibodies against P. vivax antigens in a multiplexed bead-based assay using Luminex® technology (Bio-Plex® 200 or MAGPIX®) (preprint)

Multiplexed bead-based assays that use Luminex xMAP ® technology have become popular for measuring antibodies against proteins of interest in many fields, including malaria and more recently SARS-CoV-2/COVID-19. There are currently two formats that are widely used: non-magnetic beads or magnetic beads. Data is lacking regarding the comparability of results obtained using these two types of beads, and for assays run on different instruments. Whilst non-magnetic beads can only be run on flow-based instruments (such as the Luminex ® 100/200™ or Bio-Plex ® 200), magnetic beads can be run on both these and the newer MAGPIX ® instruments. In this study we utilized a panel of purified recombinant Plasmodium vivax proteins and samples from malaria-endemic areas to measure P. vivax -specific IgG responses using different combinations of beads and instruments. We directly compared: i) non-magnetic versus magnetic beads run on a Bio-Plex ® 200, ii) magnetic beads run on the Bio-Plex ® 200 versus MAGPIX ® and iii) non-magnetic beads run on a Bio-Plex ® 200 versus magnetic beads run on the MAGPIX ® . We also performed an external validation of our optimized assay. We observed that IgG antibody responses, measured against our panel of P. vivax proteins, were strongly correlated in all three of our comparisons, however higher amounts of protein were required for coupling to magnetic beads. Our external validation indicated that results generated in different laboratories using the same coupled beads are also highly comparable, particularly if a reference standard curve is used.

Serological signatures of SARS-CoV-2 infection: Implications for antibody-based diagnostics (preprint)

Background Infection with SARS-CoV-2 induces an antibody response targeting multiple antigens that changes over time. This complexity presents challenges and opportunities for serological diagnostics. Methods A multiplex serological assay was developed to measure IgG and IgM antibody responses to seven SARS-CoV-2 spike or nucleoprotein antigens, two antigens for the nucleoproteins of the 229E and NL63 seasonal coronaviruses, and three non-coronavirus antigens. Antibodies were measured in serum samples from patients in French hospitals with RT-qPCR confirmed SARS-CoV-2 infection (n = 259), and negative control serum samples collected before the start of the SARS-CoV-2 epidemic (n = 335). A random forests algorithm was trained with the multiplex data to classify individuals with previous SARS-CoV-2 infection. A mathematical model of antibody kinetics informed by prior information from other coronaviruses was used to estimate time-varying antibody responses and assess the potential sensitivity and classification performance of serological diagnostics during the first year following symptom onset. A statistical estimator is presented that can provide estimates of seroprevalence in very low transmission settings. Results IgG antibody responses to trimeric Spike protein identified individuals with previous RT-qPCR confirmed SARS-CoV-2 infection with 91.6% sensitivity (95% confidence interval (CI); 87.5%, 94.5%) and 99.1% specificity (95% CI; 97.4%, 99.7%). Using a serological signature of IgG and IgM to multiple antigens, it was possible to identify infected individuals with 98.8% sensitivity (95% CI; 96.5%, 99.6%) and 99.3% specificity (95% CI; 97.6%, 99.8%). Informed by prior data from other coronaviruses, we estimate that one year following infection a monoplex assay with optimal anti-Stri IgG cutoff has 88.7% sensitivity (95% CI: 63.4%, 97.4%), and that a multiplex assay can increase sensitivity to 96.4% (95% CI: 80.9%, 100.0%). When applied to population-level serological surveys, statistical analysis of multiplex data allows estimation of seroprevalence levels less than 1%, below the false positivity rate of many other assays. Conclusion Serological signatures based on antibody responses to multiple antigens can provide accurate and robust serological classification of individuals with previous SARS-CoV-2 infection. This provides potential solutions to two pressing challenges for SARS-CoV-2 serological surveillance: classifying individuals who were infected greater than six months ago, and measuring seroprevalence in serological surveys in very low transmission settings.