Correction: Vaccine Strain-Specificity of Protective HLA-Restricted Class 1 P. falciparum Epitopes.

[This corrects the article DOI: 10.1371/journal.pone.0163026.]

ELISpot and CD8+ T cell IFN-γ responses of DNA/HuAd5 and HuAd5 immunized subjects to <i>P</i>. <i>falciparum</i> strains 3D7 and 7G8 AMA1 A*03 protective epitopes.

<p>ELISpot and CD8+ T cell IFN-γ activities are shown in Panels A–D. <b>Panel A:</b> ELISpot IFN-γ response of the A*03 protected subject (v11) are positive with Ap8 and the 3D7 A*03 epitope but not the 7G8 epitope (arrow). <b>Panel B:</b> ELISpot activity of v11 is not affected by CD4+-depletion but is abolished after CD8+ depletion (arrow). <b>Panel C:</b> CD8+ T cell IFN-γ responses of v11 are much higher (p = 0.001) to the 3D7 epitope than to the 7G8 epitope (arrow). <b>Panel D:</b> ELISpot IFN-γ responses of two of four non-protected subjects from the HuAd5 trial were weakly positive with the 3D7 epitope but all four subjects were negative with the 7G8 epitope (arrows).</p

Co-expression of Interleukin-15 Enhances the Protective Immune Responses Induced by Immunization with a Murine Malaria MVA-Based Vaccine Encoding the Circumsporozoite Protein.

Malaria remains a major global public health problem with an estimated 200 million cases detected in 2012. Although the most advanced candidate malaria vaccine (RTS,S) has shown promise in clinical trials, its modest efficacy and durability have created uncertainty about the impact of RTS,S immunization (when used alone) on global malaria transmission. Here we describe the development and characterization of a novel modified vaccinia virus Ankara (MVA)-based malaria vaccine which co-expresses the Plasmodium yoelii circumsporozoite protein (CSP) and IL-15. Vaccination/challenge studies showed that C57BL/6 mice immunized with the MVA-CSP/IL15 vaccine were protected significantly better against a P. yoelii 17XNL sporozoite challenge than either mice immunized with an MVA vaccine expressing only CSP or naïve controls. Importantly, the levels of total anti-CSP IgG were elevated about 100-fold for the MVA-CSP/IL15 immunized group compared to mice immunized with the MVA-CSP construct that does not express IL-15. Among the IgG subtypes, the IL-15 expressing MVA-CSP vaccine induced levels of IgG1 (8 fold) and IgG2b (80 fold) higher than the MVA-CSP construct. The significantly enhanced humoral responses and protection detected after immunization with the MVA-CSP/IL15 vaccine suggest that this IL-15 expressing MVA construct could be considered in the development of future malaria immunization strategies

Novel malaria antigen Plasmodium yoelii E140 induces antibody-mediated sterile protection in mice against malaria challenge.

Only a small fraction of the antigens expressed by malaria parasites have been evaluated as vaccine candidates. A successful malaria subunit vaccine will likely require multiple antigenic targets to achieve broad protection with high protective efficacy. Here we describe protective efficacy of a novel antigen, Plasmodium yoelii (Py) E140 (PyE140), evaluated against P. yoelii challenge of mice. Vaccines targeting PyE140 reproducibly induced up to 100% sterile protection in both inbred and outbred murine challenge models. Although PyE140 immunization induced high frequency and multifunctional CD8+ T cell responses, as well as CD4+ T cell responses, protection was mediated by PyE140 antibodies acting against blood stage parasites. Protection in mice was long-lasting with up to 100% sterile protection at twelve weeks post-immunization and durable high titer anti-PyE140 antibodies. The E140 antigen is expressed in all Plasmodium species, is highly conserved in both P. falciparum lab-adapted strains and endemic circulating parasites, and is thus a promising lead vaccine candidate for future evaluation against human malaria parasite species

Multi-parameter flow cytometry analysis of splenocytes harvested from vaccinated or sham-vaccinated control mice (4 mice per group).

<p>Splenocytes from individual mice immunized with either the MVA-CSP/IL15 vaccine or the MVA-CSP construct and naïve controls were stimulated with recombinant CSP, stained using standard intracellular staining protocols (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0141141#sec002" target="_blank">Materials and Methods</a><i>section</i> for details) and then analyzed by multi-parameter flow cytometry. The cells were evaluated for the presence of CD3, CD4 and CD8 cell surface markers and the expression of IFN-γ, TNF-α, and/or IL-2. Only the frequencies of cells expressing IFN-γ and IFN-γ/TNF-α are shown. (*p<0.05, relative to naive).</p

Parasitemia levels in infected, vaccinated mice depleted of immune cells.

<p>Panel <b>A</b>. Mice immunized with the MVA-CSP/IL15 vaccine were depleted of CD4 T cells (closed circles) using specific antibodies and then infected with PyNL SPZ before evaluating parasitemia levels. Control mice were vaccinated but were not depleted of CD4 T cells before the PyNL infection (closed circles). Results are expressed as the mean % parasitemia ± SEM (10 mice per group); p = 0.008 (Mann Whitney test). Panel <b>B</b>. Parasitemia levels at day 16 after the PyNL SPZ infection. Mice (n = 10) were immunized with the MVA-CSP/IL15 vaccine and then were treated with specific antibodies to deplete CD4 T cells, CD8 T cells or NK cells. The control groups consisted of non-depleted, vaccinated mice or naïve animals. At day 16 when parasitemia levels for all groups were compared (Kruskal-Wallis test), a statistical significance of p<0.05 was observed. Parasitemia levels for the CD4 T cell depleted vaccinated mice were significantly higher than the naïve levels (**p = 0.007) while the difference in the extent of parasitemia approached statistical significance [*p = 0.047 for the vaccinated compared to naïve mice (Mann Whitney test)].</p

Liver sections from mice infected with GFP-expressing <i>P</i>. <i>yoelii</i> SPZ.

<p>Naïve controls (A) and mice immunized with a single dose of MVA-CSP/IL15 vaccine (B) were infected intravenously with GFP-expressing Py SPZ 4 weeks after the vaccination (6000 SPZ/mouse). Livers were harvested 24 hours post-infection and subjected to 2-photon microscopy. Fluorescent green spots represent hepatocytes harboring GFP-expressing <i>P</i>. <i>yoelii</i>.</p

Representative parasitemia curves after infection of MVA-CSP/IL15, MVA-CSP vaccinated and naïve mice with 100 PyNL SPZ.

<p>MVA-CSP/IL15 vaccinated (closed circles), MVA-CSP vaccinated (open triangle) and naïve/PBS control (open circles) mice were infected intravenously with PyNL SPZ and parasitemias were evaluated by blood smears every third day starting at day 7. Results are expressed as mean % parasitemia ± SEM for ten mice per group. Differences in parasitemia levels among groups were significant at D-16 with a *p = 0.005, D-19 with a **p = 0.010 and D-22 with a ***p = 0.009 (Kruskal-Wallis test).</p