Direct Inhibition of T-Cell Responses by the Cryptococcus Capsular Polysaccharide Glucuronoxylomannan

The major virulence factor of the pathogenic fungi Cryptococcus neoformans and C. gattii is the capsule. Glucuronoxylomannan (GXM), the major component of the capsule, is a high-molecular-weight polysaccharide that is shed during cryptococcosis and can persist in patients after successful antifungal therapy. Due to the importance of T cells in the anticryptococcal response, we studied the effect of GXM on the ability of dendritic cells (DCs) to initiate a T-cell response. GXM inhibited the activation of cryptococcal mannoprotein-specific hybridoma T cells and the proliferation of OVA-specific OT-II T cells when murine bone marrow-derived DCs were used as antigen-presenting cells. Inhibition of OT-II T-cell proliferation was observed when either OVA protein or OVA323-339 peptide was used as antigen, indicating GXM did not merely prevent antigen uptake or processing. We found that DCs internalize GXM progressively over time; however, the suppressive effect did not require DCs, as GXM directly inhibited T-cell proliferation induced by anti-CD3 antibody, concanavalin A, or phorbol-12-myristate-13-acetate/ionomycin. Analysis of T-cell viability revealed that the reduced proliferation in the presence of GXM was not the result of increased cell death. GXM isolated from each of the four major cryptococcal serotypes inhibited the proliferation of human peripheral blood mononuclear cells stimulated with tetanus toxoid. Thus, we have defined a new mechanism by which GXM can impart virulence: direct inhibition of T-cell proliferation. In patients with cryptococcosis, this could impair optimal cell-mediated immune responses, thereby contributing to the persistence of cryptococcal infections. SynopsisInfections due to the pathogenic yeast Cryptococcus are a significant cause of morbidity and mortality in persons with impaired T-cell functions, particularly those with AIDS. The major virulence factor of Cryptococcus is its capsule, which is composed primarily of the polysaccharide glucuronoxylomannan (GXM). The capsule not only surrounds the organism but also is shed during cryptococcosis. GXM is taken up by macrophages in vitro and in vivo; however, little is known about the interaction between GXM and dendritic cells, which are the most potent cells capable of activating T cells. Because of the importance of T cells in the anticryptococcal response, the authors investigated the effect of GXM on the ability of dendritic cells to initiate a T-cell response. They found the polysaccharide was internalized by dendritic cells and inhibited antigen-specific T-cell responses. Furthermore, GXM had a direct, inhibitory effect on T-cell proliferation, independent of the effect on dendritic cells. These findings may help explain the persistence of cryptococcal infections and suggest that GXM could be therapeutic in situations where suppression of T-cell responses is desired.National Institutes of Health (R01 AI25780, R01 AI066087, R01 AI37532

Report from the field - Overview of the Sixth Annual Vaccine Renaissance Conference

The Sixth Annual Vaccine Renaissance Conference, hosted by the Institute for Immunology and Informatics (iCubed) at the University of Rhode Island (URI), took place on October 15–17, 2012. This conference provides a forum for the review of current progress in the discovery and development of vaccines, and creates an environment for the exchange of ideas. Dr. Joel McCleary opened the conference with a warning about the importance of preparing for well-defined biowarfare threats, including tularemia and Staphylococcal enterotoxin B. Following the keynote address, sessions explored biodefense and preparation for pandemic and biowarfare threats; vaccines for emerging and re-emerging neglected tropical diseases; animal vaccines and human health; and vaccine vectors and the human microbiome. In this issue of Human Vaccines and Immunotherapeutics, seven Vaccine Renaissance Conference speakers will showcase their work; here, we describe a few of the conference highlights

Low immunogenicity predicted for emerging avian-origin H7N9

A new avian-origin influenza virus emerged near Shanghai in February 2013, and by the beginning of May it had caused over 130 human infections and 36 deaths. Human-to-human transmission of avian-origin H7N9 influenza A has been limited to a few family clusters, but the high mortality rate (27%) associated with human infection has raised concern about the potential for this virus to become a significant human pathogen. European, American, and Asian vaccine companies have already initiated the process of cloning H7 antigens such as hemagglutinin (HA) into standardized vaccine production vehicles. Unfortunately, previous H7 HA-containing vaccines have been poorly immunogenic. We used well-established immunoinformatics tools to analyze the H7N9 protein sequences and compare their T cell epitope content to other circulating influenza A strains as a means of estimating the immunogenic potential of the new influenza antigen. We found that the HA proteins derived from closely related human-derived H7N9 strains contain fewer T cell epitopes than other recently circulating strains of influenza, and that conservation of T cell epitopes with other strains of influenza was very limited. Here, we provide a detailed accounting of the type and location of T cell epitopes contained in H7N9 and their conservation in other H7 and circulating (A/California/07/2009, A/Victoria/361/2011, and A/Texas/50/2012) influenza A strains. Based on this analysis, avian-origin H7N9 2013 appears to be a “stealth” virus, capable of evading human cellular and humoral immune response. Should H7N9 develop pandemic potential, this analysis predicts that novel strategies for improving vaccine immunogenicity for this unique low-immunogenicity strain of avian-origin influenza will be urgently needed

Immunization with Cross-Conserved H1N1 Influenza CD4\u3csup\u3e+\u3c/sup\u3e T-Cell Epitopes Lowers Viral Burden in HLA DR3 Transgenic Mice

The emergence of the pandemic H1N1 strain of influenza in 2009 was associated with a unique w-shaped age-related susceptibility curve, with higher incidence of morbidity and mortality among young persons and lower incidence among older persons, also observed during the 1918 influenza pandemic. Pre-existing H1N1 antibodies were not cross-reactive with the prior seasonal vaccine, forcing influenza experts to scramble to develop a new vaccine specific for the pandemic virus. We hypothesized that response to T-cell epitopes that are cross-conserved between pandemic H1N1 and the 2008 seasonal influenza vaccine strains might have contributed to partial protection from clinical illness among older adults, despite the lack of cross-reactive humoral immunity. Using immunoinformatics tools, we previously identified hemagglutinin and neuraminidase epitopes that were highly conserved between seasonal and pandemic H1N1. Here, we validated predicted CD4+ T-cell epitopes for their ability to bind HLA and to stimulate interferon-γ production in peripheral blood mononuclear cells from a cohort of donors presenting with influenza-like illness during the 2009 pandemic and a separate cohort immunized with trivalent influenza vaccine in 2011. A limited-epitope heterologous DNA-prime/peptide-boost vaccine composed of these sequences stimulated immune responses and lowered lung viral loads in HLA DR3 transgenic mice challenged with pandemic 2009 H1N1 influenza. Cross-priming with conserved influenza T-cell epitopes such as these may be critically important to T cell-mediated protection against pandemic H1N1 in the absence of cross-protective antibodies

A Cross-Sectional Study to Assess HPV Knowledge and HPV Vaccine Acceptability in Mali

Despite a high prevalence of oncogenic human papilloma virus (HPV) infection and cervical cancer mortality, HPV vaccination is not currently available in Mali. Knowledge of HPV and cervical cancer in Mali, and thereby vaccine readiness, may be limited. Research staff visited homes in a radial pattern from a central location to recruit adolescent females and males aged 12–17 years and men and women aged ≥18 years (N = 51) in a peri-urban village of Bamako, Mali. Participants took part in structured interviews assessing knowledge, attitudes, and practices related to HPV, cervical cancer, and HPV vaccination. We found low levels of HPV and cervical cancer knowledge. While only 2.0% of respondents knew that HPV is a sexually transmitted infection (STI), 100% said they would be willing to receive HPV vaccination and would like the HPV vaccine to be available in Mali. Moreover, 74.5% said they would vaccinate their child(ren) against HPV. Men were found to have significantly greater autonomy in the decision to vaccinate themselves than women and adolescents (p = 0.005), a potential barrier to be addressed by immunization campaigns. HPV vaccination would be highly acceptable if the vaccine became widely available in Bamako, Mali. This study demonstrates the need for a significant investment in health education if truly informed consent is to be obtained for HPV vaccination. Potential HPV vaccination campaigns should provide more information about HPV and the vaccine. Barriers to vaccination, including the significantly lower ability of the majority of the target population to autonomously decide to get vaccinated, must also be addressed in future HPV vaccine campaigns

C3d adjuvant effects are mediated through the activation of C3d-specific autoreactive T cells

Complement fragment C3d covalently attached to antigens enhances immune responses, particularly for antigens lacking T cell epitopes. Enhancement has been attributed to receptor cross-linking between complement receptor CR2 (CD21) and polysaccharide antigen to surface IgM on naïve B cells. Paradoxically, C3d has still been shown to increase immune responses in CD21 KO mice, suggesting that an auxiliary activation pathway exists. In prior studies, we demonstrated the CD21-independent C3d adjuvant effect might be due to T cell recognition of C3d T helper epitopes processed and presented by MHC class II on the B cell surface. C3d peptide sequences containing concentrated clusters of putative human C3 T cell epitopes were identified using the epitope-mapping algorithm, EpiMatrix. These peptide sequences were synthesized and shown in vitro to bind multiple HLA-DR alleles with high affinity, and induce IFNγ responses in healthy donor PBMCs. In the present studies, we establish further correlations between HLA binding and HLA-specific lymphocyte reactions with select epitope clusters. Additionally, we show that the T cell phenotype of C3d-specific reactive T cells is CD4+CD45RO+ memory T cells. Finally, mutation of a single T cell epitope residing within the P28 peptide segment of C3d resulted in significantly diminished adjuvant activity in BALB/c mice. Collectively, these studies support the hypothesis that the paradoxical enhancement of immune responses by C3d in the absence of CD21 is due to internalization and processing of C3d into peptides that activate autoreactive CD4+ T helper cells in the context of HLA class II

Correction to: Individual and household characteristics of persons with Plasmodium falciparum malaria in sites with varying endemicities in Kinshasa Province, Democratic Republic of the Congo

Unfortunately after publication of the original article [1], it came to the author’s attention that there is an error in the caption of Fig. 2

Microscopic Plasmodium falciparum Gametocytemia and Infectivity to Mosquitoes in Cambodia

Although gametocytes are essential for malaria transmission, in Africa many falciparum-infected persons without smear-detectable gametocytes still infect mosquitoes. To see whether the same is true in Southeast Asia, we determined the infectiousness of 119 falciparum-infected Cambodian adults to Anopheles dirus mosquitoes by membrane feeding. Just 5.9% of subjects infected mosquitoes. The 8.4% of patients with smear-detectable gametocytes were >20 times more likely to infect mosquitoes than those without and were the source of 96% of all mosquito infections. In low-transmission settings, targeting transmission-blocking interventions to those with microscopic gametocytemia may have an outsized effect on malaria control and elimination

Receptor-Mediated Clearance of Cryptococcus neoformans Capsular Polysaccharide In Vivo

Cryptococcus neoformans capsular glucuronoxylomannan (GXM) is shed during cryptococcosis and taken up by macrophages. The roles of the putative GXM receptors CD14, CD18, Toll-like receptor 2 (TLR2), and TLR4 in GXM clearance from serum and deposition in the liver and spleen in receptor-deficient mice were studied. While alterations in the kinetics of GXM redistribution were seen in the mutant mice, none of the receptors was absolutely required for serum clearance or hepatosplenic accumulation

Opsonic Requirements for Dendritic Cell-Mediated Responses to Cryptococcus neoformans

The encapsulated pathogenic yeast Cryptococcus neoformans is poorly recognized by phagocytic cells in the absence of opsonins. Macrophages will bind and internalize complement- or antibody-opsonized C. neoformans; however, less is known about the role of opsonins in dendritic cell (DC)-mediated recognition of the organism. Thus, we studied the opsonic requirements for binding to C. neoformans by cultured human monocyte-derived and murine bone marrow-derived DCs and whether binding leads to antifungal activity and cytokine release. Binding of unopsonized C. neoformans to human and murine DCs was negligible. Opsonization with pooled human serum (PHS) increased binding, while heat treatment of PHS virtually abolished this binding, thus suggesting a role for heat-labile complement components. PHS plus a monoclonal anticapsular antibody, 3C2, had an additive effect on binding for most cryptococcal strains. Human and murine DCs exhibited pronounced anticryptococcal activity in the presence of the antibody at early (2-h) and late (24-h) time points; however, PHS opsonization did not supplement this anticryptococcal activity. Antifungal activity against C. neoformans opsonized in PHS and/or antibody was partially reduced in the presence of inhibitors of the respiratory burst response. Human, but not murine, DCs released modest amounts of tumor necrosis factor alpha when stimulated with C. neoformans opsonized in PHS and/or antibody. However, opsonized C. neoformans failed to stimulate detectable release of interleukin 10 (IL-10) or IL-12p70 from either DC population. Thus, human and murine DCs show maximal binding to and antifungal activity against C. neoformans via a process highly dependent on opsonization