The debate over the effector function of eosinophils in helminth infection: new evidence from studies on the regulation of vaccine immunity by IL-12

The production of Th1-type cytokines is associated with strong cell-mediated immunity while Th2-type cytokines are typically involved in the generation of humoral immune responses. In mice vaccinated a single time (1X) with attenuated cercariae of Schistosoma mansoni, the immunity induced is highly dependent on CD4+ T cells and IFN-gamma. In contrast, mice vaccinated multiple times (3X) have decreased IFN-gamma expression, develop a more dominant Th2-type cytokine response as well as protective antibodies which can passively transfer immunity to naive recipients. Previously, we demonstrated the ability of IL-12, a potent IFN-gamma-inducing cytokine to enhance (1X) schistosome cell-mediated immunity when administered during the period of immunization. More recently, we asked what effects IL-12 would have on the development humoral-based immunity. While multiply-immunized/saline-treated mice demonstrated a 70-80% reduction in parasite burden, 3X/IL-12-vaccinated animals displayed an even more striking >90% reduction in challenge infection, with many mice in the later group demonstrating complete protection. Analysis of pulmonary cytokine mRNA responses demonstrated that control challenged mice elicited a dominant Th2-type response, 3X/saline-vaccinated produced a mixed Th1/Th2-type cytokine response, while 3X/IL-12-immunized animals displayed a dominant Th1-type response. The IL-12-treated group also showed a marked reduction in total serum IgE and tissue eosinophilia while SWAP-specific IgG2a and IgG2b Abs were elevated. Interestingly, animals vaccinated with IL-12 also showed a highly significant increase in total Ig titers specific for IrV-5, a known protective antigen. More importantly, 3X/IL-12 serum alone, when transferred to naive mice reduced worm burdens by over 60% while 3X/saline serum transferred significantly less protection. Nevertheless, animals vaccinated in the presence of IL-12 also develop macrophages with enhanced nitric oxide dependent killing activity against the parasites. Together, these observations suggest that IL-12, initially described as an adjuvant for cell-mediated immunity, may also be used as an adjuvant for promoting both humoral and cell-mediated protective responses.

Glutathione S-transferases of 28kDa as major vaccine candidates against schistosomiasis

For the development of vaccine strategies to generate efficient protection against chronic infections such as parasitic diseases, and more precisely schistosomiasis, controlling pathology could be more relevant than controlling the infection itself. Such strategies, motivated by the need for a cost-effective complement to existing control measures, should focus on parasite molecules involved in fecundity, because in metazoan parasite infections pathology is usually linked to the output of viable eggs. In numerous animal models, vaccination with glutathione S-transferases of 28kDa has been shown to generate an immune response strongly limiting the worm fecundity, in addition to the reduction of the parasite burden. Recent data on acquired immunity directed to 28GST in infected human populations, and new development to draw adapted vaccine formulations, are presented.

Schistosomiasis vaccine development: progress and prospects

The undisputed, worldwide success of chemotherapy notwithstanding, schistosomiasis continues to defy control efforts in as much rapid reinfection demands repeated treatment, sometimes as often as once a year. There is thus a need for a complementary tool with effect for the longer term, notably a vaccine. International efforts in this direction have been ongoing for several decades but, until the recombinant DNA techniques were introduced, antigen production remained an unsurmountable bottleneck. Although animal experiments have been highly productive and are still much needed, they probably do not reflect the human situation adequately and real progress can not be expected until more is known about human immune responses to schistosome infection. It is well-known that irradiated cercariae consistently produce high levels of protection in experimental animals but, for various reasons, this proof of principle cannot be directly exploited. Research has instead been focussed on the identification and testing of specific schistosome antigens. This work has been quite successful and is already at the stage where clinical trials are called for. Preliminary results from coordinated in vitro laboratory and field epidemiological studies regarding the protective potential of several antigens support the initiation of such trials. A series of meetings, organized earlier this year in Cairo, Egypt, reviewed recent progress, selecteded suitable vaccine candidates and made firm recommendations for future action including pledging support for large-scale production according to good manufacturing practice (GMP) and Phase I trials. Scientists at the American Centers for Disease Control and Prevention (CDC) have drawn up a detailed research plan. The major financial support will come from USAID, Cairo, which has established a scientific advisory group of Egyptian scientists and representatives from current and previous international donors such as WHO, NIAID, the European Union and the Edna McConnell Clark Foundation.

Is there a role for autoimmunity in immune protection against malaria?

Much remains to be known about the mechanisms involved in protective immunity against malaria and the way it is acquired. This is probably the reason why, in spite of so much progress, it has not yet been possible to develop an anti-malaria vaccine able to induce parasite specific antibodies (Ab) and/or T-cells. It has been considered in the early 80s that the induction of efficient protection against the blood stage forms of Plasmodium falciparum would not be possible without simultaneously eliciting an autoimmune (AI) response against erythrocytes, even at the price of inducing an AI pathology. Despite the description of the reciprocal relationship, i.e. the protective effect of malaria on the development of AI diseases - demonstrated since 1970 - no effort has been made to verify the possible involvement of the AI response in protection against malaria. With this end in view - and in the light of the knowledge acquired in autoimmunity and the existence of the so called "natural" (not associated with pathology) autoantibodies - we propose to examine the hypothesis that the participation of the AI response (not necessarily restricted to autologous erythrocyte antigens) in the immune protection against malaria is possible or even necessary.

Leishmania cysteine proteinases: from gene to subunit vaccine.

Whole genome sequences of microbial pathogens present new opportunities for clinical application. Presently, genome sequencing of the human protozoan parasite Leishmania major is in progress. The driving forces behind the genome project are to identify genes with key cellular functions and new drug targets, to increase knowledge on mechanisms of drug resistance and to favor technology transfer to scientists from endemic countries. Sequencing of the genome is also aimed at the identification of genes that are expressed in the infectious stages of the parasite and in particular in the intracellular form of the parasite. Several protective antigens of Leishmania have been identified. In addition to these antigens, lysosomal cysteine proteinases (CPs) have been characterized in different strains of Leishmania and Trypanosoma, as new target molecules. Recently, we have isolated and characterized Type I (CPB) and Type II (CPA) cysteine proteinase encoding genes from L. major. The exact function of cysteine proteinases of Leishmania is not completely understood, although there are a few reports describing their role as virulence factors. One specific feature of CPB in Leishmania and other trypanosomatids, is the presence of a Cterminal extension (CTE) which is possibly indicative of conserved structure and function. Recently, we demonstrated that DNA immunization of genetically susceptible BALB / c mice, using a cocktail of CPB and CPA genes, induced long lasting protection against L. major infection. This review intends to give an overview of the current knowledge on genetic vaccination used against leishmaniasis and the importance of CP genes for such an approach.

HIV-1 Polymorphism: a Challenge for Vaccine Development - A Review

The perspective for the development of anti-HIV/AIDS vaccines became a target sought by several research groups and pharmaceutical companies. However, the complex virus biology in addition to a striking genetic variability and the limited understanding of the immunological correlates of protection have made this an enormous scientific challenge not overcome so far. In this review we presented an updating of HIV-1 subtypes and recombinant viruses circulating in South American countries, focusing mainly on Brazil, as one of the challenges for HIV vaccine development. Moreover, we discussed the importance of stimulating developing countries to participate in the process of vaccine evaluation, not only testing vaccines according to already defined protocols, but also working together with them, in order to take into consideration their local information on virus diversity and host genetic background relevant for the vaccine development and testing, as well as including local virus based reagents to evaluate the immunogenicity of the candidate vaccines.

Intradermal vaccination of adults with three low doses (2 µg) of recombinant hepatitis B vaccine. II. Persistence of immunity and induction of immunologic memory

Of the 110 dentists who had presented seroconversion 50 days after the intradermal application of three 2 µg doses of the Belgian recombinant vaccine against hepatitis B (HB), administered eight years before at an interval of one month between the 1st and 2nd doses and of five months between the 2nd and 3rd doses, 51 were included for the assessment of the persistence of immunity. None of the dentists had hepatitis or had received HB vaccine during this period. All subjects were submitted to serological tests for the detection of the following markers of hepatitis B virus (HBV) infection: HBsAg, anti-HBc, HBeAg, anti-HBe, and anti-HBs, with no HBsAg, anti-HBc, HBeAg or anti-HBe being detected. A microparticle enzyme immunoassay (MEIA) revealed the presence of anti-HBs at protective titers (> 10 mIU/ml) in 42 dentists (82.4%), with the anti-HBs titer being higher than 100 mIU/ml in 36 of them (70.6%) (good responders), between 10 and 100 mIU/ml in 6 (11.8%) (poor responders), and lower than 10 mIU/ml in 9 (17.6%) (non-responders). According to clinical data and serological tests, none of the dentists had presented disease or latent HBV infection during the eight years following the first vaccination. A 2 µg booster dose was administered intradermally to eight dentists with anti-HBs titers lower than 10 mIU/ml (non-responders) and to six dentists with titers ranging from 10 to 100 mIU/ml (poor responders); the determination of anti-HBs one month later demonstrated the occurrence of seroconversion in the eight non-responders and an increase in anti-HBs titer in the six poor responders. In summary, the present results demonstrated the prolonged persistence of protection against HBV infection and the development of immunologic memory provided by vaccination against HB - with intra-dermal application of three 2 µg doses of the Belgian recombinant vaccine at 0, 1, and 6 months - carried out eight years before in 51 dentists.

Transmit: Health protection service bulletin. 2011: Issue 7

This edition of Transmit leads with the arrangements for the management of seasonal flu during the 2011/12 campaign. It provides an overview of the vaccination programme, including details on who should be vaccinated, and includes a link to the updated chapter in the Green Book.The bulletin has duty room guidance on the management of invasive group A streptococcal disease (iGAS) and the news section gives an encouraging update on measles in Northern Ireland.A number of routine reports are also summarised in this edition of Transmit. Statistics and graphs are provided to outline the latest reports on:· Immunisations and vaccine preventable diseases· Respiratory pathogens, quarters one and two 2011, Northern Ireland· Quarterly reporting of MRSA and Clostridium difficile infections (CDI) · Cryptosporidium 2010

Immune response mechanisms and protection against "Plasmodium falciparum" and "Plasmodium berghei"

Malaria is one of the most important tropical and infectious diseases causing many deaths and enormous social and economic consequences, particularly in the developing countries. Despite of widely use of anti-malaria drugs and insecticide, the development of successful vaccines constitutes one of the main strategies to control malaria transmission. Several proteins expressed from blood stage such as merozoite surface proteins (MSP] or liver stage as circumsporozoite protein (CSP) are shown to be the targets of immune responses in humans and in animals. Thus, several studies have illustrated that natural infection and laboratory immunizations of humans and animals with Plasmodium sporozoite (SPZ) and its derivate-proteins (peptides) can elicit protection and control of parasite infection. However, a clear understanding of immune response against defined Plasmodium proteins should be the prerequisite conditions before any development of appropriate vaccines. In this order, our study focused on the immune responses to MSP2 (dimorphic and C-terminal fragments) in human and mice; and the mechanisms by which mouse infected hepatocytes present Plasmodium antigens to CD8+ T-cells to induce protective immunity in mice.¦The first part of this work shows that infected hepatocytes can present Plasmodium antigens to PbCSP-specific CD8+ T-cells and induce a protective immunity in mice. Here, this was addressed in vivo and showed that the infected hepatocytes were able of stimulating of primed-and naive-CD8+ T-cell clones and induced fully protective immunity against SPZ challenge. The role of infected hepatocytes in antigen presentation was illustrated here by their graft into immuno-deficient mice and depletion of cosspresenting dentritic cells (DCs) that are known to have key role in the activation of CD8+ T-cells during the liver cycle stage of Plasmodium.¦The second part of this project concerned the fine specificity of Ab responses regarding D and C regions of the two allelic families of MSP2 (3D7 and FC27). Covering of the two regions by overlapping-20 mers led to delineate the epitopes in the different endemic areas and different age groups of donors. The major epitopes characterizing D or C regions were conserved in different endemic areas (P12/P13 and P15/P16 for the 3D7-D, P23/24 and P25/26 for the FC27-D; P29/P30 for the C region). This offers thus, the possibility of a multi-epitope vaccine design including the major epitopes from the two domains of the two allelic MSP2 families. On the other, the 20 mers, particularly some major epitopes of the 3D7-Dregion (P12, P13 and P16) belonged to the epitopes that presented a high probability to be associated with protection in the children group [1 to 5 year-old). In addition, D and C LSP purified Abs (pAbs) recognized merozoite derived polypeptides and native proteins. A crossreactivity activity of homologous pAbs against the heterologous was also illustrated between the two allelic MSP2 parasites. Finally, the functional analysis of D regions pAbs showed an inhibition of Plasmodium falciparum growth suggesting the functional biological activity of the D region pAbs in the control of malaria.¦The last part of this project aimed the evaluation of the immunogenicity of the D and C region LSPs of the two allelic MSP2 families in the presence of adjuvants for the possible use in clinical trial study in humans. The MSP2 LSP mixture showed that D and C were immunogenic and defined limited epitopes (whose intensity of immune responses) depending on the adjuvants and mouse strain for the D regions. The major epitopes characterizing the C region were usually conserved in different strains of mouse and adjuvants used. Furthermore, the single region (either with D or C) immunization of mice confirmed the immunogenicity and the presence of their limited epitopes. We concluded that the possibility to finely delineate in animals the immune responses to antigens might help to select optimal antigen/adjuvant combinations to be tested later in clinical trials. Thus, formulation of glucopyranosyl-lipid A stable emulsion, GLA-SE (toll like receptor (TLR) 4 agonist) and its different combination (CpG: TLR9 agonist and GDQ: LR7 agonist) with MSP2 LSP was better than with alum, montanide ISA 720 (Mt) and virosome. Immunization of mice with allelic LSP did not show a crossreactivity between the two allelic MSP2 parasites unlike as humans, suggesting that the crossreactivity could be acquired during natural infection of the population who are usually exposed to both allelic parasite forms (3D7 and FC27).¦Nevertheless, similar epitope of D (P12, P13 and P25) and C (P29) regions have been found both in mice and human. This offers an opportunity to compare their epitopes in naïve immunized donors with LSPs and naturally infected populations in the endemic areas.

Recombinant tegumental protein Shistosoma japonicum very lowdensity lipoprotein binding protein as a vaccine candidate against Schistosoma japonicum

A polyhistidine-tagged recombinant tegumental protein Schistosoma japonicum very lowdensity lipoprotein binding protein (SVLBP) from adult Schistosoma japonicum was expressed in Escherichia coli. The affinity purified rSVLBP was used to vaccinate mice. The worm numbers and egg deposition recovered from the livers and veins of the immunized mice were 33.5% and 47.6% less than that from control mice, respectively (p<0.05). There was also a marked increase in the antibody response in vaccinated mice: the titer of IgG1 and IgG2a, IgG2b in the vaccinated group was significantly higher than that in the controls (>1:6,400 in total IgG). In a comparison of the reactivity of sera from healthy individuals and patients with rSVLBP, recognition patterns against this parasite tegumental antigen varied among different groups of the individuals. Notably, the average titres of anti-rSVLBP antibody in sera from faecal egg-negative individuals was significantly higher than that in sera from the faecal egg-positives, which may be reflect SVLBP-specific protection. These results suggested that the parasite tegumental protein SVLBP was a promising candidate for further investigation as a vaccine antigen for use against Asian schistosomiasis.

Aluminum hydroxide associated to Schistosoma mansoni 22.6 kDa protein abrogates partial protection against experimental infection but not alter interleukin-10 production

The need to develop a vaccine against schistosomiasis led several researches and our group to investigate proteins from Schistosoma mansoni as vaccine candidates. Sm22.6 is a protein from S. mansoni that shows high identity with Sj22.6 and Sh22.6 (79 and 91%, respectively). These proteins are associated with high levels of IgE and protection to reinfection. Previously, we have shown that Sm22.6 induced a partial protection of 34.5% when used together with Freund's adjuvant and produced a Th0 type of immune response with interferon-g and interleukin-4. In this work, mice were immunized with Sm22.6 alone or with aluminum hydroxide adjuvant and high levels of IgG, IgG1, and IgG2a were measured. Unfortunately, no protection was detected. Since IL-10 is a modulating cytokine in schistosomiasis, we also observed a high level of this molecule in splenocytes of vaccinated mice. In conclusion, we did not observe the adjuvant effect of aluminum hydroxide associated with rSm22.6 in protective immunity.

Schistosome vaccine testing: lessons from the baboon model

The high level of protection elicited in rodents and primates by the radiation-attenuated schistosome vaccine gives hope that a human vaccine relying on equivalent mechanisms is feasible. In humans, a vaccine would be undoubtedly administered to previously or currently infected individuals. We have therefore used the olive baboon to investigate whether vaccine-induced immunity is compromised by a schistosome infection. We showed that neither a preceding infection, terminated by chemotherapy, nor an ongoing chronic infection affected the level of protection. Whilst IgM responses to vaccination or infection were short-lived, IgG responses rose with each successive exposure to the vaccine. Such a rise was obscured by responses to egg deposition in already-infected animals. In human trials it would be necessary to use indirect estimates of infection intensity to determine vaccine efficacy. Using worm burden as the definitive criterion, we demonstrated that the surrogate measures, fecal eggs, and circulating antigens, consistently overestimated protection. Regression analysis of the surrogate parameters on worm burden revealed that the principal reason for overestimation was the threshold sensitivity of the assays. If we extrapolate our findings to human schistosomiasis mansoni, it is clear that more sensitive indirect measures of infection intensity are required for future vaccine trials.

Assessment of a DNA vaccine encoding an anchored-glycosylphosphatidylinositol tegumental antigen complexed to protamine sulphate on immunoprotection against murine schistosomiasis

Protamine sulphate/DNA complexes have been shown to protect DNA from DNase digestion in a lipid system for gene transfer. A DNA-based vaccine complexed to protamine sulphate was used to induce an immune response against Schistosoma mansoni anchored-glycosylphosphatidylinositol tegumental antigen in BALB/c mice. The protection elicited ranged from 33 to 44%. The spectrum of the elicited immune response induced by the vaccine formulation without protamine was characterized by a high level of IgG (IgG1> IgG2a). Protamine sulphate added to the DNA vaccine formulation retained the green fluorescent protein encoding-plasmid longer in muscle and spleen. The experiments in vivo showed that under protamine sulphate effect, the scope of protection remained unchanged, but a modulation in antibody production (IgG1= IgG2a) was observed.

IgG and IgG2 antibodies from cattle naturally infected with Anaplasma marginale recognize the recombinant vaccine candidate antigens VirB9, VirB10, and elongation factor-Tu

Anaplasma marginale is an important vector-borne rickettsia of ruminants in tropical and subtropical regions of the world. Immunization with purified outer membranes of this organism induces protection against acute anaplasmosis. Previous studies, with proteomic and genomic approach identified 21 proteins within the outer membrane immunogen in addition to previously characterized major surface protein1a-5 (MSP1a-5). Among the newly described proteins were VirB9, VirB10, and elongation factor-Tu (EF-Tu). VirB9, VirB10 are considered part of the type IV secretion system (TFSS), which mediates secretion or cell-to-cell transfer of macromolecules, proteins, or DNA-protein complexes in Gram-negative bacteria. EF-Tu can be located in the bacterial surface, mediating bacterial attachment to host cells, or in the bacterial cytoplasm for protein synthesis. However, the roles of VirB9, VirB10, and TFSS in A. marginale have not been defined. VirB9, VirB10, and EF-Tu have not been explored as vaccine antigens. In this study, we demonstrate that sera of cattle infected with A. marginale, with homologous or heterologous isolates recognize recombinant VirB9, VirB10, and EF-Tu. IgG2 from naturally infected cattle also reacts with these proteins. Recognition of epitopes by total IgG and by IgG2 from infected cattle with A. marginale support the inclusion of these proteins in recombinant vaccines against this rickettsia.

Comparative evaluation of phenol and thimerosal as preservatives for a candidate vaccine against American cutaneous leishmaniasis

For decades thimerosal has been used as a preservative in the candidate vaccine for cutaneous leishmaniasis, which was developed by Mayrink et al. The use of thimerosal in humans has been banned due to its mercury content. This study addresses the standardization of phenol as a new candidate vaccine preservative. We have found that the proteolytic activity was abolished when the test was conducted using the candidate vaccine added to merthiolate (MtVac) as well as to phenol (PhVac). The Montenegro's skin test conversion rates induced by MtVac and by PhVac was 68.06% and 85.9%, respectively, and these values were statistically significant (p < 0.05). The proliferative response of peripheral mononuclear blood cells shows that the stimulation index of mice immunized with both candidate vaccines was higher than the one in control animals (p < 0.05). The ability of the candidate vaccines to induce protection in C57BL/10 mice against a challenge with infective Leishmania amazonensis promastigotes was tested and the mice immunized with PhVac developed smaller lesions than the mice immunized with MtVac. Electrophoresis of phenol-preserved antigen revealed a number of proteins, which were better preserved in PhVac. These results do in fact encourage the use of phenol for preserving the immunogenic and biochemical properties of the candidate vaccine for cutaneous leishmaniasis.