Vibriosis vaccines based on various sero-subgroups of Vibrio anguillarum O2 induce specific protection in Atlantic cod (Gadus morhua L.) juveniles

The purpose of this study was to investigate the efficacy of three monovalent and a trivalent vibriosis dip vaccines in juvenile Atlantic cod (Gadus morhua L.), examine whether the responses were specific and study the expression of selected immune genes after dip vaccination. In addition, the study addressed whether the deviating isolates of Vibrio anguillarum serotype O2 belongs to another sero-subgroup than the previously established sero-subgroups O2a, O2b and O2c. Rabbit V. anguillarum serotype O2 antiserum adsorbed with V. anguillarum O2a O-antigen was shown, by both ELISA and immunoblotting, to still contain serotype O2 specific antibodies. Cod V. anguillarum serotype O2 antiserum reacted only with isolate of homologous serotype and not with heterologous sero-subgroups. This indicates that the deviating V. anguillarum O2 isolates represent a new sero-subgroup differing from sero-subgroup O2a. The monovalent vaccines included formalin inactivated cultures of V. anguillarum sero-subgroup O2a, O2b or serotype O2, while the trivalent vaccine contained all three sero-subgroups. Cod mounted high protection 7 weeks post dip vaccination with monovalent vaccines when challenged with homologous isolates and significantly lower when challenged with heterologous isolates, regardless of sero-subgroups. The trivalent vaccine resulted in efficient protection against all sero-subgroups tested. Dip vaccination of cod juveniles did not result in detectable antibody production or alteration in gene expression of the heavy chain of IgM and IgD. In the trivalent vaccine group expression of IFNγ and IL-12p40 were significantly up-regulated 3 days post vaccination. However, in groups vaccinated against V. anguillarum sero-subgroups O2b or O2, IL-12p40 and IFNγ gene expression were slightly increased 3 and 55 days post vaccination, respectively.     

Generation by Reverse Genetics of an Effective,Stable, Live-Attenuated Newcastle Disease Virus Vaccine Based on a Currently Circulating,Highly Virulent Indonesian Strain

Newcastle disease virus (NDV) can cause severe disease in chickens. Although NDV vaccines exist, there are frequent reports of outbreaks in vaccinated chickens. During 2009–2010, despite intense vaccination, NDV caused major outbreaks among commercial poultry farms in Indonesia. These outbreaks raised concern regarding the protective immunity of current vaccines against circulating virulent strains in Indonesia. In this study, we investigated whether a recombinant attenuated Indonesian NDV strain could provide better protection against prevalent Indonesian viruses. A reverse genetics system for the highly virulent NDV strain Banjarmasin/010/10 (Ban/010) isolated in Indonesia in 2010 was constructed. The Ban/010 virus is classified in genotype VII of class II NDV, which is genetically distinct from the commercial vaccine strains B1 and LaSota, which belong to genotype II, and shares only 89 and 87% amino acid identity for the protective antigens F and HN, respectively. A mutant virus, named Ban/AF, was developed in which the virulent F protein cleavage site motif “RRQKR↓F” was modified to an avirulent motif “GRQGR↓L” by three amino acid substitutions (underlined). The Ban/AF vaccine virus did not produce syncytia or plaques in cell culture, even in the presence of added protease. Pathogenicity tests showed that Ban/AF was completely avirulent. Ban/AF replicated efficiently during 10 consecutive passages in chickens and remained genetically stable. Serological analysis showed that Ban/AF induced higher neutralization and hemagglutination inhibition antibody titers against the prevalent viruses than the commercial vaccines B1 or LaSota. Both Ban/AF and commercial vaccines provided protection against clinical disease and mortality after challenge with virulent NDV strain Ban/010 (genotype VII) or GB Texas (genotype II). However, Ban/AF significantly reduced challenge virus shedding from the vaccinated birds compared to B1 vaccine. These results suggest that Ban/AF can provide better protection than commercial vaccines and is a promising vaccine candidate against NDV strains circulating in Indonesia.     

Cooperativity Between CD8+ T Cells,Non-Neutralizing Antibodies,and Alveolar Macrophages Is Important for Heterosubtypic Influenza Virus Immunity

Seasonal epidemics of influenza virus result in ∼36,000 deaths annually in the United States. Current vaccines against influenza virus elicit an antibody response specific for the envelope glycoproteins. However, high mutation rates result in the emergence of new viral serotypes, which elude neutralization by preexisting antibodies. T lymphocytes have been reported to be capable of mediating heterosubtypic protection through recognition of internal, more conserved, influenza virus proteins. Here, we demonstrate using a recombinant influenza virus expressing the LCMV GP33-41 epitope that influenza virus-specific CD8+ T cells and virus-specific non-neutralizing antibodies each are relatively ineffective at conferring heterosubtypic protective immunity alone. However, when combined virus-specific CD8 T cells and non-neutralizing antibodies cooperatively elicit robust protective immunity. This synergistic improvement in protective immunity is dependent, at least in part, on alveolar macrophages and/or other lung phagocytes. Overall, our studies suggest that an influenza vaccine capable of eliciting both CD8+ T cells and antibodies specific for highly conserved influenza proteins may be able to provide heterosubtypic protection in humans, and act as the basis for a potential “universal” vaccine.     

A Recombinant Vesicular Stomatitis Virus-Based Lassa Fever Vaccine Protects Guinea Pigs and Macaques against Challenge with Geographically and Genetically Distinct Lassa Viruses

Background

Lassa virus (LASV) is endemic in several West African countries and is the etiological agent of Lassa fever. Despite the high annual incidence and significant morbidity and mortality rates, currently there are no approved vaccines to prevent infection or disease in humans. Genetically, LASV demonstrates a high degree of diversity that correlates with geographic distribution. The genetic heterogeneity observed between geographically distinct viruses raises concerns over the potential efficacy of a “universal” LASV vaccine. To date, several experimental LASV vaccines have been developed; however, few have been evaluated against challenge with various genetically unique Lassa virus isolates in relevant animal models.

Methodologies/principle findings

Here we demonstrate that a single, prophylactic immunization with a recombinant vesicular stomatitis virus (VSV) expressing the glycoproteins of LASV strain Josiah from Sierra Leone protects strain 13 guinea pigs from infection / disease following challenge with LASV isolates originating from Liberia, Mali and Nigeria. Similarly, the VSV-based LASV vaccine yields complete protection against a lethal challenge with the Liberian LASV isolate in the gold-standard macaque model of Lassa fever.

Conclusions/significance

Our results demonstrate the VSV-based LASV vaccine is capable of preventing morbidity and mortality associated with non-homologous LASV challenge in two animal models of Lassa fever. Additionally, this work highlights the need for the further development of disease models for geographical distinct LASV strains, particularly those from Nigeria, in order to comprehensively evaluate potential vaccines and therapies against this prominent agent of viral hemorrhagic fever.     

Head-to-Head Comparison of Three Vaccination Strategies Based on DNA and Raw Insect-Derived Recombinant Proteins against Leishmania

Parasitic diseases plague billions of people among the poorest, killing millions annually, and causing additional millions of disability-adjusted life years lost. Leishmaniases affect more than 12 million people, with over 350 million people at risk. There is an urgent need for efficacious and cheap vaccines and treatments against visceral leishmaniasis (VL), its most severe form. Several vaccination strategies have been proposed but to date no head-to-head comparison was undertaken to assess which is the best in a clinical model of the disease. We simultaneously assayed three vaccination strategies against VL in the hamster model, using KMPII, TRYP, LACK, and PAPLE22 vaccine candidate antigens. Four groups of hamsters were immunized using the following approaches: 1) raw extracts of baculovirus-infected Trichoplusia ni larvae expressing individually one of the four recombinant proteins (PROT); 2) naked pVAX1 plasmids carrying the four genes individually (DNA); 3) a heterologous prime-boost (HPB) strategy involving DNA followed by PROT (DNA-PROT); and 4) a Control including empty pVAX1 plasmid followed by raw extract of wild-type baculovirus-infected T. ni larvae. Hamsters were challenged with L. infantum promastigotes and maintained for 20 weeks. While PROT vaccine was not protective, DNA vaccination achieved protection in spleen. Only DNA-PROT vaccination induced significant NO production by macrophages, accompanied by a significant parasitological protection in spleen and blood. Thus, the DNA-PROT strategy elicits strong immune responses and high parasitological protection in the clinical model of VL, better than its corresponding naked DNA or protein versions. Furthermore, we show that naked DNA coupled with raw recombinant proteins produced in insect larvae biofactories –the cheapest way of producing DNA-PROT vaccines– is a practical and cost-effective way for potential “off the shelf” supplying vaccines at very low prices for the protection against leishmaniases, and possibly against other parasitic diseases affecting the poorest of the poor.     

Identification and characterization of novel immunogenic proteins of Streptococcus suis serotype 2

Streptococcus suis, a zoonotic pathogen, caused serious outbreaks in humans with high mortality rates in the past decade. To develop safer and more effective vaccines, particularly for human protection, cell wall and extracellular proteins of S. suis serotype 2 were analyzed by an immunoproteomic approach in this study. Thirty-two proteins with high immunogenicity were identified and 22 of them were newly identified. Further analyses of 9 selected proteins revealed that (1) these 9 proteins were expressed in all tested virulent S. suis serotype 2 isolates, (2) antisera against 6 of the selected proteins efficiently killed the bacteria by opsonized phagocytosis in human blood, and (3) significantly higher levels of serum antibodies against 3 proteins were detected in both patients and infected swines. Therefore, our results suggest the 3 proteins (SSU98_0197, SSU98_1094 and SSU1664) have strong potential to be vaccine candidates.     

Estimation of the health impact and cost-effectiveness of influenza vaccination with enhanced effectiveness in Canada

Introduction

The propensity for influenza viruses to mutate and recombine makes them both a familiar threat and a prototype emerging infectious disease. Emerging evidence suggests that the use of MF59-adjuvanted vaccines in older adults and young children enhances protection against influenza infection and reduces adverse influenza-attributable outcomes compared to unadjuvanted vaccines. The health and economic impact of such vaccines in the Canadian population are uncertain.

Methods

We constructed an age-structured compartmental model simulating the transmission of influenza in the Canadian population over a ten-year period. We compared projected health outcomes (quality-adjusted life years (QALY) lost), costs, and incremental cost-effectiveness ratios (ICERs) for three strategies: (i) current use of unadjuvanted trivalent influenza vaccine; (ii) use of MF59-adjuvanted influenza vaccine adults ≥65 in the Canadian population, and (iii) adjuvanted vaccine used in both older adults and children aged < 6.

Results

In the base case analysis, use of adjuvanted vaccine in older adults was highly cost-effective (ICER = $2111/QALY gained), but such a program was “dominated” by a program that extended the use of adjuvanted vaccine to include young children (ICER = $1612/QALY). Results were similar whether or not a universal influenza immunization program was used in other age groups; projections were robust in the face of wide-ranging sensitivity analyses.

Interpretation

Based on the best available data, it is projected that replacement of traditional trivalent influenza vaccines with MF59-adjuvanted vaccines would confer substantial benefits to vaccinated and unvaccinated individuals, and would be economically attractive relative to other widely-used preventive interventions.     

Vaccines beyond antibodies: Spurred by pandemic research,are T‐cell vaccines moving closer to reality?

Lessons learned from the vaccines against SARS‐CoV‐2 has encouraged research and vaccine development aimed at mustering strong T cell responses against the pathogen. Subject Categories: Microbiology, Virology & Host Pathogen Interaction, Pharmacology & Drug Discovery

The new vaccines against SARS‐CoV‐2 elicited strong antibody responses in initial trials, which encouraged optimism amongst immunologists and public health experts who expected good efficacy. “With viral infections, it is almost unheard of to have a prophylactic vaccine that doesn’t work ultimately by generating neutralising antibody responses”, explained immunologist Kingston Mills at Trinity College Dublin in Ireland. However, the antibody response is not the whole story. “Efforts to explain how immunity is working against viruses to the general public has forced everyone to try to make things so simple that now what is left is a ridiculous oversimplified picture of the vertebrate immune system”, commented Antonio Bertoletti, infectious disease scientist at Duke‐National University of Singapore. In fact, there is increasing research focus on the role of T cells in mediating the cellular response to infections and how to stimulate these cells through vaccines.Antibodies work by recognising and attaching to surface structures of a virus or bacterium, which prevents the pathogen from infecting its target cells and mark it for destruction by other immune cells. However, pathogens can escape the antibody response via mutations that decrease the efficiency of antibodies from infection or vaccination. “You will still potentially get infected if you’re vaccinated, because the antibody response is not as strong as it was”, explained immunologist Luke O’Neill at Trinity College Dublin, Ireland. “But then the T cells will kick in and stop the virus when it is inside cells”. Simply put, antibodies tend to prevent infection, while T cells combat infection and illness. Specifically, CD4 helper T cells primarily encourage B cells to generate antibodies whereas CD8 “killer” T cells eliminate cancerous and virally infected cells.     

Vaccine outlooks

After negative publicity and a series of setbacks over HIV/AIDS and influenza, the prospects for research on new vaccines are improvingVaccine research is at a crossroads between renewed optimism created by fundamental scientific advances, and pessimism from a series of scientific and publicity setbacks over the past decade. Early successes in the field against acute viral diseases, such as smallpox and polio, raised hopes that more serious infectious diseases could be controlled or even eradicated by vaccination, just as it was once thought that penicillin would eradicate major bacterial diseases such as tuberculosis and leprosy....it became clear that many viruses were much tougher nuts to crack in terms of vaccine development...However, it became clear that many viruses were much tougher nuts to crack in terms of vaccine development than had been thought, and it also emerged that not all vaccines were equally safe. Indeed, mounting concerns over the safety of vaccines culminated in the infamous Wakefield paper published in the Lancet in 1998 that associated the MMR vaccine—against measles, mumps and rubella—with autism and inflammatory bowel disease in children. After several studies failed to reproduce these results, the Lancet eventually retracted this paper in 2010, but not before considerable damage had been done to public confidence in vaccination as a whole. Other factors have also sapped confidence in the field—notably the continuing failure to develop an effective vaccine against HIV/AIDS, 15 years after the first hopeful reports that a breakthrough might be imminent (Gorse et al, 1995).Researchers have since developed a string of HIV/AIDS vaccine candidates, some of which have entered clinical trials, but none of which have been sufficiently efficient and safe. The gloom has deepened further after the failure of a vaccine candidate developed by the pharmaceutical company Merck in 2007, that had high hopes for success. This vaccine, called V520, used a weakened adenovirus that carries three HIV genes, to stimulate host production of T cells that it was hoped would kill HIV-infected cells. Early small trials had detected cellular immune responses, but these largely failed to materialize in a subsequent phase II clinical trial. The recombinant vaccine triggered a rapid immune response against itself that actually impaired the T-cell response against the HIV antigens. As a result, the trial was halted in September 2007 (Anon, 2007).Controversially, it has since been suggested that V520 rendered some individuals more liable to subsequent infection, although views on this finding are polarized. According to Steven Patterson, a research fellow specializing in HIV at Imperial College, London, “there was a greater incidence of infection in those individuals who had immunity to adenovirus type 5 before vaccination. Some scientists argue that the numbers [of people who suffered infection during the trial] are relatively low, the results represent a statistical anomaly and that the effect gradually disappeared, suggesting that it was not a real effect. Others, including ourselves, think that in adenovirus type 5-immune individuals the vector activated pre-existing memory CD4 cells migrate to mucosal tissue. Then, because the virus preferentially replicates in activated CD4 T cells and the number of HIV susceptible cells is increased at the site of HIV infection, there is an increase in the number of infections. With time the activated cells return to a resting state which would explain why the effect of adenovirus vaccination gradually disappeared.”Whatever the truth in this case, it highlighted the setbacks in HIV/AIDS research and increased negative sentiments both among the public and, more crucially, funding agencies. “The devastating impact of HIV and its lethality have placed research on vaccines and other preventative measures under an unfamiliar spotlight,” said Colonel Jerome Kim, HIV vaccines product manager for the US Army from the Walter Reed Army Institute of Research. “This has tended to exaggerate both the incremental successes and failures of HIV-1 vaccine research.”The devastating impact of HIV and its lethality have placed research on vaccines and other preventative measures under an unfamiliar spotlightThis might have contributed to a decline in public funding that has been combined with a continuing lack of investment from the private sector (AVERT, 2010). More worryingly, there have been signs that governments are withdrawing funding from vaccine research (Médecins Sans Frontières, 2009). In fact, the US government—through the National Institutes of Health—and the Bill and Melinda Gates Foundation—the charitable trust established in 1994 by Microsoft founder Bill Gates—accounted for 79% of the world''s US$868 million funding for HIV/AIDS vaccine research in 2008 (HIV Vaccines and Microbicides Resource Tracking Working Group, 2010).Other areas of vaccination research have also contributed to negative sentiments towards the field. A universal vaccine against influenza has proved similarly elusive, given the mutability of the virus. The recent swine flu pandemic—the severity of which fell short of many pessimistic expectations—left many governments having spent huge sums on vaccines that they never needed. Furthermore, had swine flu become as virulent as it might have, these vaccine stockpiles might still have been only partially effective.More worryingly, there have been signs that governments are withdrawing funding from vaccine researchAll of these factors are fuelling groups who are opposed to vaccine research for various reasons, according to Joachim Hombach, at the World Health Organization''s Initiative for Vaccine Research in Geneva. “There are certain groups, particularly in the industrialised world, that have an anti-vaccine attitude, and these kinds of cases find very fertile ground there,” he said, referring in particular to the Lancet MMR article. “There is also a different story relating to general attitudes towards acceptance of absolutely no risk associated with vaccines or other medical interventions.” This risk-averse culture exposes vaccines to public scrutiny when side effects occur during trials or afterwards, and has even been spreading to developing countries. “This creates a challenging climate for vaccine research,” said Hombach.There are still strong grounds for optimism, as huge strides have been made in understanding the relationship between viruses and the immune response, which is more subtle and diverse than has been previously appreciated. In a sense, diseases such as polio represent the low-hanging fruit in the orchard of infectious disease; early successes in vaccine development have perhaps created a false sense of optimism. “Diseases that are more chronic, where you have a very delicate balance between the pathogen and the immune response, are very difficult to prevent with vaccines,” Hombach said. “HIV is not the only one and there is also TB for instance, which is quite difficult. It is much easier to develop vaccines against acute diseases.”There is accordingly a need to educate the public about these difficulties, commented Tomáš Hanke, Nuffield professor of medicine specializing in HIV research at Oxford University. “Public confidence is a matter of public education and understanding of the process of scientific discovery,” he said. “The more challenging the aim is, the more explaining the public needs.”Researchers disagree about the major problems hindering the development of new vaccines. In the case of HIV, Kim identified the elimination of CD4⁺ helper T cells by the virus as one of the major problems, because these cells are at the centre of immune control and influence the activities of other cells. It is this disabling of helper T cells that weakens the immune response to other diseases in those who have AIDS.Patterson believes that the greatest problem is the virus''s mutability. “The ability of the virus to quickly mutate and escape from responses that are mounted against specific domains of the virus that are recognized by the immune system is, I think, the major hurdle that we need to overcome,” he said. This is one reason why the traditional strategy of inducing protective antibodies by administering an attenuated virus has not worked for HIV.There are two further problems. First, important regions of the HIV virus are shielded by sugars and second, although antibodies that disable a broad range of HIV viruses have been identified, these tend to be produced too late in the immune response, when infection is already well established. For this reason, there has been increased focus on T-cell vaccines that can recognize and kill infected cells, rather than on efforts to prevent infection in the first place. Crucially though, as Patterson pointed out, this approach still faces the problem of mutation, because this can enable the virus to escape recognition by T cells.This, in turn, suggests that vaccines must target regions of the virus that are well conserved. “A normal T cell immune response tends to be against a small number of so-called dominant epitopes and in the case of HIV these are often against the more variable regions the virus can afford to mutate without any cost to itself,” Patterson explained. “To avoid immune escape, we probably need to induce a T cell response against a number of conserved virus epitopes that the virus could not afford to mutate without severely impairing its replication capacity. I believe this aim is achievable.”While the battle against HIV/AIDS has been catching most of the headlines, a lesser known viral disease, dengue, has been rising quickly up the research agendaMore fundamental research at the molecular level is needed to achieve this goal. Robin Weiss, professor of viral oncology at University College London, leads one team who are trying to identify antigens or targets that might one day become useful in vaccine design. Weiss does not claim to be near an imminent breakthrough, but he believes that a major step might be made soon towards developing a broad-spectrum vaccine that could prevent HIV infection in the first place. The problem, as Weiss pointed out, is not that individuals with HIV fail to produce antibodies, but that HIV elicits too many different ones, nearly all of which are ineffective. Only a few HIV-infected individuals produce potent antibodies and even then, these are often in insufficient concentrations. Now that the crystal structure of the potent antibodies has been defined, this vital molecular information could be used to design new vaccines that elicit production of these antibodies in the host.Valuable information has also come from a US Army sponsored clinical trial in Thailand that studied more than 16,000 healthy individuals between 2003 and 2006. A vaccine containing genetically engineered versions of three HIV genes was used, with an inert form of the bird virus canary pox as the vector. In December 2009, the sponsors reported that the rate of HIV infection among volunteers who received the experimental vaccine was 31% lower than among those who received a placebo (Rerks-Ngarm et al, 2009). “[The trial] showed, for the first time, that a vaccine is able to reduce the risk of HIV infection in humans,” said Kim. “Although our results were modest, they are providing a great deal of information to inform the field. For example, the protection appeared highest at 6–12 months based on post-hoc analysis. If we can sustain or increase this effect, that would be a great accomplishment.”While the battle against HIV/AIDS has been catching most of the headlines, a lesser known viral disease, dengue fever, has been rising quickly up the research agenda. Dengue fever is caused by four related strains of flavivirus that are transmitted by mosquitoes. The disease affects 50–100 million people annually, mostly in urban tropical areas, where an estimated 2.5 billion are at risk (Webster et al, 2009), largely because of growing urban populations. Attempts to develop a vaccine have been inhibited by the host immune response to the vaccine, according to Sarah Rowland-Jones, professor of immunology at the Weatherall Institute of Molecular Medicine in Oxford, UK. “It is a tough target for vaccine development principally because of the possibility that immune mechanisms contribute to pathogenesis, so researchers have to be particularly careful that a dengue vaccine does not make disease more likely because of the kind of immune response it stimulates, rather than leading to protection from infection,” she said.The fact that there are four viruses is another problem, especially as a vaccine protecting against one might actually prime individuals for another serotype. “The best hypothesis for severe dengue is that following a first infection, an individual gains immunity against that serotype but becomes more prone to a second infection with a different serotype,” said Jeremy Farrar, Director of the Wellcome Unit in Vietnam and a professor of tropical medicine at Oxford University. “With that second infection against another serotype, more severe disease develops. Obviously this makes vaccine development difficult as one worries individuals will be ‘primed'' by the vaccine and, when they get a natural infection will have more severe disease.”...the effect of each nucleotide change on virus activity is very small, but the cumulative impact of hundreds of such changes causes strong attenuationHowever, Farrar believes this problem has been fixed with the new generation of ‘chimeric vaccines'' that offer protection against all four serotypes. These are based on an exisiting vaccine against the related disease yellow fever, incorporating a part of the dengue virus that triggers an immune response. This chimeric approach has the potential to be extended to develop vaccines against other diseases.This begs the question of why a dengue virus vaccine has not been developed already, given that the related yellow fever vaccine has been available for years. Part of the reason is that dengue is a tougher target, involving four serotypes, but it is also because its mortality is much lower than the 50% rate of yellow fever. “Dengue hasn''t been top of the agenda because it hasn''t caused so much mortality, but there is a lot of morbidity, and it puts a lot of stress on health authorities and has an epidemic potential,” Hombach said. For this reason it has received more funding recently, with phase III clinical trials likely to begin soon, according to Farrar.An entirely different approach for engineering vaccines might also be emerging. Traditional vaccines use mutated virus strains with limited replication abilities, in order to stimulate the immune system. The main drawback of this approach is that many attenuated strains fail to elicit adequate immunity, and it takes a long time to develop such strains. However, Eckard Wimmer and colleagues at Stony Brook University in New York, have developed a computer-aided approach to create attenuated strains without changing the composition and amino-acid sequence of the virus''s proteins (Mueller et al, 2010).They exploit the redundancy of the genetic code; 64 codons code for just 20 amino acids. As most amino acids are coded for by several codons, it is possible to introduce single-nucleotide changes without altering which proteins are expressed, thereby retaining all antigens that might generate an adaptive immune response. Crucially, however, this alters the expression of some genes, which reduces the ability of the virus to replicate. As Wimmer pointed out, the effect of each nucleotide change on virus activity is very small, but the cumulative impact of hundreds of such changes causes strong attenuation. “We call this ‘death by a thousand cuts'',” he said.Moreover, subsequent natural mutations will probably change only one or two of these nucleotide substitutions back to the original, so the chance that the virus will regain its former virulence is very small. The greatest advantage of this approach, however, is speed. “Clearly, such recoded genomes can only be produced by chemical synthesis,” said Wimmer. “They can be designed rapidly—much faster than any other live vaccine that was isolated after long trials of selection.” This, argues Wimmer, makes the approach ideal for countering emerging pandemics when time is short, especially those caused by influenza. Indeed, Wimmer has demonstrated his approach by designing an influenza vaccine (Mueller et al, 2010).Even if this synthetic approach can generate new vaccine candidates more quickly, it will still require development time and clinical trials to assess their efficacy and safety—as is the case for other emerging techniques. New knowledge and new technologies are filtering through to vaccine development; while the disappointments of the past have provided valuable lessons.     

Plant Virus Particles Carrying Tumour Antigen Activate TLR7 and Induce High Levels of Protective Antibody

Induction of potent antibody is the goal of many vaccines targeted against infections or cancer. Modern vaccine designs that use virus-like particles (VLP) have shown efficacy for prophylactic vaccination against virus-associated cancer in the clinic. Here we used plant viral particles (PVP), which are structurally analogous to VLP, coupled to a weak idiotypic (Id) tumour antigen, as a conjugate vaccine to induce antibody against a murine B-cell malignancy. The Id-PVP vaccine incorporates a natural adjuvant, the viral ssRNA, which acts via TLR7. It induced potent protective anti-Id antibody responses in an in vivo mouse model, superior to the “gold standard” Id vaccine, with prevalence of the IgG2a isotype. Combination with alum further increased antibody levels and maintained the IgG2a bias. Engagement of TLR7 in vivo was followed by secretion of IFN-α by plasmacytoid dendritic cells and by activation of splenic CD11c^hi conventional dendritic cells. The latter was apparent from up-regulation of co-stimulatory molecules and from secretion of a wide range of inflammatory cytokines and chemokines including the Th1-governing cytokine IL-12, in keeping with the IgG2a antibody isotype distribution. PVP conjugates are a novel cancer vaccine design, offering an attractive molecular form, similar to VLP, and providing T-cell help. In contrast to VLP, they also incorporate a safe “in-built” ssRNA adjuvant.     

A Controlled Study of Disturbances Recorded Following the Use of Oral Poliomyelitis Vaccine

A double-blind study of disturbances recorded following the administration of a live trivalent oral poliomyelitis vaccine and an identical-appearing placebo was undertaken. Seven hundred and forty-one persons, 81% of the entire population of a small town, constituted the trial group. The two feedings, with a varied feeding sequence, were eight days apart, and at the second feeding and again eight days later, the participants were investigated under controlled conditions. There were no statistically significant differences in the number, nature or timing of the disturbances reported following the administration of the two vaccines. With both vaccines, significantly fewer reactions were reported among males, among the under-5 year group, among Occupational Class I, and after the first feeding as compared with the second. The Salk vaccine immunization status of the participants made no significant difference. With both active vaccine and placebo “vaccine” 14 to 15% of the participants reported some “reaction”.     

Induction of Humoral and Cell-Mediated Immune Responses by Hepatitis B Virus Epitope Displayed on the Virus-Like Particles of Prawn Nodavirus

Hepatitis B virus (HBV) is a deadly pathogen that has killed countless people worldwide. Saccharomyces cerevisiae-derived HBV vaccines based upon hepatitis B surface antigen (HBsAg) is highly effective. However, the emergence of vaccine escape mutants due to mutations on the HBsAg and polymerase genes has produced a continuous need for the development of new HBV vaccines. In this study, the “a” determinant within HBsAg was displayed on the recombinant capsid protein of Macrobrachium rosenbergii nodavirus (MrNV), which can be purified easily in a single step through immobilized metal affinity chromatography (IMAC). The purified protein self-assembled into virus-like particles (VLPs) when observed under a transmission electron microscope (TEM). Immunization of BALB/c mice with this chimeric protein induced specific antibodies against the “a” determinant. In addition, it induced significantly more natural killer and cytotoxic T cells, as well as an increase in interferon gamma (IFN-γ) secretion, which are vital for virus clearance. Collectively, these findings demonstrated that the MrNV capsid protein is a potential carrier for the HBV “a” determinant, which can be further extended to display other foreign epitopes. This paper is the first to report the application of MrNV VLPs as a novel platform to display foreign epitopes.     

A Targeted Mutation within the Feline Leukemia Virus (FeLV) Envelope Protein Immunosuppressive Domain To Improve a Canarypox Virus-Vectored FeLV Vaccine

We previously delineated a highly conserved immunosuppressive (IS) domain within murine and primate retroviral envelope proteins that is critical for virus propagation in vivo. The envelope-mediated immunosuppression was assessed by the ability of the proteins, when expressed by allogeneic tumor cells normally rejected by engrafted mice, to allow these cells to escape, at least transiently, immune rejection. Using this approach, we identified key residues whose mutation (i) specifically abolishes immunosuppressive activity without affecting the “mechanical” function of the envelope protein and (ii) significantly enhances humoral and cellular immune responses elicited against the virus. The objective of this work was to study the immunosuppressive activity of the envelope protein (p15E) of feline leukemia virus (FeLV) and evaluate the effect of its abolition on the efficacy of a vaccine against FeLV. Here we demonstrate that the FeLV envelope protein is immunosuppressive in vivo and that this immunosuppressive activity can be “switched off” by targeted mutation of a specific amino acid. As a result of the introduction of the mutated envelope sequence into a previously well characterized canarypox virus-vectored vaccine (ALVAC-FeLV), the frequency of vaccine-induced FeLV-specific gamma interferon (IFN-γ)-producing cells was increased, whereas conversely, the frequency of vaccine-induced FeLV-specific interleukin-10 (IL-10)-producing cells was reduced. This shift in the IFN-γ/IL-10 response was associated with a higher efficacy of ALVAC-FeLV against FeLV infection. This study demonstrates that FeLV p15E is immunosuppressive in vivo, that the immunosuppressive domain of p15E can modulate the FeLV-specific immune response, and that the efficacy of FeLV vaccines can be enhanced by inhibiting the immunosuppressive activity of the IS domain through an appropriate mutation.     

Development of a challenge-protective vaccine concept by modification of the viral RNA-dependent RNA polymerase of canine distemper virus

We demonstrate that insertion of the open reading frame of enhanced green fluorescent protein (EGFP) into the coding sequence for the second hinge region of the viral L (large) protein (RNA-dependent RNA polymerase) attenuates a wild-type canine distemper virus. Moreover, we show that single intranasal immunization with this recombinant virus provides significant protection against challenge with the virulent parental virus. Protection against wild-type challenge was gained either after recovery of cellular immunity postimmunization or after development of neutralizing antibodies. Insertion of EGFP seems to result in overattenuation of the virus, while our previous experiments demonstrated that the insertion of an epitope tag into a similar position did not affect L protein function. Thus, a desirable level of attenuation could be reached by manipulating the length of the insert (in the second hinge region of the L protein), providing additional tools for optimization of controlled attenuation. This strategy for controlled attenuation may be useful for a “quick response” in vaccine development against well-known and “new” viral infections and could be combined efficiently with other strategies of vaccine development and delivery systems.     

Immunization of mice with recombinant S-adenosyl-L-homocysteine hydrolase protein confers protection against Brucella melitensis infection

Due to drawbacks of live attenuated vaccines, much attention has been focused on screening Brucella-protective antigens as subunit vaccine candidates. Here, an immunoproteomic assay was used to identify the immunogenic soluble proteins of Brucella melitensis 16M. In the present study, 27 unique immunogenic proteins were identified from the two-dimensional electrophoresis immunoblot profiles by liquid chromatography tandem MS (LC-MS/MS). From this set, the gene encoding one immunodominant protein of interest, S-adenosyl-l-homocysteine hydrolase (AdoHcyase), was expressed in Escherichia coli. The recombinant AdoHcyase induced a strong antibody response in BALB/c mice, and the polyclonal antibody could recognize a band of approximately 52 kDa in the immunoblots of soluble protein extracts from five Brucella strains. rAdoHcyase significantly stimulated the production of interferon-γ and interleukin-2, and induced a high level of protection against B. melitensis 16M challenge at 4 weeks postchallenge. Our results indicated that rAdoHcyase could be a useful candidate for the development of subunit vaccines against B. melitensis.     

An ultrapotent pan-β-coronavirus lineage B (β-CoV-B) neutralizing antibody locks the receptor-binding domain in closed conformation by targeting its conserved epitope

New threats posed by the emerging circulating variants of SARS-CoV-2 highlight the need to find conserved neutralizing epitopes for therapeutic antibodies and efficient vaccine design. Here, we identified a receptor-binding domain (RBD)-binding antibody, XG014, which potently neutralizes β-coronavirus lineage B (β-CoV-B), including SARS-CoV-2, its circulating variants, SARS-CoV and bat SARSr-CoV WIV1. Interestingly, antibody family members competing with XG014 binding show reduced levels of cross-reactivity and induce antibody-dependent SARS-CoV-2 spike (S) protein-mediated cell-cell fusion, suggesting a unique mode of recognition by XG014. Structural analyses reveal that XG014 recognizes a conserved epitope outside the ACE2 binding site and completely locks RBD in the non-functional “down” conformation, while its family member XG005 directly competes with ACE2 binding and position the RBD “up”. Single administration of XG014 is effective in protection against and therapy of SARS-CoV-2 infection in vivo. Our findings suggest the potential to develop XG014 as pan-β-CoV-B therapeutics and the importance of the XG014 conserved antigenic epitope for designing broadly protective vaccines against β-CoV-B and newly emerging SARS-CoV-2 variants of concern.Supplementary InformationThe online version contains supplementary material available at 10.1007/s13238-021-00871-6.     

Non-Invasive Pneumococcal Pneumonia in Portugal—Serotype Distribution and Antimicrobial Resistance

There is limited information on the serotypes causing non-invasive pneumococcal pneumonia (NIPP). Our aim was to characterize pneumococci causing NIPP in adults to determine recent changes in serotype prevalence, the potential coverage of pneumococcal vaccines and changes in antimicrobial resistance. Serotypes and antimicrobial susceptibility profiles of a sample of 1300 isolates recovered from adult patients (≥18 yrs) between 1999 and 2011 (13 years) were determined. Serotype 3 was the most frequent cause of NIPP accounting for 18% of the isolates. The other most common serotypes were 11A (7%), 19F (7%), 19A (5%), 14 (4%), 22F (4%), 23F (4%) and 9N (4%). Between 1999 and 2011, there were significant changes in the proportion of isolates expressing vaccine serotypes, with a steady decline of the serotypes included in the 7-valent conjugate vaccine from 31% (1999–2003) to 11% (2011) (P<0.001). Taking together the most recent study years (2009–2011), the potential coverage of the 13-valent conjugate vaccine was 44% and of the 23-valent polysaccharide vaccine was 66%. While erythromycin resistance increased from 8% in 1999–2003 to 18% in 2011 (P<0.001), no significant trend was identified for penicillin non-susceptibility, which had an average value of 18.5%. The serotype distribution found in this study for NIPP was very different from the one previously described for IPD, with only two serotypes in common to the ones responsible for half of each presentation in 2009–2011 – serotypes 3 and 19A. In spite of these differences, the overall prevalence of resistant isolates was similar in NIPP and in IPD.     

Simultaneous Immunization with B.C.G., Diphtheria-tetanus,and Oral Poliomyelitis Vaccines in Children Aged 13-14

The simultaneous administration of B.C.G. vaccine, diphtheria-tetanus toxoid aluminium hydroxide adsorbed vaccine, and oral poliovaccine was studied in 628 children aged 13-14 years between 1966 and 1969 in Newham, London. The efficacy of these vaccines was unaffected by administering them at the same time; routine simultaneous administration is considered justified when organizational difficulties prevent the attainment of high immunization rates with the vaccines given separately. No adverse reactions to B.C.G. or oral poliomyelitis vaccines took place, but 8% of children had moderately severe local reactions after diphtheria-tetanus aluminium hydroxide adsorbed vaccine, which were attributed to diphtheria toxoid.Serological studies showed the need for immunization against diphtheria, tetanus, and poliomyelitis at 13-14 years of age. Because of the adverse reactions to diphtheria toxoid, however, simultaneous administration of tetanus toxoid aluminium hydroxide adsorbed, oral poliomyelitis, and B.C.G. vaccines only is recommended at present.An “adult type” diphtheria-tetanus toxoid might overcome the problem of reactions, though in two to three years'' time most children aged 13-14 years will have received diphtheria-tetanus-pertussis vaccine in infancy and reinforcement might then be accomplished by a small intradermal dose of the currently available fluid diphtheria-tetanus vaccine.Continued serological studies of diphtheria and tetanus antitoxins and polio antibody are necessary to determine the future need for reinforcement of immunity; such studies should become an essential part of the surveillance of the community immunization programme.     

-Classifying prion and prion-like phenomena

The universe of prion and prion-like phenomena has expanded significantly in the past several years. Here, we overview the challenges in classifying this data informatically, given that terms such as “prion-like”, “prion-related” or “prion-forming” do not have a stable meaning in the scientific literature. We examine the spectrum of proteins that have been described in the literature as forming prions, and discuss how “prion” can have a range of meaning, with a strict definition being for demonstration of infection with in vitro-derived recombinant prions. We suggest that although prion/prion-like phenomena can largely be apportioned into a small number of broad groups dependent on the type of transmissibility evidence for them, as new phenomena are discovered in the coming years, a detailed ontological approach might be necessary that allows for subtle definition of different “flavors” of prion / prion-like phenomena.     

Atraumatic oral spray immunization with replication-deficient viral vector vaccines

The development of needle-free vaccines is one of the recently defined “grand challenges in global health” (H. Varmus, R. Klausner, R. Klausner, R. Zerhouni, T. Acharya, A. S. Daar, and P. A. Singer, Science 302:398-399, 2003). To explore whether a natural pathway to the inductive site of the mucosa-associated lymphatic tissue could be exploited for atraumatic immunization purposes, replication-deficient viral vector vaccines were sprayed directly onto the tonsils of rhesus macaques. Tonsillar immunization with viral vector vaccines encoding simian immunodeficiency virus (SIV) antigens induced cellular and humoral immune responses. Viral RNA levels after a stringent SIV challenge were reduced, providing a level of protection similar to that observed after systemic immunization with the same vaccines. Thus, atraumatic oral spray immunization with replication-deficient vectors can overcome the epithelial barrier, deliver the vaccine antigen to the mucosa-associated lymphatic tissue, and avoid induction of tolerance, providing a novel approach to circumvent acceptability problems of syringe and needle vaccines for children and in developing countries.