Formalin-inactivated Vaccine Research Articles

Use of a modified bacterial ghost lysis system for the construction of an inactivated avian pathogenic Escherichia coli vaccine candidate

Vaccination is an effective strategy to prevent avian colibacillosis. Bacterial ghosts (BGs) are prepared by the controlled expression of the phiX174 gene E, which mediates the lysis of Gram-negative bacteria. Staphylococcal nuclease A may be used to produce BGs for further inactivation of host bacteria and elimination of residual genetic material. In this study, the double promoter lysis plasmid (pUC19-ΔcI857-E-rrnB-pL-SN) was successfully constructed and BGs were prepared at 37 °C. The cleavage efficiency of Escherichia coli BGs was 99.9%. Furthermore, to evaluate the immunological effects of the BG vaccines in chickens, a BG vaccine was prepared using the serotype O2 avian pathogenic Escherichia coli deletion strain (DE17ΔluxSΔaroA). The results showed that the BG vaccine was able to achieve over 90% immune protection against virulent challenge using the same serotype O2 strain (DE17 or CE35), while it showed poor cross-protection against serotypes O1 and O78 (data not shown). The enzyme-linked immunosorbent assay results showed that the antibody levels in the immunized groups were higher than in the control group (p < 0.05), with the BG group being the highest. The cytokine tests showed that the levels of interferon-γ in the BG immune group were higher than in the phosphate-buffered saline (PBS) control group (non-immune) (p < 0.01) and the formalin-inactivated vaccine immune group (p < 0.05), and the levels of tumor necrosis factor-α in the BG group were higher than in the formalin-inactivated vaccine (p > 0.05) and the PBS control groups (p < 0.05). In addition, pathological analysis revealed that the PBS control group showed typical fibrinous pericarditis and perihepatitis, whereas the immune group showed no obvious pathological changes. In summary, our findings provide a new strategy for the prevention and control of avian colibacillosis.

Preparation of porcine enterotoxigenic Escherichia coli (ETEC) ghosts and immunogenic analysis in a mouse model

Enterotoxignenic Escherichia coli (ETEC)-associated colibacillosis causes high levels of morbidity and mortality in neonatal piglets. Vaccination is among effective strategy to fight against ETEC-related diseases. Bacterial ghosts (BGs) are empty bacterial envelopes, which substain subtle antigenic comformation in bacterial outer membrane. In this study, a BG vaccine was generated using porcine ETEC isolated strain DQ061 and evaluated its safety and immunogenicity in a mouse model. The recombinant bacteria were constructed by transformation of lysis plasmid pHH43 and generation of BGs was conducted in a lysis rate of 99.93% by incubation of the recombinant bacteria at 42 °C for 2 h. Mice were immunized subcutaneously twice in 2-week intervals with BGs, BGs emulsified with ISA 206 adjuvant, or formalin-inactivated ETEC vaccine after safety test. Mice with either of two BG vaccines developed higher titer of antibodies, secreted higher titer of interleukin 4, gamma interferon and alpha tumor necrosis factor after 2 doses than those with formalin-inactivated ETEC vaccine or those with adjuvant placebo (P < 0.01). The quantity of CD4+ and CD8+ T lymphocyte in spleen was higher in both BG groups than that in the inactivated vaccine group or adjuvant group 2 weeks post boost immunization (P < 0.05). The vaccinated mice were challenged intraperitoneally with 10 × LD50 dose of DQ061. Mice with the BGs plus adjuvant were completely protected against challenge, compared to 60% protection of mice with the inactivated vaccine. Mice exhibited decreased tissue lesion and reduced bacterial loads in the BGs groups by comparison with those with the inactivated vaccine or adjuvant only. Our results validated that the ETEC BGs bear high safety and immunogenicity in a mouse model, suggesting a potential of further evaluation in a pig model.

A Recombinant Subunit Based Zika Virus Vaccine Is Efficacious in Non-human Primates.

Zika Virus (ZIKV), a virus with no severe clinical symptoms or sequelae previously associated with human infection, became a public health threat following an epidemic in French Polynesia 2013–2014 that resulted in neurological complications associated with infection. Although no treatment currently exists, several vaccines using different platforms are in clinical development. These include nucleic acid vaccines based on the prM-E protein from the virus and purified formalin-inactivated ZIKV vaccines (ZPIV) which are in Phase 1/2 clinical trials. Using a recombinant subunit platform consisting of antigens produced in Drosophila melanogaster S2 cells, we have previously shown seroconversion and protection against viremia in an immunocompetent mouse model. Here we demonstrate the efficacy of our recombinant subunits in a non-human primate (NHP) viremia model. High neutralizing antibody titers were seen in all protected macaques and passive transfer demonstrated that plasma from these NHPs was sufficient to protect against viremia in mice subsequently infected with ZIKV. Taken together our data demonstrate the immunogenicity and protective efficacy of the recombinant subunit vaccine candidate in NHPs as well as highlight the importance of neutralizing antibodies in protection against ZIKV infection and their potential implication as a correlate of protection.

Active prophylaxis for respiratory syncytial virus: current knowledge and future perspectives.

Respiratory syncytial virus (RSV) is the most common respiratory pathogen in infants and young children but represents also an important cause of morbidity in adults, particularly in the elderly and immunocompromised persons. Despite its global impact on human health, no effective treatment is available except for supportive care and no safe vaccine has been licensed yet. Vaccine development has been hindered by several factors including vaccine enhanced disease associated with formalin-inactivated RSV vaccine, ethical concerns and lack of consensus concerning the most appropriate target antigen. In this review, we analyze history of RSV vaccine and current approaches for preventing RSV including live-attenuated, vector-based, subunit, nucleic acid-based, particle-based vaccines and we debate about concerns on target population, correlates of protection and obstacles that are slowing the progress toward a successful RSV vaccination strategy.

Enhanced neutralizing antibody response induced by inactivated enterovirus 71 in cynomolgus monkeys.

Enterovirus 71 (EV71) is a major etiological agent of various public health issues, particularly in the Asia-Pacific region. EV71 causes hand-foot-and-mouth disease (HFMD) and is associated with serious neurological disorders in young children. A formalin-inactivated EV71 candidate vaccine (KCDC-HFMDV1-EV71) based on the C4 subgenotype was previously developed and confirmed to be a potential candidate vaccine for prevention of EV71 infection in mice. In this study, an inactivated EV71 vaccine was used for analysis of long-term immunogenicity and efficacy in cynomolgus monkeys, a common nonhuman primate model. The vaccine was immunized three times at 0, 4, and 8 weeks with either 20-μg doses of EV71 candidate vaccine formulated with aluminum hydroxide gel adjuvant or phosphate-buffered saline as a control. The group immunized with the inactivated EV71 showed significantly increased EV71-specific antibody and serum neutralizing antibody titers at 3 weeks after vaccination and maintained these elevated titers until the end of the experiment (54 weeks after vaccination). The sera from vaccinated cynomolgus monkeys showed a crossreactive neutralizing antibody response to the heterologous subtype of EV71 (B1–4, C1, and C2). These findings suggest that the inactivated EV71 candidate vaccine may be a potential vaccine candidate and valuable tool for the control of HFMD.

Vaccine-induced antibody level as the parameter of the influence of environmental salinity on vaccine efficacy in Nile tilapia

To effectively increase production and improve economic returns, the co-culture of Nile tilapia (Oreochromis niloticus) and marine shrimp has been adopted in many countries, including China. Although O. niloticus is an euryhaline fish that can tolerate elevated salinities and even full-strength seawater, fluctuations in salinity levels can undoubtedly induce stress and affect the immune response of this fish. Therefore, this study assessed the impact of salinity on vaccine efficacy in Nile tilapia, which used serum antibody level as a surrogate marker to detect vaccine efficacy. Nile tilapia were acclimatized to 0, 10, 20, or 30 ppt salinity, and then immunized with a formalin-inactivated Streptococcus agalactiae vaccine. Significantly lower levels of antibody in vaccinated fish were found at 20 and 30 ppt salinity compared to 0 and 10 ppt salinity. White blood cell counts, absolute blood lymphocyte counts, and serum bactericidal activity levels were all significantly lower in vaccinated fish at 20 and 30 ppt salinity. Elevated cortisol levels were detected in all of the fish exposure to salinity. Concentrations of serum electrolytes (Na+ and Cl−) were significantly higher in fish at 30 ppt salinity, as compared to fish at lower salinities. Furthermore, the mRNA transcription levels of three of the immune-related genes analyzed (IgM, IL-1β, and IFN-γ, but not Hsp70) were significantly inhibited in the vaccinated fish at 20 and 30 ppt salinity. A suppressed immune response and decreased vaccine efficacy were also indicated by the lower survival rate of vaccinated fish at 20 ppt salinity when challenged with S. agalactiae. Therefore, salinities ≥20 ppt negatively affected antibody production in Nile tilapia, ultimately affecting vaccine efficacy.

Orally administered adenoviral-based vaccine induces respiratory mucosal memory and protection against RSV infection in cotton rats

A vaccine against Respiratory Syncytial Virus (RSV) is a major unmet need to prevent the significant morbidity and mortality that it causes in society. In addition to efficacy, such a vaccine must not induce adverse events, as previously occurred with a formalin-inactivated vaccine (FI-RSV). In this study, the safety, immunogenicity and efficacy of a molecularly adjuvanted adenovirus serotype 5 based RSV vaccine encoding the fusion (F) protein (Ad-RSVF) is demonstrated in cotton rats. Protective immunity to RSV was induced by Ad-RSVF when administered by an oral route as well as by intranasal and intramuscular routes. Compared to FI-RSV, the Ad-RSVF vaccine induced significantly greater neutralizing antibody responses and protection against RSV infection. Significantly, oral or intranasal immunization each induced protective multi-functional effector and memory B cell responses in the respiratory tract. This study uniquely demonstrates the capacity of an orally administered adenovirus vaccine to induce protective immunity in the respiratory tract against RSV in a pre-clinical model and supports further clinical development of this oral Ad-RSVF vaccine strategy.

A virus-like particle-based tetravalent vaccine for hand, foot, and mouth disease elicits broad and balanced protective immunity

Hand, foot, and mouth disease (HFMD) is an infectious disease that mainly affects infants and children, causing considerable morbidity and mortality worldwide. HFMD is commonly caused by enterovirus 71 (EV71) and coxsackieviruses A16 (CVA16), A6 (CVA6), and A10 (CVA10). Formalin-inactivated EV71 vaccines are currently available in China; however, these vaccines fail to confer cross-protection against infections by other HFMD-causing enteroviruses, highlighting the necessity of developing a multivalent HFMD vaccine. Our previous studies demonstrated that recombinant virus-like particles (VLP) of EV71, CVA16, and CVA6 are capable of inducing protective immunity against homologous virus challenges in mice. In this study, we generated CVA10-VLP using a baculovirus-insect cell expression system and then combined CVA10-VLP with EV71-VLP, CVA16-VLP, and CVA6-VLP to formulate a tetravalent VLP vaccine. Immunogenicity and protective efficacy of tetravalent VLP vaccine was compared with that of monovalent VLP vaccines. Mouse immunization studies revealed that the tetravalent vaccine elicited antigen-specific and long-lasting serum antibody responses comparable to those elicited by its corresponding monovalent vaccines. Moreover, tetravalent vaccine immune sera strongly neutralized EV71, CVA16, CVA10, and CVA6 strains with neutralization titers similar to those of their monovalent counterparts, indicating a good compatibility among the four antigens in the combination vaccine. Importantly, passively transferred tetravalent vaccine-immunized sera conferred efficient protection against single or mixed infections with EV71, CVA16, CVA10, and CVA6 viruses in mice, whereas the monovalent vaccines could only protect mice against homotypic virus infections but not heterotypic challenges. These results demonstrate that the tetravalent VLP vaccine represents a promising broad-spectrum HFMD vaccine candidate.

Epidemiology, Pathogenesis, and Control of a Tick-Borne Disease- Kyasanur Forest Disease: Current Status and Future Directions.

In South Asia, Haemaphysalis spinigera tick transmits Kyasanur Forest Disease Virus (KFDV), a flavivirus that causes severe hemorrhagic fever with neurological manifestations such as mental disturbances, severe headache, tremors, and vision deficits in infected human beings with a fatality rate of 3–10%. The disease was first reported in March 1957 from Kyasanur forest of Karnataka (India) from sick and dying monkeys. Since then, between 400 and 500 humans cases per year have been recorded; monkeys and small mammals are common hosts of this virus. KFDV can cause epizootics with high fatality in primates and is a level-4 virus according to the international biosafety rules. The density of tick vectors in a given year correlates with the incidence of human disease. The virus is a positive strand RNA virus and its genome was discovered to code for one polyprotein that is cleaved post-translationally into 3 structural proteins (Capsid protein, Envelope Glycoprotein M and Envelope Glycoprotein E) and 7 non-structural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5). KFDV has a high degree of sequence homology with most members of the TBEV serocomplex. Alkhurma virus is a KFDV variant sharing a sequence similarity of 97%. KFDV is classified as a NIAID Category C priority pathogen due to its extreme pathogenicity and lack of US FDA approved vaccines and therapeutics; also, the infectious dose is currently unknown for KFD. In India, formalin-inactivated KFDV vaccine produced in chick embryo fibroblast is being used. Nevertheless, further efforts are required to enhance its long-term efficacy. KFDV remains an understudied virus and there remains a lack of insight into its pathogenesis; moreover, specific treatment to the disease is not available to date. Environmental and climatic factors involved in disseminating Kyasanur Forest Disease are required to be fully explored. There should be a mapping of endemic areas and cross-border veterinary surveillance needs to be developed in high-risk regions. The involvement of both animal and health sector is pivotal for circumscribing the spread of this disease to new areas.

Establishment of a Bordetella avium challenge model in turkeys

ABSTRACT Despite the importance of Bordetella avium (BA) as a respiratory pathogen of young turkeys, no infection model for the evaluation of BA-vaccine efficacy is available. The objective of this study was to evaluate the influence of route and dose of infection on the establishment of a BA-challenge model. In our first experiment, 28-day-old turkeys were either inoculated oculonasally with 105, 107 or 109 colony forming units (CFU) of BA per bird or exposed to BA by aerosol with 105–108 CFU/m3. The respiratory tract of all inoculated birds was BA-colonized, which was confirmed by choanal swabs and samples of trachea and lung, showing the highest prevalence in the aerosol-inoculated group. BA-specific humoral immune response was detected in the form of IgG in serum from five days post infection (dpi) and IgA in lacrimal fluid from seven dpi. In the second experiment, the model was tested in a vaccination trial. Twenty-one-day-old turkeys were vaccinated with a formalin-inactivated BA vaccine intramuscularly and challenged 21 days post vaccination with 107 CFU per bird oculonasally. BA-specific IgG antibodies were detected in serum and in lacrimal fluid 14 days post vaccination. As in the first experiment, secretory BA-specific antibodies of the IgA isotype were only detected in the inoculated groups from seven dpi. Despite the lack of clinical signs or pathological alterations in both experiments, vaccine efficacy was demonstrated by significant reduction in BA colonization of the trachea (P ≤ 0.05). In our study, a reliable model for BA infection has been established and has been demonstrated to be suitable for evaluation of vaccine efficacy.

Adjuvant activity of Pasteurella multocida A strain, Pasteurella multocida B strain and Salmonella typhimurium bacterial DNA on cellular and humoral immunity responses against Pasteurella multocida specific strain infections in Balb/c mice

Objective: To evaluate the effects of Pasteurella multocida (P. multocida) vaccines on the expression and release of antibodies, interleukin (IL)-6 and IL-12 by serum. Methods: Balb/c mice were immunized with two formalin and iron inactivated vaccine doses within 2 weeks. The vaccines were adjuvant with P. multocida A strain, P. multocida B strain and Salmonella typhimurium bacterial DNA (AbDNA, BbDNA and SbDNA for short, respectively). The animals were challenged 4 weeks after immunization. Blood of mice was collected to detect the change of specific antibody, IL-6, and IL-12 using ELISA. Results: The specific antibody and interleukins in the immunized group increased significantly compared to the control mice after vaccination and challenge (P

Naked DNA immunization with full-length attachment gene of human respiratory syncytial virus induces safe and protective immune response.

Development of potent vaccine for human respiratory syncytial virus (HRSV) that confers better protection than natural infection remains a global challenge. Vaccination with naked DNA is considered successful approach for the control of many viral diseases. In this study, the potential of DNA vaccination using full-length attachment gene of HRSV type A Saudi strain cloned in pcDNA3.1+ vector (pcDNA/GA) was evaluated in BALB/c mice. The expression efficiency of pcDNA/GA was first confirmed in HEp-2 cells on RNA and protein levels. Mice immunization with either pcDNA/GA or the positive control formalin-inactivated vaccine (FI-RSV) has generated significant serum antibody concentration in ELISA (7.31±0.418 and 9.76±0.006 µg/ml, respectively) with superior neutralizing activity. Similarly, both immunogens evoked robust HRSV-specific CD8+ T-cell response in ELISPOT assay compared to mice immunized with pcDNA3.1+ vector or saline (negative controls). Challenge of the immunized mice with the wild-type HRSV did not provoke clinical symptoms or mortality in any mice group. On the 7th day post-challenge, mice were euthanized and lungs were extirpated for evaluation of viral load, histopathological changes and cytokine profile. A significant diminish in the viral load and histology score were concluded in lungs of pcDNA/GA immunized mice compared to those immunized with FI-RSV and negative controls. The pulmonary cytokine profile of pcDNA/GA immunized mice displayed notable upregulation of Th1-associated cytokines while that of FI-RSV immunized mice exhibited high levels of Th2-associated cytokines. In conclusion, the DNA vaccine candidate pcDNA/GA has proven prominent efficacy and safety in mouse model, which encourages further evaluation in clinical trials.

Preliminary aggregate safety and immunogenicity results from three trials of a purified inactivated Zika virus vaccine candidate: phase 1, randomised, double-blind, placebo-controlled clinical trials

A safe, effective, and rapidly scalable vaccine against Zika virus infection is needed. We developed a purified formalin-inactivated Zika virus vaccine (ZPIV) candidate that showed protection in mice and non-human primates against viraemia after Zika virus challenge. Here we present the preliminary results in human beings. We did three phase 1, placebo-controlled, double-blind trials of ZPIV with aluminium hydroxide adjuvant. In all three studies, healthy adults were randomly assigned by a computer-generated list to receive 5 μg ZPIV or saline placebo, in a ratio of 4:1 at Walter Reed Army Institute of Research, Silver Spring, MD, USA, or of 5:1 at Saint Louis University, Saint Louis, MO, USA, and Beth Israel Deaconess Medical Center, Boston, MA, USA. Vaccinations were given intramuscularly on days 1 and 29. The primary objective was safety and immunogenicity of the ZPIV candidate. We recorded adverse events and Zika virus envelope microneutralisation titres up to day 57. These trials are registered at ClinicalTrials.gov, numbers NCT02963909, NCT02952833, and NCT02937233. We enrolled 68 participants between Nov 7, 2016, and Jan 25, 2017. One was excluded and 67 participants received two injections of Zika vaccine (n=55) or placebo (n=12). The vaccine caused only mild to moderate adverse events. The most frequent local effects were pain (n=40 [60%]) or tenderness (n=32 [47%]) at the injection site, and the most frequent systemic reactogenic events were fatigue (29 [43%]), headache (26 [39%]), and malaise (15 [22%]). By day 57, 52 (92%) of vaccine recipients had seroconverted (microneutralisation titre ≥1:10), with peak geometric mean titres seen at day 43 and exceeding protective thresholds seen in animal studies. The ZPIV candidate was well tolerated and elicited robust neutralising antibody titres in healthy adults. Departments of the Army and Defense and National Institute of Allergy and Infectious Diseases.

Induction of lung CD8+ T cell responses by consecutive inoculations of a poly(I:C) influenza vaccine.

The cytotoxic T lymphocyte (CTL) response plays a key role in host recovery from influenza virus infection and in subsequent immunity. Compared to natural infection with influenza virus, however, intranasal vaccination with adjuvant-combined inactivated vaccine elicits only moderate CTL responses. Here we demonstrate that 5 days of consecutive, intranasal vaccination with a combination of inactivated influenza vaccine and poly(I:C) elicits a strong CTL response in the lung. Antigen-captured respiratory DCs did efficiently migrate from the lung to the mediastinal lymph node (mLN) after the 5 day series of inoculations with vaccine and poly(I:C). Importantly, formalin-inactivated whole virus vaccine and poly(I:C) adjuvant have synergic effects on consecutive vaccinations to elicit a strong CTL response in the lung. Although the CTL response was less effective against heterologous influenza virus, we show for the first time that intranasal administration of inactivated influenza virus vaccine and poly(I:C) for 5 consecutive days can elicit high levels of influenza virus-specific CD8+ T cells in the lung.

Lack of in vivo cross-protection of two different betanodavirus species RGNNV and SJNNV in European sea bass Dicentrachus labrax

Viral encephalopathy and retinopathy (VER) is a severe infective disease characterized by neuropathological changes in several fish species associated with high mortality. The etiological agent is a virus belonging to the Nodaviridae family, genus Betanodavirus. To date, four different betanodavirus species have been officially recognized by International Committee on Taxonomy of Viruses (ICTV), namely the red-spotted grouper- (RGNNV), the striped jack- (SJNNV), the barfin flounder- (BFNNV) and the tiger puffer nervous necrosis virus (TPNNV). Moreover, two reassortants RGNNV/SJNNV and SJNNV/RGNNV have been described.Betanodaviruses can be classified into three different serotypes (A, B and C) that are antigenically different, so none (between serotype A and C) or partial (between serotype B and C) cross-immunoreactivity has been detected in vitro.In this study we investigated the in vivo cross-protection of the two main betanodavirus species (RGNNV and SJNNV), which belong to distinct serotype, by immunizing intraperitoneally (IP) juvenile sea bass with formalin inactivated RGNNV and SJNNV vaccines, followed by a challenge with RGNNV.Fish IP vaccinated with inactivated RGNNV showed a high protection value (85%). Serological analyses highlighted a great specific anti-NNV immunoglobulin M (IgM) production against the homologous virus, while a good seroconversion with low neutralization property was highlighted against the heterologous virus. In fish IP vaccinated with inactivated SJNNV the protection recorded was equal to 25%, significantly lower respect to the one provided by RGNNV IP vaccine. ELISA test detected good IgM production against the homologous virus, and a lower, but still detectable IgM production against the heterologous one. By contrast, serum neutralization test highlighted a poorly detectable antibody production unable to neutralize either the homologous or the heterologous virus.These results confirm that the two serotypes are not cross-protective in vivo. According to these findings, the production of multivalent formulation, or at least the provision of different types of vaccines based on both fish and virus species requirement, should be recommended in order to broaden the range of protection.

Optimized production and purification of Coxsackievirus B1 vaccine and its preclinical evaluation in a mouse model

Coxsackie B viruses are among the most common enteroviruses, causing a wide range of diseases. Recent studies have also suggested that they may contribute to the development of type 1 diabetes. Vaccination would provide an effective way to prevent CVB infections, and the objective of this study was to develop an efficient vaccine production protocol for the generation of novel CVB vaccines. Various steps in the production of a formalin-inactivated Coxsackievirus B1 (CVB1) vaccine were optimized including the Multiplicity Of Infection (MOI) used for virus amplification, virus cultivation time, type of cell growth medium, virus purification method and formulation of the purified virus. Safety and immunogenicity of the formalin inactivated CVB1 vaccine was characterized in a mouse model. Two of the developed methods were found to be optimal for virus purification: the first employed PEG-precipitation followed by gelatin-chromatography and sucrose cushion pelleting (three-step protocol), yielding 19-fold increase in virus concentration (0.06µg/cm2) as compared to gold standard method. The second method utilized tandem sucrose pelleting without a PEG precipitation step, yielding 83-fold increase in virus concentration (0.24µg/cm2), but it was more labor-intensive and cannot be efficiently scaled up. Both protocols provide radically higher virus yields compared with traditional virus purification protocols involving PEG-precipitation and sucrose gradient ultracentrifugation. Formalin inactivation of CVB1 produced a vaccine that induced a strong, virus-neutralizing antibody response in vaccinated mice, which protected against challenge with CVB1 virus. Altogether, these results provide valuable information for the development of new enterovirus vaccines.

An inactivated hand-foot-and-mouth disease vaccine using the enterovirus 71 (C4a) strain isolated from a Korean patient induces a strong immunogenic response in mice.

Enterovirus 71 (EV71) is a major causative agent of hand-foot-and-mouth disease (HFMD) frequently occurring in children. HFMD induced by EV71 can cause serious health problems and has been reported worldwide, particularly in the Asia-Pacific region. In this study, we assessed the immunogenicity of a formalin-inactivated HFMD vaccine using an EV71 strain (FI-EV71 C4a) isolated from a Korean patient. The vaccine candidate was evaluated in mice to determine the vaccination doses and vaccine schedules. BALB/c mice were intramuscularly administered 5, 10, or 20 μg FI-EV71 vaccine, followed by a booster 2 weeks later. EV71-specific antibodies and neutralizing antibodies were induced and maintained until the end of the experimental period in all vaccinated groups. To determine the effectiveness of adjuvant for the EV71 vaccine, three adjuvants, i.e., aluminium hydroxide gel, monophosphoryl lipid A, and polyinosinic-polycytidylic acid, were administered separately with the FI-EV71 vaccine to mice via the intramuscular route. Mice administered the FI-EV71 vaccine formulated with all three adjuvants induced a significantly increased antibody response compared with that of the single adjuvant groups. The vaccinated group with triple adjuvants exhibited more rapid induction of EV71-specific and neutralizing antibodies than the other groups. These results suggested that the role of adjuvant in inactivated vaccine was important for eliciting effective immune responses against EV71. In conclusion, our results showed that FI-EV71 was a potential candidate vaccine for prevention of EV71 infection.

A formalin-inactivated vaccine provides good protection against Vibrio harveyi infection in orange-spotted grouper (Epinephelus coioides)

Vibrio harveyi is one of the most common threats to farmed grouper, so considerable efforts are in practice to control the pathogen. This study presents a highly effective vaccine against V. harveyi in the orange-spotted grouper with the help of a single intraperitoneal immunization. The vaccine candidate was in form of whole, formalin-inactivated V. harveyi cells combined with a metabolizable ISA763 AVG adjuvant. Our results indicated that the vaccine triggered a remarkably higher expression level of interleukin (IL)-1β, IL-6, IL-8, and IL-10 in the groupers' kidneys and spleens, as recorded after 1 and 3 days of immunization. Antibody titers were significantly elevated in the vaccinated fish. A pivotal observation was that the vaccine highly protected the grouper from a homologous V. harveyi strain challenge with relative percentage survival values of 100% and 91.7% at 6 and 12 weeks post-immunization, respectively. Vaccinated fish also demonstrated strong cross-protection against a heterologous bacterial isolate challenge. Therefore, the inactivated V. harveyi vaccine is a promising candidate that could stimulate good immune responses and confer remarkable protection in farmed groupers.

CD4+ T cells support establishment of RSV-specific IgG and IgA antibody secreting cells in the upper and lower murine respiratory tract following RSV infection

The RSV vaccine field suffered a major set-back when children were vaccinated with a formalin-inactivated RSV vaccine (FI-RSV). Unexpectedly, the vaccinated children fared worse than unvaccinated children when they were naturally infected with RSV. Mouse models were then developed that implicated the CD4+ T helper cell population as a contributor to adverse events. Today, the T cell is viewed with much caution in the RSV field, and its induction by vaccination is sometimes discouraged. Here we re-emphasize the beneficial role of the CD4+ T cell. Experiments were performed with RSV-infected nude mice that received CD4+ T cells by adoptive transfer. Data demonstrated that CD4+ T cells were necessary for the induction of mucosal and systemic RSV-specific antibodies, for the establishment of RSV-specific IgG and IgA antibody secreting cells in the upper and lower respiratory tract, and for RSV clearance.

Fowl adenovirus: history, emergence, biology and development of a vaccine against hydropericardium syndrome.

The poultry industry has emerged as one of the largest and fastest growing public sectors in the developed and developing countries. Unfortunately, this industry is under a major threat from diseases that are viral (Newcastle disease, infectious bursal disease, influenza, hydropericardium syndrome), bacterial (colibacillosis, pasteurellosis, salmonellosis, mycoplasmosis), parasitic (coccidiosis, histoplasmosis) or nutritional (dyschondroplasia, osteoporosis). Among these diseases, hydropericardium syndrome (HPS) is one of the important emerging diseases occurring in the specific areas of the world where broilers (chickens) are reared under intensive conditions. HPS was first observed in 1987 at Angara Goth, an area near Karachi, Pakistan, where broilers are raised. Since then, HPS has been reported in many countries of the world. From these reported cases, an adenovirus that was either isolated from or visualized electron microscopically in the liver of affected broilers has been implicated in the syndrome. The syndrome has been reproduced by inoculation of isolated fowl adenovirus (FAdV) strains, and hence, the syndrome is also called infectious hydropericardium syndrome. To our knowledge, HPS has not been observed in humans, so it is not considered a zoonotic disease, but it is of economic importance and causes huge losses to the poultry industry. Efforts have been made to develop conventional vaccines against this disease, which were formulated from infected liver homogenate. Formalin-inactivated liver organ vaccines have failed to protect the poultry industry. Hence, there is a dire need to develop a suitable vaccine to combat this disease. Currently, recombinant vaccine candidates are being developed by using molecular biology and biotechnological approaches for the prevention and control of infectious diseases, including HPS. Therefore, it is suggested that the immunogenicity of these recombinant proteins should be evaluated for their use as subunit vaccines.