Eosinophils as drivers of bacterial immunomodulation and persistence.

Traditionally, eosinophils have been linked to parasitic infections and pathological disease states. However, emerging literature has unveiled a more nuanced and intricate role for these cells, demonstrating their key functions in maintaining mucosal homeostasis. Eosinophils exhibit diverse phenotypes and exert multifaceted effects during infections, ranging from promoting pathogen persistence to triggering allergic reactions. Our investigations primarily focus on Bordetella spp., with particular emphasis on Bordetella bronchiseptica, a natural murine pathogen that induces diseases in mice akin to pertussis in humans. Recent findings from our published work have unveiled a striking interaction between B. bronchiseptica and eosinophils, facilitated by the btrS-mediated mechanism. This interaction serves to enhance pathogen persistence while concurrently delaying adaptive immune responses. Notably, this role of eosinophils is only noted in the absence of a functional btrS signaling pathway, indicating that wild-type B. bronchiseptica, and possibly other Bordetella spp., possess such adeptness in manipulating eosinophils that the true function of these cells remains obscured during infection. In this review, we present the mounting evidence pointing toward eosinophils as targets of bacterial exploitation, facilitating pathogen persistence and fostering chronic infections in diverse mucosal sites, including the lungs, gut, and skin. We underscore the pivotal role of the master regulator of Bordetella pathogenesis, the sigma factor BtrS, in orchestrating eosinophil-dependent immunomodulation within the context of pulmonary infection. These putative convergent strategies of targeting eosinophils offer promising avenues for the development of novel therapeutics targeting respiratory and other mucosal pathogens.

Comparison of the long-term and short-term protection in mouse model of Leishmania major infection following vaccination with Live Iranian Lizard Leishmania mixed with chitin microparticles.

Inducing long-term immunity is the primary goal of vaccination. Leishmanisation using non-pathogenic to human Leishmania spp. could be considered a reliable approach to immunising subjects against Leishmania infection. Here, we evaluated the long-term immune responses (14 weeks) after immunisation with either live- or killed-Iranian Lizard Leishmania (ILL) mixed with chitin microparticles (CMPs) against L. major infection in BALB/c mice. In total, nine groups of mice were included in the study. To evaluate short-term immunity, mice were immunised with live-ILL and CMPs and 3 weeks later were challenged with L. majorEGFP . To evaluate the long-term immunity, mice were immunised with either live- or killed-ILL both mixed with CMPs, and 14 weeks after immunisation, mice were challenged with L. majorEGFP . A group of healthy mice who received no injection was also included in the study. Eight weeks after the challenge with L. majorEGFP , all subjects were sacrificed and the parasite burden (quantitative real-time PCR and in vivo imaging), cytokines levels (IFN-γ, IL-4 and IL-10), Leishmania-specific antibody concentration, and total levels of IgG1 and IgG2a were measured. In addition, nitric oxide concentration and arginase activity were evaluated. Results showed that in mice that were immunised using live-ILL+CMP, the induced protective immune response lasted at least 14 weeks; since they were challenged with L. majorEGFP at the 14th -week post-immunisation, no open lesion was formed during the 8-week follow-up, and the footpad swelling was significantly lower than controls. They also showed a significant reduction in the parasite burden in splenocytes, compared to the control groups including the group that received killed-ILL+CMP. The observed protection was associated with a higher IFN-γ and a lower IL-10 production by splenocytes. Additionally, the results demonstrated that arginase activity was decreased in the ILL+CMP group compared to other groups. Immunisation with ILL alone reduced the parasite burden compared to non-immunised control; however, it was still significantly higher than the parasite burden in the ILL+CMP groups. In conclusion, the long-term immune response against L. major infection induced by Live-ILL+CMP was more competent than the response elicited by killed-ILL+CMP to protect mice against infection with L. majorEGFP .

Intrapulmonary Vaccination Induces Long-lasting and Effective Pulmonary Immunity Against Staphylococcus aureus Pneumonia.

BACKGROUND: Staphylococcus aureus causes community- and hospital-acquired pneumonia linked to a high mortality rate. The emergence and rapid transmission of multidrug-resistant S. aureus strains has become a serious health concern, highlighting the challenges associated with the development of a vaccine to combat S. aureus pneumonia. METHODS: This study evaluated the effects of intrapulmonary immunization on the immune response and protection against S. aureus lung infection in a respiratory mouse model using a subunit vaccine. RESULTS: Compared with the intranasal immunized mice, the intrapulmonarily immunized mice had lower levels of pulmonary bacterial colonization and lethality, accompanied by alleviated lung inflammation with reduced proinflammatory cytokines and increased levels of interleukin-10 and antimicrobial peptide following intrapulmonary challenge. Optimal protection was associated with increased pulmonary antibodies and resident memory T cells. Moreover, intrapulmonary immunization provided long-lasting pulmonary protection for at least 6 months, with persistent cellular and humoral immunity in the lungs. CONCLUSIONS: Vaccine reaching the deep lung by intrapulmonary immunization plays a significant role in the induction of efficacious and long-lasting immunity against S. aureus in the lung parenchyma. Hence, intrapulmonary immunization can be a strategy for the development of a vaccine against S. aureus pneumonia.Immunization through the intrapulmonary route with a subunit of S. aureus vaccine elicited tissue resident memory T cells and antigen-specific antibodies in the lungs, and provided optimal and long-term protection against S. aureus pneumonia.

Clearance of attenuated rabies virus from brain tissues is required for long-term protection against CNS challenge with a pathogenic variant.

Rabies virus is a neurotropic lyssavirus which is 100% fatal in its pathogenic form when reaching unprotected CNS tissues. Death can be prevented by mechanisms delivering appropriate immune effectors across the blood-brain barrier which normally remains intact during pathogenic rabies virus infection. One therapeutic approach is to superinfect CNS tissues with attenuated rabies virus which induces blood-brain barrier permeability and immune cell entry. Current thinking is that peripheral rabies immunization is sufficient to protect against a challenge with pathogenic rabies virus. While this is undoubtedly the case if the virus is confined to the periphery, what happens if the virus reaches the CNS is less well-understood. In the current study, we find that peripheral immunization does not fully protect mice long-term against an intranasal challenge with pathogenic rabies virus. Protection is significantly better in mice that have cleared attenuated virus from the CNS and is associated with a more robust CNS recall response evidently due to the presence in CNS tissues of elevated numbers of lymphocytes phenotypically resembling long-term resident immune cells. Adoptive transfer of cells from rabies-immune mice fails to protect against CNS challenge with pathogenic rabies virus further supporting the concept that long-term resident immune cell populations must be established in brain tissues to protect against a subsequent CNS challenge with pathogenic rabies virus.

Persistence of Wild-Type Japanese Encephalitis Virus Strains Cross-Neutralization 5 Years After JE-CV Immunization.

BACKGROUND: The live-attenuated Japanese encephalitis (JE) vaccine (JE-CV; IMOJEV) induces a protective response in children. A shift in circulating JE virus strains suggests that a genotype shift phenomenon may occur throughout Southeast Asia. We assessed the neutralization of wild-type (WT) JE virus isolates at distal time points after vaccination. METHODS: We analyzed serum samples from a subset of 47 children who had received a JE-CV booster after an inactivated JE vaccine primary immunization. We measured antibody titers (50% plaque reduction neutralization test) using a panel of WT JE strains at baseline, then after the booster at 28 days and 6 months in all subjects present at the time points and in a subset at year 5. Three additional recent isolates were tested at year 5. RESULTS: Of 47 subjects, 43 (91.5%) subjects had JE neutralizing antibody titers ≥10 (reciprocal serum dilution) against the homologous strain before JE-CV boost; all were seroprotected up to year 5 after the JE-CV boost. Baseline WT seroprotection ranged between 78.7% and 87.2%; all subjects were seroprotected against the 4 WT strains at 28 days and 6 months; year 5 seroprotection ranged between 95.7% and 97.9%. Similar rates of protection against 3 additional WT isolates were observed at year 5. CONCLUSIONS: The long-term immune responses induced after a JE-CV booster dose in toddlers were able to neutralize WT viruses from various genotypes circulating in Southeast Asia and India. CLINICAL TRIALS REGISTRATION: NCT00621764.

Long-Term Follow-Up of Children after Venom Immunotherapy: Low Adherence to Anaphylaxis Guidelines.

BACKGROUND: Data on the long-term outcome of children after specific venom immunotherapy (VIT) are limited. Therefore, we assessed sting recurrence and anaphylaxis relapse rates as well as adherence to anaphylaxis guidelines with regard to the availability of emergency equipment and education status. METHODS: For this long-term survey, data of 311 children with a history of anaphylactic reactions to hymenoptera stings were collected by chart review. We included patients who were treated with a 3-year VIT between 1993 and 2009 and had completed a questionnaire. RESULTS: Forty of the 311 patients were included. Mean VIT duration was 3.1 years. Of the 40 patients included, 29 children (72.5%) received VIT with vespid venom, 9 with bee venom, and 2 patients with both venoms. During a mean follow-up period of 13 years, 20/40 patients (50%) suffered re-stings. Six of the 20 (30%) patients developed again anaphylactic symptoms (grade 1 n = 5, grade 3 n = 1); 2 were allergic to vespid and 4 to bee venom. Of the entire cohort, only 5/40 (12.5%) had appropriate emergency kits according to the guidelines of the European Academy of Allergy and Clinical Immunology. Among the patients who had emergency kits available, one third (5/15) felt uncertain about the correct application of the medication. Less than two thirds of our population (25/40) affirmed that they have been educated in emergency management. The vast majority (95%; 38/40) of our patients did not have allergy follow-ups after VIT completion. CONCLUSIONS: Anaphylactic relapses are not uncommon, and there are considerable deficits in the emergency management of patients. Hence, comprehensive standardized anaphylaxis education programs as well as regular follow-ups of the allergy status are crucial.

Apical Organelle Secretion by Toxoplasma Controls Innate and Adaptive Immunity and Mediates Long-Term Protection.

Apicomplexan parasites have unique apical rhoptry and microneme secretory organelles that are crucial for host infection, although their role in protection against Toxoplasma gondii infection is not thoroughly understood. Here, we report a novel function of the endolysosomal T. gondii sortilin-like receptor (TgSORTLR), which mediates trafficking to functional apical organelles and their subsequent secretion of virulence factors that are critical to the induction of sterile immunity against parasite reinfection. We further demonstrate that the T. gondii armadillo repeats-only protein (TgARO) mutant, which is deficient only in apical secretion of rhoptries, is also critical in mounting protective immunity. The lack of TgSORTLR and TgARO proteins completely inhibited T-helper 1-dependent adaptive immunity and compromised the function of natural killer T-cell-mediated innate immunity. Our findings reveal an essential role for apical secretion in promoting sterile protection against T. gondii and provide strong evidence for rhoptry-regulated discharge of antigens as a key effector for inducing protective immunity.

Adverse and long-term protective effects following oil-adjuvanted vaccination against Aeromonas salmonicida in rainbow trout.

Prophylactic measures against Aeromonas salmonicida subsp. salmonicida, the causative agent of furunculosis, have been an active field of research for decades, with studies mainly focused on Atlantic salmon (Salmo salar). In the present study we have examined the protective and adverse effects of mineral oil-adjuvanted injection vaccines on rainbow trout (Oncorhynchus mykiss). A commercial vaccine and an experimental auto vaccine, as well as their respective adjuvant formulations alone were used to evaluate their individual effects, both prior to and during an experimental waterborne infection challenge. Macro- and microscopic examination revealed signs of vaccine-induced adverse effects from 10 weeks to 14 months post vaccination. Both vaccines induced statistically significant protection during the experimental challenge (P=0.018 for both vaccines), as well as significantly elevated levels of specific circulating antibodies prior to and during the experimental challenge when compared to an unvaccinated control group. During the early, critical time points of the infection, both vaccines appeared to protect against pathological changes to the liver and spleen, which provides a probable explanation for the reduced mortality seen in the vaccinated groups. A significant correlation was found between the level of A. salmonicida-specific antibodies measured prior to challenge and the endpoint survival of each group after the experimental infection, and furthermore, the levels of these antibodies remained elevated for at least 14 months post vaccination.

Circular RNA vaccines with long-term lymph node-targeting delivery stability after lyophilization induce potent and persistent immune responses.

IMPORTANCE: messenger RNA (mRNA) vaccines are a key technology in combating existing and emerging infectious diseases. However, the inherent instability of mRNA and the nonspecificity of lipid nanoparticle-encapsulated (LNP) delivery systems result in the need for cold storage and a relatively short-duration immune response to mRNA vaccines. Herein, we develop a novel vaccine in the form of circRNAs encapsulated in LNPs, and the circular structure of the circRNAs enhances their stability. Lyophilization is considered the most effective method for the long-term preservation of RNA vaccines. However, this process may result in irreversible damage to the nanoparticles, particularly the potential disruption of targeting modifications on LNPs. During the selection of lymph node-targeting ligands, we found that LNPs modified with mannose maintained their physical properties almost unchanged after lyophilization. Additionally, the targeting specificity and immunogenicity remained unaffected. In contrast, even with the addition of cryoprotectants such as sucrose, the physical properties of LNPs were impaired, leading to an obvious decrease in immunogenicity. This may be attributed to the protective role of mannose on the surface of LNPs during lyophilization. Freshly prepared and lyophilized mLNP-circRNA vaccines elicited comparable immune responses in both the rabies virus model and the SARS-CoV-2 model. Our data demonstrated that mLNP-circRNA vaccines elicit robust immune responses while improving stability after lyophilization, with no compromise in tissue targeting specificity. Therefore, mannose-modified LNP-circRNA vaccines represent a promising vaccine design strategy.

Long-Term Protection against Virulent Newcastle Disease Virus (NDV) in Chickens Immunized with a Single Dose of Recombinant Turkey Herpesvirus Expressing NDV F Protein.

Newcastle disease (ND) is a significant infectious disease in poultry, causing substantial economic losses in developing countries. To control ND, chickens must be vaccinated multiple times a year. In order to develop an improved vaccine that provides long-term protection, the F gene from genotype VII NDV was inserted into the herpesvirus of turkey (HVT) vaccine virus using CRISPR/Cas9-mediated NHEJ repair and Cre/LoxP technology. The immunogenicity and protective efficacy of the resulting recombinant vaccines were evaluated through antibody assays and virus challenge experiments. Two recombinant vaccines, rHVT-005/006-F and rHVT-US2-F, were generated, both exhibiting growth rates comparable with those of HVT in vitro and consistently expressing the F protein. One-day-old specific pathogen-free (SPF) chickens immunized with 2000 PFU/bird of either rHVT-005/006-F or rHVT-US2-F developed robust humoral immunity and were completely protected against challenge with the NDV F48E8 strain at 4 weeks post-vaccination (wpv). Furthermore, a single dose of these vaccines provided sustained protection for at least 52 wpv. Our study identifies rHVT-005/006-F and rHVT-US2-F as promising ND vaccine candidates, offering long-term protection with a single administration. Moreover, HVT-005/006 demonstrates promise for accommodating additional foreign genes, facilitating the construction of multiplex vaccines.

The Impact of Smoking on Long-Term Protection Following Hepatitis B Vaccination: A 24-Year Cohort Study.

The hepatitis B vaccination-induced immune response has been demonstrated to be associated with a number of factors, including age, sex, BMI, and the presence of comorbidities. Additionally, modifiable determinants such as smoking have been identified as influencing the response to vaccination. However, despite the evidence that smokers are at an increased risk of not responding to vaccination, the long-term effects of smoking on antibody persistence remain poorly understood. This study aims to assess the impact of smoking habits on long-term immunity following the primary vaccination cycle. Participants were required to have received a standard three-dose vaccine schedule in childhood, without subsequent doses, and to be between 18 and 24 years of age. Data on age, sex, BMI, age of administration of the first vaccine dose, and time between doses were collected. An antibody concentration < 10 IU/L was considered as non-protective. A total of 2133 individuals were included, 14.2% of whom were smokers. The mean age was 20.28 ± 0.92 years. The probability of having a non-protective antibody concentration was significantly higher in smokers than in non-smokers (AOR: 1.287; 95% CI: 1.002-1.652). The detrimental effects of smoking extend beyond the immediate effects on the vaccine response, also impairing the long-term immune response in individuals who received vaccinations during childhood.

Hybrid Ginseng-derived Extracellular Vesicles-Like Particles with Autologous Tumor Cell Membrane for Personalized Vaccination to Inhibit Tumor Recurrence and Metastasis.

Personalized cancer vaccines based on resected tumors from patients is promising to address tumor heterogeneity to inhibit tumor recurrence or metastasis. However, it remains challenge to elicit immune activation due to the weak immunogenicity of autologous tumor antigens. Here, a hybrid membrane cancer vaccine is successfully constructed by membrane fusion to enhance adaptive immune response and amplify personalized immunotherapy, which formed a codelivery system for autologous tumor antigens and immune adjuvants. Briefly, the functional hybrid vesicles (HM-NPs) are formed by hybridizing ginseng-derived extracellular vesicles-like particles (G-EVLPs) with the membrane originated from the resected autologous tumors. The introduction of G-EVLPs can enhance the phagocytosis of autologous tumor antigens by dendritic cells (DCs) and facilitate DCs maturation through TLR4, ultimately activating tumor-specific cytotoxic T lymphocytes (CTLs). HM-NPs can indeed strengthen specific immune responses to suppress tumors recurrence and metastasis including subcutaneous tumors and orthotopic tumors. Furthermore, a long-term immune protection can be obtained after vaccinating with HM-NPs, and prolonging the survival of animals. Overall, this personalized hybrid autologous tumor vaccine based on G-EVLPs provides the possibility of mitigating tumor recurrence and metastasis after surgery while maintaining good biocompatibility.

Bioprotective Respirator Assembled by Defective Carbon Nitride for Long-Term Light Triggered Health Protection.

Wearing face masks is the best way to stop the spread of respiratory infections. However, if masks are not sterilized, changing them too frequently can actually increase the risk of cross-contamination. Herein, the construction of an antipathogen photocatalytic mask with carbon vacancy-modified carbon nitride nanosheets (g-C3N4-VC Ns) coated on the non-woven fabrics of the out layer of the mask, offering effective and long-term protection against damaging pathogens when exposed to light is reported. The introduced carbon vacancies are found capable of creating energy-disordered sites and inducing energetic electric force to overcome the Coulomb interactions between electron-hole pairs, thus promoting the electron-hole separation to achieve a high generation of reactive oxygen species (ROS). Thanks to its high activity in generating ROS upon exposure to light, the as-prepared photocatalytic mask shows high pathogen sterilization performance. This, in turn, prolongs the mask's protective lifetime, decreases the need for regular replacement, and decreases medical waste production. The work demonstrated here opens new viewpoints in designing pathogens biocidal protective devices for health protection, offering significant promise in specific environment self-protection.

Age-dependent sting recurrence and outcome in immunotherapy-treated children with anaphylaxis to Hymenoptera venom.

BACKGROUND: Data on outcome of insect venom immunotherapy in children are rare. OBJECTIVE: We investigated the rate of sting recurrence and outcome of Hymenoptera venom anaphylaxis in children of different age groups treated with immunotherapy. METHODS: Data from children consecutively referred for anaphylaxis to Hymenoptera venom were collected using a standardized questionnaire. RESULTS: During mean follow-up of 7.7 years after commencement of immunotherapy, 45 of 83 children (56%) were re-stung 108 times by the insect they were allergic to. This corresponds to a rate of 0.23 stings per child and year of follow-up. The younger the subject, the higher was the prevalence of re-stings, with rates of 0.41 in children < 6 years, 0.21 at school age and 0.15 in adolescents (P = 0.001). In contrast, prevalence of systemic allergic reactions to field stings was significantly lower in pre-school (3.4%) and school-age children (4.3%) compared with adolescents (15.6%; P < 0.05). Overall, prevalence of systemic allergic reactions at re-sting was 15.6% in the honey bee venom and 5.9% in the Vespula venom allergic group (P = ns). Younger boys with anaphylaxis to honey bee venom predominated in our cohort (P = 0.019). CONCLUSION AND CLINICAL RELEVANCE: A majority of children with anaphylaxis to Hymenoptera venom (56%) in our cohort were re-stung, equally by honey bees or Vespula species. Younger children were more likely to be re-stung, but less likely to have a systemic reaction. Venom immunotherapy induces long-term protection in most children: 84.4% of subjects with anaphylaxis to honey bee and 94.1% of those to Vespula venom were completely protected at re-stings.

Recombinant BCG expressing the LTAK63 adjuvant increased memory T cells and induced long-lasting protection against Mycobacterium tuberculosis challenge in mice.

Vaccine-induced protection against Mycobacterium tuberculosis (Mtb) is usually ascribed to the induction of Th1, Th17, and CD8+ T cells. However, protective immune responses should also involve other immune cell subsets, such as memory T cells. We have previously shown improved protection against Mtb challenge using the rBCG-LTAK63 vaccine (a recombinant BCG strain expressing the LTAK63 adjuvant, a genetically detoxified derivative of the A subunit from E. coli heat-labile toxin). Here we show that mice immunized with rBCG-LTAK63 exhibit a long-term (at least until 6 months) polyfunctional Th1/Th17 response in the draining lymph nodes and in the lungs. This response was accompanied by the increased presence of a diverse set of memory T cells, including central memory, effector memory and tissue-resident memory T cells. After the challenge, the T cell phenotype in the lymph nodes and lungs were characterized by a decrease in central memory T cells, and an increase in effector memory T cells and effector T cells. More importantly, when challenged 6 months after the immunization, this group demonstrated increased protection in comparison to BCG. In conclusion, this work provides experimental evidence in mice that the rBCG-LTAK63 vaccine induces a persistent increase in memory and effector T cell numbers until at least 6 months after immunization, which correlates with increased protection against Mtb. This improved immune response may contribute to enhance the long-term protection.

Variable long-term protection by radiation-, chemo-, and genetically-attenuated Plasmodium berghei sporozoite vaccines.

Long-term protection against malaria remains one of the greatest challenges of vaccination against this deadly parasitic disease. Whole-sporozoite (WSp) malaria vaccine formulations, which target the Plasmodium parasite's pre-erythrocytic stages, include radiation-attenuated sporozoites (RAS), early- and late-arresting genetically-attenuated parasites (EA-GAP and LA-GAP, respectively), and chemoprophylaxis with sporozoites (CPS). Although all these four vaccine formulations induce protective immune responses in the clinic, data on the longevity of the antimalarial protection they afford remain scarce. We employed a mouse model of malaria to assess protection conferred by immunization with P. berghei (Pb)-based surrogates of these four WSp formulations over a 36-week period. We show that EA-GAP WSp provide the lowest overall protection against an infectious Pb challenge, and that while immunization with RAS and LA-GAP WSp elicits the most durable protection, the protective efficacy of CPS WSp wanes rapidly over the 36-week period, most notably at higher immunization dosages. Analyses of liver immune cells show that CD44hi CD8+ T cells in CPS WSp-immunized mice express increased levels of the co-inhibitory PD-1 and LAG-3 markers compared to mice immunized with the other WSp formulations. This indicates that memory CD8+ T cells elicited by CPS WSp immunization display a more exhausted phenotype, which may explain the rapid waning of protection conferred by the former. These results emphasize the need for a detailed comparison of the duration of protection of different WSp formulations in humans and suggest a more beneficial effect of RAS and LA-GAP WSp compared to EA-GAP or CSP WSp.

Pretreatment of outer membrane vesicle and subsequent infection with influenza virus induces a long-lasting adaptive immune response against broad subtypes of influenza virus.

Influenza is an acute respiratory disease and a global health problem. Although influenza vaccines are commercially available, frequent antigenic changes in hemagglutinin might render them less effective or unavailable. We previously reported that modified outer membrane vesicle (fmOMV) provided immediate and robust protective immunity against various subtypes of influenza virus. However, the effect was transient because it was innate immunity-dependent. In this study, we investigated the effects of consecutive administration of fmOMV and influenza virus on the adaptive immune response and long-term protective immunity against influenza virus. When the mice were pretreated with fmOMV and subsequently infected with influenza virus, strong influenza-specific antibody and T cell responses were induced in both systemic and lung mucosal compartments without pathogenic symptoms. Upon the secondary viral challenge at week 4, the mice given fmOMV and influenza virus exhibited almost complete protection against homologous and heterologous viral challenge. More importantly, this strong protective immunity lasted up to 18 weeks after the first infection. These results show that pretreatment with fmOMV and subsequent infection with influenza virus efficiently induces broad and long-lasting protective immunity against various virus subtypes, suggesting a novel antiviral strategy against newly-emerging viral diseases without suitable vaccines or therapeutics.

Long-term efficacy of nasal vaccination against enteric red mouth (ERM) disease and infectious hematopoietic necrosis (IHN) in juvenile rainbow trout (Oncorhynchus mykiss).

Previous research demonstrated that bacterial and viral vaccines delivered via the nasal route in rainbow trout (Oncorhynchus mykiss) at 7 and 28 days post-vaccination are highly protective (>95% protection). Long-term protection following nasal vaccination in teleosts has not been evaluated. The goal of this study was to assess efficacy and immune responses at 6 months (mo) post-vaccination (mpv), and long-lasting immune responses at 12 mpv of two different vaccines: an inactivated enteric red mouth disease (ERM) Yersinia ruckeri bacterin and a live attenuated infectious hematopoietic necrosis virus (IHNV) vaccine. Juvenile rainbow trout were vaccinated for Y. ruckeri via intraperitoneal (I.P.) and intranasal (I.N.) routes, and for IHNV by intramuscular (I.M.) and I.N. routes, then challenged at 6 mpv. Immune responses were determined at 6 and 12 mpv. ERM vaccine I.P. delivery elicited significantly higher serum IgM-specific titers that remained elevated compared to mock-vaccinated fish at 6 mpv. By 12 mpv, antibody titers to Y. ruckeri were not significantly different across all treatments. Following Y. ruckeri challenge at 6 mpv, a significant difference in cumulative percent mortality (CPM) was found for I.P.-vaccinated fish but not I.N.-vaccinated fish. I.M. and I.N. vaccination with live attenuated IHNV did not result in significant specific serum IgM titers at 6 or 12 mpv. Yet, I.N.-vaccinated fish showed the lowest CPM 6 mpv indicating long-term protection that does not correlate with systemic IgM responses. Repertoire analyses confirmed unique expansions of VH-JH rearrangements in the spleen of rainbow trout 12 mpv that varied with the type of vaccine and route of vaccination. Combined, these data demonstrate that I.N. vaccination with a live attenuated viral vaccine confers long lasting protection, but I.N. ERM vaccination does not and booster before 6 mpv is recommended.

Potential of Polyethyleneimine as an Adjuvant To Prepare Long-Term and Potent Antifungal Nanovaccine.

Background: Candida albicans infections are particularly prevalent in immunocompromised patients. Even with appropriate treatment with current antifungal drugs, the mortality rate of invasive candidiasis remains high. Many positive results have been achieved in the current vaccine development. There are also issues such as the vaccine's protective effect is not persistent. Considering the functionality and cost of the vaccine, it is important to develop safe and efficient new vaccines with long-term effects. In this paper, an antifungal nanovaccine with Polyethyleneimine (PEI) as adjuvant was constructed, which could elicit more effective and long-term immunity via stimulating B cells to differentiate into long-lived plasma cells. Materials and Methods: Hsp90-CTD is an important target for protective antibodies during disseminated candidiasis. Hsp90-CTD was used as the antigen, then introduced SDS to "charge" the protein and added PEI to form the nanovaccine. Dynamic light scattering and transmission electron microscope were conducted to identify the size distribution, zeta potential, and morphology of nanovaccine. The antibody titers in mice immunized with the nanovaccine were measured by ELISA. The activation and maturation of long-lived plasma cells in bone marrow by nanovaccine were also investigated via flow cytometry. Finally, the kidney of mice infected with Candida albicans was stained with H&E and PAS to evaluate the protective effect of antibody in serum produced by immunized mice. Results: Nanoparticles (NP) formed by Hsp90-CTD and PEI are small, uniform, and stable. NP had an average size of 116.2 nm with a PDI of 0.13. After immunizing mice with the nanovaccine, it was found that the nano-group produced antibodies faster and for a longer time. After 12 months of immunization, mice still had high and low levels of antibodies in their bodies. Results showed that the nanovaccine could promote the differentiation of B cells into long-lived plasma cells and maintain the long-term existence of antibodies in vivo. After immunization, the antibodies in mice could protect the mice infected by C. albicans. Conclusion: As an adjuvant, PEI can promote the differentiation of B cells into long-lived plasma cells to maintain long-term antibodies in vivo. This strategy can be adapted for the future design of vaccines.

Long-Term Protection Elicited by an Inactivated Vaccine Supplemented with a Water-Based Adjuvant against Infectious Hematopoietic Necrosis Virus in Rainbow Trout (Oncorhynchus mykiss).

Previous inactivated vaccines against infectious hematopoietic necrosis (IHN) usually had a strong early immune protective effect but failed to provide long-term protection in rainbow trout (Oncorhynchus mykiss). To find a method for stabilizing the desired protective effect of IHN vaccines, we assessed the immune enhancement effect of four adjuvants on formaldehyde inactivated vaccine for IHN at 60 days postvaccination (dpv). The efficacy of a two-dose vaccination with the candidate adjuvant-formaldehyde inactivated vaccine for IHN was evaluated in terms of early protection and long-term protection (30 to 285 dpv). Neutralizing antibody titers were also measured at each time point. The Montanide GEL 02 PR (Gel 02) adjuvant significantly enhanced the immune protection provided by the IHN inactivated vaccine, whereas the immune-boosting effect of the other tested adjuvants lacked statistical significance. Both tested Gel 02-adjuvanted IHN inactivated vaccine dosages had a strong immune protection effect within 2 months postvaccination, with a relative percent of survival (RPS) of 89.01% to 100%, and the higher dosage provided complete protection at 204 dpv and a RPS of 60.79% on 285 dpv by reducing viral titers in rainbow trout. The neutralizing antibodies were observed only in vaccinated fish on 30 and 60 dpv. Through compatibility with an appropriate adjuvant, the highly immune protective effect of an IHN inactivated vaccine was prolonged from 60 dpv to at least 284 dpv; this novel adjuvant-IHN inactivated vaccine has promise as a commercial vaccine that provides the best available and longest duration of protection against IHN to rainbow trout. IMPORTANCE Infectious hematopoietic necrosis virus (IHNV) is one of the most serious pathogens threatening the global salmon and trout industry. However, there is currently only one commercialized infectious hematopoietic necrosis (IHN) vaccine, and it is inadequate for solving the global IHN problem. In this study, a promising adjuvanted inactivated vaccine with long-term protection was developed and comprehensively studied. We confirmed the presence of a late antiviral response stage in vaccinated rainbow trout that lacked detectable neutralizing antibodies, which are commonly recognized to be responsible for long-term specific protection in mammals. These findings further our understanding of unique features of fish immune systems and could lead to improved prevention and control of fish diseases.