Multi-subunit Vaccine Research Articles

Target antigen screening and development of a multi-component subunit vaccine against Mycoplasma synoviae in chickens

IntroductionMycoplasma synoviae (MS) is a globally important avian pathogen causing infectious synovitis and respiratory diseases in poultry, leading to significant economic losses. Despite advances in vaccine development, a commercially viable subunit vaccine against MS remains elusive.MethodsWe sequenced whole genomes of six clinical MS strains isolated from different Chinese provinces. Common genes were analyzed using Biopython software, identifying those with high copy numbers in virulent strains and shared among all strains. Vaxign2 and IEDB Antibody Epitope Prediction were used to analyze protein properties. We assessed immune protective effects of candidate proteins and developed a multivalent subunit vaccine.ResultsTen candidate vaccine proteins were initially selected. A multivalent subunit vaccine composed of MSPB, Ppht, Cfba, and EF-G displayed the best protective effect. The optimal immunization dosage was 20μg, with each protein accounting for 25%. The immune production period was determined to be 28 days post-first immunization, lasting 180 days. The immune protection rate against highly virulent strains reached 90%∼100%.DiscussionThis study provides a new approach for screening vaccine antigens and develops an effective candidate vaccine for MS prevention. The multivalent subunit vaccine shows promising results in protecting against MS infections, potentially offering a solution to reduce economic losses in the poultry industry.

Mass Spectrometric and Artificial Intelligence-Based Identification of the Secretome of Plasmodium falciparum Merozoites to Provide Novel Candidates for Vaccine Development Pipeline.

Merozoites are the only extracellular form of blood stage parasites, making it a worthwhile target. Multiple invasins that are stored in the merozoite apical organelles, are secreted just prior to invasion, and mediates its interaction with RBC. A comprehensive identification of all these secreted invasins is lacking and this study addresses that gap. Pf3D7 merozoites were enriched and triggered to discharge apical organelle contents by exposure to ionic conditions mimicking that of blood plasma. The secreted proteins were separated from cellular contents and both the fractions were subjected to proteomic analysis. Also, the identified secreted proteins were subjected to GO, PPI network analysis, and AI-based in silico approach to understand their vaccine candidacy. A total of 63 proteins were identified in the secretory fraction with membrane and apical organellar localization. This includes various MSPs, micronemal EBAs and rhoptry bulb proteins, which play a crucial role in initial and late merozoite attachment, and majority of them qualified as vaccine candidates. We, for the first time, report the secretory repertoire of merozoite and its status for vaccine candidacy. This information can be utilized to develop better invasion blocking multisubunit vaccines, comprising of immunological epitopes from several secreted invasins.

A multistage protein subunit vaccine as BCG-booster confers protection against Mycobacterium tuberculosis infection in murine models

The eradication of tuberculosis remains a global challenge. Despite being the only licensed vaccine, Bacillus Calmette-Guérin (BCG) confers limited protective efficacy in adults and individuals with latent tuberculosis infections (LTBI). There is an urgent need to develop novel vaccines that can enhance the protective effect of BCG. Protein subunit vaccines have garnered significant research interest due to their safety and plasticity. Based on previous studies, we selected three antigens associated with LTBI (Rv2028c, Rv2029c, Rv3126c) and fused them with an immunodominant antigen Ag85A, resulting in the construction of a multistage protein subunit vaccine named A986. We evaluated the protective effect of recombinant protein A986 adjuvanted with MPL/QS21 as a booster vaccine for BCG against Mycobacterium tuberculosis (Mtb) infection in mice. The A986 + MPL/QS21 induced the secretion of antigen-specific Th1 (IL-2+, IFN-γ+ and TNF-α+) and Th17 (IL-17A+) cytokines in CD4+ and CD8+ T cells within the lung and spleen of mice, while also increased the frequency of central memory and effector memory T cells. Additionally, it also induced the enhanced production of IgG antibodies. Compared to BCG alone, A986 + MPL/QS21 boosting significantly augmented the proliferation of antigen-specific multifunctional T cells and effectively reduced bacterial load in infected mice. Taken together, A986 + MPL/QS21 formulation induced robust antigen-specific immune responses and provided enhanced protection against Mtb infection as a booster of BCG vaccine.

Immunogenicity of a Rotavirus VP8* Multivalent Subunit Vaccine in Mice.

Rotavirus remains a significant public health threat, especially in low-income countries, where it is the leading cause of severe acute childhood gastroenteritis, contributing to over 128,500 deaths annually. Although the introduction of the Rotarix and RotaTeq vaccines in 2006 marked a milestone in reducing mortality rates, approximately 83,158 preventable deaths persisted, showing ongoing challenges in vaccine accessibility and effectiveness. To address these issues, a novel subcutaneous vaccine formulation targeting multiple rotavirus genotypes has been developed. This vaccine consists of nine VP8* proteins from nine distinct rotavirus genotypes and sub-genotypes (P[4], P[6], P[8]LI, P[8]LIII, P[8]LIV, P[9], P[11], P[14], and P[25]) expressed in E. coli. Two groups of mice were immunized either with a single immunogen, the VP8* from the rotavirus Wa strain (P[8]LI), or with the nonavalent formulation. Preliminary results from mouse immunization studies showed promising outcomes, eliciting antibody responses against six of the nine immunogens. Notably, significantly higher antibody titers against VP8* P[8]LI were observed in the group immunized with the nonavalent vaccine compared to mice specifically immunized against this genotype alone. Overall, the development of parenteral vaccines targeting multiple rotavirus genotypes represents a promising strategy in mitigating the global burden of rotavirus-related morbidity and mortality, offering new avenues for disease prevention and control.

An immunoinformatics approach for design and validation of multi-subunit vaccine against Plasmodium falciparum from essential hypothetical proteins.

The online version contains supplementary material available at 10.1007/s12639-024-01696-w.

Development of multi epitope subunit vaccines against emerging carp viruses Cyprinid herpesvirus 1 and 3 using immunoinformatics approach

Cyprinid herpesvirus is a causative agent of a destructive disease in common and koi carp (Cyprinus carpio), which leads to substantial global financial losses in aquaculture industries. Among the strains of C. herpesvirus, C. herpesvirus 1 (CyHV-1) and C. herpesvirus 3 (CyHV-3) are known as highly pathogenic to carp fishes in Europe, Asia, and Africa. To date, no effective vaccine has been developed to combat these viruses. This study aimed to develop unique multi-epitope subunit vaccines targeting the CyHV-1 and CyHV-3 using a reverse vaccinology approach. The study began with a comprehensive literature review to identify the most critical proteins, which were then subjected to in silico analyses to predict highly antigenic epitopes. These analyses involved assessing antigenicity, transmembrane topology screening, allergenecity, toxicity, and molecular docking approaches. We constructed two multi-epitope-based vaccines incorporating a suitable adjuvant and appropriate linkers. It revealed that both the vaccines are non-toxic and immunogenic. The tertiary structures of the vaccine proteins were generated, refined, and validated to ensure their suitability. The binding affinity between the vaccine constructs and TLR3 and TLR5 receptors were assessed by molecular docking studies. Molecular dynamics simulations indicated that vaccine construct V1 exhibited greater stability with both TLR3 and TLR5 based on RMSD analysis. Hydrogen bond analysis revealed a stronger binding affinity between the vaccine constructs and TLR5 compared to TLR3. Furthermore, MM-PBSA analysis suggested that both vaccine constructs exhibited a better affinity for TLR5. Considering all aspects, the results suggest that in silico development of CyHV vaccines incorporating multiple epitopes holds promise for management of diseases caused by CyHV-1 and CyHV-3. However, further in vivo trials are highly recommended to validate the efficacies of these vaccines.

An inclusive approach to designing a multi-epitope chimeric vaccine for Taenia infections by integrating proteomics and reverse vaccinology

BackgroundSoil- and water-transmitted helminths are a major concern in the developing world due to their high prevalence. More than a quarter of the population were estimated to be infected with helminths in these endemic zones.Research designAn in silico approach was used to design a vaccine construct against the Taenia genus utilizing the proteomic information and evaluation of the construct using immune-informatics.ResultsOur study identified 451 conserved proteins in Taenia spp. using the existing proteome; out of these, 141 were found to be expressed in cysticerci. These proteins were screened for antigenic epitopes and a multi-subunit vaccine was constructed. The constructed vaccine was assessed for its efficacy in mounting the appropriate immune response. Our constructed vaccine showed stability and optimal performance against the TLR 4 receptor, which is reported to be upregulated in Taenia infections in hosts.ConclusionImmune-informatics tools help design vaccines for neglected diseases such as those attributed to helminths, which are known to cause widespread morbidity. Our vaccine construct holds tremendous potential in conferring protection against all Taenia spp. of clinical relevance to human.

BCG-booster vaccination with HSP90-ESAT-6-HspX-RipA multivalent subunit vaccine confers durable protection against hypervirulent Mtb in mice

The quest for effective and enhanced multiantigenic tuberculosis (TB) subunit vaccine necessitates the induction of a protective pathogen-specific immune response while circumventing detrimental inflammation within the lung milieu. In line with this goal, we engineered a modified iteration of the quadrivalent vaccine, namely HSP90-ESAT-6-HspX-RipA (HEHR), which was coupled with the TLR4 adjuvant, CIA09A. The ensuing formulation was subjected to comprehensive assessment to gauge its protective efficacy against the hypervirulent Mycobacterium tuberculosis (Mtb) Haarlem clinical strain M2, following a BCG-prime boost regimen. Regardless of vaccination route, both intramuscular and subcutaneous administration with the HEHR vaccine exhibited remarkable protective efficacy in significantly reducing the Mtb bacterial burden and pulmonary inflammation. This underscores its notably superior protective potential compared to the BCG vaccine alone or a former prototype, the HSP90-E6 subunit vaccine. In addition, this superior protective efficacy was confirmed when testing a tag-free version of the HEHR vaccine. Furthermore, the protective immune determinant, represented by durable antigen-specific CD4+IFN-γ+IL-17A+ T-cells expressing a CXCR3+KLRG1- cell surface phenotype in the lung, was robustly induced in HEHR-boosted mice at 12 weeks post-challenge. Collectively, our data suggest that the BCG-prime HEHR boost vaccine regimen conferred improved and long-term protection against hypervirulent Mtb strain with robust antigen-specific Th1/Th17 responses.

Development of a nano-emulsion based multivalent protein subunit vaccine against Pseudomonas aeruginosa.

Pseudomonas aeruginosa (Pa) is an opportunistic bacterial pathogen responsible for severe hospital acquired infections in immunocompromised and elderly individuals. Emergence of increasingly drug resistant strains and the absence of a broad-spectrum prophylactic vaccine against both T3SA+ (type III secretion apparatus) and ExlA+/T3SA- Pa strains worsen the situation in a post-pandemic world. Thus, we formulated a candidate subunit vaccine (called ExlA/L-PaF/BECC/ME) against both Pa types. This bivalent vaccine was generated by combining the C-terminal active moiety of exolysin A (ExlA) produced by non-T3SA Pa strains with our T3SA-based vaccine platform, L-PaF, in an oil-in-water emulsion. The ExlA/L-PaF in ME (MedImmune emulsion) was then mixed with BECC438b, an engineered lipid A analogue and a TLR4 agonist. This formulation was administered intranasally (IN) to young and elderly mice to determine its potency across a diverse age-range. The elderly mice were used to mimic the infection seen in elderly humans, who are more susceptible to serious Pa disease compared to their young adult counterparts. After Pa infection, mice immunized with ExlA/L-PaF/BECC/ME displayed a T cell-mediated adaptive response while PBS-vaccinated mice experienced a rapid onset inflammatory response. Important genes and pathways were observed, which give rise to an anti-Pa immune response. Thus, this vaccine has the potential to protect aged individuals in our population from serious Pa infection.

De novo identification of bacterial antigens of a clinical isolate by combining use of proteosurfaceomics, secretomics, and BacScan technologies.

Emerging infectious diseases pose a significant threat to both human and animal populations. Rapid de novo identification of protective antigens from a clinical isolate and development of an antigen-matched vaccine is a golden strategy to prevent the spread of emerging novel pathogens. Here, we focused on Actinobacillus pleuropneumoniae, which poses a serious threat to the pig industry, and developed a general workflow by integrating proteosurfaceomics, secretomics, and BacScan technologies for the rapid de novo identification of bacterial protective proteins from a clinical isolate. As a proof of concept, we identified 3 novel protective proteins of A. pleuropneumoniae. Using the protective protein HBS1_14 and toxin proteins, we have developed a promising multivalent subunit vaccine against A. pleuropneumoniae. We believe that our strategy can be applied to any bacterial pathogen and has the potential to significantly accelerate the development of antigen-matched vaccines to prevent the spread of an emerging novel bacterial pathogen.

Prediction of an immunogenic peptide ensemble and multi-subunit vaccine for Visceral leishmaniasis using bioinformatics approaches

Visceral Leishmaniasis (VL) is a neglected tropical disease of public health importance in the Indian subcontinent. Despite consistent elimination initiatives, the disease has not yet been eliminated and there is an increased risk of resurgence from active VL reservoirs including asymptomatic, post kala azar dermatitis leishmaniasis (PKDL) and HIV-VL co-infected individuals. To achieve complete elimination and sustain it in the long term, a prophylactic vaccine, which can elicit long lasting immunity, is desirable. In this study, we employed immunoinformatic tools to design a multi-subunit epitope vaccine for the Indian population by targeting antigenic secretory proteins screened from the Leishmania donovani proteome. Out of 8014 proteins, 277 secretory proteins were screened for their cellular location and proteomic evidence. Through NCBI BlastP, unique fragments of the proteins were cropped, and their antigenicity was evaluated. B-cell, HTL and CTL epitopes as well as IFN-ɣ, IL-17, and IL-10 inducers were predicted, manually mapped to the fragments and common regions were tabulated forming a peptide ensemble. The ensemble was evaluated for Class I MHC immunogenicity and toxicity. Further, immunogenic peptides were randomly selected and used to design vaccine constructs. Eight vaccine constructs were generated by linking random peptides with GS linkers. Synthetic TLR-4 agonist, RS09 was used as an adjuvant and linked with the constructs using EAAK linkers. The predicted population coverage of the constructs was ∼99.8 % in the Indian as well as South Asian populations. The most antigenic, nontoxic, non-allergic construct was chosen for the prediction of secondary and tertiary structures. The 3D structures were refined and analyzed using Ramachandran plot and Z-scores. The construct was docked with TLR-4 receptor. Molecular dynamic simulation was performed to check for the stability of the docked complex. Comparative in silico immune simulation studies showed that the predicted construct elicited humoral and cell-mediated immunity in human host comparable to that elicited by Leish-F3, which is a promising vaccine candidate for human VL.

A novel multistage antigens ERA005f confer protection against Mycobacterium tuberculosis by driving Th-1 and Th-17 type T cell immune responses.

Tuberculosis (TB) is a major threat to human health. In 2021, TB was the second leading cause of death after COVID-19 among infectious diseases. The Bacillus Calmette-Guérin vaccine (BCG), the only licensed TB vaccine, is ineffective against adult TB. Therefore, there is an urgent need to develop new effective vaccines. In this study, we developed a novel multistage subunit vaccine (ERA005f) comprising various proteins expressed in metabolic states, based on three immunodominant antigens (ESAT-6, Rv2628, and Ag85B). We utilized the E. coli prokaryotic expression system to express ERA005f and subsequently purified the protein using nickel affinity chromatography and anion exchange. Immunogenicity and protective efficacy of ERA005f and ERA005m were evaluated in BALB/c mice. ERA005f was consistently expressed as an inclusion body in a prokaryotic expression system, and a highly pure form of the protein was successfully obtained. Both ERA005f and ERA005m significantly improved IgG titers in the serum. In addition, mice immunized with ERA005f and ERA005m generated higher titers of antigen-specific IgG2a than the other groups. Elispot results showed that, compared with other groups, ERA005f increased the numbers of IFN-γ-secreting and IL-4-secreting T cells, especially the number of IFN-γ-secreting T cells. Meanwhile, ERA005f induced a higher number of IFN-γ+ T lymphocytes than ERA005m did. In addition, ERA005f improved the expression of cytokines, including IFN-γ, IL-12p70, TNF-α, IL-17, and GM-CSF and so on. Importantly, both ERA005f and ERA005m significantly inhibited the growth of Mtb. The novel multistage antigen ERA005f elicited a strong antigen-specific humoral response and Th-1 and Th-17 cell-mediated immunity in mice. Meanwhile, it can effectively inhibit H37Rv growth in vitro, and represents a correlate of protection in vivo, indicating that ERA005f may exhibit excellent protective efficacy against Mycobacterium tuberculosis H37Rv infection. Our study suggests that ERA005f has the potential to be a promising multistage tuberculosis vaccine candidate.

Characterization of exclusively non-commensal Neisseria gonorrhoeae pangenome to prioritize globally conserved and thermodynamically stable vaccine candidates using immune-molecular dynamic simulations

Neisseria gonorrhoeae (Ngo) has emerged as a global threat leading to one of the most common sexually transmitted diseases in the world. It has also become one of the leading antimicrobial resistant organisms, resulting in fewer treatment options and an increased morbidity. Therefore, in recent years, there has been an increased focus on the development of new treatments and preventive strategies to combat its infection. In this study, we have combined the most conserved epitopes from the completely assembled strains of Ngo to develop a universal and a thermodynamically stable vaccine candidate. For our vaccine design, the epitopes were selected for their high immunogenicity, non-allergenicity and non-cytotoxicity, making them the ideal candidates for vaccine development. For the screening process, several reverse vaccinology tools were employed to rigorously extract non-homologous and immunogenic epitopes from the selected proteins. Consequently, a total number of 3 B-cell epitopes and 6 T-cell epitopes were selected and joined by multiple immune-modulating adjuvants and linkers to generate a promiscuous immune response. Additionally, the stability and flexible nature of the vaccine construct was confirmed using various molecular dynamic simulation tools. Overall, the vaccine candidate showed promising binding affinity to various HLA alleles and TLR receptors; however, further studies are needed to assess its efficacy in-vivo. In this way, we have designed a multi-subunit vaccine candidate to potentially combat and control the spread of N. gonorrhoeae.

Multi-epitope vaccine against drug-resistant strains of Mycobacterium tuberculosis: a proteome-wide subtraction and immunoinformatics approach.

Mycobacterium tuberculosis (Mtb) is the causative agent of tuberculosis, one of the most deadly infections in humans. The emergence of multidrug-resistant and extensively drug-resistant Mtb strains presents a global challenge. Mtb has shown resistance to many frontline antibiotics, including rifampicin, kanamycin, isoniazid, and capreomycin. The only licensed vaccine, Bacille Calmette-Guerin, does not efficiently protect against adult pulmonary tuberculosis. Therefore, it is urgently necessary to develop new vaccines to prevent infections caused by these strains. We used a subtractive proteomics approach on 23 virulent Mtb strains and identified a conserved membrane protein (MmpL4, NP_214964.1) as both a potential drug target and vaccine candidate. MmpL4 is a non-homologous essential protein in the host and is involved in the pathogen-specific pathway. Furthermore, MmpL4 shows no homology with anti-targets and has limited homology to human gut microflora, potentially reducing the likelihood of adverse effects and cross-reactivity if therapeutics specific to this protein are developed. Subsequently, we constructed a highly soluble, safe, antigenic, and stable multi-subunit vaccine from the MmpL4 protein using immunoinformatics. Molecular dynamics simulations revealed the stability of the vaccine-bound Toll-like receptor-4 complex on a nanosecond scale, and immune simulations indicated strong primary and secondary immune responses in the host. Therefore, our study identifies a new target that could expedite the design of effective therapeutics, and the designed vaccine should be validated. Future directions include an extensive molecular interaction analysis, in silico cloning, wet-lab experiments, and evaluation and comparison of the designed candidate as both a DNA vaccine and protein vaccine.

Designing and Expression of Recombinant Chimeric Spike Protein from SARS-CoV-2 in Escherichia coli and Its Immunogenicity Assessment.

Since December 2019, the world has been grappling with an ongoing global COVID-19 pandemic. Various virus variants have emerged over the past two years, each posing a greater threat than its predecessors. The recent appearance of the omicron variant (B.1.1.529) has raised significant alarm within the field of epidemiology due to its highly contagious nature and rapid transmission rate. The omicron variant possessed mutations in the key receptor-binding domain (RBD) region, the S region, and these modifications have shown a notable impact on the strain's susceptibility to neutralizing antibodies. Developing safe and efficient vaccines to prevent a future severe acute respiratory outbreak of coronavirus syndrome 2 (SARS-CoV-2) is significant. Viral surface spike proteins are ideal targets for vaccines. This study aimed to find a multi-subunit chimeric vaccine. After conducting bioinformatics analysis, the recombinant spike (RS) protein of SARS-CoV-2 was deliberately designed and subsequently produced using E. coli expression systems. The immunogenicity of RS and neutralizing antibody responses were evaluated on immunized BALB/c mice. There was a significant difference in antibody titers between RS-immunized mice and control groups. The endpoint of the serum antibody titer of mice immunized with our chimeric protein was 2.5 times higher than that of the negative control. The chimeric construct could present multiple antigens simultaneously, influentially affecting immunization. Sera from mice vaccinated by RS could recognize the SARS-CoV-2 virus and neutralize antibodies. Our chimeric peptide could bind to antibodies in the serum of patients infected with different serotypes of the SARS-CoV-2 virus, such as alpha, delta, and omicron variants. The results indicated that the RS protein would be a potential novel antigenic candidate for subunit vaccine development and could be used as a useful alternative to generate diagnostic serological tests for SARS-CoV-2 infection.

The leptospiral OmpA-like protein (Loa22) is a surface-exposed antigen that elicits bactericidal antibody against heterologous Leptospira

Leptospirosis is the most widespread zoonosis, affecting over 1 million humans each year, with more than 60,000 deaths worldwide. Leptospirosis poses a significant health threat to dogs, horses, cattle, and wildlife. The disease may be self-limiting or progress to a life-threatening multi-system disorder affecting the kidneys, liver, and lungs. Currently, bacterin vaccine formulations that consist of one or more laboratory-cultivated strains are used for prevention. However, the antibody response elicited by these vaccines is directed primarily at lipopolysaccharide and is generally serovar-specific. The development of broadly protective subunit vaccines for veterinary and human applications would be a significant step forward in efforts to combat this emerging and antigenically variable pathogen. This study assessed the properties and potential utility of the Leptospira Loa22 (Leptospira OmpA-like 22 kDa protein) protein as a vaccine antigen. Loa22 is a virulence factor that is predicted to transverse the outer membrane and present its N-terminal domain on the cell surface. This report demonstrates that diverse Leptospira strains express Loa22 in vitro and that the protein is antigenic during infection in dogs. Immunoblot and size exclusion chromatography revealed that Loa22 exists in monomeric and trimeric forms. Immunization of rats with recombinant Loa22 elicited bactericidal antibodies against diverse Leptospira strains. The immunodominant bactericidal epitopes were localized within the N-terminal domain using protein-blocking bactericidal assays. This study supports the utility of Loa22, or subfragments thereof, in developing a multivalent chimeric subunit vaccine to prevent leptospirosis and sheds new light on the cellular localization of Loa22.

In silico designing of a novel polyvalent multi-subunit peptide vaccine leveraging cross-immunity against human visceral and cutaneous leishmaniasis: an immunoinformatics-based approach

Leishmaniasis is a group of vector-borne infectious diseases caused by over 20 pathogenic Leishmania species that are endemic in many tropical and subtropical countries. The emergence of drug-resistant strains, the adverse side effects of anti-Leishmania drugs, and the absence of a preventative vaccination strategy threaten the sensitive population. Recently, many groups of researchers have exploited the field of reverse vaccinology to develop vaccines, focusing chiefly on inducing immunity against either visceral or cutaneous leishmaniasis. This present work involves retrieving twelve experimentally validated leishmanial antigenic protein sequences from the UniProt database, followed by their antigenicity profiling employing ANTIGENpro and Vaxijen 2.0 servers. MHC-binding epitopes for the same were predicted using both NetCTL 1.2 and SYFPEITHI servers, while epitopes for B cell were computed using ABCpred and BepiPred 2.0 servers. The screened epitopes with significantly higher scores were utilized for designing the vaccine construct with appropriate linkers and natural adjuvant. The secondary and tertiary structures of the synthetic peptide were determined by conditional random fields, shallow neural networks, and profile-profile threading alignment with iterative structure assembly simulations, respectively. The 3-D vaccine model was validated through CASP10-tested refinement and the MolProbity web server. Molecular docking and multi-scale normal mode analysis simulation were performed to analyze the best vaccine-TLR complex. Finally, computational immune simulation findings revealed promising cellular and humoral immune responses, suggesting that the engineered chimeric peptide is a potential broad-spectrum vaccine against visceral and cutaneous leishmaniasis.

Antigenicity and immunogenicity of SARS-CoV-2 surface glycoprotein fragment in CHO cells.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) glycoprotein that projects from the virus surface is highly immunogenic. It is considered to be the target of many neutralizing antibodies as well as a target in vaccine design efforts. Evaluation the immunogenicity of a recombinant fragment of the spike protein (rfsp) that is comprised of Receptor Binding Domain (RBD), S1/S2 cleavage site, and fusion peptide (FP) as immunogenic proteins of SARS-COV-2, in BALB/c mice and evaluation of the efficacy of epitopes rfsp as a multi-subunit chimeric vaccine. The present study made use of CHO-K1 (Chinese hamster ovary K1) cells to create a cell line for constant expression rfsp. The rfsp was purified with Ni-NTA chromatography and confirmed by Western blotting. The immunogenicity and neutralizing antibody efficacy of rfsp were evaluated in BALB/c mice. ELISA was employed to test rfsp via sera of COVID-19 convalescent patients infected with SARS-CoV-2 alpha and delta variants. Our results showed significant differences in antibody titers in immunized mice compared to the control groups and neutralizing antibodies were positive, sera from mice immunized are capable of bound SARS-CoV-2 virus, chimer peptide is capable bound antibodies patients infected with SARS-CoV-2 and patients infected with delta variant SARS-CoV-2. Overall, these results indicate that rfsp protein would be a novel potential antigen candidate for the development of a subunit SARS CoV-2 vaccine and rfsp has the potential to be a useful option for the development of the assays for serodiagnosis of SARS-CoV-2 infection.

Efficacy of Pgp3 vaccination for Chlamydia urogenital tract infection depends on its native conformation.

Urogenital tract infections with Chlamydia trachomatis have frequently been detected among patients diagnosed with sexually transmitted infections, and such infections lead to inflammatory complications. Currently, no licensed chlamydial vaccine is available in clinical practice. We previously reported that immunization with recombinant C. trachomatis plasmid-encoded virulence factor Pgp3 provided cross-serovar protection against C. muridarum genital tract infection. Because Pgp3 is a homotrimer and human antisera only recognize the trimeric form of Pgp3, we compared the effects of the native conformation of Pgp3 (trimer) and heat-denatured Pgp3 (monomer) to determine whether the native conformation is dispensable for the induction of protective immunity against chlamydial vaginal challenge. Both Pgp3 trimer and monomer immunization induced corresponding specific antibody production, but only trimer-induced antibody recognized endogenous Pgp3, and trimer-immunized mouse splenocytes showed the highest IFN-γ production upon restimulation with the chlamydial elementary body or native Pgp3 in vitro. Importantly, only Pgp3 trimer-immunized mice showed shortened lower genital tract chlamydial shedding and decreased upper genital tract pathology. Thus, Pgp3-induced protective immunity against Chlamydia urogenital tract infection is highly dependent on the native conformation, which will guide the design of Pgp3-based polypeptides and multi-subunit chlamydial vaccines.

Development and evaluation of a multi-epitope subunit vaccine against group B Streptococcus infection

ABSTRACT Streptococcus agalactiae (Group B Streptococcus, GBS) is a multi-host pathogen, even causing life-threatening infections in newborns. Vaccination with GBS crossed serotypes vaccine is one of the best options for long-term infection control. Here we built a comprehensive in silico epitope-prediction workflow pipeline to design a multivalent multiepitope-based subunit vaccine containing 11 epitopes against Streptococcus agalactiae (MVSA). All epitopes in MVSA came from the proteins which were antigenic-confirmed, virulent-associated, surface-exposed and conserved in ten GBS serotypes. The in-silico analysis showed MVSA had potential to evoke strong immune responses and enable worldwide population coverage. To validate MVSA protection efficacy against GBS infection, immune protection experiments were performed in a mouse model. Importantly, MVSA induced a high titre of antibodies, significant proliferation of mice splenocytes and elicited strong protection against lethal-dose challenge with a survival rate of 100% in mice after three vaccinations. Meanwhile, the polyclonal antibody against MVSA did not only inhibit for growth of GBS from six crucial serotypes in vitro, but also protect 100% naive mice from GBS lethal challenge. These active and passive immunity assay results suggested that MVSA could therefore be an efficacious multi-epitope vaccine against GBS infection.