Background: Herpes zoster (HZ), caused by the reactivation of varicella-zoster virus (VZV), primarily affects elderly populations worldwide. Although current recombinant HZ vaccines show strong immunogenicity, their high cost and potential side effects may limit their widespread use. Therefore, developing a cost-effective HZ vaccine with improved safety profiles would have significant clinical and public health implications. Methods: Building upon our previously optimized truncated gE (tgE350) from VZV, we developed the tgE350 + Fe nanoparticle vaccine using SpyTag/SpyCatcher covalent conjugation. The tgE350 protein (with a SpyTag tag) and the Fe protein (with a SpyCatcher tag) were expressed in HEK293F and E. coli BL21, respectively, enabling spontaneous nanoparticle assembly. Protein expression and nanoparticle formation were confirmed through SDS-PAGE and negative-stain electron microscopy. BALB/c mice were inoculated with either tgE350 + Fe or tgE350 combined with Al and CpG adjuvants. Immune responses were evaluated using ELISpot and flow cytometry for cellular immunity, along with ELISA, VZV microneutralization, and fluorescent antibody membrane antigen (FAMA) assays for antibody titers. Histopathological examination of major organs ensured vaccine safety. Results: Compared with the truncated vaccine tgE350, the nanoparticle vaccine tgE350 + Fe significantly enhanced VZV neutralizing antibodies and specific antibody responses in mice without causing significant changes in lymphocyte populations (no difference from the control group). Moreover, the tgE350 + Fe group had significantly more lymphocytes secreting IFN-γ, IL-2, and IL-4 than the tgE350 group. No apparent pathological damage was observed in the heart, liver, spleen, or lungs of mice in any experimental group. Conclusions: This experiment successfully developed the HZ nanoparticle vaccine tgE350 + Fe. It enhanced VZV-specific neutralizing antibodies, generated better cellular and humoral immune responses, and demonstrated good safety.

Introduction:In China, no standardized single-nucleotide polymorphism (SNP) scheme exists for varicella-zoster virus (VZV) genotyping. The 5-SNP scheme with two amplicon gene fragments lacks systematic validation. This study aimed to evaluate the accuracy and applicability of this genotyping method. Methods:A total of 280 complete genomes were genotyped using 5 SNPs extracted from ORF22 (four SNPs) and ORF38 (one SNP) fragments. The results were compared with those of the phylogenetic clustering method as a reference. Concordance with reference was used to estimate the accuracy of the SNP scheme. The evaluated SNP scheme was applied to a national VZV surveillance screen containing 549 clinical samples from 17 Chinese provincial-level administrative divisions (2017-2026). Results:A 97.9% concordance was observed between the 5-SNP scheme and phylogenetic clustering methods. The 2.1% discordance was mostly attributed to putative recombination and early circulation of strains from patients with herpes zoster. During the national VZV surveillance screening, 434 samples were amplified, sequenced, and genotyped using a 5-SNP scheme. Of the genotyped samples, 90.3% and 8.8% were identified as clades 2 and 5, respectively, using four ORF22 SNPs, and the remaining 0.9% as clade 4, using ORF38 SNP. All samples showed 100% intragenotypic SNP profile consistency. Conclusion:The unified 5-SNP scheme is accurate and practical for VZV surveillance in China; however, periodic evaluation is required.

IntroductionThe fusion glycoprotein (F protein) of human respiratory syncytial virus (HRSV) critically determines viral infectivity and host immune recognition. This study evaluated the effects of F protein genetic variations from the globally dominant HRSV genotypes ON1 and BA9 on viral replication dynamics and pathogenicity.MethodsUsing a reverse genetics system, two recombinant HRSV strains, rLong-BJ1903-AF and rLong-SY2103-BF, were generated by replacing the F gene in the isogenic Long-bacterial artificial chromosome (BAC) backbone with F genes derived from the clinical isolates BJ19-03 (ON1 genotype) and SY21-03 (BA9 genotype), respectively. Their phenotypes were assessed in cellular and animal models. ResultsIn vitro, both recombinant viruses showed attenuated replication kinetics compared with the parental Long-BAC strain, with peak viral titers delayed by at least 12 h post-infection. In vivo, both recombinant viruses caused less severe disease than Long-BAC. Among them, rLong-SY2103-BF displayed greater pathogenicity than rLong-BJ1903-AF, as shown by slower body weight recovery, higher lung viral loads, elevated proinflammatory cytokine levels, and marked pulmonary pathological alterations.DiscussionThese findings indicate that both recombinant viruses exhibit reduced replication capacity and less severe pathogenic phenotypes compared with the Long-BAC parental strain in this F-gene replacement system, providing a foundation for antiviral and immunological strategies targeting HRSV.

ETHNOPHARMACOLOGICAL RELEVANCE:Viral pneumonia (VP), particularly that induced by influenza A virus (IAV), remains a major global health challenge and a leading cause of acute respiratory infections. Ma Xing Shi Gan Decoction (MXSG), a classical traditional Chinese medicine formula recorded in the ancient text Treatise on Cold Damage, has been widely used for centuries in China to treat febrile respiratory diseases and pneumonia. However, the molecular basis underlying its therapeutic effects remains incompletely defined. This study provides experimental evidence linking MXSG to modulation of VP and host metabolic-autophagy pathways. AIM OF THE STUDY:To investigate whether MXSG is associated with reduced IAV nucleoprotein (NP) levels and concurrent modulation of key glycolytic enzymes, HMGB1 lactylation, and autophagosome accumulation. Buddleoside was evaluated as an exploratory constituent. METHODS:A mouse model of VP was established by intranasal inoculation with IAV, while A549 cells were infected with IAV for in vitro studies. In vivo effects on glycolysis and autophagy were evaluated by assessing lung inflammatory cytokines, viral NP levels, glycolytic enzymes, and autophagy markers. Oxamate and diphenyleneiodonium chloride (DPI) were used as tools to interfere with lactate production. UPLC-MS/MS and molecular docking were performed to identify potential pathway-relevant constituents. In vitro, validation included lactate supplementation, oxamate treatment, buddleoside exposure, and HMGB1 overexpression. RESULTS:In vivo, MXSG reduced IAV NP levels and alleviated lung inflammatory injury. MXSG treatment was associated with decreased expression of glycolysis-related enzymes, reduced lactate accumulation, lower HMGB1 lactylation levels, and reduced autophagosome accumulation. Under DPI-mediated lactate accumulation, the inhibitory effect of MXSG on autophagosome accumulation was partially attenuated. Docking analysis suggested potential interaction between buddleoside and LDHA. In vitro, MXSG modulated lactate levels, HMGB1 lactylation, and autophagy-related markers, accompanied by decreased NP levels. CONCLUSION:MXSG reducing NP levels is associated with modulation of expression levels of glycolytic enzymes and reduced lactate production, and was accompanied by decreases in HMGB1 lactylation with expression and HMGB1-associated autophagosome accumulation providing a potential metabolic basis for MXSG-associated regulation of autophagy-related processes. These findings suggest a possible association between metabolic-autophagy signaling and MXSG-associated reductions in viral NP levels, but remain correlative and require further validation.

Background: Respiratory syncytial virus (RSV) remains a major etiologic agent of acute lower respiratory tract infection (ALRTI). Currently licensed RSV vaccines are administered by intramuscular injection and induce limited immunity at the respiratory mucosal interface, underscoring the need for effective mucosal vaccination strategies. Methods: To enhance mucosal immune responses, we used prefusion F protein (Pre-F) as the antigen and performed intranasal immunization in BALB/c mice. Four mucosal adjuvants (CpG-ODN, CTA1-DD, IFN-α, and PEI) were systematically compared across different dose levels to evaluate their immunological and protective efficacy. Results: Both adjuvant type and dose helped shape the magnitude and quality of the immune response and the level of protection. CpG-ODN showed a dose-restricted immunopotentiating effect: an intermediate dose (10 µg) significantly increased neutralizing antibody titers and nasal mucosal IgA responses, improved post-challenge body weight recovery, and reduced lung viral load, whereas higher doses provided no additional benefit and were associated with aggravated lung pathology. PEI and IFN-α exhibited dose-dependency within a certain range, but increasing doses did not result in further improvements in immune responses or protection; an intermediate dose (10 µg) was sufficient to elicit robust systemic and mucosal immunity. CTA1-DD improved selected immune parameters at appropriate doses, yet its overall immunopotentiating effects remained modest. Direct comparative analysis using the representative doses selected from the three dose levels for each adjuvant indicated that 10 µg CpG-ODN or PEI provided superior immunogenicity and protection, whereas PEI induced a Th2-biased immune profile at both humoral and cellular levels. Conclusions: These findings highlight that favorable immunogenicity and protection are achieved within defined dose windows rather than at maximal doses. Among the adjuvants studied, low-to-intermediate doses of CpG-ODN, particularly 10 µg, show strong potential for intranasal mucosal immunization with recombinant RSV Pre-F protein. By systematically comparing dose–effect profiles across multiple mucosal adjuvants, this study offers comparative insights into adjuvant selection and dose selection for intranasal RSV vaccine development.

IntroductionHuman respiratory syncytial virus (HRSV) has a high disease burden in infants and elderly individuals. In this study, the adjuvants AlOH, AlOH+CpG and BFA03 were used to compare the protective effect of HRSV prefusion protein (Pre-F) in BALB/c mice.MethodsWe divided BALB/c mice into three experimental groups (Pre-F+AlOH+CpG, Pre-F+AlOH, and Pre-F+BFA03) and three adjuvant control groups (AlOH+CpG, AlOH, and BFA03). After two intramuscular immunizations, we measured serum neutralizing antibody titers and quantified the numbers of IFN-γ- and IL-4-secreting lymphocytes. After viral challenge, we monitored body weight changes, determined lung viral loads (Ct values), and scored lung pathological damage.ResultsThe mice in the experimental groups exhibited high titres of neutralizing antibodies and increased numbers of IFN-γ- and IL-4-secreting lymphocytes. The mice began to regain weight on the third day after challenge, but the mice in the adjuvant groups continued to lose weight. The mice immunized with Pre-F+BFA03 elicited the highest neutralizing antibody titre (1716), the lowest viral load in the lung, and milder pathological damage. The mice immunized with Pre-F+AlOH had the most severe lung pathological injury (score: 2.83) and the highest viral load in the lung (Ct value: 32.2). Compared with the BFA03 adjuvant, the AlOH adjuvant induced a Th2-biased humoral immune response in mice. The Pre-F+AlOH+CpG group had the least pathological damage in the lung (score 2.16), and the ability to induce neutralizing antibodies and cellular immune responses was comparable with that of the BFA03 adjuvant.DiscussionThese findings indicate that Pre-F protein combined with AlOH+CpG or BFA03 adjuvant provided similar protection in mice and both were superior to the AlOH adjuvant, providing a reference for adjuvant selection and formulation strategies in HRSV Pre-F protein vaccine development.

The concurrent circulation of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza virus, and respiratory syncytial virus (RSV) has caused severe coinfections, posing significant public health threats. Developing individual vaccines for these respiratory viruses is hindered by waning immunity and rapid viral mutations. A combination mRNA vaccine streamlines development and administration, reduces healthcare burdens, and offers convenient, compliance-enhancing protection against seasonal illnesses. Here, we present SIR, an 8-valent combination mRNA vaccine designed to simultaneously target SARS-CoV-2, influenza, and RSV. SIR was further optimized into SIRC by incorporating the TLR-9 agonist CpG oligodeoxynucleotides (CpG-ODN) 1018. SIRC induces robust binding and neutralizing antibodies against all three viruses, stimulates Th1-biased antigen-specific T-cell responses, and generates CD4+ and CD8+ memory T cells. In mice model, SIRC effectively protects against four influenza subtypes (H1N1, H3N2, B/V, and B/Y) and the RSV. Overall, SIRC demonstrates superior antibodies and T cell responses, and provides greater protection in vivo compared to both the individual and SIR vaccines, without causing immune interference. It also maintains a favorable safety and development potential. These findings establish the SIRC as a promising candidate for preventing coinfections involving SARS-CoV-2, influenza, and RSV.

Neutralizing antibody titres have been shown to correlate with immune protection against COVID-19 and can be used to estimate vaccine effectiveness. Numerous studies have explored the relationship between neutralizing antibodies and protection. However, there remains a lack of quantitative data directly assessing the minimum effective protective neutralizing antibody titre in in vivo. In this study, we utilized eight cohorts of participants with diverse immune backgrounds for evaluation of protective antibody response. To precisely assess the lower threshold of neutralizing antibody titres required for effective protection against SARS-CoV-2 infections, we employed plasma adoptive transfer from different cohorts into mice. This study demonstrated that neutralizing titres in the plasma of recipient mice correlated well with those in human donors, and a positive linear correlation was observed between the human and mouse recipients of transferred plasma neutralizing titre. A pseudotyped virus neutralizing titres greater than 7 was identified as the minimum threshold necessary to reduce viral titres in infected mice, establishing a crucial baseline for effective protection. Furthermore, despite the variability in immune backgrounds, these diverse cohorts' plasma exhibited a similar neutralizing antibody threshold necessary for protection. This finding has significant implications for vaccine design and the assessment of immune competence.

The authors would like to make the following corrections to this published paper [...]

In recent years, coxsackievirus A6 (CVA6) has surpassed enterovirus A71 to become the main pathogen causing severe Hand, Foot, and Mouth disease (HFMD) in China with a substantial disease burden. However, there is currently no commercial CVA6 vaccine. The D3a genotype of CVA6 is the predominant genotype in China. In this study, virus-like particles (VLPs) and mRNA vaccines based on the CVA6 sub-genotype D3a were successfully developed. The immunogenicity and protective effects of the VLP of CVA6 combined with Al(OH)3 and CpG adjuvant indicated that VLP-induced neutralizing antibodies against three CVA6 sub-genotype (D2, D3a, and D3b) strains in Institute of Cancer Research (ICR) mice, and the combination of the two adjuvants enhanced cellular immunity. Passive immunization with serum from mice immunized with VLPs protected suckling mice against CVA6 lethal challenge in both antiserum transfer and maternal immunization experiments. The immunogenicity and protective effects of the mRNA vaccine of CVA6 indicate that it induces robust T-cell immunity. T-cell immunity was found to cross-protect against coxsackievirus A10 infection in mice. This is the first trial of a CVA6 mRNA vaccine worldwide and the first comparison of the immunogenicity and protective effects of VLP and mRNA vaccines based on D3a CVA6. The study provides a theoretical basis for the development of enteroviruses vaccines and the formulation of immunization strategies.

Purpose To evaluate the immunogenic potential of three different nanoparticle (NP) platforms for respiratory syncytial virus (RSV) prefusion (pre-F) protein vaccines. Methods Three NP platforms-24-mer ferritin (Fe), 60-mer lumazine synthase (LuS), and 120-subunit I53-50-were engineered to display RSV pre-F trimers (DS2) via SpyTag-SpyCatcher (ST-SC) conjugation (DS2-Fe, DS2-LuS) or direct genetic fusion (DS2-I53-50). The assembled particles were characterized using size-exclusion chromatography (SEC), SDS-PAGE, electron microscopy (EM), and dynamic light scattering (DLS). Antigenicity was evaluated using enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR) with prefusion-specific neutralizing antibodies. Immunogenicity and protective efficacy were evaluated in BALB/c mice following a prime-boost immunization, with analyses of humoral and cellular immune responses as well as post-challenge protection. Results All three NP platforms successfully displayed the DS2 antigen while preserving its prefusion conformation. Notably, DS2-I53-50 demonstrated superior assembly quality and particle homogeneity relative to DS2-Fe and DS2-LuS. Compared to soluble DS2, all three DS2-NPs exhibited enhanced binding affinity (7- to 12-fold increase) to prefusion-specific antibodies (D25, AM14). In vivo, all DS2-NPs elicited higher levels of RSV-specific neutralizing antibodies and induced a more balanced Th1/Th2 immune response, with DS2-I53-50 generating significantly greater neutralizing antibody titers (1.7- to 2.4-fold increase) against both prototype RSV strains (LONG, 18537) and circulating genotypes (ON1, BA9). Immune cell profiling further revealed that all three DS2-NPs enhanced germinal center formation, facilitated follicular dendritic cell recruitment, and expanded memory T cell populations. Following RSV challenge, all DS2-NPs vaccines conferred significant protection, evidenced by accelerated weight recovery, reduced lung viral loads, and mitigated pulmonary pathology. Among them, DS2-I53-50 provided the most robust protection, achieving a 3.7-log reduction in viral titers and minimal lung pathology. Conclusion NP platforms significantly enhanced the immunogenicity of RSV DS2 antigens, with DS2-I53-50 eliciting the strongest immune responses and protective efficacy. These findings underscore the potential of rationally designed NP-based vaccines for RSV.

Human respiratory syncytial virus (HRSV) poses a significant disease burden on global health. To date, two vaccines that primarily induce humoral immunity to prevent HRSV infection have been approved, whereas vaccines that primarily induce T-cell immunity have not yet been well-represented. To address this gap, 25 predicted T-cell epitope peptides derived from the HRSV fusion protein with high human leukocyte antigen (HLA) binding potential were synthesized, and their ability to be recognized by PBMC from previously infected HRSV cases was assessed using an ELISpot assay. Finally, nine T-cell epitope peptides were selected, each of which was recognized by at least 20% of different donors’ PBMC as potential vaccine candidates to prevent HRSV infection. The protective efficacy of F-9PV, a combination of nine peptides along with CpG-ODN and aluminum phosphate (Al) adjuvants, was validated in both HLA-humanized mice (DR1-TCR transgenic mice, Tg mice) and wild-type (WT) mice. The results show that F-9PV significantly enhanced protection against viral challenge as evidenced by reductions in viral load and pathological lesions in mice lungs. In addition, F-9PV elicits robust Th1-biased response, thereby mitigating the potential safety risk of Th2-induced respiratory disease during HRSV infection. Compared to WT mice, the F-9PV mice exhibited superior protection and immunogenicity in Tg mice, underscoring the specificity for human HLA. Overall, our results demonstrate that T-cell epitope peptides provide protection against HRSV infection in animal models even in the absence of neutralizing antibodies, indicating the feasibility of developing an HRSV T-cell epitope peptide-based vaccine.

人呼吸道合胞病毒(Human respiratory syncytial virus,HRSV)是引起全球婴幼儿急性下呼吸道感染的重要病原体.本研究旨在建立HRSV原型株Long毒株的反向遗传操作系统.通过利用细菌人工染色体(Bacterial artificial chromosome,BAC)为载体骨架构建HRSV的Long毒株全长质粒(Long-BAC),以pcDNA3.1为载体构建分别表达 HRSVN、P、M2-1 和 L 蛋白(pcDNA3.1-N,pcDNA3.1-P,pcDNA3.1-M2-1 和 pcDNA3.1-L)的四个辅助质粒,将五个质粒共转染于表达T7RNA聚合酶的BSRT7/9细胞系进行病毒拯救.采用间接免疫荧光(Immune fluorescence assay,IFA)和测序方法对拯救的病毒进行鉴定,并评价其在细胞和动物水平的生物学特征.本研究首次成功构建以BAC载体为骨架的HRSV原型株Long感染性克隆,拯救的病毒在不同代数测序结果显示无突变,能够稳定传代,病毒滴度为3×106 PFU/mL;与实验室保存野生型Long株(Long-WT)相比具有相似的细胞生长动力学特性,在12～36h病毒呈指数增长,在36h复制达到最高峰;在感染小鼠后能引起明显的肺部病理变化以及炎性相关细胞因子升高.本研究建立了高效、稳定的HRSV反向遗传学系统,为我国HRSV致病机制研究、疫苗和抗体药物研发提供技术平台.

本研究针对新冠病毒Omicron BA.4/5的受体结合域(Receptor binding domain,RBD)构建一个连接Fc受体的二聚体蛋白BA.4/5-RBD-Fc(BRF)并评价其免疫原性.结果显示,两种佐剂组小鼠的二免后血清均可产生高滴度的IgG抗体且显著高于一免后血清(P＜0.0001),BRF+A1OH/CpG组小鼠血清产生较为平衡的IgG1和IgG2a抗体应答,而BRF+BFA03组小鼠血清能产生更多的偏向Th2应答的IgG1抗体,且IgG1/IgG2a比值显著高于BRF+AlOH/CpG组(P＜0.0001).BRF+AlOH/CpG组产生的 Thl应答的细胞因子干扰素 γ(Interferon-γ,IFN-γ)高于BRF+BFA03组(P＜0.01),而产生的Th2应答细胞因子白介素4(Interleukin-4,IL-4)IL-4显著低于BRF+BFA03组(P＜0.01).BRF蛋白结合不同佐剂两次免疫小鼠后的血清均可以有效中和目前Omicron主要的流行亚型BA.2与BA.4活病毒,产生高达19 334598和17 224096的中和抗体滴度.因此,BRF蛋白诱导小鼠血清产生的中和抗体针对Omicron系列变异株具有一定的广谱性,AlOH/CpG佐剂可以使BRF蛋白产生偏向Th1的免疫应答反应,而BFA03佐剂产生明显偏向Th2的免疫应答反应.本研究为新冠Omicron变异株亚单位疫苗的研发提供有效的科学依据.

Objective:This study comprehensively analyzed the genomic characterizations of human parainfluenza virus type 3 (HPIV3) strains circulating in six provinces and cities of China (Beijing, Henan, Jilin, Anhui, Gansu, and Shandong) during the period of 2019-2020. The aim was to elucidate the intricate genetic variations and molecular evolutionary trends within the HPIV3 genome.Methods:Based on genotypic differentiation, genetic divergence, and spatial and temporal distribution, 12 representative HPIV3 strains (including 7 of C3a subtype, 2 of C3b subtype and 3 of C3f subtype) were selected from the aforementioned provinces, and the complete genome sequence was successfully obtained by overlapping amplification of fragments using nested RT-PCR. Subsequently, a complete genome database of global representative HPIV3 strains was constructed and analyzed using bioinformatics tools.Results:The length of complete genome of the 12 HPIV3 strains in the present study varied between 15 227 bp and 15 370 bp, the G+ C content ranged from 35.1% to 35.3% and the nucleotide identity intermediated from 97.6% to 99.6%. Compared with the prototype strain (GenBank accession number: NC_001796.2), the nucleotide identity of 12 HPIV3 strains ranged from 94.2% to 94.5%. Analysis of the complete genome of HPIV3 available in China and globally showed that the genomic variation of HPIV3 was mainly shaped by substitution mutations, and no base deletions or gene recombination were observed.Only a six-base insertion (ATTAAA) was found in the F gene’s 3′UTR region of a representative strain originating from Jilin province (CHN/Jilin036/2019/C3b) in this study, and its potential pathogenic significance needs to be further investigated. Amino acid analysis of the encoded proteins revealed that the C3a lineage of HPIV3, widely prevalent both in China and worldwide, exhibits lineage-specific mutation sites in the N, P and L proteins. Furthermore, within the Chinese prevalent C3a strains, a distinctive mutation site (N216S) in L protein was also identified. Notably, specific variant sites have not been found in Chinese C3b and C3f branch strains. Based on the complete genome, the comprehensive evolutionary analysis showed that the time to the most recent common ancestor (tMRCA) of global HPIV3 strains was estimated to 1927 (95% HPD: 1901-1945), with an average molecular evolutionary rate of 5.29 × 10 -4 substitutions/site/year, while the average molecular evolutionary rate of HPIV3 strains in China is 5.24 × 10 -4 substitutions/site/year. In addition, each gene of HPIV3 was subjected to negative selection pressure, with the P, HN and F genes showing the most significant nucleotide variation and higher rates of molecular evolution than the other genes. Conclusions:This study reveals that the complete genome of HPIV3 strains circulating in six provinces and cities of China tend to evolve conservatively. Moreover, substitution emerge as the main driving force for molecular evolution of HPIV3.

Continuous emergence of adaptive mutations of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to challenge the use and development of existing vaccines and drugs. The value of variant-based vaccines that are capable of inducing a higher and broader protection immune response against SARS-CoV-2 variants is currently being evaluated.

The development of a vaccine against human respiratory syncytial virus (HRSV) has been hampered by enhanced respiratory disease due to the Th2-biased immune response. In the present study, MA103 and aluminum phosphate (Adju-Phos) adjuvants were used to verify the immunogenicity of the recombinant fusion (RBF) protein (F protein expressed by Escherichia coli). Both adjuvants significantly increased the neutralizing antibody titer and number of interferon gamma (IFN-γ)-secreting CD4+ T cells in mice. Based on the immunoglobulin G1 (IgG1)/IgG2a and IFN-γ/interleukin 4-secreting CD4+ T cell ratio, however, MA103 significantly enhanced the Th1-biased immune response. The pathological damage to the lung in the RBF/MA103 group was less than what was seen in the RBF/Adju-Phos group. Additionally, the number of HRSV copies in the lungs of the RBF/MA103 group decreased by approximately 3 × log10. These results suggested that MA103 provides better protection against HRSV in mice.

人呼吸道合胞病毒(Human respiratory syncytial virus,hRSV)是全球婴幼儿和老年人严重呼吸道疾病的主要原因.hRSV感染主要局限于呼吸道,当鼻黏膜中特异性IgA抗体滴度较低时容易引起hRSV反复感染,理想的hRSV疫苗应诱导全身免疫应答,尤其是黏膜免疫.本研究应用CHO细胞表达融合蛋白F-Fc(含有hRSVF蛋白和人IgG1抗体的Fc片段),F-Fc蛋白结合CpG佐剂两次免疫小鼠,比较滴鼻免疫(Intranasal,in)和肌肉注射(Intramuscular,im)免疫安全性和有效性的差异.与佐剂对照组(CpG)相比,四种免疫方式(CpG+F-Fc/in+im,CpG+F-Fc/im+in,CpG+F-Fc/im+im和CpG+F-Fc/in+in)均能诱导高滴度中和抗体,高水平及Th1偏向的细胞免疫应答,减少肺脏病毒的滴度,但是两次滴鼻免疫组小鼠效果是最好的.同时,两次滴鼻免疫组小鼠诱导的IgA抗体最多,小鼠体重恢复速度最快,并且可以显著降低肺脏病理损伤.综上所述,在以上四种免疫方案中,两次滴鼻免疫诱导产生的免疫效果最优.

Objective:To evaluate the effect of synthetic CpG oligodeoxynucleotide (CpG-ODN) as adjuvant on immune response induced by inactivated human adenovirus (HAdV)-55 antigen in BALB/c mice.Methods:HAdV-55 virus QS prototype strain was purified by plaque to construct a seed bank of vaccine candidate strain. The amplified product of vaccine candidate strain was inactivated by 0.05%β-propiolactone, and purified to prepare perfect virus particle antigen. The purified HAdV-55 antigen was mixed with the same volume CPG-ODN and aluminum hydroxide adjuvant in low-dose group (0.2 mg/ml) and high-dose group (1 mg/ml), respectively, and inoculated BALB/c mice after emulsification. Meanwhile, the control group was set with PBS, and the immunization was enhanced once every 21 days. Respectively on primary immune 21 and 35 days after collecting venous blood in mice and separation of serum, serum was collected at the end of the time of separating spleen lymphocytes in mice. The levels of HAdV-55 specific IgG antibody and neutralization antibody in serum of immunized mice were observed by ELISA and micro-neutralization test, and the levels of lymphocytes secreting IL-4 and IFN-γ cytokines were detected by ELISpot.Results:No matter with or without adjuvant, along with the increase of the number of immunization and vaccination dose of inactivated HAdV-55 antigen induced BALB/c mice virus specific IgG antibody was also significantly increased. However, neutralizing antibody can reach detectable level only after enhanced immunity, and the geometric mean titer (GMT) of neutralizing antibody is between 1: 11 and 1: 23. Different adjuvants have significant effects on the immune response of mice. Low dose antigen combined with CPG-ODN and aluminum hydroxide mixed adjuvant can induce higher humoral and cellular immune responses in mice, and the levels of specific IgG antibody and neutralizing antibody are 2.2 and 1.8 times higher than those in the aluminum hydroxide adjuvant group, respectively. The number of lymphocytes secreting IFN-γ was 2.3 times that of the group immunized with aluminum hydroxide adjuvant.Conclusions:The novel CPG-ODN adjuvant significantly increased the immunogenicity of the inactivated HAdV-55 whole virus antigen in BALB/c mice and directed the cellular immune response toward Th1 type.

Human respiratory syncytial virus (HRSV) is a leading cause of lower respiratory tract infections in elderly individuals and young children/infants and can cause bronchiolitis and even death. There is no licensed HRSV vaccine. An ideal vaccine should induce high titers of neutralizing antibodies and a Th1-biased immune response. In this study, we used EXPI293 cells to express the fusion (F) protein with a prefusion conformation (PrF) and compared the safety and efficacy of intranasal immunization with PrF in combination with two mucosal adjuvants (CpG ODN and liposomes) in mice. After two intranasal administrations, mice in the PrF + CpG group produced high titers of neutralizing antibodies (4961) and a Th1-biased immune response compared with the PrF + Lipo group. The lung viral load of mice in the PrF + CpG group was significantly reduced (3.5 log) compared with that in the adjuvant control group, and the survival rate was 100 %, while the survival rate of mice in the PrF + Lipo group was only 67 %. At the same time, this immunization strategy reduced the pathological damage to the lungs in mice. In conclusion, the combination of PrF and CpG adjuvant is immunogenic, elicits a Th1 type immune response, and completely protects mice from a lethal HRSV challenge. It is worthy of further evaluation as an HRSV vaccine in clinical trials. Clinical trial registration. This study was not related to human participation or experimentation.