Fully human monoclonal antibodies against Ebola virus possess complete protection in a hamster model.

Ebola disease is a lethal viral hemorrhagic fever caused by ebolaviruses within the Filoviridae family with mortality rates of up to 90%. Monoclonal antibody (mAb) based therapies have shown great potential for the treatment of EVD. However, the potential emerging ebolavirus isolates and the negative effect of decoy protein on the therapeutic efficacy of antibodies highlight the necessity of developing novel antibodies to counter the threat of Ebola. Here, 11 fully human mAbs were isolated from transgenic mice immunized with GP protein and recombinant vesicular stomatitis virus-bearing GP (rVSV-EBOV GP). These mAbs were divided into five groups according to their germline genes and exhibited differential binding activities and neutralization capabilities. In particular, mAbs 8G6, 2A4, and 5H4 were cross-reactive and bound at least three ebolavirus glycoproteins. mAb 4C1 not only exhibited neutralizing activity but no cross-reaction with sGP. mAb 7D8 exhibited the strongest neutralizing capacity. Further analysis on the critical residues for the bindings of 4C1 and 8G6 to GPs was conducted using antibodies complementarity-determining regions (CDRs) alanine scanning. It has been shown that light chain CDR3 played a crucial role in binding and neutralization and that any mutation in CDRs could not improve the binding of 4C1 to sGP. Importantly, mAbs 7D8, 8G6, and 4C1 provided complete protections against EBOV infection in a hamster lethal challenge model when administered 12 h post-infection. These results support mAbs 7D8, 8G6, and 4C1 as potent antibody candidates for further investigations and pave the way for further developments of therapies and vaccines.

A Bacterium-like Particle Vaccine Displaying Envelope Proteins of Canine Distemper Virus Can Induce Immune Responses in Mice and Dogs.

Canine distemper virus (CDV) can cause fatal infections in giant pandas. Vaccination is crucial to prevent CDV infection in giant pandas. In this study, two bacterium-like particle vaccines F3-GEM and H4-GEM displaying the trimeric F protein or tetrameric H protein of CDV were constructed based on the Gram-positive enhanced-matrix protein anchor (GEM-PA) surface display system. Electron microscopy and Western blot results revealed that the F or H protein was successfully anchored on the surface of GEM particles. Furthermore, one more bacterium-like particle vaccine F3 and H4-GEM was also designed, a mixture consisting of F3-GEM and H4-GEM at a ratio of 1:1. To evaluate the effect of the three vaccines, mice were immunized with F3-GEM, H4-GEM or F3 and H4-GEM. It was found that the level of IgG-specific antibodies and neutralizing antibodies in the F3 and H4-GEM group was higher than the other two groups. Additionally, F3 and H4-GEM also increased the secretion of Th1-related and Th2-related cytokines. Moreover, F3 and H4-GEM induce IgG and neutralizing antibodies' response in dogs. Conclusions: In summary, F3 and H4-GEM can provoke better immune responses to CDV in mice and dogs. The bacterium-like particle vaccine F3 and H4-GEM might be a potential vaccine candidate for giant pandas against CDV infection.

Viral vectored vaccines: design, development, preventive and therapeutic applications in human diseases.

Human diseases, particularly infectious diseases and cancers, pose unprecedented challenges to public health security and the global economy. The development and distribution of novel prophylactic and therapeutic vaccines are the prioritized countermeasures of human disease. Among all vaccine platforms, viral vector vaccines offer distinguished advantages and represent prominent choices for pathogens that have hampered control efforts based on conventional vaccine approaches. Currently, viral vector vaccines remain one of the best strategies for induction of robust humoral and cellular immunity against human diseases. Numerous viruses of different families and origins, including vesicular stomatitis virus, rabies virus, parainfluenza virus, measles virus, Newcastle disease virus, influenza virus, adenovirus and poxvirus, are deemed to be prominent viral vectors that differ in structural characteristics, design strategy, antigen presentation capability, immunogenicity and protective efficacy. This review summarized the overall profile of the design strategies, progress in advance and steps taken to address barriers to the deployment of these viral vector vaccines, simultaneously highlighting their potential for mucosal delivery, therapeutic application in cancer as well as other key aspects concerning the rational application of these viral vector vaccines. Appropriate and accurate technological advances in viral vector vaccines would consolidate their position as a leading approach to accelerate breakthroughs in novel vaccines and facilitate a rapid response to public health emergencies.

Bif-1c Attenuates Viral Proliferation by Regulating Autophagic Flux Blockade Induced by the Rabies Virus CVS-11 Strain in N2a Cells.

Bax-interacting factor-1 (Bif-1) is a multifunctional protein involved in apoptosis, autophagy, and mitochondrial morphology. However, the associations between Bif-1 and viruses are poorly understood. As discrete Bif-1 isoforms are selectively expressed and exert corresponding effects, we evaluated the effects of neuron-specific/ubiquitous Bif-1 isoforms on rabies virus (RABV) proliferation. First, infection with the RABV CVS-11 strain significantly altered Bif-1 expression in mouse neuroblastoma (N2a) cells, and Bif-1 knockdown in turn promoted RABV replication. Overexpression of neuron-specific Bif-1 isoforms (Bif-1b/c/e) suppressed RABV replication. Moreover, our study showed that Bif-1c colocalized with LC3 and partially alleviated the incomplete autophagic flux induced by RABV. Taken together, our data reveal that neuron-specific Bif-1 isoforms impair the RABV replication process by abolishing autophagosome accumulation and blocking autophagic flux induced by the RABV CVS-11 strain in N2a cells. IMPORTANCE Autophagy can be triggered by viral infection and replication. Autophagosomes are generated and affect RABV replication, which differs by viral strain and infected cell type. Bax-interacting factor-1 (Bif-1) mainly has a proapoptotic function but is also involved in autophagosome formation. However, the association between Bif-1-involved autophagy and RABV infection remains unclear. In this study, our data reveal that a neuron-specific Bif-1 isoform, Bif-1c, impaired viral replication by unchoking autophagosome accumulation induced by RABV in N2a cells to a certain extent. Our study reveals for the first time that Bif-1 is involved in modulating autophagic flux and plays a crucial role in RABV replication, establishing Bif-1 as a potential therapeutic target for rabies.

Taurolidine improved protection against highly pathogenetic avian influenza H5N1 virus lethal-infection in mouse model by regulating the NF-κB signaling pathway.

Taurolidine (TRD), a derivative of taurine, has anti-bacterial and anti-tumor effects by chemically reacting with cell-walls, endotoxins and exotoxins to inhibit the adhesion of microorganisms. However, its application in antiviral therapy is seldom reported. Here, we reported that TRD significantly inhibited the replication of influenza virus H5N1 in MDCK cells with the half-maximal inhibitory concentration (EC50) of 34.45 â€‹µg/mL. Furthermore, the drug inhibited the amplification of the cytokine storm effect and improved the survival rate of mice lethal challenged with H5N1 (protection rate was 86%). Moreover, TRD attenuated virus-induced lung damage and reduced virus titers in mice lungs. Administration of TRD reduced the number of neutrophils and increased the number of lymphocytes in the blood of H5N1 virus-infected mice. Importantly, the drug regulated the NF-κB signaling pathway by inhibiting the separation of NF-κB and IκBa, thereby reducing the expression of inflammatory factors. In conclusion, our findings suggested that TRD could act as a potential anti-influenza drug candidate in further clinical studies.

A bacterium-like particle vaccine displaying Zika virus prM-E induces systemic immune responses in mice.

The emergence of Zika virus (ZIKV) infection, which is unexpectedly associated with congenital defects, has prompted the development of safe and effective vaccines. The Gram-positive enhancer matrix-protein anchor (GEM-PA) display system has emerged as a versatile and highly effective platform for delivering target proteins in vaccines. In this study, we developed a bacterium-like particle vaccine, ZI-△-PA-GEM, based on the GEM-PA system. The fusion protein ZI-△-PA, which contains the prM-E-△TM protein of ZIKV (with a stem-transmembrane region deletion) and the protein anchor PA3, was expressed. The fusion protein was successfully displayed on the GEM surface to form ZI-△-PA-GEM. Moreover, the intramuscular immunization of BALB/c mice with ZI-△-PA-GEM combined with ISA 201 VG and poly(I:C) adjuvants induced durable ZIKV-specific IgG and protective neutralizing antibody responses. Potent B-cell/DC activation was also stimulated early after immunization. Notable, splenocyte proliferation, the secretion of multiple cytokines, T/B-cell activation and central memory T-cell responses were elicited. These data indicate that ZI-△-PA-GEM is a promising bacterium-like particle vaccine candidate for ZIKV.

Molecular Characteristics and Potent Immunomodulatory Activity of Fasciola hepatica Cystatin.

Cystatin, a cysteine protease inhibitor found in many parasites, plays important roles in immune evasion. This study analyzed the molecular characteristics of a cystatin from Fasciola hepatica (FhCystatin) and expressed recombinant FhCystatin (rFhcystatin) to investigate the immune modulatory effects on lipopolysaccharide-induced proliferation, migration, cytokine secretion, nitric oxide (NO) production, and apoptosis in mouse macrophages. The FhCystatin gene encoded 116 amino acids and contained a conserved cystatin-like domain. rFhCystatin significantly inhibited the activity of cathepsin B. rFhCystatin bound to the surface of mouse RAW264.7 cells, significantly inhibited cell proliferation and promoted apoptosis. Moreover, rFhCystatin inhibited the expression of cellular nitric oxide, interleukin-6, and tumor necrosis factor-α, and promoted the expression of transforming growth factor-ß and interleukin-10. These results showed that FhCystatin played an important role in regulating the activity of mouse macrophages. Our findings provide new insights into mechanisms underlying the immune evasion and contribute to the exploration of potential targets for the development of new drug to control F. hepatica infection.

Recombinant adeno-associated virus serotype 9 AAV-RABVG expressing a Rabies Virus G protein confers long-lasting immune responses in mice and non-human primates.

Three or four intramuscular doses of the inactivated human rabies virus vaccines are needed for pre- or post-exposure prophylaxis in humans. This procedure has made a great contribution to prevent human rabies deaths, which bring huge economic burdens in developing countries. Herein, a recombinant adeno-associated virus serotype 9, AAV9-RABVG, harbouring a RABV G gene, was generated to serve as a single dose rabies vaccine candidate. The RABV G protein was stably expressed in the 293T cells infected with AAV9-RABVG. A single dose of 2 × 1011 v.p. of AAV9-RABVG induced robust and long-term positive seroconversions in BALB/c mice with a 100% survival from a lethal RABV challenge. In Cynomolgus Macaques vaccinated with a single dose of 1 × 1013 v.p. of AAV9-RABVG, the titres of rabies VNAs increased remarkably from 2 weeks after immunity, and maintained over 31.525 IU/ml at 52 weeks. More DCs were activated significantly for efficient antigen presentations of RABV G protein, and more B cells were activated to be responsible for antibody responses. Significantly more RABV G specific IFN-γ-secreting CD4+ and CD8+ T cells, and IL-4-secreting CD4+ T cells were activated, and significantly higher levels of IL-2, IFN-γ, IL-4, and IL-10 were secreted to aid immune responses. Overall, the AAV9-RABVG was a single dose rabies vaccine candidate with great promising by inducing robust, long-term humoral responses and both Th1 and Th2 cell-mediated immune responses in mice and non-human primates.

Ebola virus VP35 hijacks the PKA-CREB1 pathway for replication and pathogenesis by AKIP1 association.

Ebola virus (EBOV), one of the deadliest viruses, is the cause of fatal Ebola virus disease (EVD). The underlying mechanism of viral replication and EBOV-related hemorrhage is not fully understood. Here, we show that EBOV VP35, a cofactor of viral RNA-dependent RNA polymerase, binds human A kinase interacting protein (AKIP1), which consequently activates protein kinase A (PKA) and the PKA-downstream transcription factor CREB1. During EBOV infection, CREB1 is recruited into EBOV ribonucleoprotein complexes in viral inclusion bodies (VIBs) and employed for viral replication. AKIP1 depletion or PKA-CREB1 inhibition dramatically impairs EBOV replication. Meanwhile, the transcription of several coagulation-related genes, including THBD and SERPINB2, is substantially upregulated by VP35-dependent CREB1 activation, which may contribute to EBOV-related hemorrhage. The finding that EBOV VP35 hijacks the host PKA-CREB1 signal axis for viral replication and pathogenesis provides novel potential therapeutic approaches against EVD.

Molecular characterization of a novel GSTO2 of Fasciola hepatica and its roles in modulating murine macrophages.

Fascioliasis is an important zoonotic helminthic disease caused by Fasciola hepatica and poses a serious threat to global public health. To evade the immune response of its host (humans or animals), F. hepatica secretes various antioxidant enzymes such as glutathione transferase (GST) to facilitate its invasion, migration and parasitism in vivo. To investigate the biological functions of a novel omega-class GST (GSTO), the molecular features of GSTO2 of F. hepatica were analyzed by online software, and the biochemical properties in vitro of recombinant GSTO2 (rGSTO2) were dissected. Then, the regulatory roles of rGSTO2 protein in murine macrophages in vitro were further explored. The results revealed that the GSTO2 gene encodes 254 amino acids, which harbor the characteristic N-terminal domain (ßαßαßßα) and C-terminal domain (α-helical) of the cytoplasmic GST superfamily. GSTO2 was mainly expressed in F. hepatica vitelline follicles, intestinal tract, excretory pores and vitelline cells, with thioltransferase and dehydroascorbate reductase activities. Moreover, rGSTO2 protein could be taken up by murine macrophages and significantly inhibit the viability of macrophages. In addition, rGSTO2 protein could significantly promote apoptosis and modulate the expression of cytokines in macrophages. These findings suggested that F. hepatica GSTO2 plays an important role in modulating the physiological functions of macrophages, whereby this protein might be involved in immunomodulatory and anti-inflammatory roles during infection. This study provided new insights into the immune-evasion mechanism of F. hepatica and may contribute to the development of a potential anti-inflammatory agent.

Title: Caractérisation moléculaire d'une nouvelle GSTO2 de Fasciola hepatica et ses rôles dans la modulation des macrophages murins. Abstract: La fasciolase est une importante maladie helminthique zoonotique causée par Fasciola hepatica, qui constitue une menace sérieuse pour la santé publique mondiale. Pour échapper à la réponse immunitaire de son hôte (humain ou animal), F. hepatica sécrète diverses enzymes antioxydantes telles que la glutathion transférase (GST) pour faciliter son invasion, sa migration et son parasitisme in vivo. Pour étudier les fonctions biologiques d'une nouvelle GST de classe oméga (GSTO), les caractéristiques moléculaires de la GSTO2 de F. hepatica ont été analysées par un logiciel en ligne et les propriétés biochimiques in vitro de sa protéine recombinante (rGSTO2) ont été disséquées. Ensuite, les rôles régulateurs de la protéine rGSTO2 sur les macrophages murins in vitro ont été explorés plus avant. Les résultats ont révélé que le gène GSTO2 code pour 254 acides aminés, qui abritent le domaine N-terminal caractéristique (ßαßαßßα) et le domaine C-terminal (α-hélicoïdal) de la superfamille GST cytoplasmique. Chez F. hepatica, GSTO2 était principalement exprimée dans les follicules vitellins, le tractus intestinal, les pores excréteurs et les cellules vitellines, avec des activités de thioltransférase et de déhydroascorbate réductase. De plus, la protéine rGSTO2 a pu être absorbée par les macrophages murins et inhiber de manière significative la viabilité des macrophages. Enfin, la protéine rGSTO2 a pu favoriser de manière significative l'apoptose et moduler l'expression des cytokines dans les macrophages. Ces résultats suggèrent que la GSTO2 de F. hepatica joue un rôle important dans la modulation des fonctions physiologiques des macrophages, cette protéine pouvant être impliquée dans des rôles immunomodulateurs et anti-inflammatoires au cours de l'infection. Cette étude a fourni de nouvelles informations sur le mécanisme d'évasion immunitaire de F. hepatica et pourrait contribuer au développement d'un agent anti-inflammatoire potentiel.

Inactivated Rabies Virus Vectored MERS-Coronavirus Vaccine Induces Protective Immunity in Mice, Camels, and Alpacas.

Middle East respiratory syndrome coronavirus (MERS-CoV) is an emergent coronavirus that has caused frequent zoonotic events through camel-to-human spillover. An effective camelid vaccination strategy is probably the best way to reduce human exposure risk. Here, we constructed and evaluated an inactivated rabies virus-vectored MERS-CoV vaccine in mice, camels, and alpacas. Potent antigen-specific antibody and CD8+ T-cell responses were generated in mice; moreover, the vaccination reduced viral replication and accelerated virus clearance in MERS-CoV-infected mice. Besides, protective antibody responses against both MERS-CoV and rabies virus were induced in camels and alpacas. Satisfyingly, the immune sera showed broad cross-neutralizing activity against the three main MERS-CoV clades. For further characterization of the antibody response induced in camelids, MERS-CoV-specific variable domains of heavy-chain-only antibody (VHHs) were isolated from immunized alpacas and showed potent prophylactic and therapeutic efficacies in the Ad5-hDPP4-transduced mouse model. These results highlight the inactivated rabies virus-vectored MERS-CoV vaccine as a promising camelid candidate vaccine.

Whole-genome resequencing to investigate the determinants of the multi-lumbar vertebrae trait in sheep.

Multi-lumbar vertebrae trait is a beneficial mutation that can significantly improve livestock meat production. However, the genetic basis of the multi-lumbar vertebrae in sheep is still unclear. Here, we analysed the number of lumbar vertebrae of Duolang sheep and found three different traits of lumbar vertebrae number. Compared with the normal sheep, the length and weight of animal carcass from the multi-lumbar vertebrae sheep increased by 2.21 cm and 0.78 kg, respectively. We performed high-throughput genome resequencing on multi-lumbar vertebrae (n = 18) and normal (n = 11) Duolang sheep and obtained a total of more than 528.87 GB data. We found that the most significantly selective region were located in the 49.68-49.74 MB of chromosome 4 by selective-sweep analysis. We annotated this region and found that it contains SFRP4 which is known to regulate bone development. We further used the PCR-SSCP technology to detect the single nucleotide polymorphism (SNP) of the putative candidate SFRP4 and found that the two SNPs (rs600370085:C > T and rs415133338: A > G) of this gene were significantly associated with the multi-lumbar vertebrae of Duolang sheep. Our study indicates that the SFRP4 may be a potential major gene that affects the number of lumbar vertebrae in Duolang sheep, and has the potential to be utilized for sheep breeding in the future.

A recombinant rabies virus expressing Echinococcus granulosus EG95 induces protective immunity in mice.

Cystic echinococcosis (CE), caused by Echinococcus granulosus (E, is a zoonosis with a worldwide distribution, resulting in heavy impact to public health and social economics. In this study, we generated a recombinant rabies virus (RABV) expressing EG95 protein of E. granulosus (LBNSE-EG95) as a bivalent candidate vaccine for use in sheep and cattle against CE and rabies, which is another severe health threat in CE-endemic areas. It was found that EG95 was successfully expressed without altering the pathogenicity of parent LBNSE vector. Further study showed that LBNSE-EG95 immunization in mice elicited activation of dendric cells (DCs) and B cells and induced Th1-/Th2-mediated cellular immune responses, leading to robust production of RABV neutralizing antibodies and high level of EG95-sepecific antibodies with more than 90% protection against CE. In addition, single dose of LBNSE-EG95 conferred full protection against lethal RABV challenge in mice. Collectively, these results suggest that the recombinant LBNSE-EG95 has the potential to be developed as an efficient bivalent vaccine for sheep and cattle use in endemic areas of CE and rabies.

Viral and Host Transcriptomes in SARS-CoV-2-Infected Human Lung Cells.

Coronaviruses are commonly characterized by a unique discontinuous RNA transcriptional synthesis strategy guided by transcription-regulating sequences (TRSs). However, the details of RNA synthesis in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have not been fully elucidated. Here, we present a time-scaled, gene-comparable transcriptome of SARS-CoV-2, demonstrating that ACGAAC functions as a core TRS guiding the discontinuous RNA synthesis of SARS-CoV-2 from a holistic perspective. During infection, viral transcription, rather than genome replication, dominates all viral RNA synthesis activities. The most highly expressed viral gene is the nucleocapsid gene, followed by ORF7 and ORF3 genes, while the envelope gene shows the lowest expression. Host transcription dysregulation keeps exacerbating after viral RNA synthesis reaches a maximum. The most enriched host pathways are metabolism related. Two of them (cholesterol and valine metabolism) affect viral replication in reverse. Furthermore, the activation of numerous cytokines emerges before large-scale viral RNA synthesis. IMPORTANCE SARS-CoV-2 is responsible for the current severe global health emergency that began at the end of 2019. Although the universal transcriptional strategies of coronaviruses are preliminarily understood, the details of RNA synthesis, especially the time-matched transcription level of each SARS-CoV-2 gene and the principles of subgenomic mRNA synthesis, are not clear. The coterminal subgenomic mRNAs of SARS-CoV-2 present obstacles in identifying the expression of most genes by PCR-based methods, which are exacerbated by the lack of related antibodies. Moreover, SARS-CoV-2-related metabolic imbalance and cytokine storm are receiving increasing attention from both clinical and mechanistic perspectives. Our transcriptomic research provides information on both viral RNA synthesis and host responses, in which the transcription-regulating sequences and transcription levels of viral genes are demonstrated, and the metabolic dysregulation and cytokine levels identified at the host cellular level support the development of novel medical treatment strategies.

An inactivated recombinant rabies virus displaying the Zika virus prM-E induces protective immunity against both pathogens.

The global spread of Zika virus (ZIKV), which caused a pandemic associated with Congenital Zika Syndrome and neuropathology in newborns and adults, prompted the pursuit of a safe and effective vaccine. Here, three kinds of recombinant rabies virus (RABV) encoding the prM-E protein of ZIKV were constructed: ZI-D (prM-E), ZI-E (transmembrane domain (TM) of prM-E replaced with RABV G) and ZI-F (signal peptide and TM domain of prM-E replaced with the region of RABV G). When the TM of prM-E was replaced with the region of RABV G (termed ZI-E), it promoted ZIKV E protein localization on the cell membrane and assembly on recombinant viruses. In addition, the change in the signal peptide with RABV G (termed ZI-F) was not conducive to foreign protein expression. The immunogenicity of recombinant viruses mixed with a complex adjuvant of ISA 201 VG and poly(I:C) was tested in BALB/c mice. After immunization with ZI-E, the anti-ZIKV IgG antibody lasted for at least 10 weeks. The titers of neutralizing antibodies (NAbs) against ZIKV and RABV at week 6 were all greater than the protective titers. Moreover, ZI-E stimulated the proliferation of splenic lymphocytes and promoted the secretion of cytokines. It also promoted the production of central memory T cells (TCMs) among CD4+/CD8+ T cells and stimulated B cell activation and maturation. These results indicate that ZI-E could induce ZIKV-specific humoral and cellular immune responses, which have the potential to be developed into a promising vaccine for protection against both ZIKV and RABV infections.

A highly efficient recombinant canarypox virus-based vaccine against canine distemper virus constructed using the CRISPR/Cas9 gene editing method.

Canine distemper virus (CDV) is the causative agent of canine distemper (CD), which is one of the most important infectious diseases affecting wild and domestic carnivores. Vaccination represents an effective approach to prevent CDV infection among domestic carnivores. Canarypox-vectored recombinant CD vaccines (such as Recombitek CDV, PureVax Ferret Distemper, and Merial) with the CDV hemagglutinin (H) and fusion (F) genes can induce a potent immune response in dogs and ferrets. However, the vaccine's effectiveness varies with the species. In the current study, we developed a highly efficient recombinant canarypox virus termed as "ALVAC-CDV-M-F-H/C5-" that contained CDV virus-like particles (VLPs) by using the CRISPR/Cas9 gene editing method, which enabled concurrent expression of the matrix (M), H, and F genes. The recombinant strain provided faster seroconversion than the parent strain among minks as well as provided higher rates of antibody positivity than the parent strain among foxes and minks even before the administration of a second booster vaccination. We demonstrated, for the first time, that the CRISPR/Cas9 system can be applied for the rapid and efficient modification of the ALVAC-CDV-F-H genome and also that a high-dose new recombinant strain that produces CDV VLPs may present good outcomes in the prevention of CD among foxes and minks.

Characterization of Two Neutralizing Antibodies against Rift Valley Fever Virus Gn Protein.

The Rift Valley fever virus (RVFV) is an arthropod-borne virus that can not only cause severe disease in domestic animals but also in humans. However, the licensed vaccines or available therapeutics for humans do not exist. Here, we report two Gn-specific neutralizing antibodies (NAbs), isolated from a rhesus monkey immunized with recombinant human adenoviruses type 4 expressing Rift Valley fever virus Gn and Gc protein (rHAdV4-GnGcopt). The two NAbs were both able to protect host cells from RVFV infection. The interactions between NAbs and Gn were then characterized to demonstrate that these two NAbs might preclude RVFV glycoprotein rearrangement, hindering the exposure of fusion loops in Gc to endosomal membranes after the virus invades the host cell. The target region for the two NAbs is located in the Gn domain III, implying that Gn is a desired target for developing vaccines and neutralizing antibodies against RVFV.

A Chimeric Sudan Virus-Like Particle Vaccine Candidate Produced by a Recombinant Baculovirus System Induces Specific Immune Responses in Mice and Horses.

Ebola virus infections lead to severe hemorrhagic fevers in humans and nonhuman primates; and human fatality rates are as high as 67%-90%. Since the Ebola virus was discovered in 1976, the only available treatments have been medical support or the emergency administration of experimental drugs. The absence of licensed vaccines and drugs against the Ebola virus impedes the prevention of viral infection. In this study, we generated recombinant baculoviruses (rBV) expressing the Sudan virus (SUDV) matrix structural protein (VP40) (rBV-VP40-VP40) or the SUDV glycoprotein (GP) (rBV-GP-GP), and SUDV virus-like particles (VLPs) were produced by co-infection of Sf9 cells with rBV-SUDV-VP40 and rBV-SUDV-GP. The expression of SUDV VP40 and GP in SUDV VLPs was demonstrated by IFA and Western blot analysis. Electron microscopy results demonstrated that SUDV VLPs had a filamentous morphology. The immunogenicity of SUDV VLPs produced in insect cells was evaluated by the immunization of mice. The analysis of antibody responses showed that mice vaccinated with SUDV VLPs and the adjuvant Montanide ISA 201 produced SUDV GP-specific IgG antibodies. Sera from SUDV VLP-immunized mice were able to block infection by SUDV GP pseudotyped HIV, indicating that a neutralizing antibody against the SUDV GP protein was produced. Furthermore, the activation of B cells in the group immunized with VLPs mixed with Montanide ISA 201 was significant one week after the primary immunization. Vaccination with the SUDV VLPs markedly increased the frequency of antigen-specific cells secreting type 1 and type 2 cytokines. To study the therapeutic effects of SUDV antibodies, horses were immunized with SUDV VLPs emulsified in Freund's complete adjuvant or Freund's incomplete adjuvant. The results showed that horses could produce SUDV GP-specific antibodies and neutralizing antibodies. These results showed that SUDV VLPs demonstrate excellent immunogenicity and represent a promising approach for vaccine development against SUDV infection. Further, these horse anti-SUDV purified immunoglobulins lay a foundation for SUDV therapeutic drug research.

RNA sequencing analyses of gene expressions in a canine macrophages cell line DH82 infected with canine distemper virus.

Virulent morbillivirus infections, including Meals Virus (MeV) and Canine Distemper Virus (CDV), caused severe immune suppression and leukopenia, while attenuated vaccine strains developed protective host immune responses. However, the detailed molecular foundations of host antiviral responses were poorly characterized. In order to better understand the interactions between attenuated vaccine and host antiviral responses, the global gene expression changes in CDV-11-infected DH82 cells, a macrophage-derived cell line from canine, were investigated by transcriptomic analysis, and portions of results were confirmed with quantitative RT-PCR. The results exhibited that 372 genes significantly up-regulated (p < .01) and 119 genes were significantly down-regulated (p < .01) in CDV-infected macrophages DH82 at 48 h p.i.. The enriched functions of the significantly up-regulated (p < .01) genes were closely associated with interferon stimulated genes (ISGs), chemokine genes and pro-inflammatory factor genes. Gene ontology and pathway analysis of differentially expressed genes (DEGs) revealed that the most significantly involved pathways in CDV-infected DH82 cells were NF-κB and TNF signaling pathway, cytokine-cytokine receptor interaction, and pathogen associated molecular patterns (PAMPs), such as Toll-like, RIG-I-like and NOD-like receptor signalings. Thus, the findings indicated that pattern recognition receptors (PRRs) possibly mediated host innate and protective antiviral immune responses in CDV-11 infected DH82 cells.

Ebola virus replication is regulated by the phosphorylation of viral protein VP35.

Ebola virus (EBOV) is a zoonotic pathogen, the infection often results in severe, potentially fatal, systematic disease in human and nonhuman primates. VP35, an essential viral RNA-dependent RNA polymerase cofactor, is indispensable for Ebola viral replication and host innate immune escape. In this study, VP35 was demonstrated to be phosphorylated at Serine/Threonine by immunoblotting, and the major phosphorylation sites was S187, S205, T206, S208 and S317 as revealed by LC-MS/MS. By an EBOV minigenomic system, EBOV minigenome replication was shown to be significantly inhibited by the phosphorylation-defective mutant, VP35 S187A, but was potentiated by the phosphorylation mimic mutant VP35 S187D. Together, our findings demonstrate that EBOV VP35 is phosphorylated on multiple residues in host cells, especially on S187, which may contribute to efficient viral genomic replication and viral proliferation.