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1.
J Virol ; 96(16): e0062722, 2022 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-35924923

RESUMO

Rotavirus live-attenuated vaccines, both mono- and pentavalent, generate broadly heterotypic protection. B-cells isolated from adults encode neutralizing antibodies, some with affinity for VP5*, that afford broad protection in mice. We have mapped the epitope of one such antibody by determining the high-resolution cryo-EM structure of its antigen-binding fragment (Fab) bound to the virion of a candidate vaccine strain, CDC-9. The Fab contacts both the distal end of a VP5* ß-barrel domain and the two VP8* lectin-like domains at the tip of a projecting spike. Its interactions with VP8* do not impinge on the likely receptor-binding site, suggesting that the mechanism of neutralization is at a step subsequent to initial attachment. We also examined structures of CDC-9 virions from two different stages of serial passaging. Nearly all the VP4 (cleaved to VP8*/VP5*) spikes on particles from the earlier passage (wild-type isolate) had transitioned from the "upright" conformation present on fully infectious virions to the "reversed" conformation that is probably the end state of membrane insertion, unable to mediate penetration, consistent with the very low in vitro infectivity of the wild-type isolate. About half the VP4 spikes were upright on particles from the later passage, which had recovered substantial in vitro infectivity but had acquired an attenuated phenotype in neonatal rats. A mutation in VP4 that occurred during passaging appears to stabilize the interface at the apex of the spike and could account for the greater stability of the upright spikes on the late-passage, attenuated isolate. IMPORTANCE Rotavirus live-attenuated vaccines generate broadly heterotypic protection, and B-cells isolated from adults encode antibodies that are broadly protective in mice. Determining the structural and mechanistic basis of broad protection can contribute to understanding the current limitations of vaccine efficacy in developing countries. The structure of an attenuated human rotavirus isolate (CDC-9) bound with the Fab fragment of a broadly heterotypic protective antibody shows that protection is probably due to inhibition of the conformational transition in the viral spike protein (VP4) critical for viral penetration, rather than to inhibition of receptor binding. A comparison of structures of CDC-9 virus particles at two stages of serial passaging supports a proposed mechanism for initial steps in rotavirus membrane penetration.


Assuntos
Anticorpos Amplamente Neutralizantes , Proteínas do Capsídeo , Epitopos de Linfócito B , Rotavirus , Vacinas Atenuadas , Vírion , Animais , Anticorpos Amplamente Neutralizantes/imunologia , Anticorpos Amplamente Neutralizantes/ultraestrutura , Proteínas do Capsídeo/química , Proteínas do Capsídeo/imunologia , Proteínas do Capsídeo/ultraestrutura , Microscopia Crioeletrônica , Epitopos de Linfócito B/imunologia , Epitopos de Linfócito B/ultraestrutura , Humanos , Fragmentos Fab das Imunoglobulinas/imunologia , Fragmentos Fab das Imunoglobulinas/ultraestrutura , Camundongos , Conformação Proteica , Ratos , Rotavirus/química , Rotavirus/classificação , Rotavirus/imunologia , Rotavirus/fisiologia , Inoculações Seriadas , Vacinas Atenuadas/química , Vacinas Atenuadas/imunologia , Vacinas Atenuadas/metabolismo , Vírion/imunologia , Vírion/metabolismo , Vírion/ultraestrutura
2.
Proc Natl Acad Sci U S A ; 119(33): e2201616119, 2022 08 16.
Artigo em Inglês | MEDLINE | ID: mdl-35895717

RESUMO

With the rapid increase in SARS-CoV-2 cases in children, a safe and effective vaccine for this population is urgently needed. The MMR (measles/mumps/rubella) vaccine has been one of the safest and most effective human vaccines used in infants and children since the 1960s. Here, we developed live attenuated recombinant mumps virus (rMuV)-based SARS-CoV-2 vaccine candidates using the MuV Jeryl Lynn (JL2) vaccine strain backbone. The soluble prefusion SARS-CoV-2 spike protein (preS) gene, stablized by two prolines (preS-2P) or six prolines (preS-6P), was inserted into the MuV genome at the P-M or F-SH gene junctions in the MuV genome. preS-6P was more efficiently expressed than preS-2P, and preS-6P expression from the P-M gene junction was more efficient than from the F-SH gene junction. In mice, the rMuV-preS-6P vaccine was more immunogenic than the rMuV-preS-2P vaccine, eliciting stronger neutralizing antibodies and mucosal immunity. Sera raised in response to the rMuV-preS-6P vaccine neutralized SARS-CoV-2 variants of concern, including the Delta variant equivalently. Intranasal and/or subcutaneous immunization of IFNAR1-/- mice and golden Syrian hamsters with the rMuV-preS-6P vaccine induced high levels of neutralizing antibodies, mucosal immunoglobulin A antibody, and T cell immune responses, and were completely protected from challenge by both SARS-CoV-2 USA-WA1/2020 and Delta variants. Therefore, rMuV-preS-6P is a highly promising COVID-19 vaccine candidate, warranting further development as a tetravalent MMR vaccine, which may include protection against SARS-CoV-2.


Assuntos
Vacinas contra COVID-19 , COVID-19 , Vacina contra Sarampo-Caxumba-Rubéola , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Eficácia de Vacinas , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , COVID-19/prevenção & controle , Vacinas contra COVID-19/genética , Vacinas contra COVID-19/imunologia , Imunogenicidade da Vacina , Vacina contra Sarampo-Caxumba-Rubéola/genética , Vacina contra Sarampo-Caxumba-Rubéola/imunologia , Mesocricetus , Camundongos , Vírus da Caxumba/genética , Vírus da Caxumba/imunologia , Prolina/genética , SARS-CoV-2/genética , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia
3.
Front Immunol ; 13: 832264, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35558083

RESUMO

African swine fever (ASF) is among the most devastating viral diseases of pigs and wild boar worldwide. In recent years, the disease has spread alarmingly. Despite intensive research activities, a commercialized vaccine is still not available, and efficacious live attenuated vaccine candidates raise safety concerns. From a safety perspective, inactivated preparations would be most favourable. However, both historical and more recent trials with chemical inactivation did not show an appreciable protective effect. Under the assumption that the integrity of viral particles could enhance presentation of antigens, we used gamma irradiation for inactivation. To this means, gamma irradiated ASFV "Estonia 2014" was adjuvanted with either Polygen™ or Montanide™ ISA 201 VG, respectively. Subsequently, five weaner pigs per preparation were immunized twice with a three-week interval. Six weeks after the first immunization, all animals were challenged with the highly virulent ASFV strain "Armenia 2008". Although ASFV p72-specific IgG antibodies were detectable in all vaccinated animals prior challenge, no protection could be observed. All animals developed an acute lethal course of ASF and had to be euthanized at a moderate humane endpoint within six days. Indeed, the vaccinated pigs showed even higher clinical scores and a higher inner body temperature than the control group. However, significantly lower viral loads were detectable in spleen and liver of immunized animals at the time point of euthanasia. This phenomenon suggests an immune mediated disease enhancement that needs further investigation.


Assuntos
Febre Suína Africana , Vacinas Virais , Febre Suína Africana/prevenção & controle , Vírus da Febre Suína Africana , Animais , Raios gama , Imunogenicidade da Vacina , Suínos , Vacinação , Vacinas Atenuadas/imunologia , Proteínas Virais , Vacinas Virais/imunologia
4.
Viruses ; 14(5)2022 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-35632736

RESUMO

Live-attenuated SARS-CoV-2 vaccines received relatively little attention during the COVID-19 pandemic. Despite this, several methods of obtaining attenuated coronaviruses are known. In this systematic review, the strategies of coronavirus attenuation, which may potentially be applied to SARS-CoV-2, were identified. PubMed, Scopus, Web of Science and Embase databases were searched to identify relevant articles describing attenuating mutations tested in vivo. In case of coronaviruses other than SARS-CoV-2, sequence alignment was used to exclude attenuating mutations that cannot be applied to SARS-CoV-2. Potential immunogenicity, safety and efficacy of the attenuated SARS-CoV-2 vaccine were discussed based on animal studies data. A total of 27 attenuation strategies, used to create 101 different coronaviruses, have been described in 56 eligible articles. The disruption of the furin cleavage site in the SARS-CoV-2 spike protein was identified as the most promising strategy. The replacement of core sequences of transcriptional regulatory signals, which prevents recombination with wild-type viruses, also appears particularly advantageous. Other important attenuating mutations encompassed mostly the prevention of evasion of innate immunity. Sufficiently attenuated coronaviruses typically caused no meaningful disease in susceptible animals and protected them from challenges with virulent virus. This indicates that attenuated COVID-19 vaccines may be considered as a potential strategy to fight the threat posed by SARS-CoV-2.


Assuntos
Vacinas contra COVID-19 , COVID-19 , Desenvolvimento de Vacinas , Animais , COVID-19/prevenção & controle , Vacinas contra COVID-19/imunologia , Humanos , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/genética , Vacinas Atenuadas/imunologia
5.
J Virol ; 96(9): e0006422, 2022 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-35416719

RESUMO

Alphaviruses infect cells by a low pH-dependent fusion reaction between viral and host cell membranes that is mediated by the viral E1 glycoprotein. Most reported alphavirus E1 sequences include two phenylalanines (F87 and F95) in the fusion loop, yet the role of these residues in viral infectivity remains to be defined. Following introduction of wild type (WT), E1-F87A, and E1-F95A chikungunya virus (CHIKV) RNA genomes into cells, viral particle production was similar in magnitude. However, CHIKV E1-F87A and E1-F95A virions displayed impaired infectivity compared with WT CHIKV particles. Although WT, E1-F87A, and E1-F95A particles bound cells with similar efficiencies, E1-F87A and E1-F95A particles were unable to undergo fusion and entry into cells. Introduction of an F95A mutation in the E1 fusion loop of Mayaro virus or Venezuelan equine encephalitis virus also resulted in poorly infectious virions. We further tested whether an E1-F87A or E1-F95A mutation could be incorporated into a live-attenuated vaccine strain, CHIKV 181/25, to enhance vaccine safety. Infection of immunocompromised Ifnar1-/- and Irf3-/-Irf5-/-Irf7-/- mice with 181/25E1-F87A or 181/25E1-F95A resulted in 0% mortality, compared with 100% mortality following 181/25 infection. Despite this enhanced attenuation, surviving Ifnar1-/- and Irf3-/-Irf5-/-Irf7-/- mice were protected against virulent virus re-challenge. Moreover, single-dose immunization of WT mice with either 181/25, 181/25E1-F87A, or 181/25E1-F95A elicited CHIKV-specific antibody responses and protected against pathogenic CHIKV challenge. These studies define a critical function for residues E1-F87 and E1-F95 in alphavirus fusion and entry into target cells and suggest that incorporation of these mutations could enhance the safety of live-attenuated alphavirus vaccine candidates. IMPORTANCE Alphaviruses are human pathogens that cause both debilitating acute and chronic musculoskeletal disease and potentially fatal encephalitis. In this study, we determined that two highly conserved phenylalanine residues in the alphavirus E1 glycoprotein are required for fusion of viral and host cell membranes and viral entry into target cells. We further demonstrated that mutation of these phenylalanines results in a substantial loss of viral virulence but not immunogenicity. These data enhance an understanding of the viral determinants of alphavirus entry into host cells and could contribute to the development of new antivirals targeting these conserved phenylalanines or new live-attenuated alphavirus vaccines.


Assuntos
Febre de Chikungunya , Vírus Chikungunya , Proteínas do Envelope Viral , Vacinas Virais , Animais , Anticorpos Antivirais , Febre de Chikungunya/virologia , Vírus Chikungunya/patogenicidade , Vírus Chikungunya/fisiologia , Fatores Reguladores de Interferon/metabolismo , Camundongos , Camundongos Knockout , Fenilalanina/química , Domínios Proteicos , Vacinas Atenuadas/imunologia , Proteínas do Envelope Viral/química , Vacinas Virais/imunologia , Replicação Viral
6.
J Virol ; 96(8): e0020922, 2022 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-35343792

RESUMO

Several highly pathogenic mammarenaviruses cause severe hemorrhagic and neurologic disease in humans for which vaccines and antivirals are limited or unavailable. New World (NW) mammarenavirus Machupo virus (MACV) infection causes Bolivian hemorrhagic fever in humans. We previously reported that the disruption of specific N-linked glycan sites on the glycoprotein (GPC) partially attenuates MACV in an interferon alpha/beta and gamma (IFN-α/ß and -γ) receptor knockout (R-/-) mouse model. However, some capability to induce neurological pathology still remained. The highly pathogenic Junin virus (JUNV) is another NW arenavirus closely related to MACV. An F427I substitution in the GPC transmembrane domain (TMD) rendered JUNV attenuated in a lethal mouse model after intracranial inoculation. In this study, we rationally designed and rescued a MACV containing mutations at two glycosylation sites and the corresponding F438I substitution in the GPC TMD. The MACV mutant is fully attenuated in IFN-α/ß and -γ R-/- mice and outbred guinea pigs. Furthermore, inoculation with this mutant MACV completely protected guinea pigs from wild-type MACV lethal challenge. Last, we found the GPC TMD F438I substitution greatly impaired MACV growth in neuronal cell lines of mouse and human origins. Our results highlight the critical roles of the glycans and the TMD on the GPC in arenavirus virulence, which provide insight into the rational design of potential vaccine candidates for highly pathogenic arenaviruses. IMPORTANCE For arenaviruses, the only vaccine available is the live attenuated Candid#1 vaccine, a JUNV vaccine approved in Argentina. We and others have found that the glycans on GPC and the F427 residue in the GPC TMD are important for virulence of JUNV. Nevertheless, mutating either of them is not sufficient for full and stable attenuation of JUNV. Using reverse genetics, we disrupted specific glycosylation sites on MACV GPC and also introduced the corresponding F438I substitution in the GPC TMD. This MACV mutant is fully attenuated in two animal models and protects animals from lethal infection. Thus, our studies highlight the feasibility of rational attenuation of highly pathogenic arenaviruses for vaccine development. Another important finding from this study is that the F438I substitution in GPC TMD could substantially affect MACV replication in neurons. Future studies are warranted to elucidate the underlying mechanism and the implication of this mutation in arenavirus neural tropism.


Assuntos
Arenavirus do Novo Mundo , Febre Hemorrágica Americana , Vacinas Virais , Animais , Arenavirus do Novo Mundo/genética , Arenavirus do Novo Mundo/imunologia , Modelos Animais de Doenças , Glicoproteínas/metabolismo , Glicosilação , Cobaias , Febre Hemorrágica Americana/imunologia , Febre Hemorrágica Americana/virologia , Vírus Junin/genética , Vírus Junin/imunologia , Mutação , Vacinas Atenuadas/imunologia , Vacinas Virais/imunologia
7.
Nat Commun ; 13(1): 824, 2022 02 11.
Artigo em Inglês | MEDLINE | ID: mdl-35149692

RESUMO

Varicella caused by the primary infection of varicella-zoster virus (VZV) exerts a considerable disease burden globally. Current varicella vaccines consisting of the live-attenuated vOka strain of VZV are generally safe and effective. However, vOka retains full neurovirulence and can establish latency and reactivate to cause herpes zoster in vaccine recipients, raising safety concerns. Here, we rationally design a live-attenuated varicella vaccine candidate, v7D. This virus replicates like wild-type virus in MRC-5 fibroblasts and human PBMCs, the carrier for VZV dissemination, but is severely impaired for infection of human skin and neuronal cells. Meanwhile, v7D shows immunogenicity comparable to vOka both in vitro and in multiple small animal species. Finally, v7D is proven well-tolerated and immunogenic in nonhuman primates. Our preclinical data suggest that v7D is a promising candidate as a safer live varicella vaccine with reduced risk of vaccine-related complications, and could inform the design of other herpes virus vaccines.


Assuntos
Vacina contra Varicela/imunologia , Varicela/imunologia , Pele/imunologia , Vacinas Atenuadas/imunologia , Animais , Linhagem Celular , Varicela/prevenção & controle , Feminino , Fibroblastos , Cobaias , Herpes Zoster/virologia , Herpesvirus Humano 3 , Humanos , Imunogenicidade da Vacina , Pulmão , Masculino , Camundongos , Neurônios/patologia , Coelhos , Ratos , Pele/patologia , Vacinação , Vacinas Virais
8.
Front Immunol ; 13: 787021, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35173716

RESUMO

Vaccination is the best form of protecting fish against viral diseases when the pathogen cannot be contained by biosecurity measures. Vaccines based on live attenuated viruses seem to be most effective for vaccination against challenging pathogens like Cyprinid herpesvirus 3. However, there are still knowledge gaps how these vaccines effectively protect fish from the deadly disease caused by the epitheliotropic CyHV-3, and which aspects of non-direct protection of skin or gill integrity and function are important in the aquatic environment. To elucidate some elements of protection, common carp were vaccinated against CyHV-3 using a double deletion vaccine virus KHV-T ΔDUT/TK in the absence or presence of a mix of common carp beta-defensins 1, 2 and 3 as adjuvants. Vaccination induced marginal clinical signs, low virus load and a minor upregulation of cd4, cd8 and igm gene expression in vaccinated fish, while neutralisation activity of blood serum rose from 14 days post vaccination (dpv). A challenge infection with CyHV-3 induced a severe disease with 80-100% mortality in non-vaccinated carp, while in vaccinated carp, no mortality was recorded and the virus load was >1,000-fold lower in the skin, gill and kidney. Histological analysis showed strongest pathological changes in the skin, with a complete destruction of the epidermis in non-vaccinated carp. In the skin of non-vaccinated fish, T and B cell responses were severely downregulated, inflammation and stress responses were increased upon challenge, whereas vaccinated fish had boosted neutrophil, T and B cell responses. A disruption of skin barrier elements (tight and adherence junction, desmosomes, mucins) led to an uncontrolled increase in skin bacteria load which most likely exacerbated the inflammation and the pathology. Using a live attenuated virus vaccine, we were able to show that increased neutrophil, T and B cell responses provide protection from CyHV-3 infection and lead to preservation of skin integrity, which supports successful protection against additional pathogens in the aquatic environment which foster disease development in non-vaccinated carp.


Assuntos
Doenças dos Peixes/imunologia , Doenças dos Peixes/prevenção & controle , Infecções por Herpesviridae/veterinária , Herpesviridae/imunologia , Vacinas Virais/imunologia , Animais , Carpas , Herpesviridae/genética , Infecções por Herpesviridae/imunologia , Vacinação , Vacinas Atenuadas/imunologia , Vacinas Virais/genética
9.
PLoS Pathog ; 18(2): e1010282, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35108339

RESUMO

Immunization with radiation-attenuated sporozoites (RAS) can confer sterilizing protection against malaria, although the mechanisms behind this protection are incompletely understood. We performed a systems biology analysis of samples from the Immunization by Mosquito with Radiation Attenuated Sporozoites (IMRAS) trial, which comprised P. falciparum RAS-immunized (PfRAS), malaria-naive participants whose protection from malaria infection was subsequently assessed by controlled human malaria infection (CHMI). Blood samples collected after initial PfRAS immunization were analyzed to compare immune responses between protected and non-protected volunteers leveraging integrative analysis of whole blood RNA-seq, high parameter flow cytometry, and single cell CITEseq of PBMCs. This analysis revealed differences in early innate immune responses indicating divergent paths associated with protection. In particular, elevated levels of inflammatory responses early after the initial immunization were detrimental for the development of protective adaptive immunity. Specifically, non-classical monocytes and early type I interferon responses induced within 1 day of PfRAS vaccination correlated with impaired immunity. Non-protected individuals also showed an increase in Th2 polarized T cell responses whereas we observed a trend towards increased Th1 and T-bet+ CD8 T cell responses in protected individuals. Temporal differences in genes associated with natural killer cells suggest an important role in immune regulation by these cells. These findings give insight into the immune responses that confer protection against malaria and may guide further malaria vaccine development. Trial registration: ClinicalTrials.gov NCT01994525.


Assuntos
Imunidade , Inflamação , Vacinas Antimaláricas/imunologia , Malária Falciparum/imunologia , Plasmodium falciparum/imunologia , Esporozoítos/imunologia , Adulto , Animais , Anopheles/parasitologia , Feminino , Humanos , Imunização/métodos , Mordeduras e Picadas de Insetos/imunologia , Malária Falciparum/parasitologia , Masculino , Mosquitos Vetores/parasitologia , Linfócitos T/imunologia , Vacinação/métodos , Vacinas Atenuadas/imunologia
10.
Microbiol Spectr ; 10(1): e0143621, 2022 02 23.
Artigo em Inglês | MEDLINE | ID: mdl-35107364

RESUMO

Hepatitis-hydropericardium syndrome (HHS) caused by the highly pathogenic fowl adenovirus serotype 4 (FAdV-4) has resulted in huge economic losses to the poultry industry globally. The fiber-2 gene, as a major virulence determiner, is also an important vaccine target against FAdV-4. In this study, we used a CRISPR/Cas9-based homology-dependent recombinant technique to replace the fiber-2 gene with egfp and generate a novel recombinant virus, designated FAdV4-EGFP-rF2. Although FAdV4-EGFP-rF2 showed low replication ability compared to the wild-type FAdV-4 in LMH cells, FAdV4-EGFP-rF2 could effectively replicate in LMH-F2 cells with the expression of Fiber-2. Moreover, FAdV4-EGFP-rF2 was not only highly attenuated in chickens, but also could provide efficient protection against a lethal challenge of FAdV-4. Moreover, FAdV4-EGFP-rF2 without fiber-2 could induce neutralizing antibodies at the same level as FA4-EGFP with fiber-2. These results clearly demonstrate that although fiber-2 affects the viral replication and pathogenesis of FAdV-4, it is not necessary for virus replication and induction of neutralizing antibodies; these findings provide novel insights into the roles of fiber-2 and highlight fiber-2 as an insertion site for generating live-attenuated FAdV-4 vaccines against FAdV-4 and other pathogens. IMPORTANCE Among all serotypes of fowl adenovirus, serotypes FAdV-1, FAdV-4, and FAdV-10 are unique members with two fiber genes (fiber-1 and fiber-2). Recent studies reveal that Fiber-1, not Fiber-2, directly triggers viral infection of FAdV-4, whereas Fiber-2, but not Fiber-1, has been identified as the major virulence determiner and an efficient protective immunogen for subunit vaccines. Here, we replaced fiber-2 with egfp to generate a novel recombinant virus, designated FAdV4-EGFP-rF2. In vitro and in vivo studies on FAdV4-EGFP-rF2 revealed that fiber-2 was not necessary for either virus replication or efficient protection for FAdV-4; these results not only provide a novel live-attenuated vaccine candidate against HHS, but also give new ideas for generating a FAdV-4 based vaccine vector against other pathogens.


Assuntos
Infecções por Adenoviridae/veterinária , Aviadenovirus/imunologia , Doenças das Aves Domésticas/prevenção & controle , Proteínas Virais/imunologia , Vacinas Virais/imunologia , Infecções por Adenoviridae/imunologia , Infecções por Adenoviridae/prevenção & controle , Infecções por Adenoviridae/virologia , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Aviadenovirus/genética , Aviadenovirus/fisiologia , Galinhas , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/virologia , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , Proteínas Virais/administração & dosagem , Proteínas Virais/genética , Vacinas Virais/administração & dosagem , Vacinas Virais/genética
11.
Cell Rep ; 38(6): 110341, 2022 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-35139383

RESUMO

The tetravalent dengue vaccine candidate, TAK-003, induces a functional antibody response, but the titers of antibodies against the four serotypes of the dengue virus (DENV) can vary. Here, through a transcriptomic analysis on whole blood collected from recipients of a two-dose schedule of TAK-003, we examine gene expression, splicing, and transcript isoform-level changes for both protein-coding and noncoding genes to broaden our understanding of the immune response. Our analysis reveals a dynamic pattern of vaccine-associated regulation of long noncoding RNAs (lncRNAs), differential splicing of interferon-stimulated gene exons, and gene expression changes related to multiple signaling pathways that detect viral infection. Co-expression networks isolate immune cell-type-related and interferon-response modules that represent specific biological processes that correlate with more robust antibody responses. These data provide insights into the early determinants of the variable immune response to the vaccine, highlighting the significance of splicing and isoform-level gene regulatory mechanisms in defining vaccine immunogenicity.


Assuntos
Anticorpos Antivirais/imunologia , Vacinas contra Dengue/imunologia , Vírus da Dengue/patogenicidade , RNA Longo não Codificante/genética , Transcriptoma/genética , Anticorpos Neutralizantes/imunologia , Dengue/virologia , Vírus da Dengue/genética , Humanos , Imunogenicidade da Vacina/imunologia , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/imunologia , Vacinas Virais/imunologia , Vacinas Virais/farmacologia
12.
Proc Natl Acad Sci U S A ; 119(8)2022 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-35177472

RESUMO

While influenza infection induces robust, long-lasting, antibody responses and protection, including the T follicular helper cells (TFH) required to drive B cell germinal center (GC) responses, most influenza vaccines do not. We investigated the mechanisms that drive strong TFH responses during infection. Infection induces viral replication and antigen (Ag) presentation lasting through the CD4 effector phase, but Ag and pathogen recognition receptor signals are short-lived after vaccination. We analyzed the need for both infection and Ag presentation at the effector phase, using an in vivo sequential transfer model to time their availability. Differentiation of CD4 effectors into TFH and GC-TFH required that they recognize Ag locally in the site of TFH development, at the effector phase, but did not depend on specific Ag-presenting cells (APCs). In addition, concurrent signals from infection were necessary even when sufficient Ag was presented. Providing these signals with a second dose of live attenuated influenza vaccine at the effector phase drove TFH and GC-TFH development equivalent to live infection. The results suggest that vaccine approaches can induce strong TFH development that supports GC responses akin to infection, if they supply these effector phase signals at the right time and site. We suggest that these requirements create a checkpoint that ensures TFH only develop fully when infection is still ongoing, thereby avoiding unnecessary, potentially autoimmune, responses.


Assuntos
Vacinas contra Influenza/imunologia , Influenza Humana/imunologia , Células T Auxiliares Foliculares/imunologia , Animais , Anticorpos Antivirais/imunologia , Formação de Anticorpos/imunologia , Antígenos , Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Feminino , Centro Germinativo/imunologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Células T Auxiliares Foliculares/metabolismo , Linfócitos T Auxiliares-Indutores/imunologia , Vacinas Atenuadas/imunologia
13.
Viruses ; 14(1)2022 01 09.
Artigo em Inglês | MEDLINE | ID: mdl-35062317

RESUMO

Expansion of genotype I (GI) Japanese encephalitis viruses (JEV) has resulted in the replacement of the dominant genotype III (GIII) viruses, raising serious public health concerns for using GIII virus-derived vaccines to effectively control JEV epidemics. Therefore, this study used swine as the model to estimate the effectiveness of GIII live-attenuated vaccine against GI virus infection by comparing the incidence of stillbirth/abortion in gilts from vaccinated and non-vaccinated pig farms during the GI-circulation period. In total, 389 and 213 litters of gilts were recorded from four vaccinated and two non-vaccinated pig farms, respectively. All viruses detected in the aborted fetuses and mosquitoes belonged to the GI genotype during the study period. We thus estimated that the vaccine effectiveness of GIII live-attenuated vaccine against GI viruses in naive gilts based on the overall incidence of stillbirth/abortion and incidence of JEV-confirmed stillbirth/abortion was 65.5% (50.8-75.7%) and 74.7% (34.5-90.2%), respectively. In contrast to previous estimates, the GIII live-attenuated vaccine had an efficacy of 95.6% (68.3-99.4%) to prevent the incidence of stillbirth/abortion during the GIII-circulating period. These results indicate that the vaccine effectiveness of GIII live-attenuated JEV vaccine to prevent stillbirth/abortion caused by GI viruses is lower than that against GIII viruses.


Assuntos
Encefalite Japonesa/virologia , Genótipo , Doenças dos Suínos/virologia , Vacinas Virais/imunologia , Animais , Anticorpos Neutralizantes/imunologia , Culicidae , Vírus da Encefalite Japonesa (Espécie)/genética , Feminino , Imunização , Vacinas contra Encefalite Japonesa/imunologia , Filogenia , Suínos , Taiwan , Vacinas Atenuadas/imunologia
14.
Nat Commun ; 13(1): 549, 2022 01 27.
Artigo em Inglês | MEDLINE | ID: mdl-35087093

RESUMO

Adjuvants hold great potential in enhancing vaccine efficacy, making the understanding and improving of adjuvants critical goals in vaccinology. The TLR7/8 agonist, 3M-052, induces long-lived humoral immunity in non-human primates and is currently being evaluated in human clinical trials. However, the innate mechanisms of 3M-052 have not been fully characterized. Here, we perform flow cytometry, single cell RNA-seq and ATAC-seq to profile the kinetics, transcriptomics and epigenomics of innate immune cells in murine draining lymph nodes following 3M-052-Alum/Ovalbumin immunization. We find that 3M-052-Alum/OVA induces a robust antiviral and interferon gene program, similar to the yellow fever vaccine, which is known to confer long-lasting protection. Activation of myeloid cells in dLNs persists through day 28 and single cell analysis reveals putative TF-gene regulatory programs in distinct myeloid cells and heterogeneity of monocytes. This study provides a comprehensive characterization of the transcriptomics and epigenomics of innate populations in the dLNs after vaccination.


Assuntos
Adjuvantes Imunológicos/química , Imunidade Humoral/imunologia , Imunidade Inata , Vacinas Atenuadas/imunologia , Imunidade Adaptativa , Adjuvantes Imunológicos/farmacologia , Compostos de Alúmen , Animais , Anticorpos Antivirais/imunologia , Epigenômica , Feminino , Humanos , Imunidade Inata/efeitos dos fármacos , Imunização , Glicoproteínas de Membrana/agonistas , Camundongos , Camundongos Endogâmicos C57BL , Monócitos , Células Mieloides , Ovalbumina , Receptor 7 Toll-Like/agonistas , Receptor 8 Toll-Like/agonistas , Vacinação
15.
Lancet Infect Dis ; 22(1): e2-e12, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34506734

RESUMO

Mycobacterium bovis bacille Calmette-Guérin (BCG), an experimental vaccine designed to protect cattle from bovine tuberculosis, was administered for the first time to a newborn baby in Paris in 1921. Over the past century, BCG has saved tens of millions of lives and has been given to more humans than any other vaccine. It remains the sole tuberculosis vaccine licensed for use in humans. BCG provides long-lasting strong protection against miliary and meningeal tuberculosis in children, but it is less effective for the prevention of pulmonary tuberculosis, especially in adults. Evidence mainly from the past two decades suggests that BCG has non-specific benefits against non-tuberculous infections in newborn babies and in older adults, and offers immunotherapeutic benefit in certain malignancies such as non-muscle invasive bladder cancer. However, as a live attenuated vaccine, BCG can cause localised or disseminated infections in immunocompromised hosts, which can also occur following intravesical installation of BCG for the treatment of bladder cancer. The legacy of BCG includes fundamental discoveries about tuberculosis-specific and non-specific immunity and the demonstration that tuberculosis is a vaccine-preventable disease, providing a foundation for new vaccines to hasten tuberculosis elimination.


Assuntos
Vacina BCG/história , Vacina BCG/imunologia , Mycobacterium bovis/imunologia , Tuberculose Bovina/epidemiologia , Tuberculose Bovina/imunologia , Animais , Vacina BCG/efeitos adversos , Bovinos , História do Século XIX , História do Século XX , História do Século XXI , Humanos , Mycobacterium bovis/patogenicidade , Tuberculose Bovina/microbiologia , Tuberculose Bovina/prevenção & controle , Vacinas Atenuadas/imunologia
16.
J Med Virol ; 94(1): 82-87, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34524688

RESUMO

The rapid spread of the Delta variant suggests that SARS-CoV-2 will likely be rampant for months or years and could claim millions of more lives. All the known vaccines cannot well defeat SARS-CoV-2 due to their limited efficacy and production efficiency, except for the neglected live-attenuated vaccines (LAVs), which could have a much higher efficacy and much higher production efficiency than other vaccines. LAVs, like messiahs, have defeated far more pathogenic viruses than other vaccines in history, and most current human vaccines for viral diseases are safe LAVs. LAVs can block completely infection and transmission of relevant viruses and their variants. They can hence inhibit the emergence of vaccine-escape and virulence-enhancing variants and protect immunologically abnormal individuals better in general. The safety of COVID-19 LAVs, which could save millions of more lives, can be solidly guaranteed through animal experiments and clinical trials. The safety of COVID-19 LAVs could be greatly enhanced with intramuscular or oral administration, or administration along with humanized neutralizing monoclonal antibodies. Together, extensive global collaboration, which can greatly accelerate the development of safe COVID-19 LAVs, is imminently needed.


Assuntos
Vacinas contra COVID-19/imunologia , COVID-19/prevenção & controle , SARS-CoV-2/imunologia , Vacinas Atenuadas/imunologia , Desenvolvimento de Medicamentos , Humanos , Vacinação em Massa
17.
Methods Mol Biol ; 2410: 229-263, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-34914050

RESUMO

Vaccines are one of mankind's greatest medical advances, and their use has drastically reduced and in some cases eliminated (e.g., smallpox) disease and death caused by infectious agents. Traditional vaccine modalities including live-attenuated pathogen vaccines, wholly inactivated pathogen vaccines, and protein-based pathogen subunit vaccines have successfully been used to create efficacious vaccines against measles, mumps, rubella, polio, and yellow fever. These traditional vaccine modalities, however, take many months to years to develop and have thus proven less effective for use in creating vaccines to emerging or reemerging infectious diseases (EIDs) including influenza, Human immunodeficiency virus (HIV), dengue virus (DENV), chikungunya virus (CHIKV), West Nile virus (WNV), Middle East respiratory syndrome (MERS), and the severe acute respiratory syndrome coronaviruses 1 and 2 (SARS-CoV and SARS-CoV-2). As factors such as climate change and increased globalization continue to increase the pace of EID development, newer vaccine modalities are required to develop vaccines that can prevent or attenuate EID outbreaks throughout the world. One such modality, DNA vaccines, has been studied for over 30 years and has numerous qualities that make them ideal for meeting the challenge of EIDs including; (1) DNA vaccine candidates can be designed within hours of publishing of a pathogens genetic sequence; (2) they can be manufactured cheaply and rapidly in large quantities; (3) they are thermostable and have reduced requirement for a cold-chain during distribution, and (4) they have a remarkable safety record in the clinic. Optimizations made in plasmid design as well as in DNA vaccine delivery have greatly improved the immunogenicity of these vaccines. Here we describe the process of making a DNA vaccine to an EID pathogen and describe methods used for assessing the immunogenicity and protective efficacy of DNA vaccines in small animal models.


Assuntos
Doenças Transmissíveis Emergentes , Vacinas de DNA , Vacinas Virais , Animais , COVID-19 , Doenças Transmissíveis Emergentes/prevenção & controle , Humanos , Imunidade , SARS-CoV-2 , Vacinas Atenuadas/imunologia , Vacinas de DNA/imunologia , Vacinas de Produtos Inativados/imunologia , Vacinas Sintéticas/imunologia , Vacinas Virais/genética , Vacinas Virais/imunologia
18.
Methods Mol Biol ; 2411: 77-92, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-34816399

RESUMO

Vaccines are the most effective and economic way of combating poultry viruses. However, the use of traditional live-attenuated poultry vaccines has problems such as antigenic differences with the currently circulating strains of viruses and the risk of reversion to virulence. In veterinary medicine, reverse genetics is applied to solve these problems by developing genotype-matched vaccines, better attenuated and effective live vaccines, broad-spectrum vaccine vectors, bivalent vaccines, and genetically tagged recombinant vaccines that facilitate the serological differentiation of vaccinated animals from infected animals. In this chapter, we discuss reverse genetics as a tool for the development of recombinant vaccines against economically devastating poultry viruses.


Assuntos
Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/prevenção & controle , Aves Domésticas/imunologia , Genética Reversa/métodos , Vacinas Sintéticas/genética , Vacinas Virais/genética , Animais , Anticorpos Antivirais/imunologia , Galinhas/imunologia , Vacinas Atenuadas/imunologia , Vacinas Sintéticas/imunologia , Vacinas Virais/imunologia
19.
J Virol ; 96(2): e0176821, 2022 01 26.
Artigo em Inglês | MEDLINE | ID: mdl-34730400

RESUMO

Classical swine fever (CSF) is an economically important disease of pigs caused by classical swine fever virus (CSFV). The live attenuated vaccine C-strain (also called HCLV strain) against CSF was produced by multiple passages of a highly virulent strain in rabbits. However, the molecular determinants for its attenuation and protection remain unclear. In this study, we identified a unique glycosylation at position 986 (986NYT988) on the E2 glycoprotein Domain IV of C-strain but not (986NYA988) the highly virulent CSFV Shimen strain. We evaluated the infectivity, virulence, and protective efficacy of the C-strain-based mutant rHCLV-T988A lacking the glycosylation and Shimen strain mutant rShimen-A988T acquiring an additional glycosylation at position 986. rShimen-A988T showed a significantly decreased viral replication ability in SK6 cells, while rHCLV-T988A exhibited a growth kinetics indistinguishable from that of C-strain. Removal of the C-strain glycosylation site does not affect viral replication in rabbits and the attenuated phenotype in pigs. However, rShimen-A988T was attenuated and protected the pigs from a lethal challenge at 14 days postinoculation. In contrast, the rHCLV-T988A-inoculated pigs showed transient fever, a few clinical signs, and pathological changes in the spleens upon challenge with the Shimen strain. Mechanistic investigations revealed that the unique glycosylation at position 986 influences viral spreading, alters the formation of E2 homodimers, and leads to increased production of neutralizing antibodies. Collectively, our data for the first time demonstrate that the unique glycosylation at position 986 on the E2 glycoprotein is responsible for viral attenuation and protection. IMPORTANCE Viral glycoproteins involve in infectivity, virulence, and host immune responses. Deglycosylation on the Erns, E1, or E2 glycoprotein of highly virulent classical swine fever virus (CSFV) attenuated viral virulence in pigs, indicating that the glycosylation contributes to the pathogenicity of the highly virulent strain. However, the effects of the glycosylation on the C-strain E2 glycoprotein on viral infectivity in cells, viral attenuation, and protection in pigs have not been elucidated. This study demonstrates the unique glycosylation at position 986 on the C-strain E2 glycoprotein. C-strain mutant removing the glycosylation at the site provides only partial protection against CSFV challenge. Remarkably, the addition of the glycan to E2 of the highly virulent Shimen strain attenuates the viral virulence and confers complete protection against the lethal challenge in pigs. Our findings provide a new insight into the contribution of the glycosylation to the virus attenuation and protection.


Assuntos
Vírus da Febre Suína Clássica/imunologia , Vírus da Febre Suína Clássica/patogenicidade , Peste Suína Clássica/prevenção & controle , Proteínas do Envelope Viral/metabolismo , Vacinas Virais/imunologia , Animais , Anticorpos Neutralizantes/sangue , Peste Suína Clássica/virologia , Vírus da Febre Suína Clássica/genética , Vírus da Febre Suína Clássica/metabolismo , Glicosilação , Imunização/veterinária , Mutação , Multimerização Proteica , Coelhos , Suínos , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , Vacinas Atenuadas/metabolismo , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/imunologia , Vacinas Virais/administração & dosagem , Vacinas Virais/genética , Vacinas Virais/metabolismo , Virulência , Replicação Viral
20.
Vet Immunol Immunopathol ; 243: 110365, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34920287

RESUMO

Aujeszky's disease (AD) is a viral infectious disease caused by Suid herpesvirus 1 (SuHV-1). Vaccination and eradication of AD in domestic pigs is possible using marker vaccines with attenuated or inactivated SuHV-1, or subunit vaccines. However, vaccines with attenuated SuHV-1 have shown to be more potent in inducing strong cell-mediated immune response. The studies have shown that Parapoxvirus ovis, as well as Propionibacterium granulosum with lipopolysacharides (LPS) of Escherichia coli have pronounced immunomodulatory effects and that in combination with the vaccines can induce stronger humoral and cellular immune responses than use of vaccines alone. In our study distribution of peripheral blood T cell subpopulations was analysed after administration of vaccine alone (attenuated SuHV-1), immunostimulators (inactivated Parapoxvirus ovis or combination of an inactivated P. granulosum and detoxified LPS of E. coli) and combinations of vaccine with each immunostimulator to the 12-week old piglets. Throughout the study no significant changes were found in the proportions of γδ and most αß T cell subpopulations analysed. However, on the seventh day of the study combination of an inactivated P. granulosum and LPS of E. coli with vaccine induced transient but significant increase of the proportions of CD4+CD8α+ and CD4-CD8α+ αß T cells, that have been strongly associated with early protection of SuHV-1 infected pigs. Our findings indicate that combination of inactivated P. granulosum and detoxified E. coli LPS could be used for enhancement of a cellular immune response induced by vaccines against AD.


Assuntos
Adjuvantes Imunológicos/farmacologia , Pseudorraiva , Doenças dos Suínos , Linfócitos T/efeitos dos fármacos , Vacinas Virais , Animais , Anticorpos Antivirais , Escherichia coli , Herpesvirus Suídeo 1/imunologia , Imunidade Celular , Lipopolissacarídeos , Pseudorraiva/prevenção & controle , Suínos , Doenças dos Suínos/prevenção & controle , Vacinação/veterinária , Vacinas Atenuadas/imunologia , Vacinas Virais/imunologia
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