-
Molecular Immunology Jul 2022Viral inactivation for antibody induction purposes, among other applications, should ensure biosafety, completely avoiding the risk of infectivity, and preserving viral...
Viral inactivation for antibody induction purposes, among other applications, should ensure biosafety, completely avoiding the risk of infectivity, and preserving viral immunogenicity. β-propiolactone (BPL) is one of the most used reagents for viral inactivation, despite its high toxicity and recent difficulties related to importation, experienced in Brazil during the SARS-CoV-2 pandemic. In this context, the main objectives of this work were to test different inactivation procedures for SARS-CoV-2 and to evaluate the induction of neutralizing antibodies in mice immunized with antigenic preparations obtained after viral treatment with formaldehyde (FDE), glutaraldehyde (GDE), peroxide hydrogen (HO), as well as with viral proteins extract (VPE), in parallel with BPL. Verification of viral inactivation was performed by subsequent incubations of the inactivated virus in Vero cells, followed by cytopathic effect and lysis plaques observation, as well as by quantification of RNA load using reverse transcription-quantitative real time polymerase chain reaction. Once viral inactivation was confirmed, cell culture supernatants were concentrated and purified. In addition, an aliquot inactivated by BPL was also subjected to viral protein extraction (VPE). The different antigens were prepared using a previously developed microemulsion as adjuvant, and were administered in a four-dose immunization protocol. Antibody production was comparatively evaluated by ELISA and Plaque Reduction Neutralization Tests (PRNT). All immunogens evaluated showed some level of IgG anti-SARS-CoV-2 antibodies in the ELISA assay, with the highest levels presented by the group immunized with FDE-inactivated viral antigen. In the PRNT results, except for VPE-antigen, all other immunogens evaluated induced some level of neutralizing anti-SARS-CoV-2 antibodies, and the FDE-antigen stood out again with the most expressive values. Taken together, the present work shows that FDE can be an efficient and affordable alternative to BPL for the production of inactivated SARS-CoV-2 viral antigen.
Topics: Animals; Antibodies, Viral; Antigens, Viral; COVID-19; Chlorocebus aethiops; Disease Models, Animal; Hydrogen Peroxide; Mice; SARS-CoV-2; Vero Cells
PubMed: 35644072
DOI: 10.1016/j.molimm.2022.05.012 -
Journal of Extracellular Vesicles Dec 2022The interaction of SARS-CoV-2 infection with extracellular vesicles (EVs) is of particular interest at the moment. Studying SARS-CoV-2 contaminated-EV isolates in...
The interaction of SARS-CoV-2 infection with extracellular vesicles (EVs) is of particular interest at the moment. Studying SARS-CoV-2 contaminated-EV isolates in instruments located outside of the biosafety level-3 (BSL-3) environment requires knowing how viral inactivation methods affect the structure and function of extracellular vesicles (EVs). Therefore, three common viral inactivation methods, ultraviolet-C (UVC; 1350 mJ/cm ), β-propiolactone (BPL; 0.005%), heat (56°C, 45 min) were performed on defined EV particles and their proteins, RNAs, and function. Small EVs were isolated from the supernatant of SARS-CoV-2-infected human lung epithelial Calu-3 cells by stepwise centrifugation, ultrafiltration and qEV size-exclusion chromatography. The EV isolates contained SARS-CoV-2. UVC, BPL and heat completely abolished SARS-CoV-2 infectivity of the contaminated EVs. Particle detection by electron microscopy and nanoparticle tracking was less affected by UVC and BPL than heat treatment. Western blot analysis of EV markers was not affected by any of these three methods. UVC reduced SARS-CoV-2 spike detectability by quantitative RT-PCR and slightly altered EV-derived β-actin detection. Fibroblast migration-wound healing activity of the SARS-CoV-2 contaminated-EV isolate was only retained after UVC treatment. In conclusion, specific viral inactivation methods are compatible with specific measures in SARS-CoV-2 contaminated-EV isolates. UVC treatment seems preferable for studying functions of EVs released from SARS-CoV-2 infected cells.
Topics: Humans; SARS-CoV-2; Virus Inactivation; COVID-19; Extracellular Vesicles; Lung; Epithelial Cells
PubMed: 36468940
DOI: 10.1002/jev2.12291 -
Journal of Clinical Microbiology Jan 1976The replication of seven rabies virus strains (CVS, HEP, PV, ERA, WIRAB, CPZ and BOLIVAR) in BHK cells and the inactivation dynamics of these strains by... (Comparative Study)
Comparative Study
The replication of seven rabies virus strains (CVS, HEP, PV, ERA, WIRAB, CPZ and BOLIVAR) in BHK cells and the inactivation dynamics of these strains by beta-propiolactone, acetylethylenimine, and ethylenimine were studied to find the most immunogenic strain and the most economic and stable inactivating agent for the production of an inactivated tissue culture rabies vaccine for animal use. The seven strains reached the peak of virus production 3 to 5 days after inoculation of the cell culture; PV yielded the highest virus titer (10(9) plaque-forming units/ml). The infectivity of virus suspensions containing 10(7) to 10(8) plaque-forming units/0.1 ml was inactivated by beta-propiolactone in 0.5 h, acetylethylenimine in 3.0 h, and ethylenimine in 1.0 h. Most of the vaccine lots prepared with the different strains and inactivating agents passed a modified National Institutes of Health potency test. The vaccines prepared with the PV strain had consistently higher antigenic values (equal or better than four) than the other six strains. This difference was highly significant (F6,12=59.8), whereas there were no statistically significant differences among the antigenic values of the vaccine lots prepared with the three inactivating agents. Batches of lyophilized and liquid vaccine stored at 4 C maintained potency for over 1 year. Ten dogs vaccinated with a vaccine prepared with the PV strain and inactivated with ethylenimine developed a good antibody response and resisted challenge 60 days after vaccination, while seven of eight nonvaccinated controls died of rabies. This information indicates that an inactivated, stable, economic, and easy-to-prepare rabies vaccine can be produced in BHK cells by using the PV strain and ethylenimine as an inactivating agent.
Topics: Animals; Antibody Formation; Aziridines; Culture Techniques; Dogs; Immunization; Mice; Propiolactone; Rabies virus; Virus Replication
PubMed: 1254701
DOI: 10.1128/jcm.3.1.26-33.1976 -
The Journal of Hygiene Jun 1970Vaccines were prepared from a single pool of high-titred vaccinia virus and inactivated by six methods, namely heat, formalin, hydroxylamine, beta-propiolactone,...
Vaccines were prepared from a single pool of high-titred vaccinia virus and inactivated by six methods, namely heat, formalin, hydroxylamine, beta-propiolactone, ultraviolet irradiation, and visible light and methylene blue. Large doses of the vaccines were required to protect mice against intracerebral challenge. Differences in protection were not attributable to the method of their inactivation. The vaccines also induced similar degrees of skin immunity in rabbits which showed no severe dermal reactions when challenged with either homologous killed vaccine or live virus. The virus-neutralizing, haemagglutinin-inhibiting and complement fixing antibody responses to the vaccines differed; heat-inactivation preserved these antigens least well and beta-propiolactone apparently the best. In both rabbits and mice there was little association between the different antibody responses to each vaccine or between the degrees of antibody response and the protection they induced. The relation of these findings to pox-virus immunity and the use of inactivated smallpox vaccine in man is discussed.
Topics: Animals; Antibody Formation; Chick Embryo; Complement Fixation Tests; Formaldehyde; Hemagglutination Inhibition Tests; Heterocyclic Compounds; Hot Temperature; Hydroxylamines; Immunity; Immunization; Immunodiffusion; Interferons; Lactones; Light; Male; Methods; Methylene Blue; Mice; Neutralization Tests; Rabbits; Skin Tests; Smallpox Vaccine; Ultraviolet Rays; Vaccinia virus
PubMed: 4988047
DOI: 10.1017/s0022172400028679 -
Food and Environmental Virology Mar 2020Influenza A virus (IAV) infection is perennially one of the leading causes of death worldwide. Effective therapy and vaccination are needed to control viral expansion....
Influenza A virus (IAV) infection is perennially one of the leading causes of death worldwide. Effective therapy and vaccination are needed to control viral expansion. However, current anti-IAV drugs risk inducing drug-resistant virus emergence. Although intranasal administration of whole inactivated virus vaccine can induce efficient protective immunity, formalin and β-propiolactone are the currently used and harmful inactivating agents. Here, we analyzed the antiviral activity of hibiscus (Hibiscus sabdariffa L.) tea extract against human IAV and evaluated its potential as a novel anti-IAV drug and a safe inactivating agent for whole inactivated vaccine. The in vitro study revealed that the pH of hibiscus tea extract is acidic, and its rapid and potent antiviral activity relied largely on the acidic pH. Furthermore, the mouse study showed that the acidic extract was not effective for either therapeutic or vaccination purposes. However, hibiscus tea extract and protocatechuic acid, one of the major components of the extract, showed not only potent acid-dependent antiviral activity but also weak low-pH-independent activity. The low-pH-independent activity did not affect the conformation of immunodominant hemagglutinin protein. Although this low-pH-independent activity is very limited, it may be suitable for the application to medication and vaccination because this activity is not affected by the neutral blood environment and does not lose antigenicity of hemagglutinin. Further study of the low-pH-independent antiviral mechanism and attempts to enhance the antiviral activity may establish a novel anti-IAV therapy and vaccination strategy.
Topics: Animals; Antiviral Agents; Female; Hibiscus; Humans; Hydrogen-Ion Concentration; Influenza A virus; Influenza, Human; Mice; Mice, Inbred BALB C; Plant Extracts
PubMed: 31620998
DOI: 10.1007/s12560-019-09408-x -
Vaccines Apr 2022Seneca Valley virus (SVV), also known as Senecavirus A (SVA), is a non-enveloped and single-strand positive-sense RNA virus, which belongs to the genus of within the...
Evaluation of Immunoreactivity and Protection Efficacy of Seneca Valley Virus Inactivated Vaccine in Finishing Pigs Based on Screening of Inactivated Agents and Adjuvants.
Seneca Valley virus (SVV), also known as Senecavirus A (SVA), is a non-enveloped and single-strand positive-sense RNA virus, which belongs to the genus of within the family . Porcine idiopathic vesicular disease (PIVD) caused by SVV has frequently been prevalent in America and Southeast Asia (especially in China) since the end of 2014, and has caused continuing issues. In this study, an SVV strain isolated in China, named SVV LNSY01-2017 (MH064435), was used as the stock virus for the preparation of an SVV-inactivated vaccine. The SVV culture was directly inactivated using binary ethyleneimine (BEI) and β-propiolactone (BPL). BPL showed a better effect as an SVV inactivator, according to the results of pH variation, inactivation kinetics, and the detection of VP1 content during inactivation. Then, SVV inactivated by BPL was subsequently emulsified using different adjuvants, including MONTANIDE ISA 201 VG (ISA 201) and MONTANIDE IMG 1313 VG N (IMS 1313). The immunoreactivity and protection efficacy of the inactivated vaccines were then evaluated in finishing pigs. SVV-BPL-1313 showed a better humoral response post-immunization and further challenge tests post-immunization showed that both the SVV-BPL-201 and SVV-BPL-1313 combinations could resist challenge from a virulent SVV strain. The SVV LNSY01-2017-inactivated vaccine candidate developed here represents a promising alternative to prevent and control SVV infection in swine.
PubMed: 35455380
DOI: 10.3390/vaccines10040631 -
Revue Scientifique Et Technique... Jun 1991Results are presented for the large-scale use of a tissue vaccine, inactivated with beta-propiolactone and containing aluminium hydroxide adjuvant, against viral... (Review)
Review
Results are presented for the large-scale use of a tissue vaccine, inactivated with beta-propiolactone and containing aluminium hydroxide adjuvant, against viral haemorrhagic disease of rabbits. The kinetics of haemagglutination-inhibiting antibodies over eighteen months, the response of vaccinated animals to challenge infection with field virus between six and fifteen months after vaccination, the serological response to revaccination, and the immunological mechanisms involved in primary vaccination and revaccination were investigated.
Topics: Animals; Antibodies, Viral; Hepatitis, Viral, Animal; Immunization, Secondary; Rabbits; Vaccination; Viral Vaccines
PubMed: 1760586
DOI: No ID Found -
Structure (London, England : 1993) Nov 2020The ongoing global pandemic of coronavirus disease 2019 (COVID-19) resulted from the outbreak of SARS-CoV-2 in December 2019. Currently, multiple efforts are being made...
The ongoing global pandemic of coronavirus disease 2019 (COVID-19) resulted from the outbreak of SARS-CoV-2 in December 2019. Currently, multiple efforts are being made to rapidly develop vaccines and treatments to fight COVID-19. Current vaccine candidates use inactivated SARS-CoV-2 viruses; therefore, it is important to understand the architecture of inactivated SARS-CoV-2. We have genetically and structurally characterized β-propiolactone-inactivated viruses from a propagated and purified clinical strain of SARS-CoV-2. We observed that the virus particles are roughly spherical or moderately pleiomorphic. Although a small fraction of prefusion spikes are found, most spikes appear nail shaped, thus resembling a postfusion state, where the S1 protein of the spike has disassociated from S2. Cryoelectron tomography and subtomogram averaging of these spikes yielded a density map that closely matches the overall structure of the SARS-CoV postfusion spike and its corresponding glycosylation site. Our findings have major implications for SARS-CoV-2 vaccine design, especially those using inactivated viruses.
Topics: Animals; Betacoronavirus; COVID-19 Vaccines; Chlorocebus aethiops; Coronavirus Infections; Cryoelectron Microscopy; Disinfectants; Electron Microscope Tomography; Humans; Propiolactone; SARS-CoV-2; Spike Glycoprotein, Coronavirus; Vaccines, Inactivated; Vero Cells; Viral Vaccines; Virion
PubMed: 33058760
DOI: 10.1016/j.str.2020.10.001 -
Viruses Jan 2018In recent years, enterovirus D68 (EVD68) has been reported increasingly to be associated with severe respiratory tract infections and acute flaccid myelitis (AFM) in...
In recent years, enterovirus D68 (EVD68) has been reported increasingly to be associated with severe respiratory tract infections and acute flaccid myelitis (AFM) in children all over the world. Yet, no effective vaccines or antiviral drugs are currently available for EVD68. Although several experimental animal models have been developed, immunogenicity and protective efficacy of inactivated EVD68 vaccines has not been fully evaluated. To promote the development of vaccines, we established an Institute of Cancer Research (ICR) suckling mouse model of EVD68 infection in this study. The results showed that ICR neonatal mice up to about nine days of age were susceptible to infection with EVD68 clinical strain US/MO/14-18947 by intraperitoneal injection. The infected mice exhibited progressive limb paralysis prior to death and the mortality of mice was age- and virus dose-dependent. Tissue viral load analysis showed that limb muscle and spinal cord were the major sites of viral replication. Moreover, histopathologic examination revealed the severe necrosis of the limb and juxtaspinal muscles, suggesting that US/MO/14-18947 has a strong tropism toward muscle tissues. Additionally, β-propiolactone-inactivated EVD68 vaccine showed high purity and quality and induced robust EVD68-specific neutralizing antibody responses in adult mice. Importantly, results from both antisera transfer and maternal immunization experiments clearly showed that inactivated EVD68 vaccine was able to protect against lethal viral infection in the mouse model. In short, these results demonstrate the successful establishment of the mouse model of EVD68 infection for evaluating candidate vaccines against EVD68 and also provide important information for the development of inactivated virus-based EVD68 vaccines.
Topics: Animals; Animals, Newborn; Antibodies, Neutralizing; Cell Line, Tumor; Disease Models, Animal; Enterovirus D, Human; Enterovirus Infections; Female; Humans; Immunity, Maternally-Acquired; Mice; Mice, Inbred ICR; Neutralization Tests; Vaccines, Inactivated; Viral Load; Viral Tropism; Viral Vaccines
PubMed: 29385753
DOI: 10.3390/v10020058 -
The New England Journal of Medicine Apr 2011Yellow fever is a lethal viral hemorrhagic fever occurring in Africa and South America. A highly effective live vaccine (17D) is widely used for travelers to and...
BACKGROUND
Yellow fever is a lethal viral hemorrhagic fever occurring in Africa and South America. A highly effective live vaccine (17D) is widely used for travelers to and residents of areas in which yellow fever is endemic, but the vaccine can cause serious adverse events, including viscerotropic disease, which is associated with a high rate of death. A safer, nonreplicating vaccine is needed.
METHODS
In a double-blind, placebo-controlled, dose-escalation, phase 1 study of 60 healthy subjects between 18 and 49 years of age, we investigated the safety and immunogenicity of XRX-001 purified whole-virus, β-propiolactone-inactivated yellow fever vaccine produced in Vero cell cultures and adsorbed to aluminum hydroxide (alum) adjuvant. On two visits 21 days apart, subjects received intramuscular injections of vaccine that contained 0.48 μg or 4.8 μg of antigen. Levels of neutralizing antibodies were measured at baseline and on days 21, 31, and 42.
RESULTS
The vaccine induced the development of neutralizing antibodies in 100% of subjects receiving 4.8 μg of antigen in each injection and in 88% of subjects receiving 0.48 μg of antigen in each injection. Antibody levels increased by day 10 after the second injection, at which time levels were significantly higher with the 4.8-μg formulation than with the 0.48-μg formulation (geometric mean titer, 146 vs. 39; P<0.001). Three adverse events occurred at a higher incidence in the two vaccine groups than in the placebo group: mild pain, tenderness, and (much less frequently) itching at the injection site. One case of urticaria was observed on day 3 after the second dose of 4.8 μg of vaccine.
CONCLUSIONS
A two-dose regimen of the XRX-001 vaccine, containing inactivated yellow fever antigen with an alum adjuvant, induced neutralizing antibodies in a high percentage of subjects. XRX-001 has the potential to be a safer alternative to live attenuated 17D vaccine. (Funded by Xcellerex; ClinicalTrials.gov number, NCT00995865.).
Topics: Adolescent; Adult; Animals; Antibodies, Neutralizing; Antibodies, Viral; Chlorocebus aethiops; Dose-Response Relationship, Drug; Double-Blind Method; Female; Humans; Injections, Intramuscular; Male; Middle Aged; Vaccines, Inactivated; Vero Cells; Yellow Fever; Yellow Fever Vaccine; Yellow fever virus; Young Adult
PubMed: 21470010
DOI: 10.1056/NEJMoa1009303