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BMC Research Notes Jul 2018Infection of chickens with low pathogenic avian influenza virus, such as H9N2 virus, culminates in decreased egg production and increased mortality and morbidity if...
OBJECTIVE
Infection of chickens with low pathogenic avian influenza virus, such as H9N2 virus, culminates in decreased egg production and increased mortality and morbidity if co-infection with other respiratory pathogens occurs. We have previously observed the induction of antibody- and cell-mediated immune responses after intramuscular administration of an H9N2 beta-propiolactone inactivated virus vaccine to chickens. Given the fact that in ovo vaccination represents a practical option for vaccination against H9N2 AIV in chickens, in the current study, we set out to characterize immune responses in chickens against a beta-propiolactone inactivated H9N2 virus vaccine after primary vaccination in ovo on embryonic day 18, and secondary intramuscular vaccination on day 14 post-hatch. We also included the Toll-like receptor 21 ligand, CpG ODN 2007, and an oil emulsion adjuvant, AddaVax™, as adjuvants for the vaccines.
RESULTS
Antibody-mediated immune responses were observed after administering the secondary intramuscular vaccine. Cell-mediated immune responses were observed in chickens that received the beta-propiolactone inactivated H9N2 virus combined with AddaVax™. Our results demonstrate that adaptive immune responses can be induced in chickens after a primary in ovo vaccination and secondary intramuscular vaccination.
Topics: Animals; Antibodies, Viral; Antibody Formation; Chickens; Influenza A Virus, H9N2 Subtype; Influenza Vaccines; Influenza in Birds
PubMed: 29970157
DOI: 10.1186/s13104-018-3537-9 -
Genetics Dec 1974Two genes, rad6 and rad9, that confer radiation sensitivity in the yeast Saccharomyces cerevisiae also greatly reduce the frequency of chemically-induced reversions of a...
Two genes, rad6 and rad9, that confer radiation sensitivity in the yeast Saccharomyces cerevisiae also greatly reduce the frequency of chemically-induced reversions of a tester mutant cyc1-131, which is a chain initiation mutant in the structural gene determining iso-1-cytochrome c. Mutations induced by ethyl methanesulfonate (EMS), diethyl sulfate (DES), methyl methanesulfonate (MMS), dimethyl sulfate (DMS), nitroquinoline oxide (NQO), nitrosoguanidine (NTG), nitrogen mustard (HN2), beta-propiolactone, and tritiated uridine, as well as mutations induced by ultraviolet light (UV) and ionizing radiation were greatly diminished in strains homozygous for either the rad6 or rad9 gene. Nitrous acid and nitrosoimidazolidone (NIL), on the other hand, were highly mutagenic in these repair-deficient mutants, and at low doses, these mutagens acted with about the same efficiency as in the normal RAD strain. At high doses of either nitrous acid or NIL, however, reversion frequencies were significantly reduced in the two rad mutants compared to normal strains. Although both rad mutants are immutable to about the same extent, the rad9 strains tend to be less sensitive to the lethal effect of chemical mutagens than rad6 strains. It is concluded that yeast requires a functional repair system for mutation induction by chemical agents.
Topics: Cytochrome c Group; DNA Repair; DNA Replication; Drug Resistance, Microbial; Genes; Mechlorethamine; Mesylates; Mutagens; Mutation; Nitrosoguanidines; Nitrous Acid; Peptide Chain Initiation, Translational; Quinolines; Radiation Genetics; Saccharomyces cerevisiae; Sulfates; Tritium; Uridine
PubMed: 4376097
DOI: 10.1093/genetics/78.4.1101 -
Journal of Pharmaceutical Analysis Dec 2018A simple method was established for the determination of -propiolactone (BPL) in human inactivated rabies vaccine by gas chromatography-mass spectrometry (GC-MS). The...
A simple method was established for the determination of -propiolactone (BPL) in human inactivated rabies vaccine by gas chromatography-mass spectrometry (GC-MS). The determination was performed on an Agilent HP-INNOWAX (30 m × 0.32 mm i.d., 0.25 µm) capillary column at the temperature of 80 °C. Electrospray ionization (ESI) was used by selective ion detection at 42. The temperature for ESI source and inlet was set at 230 °C and 200 °C, respectively. Helium was used as the carrier gas at a flow rate of 25.1 mL/min. The total run time was 8 min. Acetonitrile and other components in the sample did not interfere with the determination of BPL. The results showed good linearity of BPL in the range of 0.50-10.01 μg/mL, with the limit of detection and the limit of quantification of 0.015 μg/mL and 0.050 μg/mL, respectively. Satisfactory precision was achieved for the current developed method. The method was applied to detect 6 batches of vaccine samples, and the results indicated that the target analyte BPL was present in three batches of unpurified samples, but was not detected in the purified samples, indicating the test samples were qualified. The established method was proved to be simple, versatile and sensitive, which can meet the requirements of quality control of BPL in human inactivated rabies vaccine.
PubMed: 30595943
DOI: 10.1016/j.jpha.2018.06.003 -
The Lancet. Rheumatology Feb 2022We aimed to examine the immunogenicity pattern induced by the inactivated SARS-CoV-2 vaccine CoronaVac (Sinovac Life Sciences, Beijing, China) in SARS-CoV-2 seropositive...
Immunogenicity and safety of two doses of the CoronaVac SARS-CoV-2 vaccine in SARS-CoV-2 seropositive and seronegative patients with autoimmune rheumatic diseases in Brazil: a subgroup analysis of a phase 4 prospective study.
BACKGROUND
We aimed to examine the immunogenicity pattern induced by the inactivated SARS-CoV-2 vaccine CoronaVac (Sinovac Life Sciences, Beijing, China) in SARS-CoV-2 seropositive patients with autoimmune rheumatic diseases compared with seropositive controls, seronegative patients with autoimmune rheumatic diseases, and seronegative controls.
METHODS
CoronavRheum is an ongoing, prospective, controlled, phase 4 study, in which patients aged 18 years or older with autoimmune rheumatic diseases, and healthy controls were recruited from a single site (Rheumatology Division of Hospital das Clínicas, Faculdade de Medicina da Universidade de São Paulo) in São Paulo, Brazil Participants were vaccinated with two doses of CoronaVac (intramuscular injection, 3 μg in 0·5 mL of β-propiolactone inactivated SARS-CoV-2) on day 0 and on day 28. Blood samples were taken pre-vaccination on day 0, day 28, and also on day 69. For this subgroup analysis, participants were defined as being SARS-CoV-2 seropositive or seronegative prevaccination via anti-SARS-CoV-2 spike (S)1 or S2 IgG (cutoff of 15·0 arbitrary units [AU] per mL) or neutralising antibody titres (cutoff of ≥30%) and were matched for age and sex, via convenience sampling, in a 1:3:1:1 ratio (seropositive patients to seronegative patients to seropositive controls to seronegative controls). The primary outcomes were rates of anti-SARS-CoV-2 S1 and S2 IgG seropositivity and SARS-CoV-2 neutralising antibody positivity at day 28 and day 69 and immunogenicity dynamics assessed by geometric mean titres (GMTs) of IgG and median neutralising activity in seropositive patients with autoimmune rheumatic diseases compared with seronegative patients and seropositive and seronegative controls. We assessed safety in all participants randomly selected for this subgroup analysis. This study is registered with ClinicalTrials.gov, NCT04754698, and is ongoing for long-term immunogenicity evaluation.
FINDINGS
Between Feb 4 and Feb 8, 2021, 1418 patients and 542 controls were recruited, of whom 1685 received two vaccinations (1193 patients and 492 controls). After random sampling, our immunogenicity analysis population comprised 942 participants, of whom 157 were SARS-CoV-2 seropositive patients with autoimmune rheumatic diseases, 157 were seropositive controls, 471 were seronegative patients, and 157 were seronegative controls; the median age was 48 years (IQR 38-56) and 594 (63%) were female and 348 (37%) were male. For seropositive patients and controls, an increase in anti-SARS-CoV-2 S1 and S2 IgG titres (seropositive patients GMT 52·3 [95% CI 42·9-63·9] at day 0 128·9 [105·6-157·4] at day 28; seropositive controls 53·3 [45·4-62·5] at day 0 202·0 [174·8-233·4] at day 28) and neutralising antibody activity (seropositive patients 59% [IQR 39-83] at day 0 82% [54-96] at day 28; seropositive controls 58% [41-79] at day 0 92% [79-96] at day 28), was observed from day 0 to day 28, without further increases from day 28 to day 69 (at day 69 seropositive patients' GMT was 137·1 [116·2-161·9] and neutralising antibody activity was 79% [57-94]); and seropositive controls' GMT was 188·6 [167·4-212·6] and neutralising antibody activity was 92% [75-96]). By contrast, for seronegative patients and controls, the second dose was required for maximum response at day 69, which was lower in seronegative patients than in seronegative controls. GMTs in seronegative patients were 2·3 (95% CI 2·2-2·3) at day 0, 5·7 (5·1-6·4) at day 28, and 29·6 (26·4-33·3) at day 69, and in seronegative controls were 2·3 (2·1-2·5) at day 0, 10·6 (8·7-13·1) at day 28, and 71·7 (63·5-81·0) at day 69; neutralising antibody activity in seronegative patients was 15% (IQR 15-15) on day 0, 15% (15-15) at day 28, and 39% (15-65) at day 69, and in seronegative controls was 15% (15-15) at day 0, 24% (15-37) at day 28, and 61% (37-79) at day 69. Neither seronegative patients nor seronegative controls reached the GMT or antibody activity levels of seropositive patients at day 69.
INTERPRETATION
By contrast with seronegative patients with autoimmune rheumatic diseases, seropositive patients have a robust response after a single dose of CoronaVac. Our findings raise the possibility that the reduced immunogenicity observed in seronegative patients might not be the optimum response potential to SARS-CoV-2 vaccination, and therefore emphasise the importance of at least a single booster vaccination in these patients.
FUNDING
Fundação de Amparo à Pesquisa do Estado de São Paulo, Conselho Nacional de Desenvolvimento Científico e Tecnológico, and B3-Bolsa de Valores do Brasil.
TRANSLATION
For the Portuguese translation of the abstract see Supplementary Materials section.
PubMed: 34901885
DOI: 10.1016/S2665-9913(21)00327-1 -
Macromolecules Jun 2020We report here the synthesis of poly(4-ketovalerolactone) () via ring-opening transesterification polymerization (ROTEP) of the monomer 4-ketovalerolactone (, two steps...
We report here the synthesis of poly(4-ketovalerolactone) () via ring-opening transesterification polymerization (ROTEP) of the monomer 4-ketovalerolactone (, two steps from levulinic acid). The polymerization of proceeds to high equilibrium monomer conversion (up to 96% in the melt) to give the semicrystalline polyketoester with low dispersity. displays glass transition temperatures of 7 °C and two melting temperatures at 132 and 148 °C. This polyester can be chemically recycled through hydrolytic degradation. Under aqueous neutral or acidic conditions, the dominating pathway for polyester hydrolysis is through backbiting from the chain end. Under basic conditions, mid-chain cleavage, accelerated by the ketone carbonyl group in the backbone, promotes the hydrolysis of nearby backbone ester bonds. The final hydrolysis product is 5-hydroxylevulinic acid, the ring opened hydrolysis product of . was also observed to degrade under the action of a Brønsted acid to a bis-spirocyclic dilactone natural product altaicadispirolactone, which is a dimer of . This constitutes a rare example of a one-step synthesis of a secondary metabolite of non-trivial structure in which a polymer was the starting material and the sole source of matter. Analogous ROTEP of the isomeric 4-membered lactone 4-acetyl-β-propiolactone () was also explored, although this chemistry was not as well-behaved as the to polymerization.
PubMed: 33767514
DOI: 10.1021/acs.macromol.0c00787 -
The Biochemical Journal Feb 19681. The reactions of beta-propiolactone with amino acids were investigated under various conditions of pH and temperature to find those under which the reagent acted with...
1. The reactions of beta-propiolactone with amino acids were investigated under various conditions of pH and temperature to find those under which the reagent acted with specificity. 2. At pH9.0 and 22 degrees , after 15min. of reaction, at least 85% of each amino acid had reacted, methionine and cystine being the most reactive. 3. At pH7.0 and 22 degrees most amino acids reacted; methionine, cystine and histidine reacted almost entirely, and proline and lysine to a significantly smaller extent. 4. At pH3.0 and 22 degrees further specificity was obtained; methionine and cystine were the only reactive amino acids. 5. Reaction at pH3.0 and 0 degrees was specific for methionine; it was the only amino acid modified even after 145hr. of reaction.
Topics: Amino Acids; Autoanalysis; Chemical Phenomena; Chemistry; Cold Temperature; Cystine; Histidine; Hydrogen-Ion Concentration; Isoleucine; Lactones; Leucine; Lysine; Methionine; Phenylalanine; Proline; Tyrosine
PubMed: 5637366
DOI: 10.1042/bj1060829 -
International Journal of Infectious... Jan 2004The current recommended inactivating agent for the rabies virus, beta propiolactone (BPL) is very expensive and potentially carcinogenic. There is a need to evaluate...
OBJECTIVE
The current recommended inactivating agent for the rabies virus, beta propiolactone (BPL) is very expensive and potentially carcinogenic. There is a need to evaluate alternative chemicals, which will inactivate the virus without affecting its antigenicity. In this study the effect of ascorbic acid on the infectivity of the rabies virus has been investigated.
METHOD
Vero cell grown fixed rabies virus CVS strain was treated with 0.1 mg/ml, 0.5 mg/ml and 1mg/ml final concentrations of ascorbic acid and 5 microg/ml of copper sulfate and kept at 4 degrees C along with untreated virus material. Each aliquot was titrated after various intervals for viral infectivity using both mice inoculation and titration in vero cells. The antigenicity of the virus material was determined by antibody induction in mice and modified NIH tests in parallel with virus material inactivated with a 1:4000 concentration of BPL.
RESULTS
An optimal concentration of 0.5 mg/ml of ascorbic acid and 5 microg/ml of copper sulfate completely inactivated the virus after 72 hours. The inactivated virus retained good antigenicity and potency value, which was comparable with using BPL.
CONCLUSION
These findings suggest that ascorbic acid can be used as an inactivating agent for fixed rabies virus grown in cell culture particularly for the preparation of diagnostic reagents. Further studies are required to evaluate its effect on the cell associated virus, probable therapeutic potential and feasibility of replacing BPL in production of inactivated rabies vaccine.
Topics: Animals; Antibodies, Viral; Ascorbic Acid; Biological Assay; Chlorocebus aethiops; Copper Sulfate; Female; Mice; Rabies; Rabies Vaccines; Rabies virus; Vaccines, Inactivated; Vero Cells; Virus Activation; Virus Replication
PubMed: 14690777
DOI: 10.1016/j.ijid.2003.09.002 -
BioMed Research International 2019Inactivation of rabies virus is essential for rabies vaccine preparation where the inactivating compound that is currently recommended for rabies vaccine preparation is...
Inactivation of rabies virus is essential for rabies vaccine preparation where the inactivating compound that is currently recommended for rabies vaccine preparation is -propiolactone (-PL). This compound is considered better than phenol and formalin but it is expensive and potentially carcinogenic. Data revealed that Ascorbic acid (AA) with cupric ions could yield complete and irreversible inactivation of rabies virus without adversely affecting its antigenicity. Additionally, the results of testing the vaccine potency with the selected inactivating compounds were comparable (P<0.05), and ED was higher than the recommended World Health Organization (WHO) limits. The use of HemaGel (plasma substitute) for testing vaccine stabilization was compared with the currently used vaccine stabilizers (human albumin and lactose). HemaGel yielded better stability than the other tested stabilizers. Monitoring of cellular and humoral immune responses indicated that both the total IgG level against rabies vaccine and the IFN and IL5 levels obtained with the HemaGel-stabilized vaccines were higher than those obtained with human albumin- and lactose-stabilized vaccine candidates.
Topics: Albumins; Animals; Antibodies, Viral; Ascorbic Acid; Chlorocebus aethiops; Humans; Immunogenicity, Vaccine; Immunoglobulin G; Interferons; Interleukin-5; Lactose; Propiolactone; Rabies; Rabies Vaccines; Rabies virus; Vaccine Potency; Vero Cells
PubMed: 31008105
DOI: 10.1155/2019/4518163 -
PloS One 2013Antigen detection assays can play an important part in environmental surveillance and diagnostics for emerging threats. We are interested in accelerating assay...
BACKGROUND
Antigen detection assays can play an important part in environmental surveillance and diagnostics for emerging threats. We are interested in accelerating assay formulation; targeting the agents themselves to bypass requirements for a priori genome information or surrogates. Previously, using in vitro affinity reagent selection on Marburg virus we rapidly established monoclonal affinity reagent sandwich assay (MARSA) where one recombinant antibody clone was both captor and tracer for polyvalent nucleoprotein (NP). Hypothesizing that the closely related Ebolavirus genus may share the same Achilles' heel, we redirected the scheme to see whether similar assays could be delivered and began to explore their mechanism.
METHODS AND FINDINGS
In parallel we selected panels of llama single domain antibodies (sdAb) from a semi-synthetic library against Zaire, Sudan, Ivory Coast, and Reston Ebola viruses. Each could perform as both captor and tracer in the same antigen sandwich capture assay thereby forming MARSAs. All sdAb were specific for NP and those tested required the C-terminal domain for recognition. Several clones were cross-reactive, indicating epitope conservation across the Ebolavirus genus. Analysis of two immune shark sdAb revealed they also targeted the C-terminal domain, and could be similarly employed, yet were less sensitive than a comparable llama sdAb despite stemming from immune selections.
CONCLUSIONS
The C-terminal domain of Ebolavirus NP is a strong attractant for antibodies and enables sensitive sandwich immunoassays to be rapidly generated using a single antibody clone. The polyvalent nature of nucleocapsid borne NP and display of the C-terminal region likely serves as a bountiful affinity sink during selections, and a highly avid target for subsequent immunoassay capture. Combined with the high degree of amino acid conservation through 37 years and across wide geographies, this domain makes an ideal handle for monoclonal affinity reagent driven antigen sandwich assays for the Ebolavirus genus.
Topics: Amino Acid Sequence; Antibody Specificity; Antigens, Viral; Ebolavirus; Enzyme-Linked Immunosorbent Assay; Gamma Rays; Molecular Sequence Data; Nucleoproteins; Propiolactone; Protein Structure, Tertiary; Single-Domain Antibodies; Viral Proteins; Virus Inactivation
PubMed: 23577211
DOI: 10.1371/journal.pone.0061232 -
Viruses Nov 2016Inactivated vaccines are commonly produced by incubating pathogens with chemicals such as formaldehyde or β-propiolactone. This is a time-consuming process, the...
Inactivated vaccines are commonly produced by incubating pathogens with chemicals such as formaldehyde or β-propiolactone. This is a time-consuming process, the inactivation efficiency displays high variability and extensive downstream procedures are often required. Moreover, application of chemicals alters the antigenic components of the viruses or bacteria, resulting in reduced antibody specificity and therefore stimulation of a less effective immune response. An alternative method for inactivation of pathogens is ionizing radiation. It acts very fast and predominantly damages nucleic acids, conserving most of the antigenic structures. However, currently used irradiation technologies (mostly gamma-rays and high energy electrons) require large and complex shielding constructions to protect the environment from radioactivity or X-rays generated during the process. This excludes them from direct integration into biological production facilities. Here, low-energy electron irradiation (LEEI) is presented as an alternative inactivation method for pathogens in liquid solutions. LEEI can be used in normal laboratories, including good manufacturing practice (GMP)- or high biosafety level (BSL)-environments, as only minor shielding is necessary. We show that LEEI efficiently inactivates different viruses (influenza A (H3N8), porcine reproductive and respiratory syndrome virus (PRRSV), equine herpesvirus 1 (EHV-1)) and bacteria () and maintains their antigenicity. Moreover, LEEI-inactivated influenza A viruses elicit protective immune responses in animals, as analyzed by virus neutralization assays and viral load determination upon challenge. These results have implications for novel ways of developing and manufacturing inactivated vaccines with improved efficacy.
Topics: Antigens, Bacterial; Antigens, Viral; Disinfection; Electrons; Escherichia coli; Radiation, Ionizing; Vaccines, Inactivated; Viruses
PubMed: 27886076
DOI: 10.3390/v8110319