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The propagation of feline panleukopenia virus in micro-carrier cell culture and use of the inactivated virus in the protection of mink against viral enteritis.

Using microcarrier cell culture for the production of virus antigen, a formalin-inactivated feline panleukopenia virus vaccine was evaluated for protection of mink against specific mink enteritis virus infection. The vaccine showed a good immunogenic effect in mink when used either alone or in combination with Clostridium botulinum type C-toxoid and/or Pseudomonas aeruginosa vaccine. A single vaccination induced persistent immune responses for periods of at least 1 year, as evaluated by ELISA and challenge tests. Neither immunological interference between vaccine constituents nor adverse reactions were observed.

Animals↗

Detection of anti-human cell antibodies in sera from macaques immunized with whole inactivated virus.

More than 200 sera from macaques immunized with several different vaccine preparations were tested in various assays with cells of human and macaque origin. Only in instances where whole inactivated SIV preparations were used for immunization, were reactivities found with normal human cells, and this was the case in every instance. Such sera produced a marked clumping of several normal human cell lines and exhibited strong staining of the cell surface in FACS analysis. In the presence of SIVDeltaB670, these sera also enhanced infectivity and fusion formation. When similar tests were performed with macaque cells as targets, such phenomena were not easily discernible. Likewise, there was no trace of such activities in sera from normal animals, animals chronically infected with SIV, or in those from animals which received recombinant viral subunits as vaccines. Finally, we show that in several instances where whole inactivated virus was used as a vaccine, there is a strong correlation between the titer of anticellular activity with protection.

Animals↗

Intranasal immunization with formalin-inactivated virus vaccine induces a broad spectrum of heterosubtypic immunity against influenza A virus infection in mice.

It has been known that influenza A virus infection induces a cross-protective immunity against infection by viruses with different subtypes of viral envelope proteins, hemagglutinin (HA) and neuraminidase (NA). This heterosubtypic immunity is generally mediated by cytotoxic T lymphocytes (CTL) reactive to specific epitopes in the viral internal proteins, such as nucleoprotein and matrix protein. By contrast, immunization with inactivated virus antigens has been thought to be unable to generate heterosubtypic immunity, since inactivated antigens do not usually induce CTL responses. However, we show that intranasal immunization with formalin-inactivated intact virus, but not ether-split vaccines, induced a broad spectrum of heterosubtypic protective immunity in mice. The protection may be mediated by the mucosal immune response, most likely secretory IgA antibodies to the viral proteins. This approach may overcome limitations in the efficacy of inactivated influenza vaccines and confer potent immunity to humans against viruses with new pandemic potential.

Administration, Intranasal↗

Interactions between human neutrophils and vaccinia virus: induction of oxidative metabolism and virus inactivation.

The possible role of human neutrophils (PMN) as direct effector cells against free live virus particles was tested in vitro. Oxygen consumption, production of chemiluminescence and "virus killing" by PMNs was examined during and after incubation of cells and live or dead vaccinia virus with and without specific antibody. Oxygen consumption and chemiluminescence production occurred with unopsonized live but not with inactivated virus and was greatly enhanced with opsonization. Virus titers were determined in PMN-free supernatant and freeze-thawed cell pellet fractions after 15 min incubation. Reduction in titer from virus control was seen in supernatant fractions with and without opsonins, but in the cell fraction only with opsonins; the degree of reduction with opsonins was directly related to the opsonin source. PMNs from a patient with chronic granulomatous disease did not inhibit virus replication. These data suggest an active role for human PMNs in the genesis of and defense against virus infections and indicate areas for further investigation.

Antibodies, Viral↗

Neither whole inactivated virus immunogen nor passive immunoglobulin transfer protects against SIVagm infection in the African green monkey natural host.

Attempts to protect against infection with simian immunodeficiency virus (SIV)mac grown in rhesus peripheral blood mononuclear cells (PBMCs) using whole inactivated virus immunogen or passive transfer of antibody have so far universally failed. However, such experiments have succeeded in the closely related human immunodeficiency virus (HIV)-2/cynomolgus system. To determine whether the failure in the SIVmac system is typical of primate lentiviruses we performed vaccination and passive transfer experiments using SIVagm (genetically distinct from SIVmac and HIV-2) in the natural African green monkey (AGM) host. To maximize the chances of success, both immunogen and challenge material were prepared using the molecular SIVagm3 clone. Two AGMs were immunized four times with whole inactivated SIVagm3 in RIBI adjuvant and two received intravenously a high dose of immunoglobulin (Ig) purified from a mixture of plasma from AGM, either naturally infected or infected with the homologous SIVagm3 clone. All were challenged with 20 median minimum infective doses of the SIVagm3 grown in AGM PBMCs and previously titrated in AGMs. Despite a strong humoral immune response in all monkeys at the time of challenge, including measurable neutralizing and antibody-dependent cell-mediated cytotoxicity antibodies, none were protected from infection.

Amino Acid Sequence↗

Virus inactivation during intravenous immunoglobulin production.

Effects of time, temperature, pH and stabilizers (i.e. medium) on inactivation of lipid-enveloped model viruses, Semliki Forest and vesicular stomatitis viruses in the production process of intravenous immunoglobulin were investigated on a laboratory scale. The lowering of pH, the raising of temperature and the increasing of incubation time improved the inactivation effect. However, small changes in pH and stabilizer concentrations did not influence the results. Inactivation was not linear and a clear tailing off could be seen. Therefore, for complete virus inactivation incubation times longer than 20 h are necessary. Inactivation took place much more rapidly in intravenous immunoglobulin solution than in intramuscular immunoglobulin solution. Processing steps such as freeze-dying in the presence of ethanol or storage of intramuscular immunoglobulin in the liquid state at pH7 only partially inactivated these viruses.

Blood Transfusion↗

Virus inactivation procedures: an overview.

Viruses can easily be transmitted by intravenous infusion of contaminated blood or blood products. Reduction of viral transmission is achieved by suitable donor selection, screening of donations, and choice of manufacturing processes. The components of this strategy should not be considered individually but are complementary. An overview is presented of the combined strategy for reducing the likelihood of virus transmission via blood and blood products.

Antiviral Agents↗

Involvement of reactive oxygen species in hemoglobin oxidation and virus inactivation by 1,9-dimethylmethylene blue phototreatment.

The participation of reactive oxygen species (ROS) in virus inactivation by 1,9-dimethylmethylene blue (DMMB) phototreatment in stroma-free hemoglobin (SFH) was investigated with the use of scavengers, quenchers and enhancer. Virus (R17 bacteriophage) photoinactivation by either activated monomer or dimer DMMB was suppressed by sodium azide (singlet oxygen quencher) and promoted by the substitution of H2O for deuterium oxide (D2O), which is known to prolong the lifespan of singlet oxygen. There was no or little effect of mannitol (hydroxyl radical scavenger) and superoxide dismutase (superoxide scavenger) on the photoinactivation. Similar experiments were conducted to investigate the mechanism of methemoglobin (Met-Hb) formation by the activated monomer of DMMB. There was little effect of the singlet oxygen quencher, histidine, or the enhancer, D2O, on Met-Hb formation. However, rutin, which inhibits not only singlet oxygen but also other ROS, and mannitol supressed the formation of Met-Hb by activated monomer. The addition of superoxide dismutase (SOD) did not inhibit the formation. In contrast to the activity of the DMMB monomer, that of the dimer was inhibited by histidine and enhanced by D2O. The addition of neither mannitol nor SOD affected Met-Hb formation by activated dimer. These results collectively suggest that virus photoinactivation by the activated monomer and dimer of DMMB as well as Met-Hb formation by the activated dimer proceed via a singlet oxygen mediated pathway. In contrast, singlet oxygen may play a less important role in Met-Hb formation by the activated monomer.

Bacteriophages↗

Protective immune response of chickens against Newcastle disease, induced by the intranasal vaccination with inactivated virus.

Intranasal vaccination of chickens with inactivated Newcastle disease virus (NDV) induced both local and systemic antibody responses, resulting in protection against intranasal challenge with a lethal dose of a virulent NDV strain. The immune response was enhanced by the use of cholera toxin B subunit (CTB) as an adjuvant and only small amounts of the challenge virus were recovered from the birds vaccinated together with CTB. On the other hand, subcutaneous vaccination with the same antigen induced only a serum antibody response in chickens, allowing the challenge virus to replicate in the sinus. The present results indicate that secretory antibodies induced on the respiratory mucosal surface by intranasal vaccination with inactivated NDV protected chickens from lethal infection by inhibiting virus replication at the portal of entry for the virus.

Adjuvants, Immunologic↗

Comparative study of mechanisms of herpes simplex virus inactivation by sodium lauryl sulfate and n-lauroylsarcosine.

The mechanisms of herpes simplex virus (HSV) inactivation by sodium lauryl sulfate (SLS) and n-lauroylsarcosine (LS), two anionic surfactants with protein denaturant potency, have been evaluated in cultured cells. Results showed that pretreatment of HSV type 1 (HSV-1) strain F and HSV-2 strain 333 with either surfactant inhibited, in a concentration- and time-dependent manner, their infectivities on Vero cells. SLS was a more potent inhibitor of HSV-2 strain 333 infectivity than LS with respect to the concentration (4.8-fold lower) and time (2.4-fold shorter) required to completely inactivate the virus. No inhibition of both herpesvirus strains infectivities was observed when Vero cells were pretreated with either surfactant. LS prevented the binding of HSV-2 strain 333 to cells without affecting the stable attachment and the rate of penetration into cells, whereas SLS exerted the opposite effect. Both SLS and LS inhibited, in a concentration-dependent manner, the HSV-2 strain 333-induced cytopathic effect, probably by affecting newly synthesized virions that come into contact with surfactant molecules present in culture medium. The pretreatment of HSV-2 strain 333 with specific combinations of SLS and LS concentrations inhibited the viral infectivity in a synergistic manner and resulted in only a small increase in their toxicities for exponentially growing Vero cells compared with that caused by each compound alone. Taken together, these results suggest that SLS and LS, alone or combined, could represent potent candidates as microbicides in topical vaginal formulations to prevent the transmission of herpes and possibly other pathogens that cause sexually transmitted diseases, including human immunodeficiency virus type 1.

Animals↗

Clinical experience with Octagam, a solvent detergent (SD) virus inactivated intravenous gammaglobulin.

OBJECTIVE: The safety and efficacy of Octagam, a solvent detergent (SD) virus inactivated intravenous gammaglobulin, were evaluated in 54 patients who received 301 infusions of Octagam for a total amount of 9301 grams, during 24 months. METHODS: Adverse reactions during and after Octagam infusion were carefully recorded. Serum IgG and ALT levels were recorded monthly. Antibodies to HIV, HBV and HCV were recorded every 3 months. RESULTS: In 22 chronic lymphocytic leukemia patients and 4 multiple myeloma patients, the monthly infusion of 0.4 gr/ kg Octagam proved safe and effective in increasing IgG levels and potentially reducing the rate of serious bacterial infections. In 20 patients with ITP, infusion of 2 gr/kg of Octagam was safe and effective in increasing the platelet count and in arresting spontaneous bleeding or preventing bleeding prior to surgical procedures. In 8 patients with the antiphospholipid syndrome monthly infusions of Octagam were found to be safe. Adverse reactions following Octagam infusion were uncommon, generally mild and all were transient. Viral safety assays for HIV, HBV and HCV were all negative. CONCLUSION: Octagam is a safe intravenous gammaglobulin effective in patients with secondary hypogammaglobulinemia and ITP.

Adolescent↗

Enveloped virus inactivation by fatty acid derivatives.

The enveloped bacteriophage phi6 has been shown to be a valuable model system for the preliminary screening of compounds that might be expected to inactivate herpes simplex virus and other enveloped mammalian viruses. A variety of fatty acid derivatives that form fluid micelles in aqueous media have been found to be potent inactivators of phi6. The chemical nature of the polar head group, the length of the alkyl chain(s), and the extent and geometry of unsaturation are all important parameters in determining the antiviral effectiveness of this class of compounds.

Antiviral Agents↗

Detergent solubilised bovine viral diarrhea virus elicits a similar immune response as the inactivated virus.

The immune response to two detergent solubilised preparations of Bovine Viral Diarrhea Virus (BVDV) proteins were compared with the response to a chemically inactivated BVDV preparation. The antibody titres were measured with the standard immunofluorescence assay and with a newly developed enzyme linked immunosorbent assay (ELISA). The range of viral protein specificity of the antibodies was compared using SDS-polyacrylamide gel electrophoresis (SDS-PAGE) followed by Western blot analysis. The results clearly demonstrate that the detergent solubilised viral proteins are equally as effective immunogens as the killed BVDV vaccine both in terms of antibody titers and specificity.

Animals↗

Prime-boost vaccination using DNA and whole inactivated virus vaccines provides limited protection against virulent feline immunodeficiency virus.

Protection against feline immunodeficiency virus (FIV) has been achieved using a variety of vaccines notably whole inactivated virus (WIV) and DNA. However protection against more virulent isolates, typical of those encountered in natural infections, has been difficult to achieve. In an attempt to improve protection against virulent FIV(GL8), we combined both DNA and WIV vaccines in a "prime-boost" approach. Thirty cats were divided into four groups receiving vaccinations and one unvaccinated control group. Following viral challenge, two vaccinated animals, one receiving DNA alone and one the prime-boost vaccine remained free of viraemia, whilst all controls became viraemic. Animals vaccinated with WIV showed apparent early enhancement of infection at 2 weeks post challenge (pc) with higher plasma viral RNA loads than control animals or cats immunised with DNA alone. Despite this, animals vaccinated with WIV or DNA alone showed significantly lower proviral loads in peripheral blood mononuclear cells and mesenteric lymph node cells, whilst those receiving the DNA-WIV prime-boost vaccine showed significantly lower proviral loads in PBMC, than control animals, at 35 weeks pc. Therefore both DNA and WIV vaccines conferred limited protection against viral challenge but the combination of WIV and DNA in a prime-boost approach appeared to offer no significant advantage over either vaccine alone.

Animals↗

Immunofluorescence assay for human immunodeficiency virus antibody: investigation of cell fixation for virus inactivation and antigen preservation.

Four cell fixation procedures were investigated for their abilities to inactivate human immunodeficiency virus (HIV) and preserve its antigenicity for antibody detection by immunofluorescence in MOLT-4-T4 cells. Air-dried cell smears were fixed in cold acetone, in acetone-methanol (1:1), in acetone-methanol (1:1) followed by 70% ethanol and then methanol, or in paraformaldehyde-acetone. Acetone alone did not inactivate cell-associated HIV, but the other three procedures did. HIV inactivation was achieved by storage of acetone-fixed cells at -70 degrees for 40 days. Antigenicity was measured by immunofluorescence assay titrations of selected human sera, a cerebrospinal fluid, and a gp41 monoclonal antibody. Acetone provided the best fixation as measured by fluorescence intensity and antibody titers. The other fixation methods all yielded weaker fluorescence signals and/or decreased titers. Acetone fixation and storage for 40 days at -70 degrees C provides safe and accurate immunofluorescence assay reagents.

Cell Line↗

[Safety of transfusion therapy with special emphasis on inactivated viruses in products made from human blood].

The most common complication of blood transfusion therapy is the transfusion-transmitted infection. The decrease of the risk had been achieved by several measures such as: the change from paid to all voluntary donor system, education of the population and potential donor groups as regards the risk factors, and the selection of donors without the risk factors, testing of donor blood for the presence of the hepatitis B and C viruses, HIV and syphilis, fractionation of plasma with the partition of viruses and their inactivation. In spite of all these efforts zero risk blood transfusion therapy has not been achieved and it is very unlikely that it will be achieved in the future. Clinicians must have sufficient background information on different procedures for viral inactivation in order to be able to select the least risky blood product.

Blood Coagulation Factors↗

Modulation of the immune response to the severe acute respiratory syndrome spike glycoprotein by gene-based and inactivated virus immunization.

Although the initial isolates of the severe acute respiratory syndrome (SARS) coronavirus (CoV) are sensitive to neutralization by antibodies through their spike (S) glycoprotein, variants of S have since been identified that are resistant to such inhibition. Optimal vaccine strategies would therefore make use of additional determinants of immune recognition, either through cellular or expanded, cross-reactive humoral immunity. Here, the cellular and humoral immune responses elicited by different combinations of gene-based and inactivated viral particles with various adjuvants have been assessed. The T-cell response was altered by different prime-boost immunizations, with the optimal CD8 immunity induced by DNA priming and replication-defective adenoviral vector boosting. The humoral immune response was enhanced most effectively through the use of inactivated virus with adjuvants, either MF59 or alum, and was associated with stimulation of the CD4 but not the CD8 response. The use of inactivated SARS virus with MF59 enhanced the CD4 and antibody response even after gene-based vaccination. Because both cellular and humoral immune responses are generated by gene-based vaccination and inactivated viral boosting, this strategy may prove useful in the generation of SARS-CoV vaccines.

Animals↗

Field trials of an inactivated virus vaccine against porcine parvovirus.

Serological response and reproductive performance were estimated in field trials of an inactivated virus vaccine against porcine parvovirus. Experiments were carried out in 10 selected pig breeding herds. A total of 277 seronegative gilts were used. Two hundred and twenty animals were vaccinated twice before mating, fourteen days apart and revaccinated after farrowing. Blood samples were obtained from both vaccinated and non-vaccinated (57 animal) control gilts, one week after the 2nd dose of vaccination, at farrowing time and one week after revaccination. Although there were considerable variations among the herds, the number of returns to oestrus in all herds was higher in vaccinated gilts (11.81%) than in the controls (10.52%). This difference, however, was not statistically significant. The reproductive performance results revealed the absence of an increase in the total born, as pooled values, in vaccinated gilts compared to controls. However, when these results are interpreted in relation to serological data, many control gilts were already seropositive before mating, or remained seronegative at farrowing. According to our results, the duration of immunity with this vaccine is apparently short, as there is a clear decrease in the titres between the 1st and the 2nd sampling times (2.35 +/- 0.14 and 1.97 +/- 0.08, respectively).

Animals↗