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[Studies on virus inactivation by chlorine during water disinfection (author's transl)].

The studies described indicate a potential for water-borne transmission of viral diseases and the problems involved in virus inactivation by means of water chlorination. In contrast to the amount of free chlorine, the value of the oxidation-reduction potential (ORP) was found to be a criterion of virus inactivation. For virus inactivation, higher ORP values and longer periods of contact than for the killing of bacteria, respectively, were found to be necessary. To ensure the inactivation of poliovirus in water contaminated with organic substances, an ORP of + 780 mV (0.3-0.6 mg/l free chlorine) should be maintained for 15-30 min. Adenovirus has shown an almost identical resistance to inactivation. Possibilities for utilizing the mechanism of virus inactivation by the action of chemical disinfectants are discussed.

Chlorine↗

[The virus-inactivating effect of inorganic peroxy compounds].

The inactivating effect of the anorganic peroxy compounds percarbonate and H2O2 on poliovirus is described. In addition, the reaction mechanism of these peroxy compounds with poliovirus was investigated. It was shown that the inactivation of free nucleic acid required similar (H2O2) or lower concentrations (percarbonate) than the inactivation of virus particles. After incubation with anorganic peroxy compounds the virus was not destroyed completely; though sufficiently inactivated, a rest of "intact" virus particles was demonstrated by ultracentrifugation in sucrose gradients.

Carbonates↗

Effect of manufacturing process parameters on virus inactivation by dry heat treatment at 80 degrees C in factor VIII.

BACKGROUND AND OBJECTIVES: Dry heat treatment at 80 degrees C for 72 h is used as a virus inactivation step for some coagulation factor concentrates such as Bio Products Laboratory's (BPL) factor VIII 8Y. In the current study, the effect of this process has been tested on a range of viruses. In addition the effect of various manufacturing process parameters on virus inactivation has been investigated. MATERIALS AND METHODS: Samples of product intermediate were obtained from manufacturing, spiked with virus and subjected to freeze drying and dry heat treatment. Virus inactivation was determined by infectivity assay. RESULTS: Freeze drying followed by dry heat treatment was effective for inactivating a wide range of enveloped and nonenveloped viruses. Sucrose or protein concentration had no effect on virus inactivation. Product presentation or the interruption of heat treatment also had no effect. The inactivation of some of the viruses was greater at higher residual water content but under such conditions the stability of the product was reduced. CONCLUSION: This virus inactivation step was effective for a wide range of viruses and over the range of process conditions encountered in manufacturing. This demonstrates the robustness of this process step.

Decontamination↗

Induction of immune response and protection from equine viral arteritis (EVA) by formalin inactivated-virus vaccine for EVA in horses.

Thirty-nine horses included 3 pregnant mares were examined by inoculating with formalin inactivated-virus vaccine for EVA. Antibody response of horses after one dose vaccination was somewhat poor and 50% effective inoculum dose of the vaccine should be included 10(8.4) pfu of virus before inactivation. After 2 doses given at an interval of 4 weeks, the horses developed such high titer of SN antibody as up to 1:5,120. The SN titer declined rather rapidly, but supplemental administration of the vaccine at an interval of more than 2 months elicited a prompt antibody response and SN titers persisted as 1:80 to 1:320 at 6 months after the administration. Therefore, supplemental administration of the vaccine as booster every 6 months or 1 year would be capable of maintaining high titer of SN antibody. The inactivated-virus vaccine prevented horses from clinical disease of EVA and protected pregnant mares from abortion by challenge exposure with virulent virus. Fifty percent protective dose in SN titer of 1:43 was confirmed by clinical signs and viremia.

Animals↗

Strategies for evaluation of enveloped virus inactivation in red cell concentrates using hypericin.

Photodynamically induced virus inactivation appears promising in preventing transmission of enveloped virus infections in transfusible blood products. The potential for utilizing hypericin as a photosensitizer to inactivate key enveloped viruses in packed red cell concentrates (PRC) was evaluated. In addition to inactivating effectively > or = 10(6) TCID50 of human immunodeficiency virus (HIV), inactivation of bovine viral diarrhea virus (BVDV) in PRC was used as a model for hepatitis C virus to overcome the deficiency in reliable experimental systems for hepatitis C virus (HCV) inactivation. BVDV was two orders of magnitude more sensitive to inactivation by hypericin than HIV. As part of the virucidal efficacy analyses, the effects of photosensitization on hemopoietic cell lines carrying quiescent integrated HIV provirus were studied as models for evaluating virus inactivation in latently infected cells. Phorbol ester-induced virus production by these cells was effectively prevented by photosensitization with hypericin. A refinement of the illumination conditions, incorporating a monochromatic sodium light source with an emission spectrum coinciding with the absorption peak of hypericin, was highly virucidal, however, caused unacceptable levels of hemolysis. Red blood cells could be protected from phototoxic cellular damage by complexing hypericin with human serum albumin (albumin-hypericin), but the decrease in hemolysis was at the expense of virucidal efficacy. Thus, excitation of hypericin with a fluorescent source appears to be useful potentially for virus inactivation in PRC.

Animals↗

Virus inactivation in bone tissue transplants (femoral heads) by moist heat with the 'Marburg bone bank system'.

Several virus inactivation procedures like heat treatment, gamma irradiation and chemical sterilization are used to increase the safety of bone tissue transplants. In this study we present data on the virus-inactivating effect of heat disinfection on human femoral heads, using the Marburg bone bank system 'Lobator sd-2'. Three enveloped viruses (human immunodeficiency virus type 2 [HIV-2], bovine viral diarrhoea virus as a model for Hepatitis C virus [HCV], and the herpesvirus pseudorabies virus), and three non-enveloped viruses (hepatitis A virus, poliomyelitis virus, and bovine parvovirus) were investigated. In a model system the central part of human femoral heads was contaminated with the respective cell-free virus suspension, establishing a direct contact between virus and native bone tissue. The core temperature in the femoral heads during the sterilization process was determined in additional model experiments. A temperature of 82.5 degrees C, given by the manufacturer as the effective temperature for virus inactivation, was maintained for at least 15 min in decartilaged femoral heads with a diameter of < or = 56 mm. Heat treatment using the Lobator sd-2 inactivated all viruses in human femoral heads below the detection limit (at least by a factor of > or =4 log(10)). By combining a well-focussed anamnesis of the donors and serological testing for relevant infection markers (anti-HIV-1/2, HBsAg, anti-HBcore, anti-HCV, TPHA) with heat treatment of femoral heads in the Lobator sd-2 system, a high safety level is achieved. To further increase virus safety of cadaveric bone transplants, it is recommended that multi-organ donors are tested by nucleic acid testing (i.e. polymerase chain reaction) for HIV, HBV and HCV genome.

Femur↗

Multivalent inactivated virus oil emulsion vaccines in broiler breeder chickens. I. Newcastle disease virus and infectious bursal disease virus bivalent vaccines.

Inactivated Newcastle disease virus (NDV) and infectious bursal disease virus (IBDV) were incorporated into water-in-oil emulsion vaccines alone or as a bivalent vaccine. Twenty-week-old broiler breeder chickens that had received previous live virus vaccination with NDV and IBDV were injected intramuscularly with the monovalent or bivalent vaccine. The antibody titers to either the monovalent vaccine or bivalent vaccine increased rapidly and then remained at high levels for the duration of the 40-week trial. There were no practical differences in amplitude or duration of the antibody response to either antigen used alone compared to that of the bivalent combination. Progeny hatched from the vaccinated breeders possessed maternal antibody levels at one day of age comparable to those of the hens at the time the eggs were laid. The maternal antibody titers declined at a steady rate until they reached negligibly detectable levels at approximately 3 weeks of age. This trend held true without regard to the initial antibody titer.

Animals↗

Protection of mice against Aujeszky's disease virus infection by intranasal vaccination with inactivated virus.

Intranasal vaccination of mice with inactivated Aujeszky's disease virus (ADV) induced IgA and IgG antibody responses to the virus in the secretion of the respiratory tract, resulting in complete protection of the animals against intranasal challenge with virulent ADV. The immune response was enhanced by the use of the cholera toxin B subunit (CTB) as an adjuvant. On the other hand, subcutaneous vaccination of mice with inactivated ADV, even together with CTB, scarcely stimulated secretory antibody responses, resulting in only partial protection. The present results suggest that development of a vaccination procedure to stimulate the mucosal immune response should improve the protective effects of the inactivated herpesvirus vaccines, and thereby make it possible to control the infections by prohibiting virus replication at the site where primary infection takes place, as well as inhibiting subsequent latency and reactivation of the virus.

Administration, Intranasal↗

Transient inhibition of polyoma virus synthesis by sendai virus (parainfluenza I). II. Mechanism of the interference by inactivated virus.

The mechanism of the transient inhibition of polyoma virus synthesis by betapropiolactone-inactivated Sendai virus was studied. Polyoma virus early functions did not appear to be affected, although deoxyribonucleic acid (DNA) and structural protein synthesis were inhibited 60 and 35% respectively. The inhibition of macromolecular synthesis was not sufficient to account for the 90% inhibition of infectious progeny formation. Encapsidation of polyoma DNA into mature virions appears to be completely inhibited after superinfection by beta-propiolactone-inactivated Sendai virus. Ultraviolet irradiation of live or beta-propiolactone-inactivated Sendai virus preparations abolishes the interfering capacity, indicating that a functional Sendai virus ribonucleic acid molecule is the interfering component.

Animals↗

Factors influencing virus inactivation and retention of platelet properties following treatment with aminomethyltrimethylpsoralen and ultraviolet A light.

A wide variety of viruses are inactivated by psoralen compounds in the presence of ultraviolet A light (UVA). Use of aminomethyltrimethylpsoralen (AMT) and UVA is being evaluated as a method to inactivate viruses that may be present in platelet suspensions prepared for transfusion. Studies have been conducted to assess how variation in various environmental parameters influences the extent of viral inactivation and the retention of platelet properties. Most notably, it was determined that increasing levels of plasma progressively inhibited the inactivation of model viruses. As a result, experiments were routinely conducted at a plasma level of approximately 14.5%, using 40 micrograms/ml AMT, which was determined to be optimal when using this reduced plasma level. The reduced plasma level was achieved by dilution with a nonplasma medium that has been shown to be satisfactory for storage of platelets. Under these conditions, about 5 logs of vesicular stomatitis virus (VSV), pseudorabies, and phi 6 inactivation were achieved. Variation of platelet and leukocyte counts, within normal levels, had a minimal effect on extent of viral inactivation. Although oxygen level (mean levels, 97.9 mm Hg versus 19.2 mm Hg) had only a small influence on viral inactivation with 2.4, 4.8, and 7.2 J/cm2 of UVA (equivalent to 1-3 minutes of exposure), in vitro platelet properties, such as medium pH, morphology characteristics, and aggregation response, were better retained with a longer exposure time at the reduced oxygen level. With normal oxygen (97.9 mm Hg), platelet properties declined substantially relative to untreated controls (no UVA, no AMT) on exposure to 4.8 J/cm2. Our studies have identified two sets of conditions that provide about 5 logs of virus inactivation without extensively altering platelet in vitro properties.

Blood↗

Virus inactivated single-donor fresh plasma preparations.

Photodynamic virus inactivation of single units of human fresh plasma in their plastic containers can be achieved by illuminating the plasma with visible light in the presence of low concentrations of one of the phenothiazine dyes methylene blue or toluidine blue. This procedure abrogates the infectivity of numerous lipid-enveloped viruses including that of human immune deficiency virus-1. In contrast, the functional activities and immunological properties of plasma proteins are only moderately influenced. Animal studies and the first clinical data suggest that photodynamically virus inactivated fresh plasma is as well tolerated as conventional fresh frozen plasma.

Blood Banks↗

Characterisation of a novel high-purity, double virus inactivated von Willebrand Factor and Factor VIII concentrate (Wilate).

This study summarises the biochemical and functional properties of a new generation plasma-derived, double virus inactivated von Willebrand Factor/Factor VIII (VWF/FVIII) concentrate, Wilate, targeted for the treatment of both von Willebrand disease (VWD) and haemophilia A. The manufacturing process comprises two chromatographic steps based on different performance principles, ensuring a high purity of the concentrate (mean specific activity in 15 consecutive production batches: 122 IU FVIII:C/mg total protein) and, thus, minimising the administered protein load to the patient (specification: < or = 15 mg total protein per 900 IU Wilate). The optimised solvent/detergent (S/D) treatment and prolonged terminal dry-heat (PermaHeat) treatment of the lyophilised product at a specified residual moisture (RM) provide two mechanistically independent, effective and robust virus inactivation procedures for enveloped viruses and one step for non-enveloped viruses. These process steps are aggressive enough to inactivate viruses efficiently, but yet gentle enough to maintain the structural integrity and function of the VWF and FVIII molecules, as proven by state-of-the-art assays covering the diverse features of importance. The VWF multimeric pattern is close to the one displayed by normal plasma, with a consistent content of more than 10 multimers, but a relatively lower portion of the very high multimers. The multimeric triplet structure is normal, underlining the gentle and effective manufacturing process, which does not require the addition of protein stabilisers at any step. The balanced activity ratio of VWF to FVIII is close to that of plasma from healthy subjects, rendering Wilate suitable also for the safe and effective treatment of patients with VWD.

Blood Proteins↗

Protection against lethal enterovirus 71 infection in newborn mice by passive immunization with subunit VP1 vaccines and inactivated virus.

Enterovirus 71 (EV71), the newest member of Enteroviridae, is notable for its etiological role in epidemics of severe neurological diseases in children. Developing effective vaccines is considered a top choice among all control measures. We compared the inactivated virus vaccine (10 microg protein/mouse) with subunit vaccines--VP1 DNA vaccine (100 microg/mouse) or recombinant VP1 protein (10 microg/mouse)--in its ability to elicit maternal antibody and to provide protection against lethal infection of EV71 in suckling mice. Prior to gestation, all three groups of vaccinated dams possessed similar levels of neutralizing antibody. With a challenge dose of 2300 LD(50) virus/mouse, suckling mice born to dams immunized with inactivated virus showed 80% survival. The subunit vaccines provided protection only at a lower challenge dosage of 230 LD(50) per mouse, with 40% survival for DNA vaccine and 80% survival for VP1 protein. The cytokine profile produced by splenocytes showed a high level of IL-4 in the inactivated virus group, high levels of IFN-gamma and IL-12 in the DNA vaccine group, and high levels of IL-10 and IFN-gamma in the VP1 protein group. Overall, the inactivated virus elicited a much greater magnitude of immune response than the subunit vaccines, including total IgG, all four IgG subtypes, and T-helper-cell responses; these antibodies were shown to be protective against lethal infection when passively transferred to susceptible newborn mice. Our data indicated that inactivated virus is the choice of vaccine preparation capable of fulfilling the demand for effective control, and that VP1 subunit vaccines remain promising vaccine strategies that require further refinement.

Animals↗

Transient inhibition of polyoma virus synthesis by Sendai virus (parainfluenza I). I. Demonstration and nature of the inhibition by inactivated virus.

Superinfection of polyoma virus-infected mouse embryo cells by beta-propiolactone-inactivated Sendai virus resulted in a 90% inhibition of the synthesis of infectious polyoma progeny. The interference is dependent upon the time of superinfection and the concentration of the inactivated virus. The inhibition of polyoma virus synthesis is transient in nature since normal synthesis of polyoma progeny virus is seen upon prolonged incubation. Interferon does not appear to be implicated in the interference. Various aspects of the biological and synthetic capabilities of beta-propiolactone-inactivated Sendai virus are also described.

Animals↗

Comparable virus inactivation by bovine or vegetable derived Tween 80 during solvent/detergent treatment.

A mixture of Tri-n-butyl phosphate (TNBP) and Polysorbate 80 (Tween 80) is often used for virus inactivation during the manufacture of medicinal products derived from human plasma. This procedure, known as solvent/detergent treatment, is of high effectiveness for inactivation of enveloped viruses. Tween 80 can be manufactured from bovine tallow or from vegetable material. As the bovine-derived Tween 80 is normally used for the solvent/detergent treatment, the question has been raised whether vegetable-derived Tween 80 can be applied as an alternative substance for the solvent/detergent treatment. Comparable inactivation studies were therefore performed using Vesicular Stomatitis Virus (VSV), Pseudorabiesvirus (PRV), Semliki Forest Virus (SFV) and Bovine Diarrhoea Virus (BVDV). In principle, no differences were observed in the effectiveness of the solvent/detergent treatment when bovine or vegetable-derived Tween 80 was used. The comparability in the efficiency of both detergents for virus inactivation was shown to be independent of solvent/detergent concentration, of temperature (16 degrees C and 6 degrees C vs. 27 degrees C and 25 degrees C) and protein concentration (10% and 5% human albumin). In summary, vegetable-derived Tween 80 is of the same effectiveness as bovine-derived Tween 80, when used for virus inactivation by the solvent/detergent treatment.

Animals↗

Virus inactivation and protein recovery in a novel ultraviolet-C reactor.

BACKGROUND AND OBJECTIVES: Ultraviolet-C (UVC) irradiation is a viral-inactivation method that was dismissed by many plasma fractionators as a result of the potential for protein damage and the difficulty in delivering uniform doses. A reactor with novel spiral flow hydraulic mixing was recently designed for uniform and controlled UVC treatment. The objective of this study was to investigate virus inactivation and protein recovery after treatment through the new reactor. MATERIALS AND METHODS: Virus- and mock-spiked Alpha1-proteinase inhibitor (Alpha1-PI) solutions were treated with UVC. The virus samples were assayed for residual infectivity and amplified by the polymerase chain reaction (PCR). The mock-spiked samples were assayed for protein integrity. RESULTS: Greater than 4 log10 of all test viruses were inactivated, regardless of the type of nucleic acid or presence of an envelope. Unlike previous studies, viruses with the smallest genomes were found to be those most sensitive to UVC irradiation, and detection of PCR amplicons > or = 2.0 kb was correlated to viral infectivity. Doses that achieved significant virus inactivation yielded recovery of > 90% protein activity, even in the absence of quenchers. CONCLUSIONS: The results demonstrate the effectiveness of UVC treatment, in the novel reactor, to inactivate viruses without causing significant protein damage, and confirm the utility of large PCR amplicons as markers for infectious virus.

Adenoviruses, Human↗