Inactivation by gamma irradiation of animal viruses in simulated laboratory effluent

Several animal viruses were treated with gamma radiation from a 60Co source under conditions which might be found in effluent from an animal disease laboratory. Swine vesicular disease virus, vesicular stomatitis virus, and blue-tongue virus were irradiated in tissues from experimentally infected animals. Pseudorabies virus, fowl plague virus, swine vesicular disease virus, and vesicular stomatitis virus were irradiated in liquid animal feces. All were tested in animals and in vitro. The D10 values, that is, the doses required to reduce infectivity by 1 log10, were not apparently different from those expected from predictions based on other data and theoretical considerations. The existence of the viruses in pieces of tissue or in liquid feces made no difference in the efficacy of the gamma radiation for inactivating them. Under the "worst case" conditions (most protective for virus) simulated in this study, no infectious agents would survive 4.0 Mrads.     

Inactivation of enveloped and non-enveloped viruses on seeded human tissues by gamma irradiation

Human tissue allografts are widely used in a variety of clinical applications with over 1.5 million implants annually in the US alone. Since the 1990s, most clinically available allografts have been disinfected to minimize risk of disease transmission. Additional safety assurance can be provided by terminal sterilization using low dose gamma irradiation. The impact of such irradiation processing at low temperatures on viruses was the subject of this study. In particular, both human tendon and cortical bone samples were seeded with a designed array of viruses and the ability of gamma irradiation to inactivate those viruses was tested. The irradiation exposures for the samples packed in dry ice were 11.6-12.9 kGy for tendon and 11.6-12.3 kGy for bone, respectively. The viruses, virus types, and log reductions on seeded tendon and bone tissue, respectively, were as follows: Human Immunodeficiency Virus (RNA, enveloped), >2.90 and >3.20; Porcine Parvovirus (DNA, non-enveloped), 1.90 and 1.58; Pseudorabies Virus (DNA, enveloped), 3.80 and 3.79; Bovine Viral Diarrhea Virus (RNA, enveloped), 2.57 and 4.56; and Hepatitis A Virus (RNA, non-enveloped), 2.54 and 2.49, respectively. While proper donor screening, aseptic technique, and current disinfection practices all help reduce the risk of viral transmission from human allograft tissues, data presented here indicate that terminal sterilization using a low temperature, low dose gamma irradiation process inactivates both enveloped and non-enveloped viruses containing either DNA or RNA, thus providing additional assurance of safety from viral transmission.     

Inactivation of Coxiella burnetii by gamma irradiation

The gamma radiation inactivation kinetics for Coxiella burnetii at -79 degrees C were exponential. The radiation dose needed to reduce the number of infective C. burnetii by 90% varied from 0.64 to 1.2 kGy depending on the phase of the micro-organism, purity of the culture and composition of suspending menstruum. The viability of preparations containing 10(11) C. burnetii ml-1 was completely abolished by 10 kGy without diminishing antigenicity or ability to elicit a protective immune response in vaccinated mice. Immunocytochemical examinations using monoclonal antibodies and electron microscopy demonstrated that radiation doses of 20 kGy did not alter cell-wall morphology or cell-surface antigenic epitopes.     

Inactivation of laboratory animal RNA-viruses by physicochemical treatment

Eight commonly used chemical disinfectants and physical treatments (UV irradiation and heating) were applied to both enveloped RNA viruses (Sendai virus, canine distemper virus) and unenveloped RNA viruses (Theiler's murine encephalomyelitis virus, reo virus type 3) to inactivate infectious virus particles. According to the results, alcohols (70% ethanol, 50% isopropanol), formaldehyde (2% formalin), halogen compounds (52ppm iodophor, 100ppm sodium hypochlorite), quaternary ammonium chloride (0.05% benzalkonium chloride) and 1% saponated cresol showed virucidal effects giving more than 99.95% reduction in the infectivity of virus samples of Sendai virus and canine distemper after 10 minutes exposure. There was no significant difference in the effects on the two enveloped RNA viruses. The susceptibility of unenveloped RNA viruses to chemical disinfectants and physical treatments differed greatly from the enveloped viruses. The two unenveloped viruses showed distinct resistance to 50% isopropanol, 2% formalin, 1% saponated cresol and to physical treatments (heating at 45, 56, 60 degrees C, and UV irradiation). These results indicate that using physicochemical methods to inactivate RNA viruses in laboratory animal facilities should be considered in accordance with the characteristics of the target virus. For practical purposes in disinfecting enveloped RNA viruses, 70% ethanol, 0.05% quaternary ammonium chloride and 1% saponated cresol diluted in hot water (greater than 60 degrees C) are considered as effective as UV irradiation. For unenveloped RNA viruses, halogen compounds, more than 1,000 ppm sodium hypochlorite or 260 ppm iodophor are recommended over a period of 10 minutes for disinfecting particles, although these compounds result in an oxidation problem with many metals.     

Inactivation of animal viruses during sewage sludge treatment.

Using a previously developed filter adsorption technique, the inactivation of a human rotavirus, a coxsackievirus B5, and a bovine parvovirus was monitored during sludge treatment processes. During conventional anaerobic mesophilic digestion at 35 to 36 degrees C, only minor inactivation of all three viruses occurred. The k' values measured were 0.314 log10 unit/day for rotavirus, 0.475 log10 unit/day for coxsackievirus B5, and 0.944 log10 unit/day for parvovirus. However, anaerobic thermophilic digestion at 54 to 56 degrees C led to rapid inactivation of rotavirus (k' greater than 8.5 log10 units/h) and of coxsackievirus B5 (k' greater than 0.93 log10 unit/min). Similarly, aerobic thermophilic fermentation at 60 to 61 degrees C rapidly inactivated rotavirus (k' = 0.75 log10 unit/min) and coxsackievirus B5 (k' greater than 1.67 log10 units/min). Infectivity of parvovirus, however, was only reduced by 0.213 log10 unit/h during anaerobic thermophilic digestion and by 0.353 log10 unit/h during aerobic thermophilic fermentation. Furthermore, pasteurization at 70 degrees C for 30 min inactivated the parvovirus by 0.72 log10 unit/30 min. In all experiments the contribution of temperature to the total inactivation was determined separately and was found to be predominant at process temperatures above 54 degrees C. In conclusion, the most favorable treatment to render sludge hygienically safe from the virological point of view would be a thermal treatment (60 degrees C) to inactivate thermolabile viruses, followed by an anaerobic mesophilic digestion to eliminate thermostable viruses that are more sensitive to chemical and microbial inactivations.     

Inactivation of viral agents in bovine serum by gamma irradiation

Cell culture origin or suckling mouse brain origin viruses of Akabane disease, Aino, bovine ephemeral fever, swine vesicular disease, hog cholera, bluetongue, and minute virus of mice were each suspended in bovine serum. Aliquots (1 mL) were exposed to various doses of gamma radiation from a 60Co source while at -68 degrees C. Aliquots (100-mL) of serum from a steer experimentally infected with foot-and-mouth disease virus were similarly irradiated. The samples were assayed for infectivity in cell culture systems before and after irradiation, and the data points were analyzed by linear regression. The irradiation doses (in megarads) necessary to inactivate one log10 of viral infectivity (D10) was calculated for each virus. D10 is otherwise known as the slope of the regression line. The r2 value, a measure of association with 1.0 = perfect fit, was also calculated for each regression line. The values (D10, r2) for each virus were as follows: Akabane, 0.25, 0.998; Aino, 0.35, 0.997; bovine ephemeral fever, 0.29, 0.961; swine vesicular disease, 0.50, 0.969; foot-and-mouth disease, 0.53, 0.978; hog cholera, 0.55, 0.974; bluetongue, 0.83, 0.958; and minute virus of mice, 1.07, 0.935.     

Inactivation of animal viruses during sewage sludge treatment

Using a previously developed filter adsorption technique, the inactivation of a human rotavirus, a coxsackievirus B5, and a bovine parvovirus was monitored during sludge treatment processes. During conventional anaerobic mesophilic digestion at 35 to 36 degrees C, only minor inactivation of all three viruses occurred. The k' values measured were 0.314 log10 unit/day for rotavirus, 0.475 log10 unit/day for coxsackievirus B5, and 0.944 log10 unit/day for parvovirus. However, anaerobic thermophilic digestion at 54 to 56 degrees C led to rapid inactivation of rotavirus (k' greater than 8.5 log10 units/h) and of coxsackievirus B5 (k' greater than 0.93 log10 unit/min). Similarly, aerobic thermophilic fermentation at 60 to 61 degrees C rapidly inactivated rotavirus (k' = 0.75 log10 unit/min) and coxsackievirus B5 (k' greater than 1.67 log10 units/min). Infectivity of parvovirus, however, was only reduced by 0.213 log10 unit/h during anaerobic thermophilic digestion and by 0.353 log10 unit/h during aerobic thermophilic fermentation. Furthermore, pasteurization at 70 degrees C for 30 min inactivated the parvovirus by 0.72 log10 unit/30 min. In all experiments the contribution of temperature to the total inactivation was determined separately and was found to be predominant at process temperatures above 54 degrees C. In conclusion, the most favorable treatment to render sludge hygienically safe from the virological point of view would be a thermal treatment (60 degrees C) to inactivate thermolabile viruses, followed by an anaerobic mesophilic digestion to eliminate thermostable viruses that are more sensitive to chemical and microbial inactivations.     

Inactivation of plant and animal viruses by proanthocyanidins from Alpinia zerumbet extract

Alpinia zerumbet (Pers.) B.L. Burtt and R.M. Smith belongs to the Alpinia genus in the Zingiberaceae family. In East Asia, Alpinia zerumbet has been widely used as food and traditional medicine. Previously, we identified proanthocyanidins (PACs), an anti-plant-virus molecule in A. zerumbet, using Nicotiana benthamiana and tomato mosaic virus (ToMV). Here, we found that PACs from A. zerumbet, apple, and green tea effectively inhibited ToMV infection. Additionally, the PACs from A. zerumbet exhibited greater antiviral activity than those from apple and green tea. The PACs from A. zerumbet also effectively inactivated influenza A virus and porcine epidemic diarrhea virus (PEDV), which acts as a surrogate for human coronaviruses, in a dose-dependent manner. The results from the cytopathic effect assays indicated that 0.1 mg/ml PACs from A. zerumbet decreased the titer of influenza A virus and PEDV by >3 log. These findings suggested that the direct treatment of viruses with PACs from A. zerumbet before inoculation reduced viral activity; thus, PACs might inhibit infections by an influenza virus, coronaviruses, and plant viruses.     

Inactivation of Vibrio parahaemolyticus in effluent seawater by alternating-current treatment

Vibrio parahaemolyticus, the cause of gastroenteritis in humans, was inactivated by alternating low-amperage electricity. In this study, the application of alternating low-amperage electric treatment to effluent seawater was investigated for the large-scale disinfection of seawater. This method was able to overcome the problem of chlorine generation that results from treatment with continuous direct current. In conclusion, our results showed that alternating-current treatment inactivates V. parahaemolyticus in effluent seawater while minimizing the generation of chlorine and that this alternating-current treatment is therefore suitable for practical industrial applications.     

Inactivation of viruses with photoactive compounds.

The transmission of human immunodeficiency virus (HIV-1) and other enveloped virus by blood transfusion is a major concern. Photosensitive dyes such as hematoporphyrin derivative (HPD), dihematoporphyrin ether (DHE), benzoporphyrin derivatives (BPD), extended ring porphyrins, sapphyrins and texaphyrins, and various cyanines were used with viral cultures to test the feasibility of using those light-excitable dyes to kill virus. A photodynamic flow cell was used to irradiate viral suspensions or viral infected cells in culture media or in whole blood. Herpes virus (HSV-1) was used to screen compounds. Effective compounds were subsequently tested for their ability to kill HIV-1, CMV, and SIV in culture medium and in blood and proved to effectively kill free virus and infected cells at significant viremias. Irradiation was achieved with a filtered xenon light source and/or tunable dye laser. Concentrations of dyes at 10 times viral kill dose were irradiated in blood which was tested for damage to erythrocytes (RBC), platelets, and blood proteins. No damage to RBC, complement factors, and immunoglobulins was evident immediately after photodynamic treatment. Platelet condition is minimally modified with time. Photodynamic treatment of blood appears to be a feasible means of eradicating virus and some protozoans from blood.     

Inactivation of enteric viruses in wastewater sludge through dewatering by evaporation.

The effect of dewatering on the inactivation rates of enteric viruses in sludge was determined. For this study, water was evaporated from seeded raw sludge at 21 degrees C, and the loss of viral plaque-forming units was measured. Initial results with poliovirus showed that recoverable infectivity gradually decreased with the loss of water until the solids content reached about 65%. When the solids content was increased from 65 to 83%, a further, more dramatic decrease in virus titer of greater than three orders of magnitude was observed. This loss of infectivity was due to irreversible inactivation of poliovirus because viral particles were found to have released their RNA molecules which were extensively degraded. Viral inactivation in these experiments may have been at least partially caused by the evaporation process itself because similar effects on poliovirus particles were observed in distilled water after only partial loss of water by evaporation. Coxsackievirus and reovirus were also found to be inactivated in sludge under comparable conditions, which suggests that dewatering by evaporation may be a feasible method of inactivating all enteric viruses in sludge.     

Inactivation of enteric viruses in wastewater sludge through dewatering by evaporation.

The effect of dewatering on the inactivation rates of enteric viruses in sludge was determined. For this study, water was evaporated from seeded raw sludge at 21 degrees C, and the loss of viral plaque-forming units was measured. Initial results with poliovirus showed that recoverable infectivity gradually decreased with the loss of water until the solids content reached about 65%. When the solids content was increased from 65 to 83%, a further, more dramatic decrease in virus titer of greater than three orders of magnitude was observed. This loss of infectivity was due to irreversible inactivation of poliovirus because viral particles were found to have released their RNA molecules which were extensively degraded. Viral inactivation in these experiments may have been at least partially caused by the evaporation process itself because similar effects on poliovirus particles were observed in distilled water after only partial loss of water by evaporation. Coxsackievirus and reovirus were also found to be inactivated in sludge under comparable conditions, which suggests that dewatering by evaporation may be a feasible method of inactivating all enteric viruses in sludge.     

Inactivation of rubella virus by gamma radiation

The Gilchrist and M-33 strains of rubella virus exposed in the frozen state to ¹³⁷Ce or ⁶⁰Co were inactivated exponentially according to “one hit” kinetics. There was no difference in the radiosensitivity of the two strains. Experimental D₃₇ values for both strains ranged from 1.9 × 10⁵ to 2.9 × 10⁵ rads, and computed radiosensitive molecular weights ranged from 2.6 × 10⁶ to 4.0 × 10⁶ daltons.     

Inactivation of viruses and cells by mustard gas

The action of mustard gas on six animal, one plant, and two bacterial viruses; also on bacteria, yeast, and the pneumococcus-transforming principle has been studied. The viruses include Newcastle's disease of chickens, equine encephalomyelitis (Eastern strain), feline pneumonitis (Baker), rabbit papilloma (Shope), fixed rabies, rabbit myxoma, tobacco mosaic, T(2)r(+) phage of E. coli B, and a Staphylococcus muscae phage. The cells include bakers' yeast, E. coli B, Staphylococcus muscae, and swine plague bacillus. The rates of inactivation of the viruses and cells were of the same order of magnitude and faster than those of enzymes. Of the viruses examined those containing desoxyribose nucleic acid were inactivated faster than those containing ribosenucleic acid. Preparations of the pneumococcus-transforming principle which were largely desoxyribose nucleic acid have shown the greatest sensitivity to mustard gas of all systems examined. An expression was derived describing the inactivation rate when mustard gas decreases during the experiment.     

Effect of gamma irradiation on human cortical bone transplants contaminated with enveloped and non-enveloped viruses.

In the production of bone grafts intended for transplantation, basic safety measures to avoid the transmission of pathogens are selection and serological screening of donors for markers of virus infections. As an additional safety tool we investigated the effect of gamma irradiation on the sterility of human bone diaphysis transplants and evaluated its impact on the virus safety of transplants. Model viruses were included in the study to determine the dose necessary to achieve a reduction factor for the infectivity titres of at least 4 log(10) at a temperature of -30+/-5 degrees C. The following viruses were used: human immunodeficiency virus type 2 (HIV-2), hepatitis A virus (HAV), and poliovirus (PV-1), and the following model viruses: pseudorabies virus (PRV) as a model for human herpesviruses, bovine viral diarrhoea virus (BVDV) for HCV, and bovine parvovirus (BPV) for parvovirus B19. A first approach was to determine the D(10) values (kGy) for the different viruses (virus inactivation kinetics: BPV 7.3; PV-1 7.1; HIV-2 7.1; HAV 5.3; PRV 5.3; BVDV <3.0 kGy). Based on these results, inactivation of these viruses was studied in experimentally contaminated human bone transplants (femoral diaphyses). For BPV, the most resistant one of the viruses studied, a dose of approximately 34 kGy was necessary to achieve a reduction of infectivity titres of 4 log(10). We therefore recommend a dose of 34 kGy for the sterilisation of frozen bone transplants.