The Sterways process: a new approach to inactivating viruses using gamma radiation.

The Sterways process (Process) provides a means of terminally sterilizing and virally inactivating blood products. The Process uses gamma radiation in a controlled manner to achieve maximum viral kill with minimum product loss. This goal is achieved by controlling the dose rate for the particular product and formulation in question. Because the Process uses gamma radiation it not only inactivates known viruses but also emerging or mutilating viruses.

Virucidal treatment of blood protein products with UVC radiation.

The virus safety of blood derivatives continues to be of concern, especially with respect to nonenveloped and/or heat-stable viruses. Previously, we demonstrated that treatment of whole plasma, AHF concentrate or fibrinogen with short wavelength ultraviolet light (UVC) results in the inactivation of > or = 10(6) infectious doses (ID) of encephalomyocarditis virus (EMCV), hepatitis A virus (HAV) and porcine parvovirus (PPV), each of which is nonenveloped. Protein recovery was enhanced greatly by inclusion of the flavonoid, rutin, added prior to UVC exposure to quench reactive oxygen species. We now report on the treatment of albumin and intravenous immune globulin (IVIG) isolated by a previously described, integrated chromatographic method. Albumin was treated with either 0.1 or 0.2 J/cm2 UVC in the presence of 0.8 or 1.6 mM rutin; IVIG was treated with either 0.05 or 0.1 J/cm2 UVC in the presence of 0.5 or 1.0 mM rutin. Our results show that > or = 10(6.9) ID of EMCV and PPV were inactivated under each of the conditions studied except the treatment of albumin with 0.1 J/cm2 UVC in the presence of 1.6 mM rutin where 10(4.3) ID of EMCV and > or = 10(6.9) ID of PPV were killed. It appears that the sensitivity of PPV to UVC exceeds that of EMCV and that virus kill with UVC is higher in IVIG than in albumin. In the absence of rutin, UVC increased the extent of aggregation of both albumin and IVIG by two- to three-fold. With rutin present, the increase in albumin aggregation was reduced, and it was virtually eliminated by subsequent processing on Sephacryl S-200, a step in the existing procedure designed to remove aggregates. The increase in aggregation of IVIG appeared to be eliminated on inclusion of either 0.5 mM or 1 mM rutin. We conclude that both albumin and IVIG can be treated with UVC to inactivate > or = 10(6) ID of nonenveloped viruses. The inclusion of rutin during treatment helps protect against protein aggregation.

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.

Replication of DNA containing apurinic sites in human and mouse cells probed with parvoviruses MVM and H-1.

We studied the effect of apurinic sites on DNA replication in mouse and human cells, using parvoviruses MVM (minute virus of mice) and H-1 as probes. Although apurinic sites are efficient blocks to the replication of these single-stranded DNA viruses in vivo, depurinated parvoviruses can be reactivated if host cells have been preexposed to a subtoxic dose of UV light. The target of this conditional reactivation process is the conversion of depurinated input DNA into double-stranded replicative forms; the concomitant increase in viral mutagenesis strongly suggests that apurinic sites can be bypassed in mammalian cells.

The inactivation of a bovine enterovirus and a bovine parvovirus in cattle manure by anaerobic digestion, heat treatment, gamma irradiation, ensilage and composting.

A bovine enterovirus and a bovine parvovirus seeded into liquid cattle manure were rapidly inactivated by anaerobic digestion under thermophilic conditions (55 degrees C), but the same viruses survived for up to 13 and 8 days respectively under mesophilic conditions (35 degrees C). The enterovirus was inactivated in digested liquid manure heated to 70 degrees C for 30 min, but the parvovirus was not inactivated by this treatment. The enterovirus, seeded into single cell protein (the solids recovered by centrifugation of digested liquid manure), was inactivated by a gamma irradiation dose of 1.0 Mrad, but the parvovirus survived this dose. When single cell protein seeded with bovine enterovirus or bovine parvovirus was ensiled with cracked corn, the enterovirus was inactivated after a period of 30 days, while the parvovirus survived for 30 days in one of two experiments. Neither the enterovirus nor the parvovirus survived composting for 28 days in a thermophilic aerobic environment when seeded into the solid fraction of cattle manure. It was concluded that, of the procedures tested, only anaerobic digestion under thermophilic conditions appeared to be reliable method of viral inactivation to ensure the safety of single cell protein for refeeding to livestock. Composting appeared to be a suitable method for the disinfection of manure for use as a soil conditioner.

[Reactivation of gamma-ray irradiated parvovirus H-1 in cells of patients with ataxia telangiectasia and Huntington's chorea]. / Réactivation du Parvovirus H-1 irradié aux Rayons gamma dans les cellules de patients atteints d'ataxia telangiectasia et de la chorée de Huntington.

Parvovirus H-1 was used to probe the cellular radiosensitivity of two human degeneration syndromes AT and HC. No difference in the survival of gamma irradiated H-1 was detected between skin fibroblasts from such patients and from a normal individual. However, AT and normal cells were distinguished by the fact that the reactivation of irradiated H-1 could be increased by UV or X-irradiation of the latter but not of the former cells.

U.V.-enhanced reactivation of capsid protein synthesis and infectious centre formation in mouse cells infected with U.V.-irradiated Minute-Virus-of-Mice.

The effect of U.V.-irradiation of mouse A9 cells on their ability to replicate unirradiated and U.V.-irradiated Minute-Virus-of-Mice was studied. The survival of two viral functions was measured in primary infected cells: the synthesis of viral structural proteins, as detected in situ using an immunoenzymatic assay, and the production of infectious centres, as detected by plaque formation onto appropriate indicator cells. Cell irradiation prior to infection enhanced virus survival over that in control cells (U.V.-enhanced reactivation phenomenon). The magnitude of this effect was similar for both viral functions, suggesting that the step(s) of the virus cycle sensitive to the reactivation process precede(s) the release of the primary burst and secondary infection.

Direct and indirect effects of ultraviolet light on the mutagenesis of parvovirus H-1 in human cells.

U.v. radiation is directly mutagenic for the single-stranded DNA parvovirus H-1 propagated in human cells. Mutation induction in the progeny of u.v.-irradiated virus increased linearly with the dose and could be ascribed neither to an increased number of rounds of viral replication nor to the indirect activation of an inducible cellular mutator activity by the u.v.-damaged virus. The level of mutagenesis among the descendants of both unirradiated and u.v.-damaged H-1 was enhanced if the host cells had been exposed to sublethal doses of u.v. light before infection. This indirect enhancement of viral mutagenesis in pre-irradiated cells was maximal at multiplicities lower than 0.2 infectious particles/cell. The frequency of mutations resulting from cell pre-irradiation was only slightly higher for u.v.-irradiated than for intact virus. Thus, the induced cellular mutator appeared to be mostly untargeted in the dose range given to the virus. U.v.-irradiation of the cells also enhanced the mutagenesis of u.v.-irradiated herpes simplex virus, a double-stranded DNA virus ( Lytle and Knott , 1982).

UV-irradiation of related mouse hybrid cells: similar increase in capacity to replicate intact minute-virus-of-mice but differential enhancement of survival of UV-irradiated virus.

3 hybrid cell lines between mouse fibroblasts (A9) and mouse lymphoma cells (L5178YS) were compared with regard to the ability of UV-pre-irradiated cells to replicate intact (unirradiated) Minute-virus-of-Mice (MVM) and to reactivate UV-irradiated MVM. UV irradiation of cells before virus infection enhanced their ability to plaque intact virus (Enhanced Capacity) to a similar extent in the 3 hybrid cell lines. However, pretreatment of cells with UV radiation enhanced the survival of UV-damaged virus (Enhanced Reactivation) in only 1 of these hybrids. The lack of detectable Enhanced Reactivation in the other hybrids without concomitant change in their Enhanced Capacity, suggests that these processes are at least partly independent. Virus survival in unirradiated cells was similar for all 3 hybrid cell lines, indicating that the absence of detectable Enhanced Reactivation in 2 of the hybrids was not due to the constitutive expression of this process, but might rather result from its loss or extinction. The expression of both Enhanced Capacity and Enhanced Reactivation requires synthesis of protein de novo shortly after cell irradiation.

Mutagenesis of intact parvovirus H-1 is expressed co-ordinately with enhanced reactivation of ultraviolet irradiated virus in human and rat cells treated with 2-nitronaphthofurans.

The exposure of human or rat cells to non-toxic concentrations of two 2-nitronaphthofuran derivatives activated co-ordinately the transient expression of mutator and repair activities. These activities gave rise to both an increase in the mutagenesis (enhanced mutagenesis, EM) and survival (enhanced reactivation, ER) of unirradiated and u.v.-irradiated parvovirus H-1 used as respective probes. The kinetics of expression was the same for mutator and repair activities and for the two chemicals studied. The dose-responses of these activities were also parallel for a given chemical, but one of the furan derivatives exerted its inducing effect at concentrations 20-25 times lower than the other derivative. Both EM and ER were depressed by cycloheximide, and inhibitor of de novo protein synthesis. This is the first report which shows that chemicals can enhance the mutagenesis of undamaged DNA by activating the expression of mutator functions in mammalian, including human, cells. The ability of the two 2-nitronaphthofuran derivatives to trigger EM and ER was found to correlate with their reported mutagenicity in a conventional bacterial test system.

Stability of minute virus of mice to chemical and physical agents.

Minute virus of mice (MVM), a single-stranded deoxyribonucleic acid Parvovirus, was subjected to various inactivation procedures, including chemical disinfectants, heat, and ultraviolet radiation. MVM was found to be less stable than has been reported for other Parvoviruses. This virus was readily inactivated by a variety of chemical disinfectants, including alcohols, formaldehyde, glutaraldehyde, and chloroform. MVM was more sensitive to ultraviolet radiation than was Kilham's rat virus. MVM was more sensitive to heating at temperatures of 35 to 100 C than has been reported for other Parvoviruses. More than 95% of MVM infectivity was inactivated by heating (45, 60, or 100 C) for 60 min, acid (pH 2.0) treatment, or ultraviolet radiation treatment, although a small percentage (less than 2%) of the virus preparation was found to be resistant to these treatments. In addition, more than 99% of the infectivity of MVM was lost after storage at 4C for 10 weeks, although the virus was stable on storage in liquid nitrogen.