Excerpts from scientific studies on inactivation of TSE infectivity by "steam sterilization"
TSE-contaminated wastewater from the NADC's labs, necropsy buildings, and animal containment buildings are treated as follows: "Stored waste water is next transferred to one of four heat treatment cook tanks (6,500 gallons each) that uses direct steam injection, at 250 degrees F (121 degrees C), at one atmosphere of pressure with a batch retention time of 30 minutes." Source: NADC Waste Water Disposal Evaluation - Report of the Scientific Review Panel - November 21, 2006. It should be noted that the panel's report did not reference a single scientific study to support its conclusion that "the NADC waste water pretreatment facility as designed and operated should be an effective process to inactivate TSE infectivity." In fact there are no published studies evaluating the efficacy of this specific process. There are, however, numerous studies of various physical and chemical inactivation methods, including autoclaving which is the application of superheated steam under pressure. The following excerpts are taken from peer-reviewed studies and scientific literature reviews and address the limitations of autoclaving for inactivation of TSE infectivity. JW
"High-titered (greater than 10(10) LD50 [50% lethal dose[/g) preparations of scrapie virus-infected hamster brain were subjected to inactivation by various chemicals, autoclaving, and histopathologic processing....One hour in an autoclave at 121 C reduced the titer of scrapie virus by approximately 7.5 log LD50/g of brain but left 2.5 log LD50/g of residual infectivity. A combination of exposure to chemicals and autoclaving may be necessary to sterilize high-titered scrapie virus-infected tissue." P. Brown, R.G. Rohwer, E.M. Green, D.C. Gajdusek. 1982. Effect of chemicals, heat, and histopathologic processing on high-infectivity hamster-adapted scrapie virus. Journal of Infectious Diseases 145(5):683-7.
"Autoclaving at 132 degrees C for 90 min completely eliminated the detectable infectivity, and was at least 100-fold more effective than 121 degrees C for 60 min. Although earlier reports stated that the scrapie agent contained in hamster brain homogenate can be destroyed by exposure to 132 degrees C for 60 min (Brown, et al., 1986), our data show that exposure to 132 degrees C for 60 min did not eliminate all the infectivity from the infected hamster brain homogenate....
Therefore, inactivation potentials by autoclaving may vary in different situations, and can be regarded as a treatment to lower titers of agent, but on the basis of our data, it is not safe to say that autoclaving sterilizes high titers of scrapie agent." D.R. Ernst, R.E. Race. 1993. Comparative analysis of scrapie agent inactivation methods. Journal of Virological Methods 41:193-202.
"Autoclaving at 126 degrees C for 1-2 h reduced titres by 10(3)-10(7) units, depending on the strain of agent. However, total disinfection of brain containing high titres of infectivity was approached only at 136 degrees C when titre losses of about 10(6) units were obtained by autoclaving for 4-32 min. Further studies are needed before we can make simple, general recommendations for the disinfection of CJD agents in hospital practice." R.H Kimberlin, C.A. Walker, G.C. Millson, D.M. Taylor, P.A. Robertson, A.H Tomlinson, A.G Dickinson. 1983. Disinfection studies with two strains of mouse-passaged scrapie agent. Guidelines for Creutzfeldt-Jakob and related agents. Journal of the Neurological Sciences 59(3):355-69.
"The resistance of small subpopulations of scrapie infectivity to inactivation at 100°C dictates against the use of boiling for disinfection of scrapie-contaminated material. Resistant subpopulations can also exist at 121°C (1, 2, 6, 8, 9). In other experiments we have recovered small amounts of infectivity from a sample exposed to 121°C for 60 minutes in an autoclave (1). The surviving population represented only 0.000002 percent of the starting infectivity. Nevertheless, this result suggests caution in the use of autoclaves for disinfection of scrapie and other members of this class of agents (1, 3, 21)." R.G. Rohwer. 1984. Virus-Like Sensitivity of the Scrapie Agent to Heat Inactivation. Science 223:600-602.
"Macerates of bovine brain infected with bovine spongiform encephalopathy (BSE) agent, and rodent brain infected with the 263K or ME7 strains of scrapie agent, were subjected to porous-load autoclaving at temperatures between 134 and 138 degrees C for < or = 60 min. Bioassay in rodents showed that none of the regimens produced complete inactivation." D.M. Taylor, H. Fraser, I. McConnell, D.A. Brown, K.L. Brown, K.A. Lamza, G.R. Smith. 1994. Decontamination studies with the agents of bovine spongiform encephalopathy and scrapie. Archives of Virology 139(3-4):313-26.
"Prions are very resistant to inactivation, and accidental transmission has occurred through the use of inadequate decontamination procedures....Infectivity can survive autoclaving at 132-138 degrees C, and under certain conditions the effectiveness of autoclaving actually declines as the temperature is increased. The small resistant subpopulations that survive autoclaving are not inactivated by simply re-autoclaving, and they acquire biological characteristics that differentiate them from the main population." D.M. Taylor. 1999. Inactivation of prions by physical and chemical means. Journal of Hospital Infection 43 Suppl:S69-76.
"Samples of macerated mouse-brain infected with the 22A strain of scrapie agent were subjected to gravity-displacement autoclaving at 121 degrees C for 30 minutes in the presence of 2 M sodium hydroxide. No infectivity was detectable by mouse bioassay in samples which were either held for an hour at room temperature before autoclaving, or autoclaved immediately after adding the hydroxide. In contrast, all of the mice injected with a control sample, held for an hour in distilled water before autoclaving, developed scrapie." D.M. Taylor, K. Fernie, I. McConnell. 1997. Inactivation of the 22A strain of scrapie agent by autoclaving in sodium hydroxide. Veterinary Microbiology 58(2-4):87-91.
"In the studies carried out so far, the BSE agent has proved to be just as resistant as other TSE agents to inactivation by procedures such as autoclaving or exposure to sodium hydroxide that are effective with conventional microorganisms...Despite the survival of BSE infectivity after autoclaving or exposure to sodium hydroxide, it is known that combining these procedures results in a very reliable degree of inactivation for TSE agents generally." D. Taylor. 2002. Inactivation of the BSE agent. Comptes rendus biologies 325(1):75-6.
"GD (gravity-displacement) autoclaving at 121 degrees C for 90 min reduced the titre of 263K (hamster-passaged scrapie) by 6 logs, but 3.4 logs survived (Prusiner, et al., 1984). This is in good agreement with the data obtained by Ernst and Race (1993) using the same agent and the same autoclaving regime, showing a 5.7 log reduction. Autoclaving the same agent at 134 degrees C for 30 min reduced the titre by 5.3 logs, but 3.5 logs survived (Brown, et al., 1990a). Autoclaving at 126 degrees C for 2 hours inactivated 139A (mouse-passaged scrapie) but not 22A (mouse-passaged scrapie) (Kimberlin, et al., 1983), which was inactivated only after a 4 hour exposure (Dickinson 1976)." D.M. Taylor. 2000. Inactivation of Transmissible Degenerative Encephalopathy Agents: A Review. The Veterinary Journal 159, 10-17.
"TSE agent strains show variability in their sensitivity to heat (Kimberlin et al., 1983; Somerville et al., 2002) although they are all difficult to inactivate by heating in an aqueous environment such as an autoclave...Although it is possible to achieve full inactivation (Rohwer, 1984), in some circumstances significant amounts of infectivity survive (Taylor et al., 1998)...Five experimentally maintained strains of scrapie and BSE agents have been passaged in two PrP genotypes of mice. Brain macerates were autoclaved at 126 °C and the levels of surviving infectivity were measured by titration. There was a large difference in the survival properties of transmissible spongiform encephalopathy (TSE) infectivity between TSE strains." D.M. Taylor, K. Fernie, P.J. Steele, I. McConnell, R.A. Somerville. 2002. Thermostability of mouse-passaged BSE and scrapie is independent of host PrP genotype: implications for the nature of the causal agents. Journal of General Virology 83, 3199–3204.