sodium-dodecyl-sulfate and Creutzfeldt-Jakob-Syndrome

Neurodegenerative human CJD and sheep scrapie are diseases caused by several different transmissible encephalopathy (TSE) agents. These infectious agents provoke innate immune responses in the brain, including late-onset abnormal prion protein (PrP-res) amyloid. Agent particles that lack detectable PrP sequences by deep proteomic analysis are highly infectious. Yet these agents, and their unusual resistance to denaturation, are often evaluated by PrP amyloid disruption. To reexamine the intrinsic resistance of TSE agents to denaturation, a paradigm for less resistant viruses and microbes, we developed a rapid and reproducible high yield agent isolation procedure from cultured cells that minimized PrP amyloid and other cellular proteins. Monotypic neuronal GT1 cells infected with the FU-CJD or 22L scrapie agents do not have complex brain changes that can camouflage infectious particles and prevent their disruption, and there are only 2 reports on infectious titers of any human CJD strain treated with chemical denaturants. Infectious titers of both CJD and scrapie were reduced by >4 logs with Thiourea-urea, a treatment not previously tested. A mere 5 min exposure to 4M GdnHCl at 22°C reduced infectivity by >5 logs. Infectious 22L particles were significantly more sensitive to denaturation than FU-CJD particles. A protocol using sonication with these chemical treatments may effectively decontaminate complicated instruments, such as duodenoscopes that harbor additional virulent microbes and biofilms associated with recent iatrogenic infections.

Topics: Amyloid; Animals; Biofilms; Cell Line; Creutzfeldt-Jakob Syndrome; Decontamination; Detergents; Guanidine; Mice; Neurons; Prions; Protein Denaturation; Scrapie; Sodium Dodecyl Sulfate; Surgical Instruments; Thiourea; Urea

Genetic Creutzfeldt-Jakob disease, Gerstmann-Sträussler-Scheinker syndrome, fatal familial insomnia and prion protein cerebral amyloid angiopathy are clinically and neuropathologically distinct neurodegenerative diseases linked to mutations in the PRNP gene encoding the cellular prion protein (PrPC). How sequence variants of PRNP encode the information to specify these disease phenotypes is not known. It is suggested that each mutation produces a misfolded variant of PrPC with specific neurotoxic properties. However, structural studies of recombinant PrP did not detect major differences between wild-type and mutant molecules, pointing to the importance of investigating mutant PrPs from mammalian brains. We used surface plasmon resonance and a slot-blot immunoassay to analyse the antibody-binding profiles of soluble and insoluble PrP molecules extracted from the brains of transgenic mice modelling different prion diseases. By measuring the reactivity of monoclonal antibodies against different PrP epitopes, we obtained evidence of conformational differences between wild-type and mutant PrPs, and among different mutants. We detected structural heterogeneity in both monomeric and aggregated PrP, supporting the hypothesis that the phenotype of genetic prion diseases is encoded by mutant PrP conformation and assembly state.

Topics: Animals; Antibodies, Monoclonal; Brain; Creutzfeldt-Jakob Syndrome; Detergents; Epitope Mapping; Gerstmann-Straussler-Scheinker Disease; Humans; Mice; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Transgenic; Mutation, Missense; Polymorphism, Genetic; Prions; Protein Binding; Protein Structure, Secondary; Protein Structure, Tertiary; Sodium Dodecyl Sulfate; Surface Plasmon Resonance

Distinct prion strains often exhibit different incubation periods and patterns of neuropathological lesions. Strain characteristics are generally retained upon intraspecies transmission, but may change on transmission to another species. We investigated the inactivation of two related prions strains: BSE prions from cattle and mouse-passaged BSE prions, termed 301V. Inactivation was manipulated by exposure to sodium dodecyl sulfate (SDS), variations in pH, and different temperatures. Infectivity was measured using transgenic mouse lines that are highly susceptible to either BSE or 301V prions. Bioassays demonstrated that BSE prions are up to 1,000-fold more resistant to inactivation than 301V prions while Western immunoblotting showed that short acidic SDS treatments reduced protease-resistant PrP(Sc) from BSE prions and 301V prions at similar rates. Our findings argue that despite being derived from BSE prions, mouse 301V prions are not necessarily a reliable model for cattle BSE prions. Extending these comparisons to human sporadic Creutzfeldt-Jakob disease and hamster Sc237 prions, we found that BSE prions were 10- and 10(6)-fold more resistant to inactivation, respectively. Our studies contend that any prion inactivation procedures must be validated by bioassay against the prion strain for which they are intended to be used.

Topics: Animals; Cattle; Cell Line; Creutzfeldt-Jakob Syndrome; Cricetinae; Encephalopathy, Bovine Spongiform; Humans; Hydrogen-Ion Concentration; Mice; Mice, Transgenic; Prions; Protein Denaturation; Sodium Dodecyl Sulfate; Species Specificity; Temperature

A variety of disinfection procedures were tested on two strains of scrapie agent, treated either as brain macerates (autoclaving) or as 10% homogenates (chemical treatments). It is suggested that a given treatment should produce a titre loss, of both strains of scrapie, of at least 10(4) units before it be regarded as useful for the disinfection of the agents of scrapie and Creutzfeldt-Jakob disease (CJD). By this criterion, treatment at room temperature with about 4% Hycolin (0.6% chlorinated phenols), 0.2% permanganate, 5% Tego (dodecyl-di(aminoethyl)-glycine) or 5% sodium dodecyl sulphate (SDS) are unsuitable. However, data indicate that SDS might be used to reduce the heat stability of scrapie agent. Hypochlorite (Sterilex) was the only satisfactory chemical reagent tested. At least 10(4)-10(5) units of infectivity were lost by treatment with hypochlorite containing 1,000 ppm available chlorine after a 4-16 h exposure, or containing 10,000 ppm available chlorine after a half-hour exposure. The latter result points to the use of concentrated hypochlorite (about 2% available chlorine; approximately 20% Sterilex) to decontaminate surfaces. We suggest that the cleaning action of SDS, or other strong detergents, might also help to decontaminate surfaces, but studies on this are needed. 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.

Topics: Animals; Creutzfeldt-Jakob Syndrome; Disinfection; Glycine; Humans; Hypochlorous Acid; Mice; Mice, Inbred Strains; Phenols; Potassium Permanganate; Prions; Scrapie; Sheep; Sodium Dodecyl Sulfate; Sterilization

The unusual resistance of the "unconventional viruses" to inactivation by the commonly used disinfectants has led to a high degree of apprehension regarding patients with any form of dementia. The rapid adaptation of a newly acquired isolate of the agent of Creutzfeldt-Jakob Disease (CJD) to mice made possible this large scale study of its heat and chemical stability. The agent showed a decrease in titer of approximately two logs following incubation at 80 degrees C for 30 minutes with no additional loss at 80 degrees C for up to 500 minutes. There was greater than a three log decrease in titer at 100 degrees C for 30 minutes and temperatures of 115 degrees and 130 degrees C completely inactivated the agent. Treatment with sodium hypochlorite at three concentrations (0.33 per cent, 0.66 per cent and 1.31 per cent) showed inactivation of greater than 99 per cent at each. Crude agent preparations were not inactivated by sodium dodecylsulfate at detergent to protein ratios up to 4:1. These results suggest that those hospital supplies which resist autoclaving may be adequately disinfected by autoclaving for at least 30 minutes. Treatment of surfaces with solutions of sodium hypochlorite (household bleach) at concentrations of 15 to 25 per cent is also effective. Detergent treatment of contaminated surfaces or materials is inadequate for proper decontamination.

Topics: Animals; Brain; Creutzfeldt-Jakob Syndrome; Culture Techniques; Decontamination; Disinfectants; Female; Hot Temperature; Humans; Mice; Sodium Dodecyl Sulfate; Sodium Hypochlorite