Inactivation of Template-Directed Misfolding of Infectious Prion Protein by Ozone

Abstract

Misfolded prions (PrP Sc ) are well known for their resistance to conventional decontamination processes. The potential risk of contamination of the water environment, as a result of disposal of specified risk materials (SRM), has raised public concerns. Ozone is commonly utilized in the water industry for inactivation of microbial contaminants and was tested in this study for its ability to inactivate prions (263K hamster scrapie ‫؍‬ PrP Sc ). Treatment variables included initial ozone dose (7.6 to 25.7 mg/liter), contact time (

Kinetics of Ozone Inactivation of Infectious Prion Protein

“…During the ozonation phase, reaction vials were covered with plastic lids. Samples from each of the vessels were withdrawn at predetermined times and assayed for residual ozone concentration by the indigo method, as described by Ding et al (15). For the volume remaining in each of the reaction vessels, ozone activity was immediately neutralized by the addition of 20 l of 1 M sodium thiosulfate.…”

“…Three-to six-week-old female Syrian golden hamsters (Charles River Laboratories International, Inc., Wilmington, MA) were used to prepare infectious brain homogenates (IBH) and normal brain homogenates (NBH). The detailed procedure for preparation of homogenates was described previously by Ding et al (15).…”

“…The procedures for preparation of ozone demand-free (ODF) 1ϫ phosphate-buffered saline (PBS) (pH 4.4, pH 6.0, and pH 8.0), reaction vials, pipette tips, and magnetic bars were described previously by Ding et al (15). Ozone stock solutions (in 1ϫ PBS at pH 4.4, 6.0, and 8.0) were generated using ultrapure oxygen and an ozone generator (G30; PCI Wedeco).…”

“…We reported that higher ozone doses, greater contact time (5 min), low pH (4.4), and elevated temperature (20°C) resulted in the highest level of PrP Sc inactivation, as evaluated by protein misfolding cyclic amplification (PMCA). With the demonstration of the effectiveness of ozone for inactivation of PrP Sc and the foundation of preliminary derivation of CT (disinfectant concentration multiplied by contact time) under various conditions in our previous study (15), an understanding of the kinetics of ozone inactivation of PrP Sc is required for the design of competent water treatment technologies capable of inactivating infectious prions.…”

“…We recently examined the effectiveness of PrP Sc inactivation by ozone (15). PrP Sc (scrapie 263K) was inactivated by ozone under a variety of conditions, with inactivation levels being dependent upon the applied ozone dose (7.6 to 25.7 mg/liter), contact time (5 s and 5 min), pH (pH 4.4, 6.0, and 8.0), and temperature (4 and 20°C).…”