Inactivation of Transmissible Spongiform Encephalopathy (Prion) Agents by Environ LpH

Abstract

Agents causing transmissible spongiform encephalopathy (TSE) diseases are resistant to inactivation by several conventional decontamination methods. Using an animal bioassay, we compared the TSE agent disinfectant efficacy of a commercially available product referred to alternatively as LpH-SE, LpH-AG, or LpH-st to that of a similarly named but differently formulated product, Environ LpH, which was found to be an effective TSE agent disinfectant in a previous study. Here, we found LpH-SE to be at least 10⁴-fold to 10⁵-fold less effective than Environ LpH.

Transmissible spongiform encephalopathy (TSE) agents are notoriously difficult to destroy by treatments that are effective against other infectious agents. Nevertheless, several compounds with at least some anti-TSE agent activity have been reported (1-6). Of these, Environ LpH (Steris Corp., St. Louis, Mo.) was found to be particularly effective (4). Additional advantages of Environ LpH are that it is more stable and less hazardous than other chemicals commonly used for TSE agent decontamination, such as sodium hydroxide or sodium hypochlorite (6). As a result, research and diagnostic laboratories, hospitals, and other medical facilities began using Environ LpH to inactivate items contaminated by TSE agents. At the time of the previous study (4), the Environ LpH formulation was designated LpH. After the previous study, it was also referred to as Canadian LpH. Causing even more confusion, the manufacturer introduced a different formulation of LpH, variously marketed as LpH-SE, LpH-AG, or LpH-st. Though the chemical formulations of these three products are identical to each other, they differ considerably from that of Environ LpH. Some users of these products assumed that the newer LpH formulations have the same anti-TSE activity as the Environ LpH originally tested. However, since the chemical composition of the newer formulation is different from that of the originally tested formulation, it is possible that these differences might alter TSE inactivation properties (Table 1).

TABLE 1.

Chemical composition of LpH compounds

Chemical component	% by wta
	Environ LpHb	LPH-SEb, LPH-AGb, LPH-stb
o-Benzyl-p-chlorophenol	6.4
p-Tertiary-amylphenol	3.0	7.6
o-Phenyl phenol	0.5	7.7
Hexylene glycol	4.0
Glycolic acid (hydroxyacetic)	12.6
Isopropanol	8.0	7.5
Phosphoric acid		15.0
Dodecyl benzene sulfonic acid		<5

^aPercent by weight from material safety data sheet.

^bManufactured by Steris Corporation, St. Louis, Mo.

To determine if the alternative LpH compounds have anti-TSE activity equal to that of Environ LpH, we tested the effect of 1% (vol/vol) LpH-SE on a 9% (wt/vol) brain suspension derived from clinically sick, 263K scrapie agent-infected Syrian golden hamsters for a 16-h contact period as described previously (4). Groups of normal hamsters were inoculated with three successive 10-fold dilutions of the LpH-SE-treated brain suspensions. All hamsters inoculated with the LpH-SE-treated brain developed scrapie, indicating at least some resistance of the 263K agent to LpH-SE. This result differs from earlier data, where all hamsters inoculated with Environ LpH-treated brain suspensions survived when inoculated with the identical dilution series (4). LpH-SE did have some inactivating effect, however, because the incubation periods of hamsters inoculated with LpH-SE-treated brain were longer than the incubation periods of hamsters receiving similar dilutions of untreated brain (Table 2). Comparison of the incubation periods for LpH-SE-treated brain to a standard curve relating incubation period to infectivity titer for untreated brain indicated a 100- to 1,000-fold inactivation of the 263K hamster agent by LpH-SE (data not shown). In an earlier study, Environ LpH inactivated >10⁷ logs₁₀ of 263K hamster infectivity (4). Thus, Environ LpH is several orders of magnitude (≥10⁴ to 10⁵) more effective than LpH-SE as a TSE disinfectant.

TABLE 2.

Effect of LpH-SE and Environ LpH on 263K scrapie infectivity in Syrian golden hamsters^a

Treatment	% Brain inoculated	No. of Sc+ hamsters/no. inoculated	Interval to sacrifice (days) (mean ± SD)
LpH-SE	0.1	4/4	89 ± 0
	0.01	4/4	100 ± 4.5
	0.001	4/4	133 ± 38
None	0.1	4/4	82 ± 0
	0.01	4/4	89 ± 0
	0.001	4/4	92 ± 3
LpH-SE + autoclave	0.1	0/4	*b
Environ LpH	0.1	0/3	*b
Environ LpH + autoclave	0.1	0/4	*b

^aData relative to Environ LpH is taken from an earlier study (4) and is shown for comparison to LpH-SE. The Environ LpH result for 1% LpH and 0.1% brain homogenate is shown. Hamsters were inoculated intracerebrally with 50 μl of the indicated suspension of LpH-treated or untreated brain. The interval to sacrifice is the time from inoculation to definite clinical scrapie. Sc+, scrapie positive.

^b*, no clinical scrapie disease during a 15-month postinoculation observation period.

We also tested the effect of autoclaving the LpH-SE-treated brain. A 0.9% suspension of 263K-infected hamster brain was autoclaved at 121°C for 60 min in a 1% LpH-SE solution. This combination treatment removed all detectable infectivity, similar to earlier results using Environ LpH (4).

These studies show that LpH-SE has limited anti-TSE efficacy unless combined with autoclaving. Without autoclaving, Environ LpH is a far superior disinfectant for the hamster263K agent, one of the highest-titered TSE agents known (1). We did not determine which of the Environ LpH ingredients provides the anti-TSE effect. Three Environ LpH components—glycolic acid, hexylene glycol, and o-benzyl-p-chlorophenol—are not present in LpH-SE. Presumably, one of these or a combination of one or more of them provides the effect.

Based on our studies, we recommend Environ LpH, but not the other LpH products, for use as a TSE disinfectant.