Abstract
Chronic wasting disease (CWD) is an efficiently transmitted prion disease of cervids, now identified in 22 United States, 2 Canadian provinces and Korea. One hallmark of CWD is the shedding of infectious prions in saliva, as demonstrated by bioassay in deer. It is also clear that the concentration of prions in saliva, blood, urine and feces is much lower than in the nervous system or lymphoid tissues. Rapid in vitro detection of CWD (and other) prions in body fluids and excreta has been problematic due to the sensitivity limits of direct assays (western blotting, ELISA) and the presence of inhibitors in these complex biological materials that hamper detection. Here we use real-time quaking induced conversion (RT-QuIC) to demonstrate CWD prions in both diluted and prion-enriched saliva samples from asymptomatic and symptomatic white-tailed deer. CWD prions were detected in 14 of 24 (58.3%) diluted saliva samples from CWD-exposed white-tailed deer, including 9 of 14 asymptomatic animals (64.2%). In addition, a phosphotungstic acid enrichment enhanced the RT-QuIC assay sensitivity, enabling detection in 19 of 24 (79.1%) of the above saliva samples. Bioassay in Tg[CerPrP] mice confirmed the presence of infectious prions in 2 of 2 RT-QuIC-positive saliva samples so examined. The modified RT-QuIC analysis described represents a non-invasive, rapid ante-mortem detection of prions in complex biologic fluids, excreta, or environmental samples as well as a tool for exploring prion trafficking, peripheralization, and dissemination.
Highlights
Chronic Wasting Disease (CWD) is a transmissible spongiform encephalopathy (TSE), or prion disease, that affects free-ranging and captive cervids [1,2]
We identified a number of saliva samples which at 1:10 dilution in dilution buffer were routinely positive with real-time quaking induced conversion (RT-QuIC)
We demonstrate that RT-QuIC can reproducibly detect prions in saliva, a complex biological fluid containing a wide range of complex glycoproteins including proteases [38,39,40]
Summary
Chronic Wasting Disease (CWD) is a transmissible spongiform encephalopathy (TSE), or prion disease, that affects free-ranging and captive cervids [1,2]. CWD appears to be the most transmissible of the prion diseases, and is recognized in twenty-two U.S states, as well as two Canadian provinces and the Republic of Korea (http:// www.nwhc.usgs.gov/). The emerging prevalence of CWD poses a challenge to wildlife management agencies, free-ranging and captive cervid populations, the hunting and food producing animal economies, and may pose a zoonotic risk. Monitoring for CWD prions would be carried out on minimally invasive biologic samples (such as saliva, blood, urine or feces) harvested from live animals in the field. Rapid, sensitive and specific ante-mortem detection of CWD and other prion diseases remains a challenge due to the low concentrations of prions and the presence of inhibitors in body fluids and excreta [10,11,12,13]
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