Detection of Sub-Clinical CWD Infection in Conventional Test-Negative Deer Long after Oral Exposure to Urine and Feces from CWD+ Deer

Abstract

BackgroundChronic wasting disease (CWD) of cervids is a prion disease distinguished by high levels of transmissibility, wherein bodily fluids and excretions are thought to play an important role. Using cervid bioassay and established CWD detection methods, we have previously identified infectious prions in saliva and blood but not urine or feces of CWD+ donors. More recently, we identified very low concentrations of CWD prions in urine of deer by cervid PrP transgenic (Tg[CerPrP]) mouse bioassay and serial protein misfolding cyclic amplification (sPMCA). This finding led us to examine further our initial cervid bioassay experiments using sPMCA.ObjectivesWe sought to investigate whether conventional test-negative deer, previously exposed orally to urine and feces from CWD+ sources, may be harboring low level CWD infection not evident in the 19 month observation period. We further attempted to determine the peripheral PrPCWD distribution in these animals.MethodsVarious neural and lymphoid tissues from conventional test-negative deer were reanalyzed for CWD prions by sPMCA and cervid transgenic mouse bioassay in parallel with appropriate tissue-matched positive and negative controls.ResultsPrPCWD was detected in the tissues of orally exposed deer by both sPMCA and Tg[CerPrP] mouse bioassay; each assay revealed very low levels of CWD prions previously undetectable by western blot, ELISA, or IHC. Serial PMCA analysis of individual tissues identified that obex alone was positive in 4 of 5 urine/feces exposed deer. PrPCWD was amplified from both lymphoid and neural tissues of positive control deer but not from identical tissues of negative control deer.DiscussionDetection of subclinical infection in deer orally exposed to urine and feces (1) suggests that a prolonged subclinical state can exist, necessitating observation periods in excess of two years to detect CWD infection, and (2) illustrates the sensitive and specific application of sPMCA in the diagnosis of low-level prion infection. Based on these results, it is possible that low doses of prions, e.g. following oral exposure to urine and saliva of CWD-infected deer, bypass significant amplification in the LRS, perhaps utilizing a neural conduit between the alimentary tract and CNS, as has been demonstrated in some other prion diseases.

Temporal Resolution of Misfolded Prion Protein Transport, Accumulation, Glial Activation, and Neuronal Death in the Retinas of Mice Inoculated with Scrapie

“…4 The results reported herein contribute to a sizable body of work describing the time course of prion disease. A great deal has been done to identify early biomarkers of disease in cattle, 40e42 cervids, 43,44 primates, 45 sheep, 46 and rodent models of TSEs. 34,47 The retina is a tissue that is spatially separated from the rest of the brain, has a relatively simple organization, and can be readily assessed using rapid and noninvasive techniques.…”

Prion transmission

“…46 In addition, white-tailed deer orally inoculated with a mixture of urine and feces from CWD-infected deer were shown to be sub-clinically infected with CWD, again demonstrating that such excreta contain biologically relevant amounts of prion infectivity. 47 With regard to ovine scrapie, naturally infected sheep also excreted prion in their feces during both the preclinical and clinical phases of disease. rodent models and natural hosts of prion diseases.…”

Deer density and disease prevalence influence transmission of chronic wasting disease in white‐tailed deer

“…Additionally, there is a substantial incubation period between infection and clinical signs ('15 months) and death ('23 months post-infection; Williams et al 2002). CWD prions are shed in saliva (Mathiason et al 2006, Haley et al 2009), urine (Haley et al 2009), feces (Tamgü ney et al 2009), and infected carcasses , and other prions readily bind to soil and remain infective (Johnson et al 2007).…”