title: Detection of sub-clinical CWD infection in conventional test-negative deer long after oral exposure to urine and feces from CWD+ deer.
abstract: BACKGROUND
  Chronic 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.
  
  
  OBJECTIVE
  We 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 PrP(CWD) distribution in these animals.
  
  
  METHODS
  Various 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.
  
  
  RESULTS
  PrP(CWD) 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. PrP(CWD) was amplified from both lymphoid and neural tissues of positive control deer but not from identical tissues of negative control deer.
  
  
  CONCLUSIONS
  Detection 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.
authors: Department of Microbiology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado, United States of America.
authors: Nicholas J
authors: NJ
authors: Haley
authors: Candace K
authors: CK
authors: Mathiason
authors: Mark D
authors: MD
authors: Zabel
authors: Glenn C
authors: GC
authors: Telling
authors: Edward A
authors: EA
authors: Hoover
mesh_terms: D000818:Animals; D001681:Biological Assay / Q000379:methods; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism; D003670:Deer / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay; D005243:Feces; D007150:Immunohistochemistry / Q000379:methods; D008198:Lymph Nodes / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D009474:Neurons / Q000378:metabolism; D011328:Prions / Q000378:metabolism; D020033:Protein Isoforms; D034081:Wasting Disease, Chronic / Q000175:diagnosis* / Q000635:transmission* / Q000652:urine*
chemical_list: D011328:Prions; D020033:Protein Isoforms