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

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. distribution in these animals.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. was amplified from both lymphoid and neural tissues of positive control deer but not from identical tissues of negative control deer.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

Mother to offspring transmission of chronic wasting disease in Reeves' Muntjac deer

The horizontal transmission of prion diseases has been well characterized in bovine spongiform encephalopathy (BSE), chronic wasting disease (CWD) of deer and elk and scrapie of sheep, and has been regarded as the primary mode of transmission. Few studies have monitored the possibility of vertical transmission occurring within an infected mother during pregnancy. To study the potential for and pathway of vertical transmission of CWD in the native cervid species, we used a small cervid model-the polyestrous breeding, indoor maintainable, Reeves' muntjac deer-and determined that the susceptibility and pathogenesis of CWD in these deer reproduce that in native mule and white-tailed deer. Moreover, we demonstrate here that CWD prions are transmitted from doe to fawn. Maternal CWD infection also appears to result in lower percentage of live birth offspring. In addition, evolving evidence from protein misfolding cyclic amplification (PMCA) assays on fetal tissues suggest that covert prion infection occurs in utero. Overall, our findings demonstrate that transmission of prions from mother to offspring can occur, and may be underestimated for all prion diseases

Infectious Prions in Pre-Clinical Deer and Transmission of Chronic Wasting Disease Solely by Environmental Exposure

Key to understanding the epidemiology and pathogenesis of prion diseases, including chronic wasting disease (CWD) of cervids, is determining the mode of transmission from one individual to another. We have previously reported that saliva and blood from CWD-infected deer contain sufficient infectious prions to transmit disease upon passage into naïve deer. Here we again use bioassays in deer to show that blood and saliva of pre-symptomatic deer contain infectious prions capable of infecting naïve deer and that naïve deer exposed only to environmental fomites from the suites of CWD-infected deer acquired CWD infection after a period of 15 months post initial exposure. These results help to further explain the basis for the facile transmission of CWD, highlight the complexities associated with CWD transmission among cervids in their natural environment, emphasize the potential utility of blood-based testing to detect pre-clinical CWD infection, and could augur similar transmission dynamics in other prion infections

Dogs are resistant to prion infection, due to the presence of aspartic or glutamic acid at position 163 of their prion protein

Unlike other species, prion disease has never been described in dogs even though they were similarly exposed to the bovine spongiform encephalopathy (BSE) agent. This resistance prompted a thorough analysis of the canine PRNP gene and the presence of a negatively charged amino acid residue in position 163 was readily identified as potentially fundamental as it differed from all known susceptible species. In the present study, the first transgenic mouse model expressing dog prion protein (PrP) was generated and challenged intracerebrally with a panel of prion isolates, none of which could infect them. The brains of these mice were subjected to in vitro prion amplification and failed to find even minimal amounts of misfolded prions providing definitive experimental evidence that dogs are resistant to prion disease. Subsequently, a second transgenic model was generated in which aspartic acid in position 163 was substituted for asparagine (the most common in prion susceptible species) resulting in susceptibility to BSE‐derived isolates. These findings strongly support the hypothesis that the amino acid residue at position 163 of canine cellular prion protein (PrPC) is a major determinant of the exceptional resistance of the canidae family to prion infection and establish this as a promising therapeutic target for prion diseases.MINECO/FEDER. Grant Numbers: AGL2015‐65046‐C2‐1‐R, AGL2008‐05296‐C02 Interreg. Grant Number: POCTEFA EFA148/1

Tracking infectious prions in the body fluids of deer infected with chronic wasting disease

2010 Spring.Includes bibliographic references (page 111).Chronic wasting disease (CWD) is a prion disease of cervid (elk, moose and deer) with unusually high transmission efficiency. While the nidus of disease was described in a captive herd of cervid in northern Colorado/southeastern Wyoming in the late 60's, it has now been detected in both captive and free-ranging populations in 17 states and 2 Canadian provinces of North America and one Asian country. CWD is unique in being the only transmissible spongiform encephalopathy (TSE) described in a free-ranging population of animals. The etiology of CWD, like all prion diseases, is the conversion of the normal host-encoded cellular prion protein (PrPC) to an aberrantly folded protease resistant isoform (PrPRES/PrPCWD). An intriguing aspect of prion diseases is their ability to be transmitted from one organism to the next. In this dissertation work, we ask-By what means are infectious prions transmitted from one host to the next? In particular to CWD-What do infected cervids share or leave behind that contain sufficient infectious particles to initiate disease in the next cervid? We addressed this question by bioassay of body secretions and excretions- ' secreta '- (saliva, blood, urine and feces) in the native white-tailed deer host and in transgenic mice expressing the normal cervid prion protein (Tg(CerPrP) mice). Cohorts of deer were exposed by oral (PO) ingestion of 'secreta', or intraperitoneal (IP)/intravenous (IV) transfusion of blood components. To replicate a more natural/realistic exposure to CWD in which a deer might travel into a contaminated area and feed for a short period of time, an additional cohort of deer was exposed to fomites (bedding, feed and water buckets) from the suites of CWD-infected deer-without direct contact with infected deer. These variously exposed deer were monitored for a minimum of 19 months post inoculation (mo pi) for CWD infection and disease by immunohistochemical (IHC) and western blot (WB) detection of PrPCWD in serial tonsil biopsies and in multiple tissues after necropsy. Parallel studies were conducted in Tg(CerPrP) mice with the addition of an intracranial (IC) inoculation group for each body fluid. We found that sufficient infectious prions were present in the saliva, whole blood, the B cell- and platelet-enriched fractions of blood, and in fomites from infected deer premises to transmit CWD. Conversely, PrPCWD was not detected in the brain or lymphoid tissues of deer or mice inoculated with urine and feces, cell-free plasma or CD14+ monocytes from CWD-infected donor deer. The results of this work: 1) suggest that the efficient transmission of CWD may be due in part to the sharing of saliva between cervids and its deposition upon surfaces frequented by cervids; 2) establish a hematogenous dissemination of infectious prions in CWD associated with the cellular fraction of blood- in particular B cells and platelets— in CWD-infected deer; 3) extend previous work localizing PrPCWD to the interface of follicular B cells and dendritic cells; 4) provide insights to PrPCWD trafficking and CWD pathogenesis; and 5) establish saliva and blood cells as viable substrates for PrPCWD antemortem detection

Novel Use of Chlorine Dioxide Granules as an Alternative to Methyl Bromide Soil Fumigation

 Two greenhouse studies were conducted to evaluate the effectiveness of chlorine dioxide (ClO2) granules as a soil fumigation alternative for methyl bromide. The objective of the first study was to determine the efficacy of chlorine dioxide granules to inactivate Bacillus subtilis spore samples placed in two soils and positioned at two soil depths in soil tubes. The objective of the second study was to measure ClO2 gas concentrations released over multiple days in the two soils. The granules were evenly distributed in the soils with datalogger/sensors positioned at two different locations (headspace and soil matrix). The first study achieved a maximum spore log10 reduction of 4.12 and 5.82 for play sand and mixed soil, respectively, for inoculated samples placed 8 cm deep in the soil tubes with a chlorine dioxide rate of 240 g/tube. There was a 21-fold increase in percent organic matter for the mixed soil when compared to the play sand soil. The increase in organic matter in the mixed soil resulted in a 1.7 log10 reduction decrease due to the absorption of the gas onto the organic matter. In the second study, chlorine dioxide was collected at the bottom of the soil instead of volatilizing into the headspace of the soil tube because it is denser than air. The ClO2 at the bottom of the soil was 3.95x and 3.8x higher than the headspace gas concentration for the mixed and play sand soil, respectively, as averaged over all test runs and periods. Both studies show that the chlorine dioxide granules are a promising alternative to soil fumigation with methyl bromide. Further research is needed to refine the granule formulation release rates and develop more economical application rates