scholarly journals Inhibition of Protease-Resistant Prion Protein Formation in a Transformed Deer Cell Line Infected with Chronic Wasting Disease

2006 ◽  
Vol 80 (2) ◽  
pp. 596-604 ◽  
Author(s):  
Gregory J. Raymond ◽  
Emily A. Olsen ◽  
Kil Sun Lee ◽  
Lynne D. Raymond ◽  
P. Kruger Bryant ◽  
...  
Keyword(s):  
Cell Line ◽  
Prion Protein ◽  
Chronic Wasting Disease ◽  
Transmissible Spongiform Encephalopathy ◽  
Mule Deer ◽  
Simian Virus 40 ◽  
Spongiform Encephalopathy ◽  
Wasting Disease ◽  
Neuronal Marker

ABSTRACT Chronic wasting disease (CWD) is an emerging transmissible spongiform encephalopathy (prion disease) of North American cervids, i.e., mule deer, white-tailed deer, and elk (wapiti). To facilitate in vitro studies of CWD, we have developed a transformed deer cell line that is persistently infected with CWD. Primary cultures derived from uninfected mule deer brain tissue were transformed by transfection with a plasmid containing the simian virus 40 genome. A transformed cell line (MDB) was exposed to microsomes prepared from the brainstem of a CWD-affected mule deer. CWD-associated, protease-resistant prion protein (PrPCWD) was used as an indicator of CWD infection. Although no PrPCWD was detected in any of these cultures after two passes, dilution cloning of cells yielded one PrPCWD-positive clone out of 51. This clone, designated MDBCWD, has maintained stable PrPCWD production through 32 serial passes thus far. A second round of dilution cloning yielded 20 PrPCWD-positive subclones out of 30, one of which was designated MDBCWD2. The MDBCWD2 cell line was positive for fibronectin and negative for microtubule-associated protein 2 (a neuronal marker) and glial fibrillary acidic protein (an activated astrocyte marker), consistent with derivation from brain fibroblasts (e.g., meningeal fibroblasts). Two inhibitors of rodent scrapie protease-resistant PrP accumulation, pentosan polysulfate and a porphyrin compound, indium (III) meso-tetra(4-sulfonatophenyl)porphine chloride, potently blocked PrPCWD accumulation in MDBCWD cells. This demonstrates the utility of these cells in a rapid in vitro screening assay for PrPCWD inhibitors and suggests that these compounds have potential to be active against CWD in vivo.

scholarly journals Antemortem Detection of Chronic Wasting Disease Prions in Nasal Brush Collections and Rectal Biopsy Specimens from White-Tailed Deer by Real-Time Quaking-Induced Conversion

2016 ◽  
Vol 54 (4) ◽  
pp. 1108-1116 ◽  
Author(s):  
Nicholas J. Haley ◽  
Chris Siepker ◽  
W. David Walter ◽  
Bruce V. Thomsen ◽  
Justin J. Greenlee ◽  
...  
Keyword(s):  
Real Time ◽  
Large Scale ◽  
Chronic Wasting Disease ◽  
Transmissible Spongiform Encephalopathy ◽  
Rectal Biopsy ◽  
Spongiform Encephalopathy ◽  
Wasting Disease ◽  
Experimental Conditions ◽  
Biopsy Specimens

Chronic wasting disease (CWD), a transmissible spongiform encephalopathy of cervids, was first documented nearly 50 years ago in Colorado and Wyoming and has since spread to cervids in 23 states, two Canadian provinces, and the Republic of Korea. The expansion of this disease makes the development of sensitive diagnostic assays and antemortem sampling techniques crucial for the mitigation of its spread; this is especially true in cases of relocation/reintroduction of farmed or free-ranging deer and elk or surveillance studies of private or protected herds, where depopulation is contraindicated. This study sought to evaluate the sensitivity of the real-time quaking-induced conversion (RT-QuIC) assay by using recto-anal mucosa-associated lymphoid tissue (RAMALT) biopsy specimens and nasal brush samples collected antemortem from farmed white-tailed deer (n= 409). Antemortem findings were then compared to results from ante- and postmortem samples (RAMALT, brainstem, and medial retropharyngeal lymph nodes) evaluated by using the current gold standardin vitroassay, immunohistochemistry (IHC) analysis. We hypothesized that the sensitivity of RT-QuIC would be comparable to IHC analysis in antemortem tissues and would correlate with both the genotype and the stage of clinical disease. Our results showed that RAMALT testing by RT-QuIC assay had the highest sensitivity (69.8%) compared to that of postmortem testing, with a specificity of >93.9%. These data suggest that RT-QuIC, like IHC analysis, is an effective assay for detection of PrPCWDin rectal biopsy specimens and other antemortem samples and, with further research to identify more sensitive tissues, bodily fluids, or experimental conditions, has potential for large-scale and rapid automated testing for CWD diagnosis.

scholarly journals Transmission and Adaptation of Chronic Wasting Disease to Hamsters and Transgenic Mice: Evidence for Strains

2007 ◽  
Vol 81 (8) ◽  
pp. 4305-4314 ◽  
Author(s):  
Gregory J. Raymond ◽  
Lynne D. Raymond ◽  
Kimberly D. Meade-White ◽  
Andrew G. Hughson ◽  
Cynthia Favara ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Prion Protein ◽  
Chronic Wasting Disease ◽  
Transmissible Spongiform Encephalopathy ◽  
Wild Type Mouse ◽  
Mouse Strains ◽  
Spongiform Encephalopathy ◽  
Wild Type ◽  
Wasting Disease ◽  
Incubation Periods

ABSTRACT In vitro screening using the cell-free prion protein conversion system indicated that certain rodents may be susceptible to chronic wasting disease (CWD). Therefore, CWD isolates from mule deer, white-tailed deer, and elk were inoculated intracerebrally into various rodent species to assess the rodents' susceptibility and to develop new rodent models of CWD. The species inoculated were Syrian golden, Djungarian, Chinese, Siberian, and Armenian hamsters, transgenic mice expressing the Syrian golden hamster prion protein, and RML Swiss and C57BL10 wild-type mice. The transgenic mice and the Syrian golden, Chinese, Siberian, and Armenian hamsters had limited susceptibility to certain of the CWD inocula, as evidenced by incomplete attack rates and long incubation periods. For serial passages of CWD isolates in Syrian golden hamsters, incubation periods rapidly stabilized, with isolates having either short (85 to 89 days) or long (408 to 544 days) mean incubation periods and distinct neuropathological patterns. In contrast, wild-type mouse strains and Djungarian hamsters were not susceptible to CWD. These results show that CWD can be transmitted and adapted to some species of rodents and suggest that the cervid-derived CWD inocula may have contained or diverged into at least two distinct transmissible spongiform encephalopathy strains.

Rapid Procedure For Detecting Scrapie-Associated Fibrils In Chronic Wasting Disease Of Elk And Mule Deer

1999 ◽  
Vol 5 (S2) ◽  
pp. 1310-1311
Author(s):  
C.E. Hearne ◽  
J.L. Clapper ◽  
K.L. DeVries ◽  
E.S. Williams
Keyword(s):  
High Speed ◽  
Chronic Wasting Disease ◽  
Transmissible Spongiform Encephalopathy ◽  
Mule Deer ◽  
Proteinase K ◽  
Abnormal Behavior ◽  
Enzyme Treatment ◽  
Spongiform Encephalopathy ◽  
Wasting Disease ◽  
Gradual Loss

Chronic wasting disease (CWD) is a transmissible spongiform encephalopathy (TSE) of mule deer and elk. Affected animals are characterized clinically by a combination of abnormal behavior and gradual loss of body condition. The disease is invariably fatal. Diagnosis of CWD is made by histologic examination of the central nervous system and microscopic lesions of CWD are typical of the TSEs. Brains from CWD-suspect elk and mule deer can be examined by transmission electron microscopy (TEM) for the presence of scrapie-associated fibrils (SAF). Western blot methods identify abnormal prion protein (PrPres) which accumulates in the brains of animals with TSE, including CWD.Conventional SAF purification procedures for TEM examination of CWD-suspect brain tissue require both high speed and ultracentrifugation steps followed by Proteinase K enzyme treatment. Modifications used in this experiment include early Proteinase K, or Proteinase K and Collagenase treatment prior to high speed centrifugation and the elimination of the ultracentrifugation step.

scholarly journals Prion protein in cardiac muscle of elk (Cervus elaphus nelsoni) and white-tailed deer (Odocoileus virginianus) infected with chronic wasting disease

2006 ◽  
Vol 87 (11) ◽  
pp. 3443-3450 ◽  
Author(s):  
Jean E. Jewell ◽  
Jeremy Brown ◽  
Terry Kreeger ◽  
Elizabeth S. Williams
Keyword(s):  
Western Blot ◽  
Odocoileus Virginianus ◽  
Prion Protein ◽  
Cervus Elaphus ◽  
Chronic Wasting Disease ◽  
Striated Muscle ◽  
Mule Deer ◽  
Spongiform Encephalopathy ◽  
Wasting Disease ◽  
Cervus Elaphus Nelsoni

To investigate the possible presence of disease-associated prion protein (PrPd) in striated muscle of chronic wasting disease (CWD)-affected cervids, samples of diaphragm, tongue, heart and three appendicular skeletal muscles from mule deer (Odocoileus hemionus), white-tailed deer (Odocoileus virginianus), elk (Cervus elaphus nelsoni) and moose (Alces alces shirasi) were examined by ELISA, Western immunoblot and immunohistochemistry (IHC). PrPd was detected in samples of heart muscle from seven of 16 CWD-infected white-tailed deer, including one free-ranging deer, and in 12 of 17 CWD-infected elk, but not in any of 13 mule deer samples, nor in the single CWD-infected moose. For white-tailed deer, PrPd was detected by Western blot at multiple sites throughout the heart; IHC results on ventricular sections of both elk and white-tailed deer showed positive staining in cardiac myocytes, but not in conduction tissues or nerve ganglia. Levels of PrPd in cardiac tissues were estimated from Western blot band intensity to be lower than levels found in brain tissue. PrPd was not detected in diaphragm, triceps brachii, semitendinosus, latissiumus dorsi or tongue muscles for any of the study subjects. This is the first report of PrPd in cardiac tissue from transmissible spongiform encephalopathy-infected ruminants in the human food chain and the first demonstration by immunological assays of PrPd in any striated muscle of CWD-infected cervids.

scholarly journals Abundant PrPCWD in Tonsil from Mule Deer with Preclinical Chronic Wasting Disease

2003 ◽  
Vol 15 (4) ◽  
pp. 320-323 ◽  
Author(s):  
Katherine I. O'Rourke ◽  
Dongyue Zhuang ◽  
Amy Lyda ◽  
Gabriel Gomez ◽  
Elizabeth S. Williams ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
High Throughput Screening ◽  
Chronic Wasting Disease ◽  
Transmissible Spongiform Encephalopathy ◽  
Mule Deer ◽  
Screening Tests ◽  
Spongiform Encephalopathy ◽  
Dot Blot ◽  
Wasting Disease ◽  
Dot Blot Assay

A monoclonal antibody dot-blot assay was used to evaluate detergent lysates of tonsil tissue from mule deer to detect PrPCWD, the marker for the cervid transmissible spongiform encephalopathy chronic wasting disease (CWD). Samples of formalin-fixed brain and tonsil tissues from mule deer were examined for PrPCWD using immunohistochemistry (IHC) with Mab F99/97.6.1, the gold standard for diagnosis of preclinical CWD. The contralateral tonsil from each of the 143 deer was prepared for confirmatory IHC and as a 10% (wt/vol) detergent lysate without purification or enrichment steps for monoclonal antibody dot-blot assay. PrPCWD was detected by dot-blot assay in 49 of 50 samples considered positive by IHC. Forty-eight of the positive samples were evaluated with a quantitative dot-blot assay calibrated with recombinant PrP. Tonsillar PrPCWD concentrations ranged from 34 to 1,188 ng per 0.5 mg starting wet weight of tissue. The abundant PrPCWD in mule deer tonsil will facilitate development and validation of high-throughput screening tests for CWD in large populations of free-ranging deer.

scholarly journals Comparison of Abnormal Prion Protein Glycoform Patterns from Transmissible Spongiform Encephalopathy Agent-Infected Deer, Elk, Sheep, and Cattle

2002 ◽  
Vol 76 (23) ◽  
pp. 12365-12368 ◽  
Author(s):  
Richard E. Race ◽  
Anne Raines ◽  
Thierry G. M. Baron ◽  
Michael W. Miller ◽  
Allen Jenny ◽  
...  
Keyword(s):  
Bovine Spongiform Encephalopathy ◽  
Prion Protein ◽  
Chronic Wasting Disease ◽  
Transmissible Spongiform Encephalopathy ◽  
Spongiform Encephalopathy ◽  
Wasting Disease

ABSTRACT Analysis of abnormal prion protein glycoform patterns from chronic wasting disease (CWD)-affected deer and elk, scrapie-affected sheep and cattle, and cattle with bovine spongiform encephalopathy failed to identify patterns capable of reliably distinguishing these transmissible spongiform encephalopathy diseases. However, PrP-res patterns sometimes differed among individual animals, suggesting infection by different or multiple CWD strains in some species.

scholarly journals Chronic Wasting Disease: Evolution of Diagnostic Testing for a Naturally Occurring Prion Disease

2017 ◽  
Author(s):  
Nicholas J. Haley ◽  
Juergen A. Richt
Keyword(s):  
Prion Protein ◽  
Protein Misfolding ◽  
Chronic Wasting Disease ◽  
Mule Deer ◽  
The United States ◽  
Alpha Synuclein ◽  
Wasting Disease ◽  
Disease Evolution ◽  
Naturally Occurring

Since chronic wasting disease (CWD) was first identified nearly 50 years ago in a captive mule deer herd in the Rocky Mountains of the United States, it has slowly spread across North America through the natural and anthropogenic movement of cervids and their carcasses.  As the endemic areas have expanded, so has the need for rapid, sensitive, and cost effective diagnostic tests – especially those which take advantage of samples collected antemortem.  Over the past two decades, strategies have evolved from the recognition of microscopic spongiform pathology and associated immunohistochemical staining of the misfolded prion protein to enzyme-linked immunoassays capable of detecting the abnormal prion conformer in postmortem samples.  In a history that parallels the diagnosis of more conventional infectious agents, both qualitative and real-time amplification assays have recently been developed to detect minute quantities of misfolded prions in a range of biological and environmental samples.  With these more sensitive and semi-quantitative approaches has come a greater understanding of the pathogenesis and epidemiology of this disease in the native host.  Because the molecular pathogenesis of prion protein misfolding is broadly analogous to the misfolding of other pathogenic proteins, including Aβ and α-synuclein, efforts are currently underway to apply these in vitro amplification techniques towards the diagnosis of Alzheimer’s disease, Parkinson’s disease, and other proteinopathies.   Chronic wasting disease – once a rare disease of Colorado mule deer – now represents one of the few naturally occurring protein misfolding disorders which might allow continued development and implementation of novel diagnostic strategies in an animal model.

Distribution of Protease-resistant Prion Protein and Spongiform Encephalopathy in Free-ranging Mule Deer (Odocoileus hemionus) with Chronic Wasting Disease

2002 ◽  
Vol 39 (5) ◽  
pp. 546-556 ◽  
Author(s):  
T. R. Spraker ◽  
R. R. Zink ◽  
B. A. Cummings ◽  
C. J. Sigurdson ◽  
M. W. Miller ◽  
...  
Keyword(s):  
Prion Protein ◽  
Chronic Wasting Disease ◽  
Mule Deer ◽  
Odocoileus Hemionus ◽  
Palatine Tonsil ◽  
Spongiform Encephalopathy ◽  
Motor Nucleus ◽  
Serial Sections ◽  
Wasting Disease ◽  
The Brain

Serial sections of brain and palatine tonsil were examined by immunohistochemical staining (IHC) using monoclonal antibody F89/160.1.5 for detecting protease-resistant prion protein (PrPres) in 35 hunterkilled mule deer ( Odocoileus hemionus) with chronic wasting disease. Serial sections of brain were stained with hematoxylin and eosin and examined for spongiform encephalopathy (SE). Clinical signs of disease were not observed in any of these deer. On the basis of the location and abundance of IHC and the location and severity of SE, deer were placed into four categories. Category 1 ( n = 8) was characterized by IHC in the palatine tonsil with no evidence of IHC or SE in the brain. Category 2 ( n = 13) was characterized by IHC in the palatine tonsil and IHC with or without SE in the dorsal motor nucleus of the vagus nerve (DMNV). Category 3 ( n = 2) was characterized by IHC in the palatine tonsil, IHC with SE in the myelencephalon, and IHC without SE in the hypothalamus. Category 4 ( n = 12) was characterized by IHC in the palatine tonsil and IHC with SE throughout the brain. Category 1 may represent early lymphoid tissue localization of PrPres. The DMNV appears to be the most consistent single neuroanatomic site of detectable PrPres. Categories 2–4 may represent a progression of spread of PrPres and SE throughout the brain. IHC in tonsil and brain and SE in brain were not detected in 208 control deer.

scholarly journals Resistance to Chronic Wasting Disease in Transgenic Mice Expressing a Naturally Occurring Allelic Variant of Deer Prion Protein

2007 ◽  
Vol 81 (9) ◽  
pp. 4533-4539 ◽  
Author(s):  
Kimberly Meade-White ◽  
Brent Race ◽  
Matthew Trifilo ◽  
Alex Bossers ◽  
Cynthia Favara ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Prion Protein ◽  
Prion Disease ◽  
Chronic Wasting Disease ◽  
Transmissible Spongiform Encephalopathy ◽  
Brain Regions ◽  
Spongiform Encephalopathy ◽  
Wasting Disease ◽  
Intestinal Tissue ◽  
Naturally Occurring

ABSTRACT Prion protein (PrP) is a required factor for susceptibility to transmissible spongiform encephalopathy or prion diseases. In transgenic mice, expression of prion protein (PrP) from another species often confers susceptibility to prion disease from that donor species. For example, expression of deer or elk PrP in transgenic mice has induced susceptibility to chronic wasting disease (CWD), the prion disease of cervids. In the current experiments, transgenic mice expressing two naturally occurring allelic variants of deer PrP with either glycine (G) or serine (S) at residue 96 were found to differ in susceptibility to CWD infection. G96 mice were highly susceptible to infection, and disease appeared starting as early as 160 days postinfection. In contrast, S96 mice showed no evidence of disease or generation of disease-associated protease-resistant PrP (PrPres) over a 600-day period. At the time of clinical disease, G96 mice showed typical vacuolar pathology and deposition of PrPres in many brain regions, and in some individuals, extensive neuronal loss and apoptosis were noted in the hippocampus and cerebellum. Extraneural accumulation of PrPres was also noted in spleen and intestinal tissue of clinically ill G96 mice. These results demonstrate the importance of deer PrP polymorphisms in susceptibility to CWD infection. Furthermore, this deer PrP transgenic model is the first to demonstrate extraneural accumulation of PrPres in spleen and intestinal tissue and thus may prove useful in studies of CWD pathogenesis and transmission by oral or other natural routes of infection.

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