Figure 1.
Expulsion of the lypholized CWD/Mte inoculum from a transfer pipette illustrating the ultrafine texture of the inoculum.
Figure 2.
Quantification of deer CWD prion by the cervid prion cell assay.
A. Representative wells of an ELISPOT plate showing spots given by triplicate deer 5E9-S1cells exposed to 3-fold serial dilutions of CWD+ inoculate brain homogenate, deer CWD (upper) and Tg(CerPrP-M132)1536+/−-passaged CWD prion isolate 012-09442 (lower), the concentration of brain homogenates (left to right) are: 10−2, 10−2.5, 10−2.9, 10−3.4, 10−3.9, 10−4.4 and 10−4.9. The wells at right are uninfected Deer5E9-S1 cells. B. Responsiveness of Deer5E9-S1 cells to the CWD+ inoculum and Tg(CerPrP-M132)1536+/− mouse -passaged CWD prion isolate 012-09442 were between 10−2 and 10−5. The cells were infected with serial 1∶3 dilutions of homogenates of CWD+ inoculum and subjected to the CPCA. In each case, the mean is derived from experiments performed in triplicate, with error bars indicating the standard errors of the means (SEM).
Figure 3.
Lymphoid Involvement at each of the 4 time points.
No significant difference was detected in the number of affected follicles between the groups (ANOVA, p = 0. 86).
Table 1.
Detection of CWD in lymphoid follicles.
Figure 4.
Prnp genotype affects PrPCWD distribution and proportion of PrPCWD positive follicles in the lympoid system.
The GG genotype had a wider PrPCWD distribution than GS and SS. *Tissue CWD % was significantly less than the GG genotype. ♦ Tissue CWD % was significantly less than the GS genotype. (Students one tailed T-test, p≤0.05).
Figure 5.
PrPCWD immunolabeling in the retropharyngeal and submandibular lymph nodes at 200× magnification, mAb F99/97.6.1 and alkaline phosphatase detection.
Figure 6.
Visualization of GFD/Mte particles.
6A. Longitudinal cut of a deer head without GFD/Mte under a blacklight. Inset- dissected nasal turbinates. 6B. Longitudinal cut of a deer head inoculated with GFD/Mte under a black light with visible GFD/Mte deposition. Insets- dissected nasal turbinates 6C. Ethmoid nasal turbinates under a blacklight. Arrows and circles indicate small particles of GFD/Mte. 6D. Mounted nasal turbinate with 100× magnification showing GFD/Mte particles associated with the nasal mucosa under normal light and with a ultraviolet filter. OE-Olfactory epithelium, NC-Nasal Cavity. Arrows indicate GFD/Mte (6E).
Figure 7.
Tracking lyophilized prion inocula in the nasal cavity of deer.
Highly enriched, fluorescently labeled prions were mixed with Mte, lyophilized, pulverized and puffed into the nasal cavity. (A) After 45 minutes, florescent prion aggregates (red) can be seen on (white arrowheads) and within the olfactory epithelium (OE) of the nasal turbinates. Tissue sections are counterstained with DiOC18 fluorescent membrane dye (green) and the nuclear stain DAPI (blue). A small proportion of prions can be seen near serous cells of the Bowman's glands (BG) in the lamina propria. (B) By 60 min, significant amount of prions were found in the lamina propria, with some aggregates (arrowhead) associated with nerve fibers (NF) emanating from the OE. (C) We detected no red signal from negative control sections from mock-inoculated deer. (D–G) Higher magnification of a Bowman's gland stained with DAPI (D), DiOC18 (E) and decorated with prions (F) that appear to localize on serous cells (G). NB, nerve bundle; scale bar, 20 µm.