No Adaptation of the Prion Strain in a Heterozygous Case of Variant Creutzfeldt-Jakob Disease

We investigated a clinical case of variant Creutzfeldt-Jakob Disease in a person heterozygous for methionine/valine at codon 129 of the prion protein gene and identified the same strain properties in variant Creutzfeldt-Jakob disease in methionine homozygous persons and in bovine spongiform encephalopathy. These results indicate no adaptation of the agent in a different genetic background.

Clinical signs with individual incubation periods ranging from 300 to 392 days postinfection (dpi) were apparent in the mice. The major clinical signs were a loss of body weight and body condition with eye winking and gait abnormalities. Toward the end of the clinical phase, a wet genital area could also be observed. Pathologically confirmed disease developed in 14 of 16 mice (mean incubation period + SEM 341 + 6 dpi). This finding is within the range of previous transmission studies for UK vCJD in this mouse line (mean incubation 306-387 dpi) and similar to those for BSE (mean incubation 316-335 dpi).
We also generated a transmissible spongiform encephalopathy (TSE) vacuolation profile from clinically affected RIII mice and compared it with profiles from UK vCJD and BSE transmissions ( Figure 1). We observed a mild-to-moderate gray matter vacuolation in the medulla, hypothalamus, and septum and moderate vacuolation in the cochlear nucleus and dorsal raphe ( Figure 1; Figure 2, panels A, B).
We conducted an immunohistochemical analysis, which showed abnormal PrP deposition throughout the brain of both a granular and punctate nature (Figure 2, panels C-K). There was heavy staining in the brainstem, particularly the superior vestibular and cochlear nuclei, and lower midbrain, where the substantia nigra was often targeted. Most of the thalamic nuclei exhibited staining, but staining was more intense in the habenular, hypothalamus, and the CA2 region of the hippocampus (Figure 2, panels F-K). Punctate staining was also apparent in the mid-layer of the cortex throughout the brain. This pattern of staining was very similar to that observed in a vCJD and BSE transmission in the United Kingdom, with additional observations of granular deposition in the cerebellar cortex and small plaques occurring in the corpus callosum in 2 of the samples.
Biochemical analysis of the MV isolate confirmed the presence of protease-resistant PrP (PrP res ). We identified a similar type 2B-like pattern and glycosylation profile in the RIII mice. This profile is characterized by a predominance of the diglycosylated form of PrP res at ≈30 kDa, a monoglycosylated form at ≈27kDa, and an unglycosylated band at ≈19kDa (Appendix Figure, https://wwwnc.cdc.gov/EID/ article/26/6/19-1116-App1.pdf). The biochemical profile appears identical between the RIII-MV, RIII-MM, and RIII-BSE isolates tested, although the RIII-BSE isolate appeared to have less PrP res .

Conclusions
This transmission study in RIII mice provides evidence that the prion strain isolated from this confirmed case of vCJD in a 129MV person is the same as that identified in typical 129MM vCJD and BSE cases. Further characterization in a range of mouse models is ongoing. However, transmission to RIII mice in previous studies has led to definitive identification of several strains, vCJD and BSE in particular (6,7).
PRNP codon 129 genotype has been shown to be a major factor influencing disease characteristics of Creutzfeldt-Jakob disease (8), but it has not been established if the same is true of vCJD, because previous vCJD infections in 129MV persons exposed to contaminated blood products have been asymptomatic (9,10). Earlier studies using gene-targeted mice inoculated with vCJD predicted that codon 129 genotype would determine disease susceptibility and incubation periods (11), whereas other transgenic mouse studies demonstrated that BSE could transmit with a different phenotype in mice expressing 129MV than that observed in mice expressing 129MM (12).
The clinical diagnosis in the MV case we report was uncertain while the patient was alive, and it was only at autopsy that neuropathology and biochemistry confirmed vCJD. The neurologic features alone cannot be used to discriminate between sCJD and vCJD, and the MRI findings on DWI imaging favored a diagnosis of sCJD. However, the high sensitivity and specificity of MRI for vCJD were determined by analyzing FLAIR images primarily (13) and recent review suggests that DWI imaging may be less specific than FLAIR imaging in vCJD. It is possible that the phenotype of vCJD in this case may have been altered by the heterozygous PRNP background and investigations including CSF protein misfolding cyclic amplification (2), tonsil biopsy, and perhaps FLAIR MRI may contribute to accurate diagnosis of future heterozygous cases.
The identification of vCJD in a 129MV person may indicate the start of a second wave of vCJD in association with the 129MV genotype which is present in around 45% of the UK population (14), although no further cases have been reported since 2016. This case highlights the need to continue surveillance to identify new cases of vCJD and the need for autopsy and strain typing in persons with prion diseases. Changes in clinical disease phenotype could mask the true diagnosis and may be indicative of potential changes in prion disease strains and infectious properties. Strain identification and assessing the infectious properties of prion diseases are essential components in the management of these diseases and have important implications for public health and in determining the prevalence of BSE-related prion disease in humans.