Journal of Pathology
J Pathol 2004; 203: 733–739
Published online 21 May 2004 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/path.1580
Prevalence of lymphoreticular prion protein
accumulation in UK tissue samples
David A Hilton,1* Azra C Ghani,2Lisa Conyers,1Philip Edwards,1Linda McCardle,3Diane Ritchie,3
Mark Penney,1Doha Hegazy1and James W Ironside3
1Department of Histopathology, Derriford Hospital, Plymouth, UK
2Department of Infectious Disease Epidemiology, Faculty of Medicine, Imperial College, London, UK
3National CJD Surveillance Unit, University of Edinburgh, Edinburgh, UK
Dr David A Hilton, Department
of Histopathology, Derriford
Hospital, Plymouth, PL6
Received: 19 February 2004
Revised: 15 March 2004
Accepted: 22 March 2004
This study aims to provide an estimate of the number of individuals in the UK who may
be incubating variant Creutzfeldt-Jakob disease and at risk of causing iatrogenic spread
of the disease. Lymphoreticular accumulation of prion protein is a consistent feature of
variant Creutzfeldt-Jakob at autopsy and has also been demonstrated in the pre-clinical
phase. Immunohistochemical accumulation of prion protein in the lymphoreticular system
remains the only technique that has been shown to predict neurological disease reliably
in animal prion disorders. In this study, immunohistochemistry was used to demonstrate
the presence of prion protein, with monoclonal antibodies KG9 and 3F4, in surgically
removed tonsillectomy and appendicectomy specimens. The samples were collected from
histopathology departments across the UK and anonymised prior to testing. Samples were
tested from 16703 patients (14964 appendectomies, 1739 tonsillectomies), approximately
60% of whom were from the age group 20–29 years at operation. Twenty-five per cent of the
samples were excluded from the final analyses because they contained inadequate amounts
of lymphoid tissue. Three appendicectomy samples showed lymphoreticular accumulation
of prion protein, giving an estimated prevalence of 3/12674 or 237 per million (95%
CI 49–692 per million). The pattern of lymphoreticular accumulation in two of these
samples was dissimilar from that seen in known cases of variant Creutzfeldt-Jakob disease.
Although it is uncertain whether immunohistochemical accumulation of prion protein in
the lymphoreticular system is specific for variant Creutzfeldt-Jakob disease, it has not been
described in any other disease, including other forms of human prion disease or a range of
inflammatory and infective conditions. These findings reinforce the importance of measures
taken by the UK Department of Health to reduce the risk of spread of variant Creutzfeldt-
Jakob via blood products and surgical instruments, and of the urgency to proceed with
large-scale screening of fresh tonsil specimens for the presence of prion protein.
Copyright 2004 Pathological Society of Great Britain and Ireland. Published by John
Wiley & Sons, Ltd.
Keywords: Creutzfeldt-Jakob disease (CJD); prion; screening; immunohistochemistry
Variant Creutzfeldt-Jakob disease (vCJD) was first
recognized as a new and distinctive disease in the UK
in 1996 . Subsequent transmission studies and strain
typing have shown that the transmissible agent causing
vCJD exhibits identical characteristics to the bovine
spongiform encephalopathy (BSE) agent [2–4] and
there is no evidence that vCJD occurred prior to 1995
[5,6]. These data indicate that vCJD is a new disease,
almost certainly caused by exposure to the BSE agent.
This conclusion has led to concern about a possible
human epidemic of vCJD, particularly as it is likely
that over 400000 infected cattle entered the human
food chain in the UK prior to the introduction of the
specified bovine offal ban in November 1989 and as
the ban was not fully effective for several years, a large
number of infected cattle also entered the food chain
in the early 1990s . There have been a number of
attempts to predict future numbers of vCJD cases using
mathematical models and extrapolating from vCJD
cases seen to date [8–13]. Recent estimates based on
the pattern of clinical cases suggest that the epidemic
of vCJD will be relatively small, with an upper 95%
confidence interval of 540 future cases . However,
remaining uncertainties, including the possibility that
other genetic loci affect susceptibility , make
the distribution and timing of any human epidemic
unclear. Furthermore, such models are unable to
estimate the prevalence of asymptomatic infection and
hence provide any estimate of the potential number
of future infections and cases that could arise from
secondary (human-to-human) transmission of vCJD.
In addition, questions have been raised as to the
Copyright 2004 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
PrP accumulation in UK tissue samples739
22. Hill AF, Butterworth RJ, Joiner S, et al. Investigation of variant
Creutzfeldt-Jakob disease and other human prion diseases with
tonsil biopsy samples. Lancet 1999; 353: 183–189.
23. Ironside JW, McCardle L, Horsburgh A, Lim Z, Head MW.
Pathological diagnosis of variant Creutzfeldt-Jakob disease.
APMIS 2002; 110: 79–87.
24. Hilton DA, Fathers E, Edwards P, Ironside JW, Zajicek J. Prion
immunoreactivity in appendix before clinical onset of variant
Creutzfeldt-Jakob disease. Lancet 1998; 352: 703–704.
25. Hilton DA, Ghani AC, Conyers L, et al. Accumulation of prion
protein in tonsil and appendix: review of tissue samples. Br Med
J 2002; 325: 633–634.
26. Schreuder BE, vanKeulen LJ,
Smits MA. Tonsillar biopsy and PrPSc detection in the preclinical
diagnosis of scrapie. Vet Rec 1998; 142: 564–568.
27. O’Rourke KI, Baszler TV, Besser TE, et al. Preclinical diagnosis
of scrapie by immunohistochemistry of third eyelid lymphoid
tissue. J Clin Microbiol 2000; 38: 3254–3259.
28. Ironside JW, Hilton DA, Ghani A, et al. Retrospective study of
prion-protein accumulation in tonsil and appendix tissues. Lancet
2000; 355: 1693–1694.
29. Hilton DA. vCJD — predicting the future? Neuropathol Appl
Neurobiol 2000; 26: 405–407.
30. Sabattini E, Bisgaard K, Ascani S,
system: a new immunohistochemical method for diagnostics
and research. Critical comparison with the APAAP, ChemMate,
CSA, LABC, and SABC techniques. J Clin Pathol 1998; 51:
31. Muramoto T, Kitamoto T, Tateishi J, Goto I. Accumulation of
abnormal prion protein in mice infected with Creutzfeldt-Jakob
disease via intraperitoneal route: a sequential study. Am J Pathol
1993; 143: 1470–1479.
32. Ford MJ, Burton LJ, Morris RJ, Hall SM. Selective expression of
prion protein in peripheral tissues of the adult mouse. Neuroscience
2002; 113: 177–192.
33. Hilton DA, Sutak J, Smith MEF, et al. Specificity of lymphoretic-
ular accumulation of prion protein for variant Creutzfeldt-Jakob
disease. J Clin Pathol 2004; 57: 300–302.
34. Head MW, Ritchie D, Smith N, et al. Peripheral tissue involve-
ment in sporadic, iatrogenic, and variant Creutzfeldt-Jakob disease:
an immunohistochemical, quantitative, and biochemical study. Am
J Pathol 2004; 164: 143–153.
35. Parchi P, Giese A, Capellari S, et al. Classification of sporadic
Creutzfeldt-Jakob disease based on molecular and phenotypic
analysis of 300 subjects. Ann Neurol 1999; 46: 224–233.
36. Chou I. Strain of unknown prions weighs heavily on Japan, Italy.
Nature Med 2003; 9: 1442.
Vromans ME, Langeveld JP,
et al. The EnVision++
37. Brown P,
spongiform encephalopathy: the National Institutes of Health series
of 300 cases of experimentally transmitted disease. Ann Neurol
1994; 35: 513–529.
38. Korth C, Stierli B, Streit P, et al. Prion (PrPSc)-specific epitope
defined by a monoclonal antibody. Nature 1997; 390: 74–77.
39. Paramithiotis E, Pinard M, Lawton T, et al. A prion protein
epitope selective for the pathologically misfolded conformation.
Nature Med 2003; 9: 893–899.
40. Race R, Raines A, Raymond GJ, Caughey B, Chesebro B. Long-
term subclinical carrier state precedes scrapie replication and
adaptation in a resistant species: analogies to bovine spongiform
encephalopathy and variant Creutzfeldt-Jakob disease in humans.
J Virol 2001; 75: 10106–10112.
41. Asante EA, Linehan JM,Desbruslais M,
propagate as either variant CJD-like or sporadic CJD-like prion
strains in transgenic mice expressing human prion protein. EMBO
J 2002; 21: 6358–6366.
42. Bruce ME, McConnell I, Will RG, Ironside JW. Detection of
variant Creutzfeldt-Jakob disease infectivity in extraneural tissues.
Lancet 2001; 358: 208–209.
43. Taylor DM, Fraser H, McConnell I, et al. Decontamination studies
with the agents of bovine spongiform encephalopathy and scrapie.
Arch Virol 1994; 139: 313–326.
44. Brown P, Preece MA, Will RG. ‘Friendly fire’ in medicine:
hormones, homografts, and Creutzfeldt-Jakob disease. Lancet
1992; 340: 24–27.
45. Glatzel M, Abela E, Maissen M, Aguzzi A. Extraneural patho-
logic prion protein in sporadic Creutzfeldt-Jakob disease. N Engl
J Med 2003; 349: 1812–1820.
46. Kovacs GG, Lindeck-Pozza E, Chimelli L, et al. Creutzfeldt-
Jakob disease and inclusion body myositis: abundant disease-
associated prion protein in muscle. Ann Neurol 2004; 55:
47. Collins S, Law MG, Fletcher A, Boyd A, Kaldor J, Masters CL.
Surgical treatment and risk of sporadic Creutzfeldt-Jakob disease:
a case–control study. Lancet 1999; 353: 693–697.
48. Ward HJ, Everington D, Croes EA, et al. Sporadic Creutzfeldt-
Jakob disease and surgery: a case–control study using community
controls. Neurology 2002; 59: 543–548.
49. Hunter N, Foster J, Chong A, et al. Transmission of prion diseases
by blood transfusion. J Gen Virol 2002; 83: 2897–2905.
50. Llewelyn CA, Hewitt PE, Knight RSG, et al. Possible transmis-
sion of variant Creutzfeldt-Jakob disease by blood transfusion.
Lancet 2004; 363: 417–421.
51. Collinge J. Variant Creutzfeldt-Jakob disease. Lancet 1999; 354:
Gibbs CJ Jr, Rodgers-Johnson P,
et al. Human
et al. BSEprions
J Pathol 2004; 203: 733–739