Transmissible Spongiform Encephalopathies

Etiology

The TSEs are transmissible to susceptible animals by inoculation of tissues from affected subjects. Although the infectious agents replicate in some cell cultures, they do not achieve the high titers of infectivity found in brain tissues or cause recognizable cytopathic effects in cultures. Most previous studies of TSE agents have used in vivo assays, relying on the transmission of typical neurologic disease to animals as evidence that the agent was present and intact. Inoculation of susceptible recipient animals with small amounts of the infectious TSE agent results, months later, in the accumulation in tissues of large amounts of the agent with the same physical and biologic properties as the original agent. The TSE agents display a spectrum of extreme resistance to inactivation by a variety of chemical and physical treatments that is unknown among conventional pathogens. This characteristic, as well as their partial sensitivity to protein-disrupting treatments and their consistent association with abnormal isoforms of a normal host-encoded protein (prion protein or PrP), stimulated the hypothesis that the TSE agents are probably subviral in size, composed of protein, and devoid of nucleic acid.

The term prion (for proteinaceous infectious agent), coined by S.B. Prusiner, is now widely used for such agents. The prion hypothesis proposes that the molecular mechanism by which the pathogen-specific information of TSE agents is propagated involves a self-replicating change in the folding host-encoded PrP associated with a transition from an α-helix–rich structure in the native protease-sensitive conformation (cellular PrP or PrP ^C ) to a β-sheet–rich structure in the protease-resistant conformation associated with infectivity. The existence of a second host-encoded protein—termed protein X—that participates in the transformation was also postulated to explain certain otherwise puzzling findings but has never been identified.

The prion hypothesis is still not universally accepted; it relies on the postulated existence of a genome-like coding mechanism based on differences in protein folding that have not been satisfactorily explained at a molecular level. In addition, it has yet to account convincingly for the many biologic strains of TSE agent that have been observed, although strain-specific differences in the abnormal forms of the PrP have been found and proposed as providing a plausible molecular basis for the coding. It fails to explain why pure PrP uncontaminated with nucleic acid from an infected host has not transmitted a convincingly typical spongiform encephalopathy consistently associated with a serially self-propagating agent. A finding that was also troubling, in several experimental models and human illnesses, was that abnormal PrP and infectivity were not consistently associated. Particularly problematic is the finding that some illnesses associated with mutations in the PRNP gene and accompanied by abnormal PrP failed to transmit infection to animals. If the TSE agents ultimately prove to consist of protein and only protein, without any obligatory nucleic acid component, then the term prion will indeed be appropriate and the early proponents of the prion hypothesis will prove to have been prescient. If the agents are ultimately found to contain small nucleic acid genomes, then they might better be considered atypical viruses, for which the term virino has been suggested. Until the actual molecular structure of the infectious TSE pathogens and the presence or absence of a nucleic acid genome are rigorously established, it seems less contentious to continue calling them TSE agents, although most authorities have accepted the term prion (sometimes referring to the agent of a TSE and sometimes to the abnormal protein, even when nontransmissible).

The earliest evidence that abnormal proteins are associated with the TSE was morphologic: Scrapie-associated fibrils were found in extracts of tissues from patients and animals with spongiform encephalopathies but not in normal tissues. Scrapie-associated fibrils resemble but are distinguishable from the amyloid fibrils that accumulate in the brains of patients with Alzheimer disease. A group of antigenically related protease-resistant proteins (PrPs) proved to be components of scrapie-associated fibrils and to be present in the amyloid plaques found in the brains of patients and animals with TSEs. The abnormal forms of PrP are variously designated PrP ^Sc (scrapie-type PrP), PrP-res (protease-resistant PrP), PrP ^TSE (TSE-associated PrP), or PrP ^D (disease-associated PrP) by different authorities.

It remains unclear whether abnormal PrP constitutes the complete infectious particle of spongiform encephalopathies, is a component of those particles, or is a pathologic host protein not usually separated from the actual infectious entity by currently used techniques. The demonstration that PrP is encoded by a normal host gene seemed to favor the last possibility. Several studies suggest that agent-specific pathogenic information can be transmitted and replicated by different conformations of a protein with the same primary amino acid sequence in the absence of agent-specific nucleic acids. Properties of two fungal proteins were found to be heritable without encoding in nucleic acid, although those properties have not been naturally transmitted to recipient fungi as infectious elements. Whatever its relationship to the actual infectious TSE particles, PrP clearly plays a central role in the susceptibility to infection, because the normal PrP must be expressed in mice and cattle if they are to acquire a TSE or to sustain replication of the infectious agents. Furthermore, inherited normal variations in the PrP phenotype are associated with increased susceptibility to vCJD and (to a lesser extent) to sCJD and with occurrence of familial TSEs (fCJD and GSS).

PrPs are glycoproteins; protease-resistant PrPs, when aggregated, have the physical properties of amyloid proteins. The PrPs of different species of animals are very similar in their amino acid sequences and antigenicity but are not identical in structure. The primary structure of PrP is encoded by the host and is not altered by the source of the infectious agent provoking its formation. The function of the ubiquitous protease-sensitive PrP precursor (designated PrP ^C , for cellular PrP, or PrP-sen, for protease-sensitive PrP) in normal cells is unknown; it binds copper and may play some role in normal synaptic transmission, but it is not required for life or for relatively normal cerebral function in mice and cattle. As noted, animals must express PrP to develop scrapie disease and to support replication of the TSE agents. The degree of homology between amino acid sequences of PrPs in different animal species may correlate with the species barrier that affects the susceptibility of animals of one species to infection with a TSE agent adapted to grow in another species, although the degree of sequence homology does not always predict susceptibility to the same TSE agent.

Attempts to find particles resembling those of viruses or virus-like agents in brain tissues of humans or animals with spongiform encephalopathies have been unsuccessful. Peculiar tubulovesicular structures reminiscent of some viruses have been seen repeatedly in thin sections of TSE-infected brain tissues and cultured cells but not in normal cells. It has never been established that those structures are associated with infectivity.

Epidemiology

Kuru once affected many children of both sexes ≥ 4 yr of age, adolescents, and young adults (mainly females) living in one limited area of Papua New Guinea. The complete disappearance of kuru among people born after 1957 suggests that the practice of ritual cannibalism (thought to have ended that year) was probably the only mechanism by which the infection spread in Papua New Guinea.

CJD, the most common human spongiform encephalopathy, was formerly thought to occur only in older adults; however, iCJD and, much more rarely, sCJD (to date, seven reports in adolescents, one a 14 yr old female) have affected young people. A single case of sporadic fatal insomnia was recognized in a U.S. adolescent. GSS has not been diagnosed in children or adolescents. vCJD has a peculiar predilection for younger people. Of 174 cases of vCJD reported through 2010 in the United Kingdom, all except 23 were in people younger than 40 yr of age and 22 were in people younger than 20 yr of age; the youngest age at onset was 12 yr. sCJD has been recognized worldwide, at yearly rates of 0.25-2 cases/million population (not age-adjusted), with CJD foci of considerably higher incidence among Libyan Jews in Israel, in isolated villages of Slovakia, and in other limited areas. Sporadic CJD has not been convincingly linked to any common exposure, and the source of infection remains unknown. Proponents of the prion hypothesis are convinced that PrP can spontaneously misfold, becoming self-replicating and causing sCJD; skeptics favor infection with some ubiquitous TSE agent, which, fortunately, has a very low attack rate except in persons with certain mutations in the PRNP gene. Neither of those possible etiologies has been proven. Person-to-person spread has been confirmed only for iatrogenic cases. Spouses and household contacts of patients are not at risk of acquiring CJD, although two instances of conjugal CJD have been reported. However, medical personnel exposed to brains of patients with CJD may be at some increased risk; at least 20 healthcare workers have been recognized with the disease.

The striking resemblance of CJD to scrapie prompted a concern that infected sheep tissues might be a source of spongiform encephalopathy in humans. No reliable epidemiologic evidence suggests that exposure to potentially scrapie-contaminated animals, meat, meat products, or experimental preparations of the scrapie agent have transmitted a TSE to humans. The potential of the CWD agent to infect human beings has also not been demonstrated but remains under investigation; deer, elk, and moose in 15 U.S. states and 2 Canadian provinces have been naturally infected; cases of CWD were recently detected in wild reindeer and moose (European elk) in Norway and Finland. Consumption of contaminated meat, including venison from animals infected with the CWD agent, has not been implicated as a risk factor for human TSE by epidemiologic studies; however, a recent unpublished study requiring several years yielded evidence that CWD was experimentally transmitted to monkeys fed venison from overtly healthy infected deer, prompting a health advisory from Canadian authorities. The outbreak of BSE among cattle (possibly infected by eating scrapie-agent–contaminated meat-and-bone meal added to feed) was first recognized in the United Kingdom in 1986 and later reported in cattle of 27 other countries, including Canada and the United States. More than 190,000 cases of BSE have been reported to the World Organization for Animal Health (OIE), almost 97% of those from the United Kingdom. Cases in the United Kingdom progressively declined after 1992 and later in other countries; in 2016 only 2 cases worldwide were reported to OIE (from France and Spain) and none from the United Kingdom. The finding of a new TSE in ungulate and feline animals in British zoos and later in domestic cats raised a fear that the BSE agent had acquired a range of susceptible hosts broader than that of scrapie, posing a potential danger for humans. That remains the most plausible explanation for the occurrence of vCJD, first described in adolescents in Britain in 1996 and, as of May 2017, eventually affecting at least 178 people in the United Kingdom. (not counting a disturbing number of people with evidence of possible asymptomatic or “preclinical” vCJD infection) and more than 50 in 11 other countries (total 231 cases worldwide): 27 in France, 5 in Spain, 4 in Ireland, 3 in the Netherlands, 2 each in Italy and Portugal, and single cases in Japan and Saudi Arabia. Variant CJD has also occurred in former U.K. residents (>6 mo) living in Ireland (two cases), France (one case), Canada (one case), Taiwan (one case), and the United States (two cases). Two cases of vCJD—one in the United States and one in Canada—have been reported in former long-time residents of Saudi Arabia, a country that has not recognized BSE but might have imported contaminated meat products from the United Kingdom. A third case of vCJD was previously confirmed in a Saudi citizen residing in Saudi Arabia. The most recent case of vCJD diagnosed in the United States occurred in an immigrant deemed by the CDC to have most likely been infected during early years spent in Kuwait.

No case of vCJD has been confirmed in anyone born in the United Kingdom after 1989. However, examination of resected appendixes in the United Kingdom for evidence of subclinical infection with prions suggested that about 1 in 2,000 people tested had a detectable accumulation of PrP ^TSE in lymphoid follicles. It remains controversial whether those accumulations resulted from subclinical vCJD or another TSE; none of the subjects to date has presented to medical attention with overt TSE.

Iatrogenic transmissions of CJD have been recognized for more than 30 yr ( Table 304.2 ). Such accidental transmissions of CJD have been attributed to use of contaminated neurosurgical instruments (no case reported since 1980) or operating facilities, use of cortical electrodes contaminated during epilepsy surgery, injections of human cadaveric pituitary growth hormone and gonadotropin (no longer marketed in the United States), and transplantation of contaminated corneas and allografts of human dura mater, still in limited use in the United States as a surgical patching material. Pharmaceuticals and tissue grafts derived from or contaminated with human neural tissues, particularly if obtained from unselected donors and large pools of donors, pose special risks.

Studies of animals experimentally infected with TSE agents first suggested that blood and blood components from humans with preclinical CJD infections might pose a risk of transmitting disease to recipients, and since the 1980s such blood components have been withdrawn as a precaution in the United States when a donor was later found to have CJD and blood products were still in-date. A surveillance program in the United Kingdom reported vCJD in three recipients of nonleukoreduced red blood cells from donors later diagnosed with vCJD; there was autopsy evidence of a preclinical vCJD infection in a fourth red cell recipient who died of another disease. (vCJD has not occurred in any recipient of leukoreduced red blood cells from a donor who later developed vCJD.) A study conducted over more than 20 yr by the American Red Cross and CDC found no recipient of blood components obtained from donors later diagnosed with sporadic CJD (and from one donor with familial CJD) developed a TSE.

Evidence of a preclinical vCJD infection was found at autopsy in a U.K. patient with hemophilia A treated with a human plasma–derived coagulation factor VIII to which at least one vCJD-infected donor contributed; the coagulation factor involved was never licensed in the United States. U.K. authorities have described two recipients of plasma-derived coagulation factors (both having a history of a transfusion with blood components, as well) who later developed sporadic CJD, concluding that the finding, while of concern, might be coincidental.

Pathogenesis and Pathology

The probable portal of entry for the TSE agent in kuru is thought to have been either through the gastrointestinal tract or lesions in the mouth or integument incidentally exposed to the agent during cannibalism. Patients with vCJD (and animals with BSE and BSE-related TSEs) are thought to have been similarly infected with the BSE agent by consuming contaminated beef products. Except after direct introduction into the nervous system, the first site of replication of TSE agents appears to be in tissues of the reticuloendothelial system. TSE agents have been detected in low titers in the blood of experimentally infected animals (mice, monkeys, hamsters, and sheep and in the blood of persons with vCJD and perhaps sCJD); infectivity was mainly associated with nucleated cells, although the plasma contained a substantial portion of total infectivity in blood. Circulating lymphoid cells seem to be required to infect mice by peripheral routes. Limited evidence suggests that TSE agents also spread to the central nervous system by ascending peripheral nerves. Several research groups claimed to detect the CJD agent in human blood, although other attempts failed.

In human kuru, it seems probable that the only portal of exit of the agent from the body, at least in quantities sufficient to infect others, was through infected tissues exposed during cannibalism. In iatrogenically transmitted CJD, the brains and eyes of patients with CJD have been the probable sources of contamination. Experimental transmission of the agent to animals from the kidney, liver, lung, lymph node, and spleen showed that those tissues as well as the cerebrospinal fluid (CSF) sometimes contain the CJD agent; none of those sources has been implicated in accidental transmission of CJD to humans. At no time during the course of any TSE have antibodies or cell-mediated immunity to the infectious agents been convincingly demonstrated in either patients or animals. However, mice must be immunologically competent to be infected with the scrapie agent by peripheral routes of inoculation.

Typical changes in TSE include vacuolation and loss of neurons with hypertrophy and proliferation of glial cells, most pronounced in the cerebral cortex in patients with CJD and in the cerebellum in those with kuru. The central nervous system lesions are usually most severe in or even confined to gray matter, at least early in the disease. Loss of myelin appears to be secondary to the degeneration of neurons. There generally is no inflammation, but a marked increase in the number and size of astrocytes is usual. Spongiform changes are not a striking autopsy finding in patients with FFI, and neuronal degeneration and gliosis are largely restricted to thalamic nuclei.

Amyloid plaques are found in the brains of all patients with GSS and in at least 70% of those with kuru. These plaques are less common in patients with CJD. Amyloid plaques are most common in the cerebellum but occur elsewhere in the brain, as well. In brains of patients with vCJD, plaques surrounded by halos of vacuoles (described as flower-like or florid plaques) have been a consistent finding. TSE amyloid plaques react with antiserum prepared against PrP. Even in the absence of plaques, extracellular PrP can be detected in the brain parenchyma by immunostaining.