Prion
ISSN: 1933-6896 (Print) 1933-690X (Online) Journal homepage: http://www.tandfonline.com/loi/kprn20
Exploring the risks of a putative transmission of BSE to new species Enric Vidal, Natalia Fernández-Borges, Belén Pintado, Montserrat Ordóñez, Mercedes Márquez, Dolors Fondevila, Hasier Eraña, Juan María Torres, Martí Pumarola & Joaquín Castilla To cite this article: Enric Vidal, Natalia Fernández-Borges, Belén Pintado, Montserrat Ordóñez, Mercedes Márquez, Dolors Fondevila, Hasier Eraña, Juan María Torres, Martí Pumarola & Joaquín Castilla (2013) Exploring the risks of a putative transmission of BSE to new species, Prion, 7:6, 443-446, DOI: 10.4161/pri.27014 To link to this article: http://dx.doi.org/10.4161/pri.27014
Published online: 01 Nov 2013.
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Prion 7:6, 443–446; November/December 2013; © 2013 Landes Bioscience
Exploring the risks of a putative transmission of BSE to new species Centre de Recerca en Sanitat Animal (CReSA); UAB-IRTA; Campus de la Universitat Autònoma de Barcelona; Barcelona, Spain; 2CIC bioGUNE; Parque tecnológico de Bizkaia; Bizkaia, Spain; 3Centro Nacional de Biotecnología (CNB); Campus de Cantoblanco; Madrid, Spain; 4 Department of Animal Medicine and Surgery; Veterinary faculty; Universitat Autònoma de Barcelona (UAB); Barcelona, Spain; 5CISA—INIA; Madrid, Spain; 6 IKERBASQUE; Basque Foundation for Science; Bizkaia, Spain
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Keywords: prion, BSE, scrapie, PMCA, transmission barrier, prion resistant species *Correspondence to: Joaquín Castilla; Email: [email protected] Submitted: 08/16/2013 Revised: 10/01/2013 Accepted: 10/31/2013 http://dx.doi.org/10.4161/pri.27014 Addendum to : Vidal E, Fernández-Borges N, Pintado B, Ordóñez M, Márquez M, Fondevila D, Torres JM, Pumarola M, Castilla J. Bovine spongiform encephalopathy induces misfolding of alleged prion-resistant species cellular prion protein without altering its pathobiological features. J Neurosci 2013; 33:7778-86; http:// dx.doi.org/10.1523/JNEUROSCI.0244-13.2013; PMID:23637170
he prion responsible for the Bovine Spongiform Encephalopathy (BSE) shows unique features when compared with other prions. One of these features is its ability to infect almost all experimentally tested animal models. In the paper published in The Journal of Neuroscience1 we describe a series of experiments directed toward elucidating which would be the in vivo behavior of BSE if it would infect dogs and rabbits, two alleged prion resistant species. Protein misfolding cyclic amplification (PMCA) was used to generate canidae and leporidae in vitro adapted BSE prions. A characterization of their in vivo pathobiological properties showed that BSE prions were capable not only of adapting to new species but they maintained, in the case of rabbits, their ability to infect transgenic mice expressing human PrP. The remarkable adaptation ability of certain prions implies that any new host species could lead to the emergence of new infectious agents with unpredictable transmission potential. Our results suggest that caution must be taken when considering the use of any mammal-derived protein in feedstuffs.
Perspectives Bovine Spongiform Encephalopathy (BSE) is one of the most well-known animal prion diseases.2 The “mad cow disease” outbreak had unprecedented consequences regarding worldwide public
health and food safety policies. The reason for this was the ability of the agent causing BSE to transmit to species other than cattle, its original host, including human beings.3 Enormous economical efforts were directed toward controlling the disease in cattle with exceptional results. Indeed, policies such as the ban on using ruminant protein on feed drastically reduced the reported BSE cases in cattle. The compulsory testing of a large number of animals allowed to accurately monitoring the success of such measures. In every country where these policies have been implemented, the BSE cases are practically down to zero. Thus, updated risk analyses point to a relaxation of the control measures. Thanks to the research efforts directed toward this subject, many previously unknown aspects of this prion disease have came to light during the last decade, for instance, the existence of the so called atypical BSE cases.4,5 Two cattle prions strains have been identified in aged cattle and named BSE-L and BSE-H after the different electrophoretic pattern of its proteinase-K digested disease-associated BSE prion protein (PrPd) on western blotting. Epidemiology and pathological features of the diagnosed cases suggest that those are probably sporadic prion diseases and evidence suggests that they might have had a crucial role in the origin of the BSE epizooty.6,7 If these atypical presentations are indeed sporadic prion diseases, such as the ones known in humans, it implies that current BSE control measures will not be effective to eradicate them. In other
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© 2013 Landes Bioscience. Do not distribute.
Enric Vidal1, Natalia Fernández-Borges2, Belén Pintado3, Montserrat Ordóñez1, Mercedes Márquez4, Dolors Fondevila4, Hasier Eraña 2, Juan María Torres5, Martí Pumarola4 and Joaquín Castilla 2,6*
CWD prions could readily infect hamsters.21 BSE has in fact been described to change, to a certain extent, upon passage through species other than cattle, i.e., sheep. Passage through sheep resulted in an increase of BSE virulence when inoculated into TgBov mice.17 So, attention must be paid to the emergence of new prion strains or the modifications that new hosts might pose to the pathobiological properties of already known strains, particularly regarding the ability to transmit to other species, including humans. In vitro amplification of prions22 has proven to be an effective tool to predict the in vivo behavior of certain prions.23,24 Not only does it accelerate the transmission process but it also can be used to push the system, by challenging healthy brain homogenates of different species with different prion strains at high concentration and directly in a test tube, to find out what would happen in case certain cross-species transmission of prions would occur in nature. Several publications support protein misfolding cyclic amplification (PMCA) ability to mimic potential in vivo scenarios. For instance, the susceptibility of rabbits to prion diseases was predicted by PMCA and then demonstrated in its natural host23 and, likewise, other species PrP could also be misfolded in vitro including dogs and horses.25 In the paper published in The Journal of Neuroscience1 we describe a series of experiments directed toward elucidating which would be the in vivo behavior of BSE, one of the most relevant prions in Europe, in case it managed to cross certain transmission barriers. The species chosen were dogs and rabbits for several reasons, namely both are species in close contact with human beings and no reports of prion susceptibility had been published at the time the experiments were designed, thus they were species considered to be probably prion resistant. The experiments consisted in an initial in vitro phase in which dog and rabbit normal brain homogenates were subjected to PMCA with a BSE seed. In a second phase, the newly obtained prions were inoculated into TgBov mice and in a transgenic model expressing human PrP (TgHum)26 to assess their in vivo
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behavior in comparison to the original BSE seed. Several interesting findings were obtained from that set of experiments. To start with, BSE was the only prion strain that could be propagated in vitro in both alleged resistant species, a capacity which lacked other prion strains originated in other hosts, such as scrapie. The typical BSE biochemical signature (western blotting) remained unchanged in both species even after multiple in vitro amplification cycles. Interestingly, the pathobiological features of these newly in vitro obtained prions, such as their infectivity, incubation times, biochemical profile, lesion profile, and etcetera were remarkably comparable to those of BSE when bioassayed in transgenic mouse models overexpressing bovine and human PRNP. Of particular interest were the bioassay results in tgHum mice that evidenced the zoonotic potential of rabbit adapted BSE. Two out of nine inoculated TgHum mice succumbed with a prion disease as confirmed by WB and immunohistochemical techniques. It is clear that the BSE strain has unique features when compared with other prions, particularly in terms of disease phenotype stability and permeability to different hosts. One can speculate that these differences must reside in an also unique PrPSc structure; so unique that its conformation is reproducible on a considerably wide range of amino acid sequences.11 This is not the case for other prions, which upon interspecies transmission are not able to maintain its strain characteristics. For instance, mouse adapted prions such as 139A and ME7 with a single species barrier crossing to the hamster model, a rather similar species, undergo profound strain changes.27 To fully understand why certain prion conformations, such as that of the BSE strain, remain unaltered in varying amino acid scenarios it is mandatory to resolve its structure, probably with a resolution of a few nanometers, and compare it to other prion strains. Only when this is achieved, a qualitative leap will be attained to better comprehend the prion propagation mechanisms. Previous studies had already reported that BSE is able to retain its biochemical and biological properties upon interspecies
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words, even though passive surveillance and withdrawal of specified risk material should prevent transmission to humans and the ban on the use of ruminant meat and bone meal in feedstuffs should avoid horizontal transmission, sporadic cases will still occur so we will have to coexist with cattle prions and the risks associated to them. With the exception of these atypical BSE strains, all BSE cases diagnosed in cattle have been classified as classical BSE, and for many years BSE was believed to be caused by a single stable agent.2 This observation was due to the homogeneity of biological and laboratorial features of the PrPd, a pathological isoform of the host encoded cellular prion protein (PrPC), which is believed to be the ethiological agent of transmissible spongiform encephalopathies (TSEs).8 BSE associated PrPd is characterized in western blotting by a three band pattern between 18 and 30 kDa, present after Proteinase-K digestion, with a predominance of a diglycosylated moiety.9 Also, BSE showed stable pathobiological features upon transmission to wild type inbred mouse lines which allowed precise identification of the strain based on brain lesions and PrPd deposition profiles when assessed by immunohistochemistry (IHC).10-12 These experiments suggested a link between vCJD and BSE.3 Experiments using transgenic mice expressing bovine PrP (TgBov mice) further supported the idea that the BSE epizootic was caused by a single agent with no strain diversity.13-16 Additionally, after interspecies transmission when sheep passaged BSE was inoculated back into TgBov mice, the disease exhibited typical features of BSE in this model, except for a reduced incubation period when compared with cattle BSE.17 Surprisingly, inoculation with atypical BSE isolates also showed features of classical BSE in this model, suggesting a possible link between these strains.6,18,19 Interspecies transmission of prions is a phenomenon worth studying since new hosts can modify strain properties. As an example, chronic wasting disease (CWD) prions proved not to be transmissible to hamsters20 but, upon transmission to ferrets, which were indeed susceptible,
Disclosure of Potential Conflicts of Interest
The authors declare no competing financial interests.
Acknowledgments
This work was financially supported by two national grants from Spain (AGL2012-37988-C04-01 and AGL200805296-C02), Etortek Research Programs 2011/2013, EFA205/11 and CTP11-P04. The authors thank the support from IKERBasque foundation, vivarium and maintenance from CIC bioGUNE, Sierra Espinar, Marta Valle, Mariano Moreno and Paola Marco and CReSA’s biocontainment unit staff for care and maintenance. References 1. Vidal E, Fernández-Borges N, Pintado B, Ordóñez M, Márquez M, Fondevila D, Torres JM, Pumarola M, Castilla J. Bovine spongiform encephalopathy induces misfolding of alleged prion-resistant species cellular prion protein without altering its pathobiological features. J Neurosci 2013; 33:777886; PMID:23637170; http://dx.doi.org/10.1523/ JNEUROSCI.0244-13.2013 2. Wells GA, Wilesmith JW. The neuropathology and epidemiology of bovine spongiform encephalopathy. Brain Pathol 1995; 5:91-103; PMID:7767494; http:// dx.doi.org/10.1111/j.1750-3639.1995.tb00580.x 3. Bruce ME, Will RG, Ironside JW, McConnell I, Drummond D, Suttie A, McCardle L, Chree A, Hope J, Birkett C, et al. Transmissions to mice indicate that ‘new variant’ CJD is caused by the BSE agent. Nature 1997; 389:498-501; PMID:9333239; http://dx.doi. org/10.1038/39057 4. Casalone C, Zanusso G, Acutis P, Ferrari S, Capucci L, Tagliavini F, Monaco S, Caramelli M. Identification of a second bovine amyloidotic spongiform encephalopathy: molecular similarities with sporadic Creutzfeldt-Jakob disease. Proc Natl Acad Sci U S A 2004; 101:3065-70; PMID:14970340; http://dx.doi. org/10.1073/pnas.0305777101 5. Biacabe AG, Laplanche JL, Ryder S, Baron T. Distinct molecular phenotypes in bovine prion diseases. EMBO Rep 2004; 5:110-5; PMID:14710195; http:// dx.doi.org/10.1038/sj.embor.7400054 6. Espinosa JC, Andreoletti O, Lacroux C, Prieto I, Lorenzo P, Larska M, Baron T, Torres JM. Atypical H-type BSE infection in bovine-PrP transgenicm ice led to the emergence of classical BSE strain features. Prion 2010; 4:137 7. Bencsik A, Leboidre M, Debeer S, Aufauvre C, Baron T. Unique properties of the classical bovine spongiform encephalopathy strain and its emergence from H-type bovine spongiform encephalopathy substantiated by VM transmission studies. J Neuropathol Exp Neurol 2013; 72:211-8; PMID:23399901; http://dx.doi. org/10.1097/NEN.0b013e318285c7f9 8. Prusiner SB. Novel proteinaceous infectious particles cause scrapie. Science 1982; 216:13644; PMID:6801762; http://dx.doi.org/10.1126/ science.6801762 9. Collinge J, Sidle KC, Meads J, Ironside J, Hill AF. Molecular analysis of prion strain variation and the aetiology of ‘new variant’ CJD. Nature 1996; 383:685-90; PMID:8878476; http://dx.doi. org/10.1038/383685a0 10. Green R, Horrocks C, Wilkinson A, Hawkins SA, Ryder SJ. Primary isolation of the bovine spongiform encephalopathy agent in mice: agent definition based on a review of 150 transmissions. J Comp Pathol 2005; 132:117-31; PMID:15737338; http://dx.doi. org/10.1016/j.jcpa.2004.08.002
11. Bruce M, Chree A, McConnell I, Foster J, Pearson G, Fraser H. Transmission of bovine spongiform encephalopathy and scrapie to mice: strain variation and the species barrier. Philos Trans R Soc Lond B Biol Sci 1994; 343:405-11; PMID:7913758; http://dx.doi. org/10.1098/rstb.1994.0036 12. Brown DA, Bruce ME, Fraser JR. Comparison of the neuropathological characteristics of bovine spongiform encephalopathy (BSE) and variant Creutzfeldt-Jakob disease (vCJD) in mice. Neuropathol Appl Neurobiol 2003; 29:262-72; PMID:12787323; http://dx.doi. org/10.1046/j.1365-2990.2003.00462.x 13. Castilla J, Gutiérrez Adán A, Brun A, Pintado B, Ramírez MA, Parra B, Doyle D, Rogers M, Salguero FJ, Sánchez C, et al. Early detection of PrPres in BSE-infected bovine PrP transgenic mice. Arch Virol 2003; 148:677-91; PMID:12664293; http://dx.doi. org/10.1007/s00705-002-0958-4 14. Scott MR, Safar J, Telling G, Nguyen O, Groth D, Torchia M, Koehler R, Tremblay P, Walther D, Cohen FE, et al. Identification of a prion protein epitope modulating transmission of bovine spongiform encephalopathy prions to transgenic mice. Proc Natl Acad Sci U S A 1997; 94:14279-84; PMID:9405603; http://dx.doi.org/10.1073/pnas.94.26.14279 15. Buschmann A, Groschup MH. Highly bovine spongiform encephalopathy-sensitive transgenic mice confirm the essential restriction of infectivity to the nervous system in clinically diseased cattle. J Infect Dis 2005; 192:934-42; PMID:16088845; http:// dx.doi.org/10.1086/431602 16. Buschmann A, Pfaff E, Reifenberg K, Müller HM, Groschup MH. Detection of cattle-derived BSE prions using transgenic mice overexpressing bovine PrP(C). Arch Virol Suppl 2000; •••:75-86; PMID:11214936 17. Espinosa JC, Andréoletti O, Castilla J, Herva ME, Morales M, Alamillo E, San-Segundo FD, Lacroux C, Lugan S, Salguero FJ, et al. Sheep-passaged bovine spongiform encephalopathy agent exhibits altered pathobiological properties in bovine-PrP transgenic mice. J Virol 2007; 81:835-43; PMID:17079295; http://dx.doi.org/10.1128/JVI.01356-06 18. Torres JM, Andréoletti O, Lacroux C, Prieto I, Lorenzo P, Larska M, Baron T, Espinosa JC. Classical bovine spongiform encephalopathy by transmission of H-type prion in homologous prion protein context. Emerg Infect Dis 2011; 17:163644; PMID:21888788; http://dx.doi.org/10.3201/ eid1709.101403 19. Baron T, Vulin J, Biacabe AG, Lakhdar L, Verchere J, Torres JM, Bencsik A. Emergence of classical BSE strain properties during serial passages of H-BSE in wild-type mice. PLoS One 2011; 6:e15839; PMID:21264286; http://dx.doi.org/10.1371/journal. pone.0015839 20. Williams ES, Young S. Spongiform encephalopathies in Cervidae. Rev Sci Tech 1992; 11:551-67; PMID:1617203 21. Bartz JC, Marsh RF, McKenzie DI, Aiken JM. The host range of chronic wasting disease is altered on passage in ferrets. Virology 1998; 251:297301; PMID:9837794; http://dx.doi.org/10.1006/ viro.1998.9427 22. Fernández-Borges N, Castilla J. PMCA. A decade of in vitro prion replication. Current Chemical Biology 2010; 4:200-7 23. Chianini F, Fernández-Borges N, Vidal E, Gibbard L, Pintado B, de Castro J, Priola SA, Hamilton S, Eaton SL, Finlayson J, et al. Rabbits are not resistant to prion infection. Proc Natl Acad Sci U S A 2012; 109:50805; PMID:22416127; http://dx.doi.org/10.1073/ pnas.1120076109 24. Fernández-Borges N, Chianini F, Erana H, Vidal E, Eaton SL, Pintado B, Finlayson J, Dagleish MP, Castilla J. Naturally prion resistant mammals: A utopia? Prion 2012; 6:425-9; PMID:22453170
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passage through certain species such as sheep,17,28 goats,29 felidae,30 mice,11 or humans.3,31 Although other authors have also reported that some biochemical features might change after certain BSE interspecies transmissions such as in deer32 or when testing certain transgenic mouse experimental models.33 Coexistence for centuries with animal prions, such as scrapie, without any apparent risks to the human population was disturbed by the emergence of a zoonotic prion disease in cattle: BSE. Of course, this new prion strain found its way into the field leaded by mankind’s artificial husbandry practices. So at this point any further intervention should be preceded by a meticulous consideration of all the possible risks involved. Regulations regarding TSE control and surveillance are being revised, predictably less surveillance will be performed and the roadmap is leading toward a reintroduction of animal derived protein in feedstuffs, provided that cannibalism does not happen, i.e., a determined species is not to be fed with formulas containing animal protein of its own species. Thus, feeding any farmed animal with mammal-derived protein is to be banned yet, with the exception of pet-food [Regulation (EC) No 999/2001 in the Official Journal of the European Communities]. Our results evidence that BSE prions are capable not only of adapting to new species, even to those which apparently are more resistant than others, but in doing so they maintain their ability to infect human beings. And if, in an unlikely but yet possible case, this should occur in the field, the resulting prion, as any other infectious agent, might find its way to adapt and maybe evolve in a changing environment. Eventually it might even lead to the emergence of new infectious agents with unpredictable transmission potential. Given that spontaneous prions are out there in the field, results such as the ones presented here suggest that caution must be taken when considering the use of animal derived protein in feedstuffs.
33. Yokoyama T, Masujin K, Iwamaru Y, Imamura M, Mohri S. Alteration of the biological and biochemical characteristics of bovine spongiform encephalopathy prions during interspecies transmission in transgenic mice models. J Gen Virol 2009; 90:2618; PMID:19088297; http://dx.doi.org/10.1099/ vir.0.004754-0
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29. Eloit M, Adjou K, Coulpier M, Fontaine JJ, Hamel R, Lilin T, Messiaen S, Andreoletti O, Baron T, Bencsik A, et al. BSE agent signatures in a goat. Vet Rec 2005; 156:523-4; PMID:15833975 30. Eiden M, Hoffmann C, Balkema-Buschmann A, Müller M, Baumgartner K, Groschup MH. Biochemical and immunohistochemical characterization of feline spongiform encephalopathy in a German captive cheetah. J Gen Virol 2010; 91:2874-83; PMID:20660146; http://dx.doi. org/10.1099/vir.0.022103-0 31. Hill AF, Desbruslais M, Joiner S, Sidle KC, Gowland I, Collinge J, Doey LJ, Lantos P. The same prion strain causes vCJD and BSE. Nature 1997; 389:448-50, 526; PMID:9333232; http://dx.doi.org/10.1038/38925 32. Dagleish MP, Martin S, Steele P, Finlayson J, Sisó S, Hamilton S, Chianini F, Reid HW, González L, Jeffrey M. Experimental transmission of bovine spongiform encephalopathy to European red deer (Cervus elaphus elaphus). BMC Vet Res 2008; 4:17; PMID:18507844; http://dx.doi.org/10.1186/1746-6148-4-17
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25. Fernández-Borges N, de Castro J, Castilla J. In vitro studies of the transmission barrier. Prion 2009; 3:2203; PMID:20009509; http://dx.doi.org/10.4161/ pri.3.4.10500 26. Padilla D, Béringue V, Espinosa JC, Andreoletti O, Jaumain E, Reine F, Herzog L, Gutierrez-Adan A, Pintado B, Laude H, et al. Sheep and goat BSE propagate more efficiently than cattle BSE in human PrP transgenic mice. PLoS Pathog 2011; 7:e1001319; PMID:21445238; http://dx.doi.org/10.1371/journal. ppat.1001319 27. Shi Q, Zhang BY, Gao C, Zhang J, Jiang HY, Chen C, Han J, Dong XP. Mouse-adapted scrapie strains 139A and ME7 overcome species barrier to induce experimental scrapie in hamsters and changed their pathogenic features. Virol J 2012; 9:63; PMID:22400710; http://dx.doi. org/10.1186/1743-422X-9-63 28. Stack MJ, Chaplin MJ, Clark J. Differentiation of prion protein glycoforms from naturally occurring sheep scrapie, sheep-passaged scrapie strains (CH1641 and SSBP1), bovine spongiform encephalopathy (BSE) cases and Romney and Cheviot breed sheep experimentally inoculated with BSE using two monoclonal antibodies. Acta Neuropathol 2002; 104:279-86; PMID:12172914
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Volume 7 Issue 6
ISSN: 1933-6896 (Print) 1933-690X (Online) Journal homepage: http://www.tandfonline.com/loi/kprn20
Exploring the risks of a putative transmission of BSE to new species Enric Vidal, Natalia Fernández-Borges, Belén Pintado, Montserrat Ordóñez, Mercedes Márquez, Dolors Fondevila, Hasier Eraña, Juan María Torres, Martí Pumarola & Joaquín Castilla To cite this article: Enric Vidal, Natalia Fernández-Borges, Belén Pintado, Montserrat Ordóñez, Mercedes Márquez, Dolors Fondevila, Hasier Eraña, Juan María Torres, Martí Pumarola & Joaquín Castilla (2013) Exploring the risks of a putative transmission of BSE to new species, Prion, 7:6, 443-446, DOI: 10.4161/pri.27014 To link to this article: http://dx.doi.org/10.4161/pri.27014
Published online: 01 Nov 2013.
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Article views: 74
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Citing articles: 1 View citing articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=kprn20 Download by: [85.25.103.119]
Date: 25 March 2016, At: 21:02
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Prion 7:6, 443–446; November/December 2013; © 2013 Landes Bioscience
Exploring the risks of a putative transmission of BSE to new species Centre de Recerca en Sanitat Animal (CReSA); UAB-IRTA; Campus de la Universitat Autònoma de Barcelona; Barcelona, Spain; 2CIC bioGUNE; Parque tecnológico de Bizkaia; Bizkaia, Spain; 3Centro Nacional de Biotecnología (CNB); Campus de Cantoblanco; Madrid, Spain; 4 Department of Animal Medicine and Surgery; Veterinary faculty; Universitat Autònoma de Barcelona (UAB); Barcelona, Spain; 5CISA—INIA; Madrid, Spain; 6 IKERBASQUE; Basque Foundation for Science; Bizkaia, Spain
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Keywords: prion, BSE, scrapie, PMCA, transmission barrier, prion resistant species *Correspondence to: Joaquín Castilla; Email: [email protected] Submitted: 08/16/2013 Revised: 10/01/2013 Accepted: 10/31/2013 http://dx.doi.org/10.4161/pri.27014 Addendum to : Vidal E, Fernández-Borges N, Pintado B, Ordóñez M, Márquez M, Fondevila D, Torres JM, Pumarola M, Castilla J. Bovine spongiform encephalopathy induces misfolding of alleged prion-resistant species cellular prion protein without altering its pathobiological features. J Neurosci 2013; 33:7778-86; http:// dx.doi.org/10.1523/JNEUROSCI.0244-13.2013; PMID:23637170
he prion responsible for the Bovine Spongiform Encephalopathy (BSE) shows unique features when compared with other prions. One of these features is its ability to infect almost all experimentally tested animal models. In the paper published in The Journal of Neuroscience1 we describe a series of experiments directed toward elucidating which would be the in vivo behavior of BSE if it would infect dogs and rabbits, two alleged prion resistant species. Protein misfolding cyclic amplification (PMCA) was used to generate canidae and leporidae in vitro adapted BSE prions. A characterization of their in vivo pathobiological properties showed that BSE prions were capable not only of adapting to new species but they maintained, in the case of rabbits, their ability to infect transgenic mice expressing human PrP. The remarkable adaptation ability of certain prions implies that any new host species could lead to the emergence of new infectious agents with unpredictable transmission potential. Our results suggest that caution must be taken when considering the use of any mammal-derived protein in feedstuffs.
Perspectives Bovine Spongiform Encephalopathy (BSE) is one of the most well-known animal prion diseases.2 The “mad cow disease” outbreak had unprecedented consequences regarding worldwide public
health and food safety policies. The reason for this was the ability of the agent causing BSE to transmit to species other than cattle, its original host, including human beings.3 Enormous economical efforts were directed toward controlling the disease in cattle with exceptional results. Indeed, policies such as the ban on using ruminant protein on feed drastically reduced the reported BSE cases in cattle. The compulsory testing of a large number of animals allowed to accurately monitoring the success of such measures. In every country where these policies have been implemented, the BSE cases are practically down to zero. Thus, updated risk analyses point to a relaxation of the control measures. Thanks to the research efforts directed toward this subject, many previously unknown aspects of this prion disease have came to light during the last decade, for instance, the existence of the so called atypical BSE cases.4,5 Two cattle prions strains have been identified in aged cattle and named BSE-L and BSE-H after the different electrophoretic pattern of its proteinase-K digested disease-associated BSE prion protein (PrPd) on western blotting. Epidemiology and pathological features of the diagnosed cases suggest that those are probably sporadic prion diseases and evidence suggests that they might have had a crucial role in the origin of the BSE epizooty.6,7 If these atypical presentations are indeed sporadic prion diseases, such as the ones known in humans, it implies that current BSE control measures will not be effective to eradicate them. In other
www.landesbioscience.com Prion 443
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Enric Vidal1, Natalia Fernández-Borges2, Belén Pintado3, Montserrat Ordóñez1, Mercedes Márquez4, Dolors Fondevila4, Hasier Eraña 2, Juan María Torres5, Martí Pumarola4 and Joaquín Castilla 2,6*
CWD prions could readily infect hamsters.21 BSE has in fact been described to change, to a certain extent, upon passage through species other than cattle, i.e., sheep. Passage through sheep resulted in an increase of BSE virulence when inoculated into TgBov mice.17 So, attention must be paid to the emergence of new prion strains or the modifications that new hosts might pose to the pathobiological properties of already known strains, particularly regarding the ability to transmit to other species, including humans. In vitro amplification of prions22 has proven to be an effective tool to predict the in vivo behavior of certain prions.23,24 Not only does it accelerate the transmission process but it also can be used to push the system, by challenging healthy brain homogenates of different species with different prion strains at high concentration and directly in a test tube, to find out what would happen in case certain cross-species transmission of prions would occur in nature. Several publications support protein misfolding cyclic amplification (PMCA) ability to mimic potential in vivo scenarios. For instance, the susceptibility of rabbits to prion diseases was predicted by PMCA and then demonstrated in its natural host23 and, likewise, other species PrP could also be misfolded in vitro including dogs and horses.25 In the paper published in The Journal of Neuroscience1 we describe a series of experiments directed toward elucidating which would be the in vivo behavior of BSE, one of the most relevant prions in Europe, in case it managed to cross certain transmission barriers. The species chosen were dogs and rabbits for several reasons, namely both are species in close contact with human beings and no reports of prion susceptibility had been published at the time the experiments were designed, thus they were species considered to be probably prion resistant. The experiments consisted in an initial in vitro phase in which dog and rabbit normal brain homogenates were subjected to PMCA with a BSE seed. In a second phase, the newly obtained prions were inoculated into TgBov mice and in a transgenic model expressing human PrP (TgHum)26 to assess their in vivo
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behavior in comparison to the original BSE seed. Several interesting findings were obtained from that set of experiments. To start with, BSE was the only prion strain that could be propagated in vitro in both alleged resistant species, a capacity which lacked other prion strains originated in other hosts, such as scrapie. The typical BSE biochemical signature (western blotting) remained unchanged in both species even after multiple in vitro amplification cycles. Interestingly, the pathobiological features of these newly in vitro obtained prions, such as their infectivity, incubation times, biochemical profile, lesion profile, and etcetera were remarkably comparable to those of BSE when bioassayed in transgenic mouse models overexpressing bovine and human PRNP. Of particular interest were the bioassay results in tgHum mice that evidenced the zoonotic potential of rabbit adapted BSE. Two out of nine inoculated TgHum mice succumbed with a prion disease as confirmed by WB and immunohistochemical techniques. It is clear that the BSE strain has unique features when compared with other prions, particularly in terms of disease phenotype stability and permeability to different hosts. One can speculate that these differences must reside in an also unique PrPSc structure; so unique that its conformation is reproducible on a considerably wide range of amino acid sequences.11 This is not the case for other prions, which upon interspecies transmission are not able to maintain its strain characteristics. For instance, mouse adapted prions such as 139A and ME7 with a single species barrier crossing to the hamster model, a rather similar species, undergo profound strain changes.27 To fully understand why certain prion conformations, such as that of the BSE strain, remain unaltered in varying amino acid scenarios it is mandatory to resolve its structure, probably with a resolution of a few nanometers, and compare it to other prion strains. Only when this is achieved, a qualitative leap will be attained to better comprehend the prion propagation mechanisms. Previous studies had already reported that BSE is able to retain its biochemical and biological properties upon interspecies
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words, even though passive surveillance and withdrawal of specified risk material should prevent transmission to humans and the ban on the use of ruminant meat and bone meal in feedstuffs should avoid horizontal transmission, sporadic cases will still occur so we will have to coexist with cattle prions and the risks associated to them. With the exception of these atypical BSE strains, all BSE cases diagnosed in cattle have been classified as classical BSE, and for many years BSE was believed to be caused by a single stable agent.2 This observation was due to the homogeneity of biological and laboratorial features of the PrPd, a pathological isoform of the host encoded cellular prion protein (PrPC), which is believed to be the ethiological agent of transmissible spongiform encephalopathies (TSEs).8 BSE associated PrPd is characterized in western blotting by a three band pattern between 18 and 30 kDa, present after Proteinase-K digestion, with a predominance of a diglycosylated moiety.9 Also, BSE showed stable pathobiological features upon transmission to wild type inbred mouse lines which allowed precise identification of the strain based on brain lesions and PrPd deposition profiles when assessed by immunohistochemistry (IHC).10-12 These experiments suggested a link between vCJD and BSE.3 Experiments using transgenic mice expressing bovine PrP (TgBov mice) further supported the idea that the BSE epizootic was caused by a single agent with no strain diversity.13-16 Additionally, after interspecies transmission when sheep passaged BSE was inoculated back into TgBov mice, the disease exhibited typical features of BSE in this model, except for a reduced incubation period when compared with cattle BSE.17 Surprisingly, inoculation with atypical BSE isolates also showed features of classical BSE in this model, suggesting a possible link between these strains.6,18,19 Interspecies transmission of prions is a phenomenon worth studying since new hosts can modify strain properties. As an example, chronic wasting disease (CWD) prions proved not to be transmissible to hamsters20 but, upon transmission to ferrets, which were indeed susceptible,
Disclosure of Potential Conflicts of Interest
The authors declare no competing financial interests.
Acknowledgments
This work was financially supported by two national grants from Spain (AGL2012-37988-C04-01 and AGL200805296-C02), Etortek Research Programs 2011/2013, EFA205/11 and CTP11-P04. The authors thank the support from IKERBasque foundation, vivarium and maintenance from CIC bioGUNE, Sierra Espinar, Marta Valle, Mariano Moreno and Paola Marco and CReSA’s biocontainment unit staff for care and maintenance. References 1. Vidal E, Fernández-Borges N, Pintado B, Ordóñez M, Márquez M, Fondevila D, Torres JM, Pumarola M, Castilla J. Bovine spongiform encephalopathy induces misfolding of alleged prion-resistant species cellular prion protein without altering its pathobiological features. J Neurosci 2013; 33:777886; PMID:23637170; http://dx.doi.org/10.1523/ JNEUROSCI.0244-13.2013 2. Wells GA, Wilesmith JW. The neuropathology and epidemiology of bovine spongiform encephalopathy. Brain Pathol 1995; 5:91-103; PMID:7767494; http:// dx.doi.org/10.1111/j.1750-3639.1995.tb00580.x 3. Bruce ME, Will RG, Ironside JW, McConnell I, Drummond D, Suttie A, McCardle L, Chree A, Hope J, Birkett C, et al. Transmissions to mice indicate that ‘new variant’ CJD is caused by the BSE agent. Nature 1997; 389:498-501; PMID:9333239; http://dx.doi. org/10.1038/39057 4. Casalone C, Zanusso G, Acutis P, Ferrari S, Capucci L, Tagliavini F, Monaco S, Caramelli M. Identification of a second bovine amyloidotic spongiform encephalopathy: molecular similarities with sporadic Creutzfeldt-Jakob disease. Proc Natl Acad Sci U S A 2004; 101:3065-70; PMID:14970340; http://dx.doi. org/10.1073/pnas.0305777101 5. Biacabe AG, Laplanche JL, Ryder S, Baron T. Distinct molecular phenotypes in bovine prion diseases. EMBO Rep 2004; 5:110-5; PMID:14710195; http:// dx.doi.org/10.1038/sj.embor.7400054 6. Espinosa JC, Andreoletti O, Lacroux C, Prieto I, Lorenzo P, Larska M, Baron T, Torres JM. Atypical H-type BSE infection in bovine-PrP transgenicm ice led to the emergence of classical BSE strain features. Prion 2010; 4:137 7. Bencsik A, Leboidre M, Debeer S, Aufauvre C, Baron T. Unique properties of the classical bovine spongiform encephalopathy strain and its emergence from H-type bovine spongiform encephalopathy substantiated by VM transmission studies. J Neuropathol Exp Neurol 2013; 72:211-8; PMID:23399901; http://dx.doi. org/10.1097/NEN.0b013e318285c7f9 8. Prusiner SB. Novel proteinaceous infectious particles cause scrapie. Science 1982; 216:13644; PMID:6801762; http://dx.doi.org/10.1126/ science.6801762 9. Collinge J, Sidle KC, Meads J, Ironside J, Hill AF. Molecular analysis of prion strain variation and the aetiology of ‘new variant’ CJD. Nature 1996; 383:685-90; PMID:8878476; http://dx.doi. org/10.1038/383685a0 10. Green R, Horrocks C, Wilkinson A, Hawkins SA, Ryder SJ. Primary isolation of the bovine spongiform encephalopathy agent in mice: agent definition based on a review of 150 transmissions. J Comp Pathol 2005; 132:117-31; PMID:15737338; http://dx.doi. org/10.1016/j.jcpa.2004.08.002
11. Bruce M, Chree A, McConnell I, Foster J, Pearson G, Fraser H. Transmission of bovine spongiform encephalopathy and scrapie to mice: strain variation and the species barrier. Philos Trans R Soc Lond B Biol Sci 1994; 343:405-11; PMID:7913758; http://dx.doi. org/10.1098/rstb.1994.0036 12. Brown DA, Bruce ME, Fraser JR. Comparison of the neuropathological characteristics of bovine spongiform encephalopathy (BSE) and variant Creutzfeldt-Jakob disease (vCJD) in mice. Neuropathol Appl Neurobiol 2003; 29:262-72; PMID:12787323; http://dx.doi. org/10.1046/j.1365-2990.2003.00462.x 13. Castilla J, Gutiérrez Adán A, Brun A, Pintado B, Ramírez MA, Parra B, Doyle D, Rogers M, Salguero FJ, Sánchez C, et al. Early detection of PrPres in BSE-infected bovine PrP transgenic mice. Arch Virol 2003; 148:677-91; PMID:12664293; http://dx.doi. org/10.1007/s00705-002-0958-4 14. Scott MR, Safar J, Telling G, Nguyen O, Groth D, Torchia M, Koehler R, Tremblay P, Walther D, Cohen FE, et al. Identification of a prion protein epitope modulating transmission of bovine spongiform encephalopathy prions to transgenic mice. Proc Natl Acad Sci U S A 1997; 94:14279-84; PMID:9405603; http://dx.doi.org/10.1073/pnas.94.26.14279 15. Buschmann A, Groschup MH. Highly bovine spongiform encephalopathy-sensitive transgenic mice confirm the essential restriction of infectivity to the nervous system in clinically diseased cattle. J Infect Dis 2005; 192:934-42; PMID:16088845; http:// dx.doi.org/10.1086/431602 16. Buschmann A, Pfaff E, Reifenberg K, Müller HM, Groschup MH. Detection of cattle-derived BSE prions using transgenic mice overexpressing bovine PrP(C). Arch Virol Suppl 2000; •••:75-86; PMID:11214936 17. Espinosa JC, Andréoletti O, Castilla J, Herva ME, Morales M, Alamillo E, San-Segundo FD, Lacroux C, Lugan S, Salguero FJ, et al. Sheep-passaged bovine spongiform encephalopathy agent exhibits altered pathobiological properties in bovine-PrP transgenic mice. J Virol 2007; 81:835-43; PMID:17079295; http://dx.doi.org/10.1128/JVI.01356-06 18. Torres JM, Andréoletti O, Lacroux C, Prieto I, Lorenzo P, Larska M, Baron T, Espinosa JC. Classical bovine spongiform encephalopathy by transmission of H-type prion in homologous prion protein context. Emerg Infect Dis 2011; 17:163644; PMID:21888788; http://dx.doi.org/10.3201/ eid1709.101403 19. Baron T, Vulin J, Biacabe AG, Lakhdar L, Verchere J, Torres JM, Bencsik A. Emergence of classical BSE strain properties during serial passages of H-BSE in wild-type mice. PLoS One 2011; 6:e15839; PMID:21264286; http://dx.doi.org/10.1371/journal. pone.0015839 20. Williams ES, Young S. Spongiform encephalopathies in Cervidae. Rev Sci Tech 1992; 11:551-67; PMID:1617203 21. Bartz JC, Marsh RF, McKenzie DI, Aiken JM. The host range of chronic wasting disease is altered on passage in ferrets. Virology 1998; 251:297301; PMID:9837794; http://dx.doi.org/10.1006/ viro.1998.9427 22. Fernández-Borges N, Castilla J. PMCA. A decade of in vitro prion replication. Current Chemical Biology 2010; 4:200-7 23. Chianini F, Fernández-Borges N, Vidal E, Gibbard L, Pintado B, de Castro J, Priola SA, Hamilton S, Eaton SL, Finlayson J, et al. Rabbits are not resistant to prion infection. Proc Natl Acad Sci U S A 2012; 109:50805; PMID:22416127; http://dx.doi.org/10.1073/ pnas.1120076109 24. Fernández-Borges N, Chianini F, Erana H, Vidal E, Eaton SL, Pintado B, Finlayson J, Dagleish MP, Castilla J. Naturally prion resistant mammals: A utopia? Prion 2012; 6:425-9; PMID:22453170
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passage through certain species such as sheep,17,28 goats,29 felidae,30 mice,11 or humans.3,31 Although other authors have also reported that some biochemical features might change after certain BSE interspecies transmissions such as in deer32 or when testing certain transgenic mouse experimental models.33 Coexistence for centuries with animal prions, such as scrapie, without any apparent risks to the human population was disturbed by the emergence of a zoonotic prion disease in cattle: BSE. Of course, this new prion strain found its way into the field leaded by mankind’s artificial husbandry practices. So at this point any further intervention should be preceded by a meticulous consideration of all the possible risks involved. Regulations regarding TSE control and surveillance are being revised, predictably less surveillance will be performed and the roadmap is leading toward a reintroduction of animal derived protein in feedstuffs, provided that cannibalism does not happen, i.e., a determined species is not to be fed with formulas containing animal protein of its own species. Thus, feeding any farmed animal with mammal-derived protein is to be banned yet, with the exception of pet-food [Regulation (EC) No 999/2001 in the Official Journal of the European Communities]. Our results evidence that BSE prions are capable not only of adapting to new species, even to those which apparently are more resistant than others, but in doing so they maintain their ability to infect human beings. And if, in an unlikely but yet possible case, this should occur in the field, the resulting prion, as any other infectious agent, might find its way to adapt and maybe evolve in a changing environment. Eventually it might even lead to the emergence of new infectious agents with unpredictable transmission potential. Given that spontaneous prions are out there in the field, results such as the ones presented here suggest that caution must be taken when considering the use of animal derived protein in feedstuffs.
33. Yokoyama T, Masujin K, Iwamaru Y, Imamura M, Mohri S. Alteration of the biological and biochemical characteristics of bovine spongiform encephalopathy prions during interspecies transmission in transgenic mice models. J Gen Virol 2009; 90:2618; PMID:19088297; http://dx.doi.org/10.1099/ vir.0.004754-0
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29. Eloit M, Adjou K, Coulpier M, Fontaine JJ, Hamel R, Lilin T, Messiaen S, Andreoletti O, Baron T, Bencsik A, et al. BSE agent signatures in a goat. Vet Rec 2005; 156:523-4; PMID:15833975 30. Eiden M, Hoffmann C, Balkema-Buschmann A, Müller M, Baumgartner K, Groschup MH. Biochemical and immunohistochemical characterization of feline spongiform encephalopathy in a German captive cheetah. J Gen Virol 2010; 91:2874-83; PMID:20660146; http://dx.doi. org/10.1099/vir.0.022103-0 31. Hill AF, Desbruslais M, Joiner S, Sidle KC, Gowland I, Collinge J, Doey LJ, Lantos P. The same prion strain causes vCJD and BSE. Nature 1997; 389:448-50, 526; PMID:9333232; http://dx.doi.org/10.1038/38925 32. Dagleish MP, Martin S, Steele P, Finlayson J, Sisó S, Hamilton S, Chianini F, Reid HW, González L, Jeffrey M. Experimental transmission of bovine spongiform encephalopathy to European red deer (Cervus elaphus elaphus). BMC Vet Res 2008; 4:17; PMID:18507844; http://dx.doi.org/10.1186/1746-6148-4-17
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25. Fernández-Borges N, de Castro J, Castilla J. In vitro studies of the transmission barrier. Prion 2009; 3:2203; PMID:20009509; http://dx.doi.org/10.4161/ pri.3.4.10500 26. Padilla D, Béringue V, Espinosa JC, Andreoletti O, Jaumain E, Reine F, Herzog L, Gutierrez-Adan A, Pintado B, Laude H, et al. Sheep and goat BSE propagate more efficiently than cattle BSE in human PrP transgenic mice. PLoS Pathog 2011; 7:e1001319; PMID:21445238; http://dx.doi.org/10.1371/journal. ppat.1001319 27. Shi Q, Zhang BY, Gao C, Zhang J, Jiang HY, Chen C, Han J, Dong XP. Mouse-adapted scrapie strains 139A and ME7 overcome species barrier to induce experimental scrapie in hamsters and changed their pathogenic features. Virol J 2012; 9:63; PMID:22400710; http://dx.doi. org/10.1186/1743-422X-9-63 28. Stack MJ, Chaplin MJ, Clark J. Differentiation of prion protein glycoforms from naturally occurring sheep scrapie, sheep-passaged scrapie strains (CH1641 and SSBP1), bovine spongiform encephalopathy (BSE) cases and Romney and Cheviot breed sheep experimentally inoculated with BSE using two monoclonal antibodies. Acta Neuropathol 2002; 104:279-86; PMID:12172914
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