Post by Terry S. Singeltary Sr. on Apr 24, 2023 14:54:30 GMT -6
CWD TSE PrP Feces, Contagiousness, Zoonosis Potential?
Heterozygosity for cervid S138N polymorphism results in subclinical CWD in gene-targeted mice and progressive inhibition of prion conversion
HIGHLIGHT
However, prion seeding activity was detected in spleens, brains, and feces of these mice, suggesting subclinical infection accompanied by prion shedding.
Infected animals accumulate prions in lymphoreticular and other peripheral tissues, e.g., skeletal muscle, and shed infectious prions in saliva, urine and feces, contributing to direct and environmental transmission and rapidly increasing geographic distribution of CWD (9–12).
However, prion-seeding activity was detectable in the brain, spleen, and feces, indicating subclinical infection and potential for contagiousness.
Heterozygosity for cervid S138N polymorphism results in subclinical CWD in gene-targeted mice and progressive inhibition of prion conversion
Maria I. Arifin orcid.org/0000-0003-2042-3492, Lech Kaczmarczyk orcid.org/0000-0003-2747-3134, Doris Zeng orcid.org/0009-0002-2512-6227, +7, and Sabine Gilch orcid.org/0000-0001-5923-3464 sgilch@ucalgary.caAuthors Info & Affiliations Edited by Reed Wickner, NIH, Bethesda, MD;
received December 12, 2022; accepted March 6, 2023
April 4, 2023
120 (15) e2221060120
doi.org/10.1073/pnas.2221060120
Significance
Amino acid substitutions within the cervid prion protein (PrP) can decrease susceptibility to chronic wasting disease, generally with more prominent effects in homozygous animals. Using novel gene-targeted mouse models expressing S138N reindeer/caribou PrP, we demonstrate subclinical infection with prion seeding activity in spleen and fecal prion shedding in heterozygous 138SN and homozygous 138NN mice. A lower percentage of heterozygous 138SN-PrP than homozygous 138NN-PrP expressing mice harbored seeding-efficient prions in tissues. This is caused by dominant-negative interference of the PrP variants occurring only if they are coexpressed. Our findings are relevant to inform conservation efforts for caribou, an endangered species in North America. Furthermore, our study provides new mechanistic insights into genetic resistance and dominant-negative interference of conversion-competent PrP variants.
Abstract
Prions are proteinaceous infectious particles that replicate by structural conversion of the host-encoded cellular prion protein (PrPC), causing fatal neurodegenerative diseases in mammals. Species-specific amino acid substitutions (AAS) arising from single nucleotide polymorphisms within the prion protein gene (Prnp) modulate prion disease pathogenesis, and, in several instances, reduce susceptibility of homo- or heterozygous AAS carriers to prion infection. However, a mechanistic understanding of their protective effects against clinical disease is missing. We generated gene-targeted mouse infection models of chronic wasting disease (CWD), a highly contagious prion disease of cervids. These mice express wild-type deer or PrPC harboring the S138N substitution homo- or heterozygously, a polymorphism found exclusively in reindeer (Rangifer tarandus spp.) and fallow deer (Dama dama). The wild-type deer PrP-expressing model recapitulated CWD pathogenesis including fecal shedding. Encoding at least one 138N allele prevented clinical CWD, accumulation of protease-resistant PrP (PrPres) and abnormal PrP deposits in the brain tissue. However, prion seeding activity was detected in spleens, brains, and feces of these mice, suggesting subclinical infection accompanied by prion shedding. 138N-PrPC was less efficiently converted to PrPres in vitro than wild-type deer (138SS) PrPC. Heterozygous coexpression of wild-type deer and 138N-PrPC resulted in dominant-negative inhibition and progressively diminished prion conversion over serial rounds of protein misfolding cyclic amplification. Our study indicates that heterozygosity at a polymorphic Prnp codon can confer the highest protection against clinical CWD and highlights the potential role of subclinical carriers in CWD transmission.
snip...
To conclude, our study demonstrates that CWD-infected animals harboring S138N PrP might be “silent spreaders” of CWD prions and highlights the importance of lymphatic tissues in the detection of CWD, particularly in caribou, even in the absence of clinical manifestation. It is important to keep in mind that even protective genotypes may be permissive to certain minor or newly emerging CWD strains. Our results provide new mechanistic insights into dominant-negative inhibition of prion conversion, the tissue specificity of this effect, and suggests that PrPC primary structure is a determinant for tissue-specific prion replication.
www.pnas.org/doi/10.1073/pnas.2221060120
Published: 22 August 2022
Transmission of cervid prions to humanized mice demonstrates the zoonotic potential of CWD
Samia Hannaoui, Irina Zemlyankina, Sheng Chun Chang, Maria Immaculata Arifin, Vincent Béringue, Debbie McKenzie, Hermann M. Schatzl & Sabine Gilch
Acta Neuropathologica volume 144, pages767–784 (2022)
Inoculation of these mice with deer CWD isolates resulted in atypical clinical manifestation with prion seeding activity and efficient transmissible infectivity in the brain and, remarkably, in feces, but without classical neuropathological or Western blot appearances of prion diseases.
In contrast, in cervids affected with CWD, infectivity has been found in the lymphatic system, salivary gland, intestinal tract, muscles, antler velvet, blood, urine, saliva, and feces [4], which have been demonstrated to be transmissible [57].
These data demonstrate that humanized tg650 mice inoculated with CWD prions shed prion infectivity in feces able to generate transmissible PrPSc in bank voles distinct from those generated by inoculation of the Wisc-1 deer isolate directly to bank voles.
Our findings strongly suggest that CWD should be regarded as an actual public health risk. Here, we use humanized mice to show that CWD prions can cross the species barrier to humans, and remarkably, infectious prions can be excreted in feces.
The presence of infectious prions shed in feces is one argument in favor of the existence of de novo generated PrPSc in these mice.
CWD in humans might remain subclinical but with PrPSc deposits in the brain with an unusual morphology that does not resemble the patterns usually seen in different prion diseases (e.g., mouse #328; Fig. 3), clinical with untraceable abnormal PrP (e.g., mouse #327) but still transmissible and uncovered upon subsequent passage (e.g., mouse #3063; Fig. 4), or prions have other reservoirs than the usual ones, hence the presence of infectivity in feces (e.g., mouse #327) suggesting a potential for human-to-human transmission and a real iatrogenic risk that might be unrecognizable. Here, humanized mice inoculated with CWD deer isolates had an atypical onset of the disease with myoclonus (93.75%), before presenting typical clinical signs, generating prions that presented with either atypical biochemical signature (#321 and #3063), shed in feces (#327), or were undetectable by the classical detection methods.
The finding that infectious PrPSc was shed in fecal material of CWD-infected humanized mice and induced clinical disease, different tropism, and typical three banding pattern-PrPres in bank voles that is transmissible upon second passage is highly concerning for public health. The fact that this biochemical signature in bank voles resembles that of the Wisc-1 original deer isolate and is different from that of bvWisc-1, in the migration profile and the glyco-form-ratio, is valid evidence that these results are not a product of contamination in our study. If CWD in humans is found to be contagious and transmissible among humans, as it is in cervids [57], the spread of the disease within humans might become endemic. In contrast to bank voles inoculated with fecal homogenates from mouse #327, so far, we could not detect a PK-resistant PrPSc fragment in the brain homogenates of fecal homogenate-inoculated tg650 mice. The presence of PrPres in these mice will allow us to determine if the molecular signature of hCWD prions from the brain (mouse #321 and #3063) vs feces are the same. Previously, Beringue et al. found that extraneural prions, compared to neural prions, helped more to overcome the species barrier to foreign prions, in addition, different strain types emerged from such serial transmission [11]. Our data also suggest that prions found in the periphery may hold higher zoonotic potential than prions found in neural tissues. In fact, upon second passage, 50% of the tg650 mice inoculated with fecal homogenates from mouse #327 had succumbed with terminal disease compared to only 20% of brain/spinal cord homogenates inoculated-tg650 mice suggesting that hCWD prions found in feces transmit disease more efficiently. Our results also suggest that epidemiological studies [25] may have missed subclinical and atypical infections that are/might be transmissible, undetected by the gold standard tests, i.e., Western blot, ELISA, and IHC.
Overall, our findings suggest that CWD surveillance in humans should encompass a wider spectrum of tissues/organs tested and include new criteria in the diagnosis of potential patients.
link.springer.com/article/10.1007/s00401-022-02482-9
terry
Heterozygosity for cervid S138N polymorphism results in subclinical CWD in gene-targeted mice and progressive inhibition of prion conversion
HIGHLIGHT
However, prion seeding activity was detected in spleens, brains, and feces of these mice, suggesting subclinical infection accompanied by prion shedding.
Infected animals accumulate prions in lymphoreticular and other peripheral tissues, e.g., skeletal muscle, and shed infectious prions in saliva, urine and feces, contributing to direct and environmental transmission and rapidly increasing geographic distribution of CWD (9–12).
However, prion-seeding activity was detectable in the brain, spleen, and feces, indicating subclinical infection and potential for contagiousness.
Heterozygosity for cervid S138N polymorphism results in subclinical CWD in gene-targeted mice and progressive inhibition of prion conversion
Maria I. Arifin orcid.org/0000-0003-2042-3492, Lech Kaczmarczyk orcid.org/0000-0003-2747-3134, Doris Zeng orcid.org/0009-0002-2512-6227, +7, and Sabine Gilch orcid.org/0000-0001-5923-3464 sgilch@ucalgary.caAuthors Info & Affiliations Edited by Reed Wickner, NIH, Bethesda, MD;
received December 12, 2022; accepted March 6, 2023
April 4, 2023
120 (15) e2221060120
doi.org/10.1073/pnas.2221060120
Significance
Amino acid substitutions within the cervid prion protein (PrP) can decrease susceptibility to chronic wasting disease, generally with more prominent effects in homozygous animals. Using novel gene-targeted mouse models expressing S138N reindeer/caribou PrP, we demonstrate subclinical infection with prion seeding activity in spleen and fecal prion shedding in heterozygous 138SN and homozygous 138NN mice. A lower percentage of heterozygous 138SN-PrP than homozygous 138NN-PrP expressing mice harbored seeding-efficient prions in tissues. This is caused by dominant-negative interference of the PrP variants occurring only if they are coexpressed. Our findings are relevant to inform conservation efforts for caribou, an endangered species in North America. Furthermore, our study provides new mechanistic insights into genetic resistance and dominant-negative interference of conversion-competent PrP variants.
Abstract
Prions are proteinaceous infectious particles that replicate by structural conversion of the host-encoded cellular prion protein (PrPC), causing fatal neurodegenerative diseases in mammals. Species-specific amino acid substitutions (AAS) arising from single nucleotide polymorphisms within the prion protein gene (Prnp) modulate prion disease pathogenesis, and, in several instances, reduce susceptibility of homo- or heterozygous AAS carriers to prion infection. However, a mechanistic understanding of their protective effects against clinical disease is missing. We generated gene-targeted mouse infection models of chronic wasting disease (CWD), a highly contagious prion disease of cervids. These mice express wild-type deer or PrPC harboring the S138N substitution homo- or heterozygously, a polymorphism found exclusively in reindeer (Rangifer tarandus spp.) and fallow deer (Dama dama). The wild-type deer PrP-expressing model recapitulated CWD pathogenesis including fecal shedding. Encoding at least one 138N allele prevented clinical CWD, accumulation of protease-resistant PrP (PrPres) and abnormal PrP deposits in the brain tissue. However, prion seeding activity was detected in spleens, brains, and feces of these mice, suggesting subclinical infection accompanied by prion shedding. 138N-PrPC was less efficiently converted to PrPres in vitro than wild-type deer (138SS) PrPC. Heterozygous coexpression of wild-type deer and 138N-PrPC resulted in dominant-negative inhibition and progressively diminished prion conversion over serial rounds of protein misfolding cyclic amplification. Our study indicates that heterozygosity at a polymorphic Prnp codon can confer the highest protection against clinical CWD and highlights the potential role of subclinical carriers in CWD transmission.
snip...
To conclude, our study demonstrates that CWD-infected animals harboring S138N PrP might be “silent spreaders” of CWD prions and highlights the importance of lymphatic tissues in the detection of CWD, particularly in caribou, even in the absence of clinical manifestation. It is important to keep in mind that even protective genotypes may be permissive to certain minor or newly emerging CWD strains. Our results provide new mechanistic insights into dominant-negative inhibition of prion conversion, the tissue specificity of this effect, and suggests that PrPC primary structure is a determinant for tissue-specific prion replication.
www.pnas.org/doi/10.1073/pnas.2221060120
Published: 22 August 2022
Transmission of cervid prions to humanized mice demonstrates the zoonotic potential of CWD
Samia Hannaoui, Irina Zemlyankina, Sheng Chun Chang, Maria Immaculata Arifin, Vincent Béringue, Debbie McKenzie, Hermann M. Schatzl & Sabine Gilch
Acta Neuropathologica volume 144, pages767–784 (2022)
Inoculation of these mice with deer CWD isolates resulted in atypical clinical manifestation with prion seeding activity and efficient transmissible infectivity in the brain and, remarkably, in feces, but without classical neuropathological or Western blot appearances of prion diseases.
In contrast, in cervids affected with CWD, infectivity has been found in the lymphatic system, salivary gland, intestinal tract, muscles, antler velvet, blood, urine, saliva, and feces [4], which have been demonstrated to be transmissible [57].
These data demonstrate that humanized tg650 mice inoculated with CWD prions shed prion infectivity in feces able to generate transmissible PrPSc in bank voles distinct from those generated by inoculation of the Wisc-1 deer isolate directly to bank voles.
Our findings strongly suggest that CWD should be regarded as an actual public health risk. Here, we use humanized mice to show that CWD prions can cross the species barrier to humans, and remarkably, infectious prions can be excreted in feces.
The presence of infectious prions shed in feces is one argument in favor of the existence of de novo generated PrPSc in these mice.
CWD in humans might remain subclinical but with PrPSc deposits in the brain with an unusual morphology that does not resemble the patterns usually seen in different prion diseases (e.g., mouse #328; Fig. 3), clinical with untraceable abnormal PrP (e.g., mouse #327) but still transmissible and uncovered upon subsequent passage (e.g., mouse #3063; Fig. 4), or prions have other reservoirs than the usual ones, hence the presence of infectivity in feces (e.g., mouse #327) suggesting a potential for human-to-human transmission and a real iatrogenic risk that might be unrecognizable. Here, humanized mice inoculated with CWD deer isolates had an atypical onset of the disease with myoclonus (93.75%), before presenting typical clinical signs, generating prions that presented with either atypical biochemical signature (#321 and #3063), shed in feces (#327), or were undetectable by the classical detection methods.
The finding that infectious PrPSc was shed in fecal material of CWD-infected humanized mice and induced clinical disease, different tropism, and typical three banding pattern-PrPres in bank voles that is transmissible upon second passage is highly concerning for public health. The fact that this biochemical signature in bank voles resembles that of the Wisc-1 original deer isolate and is different from that of bvWisc-1, in the migration profile and the glyco-form-ratio, is valid evidence that these results are not a product of contamination in our study. If CWD in humans is found to be contagious and transmissible among humans, as it is in cervids [57], the spread of the disease within humans might become endemic. In contrast to bank voles inoculated with fecal homogenates from mouse #327, so far, we could not detect a PK-resistant PrPSc fragment in the brain homogenates of fecal homogenate-inoculated tg650 mice. The presence of PrPres in these mice will allow us to determine if the molecular signature of hCWD prions from the brain (mouse #321 and #3063) vs feces are the same. Previously, Beringue et al. found that extraneural prions, compared to neural prions, helped more to overcome the species barrier to foreign prions, in addition, different strain types emerged from such serial transmission [11]. Our data also suggest that prions found in the periphery may hold higher zoonotic potential than prions found in neural tissues. In fact, upon second passage, 50% of the tg650 mice inoculated with fecal homogenates from mouse #327 had succumbed with terminal disease compared to only 20% of brain/spinal cord homogenates inoculated-tg650 mice suggesting that hCWD prions found in feces transmit disease more efficiently. Our results also suggest that epidemiological studies [25] may have missed subclinical and atypical infections that are/might be transmissible, undetected by the gold standard tests, i.e., Western blot, ELISA, and IHC.
Overall, our findings suggest that CWD surveillance in humans should encompass a wider spectrum of tissues/organs tested and include new criteria in the diagnosis of potential patients.
link.springer.com/article/10.1007/s00401-022-02482-9
terry