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Chronic wasting disease (CWD) is a transmissible and fatal prion disease that affects cervids. While both oral and nasal routes of exposure to prions cause disease, the spatial and temporal details of how prions enter the central nervous system (CNS) are unknown. Carotid bodies (CBs) are structures that are exposed to blood-borne prions and are densely innervated by nerves that are directly connected to brainstem nuclei, known to be early sites of prion neuroinvasion. All CBs examined contained mast cells expressing the prion protein which is consistent with these cells playing a role in neuroinvasion following prionemia.

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Chronic wasting disease (CWD) is a fatal neurodegenerative disease of cervids caused by an infectious misfolded protein (prion). Several members of the Cervidae, including Rocky Mountain elk (Cervus canadensis nelsoni), are susceptible to CWD. There is no evidence of complete genetic resistance to CWD; the M132L polymorphism in the elk prion protein gene influences the incubation period: longest in 132LL, intermediate in 132ML, and shortest in 132MM elk. We retrospectively analyzed six female 132LL elk housed in an environment heavily contaminated with prions to 1) document clinical outcomes and incubation periods, 2) describe PrPSc distribution and extent in tissues, and 3) characterize their histologic lesions. In five of six elk, PrPSc was detected postmortem, with a distribution pattern distinct from that of 132MM and 132ML elk; time to clinical CWD onset CWD ranged from 73 to 117 mo (6.1-9.8 yr). Although the remaining animal was observed for 220 mo (18.3 yr), PrPSc was not detected in its tissues postmortem. This study suggests that 132LL elk infected via natural exposure may live even longer with CWD than previously thought, but ultimately remain susceptible. We also report a distinct distribution of PrPSc in 132LL genotypes and highlight unusual histologic findings. Understanding the relationship between cervid genetics and CWD is of increasing importance, especially given the growing interest in leveraging genetics that delay disease onset despite not preventing infection.

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While animal prion diseases are a threat to human health, their zoonotic potential is generally inefficient because of interspecies prion transmission barriers. New animal models are required to provide an understanding of these prion transmission barriers and to assess the zoonotic potential of animal prion diseases. To address this goal, we generated Drosophila transgenic for human or nonhuman primate prion protein (PrP) and determined their susceptibility to known pathogenic prion diseases, namely varient Creutzfeldt-Jakob disease (vCJD) and classical bovine spongiform encephalopathy (BSE), and that with unknown pathogenic potential, namely chronic wasting disease (CWD). Adult Drosophila transgenic for M129 or V129 human PrP or nonhuman primate PrP developed a neurotoxic phenotype and showed an accelerated loss of survival after exposure to vCJD, classical BSE, or CWD prions at the larval stage. vCJD prion strain identity was retained after passage in both M129 and V129 human PrP Drosophila. All of the primate PrP fly lines accumulated prion seeding activity and concomitantly developed a neurotoxic phenotype, generally including accelerated loss of survival, after exposure to CWD prions derived from different cervid species, including North American white-tailed deer and muntjac, and European reindeer and moose. These novel studies show that primate PrP transgenic Drosophila lack known prion transmission barriers since, in mammalian hosts, V129 human PrP is associated with severe resistance to classical BSE prions, while both human and cynomolgus macaque PrP are associated with resistance to CWD prions. Significantly, our data suggest that interspecies differences in the amino acid sequence of PrP may not be a principal determinant of the prion transmission barrier.

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Pathogen transmission pathways are fundamental to understanding the epidemiology of infectious diseases yet are challenging to estimate in nature, particularly in the ocean. Seagrass wasting disease (SWD), caused by Labyrinthula zosterae, impacts seagrass beds worldwide and is thought to be a contributing factor to declines; however, little is known about natural transmission of SWD. In this study, we used field and laboratory experiments to test SWD transmission pathways and temperature sensitivity. To test transmission modes in nature, we conducted three field experiments out-planting sentinel Zostera marina shoots within and adjacent to natural Z. marina beds (20 ± 5 and 110 ± 5 m from bed edge). Infection rates and severity did not differ among outplant locations, implicating waterborne transmission. The infectious dose of L. zosterae through waterborne exposure was assessed in a controlled laboratory experiment. The dose to 50% disease was 6 cells ml-1 and did not differ with the temperatures tested (7.5°C and 15°C). Our results show L. zosterae is transmissible through water without direct contact with infected plants. Understanding the transmission dynamics of this disease in the context of changing ocean conditions will improve Z. marina protection and restoration in critical coastal habitats worldwide.

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The structure of cellular prion proteins encoded by the prion protein gene (PRNP) impacts susceptibility to transmissible spongiform encephalopathies, including chronic wasting disease (CWD) in deer. The recent emergence of CWD in Northern European reindeer (Rangifer tarandus), moose (Alces alces alces) and red deer (Cervus elaphus), in parallel with the outbreak in North America, gives reason to investigate PRNP variation in European deer, to implement risk assessments and adjust CWD management for deer populations under threat. We here report PRNP-sequence data from 911 samples of German red, roe (Capreolus capreolus), sika (Cervus nippon) and fallow deer (Dama dama) as well as additional data from 26 Danish red deer close to the German border and four zoo species not native to Germany. No PRNP sequence variation was observed in roe and fallow deer, as previously described for populations across Europe. In contrast, a broad PRNP variation was detected in red deer, with non-synonymous polymorphisms at codons 98, 226 and 247 as well as synonymous mutations at codons 21, 78, 136 and 185. Moreover, a novel 24 bp deletion within the octapeptide repeat was detected. In summary, 14 genotypes were seen in red deer with significant differences in their geographical distribution and frequencies, including geographical clustering of certain genotypes, suggesting "PRNP-linages" in this species. Based on data from North American CWD and the genotyping results of the European CWD cases, we would predict that large proportions of wild cervids in Europe might be susceptible to CWD once introduced to naive populations.

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Chronic wasting disease (CWD), a prion disease affecting cervids, has been known in North America (NA) since the 1960s and emerged in Norway in 2016. Surveillance and studies have revealed that there are different forms of CWD in Fennoscandia: contagious CWD in Norwegian reindeer and sporadic CWD in moose and red deer. Experimental studies have demonstrated that NA CWD prions can infect various species, but thus far, there have been no reports of natural transmission to non-cervid species. In vitro and laboratory animal studies of the Norwegian CWD strains suggest that these strains are different from the NA strains. In this work, we describe the intracerebral transmission of reindeer CWD to six scrapie-susceptible sheep. Detection methods included immunohistochemistry (IHC), western blot (WB), enzyme-linked immunosorbent assay (ELISA), real-time quaking-induced conversion (RT-QuIC) and protein misfolding cyclic amplification (PMCA). In the brain, grey matter vacuolation was limited, while all sheep exhibited vacuolation of the white matter. IHC and WB conventional detection techniques failed to detect prions; however, positive seeding activity with the RT-QuIC and PMCA amplification techniques was observed in the central nervous system of all but one sheep. Prions were robustly amplified in the lymph nodes of all animals, mainly by RT-QuIC. Additionally, two lymph nodes were positive by WB, and one was positive by ELISA. These findings suggest that sheep can propagate reindeer CWD prions after intracerebral inoculation, resulting in an unusual disease phenotype and prion distribution with a low amount of detectable prions.

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White-tailed deer are susceptible to scrapie (WTD scrapie) after oronasal inoculation with the classical scrapie agent from sheep. Deer affected by WTD scrapie are difficult to differentiate from deer infected with chronic wasting disease (CWD). To assess the transmissibility of the WTD scrapie agent and tissue phenotypes when further passaged in white-tailed deer, we oronasally inoculated wild-type white-tailed deer with WTD scrapie agent. We found that WTD scrapie and CWD agents were generally similar, although some differences were noted. The greatest differences were seen in bioassays of cervidized mice that exhibited significantly longer survival periods when inoculated with WTD scrapie agent than those inoculated with CWD agent. Our findings establish that white-tailed deer are susceptible to WTD scrapie and that the presence of WTD scrapie agent in the lymphoreticular system suggests the handling of suspected cases should be consistent with current CWD guidelines because environmental shedding may occur.

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Since prion diseases result from infection and neurodegeneration of the central nervous system (CNS), experimental characterizations of prion strain properties customarily rely on the outcomes of intracerebral challenges. However, natural transmission of certain prions, including those causing chronic wasting disease (CWD) in elk and deer, depends on propagation in peripheral host compartments prior to CNS infection. Using gene-targeted GtE and GtQ mice, which accurately control cellular elk or deer PrP expression, we assessed the impact that peripheral or intracerebral exposures play on CWD prion strain propagation and resulting CNS abnormalities. Whereas oral and intraperitoneal transmissions produced identical neuropathological outcomes in GtE and GtQ mice and preserved the naturally convergent conformations of elk and deer CWD prions, intracerebral transmissions generated CNS prion strains with divergent biochemical properties in GtE and GtQ mice that were changed compared to their native counterparts. While CWD replication kinetics remained constant during iterative peripheral transmissions and brain titers reflected those found in native hosts, serial intracerebral transmissions produced 10-fold higher prion titers and accelerated incubation times. Our demonstration that peripherally and intracerebrally challenged Gt mice develop dissimilar CNS diseases which result from the propagation of distinct CWD prion strains points to the involvement of tissue-specific cofactors during strain selection in different host compartments. Since peripheral transmissions preserved the natural features of elk and deer prions, whereas intracerebral propagation produced divergent strains, our findings illustrate the importance of experimental characterizations using hosts that not only abrogate species barriers but also accurately recapitulate natural transmission routes of native strains.

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Disease co-occurrence in wildlife populations is common yet understudied. In the case of disease-caused mortality, the mortality attributed to one disease has the potential to buffer populations against subsequent alternative disease outbreaks by reducing populations and thus contacts needed to sustain disease transmission. However, substantial disease-driven population declines may also prevent populations from recovering, leading to localized extinctions. Hemorrhagic disease (HD), a vector-transmitted, viral disease in white-tailed deer (WTD), similar to chronic wasting disease (CWD), a prion disease, has increased in frequency and distribution in the United States. However, unlike CWD, which progresses slowly, HD can cause mortality only days after infection. Hemorrhagic disease outbreaks can result in substantial localized mortality events in WTD near vector habitats such as wetlands and may reduce local deer densities and consequent CWD transmission. The objective of our study was to evaluate the potential for HD outbreaks to buffer CWD risk where the diseases co-occur. Using an agent-based modeling approach, we found that frequent, intense HD outbreaks have the potential to mitigate CWD risk, especially if those outbreaks occur shortly after CWD introduction. However, HD outbreaks that do not result in substantial WTD mortality are unlikely to impact CWD or WTD population dynamics. Severe HD outbreaks may reduce CWD cases and could present an opportunity for managers to boost CWD control initiatives in a post-HD outbreak year.

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Aims: Redox signaling plays a key role in skeletal muscle remodeling induced by exercise and prolonged inactivity, but it is unclear which oxidant triggers myofiber hypertrophy due to the lack of strategies to precisely regulate individual oxidants in vivo. In this study, we used tetrathiomolybdate (TM) to dissociate the link between superoxide (O2•-) and hydrogen peroxide and thereby to specifically explore the role of O2•- in muscle hypertrophy in C2C12 cells and mice. Results: TM can linearly regulate intracellular O2•- levels by inhibition of superoxide dismutase 1 (SOD1). A 70% increase in O2•- levels in C2C12 myoblast cells and mice is necessary and sufficient for triggering hypertrophy of differentiated myotubes and can enhance exercise performance by more than 50% in mice. SOD1 knockout blocks TM-induced O2•- increments and thereby prevents hypertrophy, whereas SOD1 restoration rescues all these effects. Scavenging O2•- with antioxidants abolishes TM-induced hypertrophy and the enhancement of exercise performance, whereas the restoration of O2•- levels with a O2•- generator promotes muscle hypertrophy independent of SOD1 activity. Innovation and Conclusion: These findings suggest that O2•- is an endogenous initiator of myofiber hypertrophy and that TM may be used to treat muscle wasting diseases. Our work not only suggests a novel druggable mechanism to increase muscle mass but also provides a tool for precisely regulating O2•- levels in vivo.

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Prion diseases such as scrapie, bovine spongiform encephalopathy (BSE), and chronic wasting disease (CWD) affect domesticated and wild herbivorous mammals. Animals afflicted with CWD, the transmissible spongiform encephalopathy of cervids (deer, elk, and moose), shed prions into the environment, where they may persist and remain infectious for years. These environmental prions may remain in soil, be transported in surface waters, or assimilated into plants. Environmental sampling is an emerging area of TSE research and can provide more information about prion fate and transport once shed by infected animals. In this study, we have developed the first published method for the extraction and detection of prions in plant tissue using the real-time quaking-induced conversion (RT-QuIC) assay. Incubation with a zwitterionic surfactant followed by precipitation with sodium phosphotungstate concentrates the prions within samples and allows for sensitive detection of prion seeding activity. Using this protocol, we demonstrate that prions can be detected within plant tissues and on plant surfaces using the RT-QuIC assay.

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Chronic wasting disease (CWD) is a contagious prion disease that affects cervids in North America, Northern Europe, and South Korea. CWD is spread through direct and indirect horizontal transmission, with both clinical and preclinical animals shedding CWD prions in saliva, urine, and feces. CWD particles can persist in the environment for years, and soils may pose a risk for transmission to susceptible animals. Our study presents a sensitive method for detecting prions in the environmental samples of prairie soils. Soils were collected from CWD-endemic regions with high (Saskatchewan, Canada) and low (North Dakota, USA) CWD prevalence. Heat extraction with SDS-buffer, a serial protein misfolding cyclic amplification assay coupled with a real-time quaking-induced conversion assay was used to detect the presence of CWD prions in soils. In the prairie area of South Saskatchewan where the CWD prevalence rate in male mule deer is greater than 70%, 75% of the soil samples tested were positive, while in the low-prevalence prairie region of North Dakota (11% prevalence in male mule deer), none of the soils contained prion seeding activity. Soil-bound CWD prion detection has the potential to improve our understanding of the environmental spread of CWD, benefiting both surveillance and mitigation approaches.

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After detecting chronic wasting disease (CWD) in white-tailed deer (Odocoileus virginianus) in Hampshire County, West Virginia, USA, in 2005, we investigated the change of CWD apparent prevalence and potential factors influencing infection risk during the invasion front. Over eight sampling years (2006-2012 and 2017) during a 12-yr period within a 101-km2-area monitoring zone, we sampled and tested a total of 853 deer for CWD by ELISA and immunohistochemistry. Bayesian logistic regression of risk factors included collection year, age class, sex, and adjusted body weight (weight after accounting for sex, age, kidney fat index, and number of fetuses). In the whole-herd model (n=634), collection year, age, and adjusted body weight were associated with increased odds of CWD, whereas an age-weight interaction had a negative relationship. We found that males drove the positive associations with age and adjusted body weight, whereas females were responsible for the negative interaction effect. These findings suggest potential behavioral and physiological mechanisms related to sex that may influence CWD exposure. Older males exhibited higher CWD prevalence, aligning with previous studies. Notably, the novel finding of adjusted body weight as a risk factor in males warrants further investigation, and this study highlights the need for future research on social behavior and its role in CWD transmission within white-tailed deer populations.

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Chronic wasting disease (CWD) is a transmissible spongiform encephalopathy that was first detected in captive cervids in Colorado, United States (US) in 1967, but has since spread into free-ranging white-tailed deer (Odocoileus virginianus) across the US and Canada as well as to Scandinavia and South Korea. In some areas, the disease is considered endemic in wild deer populations, and governmental wildlife agencies have employed epidemiological models to understand long-term environmental risk. However, continued rapid spread of CWD into new regions of the continent has underscored the need for extension of these models into broader tools applicable for wide use by wildlife agencies. Additionally, efforts to semi-automate models will facilitate access of technical scientific methods to broader users. We introduce software (Habitat Risk) designed to link a previously published epidemiological model with spatially referenced environmental and disease testing data to enable agency personnel to make up-to-date, localized, data-driven predictions regarding the odds of CWD detection in surrounding areas after an outbreak is discovered. Habitat Risk requires pre-processing publicly available environmental datasets and standardization of disease testing (surveillance) data, after which an autonomous computational workflow terminates in a user interface that displays an interactive map of disease risk. We demonstrated the use of the Habitat Risk software with surveillance data of white-tailed deer from Tennessee, USA.

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Chronic wasting disease (CWD) is a cervid prion disease with unknown zoonotic potential that might pose a risk to humans who are exposed. To assess the potential of CWD to infect human neural tissue, we used human cerebral organoids with 2 different prion genotypes, 1 of which has previously been associated with susceptibility to zoonotic prion disease. We exposed organoids from both genotypes to high concentrations of CWD inocula from 3 different sources for 7 days, then screened for infection periodically for up to 180 days. No de novo CWD propagation or deposition of protease-resistant forms of human prions was evident in CWD-exposed organoids. Some persistence of the original inoculum was detected, which was equivalent in prion gene knockout organoids and thus not attributable to human prion propagation. Overall, the unsuccessful propagation of CWD in cerebral organoids supports a strong species barrier to transmission of CWD prions to humans.

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Solanum glaucophyllum is a toxic plant with calcinogenic effect that causes enzootic calcinosis (EC) characterized by soft tissue metastatic mineralization mainly in cattle and rarely sheep, buffaloes, pigs, horses, and goats. We describe an outbreak of EC in a herd of 64 goats due to S. glaucophyllum consumption. Thirty-four goats were affected exhibiting hirsutism, stiffening, kyphosis and emaciation. Twelve goats died. Grossly, tissue mineralization was observed in the aorta and carotid arteries, lungs, and heart. Lesions were characterized by multiple rough white plaques, and hardened tissues with loss of elasticity. Microscopically, multisystemic mineralization was observed in aorta and carotid arteries, heart, lung, abomasum, intestine, spleen, lymph nodes, kidney, spleen, and meninges, characterized by extensive granular basophilic deposits of tunica media and/or intima of blood vessels; confirmed as calcium salt deposits with Von Kossa stain. We conclude that ingestion of S. glaucophyllum can cause EC in goats. Though EC is rare in goats under some conditions such as heavy drought and abundant S. glaucophyllum exposure disease can develop.

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The first case of CWD in a Norwegian red deer was detected by a routine ELISA test and confirmed by western blotting and immunohistochemistry in the brain stem of the animal. Two different western blotting tests were conducted independently in two different laboratories, showing that the red deer glycoprofile was different from the Norwegian CWD reindeer and CWD moose and from North American CWD. The isolate showed nevertheless features similar to the classical BSE (BSE-C) strain. Furthermore, BSE-C could not be excluded based on the PrPSc immunohistochemistry staining in the brainstem and the absence of detectable PrPSc in the lymphoid tissues. Because of the known ability of BSE-C to cross species barriers as well as its zoonotic potential, the CWD red deer isolate was submitted to the EURL Strain Typing Expert Group (STEG) as a BSE-C suspect for further investigation. In addition, different strain typing in vivo and in vitro strategies aiming at identifying the BSE-C strain in the red deer isolate were performed independently in three research groups and BSE-C was not found in it. These results suggest that the Norwegian CWD red deer case was infected with a previously unknown CWD type and further investigation is needed to determine the characteristics of this potential new CWD strain.

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Chronic wasting disease (CWD) can spread among cervids by direct and indirect transmission, the former being more likely in emerging areas. Identifying subpopulations allows the delineation of focal areas to target for intervention. We aimed to assess the population structure of white-tailed deer (Odocoileus virginianus) in the northeastern United States at a regional scale to inform managers regarding gene flow throughout the region. We genotyped 10 microsatellites in 5701 wild deer samples from Maryland, New York, Ohio, Pennsylvania, and Virginia. We evaluated the distribution of genetic variability through spatial principal component analysis and inferred genetic structure using non-spatial and spatial Bayesian clustering algorithms (BCAs). We simulated populations representing each inferred wild cluster, wild deer in each state and each physiographic province, total wild population, and a captive population. We conducted genetic assignment tests using these potential sources, calculating the probability of samples being correctly assigned to their origin. Non-spatial BCA identified two clusters across the region, while spatial BCA suggested a maximum of nine clusters. Assignment tests correctly placed deer into captive or wild origin in most cases (94%), as previously reported, but performance varied when assigning wild deer to more specific origins. Assignments to clusters inferred via non-spatial BCA performed well, but efficiency was greatly reduced when assigning samples to clusters inferred via spatial BCA. Differences between spatial BCA clusters are not strong enough to make assignment tests a reliable method for inferring the geographic origin of deer using 10 microsatellites. However, the genetic distinction between clusters may indicate natural and anthropogenic barriers of interest for management.

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Integrating host movement and pathogen data is a central issue in wildlife disease ecology that will allow for a better understanding of disease transmission. We examined how adult female mule deer (Odocoileus hemionus) responded behaviorally to infection with chronic wasting disease (CWD). We compared movement and habitat use of CWD-infected deer (n = 18) to those that succumbed to starvation (and were CWD-negative by ELISA and IHC; n = 8) and others in which CWD was not detected (n = 111, including animals that survived the duration of the study) using GPS collar data from two distinct populations collared in central Wyoming, USA during 2018-2022. CWD and predation were the leading causes of mortality during our study (32/91 deaths attributed to CWD and 27/91 deaths attributed to predation). Deer infected with CWD moved slower and used lower elevation areas closer to rivers in the months preceding death compared with uninfected deer that did not succumb to starvation. Although CWD-infected deer and those that died of starvation moved at similar speeds during the final months of life, CWD-infected deer used areas closer to streams with less herbaceous biomass than starved deer. These behavioral differences may allow for the development of predictive models of disease status from movement data, which will be useful to supplement field and laboratory diagnostics or when mortalities cannot be quickly retrieved to assess cause-specific mortality. Furthermore, identifying individuals who are sick before predation events could help to assess the extent to which disease mortality is compensatory with predation. Finally, infected animals began to slow down around 4 months prior to death from CWD. Our approach for detecting the timing of infection-induced shifts in movement behavior may be useful in application to other disease systems to better understand the response of wildlife to infectious disease.

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Infectious prions are resistant to degradation and remain infectious in the environment for several years. Chronic wasting disease (CWD) has been detected in cervids inhabiting North America, the Nordic countries, and South Korea. CWD-prion spread is partially attributed to carcass transport and disposal. We employed a forensic approach to investigate an illegal carcass dump site connected with a CWD-positive herd. We integrated anatomic, genetic, and prion amplification methods to discover CWD-positive remains from six white-tailed deer (Odocoileus virginianus) and, using microsatellite markers, confirmed a portion originated from the CWD-infected herd. This approach provides a foundation for future studies of carcass prion transmission risk.

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