Sharkey
This is a lot more info that is again abstract. The word "likely" isn't very scientific.
I don't think that anyone knows at what point a miss folded prion is shed by a living host.
This info was edited from a blog run by Terry Singletary.
Grass Plants Bind, Retain, Uptake, and Transport Infectious Prions
DOI:
http://dx.doi.org/10.1016/j.celrep.2015.04.036
•Grass plants bind prions from contaminated brain and excreta •Prions from different strains and species remain bound to living plants •Hamsters fed with prion-contaminated plant samples develop prion disease •Stems and leaves from grass plants grown in infected soil contain prions.
Summary
Prions are the protein-based infectious agents responsible for prion diseases. Environmental prion contamination has been implicated in disease transmission. Here, we analyzed the binding and retention of infectious prion protein (PrPSc) to plants. Small quantities of PrPSc contained in diluted brain homogenate or in excretory materials (urine and feces) can bind to wheat grass roots and leaves. Wild-type hamsters were efficiently infected by ingestion of prion-contaminated plants. The prion-plant interaction occurs with prions from diverse origins, including chronic wasting disease. Furthermore, leaves contaminated by spraying with a prion-containing preparation retained PrPSc for several weeks in the living plant. Finally, plants can uptake prions from contaminated soil and transport them to aerial parts of the plant (stem and leaves). These findings demonstrate that plants can efficiently bind infectious prions and act as carriers of infectivity, suggesting a possible role of environmental prion contamination in the horizontal transmission of the disease.
This is an open access article under the CC BY license (
http://creativecommons.org/licenses/by/4.0/).
Received: October 31, 2014; Received in revised form: February 4, 2015; Accepted: April 15, 2015; Published Online: May 14, 2015
© 2015 The Authors. Published by Elsevier Inc.
http://www.cell.com/cell-reports/abstract/S2211-1247(15)00437-4
Prions Bind to Plants and Bound-PrPSc Efficiently Sustain Prion Replication
To study whether plants can interact with prions, we exposed wheat grass roots and leaves to brain homogenate from hamsters that have succumbed to prion disease induced by experimental inoculation with the 263K prion strain. The presence of PrPSc and infectivity attached to the plants was studied in vitro using the protein misfolding cyclic amplification (PMCA) technique and in vivo by infectivity bioassays. For in vitro analyses, the plant tissues (roots and leaves) were incubated for 16 hr with serial dilutions of 263K-brain homogenate ranging from 10 1 to 10 8. Roots and leaves were washed thoroughly and analyzed for the presence of PrPSc by serial PMCA (Morales et al., 2012). The results show that even highly diluted PrPSc can bind to roots and leaves and sustain PrPC conversion (Figure 1A). Although a direct comparison cannot be made, because of differences on the effective surface, roots appear to retain PrPSc better than leaves. However, both roots and leaves capture PrPSc efficiently, even at very small concentrations, equivalent to those present in biological fluids, such as blood and urine (Chen et al., 2010). By comparing the detection of PrPSc-bound to plants (Figure 1A) with an experiment in which the same dilutions of 263K brain homogenate were added directly to the tubes containing normal brain homogenate and an equivalent piece of leaves or roots (Figure 1B), we can estimate that a high proportion of PrPSc present in the sample was attached to the plant tissue. Importantly, no detection of PrPSc was observed when leaves and roots were exposed to normal brain homogenate (Figure 1C). However, comparing PMCA amplification in the presence (Figure 1B) or in the absence (Figure S1A) of plant tissue, it is possible to appreciate that plants (both leaves and roots) partially inhibits the PMCA reaction. This explains why in most of the experiments with plants, protease-resistant PrPSc is only observed after two rounds of PMCA. In our current PMCA settings, no false-positive PrPSc signals were ever detectable when samples did not contain PrPSc inoculum (Figure S1B). These results indicate that leaves and roots can efficiently bind PrPSc, which remains able to catalyze PrPC to PrPSc conversion, leading to prion replication. In these experiments, plant tissues were incubated with prions for 16 hr, but a similar experiment in which roots and leaves were exposed to a 10 5 dilution of 263K brain homogenate for different times, we found that as little as 2 min of incubation was sufficient for the efficient contamination of plants (Figure S2).
Animals Can Be Infected by Oral Administration of Prion-Contaminated Plants
To investigate whether prion-contaminated plants were able to infect animals by ingestion, leaves and roots previously incubated with either 263K-infected or control hamster brain homogenates were orally administered into naive hamsters. After exposure, plants were extensively washed five times with water and animals were fed with dried material. As positive controls, we orally administered 750 ml of 5% 263K brain homogenate (same material used to contaminate plant tissue). All animals that ingested prion contaminated leaves and roots developed typical prion disease. Although the incubation times were significantly longer in animals ingesting prions attached to leaves and roots as compared with those fed directly with the brain material, the differences were not as high as one could have expected (Figure 2A). Indeed, incubation periods were 147 ± 10,
159 ± 10, and 164 ± 13 days (mean ± SEM) for the groups inoculated with brain homogenate, and prion contaminated roots and leaves, respectively. Prion disease was confirmed by histological study of PrPSc deposition, astrogliosis, and brain vacuolation (Figure 2B), as well as by biochemical detection of protease-resistant PrPSc by western blot (Figure 2C). None of the animals inoculated with leaves and roots exposed to normal brain homogenate developed disease up to 550 days post-inoculation. Histological analysis did not show any PrPSc staining or disease specific alteration in control animals.
Plants Bind Prions from Different Strains and Species To analyze prion-plant interaction with other species and strains of the prion agent, we performed similar studies as described in Figure 1, by incubating leaves and roots with a preparation containing hamster, murine, cervid, and human prions corresponding to the Hyper, 301C, CWD, and vCJD prion strains, respectively. PrPSc from these strains and species showed good amplification by PMCA, using homologous substrates (Figure S3A). In all cases, leaves and roots bound prions from these species and retained the ability to replicate in vitro (Figure S3B), indicating that the interaction of PrPSc with plants is a general feature of infectious prions.
Contamination of Plants with Prions Excreted in Urine and Feces
Under natural conditions, it is likely that the main source of prions in the environment comes from secretory and excretory fluids, such as saliva, urine, and feces. We and others have shown that PrPSc is released in these fluids and excretions in various animal species (Gonzalez-Romero et al., 2008; Haley et al., 2009, 2011; Maddison et al., 2010; Terry et al., 2011; Moda et al., 2014). It has been estimated that the amount of infectious prions spread by excreta during the animals’ lifespan could match or even surpass the quantity present in the brain of a symptomatic individual (Tamgu¨ ney et al., 2009). To study whether plant tissue can be contaminated by waste products excreted from prion-infected hamsters and deer, leaves and roots were incubated with samples of urine and feces and the presence of PrPSc analyzed by serial rounds of PMCA. For these experiments, plant tissues were incubated for 1 hr with urine or feces homogenates obtained either from 263K-infected hamsters or CWD-affected cervids. This time was chosen because longer incubation with these biological fluids affected the integrity of the plant tissue. After being thoroughly washed and dried, PrPSc attached to leaves and roots was detected by PMCA. The results clearly show that PrPSc was readily detectable after three or four rounds of PMCA in samples of wheat grass leaves and roots exposed to both urine and feces from 263K sick hamsters (Figure 3A) and CWD-affected cervids (Figure 3B). Comparing these results with studies of the direct detection of PrPSc in urine and feces (Figures 3A and 3B), it seems that the majority of PrPSc present in these waste products was effectively attached to leaves and roots. No signal was observed in plant tissue exposed to urine or feces coming from non-infected hamsters.
To investigate a more natural scenario for prion contamination of living plants, we sprayed the leaves of wheat grass with a preparation containing 1% 263K hamster brain homogenate. Plants were let to grow for different times after exposure, and PrPSc was detected in the leaves by PMCA in duplicates for each time point. The results show that PrPSc was able to bind to leaves and remained attached to the living plants for at least 49 days after exposure (Figure 4). Considering that PrPSc signal was detectable normally in the second or third round of PMCA without obvious trend in relation to time, we conclude that the relative amount of PrPSc present in leaves did not appear to change substantially over time. These data indicate that PrPSc can be retained in living plants for at least several weeks after a simple contact with prion contaminated materials, and PrPSc remains competent to drive prion replication.
Plants Uptake Prions from Contaminated Soil
The experiments described above were done by exposure of the surface of leaves and roots with different solutions containing prions. To evaluate whether living plants can uptake PrPSc from contaminated soil, we grew barley grass plants on soil that was contaminated by addition of 263K brain homogenate. Plants were grown for 1 or 3 weeks under conditions that carefully prevented any direct contact of the aerial part of the plant with the soil. After this time, pieces of stem and leaves were collected and analyzed for the presence of PrPSc by PMCA. As shown in Figure 5A, all plants grown for 3 weeks in contaminated soil contained PrPSc in their stem, albeit in small quantities that required four serial rounds of PMCA for detection. One of the four plants analyzed contained a detectable amount of PrPSc in the leaves (Figure 5B), indicating that prions were uptaken from the soil and transported into the aerial parts of the plants, far from the soil. These results differ from a recent article reporting that infectious prions were not detectable in above the ground tissues of wheat plants exposed to CWD prions (Rasmussen et al., 2014). The lack of detection in this article is most likely due to the low sensitive techniques (western blots or ELISA) employed to analyze the presence of PrPSc. Indeed, as we reported previously, PMCA has a power of detection, which is several millions times higher than western blots or ELISA (Saa´ et al., 2006). In order to estimate the amount of PrPSc present in stem and leaves coming from contaminated soil, we performed a quantitative PMCA study, as previously described (Chen et al., 2010). Unfortunately, by comparing the PMCA amplification in the absence or the presence of plant tissue, it is possible to conclude that stems and leaves substantially interfered with the PMCA procedure, and thus the calculation cannot be very precise (Figure S4). Indeed, after two rounds of PMCA we cannot detect any protease-resistant PrPSc, but on the third round we observed the maximum amplification (10 9), presumably because at this round the concentration of PMCA inhibitors has been reduced enough to permit good amplification. At this point, we can estimate that the amount of PrPSc that reaches the stem and leaves from contaminated soil is equivalent to the PrPSc concentration present in a 10 6 to 10 9 dilution of sick brain homogenate. Nevertheless, this result is interesting, because it indicates that the amount of prions uptaken from soil and transported to aerial parts of the plant is within the infectious range. Indeed, titration studies showed that the last infectious dilution of a 263K brain homogenate is 10 9 (Gregori et al., 2006).
This study shows that plants can efficiently bind prions contained in brain extracts from diverse prion infected animals, including CWD-affected cervids. PrPSc attached to leaves and roots from wheat grass plants remains capable of seeding prion replication in vitro. Surprisingly, the small quantity of PrPSc naturally excreted in urine and feces from sick hamster or cervids was enough to efficiently contaminate plant tissue. Indeed, our results suggest that the majority of excreted PrPSc is efficiently captured by plants’ leaves and roots. Moreover, leaves can be contaminated by spraying them with a prion-containing extract, and PrPSc remains detectable in living plants for as long as the study wasperformed (several weeks). Remarkably, prion contaminated plants transmit prion disease to animals upon ingestion, producing a 100% attack rate and incubation periods not substantially longer than direct oral administration of sick brain homogenates. Finally, an unexpected but exciting result was that plants were able to uptake prions from contaminated soil and transport them to aerial parts of the plant tissue. Although it may seem farfetched that plants can uptake proteins from the soil and transport it to the parts above the ground, there are already published reports of this phenomenon (McLaren et al., 1960; Jensen and McLaren, 1960; Paungfoo-Lonhienne et al., 2008). The high resistance of prions to degradation and their ability to efficiently cross biological barriers may play a role in this process. The mechanism by which plants bind, retain, uptake, and transport prions is unknown. Weare currently studying the way in which prions interact with plants using purified, radioactively labeled PrPSc to determine specificity of the interaction, association constant, reversibility, saturation, movement, etc.
Epidemiological studies have shown numerous instances of scrapie or CWD recurrence upon reintroduction of animals on pastures previously exposed to prion-infected animals. Indeed, reappearance of scrapie has been documented following fallow periods of up to 16 years (Georgsson et al., 2006), and pastures were shown to retain infectious CWD prions for at least 2 years after exposure (Miller et al., 2004). It is likely that the environmentally mediated transmission of prion diseases depends upon the interaction of prions with diverse elements, including soil, water, environmental surfaces, various invertebrate animals, and plants. However, since plants are such an important component of the environment and also a major source of food for many animal species, including humans, our results may have far-reaching implications for animal and human health. Currently, the perception of the risk for animal-to-humanprion transmissionhas beenmostly limited to consumption or exposure to contaminated meat; our results indicate that plants might also be an important vector of transmission that needs to be considered in risk assessment.
http://www.cell.com/cell-reports/pdf/S2211-1247(15)00437-4.pdf
Grass Plants Bind, Retain, Uptake, and Transport Infectious Prions
http://www.cell.com/cell-reports/pdfExtended/S2211-1247(15)00437-4
Friday, September 27, 2013
Uptake of Prions into Plants
http://chronic-wasting-disease.blogspot.com/2013/09/uptake-of-prions-into-plants.html
Friday, May 15, 2015 Grass Plants Bind, Retain, Uptake, and Transport Infectious Prions
Report
http://transmissiblespongiformencephalopathy.blogspot.com/2015/05/grass-plants-bind-retain-uptake-and.html
