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Prion White Paper


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Issue:  Can the potential presence of prions in land applied biosolids result in food chain contamination with the subsequent development of animal and human disease?

What are Prions and What are the Diseases Attributed to Exposure to Prions?  
“Prion” refers to a particular kind of protein found in animal tissue. Most prions occur in a normal, harmless form, but there are abnormal or infectious forms. The normal, harmless form has the same sequence of amino acids as the abnormal form, but the abnormal, or infectious, form takes a different folded shape. (Epstein, 2005)1. In their normal, non-infectious state, it is believed that prions are involved in cell-to-cell communications and other important functions.  Unlike bacteria and viruses, prions do not contain genetic material. However, like viruses and bacteria, prions are infectious and replicate in host tissues. Throughout this white paper the word “Prion” is used to indicate the abnormal, infectious form of prions. Prions cause normal celluar proteins to convert to the abnormal or prion form.  In animals affected with prion-caused diseases, prions have been found mainly in the brain, spinal cord, lymph nodes, spleen, tonsils, eyes, pancreas, adrenal gland, and blood.  In studies with mice, prions have been observed in muscle tissue.  Prions have not been observed in manure or biosolids. 

It is now commonly accepted that prions are responsible for a number of previously known but little-understood animal (including human) diseases generally classified under transmissible spongiform encephalopathy diseases (TSEs) (Wikipedia, 2005)2.  These diseases affect the structure of brain tissue and are all fatal and untreatable.  The TSE diseases that have received the most attention recently include chronic wasting disease (CWD) that affects deer and elk, bovine spongiform encephalopathy (BSE) that affects cattle (“Mad Cow Disease”)(Collinge, 2001)3, and Creutzfeldt-Jacob Disease (CJ Disease) that affects humans.

What Are the Sources of Prions That May be Relevant to Wastewater Treatment and Biosolids Production?

  • Abattatoirs, Animal Rendering, and Meat Processing Operations - These operations, if they process BSE-contaminated cattle, can serve as a potential source of prions in wastewater treatment plants.  However, preliminary calculations on a worst case scenario in which the entire prion-infected brain of a slaughtered cattle were released into a wastewater treatment plant over the span of a day indicate that the resulting concentration of prions in the treatment plant’s effluent would be significantly less than the prion concentration necessary to infect an individual (assuming that the individual was directly consuming the treatment plant’s effluent (Pederson, 2005)4).  A recent effluent guideline (USEPA, 2004a)5 and general pretreatment standards promulgated by the United States Environmental Protection Agency (USEPA) for the Meat and Poultry Products Point Source Category, and the ability of local wastewater treatment authorities to impose these guidelines and standards on these discharging operations make it extremely unlikely that this assumed mass of prion-infected tissue would ever enter the sewer.
  • Landfill Leachates - Leachates from landfills can act as a potential source of prions to a wastewater treatment plant if the landfill accepts for disposal carcasses from BSE-contaminated cattle or CWD-contaminated deer or elk.  However, in areas where CWD is being actively managed, the disposal of contaminated deer or elk carcasses in municipal solid waste landfills appears to be an uncommon practice.  CWD management approaches in the United States in areas of known prion infectivity typically involve incineration of infected materials.  For example, the State of Wisconsin’s policy is to test potentially prion- infected deer and elk and to subsequently incinerate all prion positive deer and elk carcasses and landfill all prion negative carcasses (Kester, 2005)6.   Because prions are positively charged, and have been described as being “sticky”, they are likely to strongly sorb to solids and soils both in the landfill and to the landfill liner (Taylor, 2005)7.  Therefore, even if BSE-contaminated (prion) material were disposed in a municipal solid waste landfill that sent leachate to a wastewater treatment plant, the potential that significant concentrations of prions would be contained in the leachate is very low.  This should be confirmed once analytical methodologies are developed to determine prions in leachate. 
  • Urine, feces, and blood from CJ Disease patients - Several researchers (Gabizon, et. al. 20018; Reichl, et. al. 20029) have reported the presence of prions in the blood and urine of CJ Disease patients. Prions have not been reported in the feces of CJ Disease patients. It should be noted that the concentrations of prions in the blood and urine of CJ Disease patients would be relatively low and, after entry into the sewer with a vast amount of dilution available, even lower compared to the concentrations of prions in the neural tissue of these patients or in the neural tissue of BSE-infected meat that these patients may have consumed.  This is important to consider since the risk assessment results presented below are based on these significantly higher prion contaminated materials than the levels of prion contamination in blood, urine, or untreated wastewaters.

Have Prions Been Detected in Wastewater or Biosolids?
There are no reports in the scientific literature of the presence of prions in municipal wastewater or in biosolids.  Currently, no validated analytical methodologies are available for the determination of prions in municipal wastewater influent, treated municipal wastewater effluent, or in biosolids.  Once these analytical methods are developed, prions might be detected and quantified in these media. However, based on the discussion above, their concentrations would be expected to be extremely low and not capable of causing subsequent infection either through direct contact or indirectly through food chain contamination.  Analytical methodologies exist for the detection of prions in brain and other neurological tissue, mammalian lymphoid tissue, blood, and urine.  Prions have been detected in each of these biological materials (Pederson, 2005)4.

What are the Methods of Prion Destruction?
Prions found in environmental media or in residuals such as biosolids appear to be extremely resistant to degradation and loss of infectivity.  Current methods for denaturing prions and significantly reducing infectivity such as high temperatures and treatment with alkalis and bleach are applicable to prion-contaminated animal tissues but are not applicable to wastewater and biosolids treatment.

What Current Research and Measures Are Underway to Mitigate/Prevent Prion Entry Into Wastewater Treatment Plants?

  • Abattatoirs, Animal Rendering, and Meat Processing Operations - The U.S. EPA in 2004 published an effluent guideline (regulatory standard) for the Meat and Poultry Products Point Source Category that will result in a dramatic reduction in the amount of animal tissue that can be discharged directly into the aquatic environment. (USEPA, 2004a)5.  The technologies associated with this standard are designed to enhance ambient water quality by reducing the amount of solids such as animal tissues, biochemical oxygen demand (BOD), and ammonia that can be discharged into the aquatic environment.  Although this regulation does not pertain nationally to operations that discharge into sanitary sewers (“indirect dischargers”), local wastewater treatment authorities are free to impose the standard’s technologies on these indirect dischargers through local pretreatment limits where the potential for the processing of infected animals exist.  These operations are also subject to EPA’s General pretreatment regulations which will also reduce solids input to the sewer.   The United Kingdom has promulgated guidance for their meat processing industry on practices that minimize the amount of neurological tissue lost to the sewer (Gale and Stanfield, 2001)10.
  • Landfill Leachates - EPA has issued guidance on the operations of municipal solid waste landfills that accept prion-contaminated animal carcasses for disposal (USEPA, 2004b)11.  This guidance discusses the importance of liners and leachate collection systems and recirculation of the leachate in the landfill rather than discharge of the leachate to the wastewater treatment plant for containment of prions at the landfill site. 
  • Urine, Feces, and Blood from CJ Disease Patients - There are no current regulations, at least at the Federal level, that prohibit pathology laboratories or mortuaries from disposing of prion-contaminated tissue and fluids of CJ Disease patients into the sanitary sewer.  However, EPA has developed a draft strategy to reduce the prion contamination threat from the discharge of wastewater into the sanitary sewer from pathology/necropsy and research laboratories working with prion-contaminated tissues. (USEPA, 2005a)12.
  • Research - Currently, there is at least one research effort (the University of Wisconsin/Madison funded by EPA) underway to characterize the potential presence and fate of prions in wastewater treatment plants.  These studies will also determine the potential for prions to partition into and concentrate in biosolids (USEPA, 2005b)13.

What Are the Properties, Fate, and Transport of Prions in Wastewater Treatment and in the Land Application of Biosolids?
Very little data in this area is available.  Based on the properties of prions, it is expected that prions initially in wastewater (most likely at very small concentrations- see above discussion) will survive and most likely be attached to and be transported by solid particulates in the wastewater entering the wastewater treatment plant.  Once in the wastewater treatment process, no significant decrease in prion infectivity or prion degradation is expected to occur because of prions’ resistance to physical and chemical conditions encountered in wastewater treatment plants.

Whatever little concentration of prions in the incoming wastewater, they are expected to strongly partition to and concentrate in biosolids during wastewater treatment.  Research in progress will provide a quantitative estimate of this partitioning (USEPA, 2005b)13.  Based on the physical and chemical stability of prions, it is expected that prions will persist in biosolids, albeit at expectedly very low levels with respect to potential infectivity and the very limited number of potential environmental transport pathways available to infect animals or humans.  

Prions and their infectivity related to an animal TSE have been demonstrated to persist in soils for several years (Brown, 1991)14.  Because of their strong affinity with solid particulates and, therefore, very low concentrations in the aqueous phase, prions are not expected to threaten human-consumed or animal feed crops through root uptake in biosolids land application.  For the same reason, transport of prions to groundwater or surface waters from biosolids land application is not anticipated.  Prions have no volatility so ambient air transport can be ruled out.  The only potential significant environmental transport mechanism available for prions with subsequent exposure and potential infectivity to animals and humans is biosolids/soil ingestion by grazing ruminants and, theoretically, biosolids/soil ingestion by toddlers in a home garden scenario.  However, for these potential pathways of exposure, it is highly unlikely that prion concentration in the biosolids could ever approach an infectious dose for either animals or humans based on the extremely high dilution that occurs in wastewater treatment plants if prion-contaminated tissue were discharged to these plants and the prions subsequently partitioned to the biosolids (see discussions in previous sections and the section below).

What is the Risk to Human Health From BSE in Wastewater Treatment?
In 2001, Gale and Stanfield performed a quantitative risk assessment for BSE in biosolids for land application to cattle pasturing and vegetable crop production in the United Kingdom (UK)(Gale and Stanfield, 2001)10.  Using a worse case set of scenarios, they concluded: The risks to humans through consumption of vegetable crops are extremely low (approaches zero).  Although the risks to cattle are higher, because of their higher exposure to soil and greater susceptibility to prion infectivity, the risk assessment model demonstrates that biosolids containing trace quantities of prions alone cannot initiate or sustain a BSE epidemic in the UK cattle herd.  The conclusions are consistent with the findings from epidemiological studies, which so far, have not detected horizontal transmission of BSE (including transmission from BSE-contaminated pastures) (Gale and Stanfield, 2001).  The risk assessment demonstrates the importance of containment of neurological tissue from animal processing operations and absolutely minimizing or eliminating the amount of neurological tissue from BSE-infected animals that enter the sewer system.

Other “first order” risk assessments and estimates have demonstrated under worst-case scenarios extremely low risks to the theoretically highest exposed population, the farmer, from prions in land applied biosolids .  It should be noted that these risk assessments are performed on subpopulations that are at “bounded” maximum exposures.  In reality, compared to these subpopulations that are used for risk estimation purposes, almost all people living in countries with mature and regulated agricultural industries are exposed orders of magnitude less to prions or for that matter to any other chemical or biological agent that can be found in trace quantities in biosolids or in background soils.  This in turn results in orders of magnitude less risk to the general population from theoretical or actual exposure to these substances.

Summary
The information presented in this fact sheet strongly suggests that the risk of prion transmission directly to ruminants and indirectly to humans with subsequent infection from biosolids land application is extremely low and indeed is practically zero.  Prion transmission via biosolids land application seems less likely than other potential food chain pathways such as the consumption of prion-contaminated feed in animal raising operations and prion transmission to or between humans via contaminated surgical instruments and blood products, all of which are relatively rare, and compared to which, biosolids transmission of prions is even rarer.

There is an ongoing need for additional research in the areas described in this fact sheet to better quantify the information presented herein.  Results of this research should further expand the scientific knowledge based on the subject of prions.


References 

1. Epstein E. and N. Beecher. 2005.  Mad Cow Disease, Creutzfeld-Jakob Disease, other TSEs and Biosolids.  J. Residuals Science and Technology. 2(3): 181-187.

2. Wikipedia. 2005.  The Free Encyclopedia.  “Transmissible spongiform encephalopathy”.  Available on the Internet.

3. Collinge J. 2001.   Prion diseases of humans and animals: Their causes and molecular basis.  Ann. Rev. Neurosci. 24: 519-550.

4. Pedersen J. 2005.  Personal communication from Joel Pedersen, University of Wisconsin/Madison, to Alan B. Rubin.

5. USEPA. 2004a. Effluent Limitations Guidelines and New Source Performance Standards for the Meat and Poultry Products Point Source Category.  69 Federal Register (173):54475-54555.  September 8, 2004.

6. Kester G. 2005.  Personal communication from Greg Kester, Wisconsin Department of Natural Resources, to Alan B. Rubin.

7. Taylor D. 2005. Personal communication from David Taylor, Madison (WI) Metropolitan Sewerage District, to Alan B. Rubin.

8. Gabizon R., Shaked G.M., Shaked Y., Karn-Inbal Z., Halami M., and I. Avraham.  2001.  A protease resistant prion protein isoform is present in urine of animals and humans affected with prion diseases.  J. Biol. Chem. 276(34): 31479-31482.

9. Reichl H., Balen A., and C.A. Jansen.  2002.  Prion transmission in blood and urine: What are the implications for recombinant and urinary-derived gonadotropins?  Human Reprod. (10): 2501-2508.

10. Gale P. and G. Stanfield. 2001.  Towards a quantitative risk assessment for BSE in sewage sludge.  Journal of Applied Microbiology.  91:563-569.

11. USEPA. 2004b. Recommended Interim Practices for Disposal of Potentially Contaminated Chronic Wasting Disease Carcasses and Wastes.  Memorandum from: Robert Springer, Director, Office of Solid Waste to:  RCRA Division Directors (Regions I-X), Superfund Division Directors (Regions I-X), OSWER Office Director.  April 6, 2004.

12. USEPA. 2005a. EPA Draft Strategy Addendum to the Region 8 Local Limits Strategy.  Discharges of Wastewater to Publicly-Owned Treatment Works (POTWs) from Pathology/Necropsy and Research Laboratories Working with Prion-Contaminated Tissue.  Industrial Pretreatment Program (8P-W-P).  May 9, 2005.

13. USEPA. 2005b. Preliminary Results from the First Phase of a Two Phase Study Examining the Fate of Prions in Wastewater Treatment.  Poster presentation.  USEPA Science Forum, Washington, DC.  May 17, 2005.

14. Brown P. and D.C. Gajdusek. 1991.  Survival of scrapie virus after 3 years internment.  The Lancet. 337:269-270.