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Title: Prion infected Meat-and-Bone Meal is still infectious after biodiesel production

item Bruederle, Cathrin
item Hnasko, Robert
item Garcia, Rafael
item Haas, Michael
item Marmer, William
item Carter, John

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/24/2008
Publication Date: 8/13/2008
Citation: Bruederle, C.E., Hnasko, R.M., Kraemer, T., Garcia, R.A., Haas, M.J., Marmer, W.N., Carter, J.M. 2008. Prion infected Meat-and-Bone Meal is still Infectious After Biodiesel Production. PLoS One 3(8): e2969. doi:10.1371/journal.pone.0002969.

Interpretive Summary: The problem addressed by this research is safe disposal of slaughterhouse by-products. Some parts of cattle and sheep have low commercial value as human food. Much of this material has traditionally been processed into Meat-and-Bone Meal (MBM), a highly nutritious animal feed supplement. But MBM was linked to the spread of Bovine Spongiform Encephalopathy (BSE), and the BSE epidemic in the UK and EU was probably ended by banning MBM from feed. This has created a problem for MBM as an agricultural commodity. MBM is high in fat, and rising prices for crude oil have recently created a favorable market shift for biodiesel, which can be produced from this fat through a simple but harsh chemical reaction. Biodiesel is usually manufactured from relatively pure fat or oil, but we found the reaction worked equally well on MBM. We then used a hamster scrapie model of BSE, with two different methods to test whether the biodiesel reaction also destroyed infectivity in the MBM. When we used antibody-based detection (Western blot) we were unable to detect the proteinase-resistant protein marker for disease in the biodiesel or solid residue. This result agrees with published reports implying the method produces safe biodiesel. When we used a bioassay (intracranial inoculation) we found that the biodiesel method destroyed all infectivity in the biodiesel and 99.9999% of the infectious material in the solid residue. This suggests the biodiesel fraction is safe for use as fuel. However, this level of decontamination/disinfection is unfortunately not sufficient to allow use of the solid residue as a feed supplement for cows and sheep. We think minor changes to the biodiesel recipe, such as increased time and temperature, could make it strong enough to completely decontaminate MBM.

Technical Abstract: The epidemic of bovine spongiform encephalopathy (BSE) has led to world-wide drop in the market for beef by-products, such as Meat-and-Bone Meal (MBM), a fat-containing product traditionally used as an animal feed supplement. While normal rendering is insufficient, the production of biodiesel from MBM has been suggested to destroy infectivity from transmissible spongiform encephalopathies (TSEs). In addition to producing fuel, this method simultaneously generates a nutritious solid residue. In our study we produced biodiesel from MBM under defined conditions using a modified form of alkaline methanolysis. We evaluated the presence of prion in the three resulting phases of the biodiesel reaction (Biodiesel, Glycerol and Solid Residue) in vitro and in vivo. Analysis of the reaction products from 263K scrapie infected MBM led to no detectable immunoreactivity by Western Blot. Importantly, and in contrast to the biochemical results the solid MBM residue from the reaction retained infectivity when tested in an animal bioassay. Histochemical analysis of hamster brains inoculated with the solid residue showed typical spongiform degeneration and vacuolization. Re-inoculation of these brains into a new cohort of hamsters led to onset of clinical scrapie symptoms within 75 days, suggesting that the intrinsic properties of the prion protein were not changed during the biodiesel process. Results from our study demonstrate biochemical detection by Western Blot is insufficient to conclude an absence of TSE infectivity. Furthermore the biodiesel reaction can not be considered a viable prion decontamination method despite increased survival time of hamsters and reduced infectivity greater than 6 log orders in the solid MBM residue.