Evidence for degradation of abnormal prion protein in tissues from sheep with scrapie during composting

Abstract

This study investigated whether the abnormal prion protein (PrP(Sc)) in tissues from sheep with scrapie would be destroyed by composting. Tissues from sheep naturally infected with scrapie were placed within fiberglass mesh bags and buried in compost piles for 108 d in experiment 1 or 148 d in experiment 2. The temperature in the compost piles rose quickly; it was above 60 degrees C for about 2 wk and then slowly declined to the ambient temperature. Before composting, PrPSc was detected in all the tissues by Western blotting. In experiment 1, PrPsc was not detected after composting in the tissue remnants or the surrounding sawdust. In experiment 2, 1 of 5 specimens tested negative after composting, whereas PrP(Sc) was detected in the other 4 bags, though in reduced amounts compared with those before composting. Tissue weights were reduced during composting. Analysis of the tissue remnants for microbial 16S ribosomal DNA demonstrated that there were more diverse microbes involved in experiment 1 than in experiment 2 and that the guanine and cytosine content of the microbial 16S DNA was higher in the specimens of experiment 1 than in those of experiment 2, which suggests greater dominance of thermophilic microbes in experiment 1. These results indicate that composting may have value as a means for degrading PrP(Sc) in carcasses and other wastes.

Figure 1

Diagram of compost-pile structure for experiment 1; the structure for experiment 2 was similar. The walls and floor of the bin were bales of hay, and the interior was lined with heavy plastic. A perforated, flexible plastic drainage pipe, 10 cm in diameter, was laid towards the bottom of the pile to provide aeration. The pile was covered with vapor-barrier fabric. The fiberglass bags containing the specimens were placed 20 to 40 cm below the surface of the pile, which was a mixture of cow manure and straw. Two thermocouple probes were located close to the specimens to enable temperature monitoring by a computerized system.

Figure 2

Temperature 0 to 80 cm from the bottom of the compost pile in experiment 1; the pattern in experiment 2 was similar. RT — room temperature inside the facility.

Figure 3

Detection of thermophilic microbial DNA in specimens from bags 1 to 5 of sheep 3 in experiment 2 (lanes 1 to 5) and from bags 1 to 4 of sheep 1 in experiment 1 (lanes 6 to 9). Arrows indicate individual DNA bands.

Figure 4

Detection of abnormal prion protein (PrP^Sc) in animals with scrapie by a sensitive Western blot procedure involving enrichment by precipitation with sodium phosphotungstic acid (PTA) before and after composting. Lanes 1 to 3 are the results for bag 3 from sheep 1 in experiment 1: PrP^Sc was detected on day 0 (lane 1) but not after composting in tissue remnants (lane 2) or immediately surrounding sawdust (lane 3). The results for the other bags in experiment 1 were similar. Lanes 4 to 7 are the results for bag 5 of experiment 2: PrP^Sc was detected on day 0 in both the PTA-enriched sample, which had an excessive amount of PrP^Sc loaded onto the gel (lane 4), and the crude extract (lane 5) but not after composting in tissue remnants (lane 6) or immediately surrounding sawdust (lane 7), both PTA-enriched. Lanes 8 to 10 are the results for bag 2 of experiment 2: PrP^Sc was detected on day 0 in both the PTA-enriched sample (lane 8) and the crude extract (lane 9), as well as after composting in PTA-enriched tissue remnants (lane 10).