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Volume 31, Number 5—May 2025
Dispatch

Administration of L-type Bovine Spongiform Encephalopathy to Macaques to Evaluate Zoonotic Potential

Morikazu Imamura1Comments to Author , Ken’ichi Hagiwara, Minoru Tobiume, Minako Ohno, Hiromi Iguchi, Hanae Takatsuki, Tsuyoshi Mori, Ryuichiro Atarashi, Hiroaki Shibata, and Fumiko Ono1
Author affiliation: University of Miyazaki, Miyazaki, Japan (M. Imamura, M. Ohno, H. Iguchi, H. Takatsuki, T. Mori, R. Atarashi); National Institute of Infectious Diseases, Tokyo, Japan (K. Hagiwara, M. Tobiume); The Corporation for Production and Research of Laboratory Primates, Tsukuba, Japan (H. Shibata); Okayama University of Science, Imabari, Japan (F. Ono)

Main Article

Figure 1

Sensitivity of modified protein misfolding cyclic amplification (PMCA) to detect abnormal prion protein in study of oral transmission of L-type bovine spongiform encephalopathy (L-BSE) in intracerebrally inoculated macaques to evaluate zoonotic potential. We evaluated in macaques intracerebrally inoculated with L-BSE prions (A) and cattle intracerebrally inoculated with classical BSE prions (B) (Appendix). We serially diluted (10−3–10−11) brain homogenates (10% weight by volume) in 50 μL of human normal prion protein (PrPC) substrate and performed PMCA. We further diluted the initial PMCA product to 1:5 with a fresh PrPC substrate for subsequent rounds. We conducted 6 rounds of PMCA in duplicate. In macaques, PMCA propagated PrPSc-like proteinase K–resistant prion protein (PrPres) from a 10−7 dilution in the fifth amplification round; in cattle, PMCA propagated PrPres from a 10−8 dilution during the second amplification round. We performed Western blot for each PMCA product (2.5 μL) after proteinase K digestion using the T-2 antibody (10). NS, nonseeded control; ND, assays not done; R, round.

Figure 1. Sensitivity of modified protein misfolding cyclic amplification (PMCA) to detect abnormal prion protein in study of oral transmission of L-type bovine spongiform encephalopathy (L-BSE) in intracerebrally inoculated macaques to evaluate zoonotic potential. We evaluated in macaques intracerebrally inoculated with L-BSE prions (A) and cattle intracerebrally inoculated with classical BSE prions (B) (Appendix). We serially diluted (10−3–10−11) brain homogenates (10% weight by volume) in 50 μL of human normal prion protein (PrPC) substrate and performed PMCA. We further diluted the initial PMCA product to 1:5 with a fresh PrPC substrate for subsequent rounds. We conducted 6 rounds of PMCA in duplicate. In macaques, PMCA propagated PrPSc-like proteinase K–resistant prion protein (PrPres) from a 10−7 dilution in the fifth amplification round; in cattle, PMCA propagated PrPres from a 10−8 dilution during the second amplification round. We performed Western blot for each PMCA product (2.5 μL) after proteinase K digestion using the T-2 antibody (10). NS, nonseeded control; ND, assays not done; R, round.

Main Article

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1These authors were co–principal investigators.

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