H-Type Bovine Spongiform Encephalopathy

Figures & data

Figure 1 Molecular discrimination of H-type and typical BSE in cattle and in C57Bl/6 infected mice. Western blot detection of PrP^res in cattle (A–C) or C57Bl/6 infected mice (E–G), from H-type isolate (right lane in each panel) or typical BSE (left lane in each panel), using 12B2 (A and E), Sha31 (B and F) or SAF84 (C and G) monoclonal antibodies. In each Western blot, typical BSE is shown in the left lane and H-type BSE in the right lane. Brain equivalent quantities loaded per lane were 100 µg (left lanes) and 250 µg (right lanes) for cattle samples and 1.2 mg (left lanes) and 4.8 mg (right lanes) for mouse samples. Inset of (A) shows the N-terminal PrP^res fragment (≈7 kDa) in cattle (left lane) and mouse (right lane). (D) Interpretation of the Western blot profile observed with SAF84 monoclonal antibody. (H) Unspecific immunoreactivity of anti-mouse Ig conjugate on samples prepared from normal brain of C57Bl/6 mouse.

Figure 2 Molecular analysis of PrP^res after deglycosylation in C57Bl/6 infected mice. Western blot analysis of PrP^res from cattle (A–C) or C57Bl/6 mice (D–F) infected with H-type (lanes 3 and 4) or typical BSE (lanes 1 and 2), after deglycosylation by PNGase treatment. Monoclonal antibodies used for PrP^res detection were 12B2 (A and D), Sha31 (B and E) or SAF84 (C and F). Brain equivalent quantities loaded per lane were (i) from cattle, for typical BSE, 50 (lane 1) or 2,6 (lane 2) mg, and for H-type BSE, 50 (lane 3) or 6 (lane 4) mg, and (ii) from mice, for typical BSE, 0.5 (D and F) or 0.3 (E) mg, and for H-type BSE, 2.5 (D and F) or 1.3 mg (E).

Figure 3 Quantitative molecular features of PrP^res in C57Bl/6 infected mice. Molecular masses (A) and glycoform ratios (B) (means ± standard deviations) of PrP^res detected with Sha31 antibody, from C57Bl/6 mice infected with H-type or typical BSE isolates. PrP^res of mice infected with an experimental scrapie (C506M3) strain4 was used as a control.

Figure 4 Schematic representation of proteinase K-resistant PrP fragments identified from H-type or typical BSE. Putative regions for proteinase K cleavage are indicated, as assessed by immunoreactivities with anti-PrP antibodies, and categories of epitope locations (groups A, B and C).

Table 1 Antibodies used to characterize H-type and typical BSE

Antibodies	Sequences Recognized in the Bovine PrP Protein	Concentration	Ref.
Group A
SAF32	62 - QPHGGGW - 92	10.000 (ascitic fluid)	51
4F2	62 - QPHGGGW - 92	100 ng/ml	57
12B2	101 - WGQGG - 105	340 ng/mL	58
P4	101 - WGQGGSH - 107	100 ng/ml	52
Group B
9A2	110 - WNK -112	200 ng/ml	58
RB1	110 - WNKP - 113	1/2500 (rabbit antiserum)	53
F89/160.1.5	148 - PLIHFGSD - 155	200 ng/ml	54
Bar233	152 - FGSDYEDRYYRE - 163	1/10.000 (ascitic fluid)	51
L42	156 - YEDRYY - 161	200 ng/ml	58
Sha31	156 - YEDRYYRE - 163	Kit Biorad TeSeE sheep/goat 1/10	51
6H4	156 - YEDRYYREN - 164	100 ng/ml	55
Group C
12F10	154 - SDYEDRYYRE - 163	100 ng/ml	51
SAF84	175 - RPVDQY - 180	75 (mouse) or 150 (bovine) ng/ml	51
94B4	198 - HTVTTTTK - 205	70 ng/ml	24
F99/97.6.1	229 - YQRE - 232	540 ng/ml	54
R524	233 - SQAY - 236	1/500 (rabbit antiserum)	56

Baron TGM, Biacabe AG. Molecular analysis of the abnormal prion protein during coinfection of mice by bovine spongiform encephalopathy and a scrapie agent. J Virol 2001; 75:107 - 114

 PubMed Web of Science ®Google Scholar

Feraudet C, Morel N, Simon S, Volland H, Frobert C, Creminon D, Vilette S, Lehmann S, Grassi J. Screening of 145 anti-PrP monoclonal antibodies for their capacity to inhibit PrPSc replication in infected cells. J Biol Chem 2005; 280:11247 - 11258

 PubMed Web of Science ®Google Scholar

Thuring CM, Erkens JHF, Jacobs JG, Bossers A, van Keulen LJM, Garssen GJ, van Zijderveld FG, Ryder SJ, Groschup MH, Sweeney T, Langeveld JPM. Discrimination between scrapie and bovine spongiform encephalopathy in sheep by molecular size immunoreactivity and glycoprofile of prion protein. J Clin Microbiol 2004; 42:972 - 980

 PubMed Web of Science ®Google Scholar

Langeveld JPM, Jacobs JG, Erkens JHF, Bossers A, Van Zijderveld FG, Van Keulen LJM. Rapid and discriminatory diagnosis of scrapie and BSE in retro-pharyngeal lymph nodes of sheep. BMC Vet Res 2006; 2:19

 PubMedGoogle Scholar

Harmeyer S, Pfaff E, Groschup MH. Synthetic peptide vaccines yield monoclonal antibodies to cellular and pathological prion proteins of ruminants. J Gen Virol 1998; 79:937 - 945

 PubMed Web of Science ®Google Scholar

Madec JY, Groschup MH, Buschmann A, Belli P, Calavas D, Baron T. Sensitivity of the Western blot detection of prion protein PrPres in natural sheep scrapie. J Virol Methods 1998; 75:169 - 177

 PubMed Web of Science ®Google Scholar

O'Rourke KI, Baszler TV, Besser TE, Miller JM, Cutlip RC, Wells GAH, Ryder SJ, Parish SM, Hamir SM, Cockett NE, Jenny A, Knowles DP. Preclinical diagnosis of scrapie by immunohistochemistry of third eyelid lymphoid tissue. J Clin Microbiol 2000; 38:3254 - 3259

 PubMed Web of Science ®Google Scholar

Korth C, Stierli B, Streit P, Moser M, Schaller O, Fischer R, Schultz-Schaeffer W, Kretzschmar H, Raeber A, Braun U, Ehrensperger F, Hornemann S, Glockshuber R, Riek R, Billeter M, Wuthrich K, Oesch B. Prion (PrPsc)-specific epitope defined by a monoclonal antibody. Nature 1997; 390:74 - 77

 PubMed Web of Science ®Google Scholar

Thuring CMA, van Keulen LJM, Langeveld JPM, Vromans MEW, van Zijderveld FG, Sweeney T. Immunohistochemical distinction between preclinical bovine spongiform encephalopathy and scrapie infection in sheep. J Comp Pathol 2005; 132:59 - 69

 PubMed Web of Science ®Google Scholar

Garssen GJ, Van Keulen LJM, Farquhar CF, Smits MA, Jacobs JG, Bossers A, Meloen RH, Langeveld JM. Applicability of three anti-PrP peptide sera including staining of tonsils and brainstem of sheep with scrapie. Microsc Res Techniq 2000; 50:32 - 39

 PubMed Web of Science ®Google Scholar