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Atypical/Nor98 scrapie infectivity in sheep peripheral tissues.

Andréoletti O, Orge L, Benestad SL, Beringue V, Litaise C, Simon S, Le Dur A, Laude H, Simmons H, Lugan S, Corbière F, Costes P, Morel N, Schelcher F, Lacroux C - PLoS Pathog. (2011)

Bottom Line: It is now considered as a worldwide disease of small ruminants and currently represents a significant part of the detected transmissible spongiform encephalopathies (TSE) cases in Europe.However, based on the absence of detectable abnormal PrP in peripheral tissues of affected individuals, human and animal exposure risk to this specific TSE agent has been considered low.The results obtained indicate that dietary exposure risk to small ruminants TSE agents may be higher than commonly believed.

View Article: PubMed Central - PubMed

Affiliation: UMR INRA ENVT 1225, Interactions Hôtes Agents Pathogènes, Ecole Nationale Vétérinaire de Toulouse, Toulouse, France. o.andreoletti@envt.fr

ABSTRACT
Atypical/Nor98 scrapie was first identified in 1998 in Norway. It is now considered as a worldwide disease of small ruminants and currently represents a significant part of the detected transmissible spongiform encephalopathies (TSE) cases in Europe. Atypical/Nor98 scrapie cases were reported in ARR/ARR sheep, which are highly resistant to BSE and other small ruminants TSE agents. The biology and pathogenesis of the Atypical/Nor98 scrapie agent in its natural host is still poorly understood. However, based on the absence of detectable abnormal PrP in peripheral tissues of affected individuals, human and animal exposure risk to this specific TSE agent has been considered low. In this study we demonstrate that infectivity can accumulate, even if no abnormal PrP is detectable, in lymphoid tissues, nerves, and muscles from natural and/or experimental Atypical/Nor98 scrapie cases. Evidence is provided that, in comparison to other TSE agents, samples containing Atypical/Nor98 scrapie infectivity could remain PrP(Sc) negative. This feature will impact detection of Atypical/Nor98 scrapie cases in the field, and highlights the need to review current evaluations of the disease prevalence and potential transmissibility. Finally, an estimate is made of the infectivity loads accumulating in peripheral tissues in both Atypical/Nor98 and classical scrapie cases that currently enter the food chain. The results obtained indicate that dietary exposure risk to small ruminants TSE agents may be higher than commonly believed.

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Infectivity titre in reference brain sample from classical and Atypical/Nor98 affected ewes.(A–B) Intra-cerebral endpoint titration in a tg338 mouse model of CNS homogenate (20 µl per mice), (A) prepared from terminally scrapie affected sheep with Langlade isolate (12.5% homogenate - case 10 - ○) and with PG127 isolate (10% homogenate- case 13- △) – (B) an ARQ/ARQ (10% homogenate - case 14- ○) and an ARR/ARR (10% homogenate -case 15- △) atypical scrapie cases identified in field through active epidemiosurveillance program. Such titration allowed the determination of the infectious dose 50 (ID50) of the reference brain samples (Table 3). (D–E) Variation of the incubation period as a function of the infectious dose inoculated intra-cerebrally in tg338 mice. (C): Langlade isolate, (D): PG127 isolate and (E): atypical scrapie isolate. To establish these diagrams, the individual incubation periods and the number of ID50 inoculated in each mouse were plotted. The number of ID50 is obtained by multiplying the titre of the inoculum (ID50 IC in tg338 per gram of tissue) per the amount of tissue inoculated in each mice. On this basis, a four parameter logistic regression was computed (Sigma Plot) to provide a curve associating the incubation period and the number of ID50 inoculated with the observed incubation period.
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ppat-1001285-g004: Infectivity titre in reference brain sample from classical and Atypical/Nor98 affected ewes.(A–B) Intra-cerebral endpoint titration in a tg338 mouse model of CNS homogenate (20 µl per mice), (A) prepared from terminally scrapie affected sheep with Langlade isolate (12.5% homogenate - case 10 - ○) and with PG127 isolate (10% homogenate- case 13- △) – (B) an ARQ/ARQ (10% homogenate - case 14- ○) and an ARR/ARR (10% homogenate -case 15- △) atypical scrapie cases identified in field through active epidemiosurveillance program. Such titration allowed the determination of the infectious dose 50 (ID50) of the reference brain samples (Table 3). (D–E) Variation of the incubation period as a function of the infectious dose inoculated intra-cerebrally in tg338 mice. (C): Langlade isolate, (D): PG127 isolate and (E): atypical scrapie isolate. To establish these diagrams, the individual incubation periods and the number of ID50 inoculated in each mouse were plotted. The number of ID50 is obtained by multiplying the titre of the inoculum (ID50 IC in tg338 per gram of tissue) per the amount of tissue inoculated in each mice. On this basis, a four parameter logistic regression was computed (Sigma Plot) to provide a curve associating the incubation period and the number of ID50 inoculated with the observed incubation period.

Mentions: Two different methods were applied to detect PrPSc presence in tissue homogenate that were used for mice inoculation: WB (TeSeE WB kit – BioRad, using SHa31 as anti-PrP antibody) and ELISA (TeSeE Sheep and Goat– BioRad). Additionally when formalin fixed tissue was available, PrPSc immunohistochemistry was also carried out (using 8G8 antibody). (++) indicate a case were minimal PrPSc labelling was observed after the exam of a serial series of slides. In some cases (+), the tissue homogenates were heated (60°C −10 min) in order to destroy contaminating bacteria before bioassay. Such heat treatment might have reduced infectivity level in these samples in an unknown proportion. Mice were considered positive when abnormal PrP deposition was detected in brain. Incubation periods are presented as mean +/−SD (in days) except for that dilution with which less than 50% of mice were found positive. In that case (*) incubation periods of the positive mice are individually presented. In the different tissues, infectious titres were estimated on the basis of incubation period in mice (Figure 4). This estimation was only performed when the attack rate was 100% and data from more than 5 animals were available.


Atypical/Nor98 scrapie infectivity in sheep peripheral tissues.

Andréoletti O, Orge L, Benestad SL, Beringue V, Litaise C, Simon S, Le Dur A, Laude H, Simmons H, Lugan S, Corbière F, Costes P, Morel N, Schelcher F, Lacroux C - PLoS Pathog. (2011)

Infectivity titre in reference brain sample from classical and Atypical/Nor98 affected ewes.(A–B) Intra-cerebral endpoint titration in a tg338 mouse model of CNS homogenate (20 µl per mice), (A) prepared from terminally scrapie affected sheep with Langlade isolate (12.5% homogenate - case 10 - ○) and with PG127 isolate (10% homogenate- case 13- △) – (B) an ARQ/ARQ (10% homogenate - case 14- ○) and an ARR/ARR (10% homogenate -case 15- △) atypical scrapie cases identified in field through active epidemiosurveillance program. Such titration allowed the determination of the infectious dose 50 (ID50) of the reference brain samples (Table 3). (D–E) Variation of the incubation period as a function of the infectious dose inoculated intra-cerebrally in tg338 mice. (C): Langlade isolate, (D): PG127 isolate and (E): atypical scrapie isolate. To establish these diagrams, the individual incubation periods and the number of ID50 inoculated in each mouse were plotted. The number of ID50 is obtained by multiplying the titre of the inoculum (ID50 IC in tg338 per gram of tissue) per the amount of tissue inoculated in each mice. On this basis, a four parameter logistic regression was computed (Sigma Plot) to provide a curve associating the incubation period and the number of ID50 inoculated with the observed incubation period.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3037359&req=5

ppat-1001285-g004: Infectivity titre in reference brain sample from classical and Atypical/Nor98 affected ewes.(A–B) Intra-cerebral endpoint titration in a tg338 mouse model of CNS homogenate (20 µl per mice), (A) prepared from terminally scrapie affected sheep with Langlade isolate (12.5% homogenate - case 10 - ○) and with PG127 isolate (10% homogenate- case 13- △) – (B) an ARQ/ARQ (10% homogenate - case 14- ○) and an ARR/ARR (10% homogenate -case 15- △) atypical scrapie cases identified in field through active epidemiosurveillance program. Such titration allowed the determination of the infectious dose 50 (ID50) of the reference brain samples (Table 3). (D–E) Variation of the incubation period as a function of the infectious dose inoculated intra-cerebrally in tg338 mice. (C): Langlade isolate, (D): PG127 isolate and (E): atypical scrapie isolate. To establish these diagrams, the individual incubation periods and the number of ID50 inoculated in each mouse were plotted. The number of ID50 is obtained by multiplying the titre of the inoculum (ID50 IC in tg338 per gram of tissue) per the amount of tissue inoculated in each mice. On this basis, a four parameter logistic regression was computed (Sigma Plot) to provide a curve associating the incubation period and the number of ID50 inoculated with the observed incubation period.
Mentions: Two different methods were applied to detect PrPSc presence in tissue homogenate that were used for mice inoculation: WB (TeSeE WB kit – BioRad, using SHa31 as anti-PrP antibody) and ELISA (TeSeE Sheep and Goat– BioRad). Additionally when formalin fixed tissue was available, PrPSc immunohistochemistry was also carried out (using 8G8 antibody). (++) indicate a case were minimal PrPSc labelling was observed after the exam of a serial series of slides. In some cases (+), the tissue homogenates were heated (60°C −10 min) in order to destroy contaminating bacteria before bioassay. Such heat treatment might have reduced infectivity level in these samples in an unknown proportion. Mice were considered positive when abnormal PrP deposition was detected in brain. Incubation periods are presented as mean +/−SD (in days) except for that dilution with which less than 50% of mice were found positive. In that case (*) incubation periods of the positive mice are individually presented. In the different tissues, infectious titres were estimated on the basis of incubation period in mice (Figure 4). This estimation was only performed when the attack rate was 100% and data from more than 5 animals were available.

Bottom Line: It is now considered as a worldwide disease of small ruminants and currently represents a significant part of the detected transmissible spongiform encephalopathies (TSE) cases in Europe.However, based on the absence of detectable abnormal PrP in peripheral tissues of affected individuals, human and animal exposure risk to this specific TSE agent has been considered low.The results obtained indicate that dietary exposure risk to small ruminants TSE agents may be higher than commonly believed.

View Article: PubMed Central - PubMed

Affiliation: UMR INRA ENVT 1225, Interactions Hôtes Agents Pathogènes, Ecole Nationale Vétérinaire de Toulouse, Toulouse, France. o.andreoletti@envt.fr

ABSTRACT
Atypical/Nor98 scrapie was first identified in 1998 in Norway. It is now considered as a worldwide disease of small ruminants and currently represents a significant part of the detected transmissible spongiform encephalopathies (TSE) cases in Europe. Atypical/Nor98 scrapie cases were reported in ARR/ARR sheep, which are highly resistant to BSE and other small ruminants TSE agents. The biology and pathogenesis of the Atypical/Nor98 scrapie agent in its natural host is still poorly understood. However, based on the absence of detectable abnormal PrP in peripheral tissues of affected individuals, human and animal exposure risk to this specific TSE agent has been considered low. In this study we demonstrate that infectivity can accumulate, even if no abnormal PrP is detectable, in lymphoid tissues, nerves, and muscles from natural and/or experimental Atypical/Nor98 scrapie cases. Evidence is provided that, in comparison to other TSE agents, samples containing Atypical/Nor98 scrapie infectivity could remain PrP(Sc) negative. This feature will impact detection of Atypical/Nor98 scrapie cases in the field, and highlights the need to review current evaluations of the disease prevalence and potential transmissibility. Finally, an estimate is made of the infectivity loads accumulating in peripheral tissues in both Atypical/Nor98 and classical scrapie cases that currently enter the food chain. The results obtained indicate that dietary exposure risk to small ruminants TSE agents may be higher than commonly believed.

Show MeSH
Related in: MedlinePlus