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Comparative titration of infectivity of BSE in sheep and mice - SE1842

Bioassay of scrapie or related disease infection is necessary because there is no other method for conclusively demonstrating the infectious pathogen, nor for quantification of the infectivity.

The usual predictability of the incubation period in rodent models of scrapie, when the relevant variables are controlled, has provided highly reproducible methods for the bioassay of infectivity of scrapie-like agents. Standard conditions include the intracerebral route of inoculation. Transmission to mice, the species most extensively used for scrapie bioassay, is, however, not always successful from natural cases of sheep scrapie. Such failures are not attributable to avoidable technical defects. They involve high concentrations of inocula and have been reported to occur in 19% of attempted primary transmissions. However, it is known that demonstrable infectivity varies according to brain region sampled and that the efficiency of infection at primary interspecific transmissions of scrapie-like pathogens is reduced (the species barrier effect).

Measurement of infectivity (infectivity titre) of scrapie-like diseases is made mostly through intracerebral end-point titrations. The assumption that the dose-response kinetics of scrapie infectivity in different organs of the same host are the same, has been challenged.Differences in the dose-response relationships between infectivity in hamster spleen and brain were interpreted as indicating qualitative differences in the pathogen or the pathogenic mechanisms operating in these different organs.

In mouse models of scrapie after infection by non-neural peripheral routes and in natural sheep scrapie,studies indicate that replication occurs first in lymphoreticular tissues, fatal disease developing only if infection spreads to the central nervous system.

At the clinical stage in mouse models of scrapie, infectivity titres in the brain are about 10 times greater than anywhere in the lymphoreticular system. Similarly, in sheep and goats affected with natural scrapie, infectivity titres in neural tissues were found to be greater than in non-neural tissues by a factor of 10^10.Parallel titration of BSE tissues obtained from cattle have been carried out in mice and calves (SE1821) and also indicate that results obtained for cattle are considerably more sensitive.

BSE brain homogenate has been transmitted, passaged and titrated in mice.Titrations of BSE brain infectivity in mice have used intracerebral (i.c.) or intraperitoneal (i.p.) routes. Results of initial titration studies of BSE-affected brain in mice by the i.c. route show at least 105 i.C. ID50 mouse infectious units per gram of terminally affected cattle brain. Results of studies by the i.p. route are incomplete, but end-point titrations so far approximate to 10^3 ID50 mouse infectious units per gram of brain.These results indicate that, when titrated across a species barrier in mice, the i.p. route is less efficient than the i.c. route for detection of infectivity and therefore when, as previously, simultaneous i.c. and i.p. routes have been employed, the i.p. component of the inoculation does not contribute significantly to the estimate of titre. In cattle, where there is no species barrier effect for BSE, the efficiency of i.c. inoculation cannot be in doubt.

The results obtained for mouse bioassay of tissues from cattle naturally infected with BSE have failed to demonstrate infection in non neural tissues, although transmission of CNS tissues have been 100% successful. Mouse bioassay results for cattle experimentally challenged with BSE (SE1901) have shown limited infectivity in distal illeum, dorsal root ganglia, trigeminal ganglia and bone marrow.

There are, therefore, a variety of possible reasons or precedents for inefficiency of detection of scrapie infectivity by bioassay in mice, particularly on primary transmission. There are no data on scrapie or BSE infectivity titrations in sheep or goats for, although this approach would theoretically avoid the problems related to interspecies transmission, such experiments have previously been considered prohibitive in cost and difficult to control.

Recently, public health concerns have been expressed regarding the sensitivity of tests to determine the levels of infectivity in cattle BSE infected tissues potentially entering the human food chain and in offal. It is also possible that sheep may have been exposed to the BSE agent through contaminated meat and bonemeal. In order to aid any future assessment of the potential exposure of the human population to BSE of sheep origin it is necessary to determine infectivity titres of BSE infected sheep. Such estimates may be important for predictions regarding nvCJD. Aquisition of such data is important and justifies the use of sheep for titration purposes.

The proposed study has three main aims:

(1)To obtain a measure of the underestimation of the infectivity titre of sheep BSE tissues when titrated across a species barrier in mice.

(2)To produce an approximate dose/incubation curve for infectivity of brain
from clinically BSE-affected sheep by simultaneous titration of a primary
inoculum in sheep and mice.

(3)To compare the infectivity curves obtained for ovine BSE in CNS and spleen
of RI 11 mice.
Project Documents
• Final Report : Comparative titration of infectivity of BSE in sheep and mice   (383k)
Time-Scale and Cost
From: 2000

To: 2007

Cost: £1,292,496
Contractor / Funded Organisations
Moredun Research Institute
Animal Health              
Plants and Animals              
Fields of Study
Animal Health