3R-Project 116-09

Organotypic brain slice cultures derived from regularly slaughtered animals as in vitro alternative for the investigation of neuroinfectious diseases in ruminants

Anna Oevermann and Torsten Seuberlich
Neurocenter, Department of Clinical Research and Veterinary Public Health, Vetsuisse Faculty Bern, 3001 Bern, Switzerland
anna.oevermann@itn.unibe.ch, torsten.seuberlich@itn.unibe.ch

Keywords: live stock; brain; cns, brain disorders; veterinary disease; cell cultures: organ-specific; slices; reduction; replacement

Duration: 3 years Project Completion: 2013

Background and Aim
Infectious disorders of the central nervous system in livestock may have severe economic and public health implications and are therefore of major concern. This was demonstrated dramatically in the mid 1990s, when it became evident during the upsurge of bovine spongiform encephalopathy (BSE) that the disease was transmissible from cattle to humans. Recently, atypical variants of transmissible spongiform encephalopathies (TSEs) have been detected in ruminants, whose potential to cross over to other species including humans is not known at present. Listeriosis, caused by Listeria monocytogenes (LM), is another infectious and zoonotic CNS disease of high impact on livestock and humans.
Despite intense research activities in the field of TSEs and listeriosis during the past decades, very few in-vitro models for their neuropathogenesis, host-pathogen interactions and strain-typing exist. Studies largely depend on bioassays either in laboratory rodents or in the natural ruminant host because they reflect best the intricate pathogenesis of CNS infections. Such experiments raise fundamental ethical concerns, considering the highly invasive inoculation routes and the resulting severe disease, because of which most of these experiments are classified into the highest severity degree (Schweregrad 3). In the case of TSEs, experiments may last up to several years until the animals are sacrificed at the end. In addition, is often not clear to what extent the results of rodent models can be extrapolated to the natural hosts’ situation. On the other hand, pertinent cell models of ruminant neuroinfectious diseases do not exist. Our aim is, therefore, to develop an ethically sustainable, host specific, organotypic brain slice culture of regularly slaughtered ruminants as an in-vitro system for the identification and investigation of neuroinfectious diseases using the examples of prion diseases and listeric encephalitis.

Method and Results
in progress (present status)
We will develop organotypic brain slice cultures from young ruminants (up to 6 months of age), which are slaughtered for human consumption. Organotypic slice explants of various brain areas will be cultured and the best suitable candidate brain regions for long-term inoculation studies selected. Once we are able to maintain brain slices from slaughtered ruminants, we will validate the system for the investigation of prion diseases and listeriosis.

Conclusions and Relevance for 3R
Organotypic brain slice cultures from regularly slaughtered ruminants as a host-specific in-vitro system will not only help to considerably reduce the number of animals that are sacrificed for the study of known neuroinfectious diseases. They might also be used for the cultivation and identification of yet unidentified agents and for the production of positive reference material for diagnostic purposes. Hence, this system will allow a wide range of in vitro studies in the natural host, offering the advantage of harvesting data that are not obtainable from in-vivo studies in rodents.

Figures

Figure 1: Three organotypic hippocampal slices of a calf in culture