Transmissible spongiform encephalopathies (TSEs) are fatal neurodegenerative diseases that affect humans and animals. The hallmark lesion of TSEs is spongiform changes in the brains of affected subjects. According to the prion hypothesis TSEs are caused by proteinaceous infectious particles consisting of abnormal isoforms (PrPSc) of the normal cellular prion protein (PrPC) [1–3] although this view is not accepted universally [4–6]. Despite the disagreement in the scientific community regarding the exact nature of the infectious agent, PrPSc is considered to be a ubiquitous marker of TSE disease and it accumulates in the brains of affected subjects from the very early stages of the disease before any spongiform changes become evident [7, 8]. Therefore, techniques such as immunohistochemistry (IHC) are widely used to detect PrPSc as a marker of disease and diagnose experimental  or naturally occurring [9, 10] TSEs.
Scrapie, the archetypal TSE which affects small ruminants, is not considered to be a public health risk and is known for centuries [11–13]. In contrast, bovine spongiform encephalopathy (BSE) is a TSE of cattle first recognised in 1986 . After its discovery, novel spongiform encephalopathies in captive ruminants  and cats [16–18] and a variant of Creutzfeldt-Jakob disease (vCJD) in humans [19–21] were identified. Subsequently a link between BSE and these new emerging TSEs was established and wild type mouse bioassay contributed in determining this association .
Wild type mouse bioassay has been used to study the infectivity of TSE sources and to identify different strains of the agent [23–25]. According to established methodology, strains are identified after serial passage in a given mouse line to allow a TSE agent to adapt into the new host by overcoming the low attack rates, and prolonged and variable incubation periods which are experienced during primary isolation of the agent into mice, a property called the species or transmission barrier. For strain typing C57BL/6, RIII and VM are the most commonly used wild type mouse lines either alone or in combination .
The amino acid sequence of PrP in RIII and C57BL/6 mice (Prnp
) differs from that of VM mice (Prnp
) at codons 108 and 189 (Prnp
– leucine, threonine; Prnp
– phenylalanine, valine) [27, 28]. This difference in Prnp genotype affects both bioassay parameters which are traditionally used to study TSEs in mice: the incubation period and the lesion profile – a semi-quantitative assessment of vacuolation at specific neuroanatomical areas [23, 24, 29]. However, there are some differences in the data observed between RIII and C57BL/6 mice challenged with the same TSE source or strain attributed to genetic factors other than the PrP genotype [30, 31]. Also in RIII mice, even upon primary isolation, BSE and BSE derived TSEs behave reproducibly with consistent, albeit prolonged compared to serial passage, incubation period and exhibit a single characteristic lesion profile [22, 32–34]. In contrast to RIII mice, primary isolation of BSE in C57BL/6 and VM mice does not yield informative data according to traditional parameters. However, as for classical scrapie, the agent can be fully characterised in these mouse lines upon serial passage. Incubation period and lesion profile data obtained from the serial passage of BSE through C57BL/6 and VM mice indicate that there are two stable mouse adapted strains of BSE: 301C which is derived after serial passage through C57BL/6 mice and 301V which results after serial passage through VM mice .
The association of vCJD with BSE [22, 36, 37], the successful experimental transmission of BSE to sheep [38, 39] and the first report of BSE in a goat , raised the need to be able to confidently distinguish between BSE and scrapie, and applying traditional methods on RIII bioassays is regarded as one of the most reliable approaches to do so [22, 32]. There are some limitations, however, as it has been demonstrated by recent studies, according to which scrapie-infected sheep can produce a similar incubation period and lesion profile to BSE upon primary isolation in RIII mice [34, 41]. It was still possible, however, to confirm these cases as scrapie using histopathology, Western blot and IHC although detailed description of the PrPSc types in the mouse brain was not attempted. Other studies also indicate that an assessment of the distribution of PrPSc in the brain could aid discrimination of TSE source [42, 43]. These data, although limited, suggest that assessment of PrPSc distribution could provide an alternative approach to lesion profiling in distinguishing BSE from non-BSE related TSEs upon primary passage and the potential to diagnose BSE based on individual mice rather than groups of animals makes this method very powerful as a discriminatory tool. This approach has recently been employed to help confirm BSE in a goat in UK where incubation period and lesion profile data on primary isolation were unavailable .
IHC is used routinely as a diagnostic tool in the study of TSEs in large animals. Furthermore, IHC has been used to characterise TSE sources within sheep by the identification of different PrPSc types and the spatial distribution of each type in the brain to give specific PrPSc distribution patterns [10, 45–47]. It has been proposed that these patterns may be dependent on agent strain, host genetic parameters (the most profound of the host factors being its PrP sequence) or a combination of both . In contrast, there is limited data available regarding the systematic observation of PrPSc patterns and their neuroanatomical distribution upon primary isolation in murine TSE studies. The majority of studies involve mouse-adapted scrapie strains, such as 87V or ME7, and a systematic recording of the different PrPSc types in the murine brain was not attempted [48–50].
In the current study we systematically studied and recorded in detail the distribution of PrPSc throughout the brain of wild type mice challenged with BSE from bovine and ovine sources during primary isolation or after serial passages using IHC and PET blot. We were able to identify source- and strain-specific markers, in addition to mouse lineage-specific markers which could enhance our ability to discriminate between TSE sources and help characterise strains on primary isolation.