Neuro‐invasion by a ‘Trojan Horse’ strategy and vasculopathy during intrauterine flavivirus infection

Abstract

The central nervous system (CNS) is a major target of several important human and animal viral pathogens causing congenital infections. However, despite the importance of neuropathological outcomes, for humans in particular, the pathogenesis, including mode of neuro-invasion, remains unresolved for most congenital virus infections. Using a natural model of congenital infection with an RNA virus, bovine viral diarrhoea virus in pregnant cattle, we sought to delineate the timing and mode of virus neuro-invasion of and spread within the brain of foetuses following experimental respiratory tract infection of the dams at day 75 of pregnancy, a time of maximal risk of tissue pathology without foetal death. Virus antigen was first detected in the foetal brains 14 days postinfection of dams and was initially restricted to amoeboid microglial cells in the periventricular germinal layer. The appearance of these cells was preceded by or concurrent with vasculopathy in the same region. While the affected microvessels were negative for virus antigen, they expressed high levels of the type I interferon-stimulated protein ISG15 and eventually disappeared in parallel with the appearance of microcavitary lesions. Subsequently, the virus spread to neurons and other glial cells. Our findings suggest that the virus enters the CNS via infected microglial precursors, the amoeboid microglial cells, in a 'Trojan horse' mode of invasion and that the microcavitary lesions are associated with loss of periventricular microvasculature, perhaps as a consequence of high, unrestricted induction of interferon-regulated proteins.

Bovine Viral Diarrhea Virus Cyclically Impairs Long Bone Trabecular Modeling in Experimental Persistently Infected Fetuses

“…The subphyseal and proximal zones were not analyzed for dynamic parameters due to the complete lack of labeling and predominance of diffuse labeling, respectively. The definition of the parameters evaluated, their corresponding referents, standardized nomenclature, units of measurement, and criteria for evaluation, where applicable, are listed as follows: bone volume-trabecular bone volume including calcified cartilage cores normalized to tissue volume (BV/TV, %); cartilage volume-longitudinal calcified cartilage core volume normalized to bone volume (Cg.V/BV, %); total bone surface (BS, mm 2 ); number of osteoclasts (TRAP positive) per mm 2 bone surface (N.Oc/BS, #/mm 2 ); osteoclast surface-TRAP-positive osteoclast-covered bone surface normalized to bone surface (Oc.S/BS, %); single-labeled surface-area of bone surface with a single flurochrome label normalized to bone surface (s.LS/BS, %); double-labeled surface-area of bone surface with 2 labels normalized to bone surface (d.LS/BS, %); mineralizing surface-equal to half the area of single-labeled surface plus the area of double-labeled surface normalized to bone surface (MS/BS, %); mineral apposition rate-average distance between first and second label divided by the number of days in the interlabel period (MAR, mm/day); and bone formation rate-equal to the mineralizing surface normalized to bone surface multiplied by the mineral apposition rate (MS/BS * MAR, mm 3 /mm 2 /day). Femoral middiaphyseal apposition rates were determined by averaging the width of nonlabeled subperiosteal plexiform bone at 5 standardized sites and dividing this distance by the period (7 days) between administration of the second label and collection.…”

End-point disease investigation for virus strains of intermediate virulence as illustrated by flavivirus infections

“…Rabbit anti-GFAP (Dako) and goat anti-Iba1 (Abcam) antibodies were used to assess astrocytic and microglial activation in brain and spinal cord sections, using a previously described protocol (Miura et al, 2008). The degree of viral antigen load and glial cell activation was scored semiquantitatively as described previously (Bielefeldt-Ohmann et al, 2012;Tolnay et al, 2010) by an observer (H. B.-O.)…”

Investigation of neuropathogenesis and control of an equine-pathogenic Australian West Nile virus isolate in an established CD1 Swiss white mouse and a novel rabbit model

“…Furthermore, the function and trafficking behaviour of WNV infected leucocytes have not been examined, as noted by King et al [71] and Bielefeldt-Ohmann et al [72].…”

“…The degree of viral antigen load and glial cell activation was scored semi-quantitatively as previously described [72,256] …”

“…Studies into viral pathogenesis of bovine viral diarrhoea virus (BVDV ) [72] and Nipah virus [347] have suggested that viruses can either be trafficked inside or on the surface of leucocytes in the blood with or without productive replication. Therefore, the serum/plasma viraemia levels in Chapter 5 may not represent the true amount of virus in the blood, if WNV/MVEV were cellassociated.…”