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

Suen, Willy (2017). 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 PhD Thesis, School of Veterinary Science, The University of Queensland. doi:10.14264/uql.2017.125

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Author Suen, Willy
Thesis Title 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
School, Centre or Institute School of Veterinary Science
Institution The University of Queensland
DOI 10.14264/uql.2017.125
Publication date 2017-01-13
Thesis type PhD Thesis
Supervisor Helle Bielefeldt-Ohmann
Natalie Prow
Roy Hall
Rachel Allavena
Total pages 246
Total colour pages 27
Total black and white pages 219
Language eng
Subjects 070709 Veterinary Pathology
060506 Virology
070705 Veterinary Immunology
Formatted abstract
West Nile virus (WNV) is a globally significant mosquito-borne neurotropic virus. In 2011, a
large-scale arboviral encephalitis outbreak occurred in Australia, involving close to 1000 horses. A
virulent WNV strain, WNVNSW2011, was isolated from the brain of an encephalitic horse during this
outbreak. Preliminary assessment showed that this strain is intermediate in virulence and belongs in
the same lineage as the highly virulent North American strains. However, the pathogenesis of
WNV in general, let alone this novel Australian strain, remains largely unclear.

The first part of the project aimed to investigate the pathogenesis of lethal WNVNSW2011 infection in
the established young adult Swiss white (CD1) mouse model, since previous survival challenge
experiments demonstrated high lethality (~70%) of WNVNSW2011 in this animal model. I conducted
a pilot time-course experiment, which demonstrated only benign pathological and virological
outcomes in sacrificed mice on day 3 and 7 post-infection (pi), despite the relatively high lethality
seen in previous survival trials. In a subsequent experiment, I characterised the disease phenotype
in moribund/dead and surviving mice after WNVNSW2011 infection. Notably, I detected a high
degree of intra-group variability in the neuropathology, neural infection and glial cell activation.
Similar variability was also observed in CD1 mice challenged with another Australian WNV strain
of intermediate virulence, WNVNSW2012. These inconsistencies in the end-point disease suggest that
random sacrifice of infected mice at any given time-point may not give reliable information of the
pathogenic processes towards lethal encephalitis. Therefore, conventional sacrifice experiments
may not be the best approach for investigating pathogenesis of virus strains of intermediate
virulence. This previously unacknowledged challenge to investigating pathogenesis of virus strains
of intermediate virulence should inform the planning and execution of future studies with such virus

The second part of the project addressed the need for a resistant small animal model for studying
the in vivo control of virulent WNV infections. Since rodent models typically produce lethal
disease after infection with virulent WNV strains, they may not reflect what happens in the natural
“dead-end” hosts, humans and horses, in whom most WNV infections (> 90%) are non-lethal.
Even cases with neuroinvasion are typically not fatal, unless underlying co-morbidities or
immunosuppressive disease exist.

I chose rabbits as a candidate for a novel animal model, due to the ease of performing intra-vital
sampling and monitoring, as well as the possibility for tissue sharing in different assays, given the
larger size of rabbits than mice. Rabbits are also hind-gut fermenters, and thus may be a better
representation of a horse than a mouse. New Zealand White (NZWRs; Oryctolagus cuniculus) and
North American cottontail rabbits (CTRs; Sylvilagus sp.) were challenged with WNV strains of
different virulence (WNVNSW2011 and WNVTX8667), and the prototype Murray Valley encephalitis
virus strain (MVE1-51), by the footpad route. The rabbits were consistently resistant to developing
severe disease after virulent WNV and MVEV infection, regardless of age, gender and species,
despite productive virus replication in the draining popliteal lymph node (PLN). The resultant
viraemia for all rabbits was also of low magnitude and transient. Furthermore, establishment of
virus infection in the brain was not a feature of the disease, despite mild to moderate
neuropathology in a subset of rabbits. This capacity to control flavivirus infection may be
explained by the rapid antiviral innate immune response detectable by day 3 pi, as well as a fast
anti-WNV neutralising antibody response detectable from day 7 pi.

Comparisons of the tissue-specific transcriptional profile of important cytokine and chemokine
genes suggested that virus control in rabbits was achieved differently depending on the virus, as
well as the rabbit species. However, common transcriptional upregulation of IFNγ in the draining
PLN, TNFα in the non-draining PLN and IL6 in the spleen was detected in all rabbits on day 3 pi,
suggesting a common tissue-specific innate immune response signature against flavivirus infection.
Involvement of anti-viral, pro- and anti-inflammatory signalling molecules, such as NF-kB2,
STAT1, and STAT3, was also detected mostly in infiltrating mononuclear leucocytes in brain
sections with elevated levels of IFN-I/II, TNFα, IL6 and IL10 transcripts. Thus, immunocompetent
rabbits are able to resolve virulent flavivirus infections via a well-coordinated antiviral, pro- and
anti-inflammatory immune response. Given the similarities to the non-lethal disease profile of most
human and equine infections, rabbits are a valuable model for studying the mechanisms of virus
control and pathogenesis of non-lethal disease.

This project has therefore advanced the field of WNV research by describing a previously
unacknowledged complication for studies of the pathogenesis of infection involving virus strains of
intermediate virulence, as well as having established an alternative animal model for studying virus
control and induction of non-lethal disease.
Keyword West Nile virus
Virus control
Animal model

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Created: Thu, 12 Jan 2017, 23:27:55 EST by Willy Suen on behalf of Learning and Research Services (UQ Library)