Characterization of non-lethal West Nile Virus (WNV) infection in horses: Subclinical pathology and innate immune response Helle Bielefeldt-Ohmann a, b, c, * , Angela Bosco-Lauth d , Airn-Elizabeth Hartwig d , M. Jasim Uddin b , Jean Barcelon c , Willy W. Suen b , Wenqi Wang b , Roy A. Hall a, c , Richard A. Bowen d a Australian Infectious Diseases Research Centre, University of Queensland, Australia b School of Veterinary Science, University of Queensland, Australia c School of Chemistry and Molecular Biosciences, University of Queensland, Australia d College of Veterinary Medicine & Biomedical Sciences, Colorado State University, Fort Collins, CO, USA article info Article history: Received 22 August 2016 Received in revised form 27 November 2016 Accepted 19 December 2016 Available online 21 December 2016 Keywords: Flavivirus Arbovirus Equine Interferons Innate immune response abstract Most natural West Nile virus (WNV) infections in humans and horses are subclinical or sub-lethal and non-encephalitic. Yet, the main focus of WNV research remains on the pathogenesis of encephalitic disease, mainly conducted in mouse models. We characterized host responses during subclinical WNV infection in horses and compared outcomes with those obtained in a novel rabbit model of subclinical WNV infection (Suen et al. 2015. Pathogens, 4: 529). Experimental infection of 10 horses with the newly emerging WNV-strain, WNV NSW2011 , did not result in neurological disease in any animal but transcrip- tional upregulation of both type I and II interferon (IFN) was seen in peripheral blood leukocytes prior to or at the time of viremia. Likewise, transcript upregulation for IFNs, TNFa, IL1b, CXCL10, TLRs, and MyD88 was detected in lymphoid tissues, while IFNa, CXCL10, TLR3, ISG15 and IRF7 mRNA was upregulated in brains with histopathological evidence of mild encephalitis, but absence of detectable viral RNA or an- tigen. These responses were reproduced in the New Zealand White rabbits (Oryctolagus cuniculus) experimentally infected with WNV NSW2011, by intradermal footpad inoculation. Kinetics of the anti-WNV antibody response was similar in horses and rabbits, which for both species may be explained by the early IFN and cytokine responses evident in circulating leukocytes and lymphoid organs. Given the similarities to the majority of equine infection outcomes, immunocompetent rabbits appear to represent a valuable small-animal model for investigating aspects of non-lethal WNV infections, notably mecha- nisms involved in abrogating morbidity. © 2016 Elsevier Ltd. All rights reserved. 1. Introduction West Nile virus (WNV), a mosquito-borne flavivirus in the family Flaviviridae, is widely distributed throughout Africa, the Middle East, Asia, Southern Europe, Australia and the Americas. Since the 1990s, WNV has been the causative agent for numerous equine and human encephalitis outbreaks worldwide, highlighting it as an important re-emerging neurotropic virus. However, the current rodent (mouse and hamster) models for studying the pathogenesis of WNV infection have several shortcomings limiting their ability to accurately model equine and human infection. These include markedly different levels of virus replication in the central nervous system (CNS), as well as in peripheral tissues in rodents compared to horses [1e4]. The much exaggerated level of especially Abbreviations: CNS, central nervous system; CXCL10, chemokine (C-X-C motif) ligand 10; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; HE, hematoxylin and eosin; HIF-1a, hypoxia inducible factor 1a; IFN, Interferon; IL, interleukin; IRF, interferon regulatory factor; ISG, interferon-stimulated genes; IU, infectious units; mAb, monoclonal antibody; MyD88, myeloid differentiation 88; NF-kB, nuclear factor kappa- B; NSW, New South Wales; NZW, New Zealand white; PFU, plaque forming units; p.i., post infection; PRNT, plaque reduction neutralization test; PTX3, pentraxin-related protein 3; qRT-PCR, quantitative reverse transcriptase polymer- ase chain reaction; STAT, Signal transducer and activator of transcription; TCID 50 , 50% tissue culture infectious dose; TLR, Toll-like receptors; TNFa, tumor necrosis factor a; TRAF, TNF receptor-associated factor; VEGF, vascular endothelial growth factor; WNV, West Nile virus. * Corresponding author. School of Veterinary Science, University of Queensland, Gatton 4343, Australia. E-mail address: h.bielefeldtohmann1@uq.edu.au (H. Bielefeldt-Ohmann). Contents lists available at ScienceDirect Microbial Pathogenesis journal homepage: www.elsevier.com/locate/micpath http://dx.doi.org/10.1016/j.micpath.2016.12.018 0882-4010/© 2016 Elsevier Ltd. All rights reserved. Microbial Pathogenesis 103 (2017) 71e79