Spillover of bat borne rubulavirus in Australian horses – Horses as sentinels for emerging infectious diseases

Background: Over 1000 horses are investigated annually for Hendra virus (HeV)-like illness, of which very few (<1%) test HeV positive. In Australia, in addition to HeV, other zoonotic viruses have affected horses including Australian bat lyssavirus, West Nile Virus (Kunjin), Murray Valley encephalitis virus and Ross River virus. In 1997, Menangle virus (MenPV), family Paramyxovirus, genus Rubulavirus, caused severe reproductive failure in pigs and influenza-like illness with rash in two piggery staff. MenPV has been isolated from Australian flying fox urine along with novel related rubulaviruses and HeV. We describe evidence of natural exposure to bat-borne rubulaviruses in Australian horses as well as seroconversion suggesting causality of severe respiratory illness. Methods and materials: Three-hundred-and-seventy-four horses were tested by a multiplex microsphere-based immunoassay (MIA) for IgG against MenPV nucleocapsid (N) protein and a subset also against Tioman virus (TioPV) N protein and MenPV hemagglutinin-neuraminidase (HN) protein. Confirmatory testing comprised immunofluorescence assay (IFA) on Vero cells infected with MenPV and related rubulaviruses. Results: Median fluorescence intensities (MFI) against MenPV N and a prior prevalence estimate of 20% were used in a Bayesian latent class model to determine appropriate cut-offs for positive test classification. Assay sensitivity was estimated assuming a specificity of both 95% and 99%. MFI reflecting potentially significant IgG to MenPV N protein was demonstrated in 34% (94/274) of horses with high perceived flying fox exposure (29% in QLD and 32% in NSW) whereas horses without plausible exposure recorded insignificant MFI. IFA confirmed antibodies to three of five related flying fox rubulaviruses tested (MenPV, Yeppoon virus and Grove virus). Case presentations: Two young-adult geldings developed severe acute respiratory illness in 2016 featuring obtunded demeanour, tachypnoea, tachycardia, congested/ hy