Abstract
Ferrets are widely used to study human influenza virus infection. Their airway physiology and cell receptor distribution makes them ideal for the analysis of pathogenesis and virus transmission, and for testing the efficacy of anti-influenza interventions and vaccines. The 2009 pandemic influenza virus (H1N1pdm09) induces mild to moderate respiratory disease in infected ferrets, following inoculation with 106 plaque-forming units (pfu) of virus. We have demonstrated that reducing the challenge dose to 102 pfu delays the onset of clinical signs by 1 day, and results in a modest reduction in clinical signs, and a less rapid nasal cavity innate immune response. There was also a delay in virus production in the upper respiratory tract, this was up to 9-fold greater and virus shedding was prolonged. Progression of infection to the lower respiratory tract was not noticeably delayed by the reduction in virus challenge. A dose of 104 pfu gave an infection that was intermediate between those of the 106 pfu and 102 pfu doses. To address the hypothesis that using a more authentic low challenge dose would facilitate a more sensitive model for antiviral efficacy, we used the well-known neuraminidase inhibitor, oseltamivir. Oseltamivir-treated and untreated ferrets were challenged with high (106 pfu) and low (102 pfu) doses of influenza H1N1pdm09 virus. The low dose treated ferrets showed significant delays in innate immune response and virus shedding, delayed onset of pathological changes in the nasal cavity, and reduced pathological changes and viral RNA load in the lung, relative to untreated ferrets. Importantly, these observations were not seen in treated animals when the high dose challenge was used. In summary, low dose challenge gives a disease that more closely parallels the disease parameters of human influenza infection, and provides an improved pre-clinical model for the assessment of influenza therapeutics, and potentially, influenza vaccines.
Highlights
In April 2009, a novel H1N1 influenza A virus of swine origin emerged from North America and spread around the world, resulting in the first influenza pandemic of the 21st century [1]
Low dose challenge leads to delayed clinical disease kinetics and increases virus shedding
This study determined the effect of reducing intra-nasal dose of infectious virus on the kinetics of virus shedding and disease progression, and compared the effects of oseltamivir treatment on ferrets infected with a high or low virus dose
Summary
In April 2009, a novel H1N1 influenza A virus of swine origin emerged from North America and spread around the world, resulting in the first influenza pandemic of the 21st century [1]. Infection of ferrets via the intra-nasal route with H1N1pdm virus has typically induced a mild to moderate, nonlethal infection resulting in weight loss, transient pyrexia, and mild upper respiratory tract signs These studies have typically used a dose of 106 TCID50 per ferret in order to ensure all ferrets become infected [9,10,11,12]. Use of high dose inocula may enhance pathogenicity and accelerate infection kinetics, which may obscure the effects of antiviral interventions It has been shown, for example, that a high dose of influenza A virus (106.8 EID50) overcame the protective effect of a defective interfering virus preparation, which was fully protective in mice when a lower challenge dose (103.5 EID50 virus) was used [18]. This consideration is relevant for the ferret model of influenza, as these animals have been used extensively in demonstrating the efficacy of anti-influenza drugs such as the neuraminidase inhibitors oseltamivir, zanamivir and peramivir [12,19,20,21,22,23]
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