Heterogeneous pathological outcomes after experimental pH1N1 influenza infection in ferrets correlate with viral replication and host immune responses in the lung

Vidaña, Beatriz; Martínez, Jorge; Martínez-Orellana, Pamela; García Migura, Lourdes; Montoya, María; Martorell, Jaime; Majó, Natàlia

Ten ferrets between the ages of twelve and twenty-four months that were seronegative for influenza A virus (influenza A antibody competition multi-species ELISA, ID Screen®, France) and Aleutian disease virus were randomly selected from a stable, purposely bred colony (Isoquimen, Spain). Upon arrival at the Centre de Recerca en Sanitat Animal (CReSA), the animals were placed in biosafety level 3 (BSL-3) facilities. The ferrets were randomly assigned to different experimental groups, separated into experimental isolation rooms and then maintained for one week for acclimation. The animals were maintained in standard housing cages and provided with commercial food pellets and tap water ad libitum throughout the experiment. All experiments were performed under a protocol (n°1976) that was reviewed and approved by the Animal and Human Experimentation Ethics Commission of the Universitat Autònoma de Barcelona. All ferrets were monitored daily for clinical signs, and any animal that was determined to be in a moribund state was ethically euthanized. Any animal losing 25% of its original body weight or presenting severe respiratory signs such as severe tachypnea, nasal and conjunctival exudates and/or severe apathy (irresponsiveness to external stimulus) were considered to be in a moribund state and was humanely euthanized. Anorexia was measured daily by observing the amount of faeces and the amount of food that needed to be replaced and the attitude of the animals towards food. Depression was evaluated with reference to the activity of the animals and their response to external stimulus (such as sound or human presence). Tachypnea was considered when respiratory rate was higher than 70 breaths per minute, accompanied by abdominal breathing.