SummaryTo test the hypothesis that airborne microbial cell wall components could induce an in¯ammatory response in the lungs, measurements were made of the am ounts of bac terial endotoxin and (1! 3)-b-D-glucan in laborat ory animal bedding materials. Groups of rat s were exposed by inhalati on to airborne endotoxin, (1! 3)-b-D-glucan or a combination of the two for 5 weeks. T he results demonstrated that measurable amounts of endotoxin and (1! 3)-b-Dglucan could be detected in the different bedding materials. In contrast to animals at delivery, those kept on bedding for 5 weeks showed moderat e in¯ammatory reactions in the lung. T hese were most pronounced among animals exposed to endotoxin and (1! 3)-b-D-glucan. T he results suggest that further studies need to be undertaken to elucidat e the role of microbial cell wall products in the development of in¯ammatory lung responses am ong research animals. Keywords Lung; in¯am mation; endotoxin; (1! 3)-b-D-glucan; bedding T he need for high-quality laboratory research anim als was realized during the 1960s and 1970s, leading to the development of control of infectious agents and to animals of de®ned microbiological qualit y. In spite of always working with high-qualit y animals, we (BE and LE) have noticed over a number of years hyperreactivity in the airways among the rats in our experimental laboratory, expressed in an increased mortality in anaesthetized animals and unusual reactions in normally well-running animal models. T hese phenomena occurred particularly during autumn. Mac roscopically, the lungs showed multiple small grey nodules. Histological sections revealed mild to moderate multifocal interstitial pneumonia, alveolar histocytosis, lymphoid cuf®ng and perivascular eosinophilia. In spite of this, the animals were free from known respiratory and general infections. Similar observations have previously been reported by Elwell e t a l. (1997).During recent years, it has become evident that an in¯ammat ory response in the lungs may, apart from infections, be caused by microbial cell wall components, particularly endotoxin from Gram-negative bac teria and (1! 3)-b-D-glucan, a cell wall component of moulds, certain bac teria and plants (Fogelmark e t a l. 1994, Fogelmark & Rylander 1997

Exposure to organic dusts might cause an acute reaction with influenza like symptoms (toxic pneumonitis). Although it is well known that endotoxin, which is often present in organic dusts, may cause this reaction, there is no information about fungal cell wall agents as causative agents. The capacity of different fungal cell wall agents to induce an acute inflammation was evaluated in animal inhalation experiments. Guinea pigs were exposed to an aerosol of endotoxin, molds, and different fungal cell wall agents (FCWA) in a continuous flow exposure chamber, and the number of free lung cells was determined 24 h thereafter. Endotoxin caused a dose-dependent increase in the number of neutrophils and macrophages. None of the other agents tested caused a neutrophil response. The results suggest that toxic pneumonitis in environments with organic dusts is caused by the presence of inhaled endotoxin. Although no effect from FCWA was found, it is well known that this inhalation under chronic conditions will cause lung disease, particularly granulomatous pneumonitis.

Simple Summary:The mouse is the most commonly used mammal in scientific research, and housed in research facilities around the world. Mice are a social species, but when housing male mice together in a confined environment in the laboratory, aggression is often observed and can be problematic. Fighting or trying to avoid fighting can be stressful. Furthermore, fighting can lead to injuries which can sometimes be fatal. Mouse aggression is therefore a significant welfare problem and has implications on the 3Rs (Replacing, Reducing, and Refining animal use in scientific procedures and education). In this study, we used a survey and workshops to collect the experiences of animal technicians, veterinarians, and researchers at Swedish research animal facilities relating to mouse aggression and what methods of preventing aggression they practice. Both group housing and single-housing as a consequence of aggression was perceived as problematic and stressful for the animals. In line with current recommendations from the literature, participants perceived that aggression occurred less if mice were grouped with litter mates at an early age, that nesting material was transferred at cage cleaning, and disturbance was kept to a minimum. Experience from practice will play a valuable part in developing guidelines for group-housed male mice.Abstract: Animals used for scientific purposes are protected by EU legislation. Social animals should be kept in stable groups that enable species-typical social behavior and provide individuals with social comfort. However, when group-housing male mice, aggression within the homecage is a common husbandry and welfare problem. Excessive fighting and injuries due to aggression can cause pain and stress, resulting in individuals being euthanized or housed individually. In addition, stress can alter physiological parameters, risking scientific validity and generating larger sample sizes. Mouse aggression, and the consequences thereof, thus opposes the 3R goals of Refining the methods to Animals 2019, 9, 1010 2 of 24 minimize potential pain and suffering and Reducing the number of animals used. Animal technicians, veterinarians, and scientists using animals have valuable information on how these problems are experienced and handled in practice. We assembled these experiences from laboratory animal facilities in Sweden, mapping problems observed and identifying strategies used to prevent mouse aggression. In line with current literature, less aggression was perceived if mice were grouped before sexual maturity, re-grouping avoided and nesting material transferred at cage cleaning. Preventing aggression will minimize pain and suffering and enable housing of stable groups, leading to more reliable scientific outcomes and is thus of high 3Rs relevance.

Aggression among group-housed male mice is a major animal welfare concern often observed at animal facilities. Studies designed to understand the causes of male mice aggression have used different methodological approaches and have been heterogeneous, using different strains, environmental enrichments, housing conditions, group formations and durations. By conducting a systematic literature review based on 198 observed conclusions from 90 articles, we showed that the methodological approach used to study aggression was relevant for the outcome and suggested that home cage observations were better when studying home cage aggression than tests provoking aggression outside the home cage. The study further revealed that aggression is a complex problem; one solution will not be appropriate for all animal facilities and all research projects. Recommendations were provided on promising tools to minimize aggression, based on the results, which included what type of environmental enrichments could be appropriate and which strains of male mice were less likely to be aggressive.

Review question / Objective: By systematically reviewing articles investigating male mouse aggression we wanted to map how the literature in the field support, or not, the available recommendations on how to prevent aggression in group housed male mice, and to detect knowledge gaps that ought to be filled. We also wanted to address and describe how aggression have been measured in the literature, since this may influence the possibility to translate outcomes to normal husbandry conditions and contribute to useful recommendations. Condition being studied: Aggression between male cage mates is one of the main problems in laboratory mouse husbandry, affecting both animal welfare and scientific quality.