Health Monitoring of Laboratory Rodent Colonies—Talking about (R)evolution

The health monitoring of laboratory rodents is essential for ensuring animal health and standardization in biomedical research. Progress in housing, gnotobiotic derivation, and hygienic monitoring programs led to enormous improvement of the microbiological quality of laboratory animals. While traditional health monitoring and pathogen detection methods still serve as powerful tools for the diagnostics of common animal diseases, molecular methods develop rapidly and not only improve test sensitivities but also allow high throughput analyses of various sample types. Concurrently, to the progress in pathogen detection and elimination, the research community becomes increasingly aware of the striking influence of microbiome compositions in laboratory animals, affecting disease phenotypes and the scientific value of research data. As repeated re-derivation cycles and strict barrier husbandry of laboratory rodents resulted in a limited diversity of the animals’ gut microbiome, future monitoring approaches will have to reform—aiming at enhancing the validity of animal experiments. This review will recapitulate common health monitoring concepts and, moreover, outline strategies and measures on coping with microbiome variation in order to increase reproducibility, replicability and generalizability

Running in the wheel: Defining individual severity levels in mice.

The fine-scale grading of the severity experienced by animals used in research constitutes a key element of the 3Rs (replace, reduce, and refine) principles and a legal requirement in the European Union Directive 2010/63/EU. Particularly, the exact assessment of all signs of pain, suffering, and distress experienced by laboratory animals represents a prerequisite to develop refinement strategies. However, minimal and noninvasive methods for an evidence-based severity assessment are scarce. Therefore, we investigated whether voluntary wheel running (VWR) provides an observer-independent behaviour-centred approach to grade severity experienced by C57BL/6J mice undergoing various treatments. In a mouse model of chemically induced acute colitis, VWR behaviour was directly related to colitis severity, whereas clinical scoring did not sensitively reflect severity but rather indicated marginal signs of compromised welfare. Unsupervised k-means algorithm-based cluster analysis of body weight and VWR data enabled the discrimination of cluster borders and distinct levels of severity. The validity of the cluster analysis was affirmed in a mouse model of acute restraint stress. This method was also applicable to uncover and grade the impact of serial blood sampling on the animal's welfare, underlined by increased histological scores in the colitis model. To reflect the entirety of severity in a multidimensional model, the presented approach may have to be calibrated and validated in other animal models requiring the integration of further parameters. In this experimental set up, however, the automated assessment of an emotional/motivational driven behaviour and subsequent integration of the data into a mathematical model enabled unbiased individual severity grading in laboratory mice, thereby providing an essential contribution to the 3Rs principles

Time to Integrate to Nest Test Evaluation in a Mouse DSS-Colitis Model

<div><p>Severity assessment in laboratory animals is an important issue regarding the implementation of the 3R concept into biomedical research and pivotal in current EU regulations. In mouse models of inflammatory bowel disease severity assessment is usually undertaken by clinical scoring, especially by monitoring reduction of body weight. This requires daily observance and handling of each mouse, which is time consuming, stressful for the animal and necessitates an experienced observer. The time to integrate to nest test (TINT) is an easily applicable test detecting disturbed welfare by measuring the time interval mice need to integrate nesting material to an existing nest. Here, TINT was utilized to assess severity in a mouse DSS-colitis model. TINT results depended on the group size of mice maintained per cage with most consistent time intervals measured when co-housing 4 to 5 mice. Colitis was induced with 1% or 1.5% DSS in group-housed WT and <i>Cd14</i>-deficient mice. Higher clinical scores and loss of body weight were detected in 1.5% compared to 1% DSS treated mice. TINT time intervals showed no dose dependent differences. However, increased clinical scores, body weight reductions, and increased TINT time intervals were detected in <i>Cd14</i>^<i>-/-</i> compared to WT mice revealing mouse strain related differences. Therefore, TINT is an easily applicable method for severity assessment in a mouse colitis model detecting CD14 related differences, but not dose dependent differences. As TINT revealed most consistent results in group-housed mice, we recommend utilization as an additional method substituting clinical monitoring of the individual mouse.</p></div

Intestinal inflammation induced by DSS-treatment.

<p>Hematoxylin and eosin staining of colon tissue obtained from (A-D) wild-type and (E-H) <i>Cd14</i>-deficient mice treated with 1% DSS for seven days (C-D and G-H, respectively). Untreated controls (A-B and E-F) did not show any signs of inflammation. Colitis was characterized by the presence of mixed cell infiltrates, hyperplasia, abnormal crypt architecture, edema and erosions (see boxed magnifications D and H). Original magnification 5x and 10x. Histological score quantifying the alterations observed in the colon (J).</p

Clinical disease activity score after DSS treatment.

<p>Assessment of severity in controls (A, B) and DSS treated mice (1% DSS C, D and 1.5% DSS E, F) determined by an overall clinical disease activity score (A, C, E) and specifically by the change in body weight (B, D, F). Untreated controls exhibited low clinical scores (A) and a steady body weight (B). Mice treated with 1% (C, D) or 1.5% DSS (E, F) demonstrated increasing clinical scores and loss of body weight. <i>Cd14</i>^<i>-/-</i> mice showed significantly higher clinical scores and a significantly higher reduction of body weight than WT mice.</p

Histology score.

<p>Histology score.</p

Colitis severity assessment by utilizing TINT.

<p>TINT time intervals determined in controls (A) and 1% (B) as well as 1.5% DSS treated mice (C). TINT time intervals were significantly increased in 1% DSS treated <i>Cd14</i>^<i>-/-</i> mice compared to WT mice on day 7 post colitis-induction (B).</p

Clinical colitis scoring.

<p>Clinical colitis scoring.</p