Proper micro-environment alleviates mortality in laboratory mouse breeding induced by litter overlap and older dams.

The ongoing worldwide effort to reduce animal numbers in research often omits the issue of pre-weaning mortality in mouse breeding. A conservative estimate of 20% mortality would mean approximately 1.1 M mice die annually in the EU before scientific use. We hypothesize that pre-weaning mortality in laboratory mouse breeding is associated with cage social and macro/micro-environment conditions. Here we count pups from 509 C57BL/6J litters daily for accurate detection of mortality, and monitor cage micro-environment for 172 C57BL/6J litters. Probability of pups to die increases with the increase in dam age, number and age of older pups in the cage (of overlapped/cohabitating litters), and in small (<6 pups) and large (>11 pups) focal litters. Higher temperatures (>23.6 °C) and nest scores (>3.75) compensate for some of the socially-associated risks for pup death. These findings can be implemented in strategies for reducing pre-weaning mouse mortality, a more welfare-friendly and sustainable approach for science.

Interdisciplinary Animal Research Ethics-Challenges, Opportunities, and Perspectives.

Can nonhuman animals be used for the benefit of humans in a scientifically and morally justified manner and, if yes, how? Based on our own experiences as scholars from various academic backgrounds, we argue that this question can only be answered as an interdisciplinary and international endeavor, considering insights from research ethics and animal ethics as well as scientific and legal aspects. The aim of this article is to contribute to the foundation of the emerging field of animal research ethics. In doing so, we describe the following seven phases of animal research experiments: ethical, legal and social presumptions (phase 0), planning (phase I), review (phase II), conduct of experiments (phase III), publication/dissemination (phase IV), further exploitation of results (phase V), and evaluation (phase VI). In total, 20 key ethical, legal, and practical challenges that an ethical framework for the use of animals in research needs to address are identified and analyzed. Finally, we characterize the following four meta-challenges and opportunities associated with animal research ethics as a field: (1) moral pluralism, (2) the integration of views and positions outside the laboratory, (3) international plurality of conduct, standards, and legal norms, and (4) interdisciplinary education.

From Mating to Milk Access: A Review of Reproductive Vocal Communication in Mice.

Vocalisations play a central role in rodent communication, especially in reproduction related behaviours. In adult mice (Mus musculus) the emission of ultrasonic vocalisations (USVs) has been observed in courtship and mating behaviour, especially by males. These have been found to have distinctive individual signatures that influence female choice of mating partner. The most recent findings show that vocal communication also has a role in parental cooperation, in that female mice communicate with male partners in ultrasonic frequencies to induce paternal behaviour. Infant vocalisations form the other important part of reproductive vocal communication. Although born deaf, neonatal mice are capable of producing vocalisations since birth. As an altricial species, successful mother-infant communication is essential for survival, and these vocalisations are important modulators of maternal behaviour. Three main types of infant vocalisations have been identified and characterised. Most research has addressed pure USVs, related to stressful situations (e.g., cold, isolation, handling, presence of unfamiliar males or predators), which usually elicit maternal search and retrieval. In addition, broad-band spectrum signals, emitted post-partum during cleaning of foetal membranes, inhibit biting and injury by adults and "wriggling calls," emitted during suckling, release maternal behaviour (such as licking). Several variables have been identified to modulate vocalisations in mice, including individual characteristics such as strain/genotype, age, sex, and experimental factors such as pharmacological compounds and social context. In recent years, there has been a big increase in the knowledge about the characteristics of vocal communication in rodents due to recent technological advances as well as a growing interest from the neuroscience community. Vocalisation analysis has become an essential tool for phenotyping and evaluating emotional states. In this review, we will (i) provide a comprehensive summary of the current knowledge on mouse reproductive vocal communication and (ii) discuss the most recent findings in order to provide a broad overview on this topic.

All the Pups We Cannot See: Cannibalism Masks Perinatal Death in Laboratory Mouse Breeding but Infanticide Is Rare.

Perinatal mortality is a major issue in laboratory mouse breeding. We compared a counting method using daily checks (DAILY_CHECK) with a method combining daily checks with detailed video analyses to detect cannibalisms (VIDEO_TRACK) for estimating the number of C57BL/6 pups that were born, that died and that were weaned in 193 litters from trios with (TRIO-OVERLAP) or without (TRIO-NO_OVERLAP) the presence of another litter. Linear mixed models were used at litter level. To understand whether cannibalism was associated with active killing (infanticide), we analysed VIDEO_TRACK recordings of 109 litters from TRIO-OVERLAP, TRIO-NO_OVERLAP or SOLO (single dams). We used Kaplan-Meier method and logistic regression at pup level. For DAILY_CHECK, the mean litter size was 35% smaller than for VIDEO_TRACK (p < 0.0001) and the number of dead pups was twice lower (p < 0.0001). The risk of pup loss was higher for TRIO-OVERLAP than TRIO-NO_OVERLAP (p < 0.0001). A high number of pup losses occurred between birth and the first cage check. Analyses of VIDEO_TRACK data indicated that pups were clearly dead at the start of most of the cannibalism events and infanticide was rare. As most pups die and disappear before the first cage check, many breeding facilities are likely to be unaware of their real rates of mouse pup mortality.

Necropsy protocol for newborn mice.

Neonatal mortality is high in laboratory mouse breeding, and causes are poorly understood. Post-mortem analysis of pups is an often overlooked source of information and insight. We present a necropsy protocol for neonatal mice designed for easy practical application by animal technicians.

2,4,6-trinitrobenzenesulfonic acid-induced colitis in Rattus norgevicus: a categorization proposal.

Reproducibility in animal research is crucial for its reliance and translational relevance. The 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced model of inflammatory bowel disease (IBD) is widely used but inconsistently and incompletely characterized throughout the literature. This hinders comparisons between studies and influences the low rate of translation of effective preclinical molecules. The purpose of this study was to categorize TNBS-induced colitis, based on macroscopic and microscopic scoring systems, and to identify basic routine parameters that could anticipate those categories. We retrospectively analysed male Wistar Rattus norvegicus (n=28 for the control group and n=87 for the TNBS group) and categorized TNBS-induced colitis in three phenotypes: Mild, Moderate and Severe colitis, as for human IBD. Also, we showed that the time course of food intake and fecal excretion (but not body weight, fluid intake or welfare scores) could foresee those categories. So, routine evaluation of food intake and fecal excretion may guide researchers in planning their experiments, selecting the animals with the severity of colitis that better matches their aims, or applying early humane endpoints to animals that will not be used in the experiments. In conclusion, categorizing TNBS-induced colitis enhances the reproducibility of data gathered with this experimental model and strengths its translational relevance.

High laboratory mouse pre-weaning mortality associated with litter overlap, advanced dam age, small and large litters.

High and variable pre-weaning mortality is a persistent problem in laboratory mouse breeding. Assuming a modest 15% mortality rate across mouse strains, means that approximately 1 million more pups are produced yearly in the EU to compensate for those which die. This paper presents the first large study under practical husbandry conditions to determine the risk factors associated with mouse pre-weaning mortality. We analysed historical records from 219,975 pups from two breeding facilities, collected as part of their management routine and including information on number of pups born and weaned per litter, parents' age and identification, and dates of birth and death of all animals. Pups were counted once in their first week of life and at weaning, and once every one or two weeks, depending on the need for cage cleaning. Dead pups were recorded as soon as these were found during the daily cage screening (without opening the cage). It was hypothesized that litter overlap (i.e. the presence of older siblings in the cage when new pups are born), a recurrent social configuration in trio-housed mice, is associated with increased newborn mortality, along with advanced dam age, large litter size, and a high number and age of older siblings in the cage. The estimated probability of pup death was two to seven percentage points higher in cages with litter overlap compared to those without. Litter overlap was associated with an increase in death of the entire litter of five and six percentage points, which represent an increase of 19% and 103% compared to non-overlapped litters in the two breeding facilities, respectively. Increased number and age of older siblings, advanced dam age, small litter size (less than four pups born) and large litter size (over 11 pups born) were associated with increased probability of pup death.

Food Chain Information: Data Quality and Usefulness in Meat Inspection in Portugal.

Food chain information (FCI) is a mandatory component of meat inspection in the European Union. In Portugal there has been no assessment of FCI regarding risk-based meat inspection. The goal of the present study was to assess the quality and the usefulness of data provided by farmers. A total of 1,694 reports (989 for bovine animals, 575 for swine, and 130 for small ruminants) submitted to nine red meat slaughterhouses between September 2015 and January 2017 were analyzed. These reports covered a total of 79,889 animals. Overall, information was provided for the majority of the general identification items. To assess the quality of the information on health status, medication, disease occurrence, diagnostic tests, and results of previous ante- and postmortem exams, responses were classified as plausible, "nothing to declare," invalid, and nonresponse. Plausible information was provided by farmers on health status, medication, tests, and diseases on 82.7, 24.6, 5.4, and 0.1% of the FCI forms, respectively. More than 70% of the responses in all categories except health status were "nothing to declare." In pigs, when comparing ante- and postmortem inspection results with the corresponding FCI, no relevant contribution of the FCI was observed. Inaccurate or irrelevant information was provided for the majority of the detailed questions, impeding the formulation of appropriate risk-based meat inspection decisions. Farmers seem to have difficulty providing specific information. Better forms and better training are needed. Information on the prevalence of biological hazards relevant to public health currently is not available. Our results support the need to improve the systems used to obtain and integrate relevant information from the farm to the slaughterhouse.