Development of a novel primate welfare assessment tool for research macaques.

Primates are important species for biomedical research and ensuring their good welfare is critical for research translatability and ethical responsibility. Systematic animal welfare assessments can support continuous programme improvements and build institutional awareness of areas requiring more attention. A multi-facility, collaborative project aimed to develop and implement a novel primate welfare assessment tool (PWAT) for use with research macaques. PWAT development involved: establishing an internal focus group of primate subject matter experts, identifying animal welfare categories and descriptors based on literature review, developing a preliminary tool, beta-testing the tool to ensure practicality and final consensus on descriptors, finalising the tool in a database with semi-automated data analysis, and delivering the tool to 13 sites across four countries. The tool uses input- and outcome-based measures from six categories: physical, behavioural, training, environmental, procedural, and culture of care. The final tool has 133 descriptors weighted based upon welfare impact, and is split into three forms for ease of use (room level, site level, and personnel interviews). The PWAT was trialled across facilities in March and September 2022 for benchmarking current macaque behavioural management programmes. The tool successfully distinguished strengths and challenges at the facility level and across sites. Following this benchmarking, the tool is being applied semi-annually to assess and monitor progress in behavioural management programmes. The development process of the PWAT demonstrates that evidence-based assessment tools can be developed through collaboration and consensus building, which are important for uptake and applicability, and ultimately for promoting global improvements in research macaque welfare.

Implementation of an Alternative Training Method for Cardiac Blood Collection in Mice.

Training personnel to work with animals presents a variety of challenges, both logistically and with regard to animal welfare. These issues make training an ideal opportunity to evaluate practices and to implement the 3R principles (refinement, replacement, and reduction). Cardiac blood collection from mice is a procedure that can compromise the 3Rs by requiring repeated practice and animal euthanasia. The development of a non-animal training model would promote the 3R principles. Our goals for the development of a new training model for cardiac blood collection from mice were to reduce the number of mice needed to achieve competency, improve our culture of care, and refine the training approach by improving competency. The training model was developed using commonly available materials. The total cost of the model was less than $15 USD per model. Two training curricula were conducted concurrently over a 5-mo period: 1) a curriculum in which trainees used the model before progressing to live mice and 2) the traditional curriculum, which used euthanized mice throughout. The measured variables included the total number of mice used, proportions of trainees who reached competency, the time needed to reach competency, method comprehension, quality of skill performance, trainer and trainee feedback, and training costs. The alternative group used at least 10 fewer mice per technician as compared with the traditionally trained group. The alternative group had a higher competency rate, with 82% (9 of 11 trainees) reaching competency compared with 60% (3 of 5 trainees) in the traditional group. Skill comprehension and quality were superior in the alternative group, as evidenced by fewer gross lesions at necropsy. Overall, personnel in the alternative group provided positive feedback with regard to the use of fewer mice, acquisition of both skill and confidence, and benefits for compassion fatigue. The use of this model is now our standard approach for training personnel in cardiac blood collection in mice. Our results demonstrate that the use of models in training curricula can enhance skill development and reduce the use of mice.

Using Stakeholder Focus Groups to Refine the Care of Pigs Used in Research.

Research organizations should be proactive in regularly evaluating and refining their animal care and use programs in order to advance animal welfare and minimize distress. Pigs are often used in research, but few empirical studies have examined optimal husbandry and research use practices for pigs in a research environment. We developed the Pig Welfare Working Group (PWWG) to address the need for more formal guidelines on the management and use of pigs in research. The PWWG was a stakeholder focus group whose goal was to identify challenges and opportunities relevant to improving animal welfare through collaboration, knowledge sharing, and inclusive decision-making. Through consensus building, the PWWG developed 12 recommendations for behavioral management, housing, research procedures, transportation, and rehoming programs. The recommendations were rolled out across the contract research organization, business units, sites, and countries. Follow up will be conducted regularly to assess welfare, monitor progress toward implementing the recommendations, and recognize and reward participants making changes at their site.

Development and validation of a cynomolgus macaque grimace scale for acute pain assessment.

Cynomolgus macaques may undergo surgical procedures for scientific and veterinary purposes. Recognition and assessment of pain using validated tools is a necessary first step for adequately managing pain in these primates. Grimace scales are one means of assessing the occurance of acute pain using action units such as facial expressions and posture. The aim of this study was to create and validate a Cynomolgus Macaque Grimace Scale (CMGS). Cynomolgus macaques (n = 43) were video recorded before and after a surgical procedure. Images were extracted from videos at timepoints at which breakthrough pain might be expected based on analgesic pharmacokinetics. Using the CMGS images were scored by 12 observers blinded to animal identification, times, and conditions. To validate the tool, detailed behavioral analyses emphasizing changes to baseline activity ethograms were compared to grimace scores. Four action units were identified related to potential pain including orbital tightening, brow lowering, cheek tightening, and hunched posture. The CMGS tool was found to have moderate inter- (ICCaverage action unit mean ± SD: 0.67 ± 0.28) and good intra- (ICCsingle mean ± SD: 0.79 ± 0.14) observer reliability. Grimace scores increased significantly (p < 0.0001) in the first four post-operative timepoints compared to baseline, correlating with behavioral findings (rho range = 0.22-0.35, p < 0.001). An analgesic intervention threshold was determined and should be considered when providing additional pain relief. The CMGS was shown to be a reliable and valid tool; however, more research is needed to confirm external validity. This tool will be highly valuable for refining analgesic protocols and acute peri-procedural care for cynomolgus macaques.

Use of nonaversive handling and training procedures for laboratory mice and rats: Attitudes of American and Canadian laboratory animal professionals.

Nonaversive or low stress handling techniques can reduce fear and stress in research rodents, ultimately improving study data quality. Uptake of low stress handling has been slow in the USA and Canada. In this study we explored the understanding, experience, and attitudes toward low stress handling of rats and mice in laboratory animal professionals from the USA (US) and Canada (CA). Participants (n = 40) were recruited for a standardized interview and job categories were divided into veterinary/PhD level roles (doctoral level; DL) and non-veterinary/non-PhD level roles (non-doctoral level, NDL) (US: 23, DL: 9, NDL: 14; CA: 17, DL: 8, and NDL: 9). Interviews were transcribed and analyzed using NVIVO. Two research assistants independently coded themes for each question and consolidated responses based on commonality. Laboratory animal professionals understood the benefits of low stress handling and training techniques with rats and mice, stating reduced stress, better data, and improved welfare, with CA participants more likely to mention animal welfare as a benefit, and DL more likely to mention improved research data and reduced stress. Participants across demographic groups indicated improved job satisfaction and decreased stress as the positive impacts low stress handling would have on their positions. The primary perceived barriers to low stress handling implementation were researcher attitudes, the time needed to implement and use these techniques, and training personnel to use the techniques properly and consistently. To promote refinement of handling of rats and mice, more educational opportunities on the benefits and implementation of low stress handling techniques need to be provided to laboratory animal professionals, as well as to researchers.

Refining restraint techniques for research pigs through habituation.

Pigs are common research models and are strong animals that can be difficult to restrain. Improper restraint can put pigs and research personnel at risk for injury and induce stress, which can affect research outcomes. This study aimed to refine restraint techniques for research pigs using habituation and operant conditioning. Forty-four (22 males, 22 females; 4 months old, ~8.1 kg) Göttingen minipigs were randomly assigned to a control (C: no interventions) or a treatment group (T). Pigs in the T group received 3 min training sessions 3 days/week for the first 14 d after arrival. Training sessions included human socialization and habituation to a hammock sling for blood collection. Blood collection occurred on day 13 for all pigs by novel technicians. Pigs were placed in the sling, blood was collected from the radial vein, and serum cortisol levels were determined (ug/dL). Pig behavior was recorded and scored for duration of time spent struggling (s) and vocalizing (s). Novel human approach tests occurred on day 12, before blood collection, and day 14, after blood collection. Pigs were scored on latency to touch the human (s) and duration of time spent in contact with the human (s). Pig weight was taken upon arrival and on day 15. Separate linear models were fitted for response variables struggle duration in sling, serum cortisol, latency to touch human, time spent in contact with human, and body weight. Fixed effects were treatment and sex. Prior to blood collection, there was no difference in response to a novel human (P > 0.05) but after blood collection, T pigs were quicker to approach (estimate: -5.352, SE: 1.72, P = 0.003) and spent more time in contact with the novel human (estimate: 3.091, SE: 1.448, P = 0.039). T pigs also had lower cortisol levels during blood collection (estimate: -2.36, SE: 0.657, P = 0.001). There was no difference in behavior while in the sling (P > 0.05). The results of the study suggest that even small investments in habituation and training pigs to study procedures is beneficial in reducing stress and improving human-animal relationships, but more time would be beneficial to promote calmer behavior in the sling.

Compassion Fatigue and Coping Mechanisms of Laboratory Animal Professionals from Europe, China, and Japan.

Laboratory animal professionals (LAP) may experience situations that contribute to compassion fatigue (CF). The goal of this research was to better understand CF in LAP in and across employment categories. Surveys were distributed through LAP organizational listservs in the European Union (EU), China, and Japan, and results were analyzed to identify CF prevalence, personal and work-related factors, coping mechanisms, and beneficial work-support programs. Independent χ²-tests compared personal and work-related factors and feelings of CF. Feelings of CF and coping mechanisms were compared with personality scores using independent sample t tests. There were 302 respondents from the EU, 39 from China, and 77 from Japan. Over half of respondents from the EU (52%) and China (56%) reported experiencing CF (52%), with fewer (32%) reporting CF in Japan. No major differences were found based on employer type. Personality scores were significantly related to feelings of CF and preferred coping mechanisms. Work-related factors that contributed to feelings of CF in over half of respondents included staffing levels, workplace relationships, and availability of programs geared to address CF. Across regions, talking to someone, physical activity, getting away from work, and self-care were effective coping mechanisms in over 50% of respondents. Fewer than 30% of respondents indicated that their place of employment had CF support programs, and even fewer (8% to 28%) indicated that these programs were helpful. The study results suggest that to be effective, employer CF programs for LAP should consider providing quiet places at work and programs for self-care, promoting physical and mental health and social support systems, and establishing opportunities to memorialize animals.

The Social Life of Pigs: Changes in Affiliative and Agonistic Behaviors following Mixing.

This study investigated potentially affiliative behaviors in grow-finish pigs, how these behaviors changed over time and their relationship to agonistic behaviors. A total of 257 Yorkshire barrows were observed for agonistic (reciprocal fights, attacks) and affiliative (nosing, play, non-agonistic contact) behaviors after mixing (at 10 weeks of age), and weeks 3, 6, and 9 after mix. The least square means of affiliative behaviors were compared across time points. Relationships among affiliative and agonistic behaviors were assessed using generalized linear mixed models. Non-agonistic contact with conspecifics increased until week 6 then remained stable between weeks 6 and 9. Nosing was highest at mix, then decreased in the following weeks. Play was lowest at mix and highest at week 3. Affiliative behaviors were negatively related with aggression at mix (p < 0.001). Pigs who engaged in play and nosing behaviors were more likely to be involved in agonistic interactions in the weeks after mixing (p < 0.05), while pigs engaging in non-agonistic contact were less likely to be involved in agonistic interactions (p < 0.001). There appear to be relationships between affiliative and agonistic behaviors in pigs, with contact being the most predictive of less aggression. Future studies could focus on promoting positive non-agonistic contact in unfamiliar pigs as a way to mitigate aggressive interactions.

Developing Recommendations for Cumulative Endpoints and Lifetime Use for Research Animals.

Research animals are important for scientific advancement, and therefore, their long-term welfare needs to be monitored to not only minimize suffering, but to provide positive affective states and experiences. Currently, there is limited guidance in countries around the world on cumulative and experimental endpoints. This paper aims to explore current opinions and institutional strategies regarding cumulative use and endpoints through a scoping survey and review of current regulations and welfare assessment tools, and ultimately to provide recommendations for assessment of cumulative and lifetime use of research animals. The survey found that only 36% of respondents indicated that their institution had cumulative use endpoint policies in place, but these policies may be informal and/or vary by species. Most respondents supported more specific guidelines but expressed concerns about formal policies that may limit their ability to make case-by-case decisions. The wide diversity in how research animals are used makes it difficult for specific policies to be implemented. Endpoint decisions should be made in an objective manner using standardized welfare assessment tools. Future research should focus on robust, efficient welfare assessment tools that can be used to support planning and recommendations for cumulative endpoints and lifetime use of research and teaching animals.

Time budgets of group-housed pigs in relation to social aggression and production.

Commercial producers house growing pigs by sex and weight to allow for efficient use of resources and provide pigs the welfare benefits of interacting with their conspecifics and more freedom of movement. However, the introduction of unfamiliar pigs can cause increased aggression for 24 to 48 h as pigs establish social relationships. To address this issue, a better understanding of pig behavior is needed. The objectives of this study were to quantify time budgets of pigs following introduction into a new social group and how these changed over time and to investigate how social aggression influences the overall time budgets and production parameters. A total of 257 grow-finish Yorkshire barrows across 20 pens were introduced into new social groups at 10 wk of age (~23 kg) and observed for aggression and time budgets of behavior at four periods: immediately after introduction and 3, 6, and 9 wk later. Pigs were observed for the duration of total aggression and initiated aggression (s) for 9 h after introduction and for 4 h at 3, 6, and 9 wk later. Time budgets were created by scan sampling inactive, movement, ingestion, social, and exploration behaviors every 2 min for 4 h in the afternoon and summarizing the proportion of time each behavior was performed by period. The least square means of each behavior were compared across time points. Pigs spent most of their time inactive. In general, the greatest change in pig behavior was observed between introduction and week 3 (P < 0.003), with gradual changes throughout the study period as pigs became more inactive (week 3 vs. week 6: P = 0.209; week 6 vs. week 9: P = 0.007) and spent less time on other behaviors. Pigs' nonaggressive behavior and production parameters were compared with aggression using generalized linear mixed models. The time pigs spent on nonaggressive behaviors was negatively related to aggression (P < 0.045) with few exceptions. Initiated aggression after introduction was negatively related to loin muscle area (P = 0.003). These results show how finishing pigs spend their time in commercial facilities and indicate that behavior continues to change for up to 9 wk after introduction into a new social group. Efforts to reduce chronic levels of aggression should focus on promoting nonaggressive behaviors, such as exploration and movement, after the initial fighting that occurs immediately after introduction has waned, and should be implemented for up to 9 wk after introduction into new social groups.

Estimation of indirect social genetic effects for skin lesion count in group-housed pigs by quantifying behavioral interactions1.

Mixing of pigs into new social groups commonly induces aggressive interactions that result in skin lesions on the body of the animals. The relationship between skin lesions and aggressive behavioral interactions in group-housed pigs can be analyzed within the framework of social genetic effects (SGE). This study incorporates the quantification of aggressive interactions between pairs of animals in the modeling of SGE for skin lesions in different regions of the body in growing pigs. The dataset included 792 pigs housed in 59 pens. Skin lesions in the anterior, central, and caudal regions of the body were counted 24 h after pig mixing. Animals were video-recorded for 9 h postmixing and trained observers recorded the type and duration of aggressive interactions between pairs of animals. The number of seconds that pairs of pigs spent engaged in reciprocal fights and unilateral attack behaviors were used to parametrize the intensity of social interactions (ISI). Three types of models were fitted: direct genetic additive model (DGE), traditional social genetic effect model (TSGE) assuming uniform interactions between dyads, and an intensity-based social genetic effect model (ISGE) that used ISI to parameterize SGE. All models included fixed effects of sex, replicate, lesion scorer, weight at mixing, premixing lesion count, and the total time that the animal spent engaged in aggressive interactions (reciprocal fights and unilateral attack behaviors) as a covariate; a random effect of pen; and a random direct genetic effect. The ISGE models recovered more direct genetic variance than DGE and TSGE, and the estimated heritabilities (h^D2) were highest for all traits (P < 0.01) for the ISGE with ISI parametrized with unilateral attack behavior. The TSGE produced estimates that did not differ significantly from DGE (P > 0.5). Incorporating the ISI into ISGE, even in a small dataset, allowed separate estimation of the genetic parameters for direct and SGE, as well as the genetic correlation between direct and SGE (rs), which was positive for all lesion traits. The estimates from ISGE suggest that if behavioral observations are available, selection incorporating SGE may reduce the consequences of aggressive behaviors after mixing pigs.