Nuances related to the milieu of the gastrointestinal tract have led to investigations of environmental (or extrinsic) factors, like feed sources and fluid intake, and their influences on the gut microbiome in research animals. Water is typically provided to laboratory mice either by reusable water bottle (RWB), housing rack automatic water (RAW) delivery, or single-use disposable plastic pouch (DPP). In this study, the influence of differing water delivery methods on gut microbiome stability was evaluated in immunocompetent (n = 36 B6; 18 male [M]:18 female [F]) and immunocompromised (n = 36 NOG; 18 M:18 F) strains of mice. Mice were housed on a single IVC rack in sex-specific groups and provided with autoclaved caging and bedding, irradiated feed, and chlorinated, reverse-osmosis water provided by one of 3 delivery methods (8 cages per method). Access to the room was restricted to select personnel to conduct animal care and sample collection tasks. Fecal pellets (n = 2) were collected from each animal every other week, and water samples were collected weekly for analysis. Over the course of the study, bacteria were detected in 11% of the RWB samples (7 of 63) and 4% of the RAW samples (1 of 25). DPP samples were consistently free of bacterial contamination. Shotgun metagenomics and statistical analyses revealed overt shifts in gut microbiota in the majority of mice throughout the study (21 of 25 cages). Histologic examinations of organs from representative clinically normal study mice (n = 12) were unremarkable. With minimal exceptions, microbiome shifts were statistically significant across cage cohorts, despite attempts to control experimental variables. This study is the first to demonstrate that the water delivery method does not impart a significant influence on gut microbiota stability in research rodents and highlights the need to document water type, treatment, and delivery method as extrinsic factors in reporting animal studies.

Advancing scientific discovery requires investigators to embrace research practices that increase transparency and disclosure about materials, methods, and outcomes. Several research advocacy and funding organizations have produced guidelines and recommended practices to enhance reproducibility through detailed and rigorous research approaches; however, confusion around vocabulary terms and a lack of adoption of suggested practices have stymied successful implementation. Although reproducibility of research findings cannot be guaranteed due to extensive inherent variables in attempts at experimental repetition, the scientific community can advocate for generalizability in the application of data outcomes to ensure a broad and effective impact on the comparison of animals to translation within human research. This report reviews suggestions, based upon work with National Institutes of Health advisory groups, for improving rigor and transparency in animal research through aspects of experimental design, statistical assessment, and reporting factors to advocate for generalizability in the application of comparative outcomes between animals and humans.

The ability to apply findings from animal studies efficiently and effectively is predicated on an understanding of biology and pathobiology, how that biology relates to the human systems being modeled, and how the studies are conducted and reported. This overview discusses various factors in research within the animal environment (referred to as extrinsic factors) that the NIH now expects to be documented to foster replicability in science and expand interpretations of study outcomes. Specifically, an important extrinsic factor in research with animals is that of individual personnel who perform handling practices, participate in research interactions, and share an overall presence in the housing facility with animals, all of which can confound reproducibility efforts in biomedical science. An improved understanding of the influences and behaviors of animal research personnel on animal responses is critical with regard to research results and the interpretation of data collected from animal models of biomedical disease.

An essential aspect of animal resource programs is the storage and provision of food for a variety of species. Environmental parameters for feed storage conditions (temperature less than 70 °F; relative humidity less than 50%) are recommended in the Guide for the Care and Use of Laboratory Animals, along with aspects of nutrition such as palatability, vermin-control measures, diet quality, and integrity of feed bags. After receiving a suggestion for improvement for environmental conditions in 2 feed storage locations during an AAALAC accreditation visit, we hypothesized that the packaging of contemporary rodent feed could sustain wider environmental variations in temperature and humidity without adverse impact on integrity and palatability. This study evaluated representative feed storage sites across campus buildings to capture the variation in environmental conditions that are inherent to large and diverse animal care programs. Each test storage location held 2 identical bags of feed (same type, lot, and expiration date) that were stored from June to September of 2021; some aspects of the project were repeated during summer 2022 with a similar rodent feed. Baseline nutrients were analyzed from feed samples collected at time 0 (control) and again after 1 and 3mo of storage. The overall nutritional values measured in feed at the end of the study were not significantly different from control values, regardless of test site and variation in environmental parameters. Retinol (as a measure of Vitamin A) was the only component that decreased significantly; however, final retinol levels were consistently above those necessary for appropriate nutrition for mice. Our animal care program stakeholders were briefed on the outcomes of this study with the intent to verify at future AAALAC site visits that our storage conditions are adequate for maintaining the nutritional quality of packaged rodent feed.

This chapter provides general periprocedural suggestions for rodent, nonrodent larger species and agricultural species used in biomedical, agricultural, and production research.

Surgical procedures are commonly performed using mice but can have major effects on their core body temperature, including development of hypothermia. In this study, we evaluated active perioperative warming with and without surgical draping with adherent plastic wrap to refine practices, improve animal welfare, and optimize research experiments. Mice were randomized into treatment groups (n = 6; 8 CD1 mice per group). Treatments included placement within a small-animal forced-air incubator at 38 ° C for 30 min before surgery (Pre), after surgery (Post), or before and after surgery (Both). To explore the effect of surgical draping, one group received incubator warming before and after surgery in addition to surgical draping (Both/ Drape), whereas another group received surgical draping only without incubator warming (Control/Drape). The final group of mice received neither warming nor draping (Control). Subcutaneous temperature transponders were placed in all mice. Approximately 5 d after transponder placement, mice were anesthetized with ketamine-xylazine and underwent laparotomy. Subcutaneous body temperatures were collected perioperatively from transponders, and rectal temperatures were taken every minute during surgery. For recovery from anesthesia, mice were placed either in a standard cage on a warm water blanket set to 38 °C (100.4 °F) or in the incubator. Subcutaneous body temperatures were significantly higher in mice prewarmed for 30 min (Pre, Both, Both/Drape) as compared with mice that were not prewarmed. Anesthetic recovery times were significantly longer for mice placed in the incubator (Pre, Post, Both, Both/Drape) than for those that did not receive incubator warming (Control, Control/Drape). Mean intraoperative rectal temperatures of Both/Drape mice tended to be greater than those of mice in the Both group, suggesting a warming benefit of surgical draping. Using a forced air incubator and adherent plastic draping mitigated body temperature loss in mice during both surgery and postoperative recovery.

Rodents are frequently used for models that require surgical procedures. At our institution, laboratory rats are increasingly preferred for investigations of neurologic disorders, cardiovascular interventions, and assessment and treatment of addictive and depressive behaviors. For these types of studies, surgical preparations of the head and neck areas are necessary for catheterization and instrumentation. Based upon our former work in laboratory mice, we sought to improve rat surgery outcomes and confirm the efficacy of a waterless alcohol-based (WAB) antiseptic for skin disinfection prior to incision. In addition, we wanted to investigate whether active warming efforts improved perioperative body temperatures for rats to aid in return to consciousness. Prior to cranial surgical incision and placement in stereotactic equipment, rats were assessed after skin preparation with WAB and after thermal interventions, including prewarming cages for 30 min before anesthesia and delivery of warmed fluid (NaCl) supplementation. Core temperatures were recorded and aerobic culture swabs collected from surgical sites at multiple time points. As previously shown in mice, bacterial counts in rats were effectively diminished by WAB agents. Assessment of intraoperative body temperature trajectories did not identify appreciable differences between control rats and rats that were exposed to prewarming or warmed fluid supplementation or both. However, heavier male rats recovered more rapidly from isoflurane anesthesia than did lighter male and female rats. Although these thermal support measures did not significantly improve anesthetic recovery times in rats, animals warmed for 30 min trended toward a faster return to righting reflex after exposure to isoflurane. These findings confirm that WAB antiseptic is an acceptable option for skin preparation in rats and suggest that continued evaluation of thermal interventions remains of interest for improved outcomes in rat surgery.

Introduction:During the COVID-19 pandemic, health care systems and safety providers have faced an unprecedented challenge of limited access to personal protective equipment (PPE) to conduct patient and public care. In federal emergencies, reuse of PPE after disinfection can occur by processes, like vaporized hydrogen peroxide (VHP), recommended by the Centers for Disease and Control and Prevention. We identified a vacant animal holding facility at our institution to repurpose into a regional VHP decontamination center.Methods:The facility is a multiroom, 20 000 ft2 building with control of HVAC to adjust to VHP conditional requirements. H2O2 was delivered to rooms using robotic HaloFoggers, dispersing H2O2 vapor and increasingly concentrated microdroplets as a fog for a timed period based on cubic footage of rooms.Results:Fogging cycles eliminated 6-log Geobacillus stearothermophilus up to 7 days postcycle. Functional efficacy of treated N95s was confirmed by fit tests of institutional personnel. Signage, process flow mapping, and training materials facilitated ease of workflow and adherence to safety expectations within the building.Discussion and Conclusion:Our study determined that a variety of N95 respirator types and sizes were able to be cleared of potential bacterial and viral agents using VHP in a controlled fog/dwell/exhaust cycle. This repurposed animal facility has the capacity to decontaminate up to 6700 respirators daily, which will address the predicted surge of COVID-19 cases in the state, and ultimately allow each respirator to be reused multiple times. There is no other public site in the region with our capacity to offset the continued supply chain issues for PPE needs.

The authors discuss approaches to bolster investigator engagement, inviting investigators to be partners within the Animal Care Program. Regulatory burden in animal research endeavors continues to be reviewed and critiqued; therefore, this article intends to encourage Animal Care Programs to promote transparency and incorporation of unique educational training initiatives to tailor and focus compliance efforts across research programs. Borrowing from concepts of patient engagement, adherence, and enrollment efforts within the medical profession, it is likely that gains in trust, understanding, and communication between stakeholders within animal programs can be achieved without excessive efforts to alter existing approaches. Institutions will continue to be challenged to balance animal welfare expectations with promotion of research missions. This article provides a framework for somewhat radical ideas, including the use of collaborative orientations, assistance with self-evaluations, timely self-reporting, and meaningful and directed trainings, that are all aimed to resonate in contemporary animal care programs and foster investigator engagement in ongoing compliance efforts.

Review of the use of nonexperimental xenobiotics in terrestrial animal models and the potential unintended consequences of these compounds, including drug-related side effects and adverse reactions.

Preparing the skin of rodents for surgery often involves multiple applications of antiseptic agents. However, fewer applications may achieve the same antiseptic outcome. We evaluated the antimicrobial efficacy and effects on intraoperative body temperature of various surgical scrub agents, including novel waterless alcohol-based (WAB) options. Prior to ventral laparotomy, female C57BL/6 mice were treated with 0.9% saline (control); 70% ethanol; 10% povidone-iodine alternated with saline or 70% ethanol; 2% chlorhexidine digluconate alternated with saline or 70% ethanol; or 1 of 3 WAB products-commercial surgical scrub A, commercial surgical scrub B, or a common commercial hand sanitizer. Core temperatures were recorded, and aerobic culture swabs were collected from the surgical site at multiple time points. Intraoperative temperature trajectories for animals treated with scrub B, 10% povidone-iodine with saline, or hand sanitizer did not differ from saline (control). Temperature trajectories of mice treated with other scrub agents did differ significantly from saline. Bacteria were not detected at the operative site after 3 scrubs of 70% ethanol or 10% povidone-iodine alternated with ethanol, 2 scrubs of scrub A or B, 1 scrub of hand sanitizer, and both 1 and 3 scrubs of 2% chlorhexidine alternated with ethanol. Scrub B and 2% chlorhexidine-ethanol demonstrated prolonged antibacterial efficacy. Histology of corresponding haired skin sections revealed no differences in postoperative healing between groups, and no postoperative infections occurred. These results indicate that various novel WAB disinfectants, particularly scrub B (61% ethanol and 1% chlorhexidine gluconate), mitigate intraoperative temperature effects associated with several traditional agents and combinations. Furthermore, reduction of skin bacterial load without adverse effects on healing was seen with fewer than triplicate applications of most tested agents. Ultimately effective skin preparation can be achieved by using only 1 or 2 applications of scrub, thus rendering the triplicate skin-prep method unnecessary in laboratory mice.

To best promote animal wellbeing and the efficacy of biomedical models, scientific, husbandry, and veterinary professionals must consider the mechanisms, influences, and outcomes of rodent thermoregulation in contemporary research environments. Over the last 2 decades, numerous studies have shown that laboratory mice and rats prefer temperatures that are several degrees warmer than the environments in which they typically are housed within biomedical facilities. Physiologic changes to rodents that are cage-housed under standard temperatures (20 to 26 °C) are attributed to 'cold stress' and include alterations in metabolism, cardiovascular parameters, respiration, and immunologic function. This review article describes common behavioral and physiologic adaptations of laboratory mice and rats to cold stress within modern vivaria, with emphasis on environmental enrichment and effects of anesthesia and procedural support efforts. In addition, potential interventions and outcomes for rodents are presented, relative to the importance of repeating and reproducing experiments involving laboratory rodent research models of human disease.

Laboratory mice (Mus musculus) are prone to develop hypothermia during anesthesia for surgery, thus potentially impeding anesthetic recovery, wound healing, and future health. The core body temperatures of isoflurane-anesthetized mice are influenced by the choice of supplemental heat sources; however, the contribution of various surgical scrubs on the body temperatures of mice under gas anesthesia has not been assessed. We sought to quantify the effect of using alcohol (70% isopropyl alcohol [IPA]) compared with saline to rinse away surgical scrub on the progression of hypothermia in anesthetized mice (n = 47). IPA, room-temperature saline, or warmed saline (37 °C) was combined with povidone-iodine and then assessed for effects on core (rectal) and surface (infrared) temperatures. Agents were applied to a 2×2-cm shaved abdominal area of mice maintained on a water-recirculating blanket (at 38 °C) under isoflurane anesthesia (1.5% to 2.0% at 0.6 L/min) for 30 min. Although all scrub regimens significantly decreased body temperature at the time of application, treatments that included povidone-iodine led to the coldest core temperatures, which persisted while mice were anesthetized. Compared with room-temperature saline and when combined with povidone-iodine, warming of saline did not ameliorate heat loss. IPA alone demonstrated the most dramatic cooling of both surface and core readings at application but generated an unanticipated warming (rebound) phase during which body temperatures equilibrated with those of controls within minutes of application. Although alcohol is inappropriate as a stand-alone agent for surgical skin preparation, IPA is a viable alternative to saline-based rinses in this context, and its use should be encouraged within institutional guidance for rodent surgical procedures without concern for prolonged hypothermia in mice.

826 Introduction This position statement was developed by the American College of Laboratory Animal Medicine (ACLAM) to assist in the provision and evaluation of veterinary care for laboratory animals used, or intended for use, in research, teaching, testing or production. While this document aims to deliver independent guidance for animal programs, ACLAM recognizes that both regulatory and scientific sponsoring agencies such as the United States Department of Agriculture (USDA) and the Public Health Service of the United States Department of Health and Human Services (PHS/DHHS), through their respective missions, sanction the provision of adequate veterinary care; further elaboration on this concept can be found within the NRC Guide for the Care and Use of Laboratory Animals (2011). Additional animal care guidance is supplied by allied laboratory animal organizations, including the Association of Primate Veterinarians (APV), AAALAC International, the American Association for Laboratory Animal Science (AALAS) and the American Society of Laboratory Animal Practitioners (ASLAP). While these organizations may be founded in the United States, the shared principles regarding veterinary care are global in scope. Importantly, the professional judgment of a trained and experienced veterinarian is essential in the delivery of adequate veterinary care to laboratory animals.

Intraperitoneal injectable anesthetics are often used to achieve surgical anesthesia in laboratory mice. Because bolus redosing of injectable anesthetics can cause unacceptably high mortality, we evaluated intraperitoneal continuous-rate infusion (CRI) of ketamine with or without xylazine for maintaining surgical anesthesia for an extended period of time. Anesthesia was induced in male C57BL/6J mice by using ketamine (80 mg/kg) and xylazine (8 mg/kg) without or with acepromazine at 0.1 mg/kg or 0.5 mg/kg. At 10 min after induction, CRI for 90 min was initiated and comprised 25%, 50%, or 100% of the initial ketamine dose per hour or 50% of the initial doses of both ketamine and xylazine. Anesthetic regimens were compared on the basis of animal immobility, continuous surgical depth of anesthesia as determined by the absence of a pedal withdrawal reflex, and mortality. Consistent with previous studies, the response to anesthetics was highly variable. Regimens that provided the longest continuous surgical plane of anesthesia with minimal mortality were ketamine-xylazine-acepromazine (0.1 mg/kg) with CRI of 100% of the initial ketamine dose and ketamine-xylazine-acepromazine (0.5 mg/kg) with CRI of 50% of the initial ketamine and xylazine doses. In addition, heart rate and respiratory rate did not increase consistently in response to a noxious stimulus during CRI anesthesia, even when mice exhibited a positive pedal withdrawal reflex, suggesting that these parameters are unreliable indicators of anesthetic depth during ketamine-xylazine anesthesia in mice. We conclude that intraperitoneal CRI anesthesia in mice prolongs injectable anesthesia more consistently and with lower mortality than does bolus redosing.

Retroorbital blood collection is a common technique in laboratory rodents due to the ease with which it can be performed and the sample volumes obtained for subsequent blood analyses. However, its use has been discouraged recently due to aesthetic discomfort and anecdotal reports of potential for ocular injury during blood collection. We hypothesized that a single standardized session of in-person training would be sufficient to learn the appropriate technique and minimize the likelihood for adverse outcomes. Experienced instructors (n = 2) conducted hands-on training classes to teach novice personnel (n = 40) to perform this procedure. Blood was collected from anesthetized mice (n = 40) via a capillary tube first placed at the medial canthus of the right eye and then advanced into the retroorbital space; the left retroorbital spaces served as unmanipulated controls. For comparison, the experienced instructors similarly collected blood from 40 additional mice. The tube could be inserted only once in each mouse, with the goal of obtaining 50 to 100 μL blood. Overall, 79 of 80 mice (98.8%) showed normal body condition, posture, and behavior throughout the 14-d study. Thus, any clinical observation scores pertained specifically to ocular lesions, which occurred at least once after sampling in 43 (53.8%) of the mice. Clinical and histopathologic scores of mice after bleeding did not differ between experienced and novice personnel. We conclude that a coordinated hands-on training program can provide consistent and sufficient instruction for research personnel to conduct retroorbital blood collection with competence in anesthetized laboratory mice.