Intrauterine sperm injection for artificial insemination is difficult in cynomolgus macaques (Macaca fascicularis) and rhesus macaques (M. mulatta) due to the complex structure of the cervical canal, which differs from that of humans. Despite the availability of several artificial insemination methods for macaques, pregnancy rates are inconsistent, and details regarding ovulation are unclear, thus warranting more effective methods. Therefore, we developed an effective, ultrasound-guided, transabdominal intrauterine artificial insemination method for cynomolgus macaques that involves timing sperm injection to coincide with the periovulation phase estimated according to rapid hormone measurement. We performed our intrauterine artificial insemination on 6 female macaques; 4 of the 5 animals that were predicted to have ovulated soon after insemination became pregnant, whereas the 1 macaque that was predicted not to have ovulated did not. Furthermore, we saw no evidence of injury, such as a conspicuous needle hole or bleeding on the surface of or inside the uterus, nor did our method result in any abnormalities in the mothers or their offspring. Thus, our ultrasound-guided, transabdominal, intrauterine artificial insemination method is rapid, safe, and effective in cynomolgus macaques.

Little research has been conducted to examine the influence of various methods of providing nest materials—such as dispersing them, providing them as single units, or clustering them—on the behavior and welfare of group-housed mice. In this study, 6 wk-old C57BL/6NCrl mice were housed 3 per cage and randomized into 1 of 3 nest-material groups: 1) one facial tissue per cage (control; female mice, 3 cages; male mice, 3 cages); 2) an 8-g 'puck' of compressed nesting material and a facial tissue (females, 3 cages; males, 3 cages); or 3) 8 g of dispersed paper strips and a facial tissue (females, 3 cages; males, 3 cages). Mouse behavior (agonistic, stereotypic, nesting), physical examination data, and nest scores were evaluated over 16 d. The results showed that mice in the puck and control groups spent more time manipulating nest materials after cage changes than did mice in the paper-strip group. Average nest scores were highest in the paper-strip group compared with controls and puck cages. Female cages with pucks showed no barbering, whereas all other female mice cages demonstrated barbering. Overall, nest pucks may provide a time-consuming activity for mice and may help protect female C57BL/6 mice from barbering. However, more research is needed to replicate and expand these study results.

The quality of research animal welfare is undeniably linked to the quality of scientific results generated from the animals. Although mice are the most commonly used mammalian species in biomedical research, little information is available about what factors should be considered to promote future progress. To address this issue, the Animal Welfare Committee of the American Society of Laboratory Animal Practitioners (ASLAP) surveyed laboratory animal veterinarians to obtain their opinions about the welfare of mice and to consider the roles of 5 factors that significantly affect animal welfare in biomedical research: husbandry, clinical care, experimental use, regulatory oversight, and training. The survey revealed that 95% of veterinarians scored mouse welfare as acceptable to excellent, although areas for improvement remain. These areas include: 1) training of researchers performing experimental procedures; 2) the frequency of monitoring mice likely to experience pain and distress due to experimental manipulation; 3) inclusion of the institutional veterinary staff in the monitoring of mice likely to experience pain and distress; 4) continued improvement in the environmental enrichment provided to mice; 5) the ability of the IACUC to ensure that instances of noncompliance are fully addressed in order to prevent reoccurrence both within laboratories and among other research groups at the institution; and 6) reliance on non-veterinarians to perform examinations, diagnose disease, and prescribe the treatment of sick or injured mice.

Spironucleus muris is an intestinal protozoal pathogen that can infect various species of rodents. The infection can have a wide range of clinical presentations, from no signs of disease to death. In addition, this pathogen can adversely affect research results, especially immunologic and gastrointestinal studies. For these reasons, institutions may exclude Spironucleus muris. However, despite rigorous efforts to keep this pathogen out, it can be common in rodent colonies. The current recommended approach to eradicating this pathogen is by testing and culling positive animals. A similar organism, Giardia muris, has been effectively eliminated by using chemotherapeutics. Therefore, the objective of this study was to determine whether S. muris is also susceptible to chemotherapeutics. Naturally infected mice were randomized to treatment groups after confirmation of positive infection via PCR. Mice received either metronidazole, fenbendazole, a combination of metronidazole-fenbendazole, or acidified water (control) treatments for a period of 4 wk. Each week fecal testing of S. muris was performed via PCR to evaluate the effectiveness of the treatments. At the end of the 4 wk period, mice were euthanized via CO₂ inhalation and segments of the proximal gastrointestinal tract were submitted for histopathologic analysis. Treatment with metronidazole or fenbendazole alone or in combination, failed to clear S. muris infected mice. After 4 wk of treatment, none of the mice given fenbendazole via sucralose medicated gel were positive by either PCR or histopathology; however, this finding is most likely due to intermittent shedding rather than chemotherapeutic success. Therefore, the recommendation remains to test-and-cull or rederive mice as necessary to eliminate S. muris from laboratory animal facilities.

Sentinel animals remain a common means of evaluating rodent health in research colonies. An evaluation of our sentinel program revealed that using Crl:CD1(ICR)-Elite (CD1-E) mice was expensive, occasionally disrupted by limited supply, and minimally responsive to the 3Rs. This evaluation prompted us to explore the use of CRL:NU-Foxn1^nu/+ (Het-nude) mice as soiled-bedding sentinel (SBS) animals. Het-nude mice are a byproduct of breeding outbred athymic nude mice and are reared in isolators, with similar health status as CD1-E. Het-nude mice have a thymus, but may have smaller thymic size and fewer bone marrow stem cells than do wildtype controls, suggesting that Het-nude mice might not be immunologically normal. This study compared the antibody titer and seroconversion kinetics of Het-nude and CD1-E SBS to murine norovirus (MNV) and mouse hepatitis virus (MHV). Het-nude and CD1-E female SBS (n = 22 mice of each stock) were housed continuously on soiled bedding collected from MNV-positive or MNV- and MHV-positive colonies at cage changes. Blood was collected for serology at 3, 9 and 12 to 19 wk after the start of soiled bedding exposure. Antibody titers to MNV or MHV did not differ significantly between Het-nude and CD1-E mice. A significant relationship was found between weeks of exposure and titer levels with an increase in titer over the testing period. This study supports the possible use of Het-nude mice as SBS, given that their antibody responses to MNV and MHV are equivalent to those of CD1-E mice.

Recent studies have evaluated alternatives to the use of live animals in colony health monitoring. Currently, an alternative method that is suitable for all rack types and that has been verified to detect the infectious agents most commonly excluded from mouse colonies is unavailable. We compared the use of filter paper placed on the inside floor of mouse cages to the traditional use of sentinel mice in the detection of several prevalent murine pathogens including mouse hepatitis virus (MHV), murine norovirus (MNV), minute virus of mice (MVM), mouse parvovirus (MPV), Theiler murine encephalomyelitis virus (TMEV), Helicobacter spp., Syphacia obvelata, and Aspiculuris tetraptera. Experimental groups comprised 7 cages containing either 2 pieces of filter paper on the cage floor or 2 ICR sentinel mice. Soiled bedding from pet-store mice was transferred to the experimental cages weekly for 8 wk. At 1 and 2 mo after bedding transfer, the filter papers were evaluated by PCR and sentinel mice were tested by serology and fecal PCR. Filter papers detected all pathogens as effectively (MHV, MNV, MPV, MVM, TMEV S. obvelata, and A. tetraptera) or more effectively (Helicobacter spp.) than sentinel mice at both time points. Filter papers more readily detected pathogens with a high copy number per RT-PCR analysis than a low copy number. Helicobacter spp. were not detected by sentinel mice at either time point. These results indicate that the use of filter paper placed on the interior floor of empty mouse cages and exposed to soiled bedding is efficient in detecting bacteria, endoparasites, and most of the common mouse viruses included in an animal health monitoring program.

West Nile virus (WNV) was first detected in Florida in July 2001, with 404 human cases reported to the Centers for Disease Control and Prevention as of February 2020. The subtropical climate of Florida is ideal for the mosquitoes that transmit WNV. We investigated the WNV seroprevalence in 3 NHP species housed outdoors at The Mannheimer Foundation in South Florida. From January to December 2016, 520 3 to 30 y old NHP were sampled at our 2 closed sites in Homestead and LaBelle: 200 rhesus macaques (Macaca mulatta), 212 cynomolgus macaques (Macaca fascicularis), and 108 hamadryas baboons (Papio hamadryas hamadryas). The presence of WNV IgG antibodies in these animals was determined by serum neutralization assays, which found a total seroprevalence of 14%. Seroprevalence was significantly higher in the baboons (29%) than the rhesus (11%) and cynomolgus (9%) macaques. The probability of seropositivity significantly increased with age, but sex and site did not significantly affect seroprevalence. The frequency of WNV seropositivity detected in these outdoor-housed NHP suggests that screening for WNV and other vector-borne diseases may be necessary prior to experimental use, particularly for infectious disease studies in which viremia or viral antibodies could confound results, and especially for populations housed outdoors in warm, wet climates. As no seropositive subjects demonstrated clinical signs of WNV and WNV exposure did not appear to significantly impact colony health, routine testing is likely unnecessary for most NHP colonies. However, WNV infection should still be considered as a differential diagnosis for any NHP presenting with nonspecific neurologic signs. Mosquito abatement plans and vigilant sanitation practices to further decrease mosquito and avian interaction with research NHP should also be considered.

The sustained-release formulation of meloxicam (MSR) is a compounded NSAID that may provide pain relief for as long as 72 h after administration. MSR injection-site skin reactions have occurred in several species but have not previously been observed in mice. We investigated the development and progression of localized skin reactions after a single injection of MSR in Crl:CD1(ICR), C57BL/6J, and BALB/cJ mice. Each mouse received a subcutaneous injection of MSR (n = 60), standard-formulation meloxicam (MEL; n = 24) or saline (control; SC; n = 24) and was scored daily according to a 5-point system for erythema and mass characteristics. Mice were euthanized at either 7 or 14 d after injection and underwent postmortem analysis. MSR-treated mice had more erythematous and mass reactions than did MEL and SC mice. Mass lesions developed in 49 MSR mice (82%; 95% CI, 70% to 90%), 5 MEL animals (21%; 95% CI, 7% to 42%), and 1 SC mouse. MSR-treated BALB/cJ developed erythematous lesions less frequently than similarly treated Crl:CD1(ICR) or C57BL/6J. Lesions often were ventrolateral to the injection site. The median times to the appearance of mass and erythematous lesions were 2 d and 3 d, respectively. Histologically, the erythematous and mass reactions correlated with necrotizing to pyogranulomatous injection-site panniculitis. Inflammation severity scores at 7 and 14 d after injection were greater in the MSR-treated group than the other 2 groups. No strain- or sex-associated differences emerged except that inflammation severity scores at day 14 were higher in Crl:CD1(ICR) females than males. The character of the inflammatory response in MSR-treated mice did not differ between 7 and 14 d after injection, indicating that MSR-induced inflammation is slow to resolve. The ventral migration and delayed onset of MSR injection-site reactions could result in their being attributed to another cause or not being identified. Researchers and clinicians should be aware of the potential for slowly resolving injection-site reactions with MSR.

Compared with intravenous and intramuscular methods, intranasal administration of sedatives is a less invasive and nonpainful technique. In this prospective, randomized, crossover study, we evaluated the sedative characteristics of 2 doses (1 and 2 mg/kg) of alfaxalone administered intranasally to 7 adult Yucatan swine. We compared sedation scores before and after administration of alfaxalone and between groups by using a composite sedation scoring system (range, 0 to 12, with 12 being the highest level of sedation)). Pigs were randomly assigned to receive 2 doses of intranasal alfaxalone (1 mg/kg [A1]); 2 mg/kg [A2]) as 2 separate events in a crossover design with a 60-d washout period. Categories scored were posture, palpebral droop, uninhibited behavior, drowsiness, and acceptance of anesthetic facemask. Sedation scores were collected before sedation was administered and then every 3 min for 30 min afterward. Instilled volumes (mean ± 1 SD) were 5.7 ± 0.5 and 11.3 ± 0.8 mL for A1 and A2, respectively. Both alfaxalone doses produced significant increases in sedation scores compared with baseline. Median sedation scores for A1 (6; range, 4–12) were not different from those for A2 (6; range, 6 to 12). Intranasal administration of alfaxalone as the sole sedative agent increased sedation scores from baseline, achieving peak sedation at 6 to 9 min after instillation of A2. However, sedation scores were similar between the 2 groups, and neither dose produced sufficient sedation to facilitate handling or the performance of any clinical procedures. Given the concentration of alfaxalone solution currently available, volume is the major limiting factor regarding testing higher doses of this drug for its use as a sole sedative agent in swine.

Buprenorphine is an essential component of analgesic protocols in common marmosets (Callithrix jacchus). The use of buprenorphine HCl (BUP) and sustained-release buprenorphine (BSR) formulations has become commonplace in this species, but the pharmacokinetics have not been evaluated. Healthy adult (age, 2.4 to 6.8 y; 6 female and 6 male) common marmosets were enrolled in this study to determine the pharmacokinetic parameters, plasma concentration–time curves, and any apparent adverse effects of these compounds. Equal numbers of each sex were randomly assigned to receive BUP (0.02 mg/kg IM) or BSR (0.2 mg/kg SC), resulting in peak plasma concentrations (mean ± 1 SD) of 15.2 ± 8.1 and 2.8 ± 1.2 ng/mL, terminal phase t_1/2 of 2.2 ± 1.0 and 32.6 ± 9.6 h, and AUC_0-last of 16.1 ± 3.7 and 98.6 ± 42.7 ng × h/mL. The plasma concentrations of buprenorphine exceeded the proposed minimal therapeutic threshold (0.1 ng/mL) at 5 and 15 min after BUP and BSR administration, showing that both compounds are rapid-acting, and remained above that threshold through the final time points of 8 and 72 h. Extrapolation of the terminal elimination phase of the mean concentration–time curves was used to develop the clinical dosing frequencies of 6 to 8 h for BUP and 3.0 to 3.5 d for BSR. Some adverse effects were observed after the administration of BUP to common marmosets in this study, thus mandating judicious use in clinical practice. BSR provided a safe, long-acting option for analgesia and therefore can be used to refine analgesic protocols in this species.

General anesthesia induces many systemic effects, including thermoregulatory impairment and subsequent perioperative hypothermia. Due to the animals' small size, monitoring and maintaining body temperatures in laboratory rodents during anesthesia is important for successful surgical outcomes and prompt anesthetic recovery. Draping materials have the potential to aid in thermal support during surgical anesthesia. In this study, rectal and surface (infrared) temperatures were measured in C57BL/6 mice under isoflurane anesthesia every 5 min for the duration of a 35-min sham surgery. In addition to placement on a circulating water bath, mice (n = 6/group) were draped with commercial cling film (CF; Press'n Seal, Glad, Oakland, CA), a conventional paper drape (PD), or no drape (ND) during surgery. Results demonstrated that CF-draped animals had significantly higher rectal temperatures than nondraped animals. Furthermore, surface temperatures of CF-draped mice were considerably higher than those of both paper-draped and undraped animals. The data indicate that cling film is an effective material to help minimize hypothermia in mice and potentially in other laboratory rodents requiring general anesthesia.

Mouse handling and restraint affect behavior, physiology, and animal welfare, yet little information is available on how various mouse restraint methods affect cardiovascular parameters. We validated the use of a smartphone-based ECG sys- tem in mice by performing simultaneous smartphone and telemetry ECG recordings in conscious, restrained mice and in anesthetized mice. We observed that mice held in standard immobilizing restraint ("scruffing") experienced severe bradycardia. Mice of both sexes and 4 different strains (BALB/cJ, C57BL/6J, DBA/2J, and FVB/nJ) were restrained by 3 handlers using 3 different restraint methods: light restraint; 3-finger restraint, which creates a dorsal transverse fold of skin; and the standard immobilizing restraint, which creates a dorsal longitudinal fold of skin that results in a crease on the ventral neck. Regardless of the handler, immobilizing restraint, but not 3-finger restraint, produced severe bradycardia with irregular rhythm in all 4 strains and both sexes, with an average decrease in heart rate of 31%, or 211 bpm, and a maximal decrease of 79%, or 542 bpm. When evaluated using telemetry, immobilizing restraint produced severe arrhythmias such as junctional and ventricular escape rhythms, and second- and third-degree atrioventricular block. Sinus pauses were observed for an average of 4 min, but up to 6.8 min after release from immobilizing restraint. Atropine administration to C57BL/6J mice attenuated immobilizing restraint-induced bradycardia, supporting the hypothesis that pressure on cervical baroreceptors during stretching of the neck skin results in a vagally-mediated reflex bradycardia. Because of these profound cardiovascular effects, we recommend using the light or 3-finger restraint and avoiding or minimizing the use of immobilization restraint while handling mice.

Rats commonly undergo surgery for research purposes. However, the effects of different methods of hair removal on wound healing and surgical site infections (SSI) in rats has not been evaluated. The current study evaluated 2 hair removal methods, clipping with an electric clipper and using a depilatory agent, and their effect on wound healing and SSI. Swabs for bacterial culture were obtained on Day 0 just after hair removal, after aseptic skin preparation, and on Days 1 and 3 before conducting skin biopsies to assess bacterial load and recolonization. Full-thickness punch biopsies were taken for histopathologic evaluation on Days 0, 1, 3, 7, and 10. The surgical incisions were assigned an ASEPSIS score on Days 1 and 3. The data revealed that the bacterial load was significantly higher with the depilatory method as compared with the clipper method, but only on Day 1. The histopathologic evaluation found no significant difference in wound healing between the 2 methods. Although the ASEPSIS score was significantly higher for the clipping method than for the depilatory method on Day 1, both techniques were equivalent by Day 3. We conclude that both hair removal methods are safe and efficacious components of aseptic technique in rats.

Due to their effective analgesic properties, opioids are worthy of consideration for pain management in rabbits. However, this class of drugs causes undesirable effects including reduced gastrointestinal (GI) motility, reduced fecal output, and delays GI transit times and thus increases the risk of GI stasis. The risk of stasis discourages the use of opioids in rabbits, which could affect animal welfare. Gastroprokinetic agents such as cisapride are effective in promoting gastric emptying in many species, but whether this effect occurs in rabbits is unknown. This study assessed the efficacy of cisapride when administered as a single agent and in combination with buprenorphine in rabbits; efficacy was assessed by measuring GI transit times, fecal output, body weight, and food and water intake. Female New Zealand White rabbits (n = 10) were studied in a crossover, randomized design and received either vehicle and buprenorphine, cisapride and saline, cisapride and buprenorphine, or vehicle and saline (control) every 8 h for 2 d. Rabbits were anesthetized and administered radio-opaque, barium-filled spheres via orogastric tube. Feces was assessed via radiography for detection of the barium-spheres to determine GI transit time. GI transit time was significantly longer in buprenorphine groups than in control groups, regardless of the use of cisapride. Fecal output and food and water intake were lower for buprenorphine groups than control groups. Cisapride did not significantly alter GI transit, fecal output, or food and water intake. In addition, treatment group did not significantly affect body weight. In conclusion, buprenorphine treatment (0.03 mg/kg TID) prolonged GI transit time and reduced fecal output and food and water consumption in rabbits. Coadministration of buprenorphine and cisapride (0.5 mg/kg) did not ameliorate these effects, and the administration of cisapride at this dose did not appear to affect GI motility in female rabbits.

Quantitative blood pressure measurement is a critical parameter for assessing cardiovascular health, monitoring physiologic status under anesthesia, and making clinical decisions. The placement of an arterial catheter is the most accurate way to measure blood pressure, but is invasive and perhaps uncomfortable for the patient, requires sedation or anesthesia, and is technically challenging for staff. Noninvasive devices and methods, including oscillometry, high-definition oscillometry, and Doppler flow can be used to measure blood pressure. However, the American College of Veterinary Internal Medicine guidelines state that blood pressure should be measured using devices that have been validated in the species of interest and under the circumstances in which the measurement is being made. The alternatives listed above have not been validated in this manner. The objective of this study was to compare indirect ultrasonic Doppler flow detection and oscillometric blood pressure measures with direct arterial measurement in 8 healthy, anesthetized African green monkeys using the methods of Bland-Altman to assess agreement. Our results indicated that neither Doppler flow nor oscillometry consistently estimates direct blood pressure measurements in anesthetized African green monkeys. In 6 female monkeys, which weighed less than the 2 male subjects, the indirect Doppler measurement more closely predicted direct mean arterial pressure, indicating Dop- pler flow may be useful for monitoring mean arterial pressure in small patients.