Interest in the genetic composition of cynomolgus macaques (Macaca fascicularis) has increased due to the rising demand for NHP models in human biomedical research. Significant genetic differences among regional populations of cynomolgus macaques can confound interpretations of research results because they do not solely reflect differences in experimental treatment effects. Therefore, the common origin of cynomolgus macaques used as research subjects should be verified by using region-specific genetic markers to minimize the influence of underlying genetic variation among animals selected as research subjects on phenotypes under study. We compared the effectiveness of 18 short tandem repeat (STR) markers with that of 83 single-nucleotide polymorphism (SNP) markers to differentiate the ancestry of cynomolgus macaques from 6 different populations (Cambodia, Sumatra, Mauritius, Singapore, and the islands of Luzon and Zamboanga in the Philippines). Genetic diversity indices such as allele numbers and expected heterozygosity based on SNP were lower and exhibited lower standard errors than those provided by STR, probably because, unlike STR, most SNP are biallelic and consequently exhibit maximal expected heterozygosity values of 0.50. However, the standard error of estimates of observed heterozygosity based on SNP was higher than that for STR, perhaps reflecting sampling errors. Only 27 SNP were required to match the resolving power of 17 STR to detect population structure, that is, 1.6 SNP:1 STR. Whereas STR only differentiated the Mauritian population from all other populations, SNP detected 4 genetically distinct groups (Cambodia, Singapore-Sumatra, Mauritius, and Zamboanga). SNP are poised to become as valuable as STR for understanding and detecting genetic structure among cynomolgus macaques. Although STR will remain an important tool for cynomolgus macaque population studies, SNP have the potential to become the mainstream marker type.

Components of bedding might interact with experimental treatments and affect the outcome of various experiments. Here we studied the biochemical effects of 3 rodent bedding materials that are commonly used in Egypt. Male and female rats and mice were assigned randomly into 4 single-sex and single-species groups (10 animals per group). Three types of bedding—rice straw, wheat straw, and pine wood shavings—were evaluated. After 4 wk, animals were euthanized, and biochemical parameters were measured. In male and female rats given wood shavings, serum ALT activity and malondialdehyde concentration increased whereas catalase activity decreased compared with levels in the wheat straw group. In contrast, ALT activity and malondialdehyde concentrations decreased but CAT activity increased in rats housed on rice straw compared with wheat straw. Serum AST and ALT activities increased in male and female mice exposed to rice straw, whereas the malondialdehyde concentration increased and catalase decreased in the wood shavings group relative to the wheat straw group. In mice exposed to wheat straw, AST and ALT activities and malondialdehyde concentrations decreased and CAT activity increased compared with the other groups. Because our results showed that exposure to wood shavings affects some biochemical parameters of rats and mice, we do not recommend its use as laboratory animal bedding. We consider that, of the materials tested, rice straw bedding is the best bedding material for rats, whereas wheat straw is best for mice.

Despite documented adverse effects, limits for rodent exposure to vibration in the laboratory animal facility have not been established. This study used female C57BL/6 mice to determine the frequencies of vibration at which mice were most sensitive to behavioral changes, the highest magnitude of vibration that would not cause behavioral changes, the behavioral changes that occur in response to vibration, and the extent to which mice habituate to vibration. Mice were exposed to frequencies of vibration between 20 and 190 Hz at accelerations of 0.05 to 1.0 m/s². Behavioral responses were videorecorded and subsequently scored. Mice showed the most behavioral responses at 1.0 m/s². At intermediate accelerations of 0.5 and 0.75 m/s², behavioral responses were most prevalent at frequencies of 70 to 100 Hz. In contrast, at an acceleration of 0.05 m/s², mice did not show any discernible behavioral response. Behavioral responses induced by the initiation of vibration were transient, generally lasting only 2 to 10 s. Behaviors in awake mice included abrupt freezing of motion, hunched posture, and surveying the cage environment. In mice that were asleep, responses consisted of lifting the head suddenly with or without prior shifting of body position. When exposed to multiple periods of vibration over a short time, responses seemed to decrease. In summary, mice were particularly sensitive to vibration between 70 to 100 Hz, did not respond to the slowest acceleration (0.05 m/s²), and exhibited transient responses at the initiation of vibration.

Commonly used in biomedical research, vervets (Chlorocebus aethiops) are omnivorous but primarily meet their vitamin A requirements from provitamin A carotenoids. Hypervitaminosis A has occurred in vervets that consume feed high in preformed vitamin A. We investigated the vitamin A status of vervets supplemented daily with various antioxidants derived from palm oil. Male vervets (n = 40) were placed for 23 wk on a high-fat diet (34.9% energy) containing 645 μ g retinol activity equivalents (RAE), with 515 μ g RAE from preformed vitamin A. Vervets were randomized to 5 treatments (duration, 20 mo): control; 100 mg d-α-tocopheryl acetate; 100 mg oil palm (Elaeis guineensis)-derived vitamin E; 50 mg oil palm-derived vitamin E + 50 mg carotenoid complex + unrestricted palm-derived water-soluble antioxidants; and 5) unrestricted water-soluble antioxidants. Livers (n = 38) were analyzed for vitamin A, α-retinol (α-vitamin A), and carotenoids. Median hepatic vitamin A and total carotenoid concentrations were 6.49 μ mol/g and 4.30 nmol/g, respectively. Compared with controls, vervets fed the carotenoid complex had higher hepatic vitamin A (11.9 ± 5.1 μ mol/g), α -vitamin A (1.3 ± 0.7 μ mol/g), α -carotene (11.5 ± 5.3 nmol/g), β-carotene (15.6 ± 8.6 nmol/g), and total carotenoids (28.1 ± 13.9 nmol/g) but lower lutein (0.66 ± 0.28 nmol/g) and zeaxanthin (0.24 ± 0.06 nmol). NHP may benefit from replacement of preformed vitamin A with carotenoids in feeds; however, bioconversion efficiency in these models should be investigated to determine optimal levels.

Facility-wide Corynebacterium bovis eradication was established using vaporized hydrogen peroxide (VHP) decontamination guided by C. bovis PCR surveillance. Prior attempts limited to culling PCR-positive mice and decontaminating affected rooms were ineffective in preventing recurrence. Because research aims often require trafficking to and use of procedural cores, a 12-mo facility-wide C. bovis PCR surveillance of 2064 specimens was performed and documented that, despite the presence of few clinically hyperkeratotic mice, 35% of the murine housing and use space was contaminated by C. bovis. The airways of IVC racks and air-handling units (AHU) provided a substantive niche for C. bovis survival, comparable to the primary enclosure, with 26% of murine and 22% of airway specimens PCR-positive for C. bovis. Equipment airway VHP sterilization in a 'flex room' required an 'active–closed' setting with the IVC rack connected to the AHU set to the VHP cycle, because 12% of specimens from 'static–open' VHP-exposed airways remained PCR-positive for C. bovis, whereas 0% of specimens from active–closed VHP exposures were positive. VHP decontamination of the 29,931-ft² facility was completed in 2 mo. C. bovis PCR testing of IVC exhaust plenums for 200 d in previously C. bovis-affected rooms confirmed that none of the 259 specimens tested were PCR-positive for the organism. Monthly surveillance identified a single recurrence during June 2017 (month 9), ensuring rapid culling of C. bovis PCR-positive mice and acute VHP decontamination of equipment and rooms. Molecular persistence of C. bovis was resolved in procedural and personnel areas, and no murine or housing specimens tested C. bovis PCR-positive during study months 11 and 12. Furthermore, since the conclusion of the 12-mo study, none of the 452 additional murine, cell biologic, environmental, and monthly equipment surveillance specimens tested were C. bovis PCR-positive, documenting an 11-mo period of facility-wide C. bovis eradication to date. Study invalidation due to C. bovis can be avoided through PCR surveillance for the organism, immediate culling of PCR-positive mice, and acute VHP decontamination of affected areas.

Testing sentinel animals exposed to soiled bedding from colony animals is the most common method used for health monitoring in rodent facilities. Although environmental sampling is being explored—and, in many cases, has been implemented—as an alternative, exhaust plenum sampling is not effective for all ventilated rack designs. This study evaluated PCR testing of filter paper from sentinel cages on ventilated racks. We hypothesized that testing filter paper from cages containing soiled bedding would be as effective as testing sentinel mice and that periodic shaking of cages would generate sufficient particulate movement to substitute for the presence of live animals. Three cages containing soiled bedding were maintained in each of 8 rooms; one cage contained 2 Cr:NIH(S) mice, one had no mice and was shaken twice weekly, and the remaining one had no mice and was left undisturbed. For 3 consecutive months, a piece of filter paper from the undersurface of the cage lid was tested monthly for adventitial agents and then replaced. A second piece remained on the cage undersurface for 3 mo. Fecal pellets and oral and fur swabs were collected from sentinel mice at months 1 and 3 and tested for the same agents. At month 3, serology was performed on the sentinel mice; feces and oral and fur swabs from colony animals were tested concurrently for comparison. Filter paper from cages without mice and shaken were at least as effective than all other methods in detecting the presence of endemic agents, including mouse norovirus, Helicobacter spp., Pasteurella pneumotropica, Entamoeba muris, and Spironucleus muris. For IVC systems where exhaust plenum testing is ineffective, PCR testing of IVC filter tops should be considered as an alternative to soiled bedding sentinels. Environmental sampling may provide increased reliability and reduce the number of rodents used for routine health surveillance.

Demodex musculi is a prostigmatid follicular mite that has rarely been reported in laboratory mice. Although prevalence of this species has not been assessed formally, we have found that many imported mouse strains from noncommercial sources harbor Demodex mites. To assess whether an acaricide can be used to eradicate this mite, infested immunocompromised mice were provided ivermectin-compounded (12 ppm) feed without restriction for 8 wk (n = 10), were treated topically with moxidectin and imidacloprid (MI; 3 and 13 mg/kg, respectively) weekly for 8 wk (n = 10), or remained untreated (n = 10). Mice were confirmed to be mite-infested before treatment and were tested after treatment by using fur plucks (FP), deep skin scrapes (DSS), and PCR analysis of fur swabs. In addition, the presence of mites was confirmed through skin biopsies at 2 study endpoints (1 wk [n = 5] and 12 wk [ n = 5] after treatment). Samples collected before treatment and from untreated mice were positive for D. musculi at all time points and by all test modalities. After treatment, all ivermectin-treated animals remained infested, whereas mice treated with MI were repeatedly negative by all test modalities. An additional shortened treatment trial revealed that 4 wk of MI (n = 7) was insufficient to eradicate mites. Neither treatment produced any evidence of adverse effects according to hematology, serum chemistry parameters, behavior, body weight, and histopathology. Of the 70 PCR assays performed in treated mice, 14 were positive when FP+DSS was negative. In 6 cases where PCR results were negative, 5 were positive by FP+DSS and a single sample was positive on skin biopsy. Although PCR analysis has a high detection rate for D. musculi, FP+DSS can further enhance the detection rate. In conclusion, topical MI administered for 8 consecutive weeks can safely eradicate D. musculi from an immunocompromised mouse strain.

Managing postoperative pain in rodents is an important part of any animal care and use program, and identifying an optimal analgesic plan for a surgical procedure is critical to providing for animal welfare. Opioids and NSAID are commonly used in rodents, but few studies have evaluated their efficacy in surgical models. The current study aimed to evaluate the therapeutic efficacy of clinically relevant doses of buprenorphine (2 formulations) or meloxicam used in combination with ketamine and xylazine anesthesia in a Sprague–Dawley rat ovariohysterectomy surgical model. Rats received either subcutaneous saline once daily for 3 d, low-dose (0.05 mg/kg SC) or high-dose (0.1 mg/kg SC) buprenorphine twice daily for 3 d, a single injection of sustained-release buprenorphine (1.2 mg/kg SC), or low-dose (1 mg/kg SC) or high-dose (2 mg/kg SC) meloxicam once daily for 3 d. Clinical analgesic efficacy was assessed over 8 d according to cageside observation scoring, body weight, and behavioral testing. Ovariohysterectomy was associated with 2 d of postoperative pain, and all 3 buprenorphine dosing strategies and both doses of meloxicam demonstrated varying amounts of analgesia. Given the results of the current study, we recommend 0.05 mg/kg SC buprenorphine at least twice daily or a single dose of 1.2 mg/kg SC of sustained-release buprenorphine for rats undergoing midline laparotomy with ovariohysterectomy. Alternatively, meloxicam at 1 to 2 mg/kg SC once daily could be used for this indication.

Curcumin, a polyphenol derived from turmeric, has a wide variety of therapeutic benefits including antiinflammatory, antioxidative, and chemopreventative effects. Oral gavage is widely performed to administer curcumin in laboratory rodents in several experimental models. Although effective, this method can increase stress in the animal, potentially influencing experimental results. Moreover, oral gavage can result in mortality due to accidental instillation of fluid into the lungs, serious mechanical damage, and gavage-related reflux. Here we describe a method for the administration of fixed dosages of curcumin to rats through voluntary consumption of peanut butter, to reduce gavage-related morbidity and distress to animals and to provide environmental enrichment. Fischer 344 (n = 6) rats received 1100 mg/kg of a commercial curcumin product (equivalent to approximately 200 mg/kg of curcumin) in 8 g/kg of peanut butter daily for 5 wk. Curcumin concentrations in rat plasma were measured by using UPLC–MS at 2 to 4 h after administration. All rats voluntarily consumed the peanut butter–curcumin mixture consistently over the 5-wk period. Total curcumin concentrations in plasma samples collected 2 to 4 h after curcumin consumption were 171 ± 48.4 ng/mL (mean ± 1 SD; range, 103 to 240 ng/mL). This noninvasive curcumin delivery method was effective, eliminated the stress caused by daily oral gavage, and added environmental enrichment.

The 2013 edition of the AVMA Guidelines for the Euthanasia of Animals recommends a 10% to 30% volume displacement rate (VDR) per minute for CO₂ euthanasia of rodents. Here we sought to evaluate behavior and plasma catecholamine levels in multiple strains of male and female mice, euthanized individually or in a group, with CO₂ VDR of 10% to 100%. Behavioral observations included ataxia, labored breathing, time to recumbency, time to surgical plane of anesthesia, and the number of jumps or paws at the face during the euthanasia process. Behaviors did not differ significantly between male and female mice at any of the VDR, but interstrain differences occurred. Slower VDR resulted in longer periods of ataxia and labored breathing regardless of euthanasia as a group or as an individual. In addition, mice jumped and pawed at the face more often with slower VDR than higher. At all VDR, mice euthanized as a group had lower catecholamine levels than mice euthanized individually, but there were no significant differences between VDR. Time to recumbency and time to surgical plane anesthesia were longer with slower displacement rates; in addition, these parameters were prolonged for mice euthanized as a group compared with individually. Overall, faster VDR do not appear to be more distressful than slower rates. In fact, faster VDR shorten the time during which mice might experience distress prior to recumbency.

Preclinical studies in animals often require frequent blood sampling over prolonged periods. A preferred method in rats is the implantation of a polyurethane catheter into the jugular vein, with heparinized glycerol as a lock solution. However, analysis of various biologic compounds (for example, microRNA) precludes the use of heparin. We used sodium citrate as an alternative to heparin but observed more frequent loss of catheter patency. We hypothesized that this effect was due to evaporation of lock solution at the exteriorized portion of the catheter, subsequent blood infiltration into the catheter, and ultimately clot formation within the catheter. We therefore tested evaporation and its variables in vitro by using 5 common catheter materials. We used the migration of dye into vertically anchored catheters as a measure of lock displacement due to evaporation. Exposure to dry room-temperature air was sufficient to cause dye migration against gravity, whereas a humid environment and adding glycerol to the lock solution mitigated this effect, thus confirming loss of the lock solution from the catheter by evaporation. We tested 4 catheter treatments for the ability to reduce lock evaporation. Results were validated in vivo by using male Sprague–Dawley rats (n = 12) implanted with polyurethane jugular vein catheters and randomized to receive a nitrocellulose-based coating on the exteriorized portion of the catheter. Coating the catheters significantly improved patency, as indicated by a Kaplan–Meier log-rank hazard ratio greater than 5 in untreated catheters. We here demonstrate that a simple nitrocellulose coating reduces evaporation from and thus prolongs the patency of polyurethane catheters in rats.

Over a 4-wk period in 2017, we received notification from 7 different institutions that mice exported from our SPF barrier facilities had tested positive for mouse rotavirus (MRV). The exports originated from several different buildings across multiple campuses. Our institution excludes MRV in all of our barrier facilities and has historically been free of this virus. Extensive testing of our rooms from which the exported mice originated did not detect the presence of rotavirus. The single commonality among the 7 shipments was the use of shipping boxes acquired from one vendor. These shipping boxes arrived at our institution prepackaged with unsterilized feed and bedding, which we hypothesized was the source of the rotavirus. To test this hypothesis, we housed naïve sentinel mice in clean cages with feed and bedding transferred from 29 unopened, unused shipping boxes. Sentinel mice were exposed to this bedding and feed for 14 d and then evaluated through MRV serology and PCR assay. Of the 29 sentinels, 24 were seropositive for MRV, and 14 of the 29 were PCR positive. These results provided direct evidence that MRV detected by recipient institutions originated from the contaminated feed or bedding within the shipping boxes. To our knowledge, this report is the first description of contaminated materials in shipping boxes resulting in rotaviral infection of mice during export.