Personnel working with laboratory animals are required by laws and guidelines to be trained and qualified to perform biomethodologic procedures. The assessment of competency and proficiency is a vital component of a laboratory animal training program, because this process confirms that the trainees have met the learning objectives for a particular procedure. The approach toward qualification assessment differs between organizations because laws and guidelines do not outline how the assessment should be performed or which methods and tools should be used. Assessment of clinical and surgical medicine has received considerable attention over the last few decades and has progressed from simple subjective methods to well-defined and objective methods of assessing competency. Although biomethodology competency and proficiency assessment is discussed in the literature, a standard and objective assessment method has not yet been developed. The development and implementation of an objective and standardized biomethodologic assessment program can serve as a tool to improve standards, ensure consistent training, and decrease research variables yet ensure animal welfare. Here we review the definition and goals of training and assessment, review assessment methods, and propose a method to develop a standard and objective assessment program for the laboratory animal science field, particularly training departments and IACUC.

A compact facility for SPF mice that was not equipped with a large autoclave used disposable mouse cages instead. The SPF clean room was 5.7 × 8.1 × 2.7 m³, with a breeding capacity of 1008 cages (168 cages on each of 6 racks). We evaluated cleanliness in the SPF clean room under the conditions of an occupation rate of 60% to 70% and typically 1 to 3 personnel (maximum, 4 to 6) daily on weekdays. Personnel were taught standard procedures and received training beforehand. During the 15-mo study period, the maximal concentration of airborne particles 0.5 μm or larger was 1.0 × 10⁴ particles/m3 and that of particles 5.0 μm or larger was 5.0 × 10² particles/m³—well below the maximal permissible concentrations of 3.52 × 10⁵ and 2.93 × 10³ particles/m³, respectively. During the study period, no mice exhibited clinical symptoms of infection. Testing of 2 representative, overtly healthy mice for 16 pathogens including Staphylococcus aureus, Pseudomonas aeruginosa, and Helicobacter bilis failed to detect any of the target agents. The current study demonstrates the feasibility of the compact facility for breeding SPF mice in the academic environment.

Little is known about the prevalence of zoonotic infections among laboratory animal care technicians (LAT). Q fever, a disease caused by Coxiella burnetii, is a known occupational hazard for persons caring for livestock. We sought to determine the seroprevalence of C. burnetii antibodies among LAT and to identify risk factors associated with C. burnetii seropositivity. A survey was administered and serum samples collected from a convenience sample of 97 LAT. Samples were screened by using a Q fever IgG ELISA. Immunofluorescent antibody assays for phase I and phase II IgG were used to confirm the status of samples that were positive or equivocal by ELISA; positive samples were titered to endpoint. Antibodies against C. burnetii were detected in 6 (6%) of the 97 respondents. In our sample of LAT, seropositivity to C. burnetii was therefore twice as high in LAT as compared with the general population. Age, sex, and working with sheep regularly were not associated with seropositivity. Risk factors associated with seropositivity included breeding cattle within respondent's research facility, any current job contact with waste from beef cattle or goats, and exposure to animal waste during previous jobs or outside of current job duties. Only 15% of responding LAT reported being aware that sheep, goats, and cattle can transmit Q fever. Research facilities that use cattle or goats should evaluate their waste-management practices and educational programs in light of these findings. Additional efforts are needed to increase awareness among LAT regarding Q fever and heightened risk of exposure to infectious materials. Physicians should consider the risk of infection with C. burnetii when treating LAT with potential occupational exposures.

Sperm preservation protocols differ among animal species because of different sperm characteristics among species. Rat sperm have extreme sensitivity to suboptimal conditions in centrifugation, pipetting and chilling due to their longer tail, the shape and size of the sperm head, and membrane composition. The aim of this study was to determine optimal conditions for short-term storage of rat sperm by evaluating their motility and membrane and acrosomal integrity in response to various extender solutions, temperatures, and durations. Motility of rat sperm was highest when stored at 22 °C; motility was 28% and 14% at 72 h in TL-HEPES and PBS extenders, respectively. The motility and membrane integrity of rat sperm fell significantly within 24 h at 4 and 37 °C. Although cold storage did not have a detrimental effect on acrosomal integrity of sperm, room temperature storage reduced acrosomal integrity after 24 h. LEY extender caused the highest loss in acrosomal integrity at 48 and 72 h. In conclusion, storage at 4 or 37 ° C reduced the motility and membrane integrity of rat sperm even with short incubation periods. Rat sperm stored in TL-HEPES or PBS remained motile for at least 3 d when held at 22 °C.

Individual ventilated cages (IVC) are increasing in popularity. Although mice avoid IVC in preference testing, they show no aversion when provided additional nesting material or the cage is not ventilated. Given the high ventilation rate in IVC, we developed 3 hypotheses: that mice housed in IVC experience more cold stress than do mice housed in static cages; that IVC-induced cold stress affects the results of experiments using mice; and that, when provided shelters, mice behaviorally thermoregulate and thereby rescue the cold-stress effects of IVC. To test these hypotheses, we housed mice in IVC, IVC with shelters, and static cages maintained at 20 to 21 °C. We quantified the cold stress of each housing system on mice by assessing nonshivering thermogenesis and brown adipose vacuolation. To test housing effects in a common, murine model of human disease, we implanted mice with subcutaneous epidermoid carcinoma cells and quantified tumor growth, tumor metabolism, and adrenal weight. Mice housed in IVC had histologic signs of cold stress and significantly higher nonshivering thermogenesis, smaller subcutaneous tumors, lower tumor metabolism, and larger adrenal weights than did mice in static cages. Shelters rescued IVC-induced nonshivering thermogenesis, adrenal enlargement, and phenotype-dependent cold-mediated histologic changes in brown adipose tissue and tumor size. IVC impose chronic cold stress on mice, alter experimental results, and are a source of systemic confounders throughout rodent-dependent research. Allowing mice to exhibit behavioral thermoregulation through seeking shelter markedly rescues the experiment-altering effects of housing-imposed cold stress, improves physiologic uniformity, and increases experimental reproducibility across housing systems.

Light entrains normal circadian rhythms of physiology and metabolism in all mammals. Previous studies from our laboratory demonstrated that spectral transmittance (color) of light passing through cages affects these responses in rats. Here, we addressed the hypothesis that red tint alters the circadian nocturnal melatonin signal and circadian oscillation of other metabolic and physiologic functions. Female nude rats (Hsd:RH-Foxn1^rnu; n = 12 per group) were maintained on a 12:12-h light (300 lx; 123.0 μW/cm²; lights on 0600):dark regimen in standard polycarbonate translucent clear or red-tinted cages. After 1 wk, rats underwent 6 low-volume blood draws via cardiocentesis over a 4-wk period. Plasma melatonin levels were low during the light phase (1.0 ± 0.2 pg/mL) in rats in both types of cages but were significantly lower in red-tinted (105.0 ± 2.4 pg/mL) compared with clear (154.8 ± 3.8 pg/mL) cages during the dark. Normal circadian rhythm of plasma total fatty acid was identical between groups. Although phase relationships of circadian rhythms in glucose, lactic acid, pO₂, and pCO₂ were identical between groups, the levels of these analytes were lower in rats in red-tinted compared with clear cages. Circadian rhythms of plasma corticosterone, insulin, and leptin were altered in terms of phasing, amplitude, and duration in rats in red-tinted compared with clear cages. These findings indicate that spectral transmittance through red-colored cages significantly affects circadian regulation of neuroendocrine, metabolic, and physiologic parameters, potentially influencing both laboratory animal health and wellbeing and scientific outcomes.

The common marmoset (Callithrix jacchus), a laboratory nonhuman primate, is a well-known model of several human diseases and conditions, but the nutritional needs of these animals are not fully understood. Here we describe a 4-mo controlled study in which we increased the dietary fat and protein of subadult male common marmosets by using healthy snacks. Six male marmosets received their normal diet (control), and an additional 6 were given their normal diet supplemented daily with a 14-kcal snack. Cashews and waxworms were used as the snack, given their high-fat content. Although body weight did not differ between the 2 groups, only control male marmosets showed increased chest circumferences over the course of the study. Glucoregulatory function remained consistent in the snack-fed marmosets, whereas control animals had progressed toward higher insulin. Other indices of glucoregulation indicated significant differences in adiponectin and the cortisol:cortisone ratio between the 2 groups, but no differences in lipid concentration were detected. Therefore, the most notable difference attributable to the snack feeding was improved glucoregulation. Because the snacks we used had a high proportion of unsaturated compared with saturated fat, we suggest that these healthy high-fat–high-protein snacks provide an important contribution to the nutrition of this laboratory species. This study also demonstrates the utility of marmosets as a model for understanding the implications of dietary fats in humans.

We used a high-density array of real-time PCR assays for commonly reported rodent infectious agents (PRIA) to test naturally infected index mice and sentinel mice exposed by contact and soiled-bedding transfer. PRIA detected 14 pathogens—including viruses, bacteria, fur mites, pinworms, and enteric protozoa—in 97.2% of 28 pooled fecal samples, fur–perianal swabs, and oral swabs from 4 cages containing a total of 10 index mice. Among these pathogens, PRIA (like conventional health monitoring methods) failed to detect Mycoplasma pulmonis, Pasteurella pneumotropica, and Giardia spp. in all of the 9 contact and 9 soiled-bedding sentinels. PRIA demonstrated murine adenovirus and Cryptosporidium and Spironucleus spp. in contact but not soiled-bedding sentinels and detected Helicobacter and pinworms in fewer than half of the soiled-bedding sentinels. Of the 4 species of Helicobacter that species-specific PCR assays identified in index mice, only H. ganmani was found in soiled-bedding and contact sentinels. PRIA detected all of the pathogens in sentinels that were identified by conventional methods. Myobia musculi was detected by PCR in index and sentinel mice but missed by conventional parasitologic examinations. In summary, PRIA reproducibly detected diverse pathogens in heavily pooled specimens collected noninvasively from infected index mice antemortem. The inability of PRIA and conventional health monitoring methods (that is, parasitology, micro-biology, and serology) to demonstrate transmission of some pathogens to contact sentinels and the inefficient transmission of others to soiled-bedding sentinels underscores the importance of direct PCR testing to determine the pathogen status of rodents in quarantine and during routine colony surveillance.

Detecting and controlling murine fur mites continues to be challenging. Here we compared the efficacy of fur-pluck, cage PCR, and fur PCR testing of mice naturally infested with Myocoptes musculinus and make recommendations regarding the application of these diagnostic strategies in aged or treated mice. We compared all 3 diagnostic methods in groups of infested and noninfested control mice over time. For fur plucks, we used a scoring system to quantitatively compare mite infestations across ages. Mice that were 4 wk old had higher egg and mite scores than did older mice, with average scores at 4 wk corresponding to 40 to 100 individual fur mites and eggs per sample. Furthermore, 15% and 20% of samples from infested mice at 24 and 28 wk of age, respectively, lacked all fur mites and eggs. Cage PCR results varied as mice grew older. Fur PCR testing was the most sensitive and specific assay in untreated infested mice, particularly when mite densities were low. In addition, we compared fur-pluck and fur PCR tests for evaluating the efficacy of selamectin treatment. Two treatments with selamectin eliminated Myocoptes fur-mite infestations. At 8 wk after treatment, all fur-pluck samples were negative, but one-third of treated infested cages remained positive by fur PCR assay; at 16 wk after treatment, all cages were negative by fur PCR assay. Because offspring of infested mice were invariably heavily infested, breeding of suspected infested mice with subsequent testing of offspring was the definitive testing strategy when fur-pluck and PCR results conflicted.

A detailed epidemiologic survey of spontaneous diseases of mice used in biomedical research has not been performed in more than 4 decades. The current study examined all mouse disease reports for a subset of the University of Pennsylvania vivaria from October 2010 through September 2011. Mortality logs were examined over the same period of time. After eliminating protocol-related cases, the incidence rates for more than 30 diseases were calculated in terms of number of cases per 1000 cages per month. The average daily census for the facilities analyzed exceeded 29,000 cages and included more than 180 research groups. No single research group accounted for more than 4% of the total number of cases reported, indicating that this study did not simply quantify the spontaneous disease incidence in a limited number of research groups. Spontaneous mortality with unknown cause in adult and neonatal mice without prior reported illness was the most commonly reported issue, followed by dermatitis, ocular disease, and nonspecific clinical signs including lethargy, poor hair coat, and muscle wasting. These results indicate that improving the ability to identify sick mice is important in refining the care and use of mice in biomedical research. The information provided in the current study can help to provide a baseline for comparison, guide the field in directing mouse welfare research toward areas of need, and identify optimal methods of care for mice in biomedical research.

Investigations into the biology of aquatic and semiaquatic species, including those involving sensory specialization, often require creative solutions to novel questions. We developed a technique for safely anesthetizing a semiaquatic chelonian species, the diamondback terrapin (Malaclemys terrapin), for measurement of auditory evoked potentials while animals were completely submerged in water. Custom-modified endotracheal tubes were used to obtain a watertight seal on both sides of the glottis and prevent aspiration of water during testing. No adverse effects were seen after the procedures, and assessment of venous blood-gas partial pressures and lactate concentrations indicated that sufficient gas exchange was maintained under anesthesia through manual ventilation.