Equipment and Methods for Concurrently Housing Germfree and Gnotobiotic Mice in the Same Room.

Here, we combined the use of 2 technologies that have not previously been used together-a positively pressurized isolator IVC (IsoIVC-P) and a modular isolator with integrated vaporized hydrogen peroxide (VHP) technology???to develop highly tractable and scalable methods to support long-term maintenance of germfree mouse colonies and the concurrent use of germfree and gnotobiotic mice in the same room. This space-efficient system increases the practicality of microbiome studies. Specifically, the exterior surfaces of microbially similar IsoIVC-P were sterilized by using VHP prior to opening the cages and handling the mice therein. This space-efficient system increases the feasibility of microbiome studies. After over 74 wk of experimentation and handling equivalent to more than 1,379,693 germfree mouse-days, we determined that the method and practices we developed have a weekly performance metric of 0.0001 sterility breaks per husbandry unit; this rate is comparable to the isolator 'gold standard.' These data were achieved without adverse incidents while maintaining an Altered Schaedler Flora colony and multiple gnotobiotic studies involving fecal microbial transplants in the same room. Our novel IsoIVC-P???VHP workstation housing system thus improves microbiome research efficiency, eliminates hazards, and reduces risks associated with traditional methods.

Housing and Husbandry Alternatives for Naked Mole Rat Colonies Used in Research Settings.

Naked mole rats (Heterocephalus glaber) are a unique rodent species originating in Africa and are increasingly being used in research. Their needs and characteristics differ from those of other rodents used in research. Unique housing systems are necessary to address the special macro- and microenvironmental requirements of NMRs. Naked mole rats are one of the 2 known eusocial mammalian species, are extremely long-living, are active burrowers, and are accustomed to a subterranean environment. Unlike typical rats and mice, naked mole rats need specific, unique housing systems that mimic their natural subterranean environment to support health and longevity. Here we provide an overview of naked mole rats and a housing method that can be used in research settings.

Multiregion transcriptomic profiling of the primate brain reveals signatures of aging and the social environment.

Aging is accompanied by a host of social and biological changes that correlate with behavior, cognitive health and susceptibility to neurodegenerative disease. To understand trajectories of brain aging in a primate, we generated a multiregion bulk (N = 527 samples) and single-nucleus (N = 24 samples) brain transcriptional dataset encompassing 15 brain regions and both sexes in a unique population of free-ranging, behaviorally phenotyped rhesus macaques. We demonstrate that age-related changes in the level and variance of gene expression occur in genes associated with neural functions and neurological diseases, including Alzheimer's disease. Further, we show that higher social status in females is associated with younger relative transcriptional ages, providing a link between the social environment and aging in the brain. Our findings lend insight into biological mechanisms underlying brain aging in a nonhuman primate model of human behavior, cognition and health.

Social connections predict brain structure in a multidimensional free-ranging primate society.

Reproduction and survival in most primate species reflects management of both competitive and cooperative relationships. Here, we investigated the links between neuroanatomy and sociality in free-ranging rhesus macaques. In adults, the number of social partners predicted the volume of the mid-superior temporal sulcus and ventral-dysgranular insula, implicated in social decision-making and empathy, respectively. We found no link between brain structure and other key social variables such as social status or indirect connectedness in adults, nor between maternal social networks or status and dependent infant brain structure. Our findings demonstrate that the size of specific brain structures varies with the number of direct affiliative social connections and suggest that this relationship may arise during development. These results reinforce proposed links between social network size, biological success, and the expansion of specific brain circuits.

Fecal Bacterial Microbiota of Healthy Free-Ranging, Healthy Corralled, and Chronic Diarrheic Corralled Rhesus Macaques (Macaca mulatta).

A clinical challenge to nearly every primate facility in North America is chronic idiopathic diarrhea (CID), the pathogenesis of which has yet to be fully elucidated. However, wild macaques appear resistant to CID, a trend that we observed in the free-ranging population of the Caribbean Primate Research Center. The gastrointestinal microbiota has been shown to have a significant role in the pathogenesis of disease and in maintaining normal health and development of the gut. In humans, chronic diarrhea is associated with alteration of the gut microbiota, which has lower bacterial diversity than does the microbiota of healthy humans. The current study was designed to describe and compare the fecal bacterial microbiota of healthy corralled, CID corralled, and healthy, free-ranging macaques. Fresh fecal samples were collected from healthy corralled (HC; n = 30) and CID (n = 27) rhesus macaques and from healthy macaques from our free-ranging colony (HF; n = 43). We excluded macaques that had received antibiotics during the preceding 60 d (90 d for healthy animals). Bacterial DNA was extracted, and the V4 region of the 16S rRNA gene was sequenced and compared with known databases. The relative abundance of Proteobacteria was higher in CID animals than HC animals, but otherwise few differences were found between these 2 groups. HF macaques were differentially enriched with Christensenellaceae and Helicobacter, which are highly associated with a 'healthy' gut in humans, as compared to corralled animals, whereas CID animals were enriched with Proteobacteria, which are associated with dysbiosis in other species. These results indicate that environment has a greater influence than health status on the gut microbiota. Furthermore, the current data provided targets for future studies on potential clinical interventions, such as probiotics and fecal transplants.

Sociality predicts individual variation in the immunity of free-ranging rhesus macaques.

Social integration and social status can substantially affect an individual's health and survival. One route through which this occurs is by altering immune function, which can be highly sensitive to changes in the social environment. However, we currently have limited understanding of how sociality influences markers of immunity in naturalistic populations where social dynamics can be fully realized. To address this gap, we asked if social integration and social status in free-ranging rhesus macaques (Macaca mulatta) predict anatomical and physiological markers of immunity. We used data on agonistic interactions to determine social status, and social network analysis of grooming interactions to generate measures of individual variation in social integration. As measures of immunity, we included the size of two of the major organs involved in the immune response, the spleen and liver, and counts of three types of blood cells (red blood cells, platelets, and white blood cells). Controlling for body mass and age, we found that neither social status nor social integration predicted the size of anatomical markers of immunity. However, individuals that were more socially connected, i.e., with more grooming partners, had lower numbers of white blood cells than their socially isolated counterparts, indicating lower levels of inflammation with increasing levels of integration. These results build upon and extend our knowledge of the relationship between sociality and the immune system in humans and captive animals to free-ranging primates, demonstrating generalizability of the beneficial role of social integration on health.

Outlining key inflammation-associated parameters during early phase of an experimental gram-negative sepsis model in rhesus macaques (Macaca mulatta).

The aim of this study was to identify inflammation-associated markers during the early phase of sepsis in rhesus macaque. Four rhesus macaques were given an intravenous dose of 1010 CFU/kg of E. coli. Blood samples were collected before, or 30 minutes, 2, 4, 6 and 8 hours after E. coli infusion. Physiological parameters, bacteremia, endotoxemia, C-reactive protein (CRP), procalcitonin (PCT), and plasma cytokines/chemokines were determined for each animal. Bacteremia was present in all animals from 30 minutes to 3 hours after E. coli infusion whereas endotoxin was detected during the full-time course. CRP and PCT levels remained at detectable levels during the whole experimental window suggesting an ongoing inflammatory process. Signature cytokines and chemokines such as TNF-α, MIP-1α, and MIP-1ß peaked about 2 hours after E. coli infusion and decreased thereafter. Plasma IL-6, IL-12p40, IFN-γ, and IL-1Ra, as well as I-TAC, MIG, IP-10 and MCP-1, remained at detectable levels after 4 hours of E. coli infusion. This nonhuman primate model could be useful for the assessment of new therapeutics aiming to suppress key inflammatory markers throughout sepsis early phases.

Fecal bacterial microbiota of Canadian commercial mink (Neovison vison): Yearly, life stage, and seasonal comparisons.

The gastrointestinal microbiome is known to play a critical role in animal health but has been relatively poorly characterized in commercial mink, an obligate carnivore. Whether the microbiota can be manipulated in mink to improve pelt quality, health, and well-being is unknown. The objectives of this study were to characterize the fecal microbiota of commercial mink, and to evaluate potential changes due to year (2014 vs 2015), life stage (adult female vs weaned kit), season (summer vs winter), and between Canadian farms. Pooled fecal samples were collected from adult females and weaned kits in the summers of 2014 (n = 173) and 2015 (n = 168), and from females in the winter of 2016 (n = 39), a time when females undergo marked calorie restriction, from 49 mink farms in Ontario. Bacterial DNA was extracted and the V4 region of the 16S rRNA gene was amplified. Approximately 22 million sequences were identified following quality control filtering. A total of 31 bacterial phyla were identified; however, only 3 comprised >1% of the total sequences identified, with Firmicutes and Proteobacteria together comprising 95% of the total sequences. Comparisons were made by life stage, season and year; no differences were found in the relative abundance of any taxa between samples collected from adult females and weaned kits from the same year and the greatest number of differences at each taxonomic level were noted between 2014 and 2015. Significantly more operational taxonomic units (OTUs) were found in 2014 than 2015 or 2016 (p<0.05) and samples from 2014 were more even, but less diverse than in 2015 (p = 0.002 and 0.001, respectively). There were significant differences in community population and structure by year and season (all p-values <0.001). The predominant phyla and genera at the farm level were similar from year to year. Together, these indicate that mink environment, season, and time are important factors in the stability of gastrointestinal microbiota, once mink reach maturity.

On-farm biosecurity practices and causes of preweaning mortality in Canadian commercial mink kits.

BACKGROUND: Mink are an important animal commodity group in Canada and excessive kit mortality represents a significant loss to production. National biosecurity standards have been developed for Canadian mink farms, but it is unclear how well these standards have been implemented as there are no studies correlating management practices of mink producers with causes of death in mink kits. To that end, we surveyed Ontario mink producers on their biosecurity and management practices and conducted almost 5660 post mortem examinations on found-dead, preweaned kits to characterize mink farm biosecurity practices and causes of death in preweaned kits. RESULTS: We found that very few biosecurity and management practices were uniformly used by producers, despite good awareness of appropriate practices. Use of personal protective equipment was implemented by fewer than 50% of respondents, while control of mink shed access, disinfection of feed containers after use, and use of a rodent control program were the only practices implemented by greater than 70% of respondents. Only 18% of producers reported regular use of antimicrobials in feed or water, although 91% stated they used antimicrobials for treatment of bacterial diseases on a regular basis. On post mortem examination, no gross abnormalities were noted in 71% of the kits, 45% were thought to be stillborn or aborted, 27% had some form of abnormal fluid distribution in the body, and 2% had a congenital malformation. A subset of 69 gastrointestinal tract samples was submitted for bacterial culture, of which 45 samples yielded sufficient growth. Most interesting was the identification of Salmonella enterica serovar Heidelberg in 11% of samples. CONCLUSIONS: The results of this study will provide a benchmark for Canadian mink producers and their veterinarians, defining the areas to which greater attention should be given to ensure more rigorous biosecurity practices are in place. Ultimately, these improvements in practices may contribute to increased mink production and animal well-being.

Generalized Tetanus in a Gyrfalcon ( Falco rusticolus ) with Pododermatitis.

A 2-yr-old male gyrfalcon ( Falco rusticolus ) was presented for severe and generalized muscle spasticity and pododermatitis. The falcon had been treated for pododermatitis over the previous 4 mo. Muscle rigidity and spasms involved the entire bird but were more severe on the right leg. The bird was also tachypneic and hyperthermic at 45 C. While the plantar pododermatitis lesions had healed, there was still a small abscess on the lateral aspect of the right foot. Clinical signs were consistent with tetanus. Several bacteria were isolated from the abscess including Clostridium tetani . The isolate was confirmed to be toxigenic by PCR. Attempts to detect tetanus toxin in the bird's plasma were unsuccessful. The abscess was debrided. The gyrfalcon received equine tetanus antitoxin, intravenous metronidazole, methocarbamol, midazolam, a constant-rate infusion of Fentanyl, active cooling, and supportive care. Inhalant anesthesia with isoflurane was the only treatment that would lower the body temperature and reduce the clinical signs. The gyrfalcon died a few hours after admission. The characteristic clinical signs and isolation of toxigenic C. tetani from a wound were strong supportive evidence for a diagnosis of tetanus. This case constitutes the first reported natural occurrence of tetanus in an avian species. Further information is needed to determine whether gyrfalcons are more susceptible to tetanus than are other avian species and whether pododermatitis lesions may be risk factors.