Depletion of the Microbiota Has a Modest but Important Impact on the Fungal Burden of the Heart and Lungs during Early Systemic Candida auris Infection in Neutropenic Mice.

The progression and systemic pathobiology of C. auris in the absence of a microbiota have not been described. Here, we describe the influence of the microbiota during the first 5 days of C. auris infection in germ-free or antibiotic-depleted mice. Depletion of the bacterial microbiota in both germ-free and antibiotic-depleted models results in a modest but important increase in the early stages of C. auris infection. Particularly the heart and lungs, followed by the cecum, uterus, and stomach, of intravenously (i.v.) infected neutropenic mice showed significant fungal organ burden. Understanding disease progression and pathobiology of C. auris in individuals with a depleted microbiota could potentially help in the development of care protocols that incorporate supplementation or restoration of the microbiota before invasive procedures, such as transplantation surgeries.

Minimum habitat thresholds required for conserving mountain lion genetic diversity.

Jointly considering the ecology (e.g., habitat use) and genetics (e.g., population genetic structure and diversity) of a species can increase understanding of current conservation status and inform future management practices. Previous analyses indicate that mountain lion (Puma concolor) populations in California are genetically structured and exhibit extreme variation in population genetic diversity. Although human development may have fragmented gene flow, we hypothesized the quantity and quality of remaining habitat available would affect the genetic viability of each population. Our results indicate that area of suitable habitat, determined via a resource selection function derived using 843,500 location fixes from 263 radio-collared mountain lions, is strongly and positively associated with population genetic diversity and viability metrics, particularly with effective population size. Our results suggested that contiguous habitat of ≥10,000 km2 may be sufficient to alleviate the negative effects of genetic drift and inbreeding, allowing mountain lion populations to maintain suitable effective population sizes. Areas occupied by five of the nine geographic-genetic mountain lion populations in California fell below this habitat threshold, and two (Santa Monica Area and Santa Ana) of those five populations lack connectivity to nearby populations. Enhancing ecological conditions by protection of greater areas of suitable habitat and facilitating positive evolutionary processes by increasing connectivity (e.g., road-crossing structures) might promote persistence of small or isolated populations. The conservation status of suitable habitat also appeared to influence genetic diversity of populations. Thus, our results demonstrate that both the area and status (i.e., protected or unprotected) of suitable habitat influence the genetic viability of mountain lion populations.

Impact of Candida auris Infection in a Neutropenic Murine Model.

Candida auris has become a global public health threat due to its multidrug resistance and persistence. Currently, there are limited murine models to study C. auris infection. Those models use a combination of cyclophosphamide and cortisone acetate, suppressing both innate and adaptive immunity. Here, we compare C. auris infection in two neutrophil-depleted murine models in which innate immunity is targeted using the monoclonal antibodies 1A8 and RB6-8C5.

Candida albicans Elicits Pro-Inflammatory Differential Gene Expression in Intestinal Peyer's Patches.

The details of how gut-associated lymphoid tissues such as Peyer's patches (PPs) in the small intestine play a role in immune surveillance, microbial differentiation and the mucosal barrier protection in response to fungal organisms such as Candida albicans are still unclear. We particularly focus on PPs as they are the immune sensors and inductive sites of the gut that influence inflammation and tolerance. We have previously demonstrated that CD11c+ phagocytes that include dendritic cells and macrophages are located in the sub-epithelial dome within PPs sample C. albicans. To gain insight on how specific cells within PPs sense and respond to the sampling of fungi, we gavaged naïve mice with C. albicans strains ATCC 18804 and SC5314 as well as Saccharomyces cerevisiae. We measured the differential gene expression of sorted CD45+ B220+ B-cells, CD3+ T-cells and CD11c+ DCs within the first 24 h post-gavage using nanostring nCounter® technology. The results reveal that at 24 h, PP phagocytes were the cell type that displayed differential gene expression. These phagocytes were able to sample C. albicans and discriminate between strains. In particular, strain ATCC 18804 upregulated fungal-specific pro-inflammatory genes pertaining to innate and adaptive immune responses. Interestingly, PP CD11c+ phagocytes also differentially expressed genes in response to C. albicans that were important in the protection of the mucosal barrier. These results highlight that the mucosal barrier not only responds to C. albicans, but also aids in the protection of the host.

Assessment of environmental and occupational exposure while working with multidrug resistant (MDR) fungus Candida auris in an animal facility.

In less than a decade since its identification in 2009, the emerging fungal pathogen Candida auris has become a major public health threat due to its multidrug resistant (MDR) phenotype, high transmissibility, and high mortality. Unlike other Candida species, C. auris has acquired high levels of resistance to an already limited arsenal of antifungals. As an emerging pathogen, there are currently a limited number of documented murine models of C. auris infection. These animal models use inoculums as high as 107-108 cells per mouse, and the environmental and occupational exposure of working with these models has not been clearly defined. Using real-time quantitative polymerase chain reaction (PCR) and culture, we monitored the animal holding room as well as the procedure room for up to 6 months while working with an intravenous model of C. auris infection. This study determined that shedding of the organism is dose-dependent, as detectable levels of C. auris were detected in the cage bedding when mice were infected with 107 and 108 cells, but not with doses of 105 and 106 cells. Autoclaving bedding in closed micro-isolator cages was found to be an effective way to minimize exposure for animal caretakers. We found that tissue necropsies of infected mice were also an important source of potential source exposure to C. auris. To mitigate these potential exposures, we implemented a rigorous "buddy system" workflow and a disinfection protocol that uses 10% bleach followed by 70% ethanol and can be used in any animal facility when using small animal models of C. auris infection.

PSOROPTES INFESTATION AND TREATMENT IN AN ISOLATED POPULATION OF BIGHORN SHEEP (OVIS CANADENSIS).

The authors captured bighorn sheep (Ovis canadensis) comprising a small population in the San Bernardino Mountains of California and evaluated the degree of infestation by mites of the genus Psoroptes for each individual. The animals were treated with two novel methods: amitraz-impregnated collars and cyfluthrin-impregnated ear tags and recaptured the following year to evaluate the effect of treatment. The authors compared data on degree of infestation for animals recaptured in the posttreatment year, detected no significant interyear differences in infestation severity scores among animals treated with amitraz or cyfluthrin, and could not detect any differences between treatment types. However, a significant (P<0.10) decreased pattern in severity scores from the beginning to the end of treatments was detected, suggesting a cumulative therapeutic value in repeated annual treatments across the 3-yr period. Additionally, the authors detected a lower median mite severity score between 2000 and a later capture in 2006. These positive outcomes may be the result of previous treatments during 2000-2002, but environmental covariates not accounted for could have been contributing factors. Avermectin drugs with longer release profiles may be a more effective treatment option in this and other small bighorn sheep populations that are compromised with mite infestations.

Comparison of trace mineral concentrations in tail hair, body hair, blood, and liver of mule deer (Odocoileus hemionus) in California.

Measuring trace mineral concentrations can be an important component of assessing the health of free-ranging deer. Trace mineral concentrations in liver most accurately reflect the trace mineral status of an individual, but, in live animals, whole blood or serum are the most commonly used sample types. Trace minerals measured in serum, such as copper, zinc, and iron, do not always accurately correlate to liver concentrations, and supplementary samples for evaluating the trace mineral status in live deer would be useful. We evaluated the utility of body and tail hair for measuring selenium, copper, zinc, iron, and manganese in free-ranging mule deer (Odocoileus hemionus) by using Spearman rank correlations and linear regression. Correlations were strongest at the time of or shortly after growth of the winter coat and in resident deer. In live deer, strong correlations and moderate linear associations (R (2) = 0.57) were detected between body and tail hair and whole blood selenium in December. In postmortem-sampled deer, a strong correlation and linear association (R (2) = 0.80) were found between liver and body hair selenium in August-November. Results indicate that body hair, if collected during or shortly after growth of the winter coat, can be used as a supplementary sample for measuring selenium concentrations in deer. None of the other correlations and linear associations were found to be sufficiently strong to conclude that hair can reliably be utilized as a complementary sample for measuring these trace mineral concentrations.

Serosurveillance for livestock pathogens in free-ranging mule deer (Odocoileus hemionus).

Routine disease surveillance has been conducted for decades in mule deer (Odocoileus hemionus) in California for pathogens shared between wildlife and domestic ruminants that may have implications for the animal production industry and wildlife health. Deer sampled from 1990 to 2007 (n = 2,619) were tested for exposure to six pathogens: bluetongue virus (BTV), epizootic hemorrhagic disease virus (EHDV), bovine viral diarrhea virus (BVDV), Leptospira spp., Anaplasma spp. and Brucella spp. We evaluated the relationship between exposure to these pathogens and demographic risk factors to identify broad patterns in seroprevalence across a large temporal and spatial scale. The overall seroprevalence for the entire study period was 13.4% for BTV, 16.8% for EHDV, 17.1% for BVDV, 6.5% for Leptospira spp., 0.2% for Brucella spp., and 17% for Anaplasma spp. Antibodies against BTV and EHDV were most prevalent in the deer populations of southern California. Antibodies against Leptospira spp. and Anaplasma spp. were most prevalent in coastal and central northern California whereas antibodies against BVDV were most prevalent in central-eastern and northeastern California. The overall seroprevalence for Anaplasma spp. was slightly lower than detected in previous studies. North and central eastern California contains large tracts of federal land grazed by livestock; therefore, possible contact between deer and livestock could explain the high BVDV seroprevalence found in these areas. Findings from this study will help to establish baseline values for future comparisons of pathogen exposure in deer, inform on long-term trends in deer population health and provide relevant information on the distribution of diseases that are shared between wildlife and livestock.

Zoonotic vector-borne bacterial pathogens in California mountain lions (Puma concolor), 1987-2010.

Sera collected from 442 mountain lions in 48 California counties between the years of 1987 and 2010 were tested using immunofluorescence assays and agglutination tests for the presence of antibodies reactive to Yersinia pestis, Francisella tularensis, Bartonella henselae, Borrelia burgdorferi, and Anaplasma phagocytophilum antigens. Data were analyzed for spatial and temporal trends in seropositivity. Seroprevalences for B. burgdorferi (19.9%) and B. henselae (37.1%) were relatively high, with the highest exposure in the Central Coast region for B. henselae. B. henselae DNA amplified in mountain lion samples was genetically similar to human-derived Houston-1 and domestic cat-derived U4 B. henselae strains at the gltA and ftsZ loci. The statewide seroprevalences of Y. pestis (1.4%), F. tularensis (1.4%), and A. phagocytophilum (5.9%), were comparatively low. Sera from Y. pestis- and F. tularensis-seropositive mountain lions were primarily collected in the Eastern and Western Sierra Nevada, and samples reactive to Y. pestis antigen were collected exclusively from adult females. Adult age (≥ 2 years) was a risk factor for B. burgdorferi exposure. Over 70% of tested animals were killed on depredation permits, and therefore were active near areas with livestock and human residential communities. Surveillance of mountain lions for these bacterial vector-borne and zoonotic agents may be informative to public health authorities, and the data are useful for detecting enzootic and peridomestic pathogen transmission patterns, particularly in combination with molecular characterization of the infecting organisms.

Genetic structure of mountain lion (Puma concolor) populations in California.

Analysis of 12 microsatellite loci from431 mountain lions (Puma concolor)revealed distinct genetic subdivision that wasassociated with geographic barriers andisolation by distance in California. Levels ofgenetic variation differed among geographicregions, and mountain lions that inhabitedcoastal areas exhibited less heterozygositythan those sampled inland. The San FranciscoBay and Sacramento-San Joaquin River Delta, theCentral Valley, and the Los Angeles Basinappeared to be substantial barriers to geneflow, and allele frequencies of populationsseparated by those features differedsubstantially. A partial barrier to gene flowappeared to exist along the crest of the SierraNevada. Estimated gene flow was high amongmountain lions inhabiting the Modoc Plateau,the western Sierra Nevada, and northern sectionof the eastern Sierra Nevada. SouthernCalifornia mountain lion populations mayfunction as a metapopulation; however, humandevelopments threaten to eliminate habitat andmovement corridors. While north-south geneflow along the western Sierra Nevada wasestimated to be very high, projected loss andfragmentation of foothill habitat may reducegene flow and subdivide populations. Preservation of existing movement corridorsamong regions could prevent population declinesand loss of genetic variation. This studyshows that mountain lion management andconservation efforts should be individualizedaccording to region and incorporatelandscape-level considerations to protecthabitat connectivity.