Bradley J. Swanson

Impact of Natural and Artificial Barriers to Dispersal on the Population Structure of Bobcats

Abstract We investigated population structure and genetic diversity for bobcats (Lynx rufus) in Michigan, USA, which are distributed throughout the upper peninsula (UP) and the northern half of the lower peninsula (LP) of Michigan. Specifically, we assessed the influence of natural and artificial barriers to dispersal on the genetic population structure of the bobcat across Michigan, as well as in each peninsula. We used 5 microsatellite markers and the statistical package STRUCTURE to identify populations and assign individuals to their population of origin. STRUCTURE identified one population in each peninsula, indicating that the UP and LP are genetically isolated by the Straits of Mackinac which divide the UP and LP. Despite a greater density of roads in the LP, we found no evidence that they have led to intra-peninsular population structure. Our results suggest that, from a genetic standpoint, management agencies do not need to be concerned about the fragmenting effects of roads when producing management plans for bobcats.

Elevational isolation of red fox populations in the Greater Yellowstone Ecosystem

Foxes in the Greater Yellowstone Ecosystem are reported to show high frequencies of blonde and gray coat colors. A survey of park sighting records showed that the frequency of the novel coat colors significantly increases at elevations greater than 2300 m, suggesting some form of elevational isolation. We evaluated the degree of genetic separation between the high-elevation foxes (>2300 m) and contiguous populations of foxes at mid-elevations (1600–2300m). Low-elevation (>1600 m) foxes from North Dakota, >1000 km straight line distance from our populations, were used as a control group. We genotyped 15 high-elevation, 15 mid-elevation, and 10 low-elevation foxes at 10 microsatellite loci each. Heterozygosity was significantly lower in both the high-elevation and mid-elevation populations compared to the low-elevation foxes. The genetic differentiation was significantly greater between the high-elevation and mid-elevation foxes than between the mid-elevation and low-elevation foxes. Similarly, estimates of RST and FST suggest less gene flow occurs between the contiguous high- and mid-elevation fox populations than between the mid- and low-elevation fox populations separated by > 1000 km. The assignment test further supports this hypothesis. Although further work is needed, we suggest that the high-elevation foxes are remnant populations from the Wisconsin glaciation and should be managed as a unique population.

RNA expression profiling of human iPSC-derived cardiomyocytes in a cardiac hypertrophy model.

Cardiac hypertrophy is an independent risk factor for cardiovascular disease and heart failure. There is increasing evidence that microRNAs (miRNAs) play an important role in the regulation of messenger RNA (mRNA) and the pathogenesis of various cardiovascular diseases. However, the ability to comprehensively study cardiac hypertrophy on a gene regulatory level is impacted by the limited availability of human cardiomyocytes. Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) offer the opportunity for disease modeling. Here we utilize a previously established in vitro model of cardiac hypertrophy to interrogate the regulatory mechanism associated with the cardiac disease process. We perform miRNA sequencing and mRNA expression analysis on endothelin 1 (ET-1) stimulated hiPSC-CMs to describe associated RNA expression profiles. MicroRNA sequencing revealed over 250 known and 34 predicted novel miRNAs to be differentially expressed between ET-1 stimulated and unstimulated control hiPSC-CMs. Messenger RNA expression analysis identified 731 probe sets with significant differential expression. Computational target prediction on significant differentially expressed miRNAs and mRNAs identified nearly 2000 target pairs. A principal component analysis approach comparing the in vitro data with human myocardial biopsies detected overlapping expression changes between the in vitro samples and myocardial biopsies with Left Ventricular Hypertrophy. These results provide further insights into the complex RNA regulatory mechanism associated with cardiac hypertrophy.

DEMOGRAPHIC AND GENETIC EVALUATION OF AN AMERICAN MARTEN REINTRODUCTION

Abstract Reintroduced populations are generally smaller and more isolated than native populations; thus even when reintroduced populations are demographically stable, a lack of genetic variation may present a threat to long-term persistence. We examined the demographic structure and genetic variation of the marten reintroduction into the Upper Peninsula of Michigan. Male : female and juvenile : adult female ratios indicate that the Michigan population is demographically stable. Michigan martens had higher allelic diversity (A = 7.4) compared to the average diversity found among Canadian populations (A = 5.8) and similar levels of observed heterozygosity (HCanadian = 0.64, HMichigan = 0.63), excluding Newfoundland martens. We found no significant differences in the allelic diversity or heterozygosity between the reintroduced Michigan population and the source population for the reintroduction, that of Chapleau, Ontario. Surprisingly, we found no evidence of a genetic bottleneck in the Michigan population. We suggest that the genetic success of this reintroduction is a result of the multiple reintroductions and subsequent intrastate translocations that mimicked gene flow. The success was further aided by the presence of small remnant populations that remained in Michigan, as evidenced by the presence of private alleles in Michigan.

Combined genetic and telemetry data reveal high rates of gene flow, migration, and long-distance dispersal potential in Arctic ringed seals (Pusa hispida).

Ringed seals (Pusa hispida) are broadly distributed in seasonally ice covered seas, and their survival and reproductive success is intricately linked to sea ice and snow. Climatic warming is diminishing Arctic snow and sea ice and threatens to endanger ringed seals in the foreseeable future. We investigated the population structure and connectedness within and among three subspecies: Arctic (P. hispida hispida), Baltic (P. hispida botnica), and Lake Saimaa (P. hispida saimensis) ringed seals to assess their capacity to respond to rapid environmental changes. We consider (a) the geographical scale of migration, (b) use of sea ice, and (c) the amount of gene flow between subspecies. Seasonal movements and use of sea ice were determined for 27 seals tracked via satellite telemetry. Additionally, population genetic analyses were conducted using 354 seals representative of each subspecies and 11 breeding sites. Genetic analyses included sequences from two mitochondrial regions and genotypes of 9 microsatellite loci. We found that ringed seals disperse on a pan-Arctic scale and both males and females may migrate long distances during the summer months when sea ice extent is minimal. Gene flow among Arctic breeding sites and between the Arctic and the Baltic Sea subspecies was high; these two subspecies are interconnected as are breeding sites within the Arctic subspecies.

An Application of Manel's Model: Detecting Bobcat Poaching in Michigan

Abstract The illegal harvest of natural resources (i.e., poaching) has the potential to threaten the persistence of many plant and animal species. In Michigan bobcats (Lynx rufus) are distributed throughout the Upper Peninsula (UP) and the northern half of the Lower Peninsula (LP) and are a biologically and economically important species. The popularity of bobcat hunting and trapping in Michigan, along with different harvest regulations between the 2 peninsulas, has created the need for a reliable method of identifying incidences of poaching. Because the bag limit is higher in the UP, we hypothesized that some bobcats harvested in the LP are being registered as originating from the UP. We used 8 polymorphic microsatellite markers and the statistical package STRUCTURE to assign individuals to the population in which they had the highest likelihood of occurrence based on their genotype. We evaluated the influence of using posterior probability threshold values from T > 0.9–0.999 on the number of animals classified as poached. Based on this range, STRUCTURE produced correct assignment rates of 53–82%. All instances of genetic re-assignment involved bobcats claimed as harvested in the UP but genetically assigned to the LP following the suspected method of bobcat poaching in Michigan. This approach provides a reliable method of determining the source population for bobcats harvested in the state and should provide enforcement agencies with a useful way of identifying potential poaching cases.

Genetic analysis and captive breeding program design for the eastern massasauga Sistrurus catenatus catenatus

Abstract The eastern massasauga Sistrurus catenatus catenatus is a declining species for which a captive breeding program was established in 2006. To effectively manage wild and captive populations, an understanding of genetic diversity within the species is necessary. We analyzed mitochondrial DNA sequences of 186 individuals: 109 wild snakes from 34 U.S. and Canadian counties and districts, all 52 breeding program members (23 of known and 29 of unknown origin), 18 other captives of unknown origin, and 7 outgroup representatives of desert massasauga S. c. edwardsii, and western massasauga, S. c. tergeminus. Statistical parsimony, maximum likelihood, and maximum parsimony analyses all identified eastern massasaugas as divergent from western and desert massasaugas. We found 18 different haplotypes among eastern massasaugas, comprising three geographically and genetically differentiated NADH dehydrogenase II (ND2) subunits that potentially reflect post-Pleistocene range expansion from unglaciated into forme...

Detection and Classification of Cougars in Michigan Using Low Copy DNA Sources

Abstract Sporadic reports of cougars (Puma concolor) have occurred in Michigan since its official classification as extirpated in the 1930s. We collected 297 scats from 12 areas in Michigan with heavy sighting reports of cougars. Ten scats produced DNA profiles consistent with cougars. One scat was identified as having a North American origin; the other nine scats produced no useable sequences. One pre-Columbian sample, from a Native American burial site; also matched the current North American genotype. Based on the distance between cougar scats, we conclude that there were at least eight cougars in Michigan during the 3 y of this study. The mtDNA sequences also suggest that at least some of the matrilines currently and historically found in Michigan are the same as those found in current and historical western populations.

Genetic evaluation of a reintroduced population of black-footed ferrets (Mustela nigripes)

The black-footed ferret (Mustela nigripes) went extinct in the wild when the last 18 known ferrets were captured for a captive-breeding program. Following the success of the captive-breeding program, 146 genetically nonessential ferrets were released at the Conata Basin, South Dakota, from 1996 to 1999. We conducted a genetic analysis of the Conata Basin black-footed ferret population from 2001 to 2003 to determine if genetic variation had been lost since the cessation of reintroductions and if demographic- and genetic-based estimates of effective population size (Ne) accurately predicted observed levels of heterozygosity. We used DNA from wild-born kits (n  =  254) in the Conata Basin population (2001–2003) to calculate current genetic diversity levels. Both allelic diversity (A  =  2, both subpopulations) and mean heterozygosity were low for both subpopulations—0.39 ± 0.12 SE in Agate-Sage Creek and 0.39 ± 0.16 SE in Heck Table—but not significantly different from estimates made in 1999. We found no significant difference between observed and expected heterozygosity levels. Demographic-based estimates of Ne were an order of magnitude higher than genetic-based estimates of Ne, but the 2 estimates provide a range of Ne values for the population. This study shows that the Conata Basin ferret population is able to maintain its genetic diversity over time despite its population history.

Human Induced Pluripotent Stem Cell-Derived Cardiac Progenitor Cells in Phenotypic Screening: A Transforming Growth Factor-β Type 1 Receptor Kinase Inhibitor Induces Efficient Cardiac Differentiation

Several progenitor cell populations have been reported to exist in hearts that play a role in cardiac turnover and/or repair. Despite the presence of cardiac stem and progenitor cells within the myocardium, functional repair of the heart after injury is inadequate. Identification of the signaling pathways involved in the expansion and differentiation of cardiac progenitor cells (CPCs) will broaden insight into the fundamental mechanisms playing a role in cardiac homeostasis and disease and might provide strategies for in vivo regenerative therapies. To understand and exploit cardiac ontogeny for drug discovery efforts, we developed an in vitro human induced pluripotent stem cell‐derived CPC model system using a highly enriched population of KDRpos/CKITneg/NKX2.5pos CPCs. Using this model system, these CPCs were capable of generating highly enriched cultures of cardiomyocytes under directed differentiation conditions. In order to facilitate the identification of pathways and targets involved in proliferation and differentiation of resident CPCs, we developed phenotypic screening assays. Screening paradigms for therapeutic applications require a robust, scalable, and consistent methodology. In the present study, we have demonstrated the suitability of these cells for medium to high‐throughput screens to assess both proliferation and multilineage differentiation. Using this CPC model system and a small directed compound set, we identified activin‐like kinase 5 (transforming growth factor‐β type 1 receptor kinase) inhibitors as novel and potent inducers of human CPC differentiation to cardiomyocytes.