Hugh B. Britten

The absence of concordant population genetic structure in the black-tailed prairie dog and the flea, Oropsylla hirsuta, with implications for the spread of Yersinia pestis.

The black‐tailed prairie dog (Cynomys ludovicianus) is a keystone species on the mid‐ and short‐grass prairies of North America. The species has suffered extensive colony extirpations and isolation as a result of human activity including the introduction of an exotic pathogen, Yersinia pestis, the causative agent of sylvatic plague. The prairie dog flea, Oropsylla hirsuta, is the most common flea on our study colonies in north‐central Montana and it has been shown to carry Y. pestis. We used microsatellite markers to estimate the level of population genetic concordance between black‐tailed prairie dogs and O. hirsuta in order to determine the extent to which prairie dogs are responsible for dispersing this potential plague vector among prairie dog colonies. We sampled fleas and prairie dogs from six prairie dog colonies in two regions separated by about 46 km. These colonies were extirpated by a plague epizootic that began months after our sampling was completed in 2005. Prairie dogs showed significant isolation‐by‐distance and a tendency toward genetic structure on the regional scale that the fleas did not. Fleas exhibited higher estimated rates of gene flow among prairie dog colonies than the prairie dogs sampled from the same colonies. While the findings suggested black‐tailed prairie dogs may have contributed to flea dispersal, we attributed the lack of concordance between the population genetic structures of host and ectoparasite to additional flea dispersal that was mediated by mammals other than prairie dogs that were present in the prairie system.

Genetic Delineation of Management Units for the Desert Tortoise, Gopherus agassizii, in Northeastern Mojave Desert

Blood samples from 236 desert tortoises, Gopherus agassizii, representing 15 sites in northeastern Mojave Desert (Nevada, Utah), were assayed for variation in allozymes and mitochondrial DNA restriction sites. Morphometric measurements from 257 tortoises from the same locations were subjected to sheared principal component analysis. The 15 localities were partitioned into five management units. Geographic concordance between the five management units delineated in this study and proposed Desert Wildlife Management Areas (DWMAs), delineated ecologically, was assessed. Suggestions for improving placement and boundaries of DWMAs are discussed. THE desert tortoise, Gopherus agassizii, is one of four extant species of testudinid tortoises found in desert and semitropical regions of North America. Its range includes the Mojave Desert of southern Nevada, California, and Utah and the Sonoran Desert of western Arizona, western Sonora, Mexico, and northern Sinaloa, Mexico (Lamb et al., 1989). Beginning in the early 1970s, declines of from 3-59% in desert tortoise populations were noted over much of its range in the United States (USFWS Recovery Plan, 1994, unpubl.). De

Sylvatic plague reduces genetic variability in black-tailed prairie dogs.

Small, isolated populations are vulnerable to loss of genetic diversity through inbreeding and genetic drift. Sylvatic plague due to infection by the bacterium Yersinia pestis caused an epizootic in the early 1990s resulting in declines and extirpations of many black-tailed prairie dog (Cynomys ludovicianus) colonies in north-central Montana, USA. Plague-induced population bottlenecks may contribute to significant reductions in genetic variability. In contrast, gene flow maintains genetic variability within colonies. We investigated the impacts of the plague epizootic and distance to nearest colony on levels of genetic variability in six prairie dog colonies sampled between June 1999 and July 2001 using 24 variable randomly amplified polymorphic DNA (RAPD) markers. Number of effective alleles per locus (ne) and gene diversity (h) were significantly decreased in the three colonies affected by plague that were recovering from the resulting bottlenecks compared with the three colonies that did not experience plague. Genetic variability was not significantly affected by geographic distance between colonies. The majority of variance in gene frequencies was found within prairie dog colonies. Conservation of genetic variability in black-tailed prairie dogs will require the preservation of both large and small colony complexes and the gene flow among them.

Nonlethal Tissue Sampling Techniques and Microsatellite Markers Used for First Report of Genetic Diversity in Two Populations of the Endangered Somatochlora hineana (Odonata: Corduliidae)

ABSTRACT Techniques for obtaining DNA noninvasively or nonlethally are highly desirable in molecular genetic studies of protected species, and several advances have been made in these types of sampling and extraction techniques. Insects present a unique set of difficulties in this regard that are not present when working with most vertebrates. This study evaluated the effectiveness of several nonlethal sampling techniques for larval and adults of the federally listed endangered dragonfly Somatochlora hineana (Williamson) (Odonata: Corduliidae). Fecal pellets and shed exuviae from captive S. hineana larvae did not provide high enough quality DNA for microsatellite analyses. Invasive, but nonlethal, wing clips from adults and tarsi from larvae provided high-quality DNA that amplified 10 microsatellite markers for this species. Ten loci were polymorphic in 94 specimens with four to 14 alleles per locus. Two populations in WI had average observed heterozygosity of 0.47, which is within the range reported for other odonates. Our sampling techniques and these new microsatellite markers provide an essential tool for determining the genetic structure of S. hineana populations throughout its range.

Genetic population structure of the Dakota skipper (Lepidoptera: Hesperia dacotae): A North American native prairie obligate

A range-wide survey of Dakotaskipper (Hesperia dacotae) populationsassessed levels of genetic variability andgeographic scale of population structure inthis species of conservation concern. Thisspecies exists on isolated patches of nativetall- and mixed-grass prairie within a highlymodified landscape dominated by agriculture. It has been extirpated from the southernportion of its range and has sufferedrange-wide declines. Nine populations weresampled from western Minnesota, eastern SouthDakota, and southern Manitoba. Starch gelelectrophoresis was used to resolve 21 isozymeloci in 278 Dakota skippers. Dakota skipperpopulations were approximately as variable asother lepidopterans found in isolated habitats. Genetic distances indicated that Manitobapopulations were somewhat distinct from ones inMinnesota and South Dakota. Isolation-by-distance was detected range-wideand among the seven southern-most populations. Genetically effective immigration rates weresmall at both range-wide and regional scalesand effective populations sizes were lowsuggesting that Dakota skipper populations aregenetically isolated from one another, althoughthey were likely more connected in the recentpast. Genotype assignment tests revealed twoclusters of populations in Minnesota and SouthDakota that were not apparent from theisolation-by-distance results. Significantheterozygote deficiencies relative toHardy-Weinberg expectations and high inbreedingcoefficients suggest structure within samplelocations. Management recommendations includethe maximization of effective population sizein each Dakota skipper population to offset theeffects of drift and habitat corridors in somecases. Habitat management should consider thewithin-site population structure and possibletemporal population structure detected in thisstudy.

Conservation in Hine’s sight: the conservation genetics of the federally endangered Hine’s emerald dragonfly, Somatochlora hineana

Hine’s emerald dragonfly (Somatochlora hineana) is distributed in discrete fen and wet meadow habitats over its range from Ontario, Canada, to Missouri, USA. Habitat destruction in the vicinity of Chicago, IL, and other areas lead to its designation as an US federal endangered species in 1995. Our main goal was to delineate the population genetic structure of the species within the northern recovery unit centered on the Door Peninsula in Wisconsin and the southern recovery unit in the Des Plaines River Valley near Chicago, IL. Sites on the Door Peninsula, WI, are in a matrix of agricultural development and second-growth forest and were used as a best available approximation of a pristine system for the dragonfly. We nondestructively sampled 557 adults and larvae from 16 sites in Illinois, Michigan, and Wisconsin from 2008 through 2011 and used ten microsatellite markers to estimate levels of genetic variability, and genetic structure. Mean allelic richness across all sites and years was 5.03 (±0.64) and expected heterozygosity was 0.52 (±0.032). Northern and southern recovery units as designated in the original recovery plan were genetically distinct. We delineated two genetic populations in the northern unit and three within the southern including two disjunct sites.

Deltamethrin flea-control preserves genetic variability of black-tailed prairie dogs during a plague outbreak

Genetic variability and structure of nine black-tailed prairie dog (BTPD, Cynomys ludovicianus) colonies were estimated with 15 unlinked microsatellite markers. A plague epizootic occurred between the first and second years of sampling and our study colonies were nearly extirpated with the exception of three colonies in which prairie dog burrows were previously dusted with an insecticide, deltamethrin, used to control fleas (vectors of the causative agent of plague, Yersinia pestis). This situation provided context to compare genetic variability and structure among dusted and non-dusted colonies pre-epizootic, and among the three dusted colonies pre- and post-epizootic. We found no statistical difference in population genetic structures between dusted and non-dusted colonies pre-epizootic. On dusted colonies, gene flow and recent migration rates increased from the first (pre-epizootic) year to the second (post-epizootic) year which suggested dusted colonies were acting as refugia for prairie dogs from surrounding colonies impacted by plague. Indeed, in the dusted colonies, estimated densities of adult prairie dogs (including dispersers), but not juveniles (non-dispersers), increased from the first year to the second year. In addition to preserving BTPDs and many species that depend on them, protecting colonies with deltamethrin or a plague vaccine could be an effective method to preserve genetic variability of prairie dogs.

Detections of Yersinia pestis East of the Known Distribution of Active Plague in the United States

We examined fleas collected from black-tailed prairie dog (Cynomys ludovicianus) burrows from 2009 through 2011 in five national park units east of the known distribution of active plague across the northern Great Plains for the presence of Yersinia pestis. Across all national park units, Oropsylla tuberculata and Oropsylla hirsuta were the most common fleas collected from prairie dog burrows, 42.4% and 56.9%, respectively, of the 3964 fleas collected from burrow swabbing. Using a nested PCR assay, we detected 200 Y. pestis-positive fleas from 3117 assays. In total, 6.4% of assayed fleas were Y. pestis positive and 13.9% of prairie dog burrows swabbed contained Y. pestis-positive fleas. Evidence of the presence of Y. pestis was observed at all national park units except Devils Tower National Monument in Wyoming. We detected the presence of Y. pestis without large die-offs, i.e., enzootic sylvatic plague, east of the known distribution of active plague and near the eastern edge of the present distribution of black-tailed prairie dogs. This study, in combination with previous work suggests that sylvatic plague likely occurs across the range of black-tailed prairie dogs and should now be treated as endemic across this range.

Effects of Host-Plant Distribution on Genetic Structuring of Two Tortoise Beetles (Coleoptera: Chrysomelidae: Cassidinae) in the Northern Great Plains

Abstract Physonota helianthi Randall feeds exclusively on a native sunflower, Helianthus grosseserratus Martens, in southeastern South Dakota. Helianthus grosseserratus populations are highly fragmented by agriculture and their patchy distribution presents the hypothesis that host-plant fragmentation results in genetic structuring in P. helianthi. In contrast, another cassidine beetle, Chelymorpha cassidea F., feeds on common bindweeds (Convolvulus arvensis L. and Calystegia sepium (L.) R. Br.) and therefore may not exhibit such genetic structuring. We sampled patches of host plants for P. helianthi and C. cassidea. Families were reared in the laboratory through adult emergence and assayed with starch-gel electrophoresis. Genetic variability within and among beetle families from different patches was compared for both species. For P. helianthi, 15 loci were used to evaluate allele frequencies in 30 families from five sites. For C. cassidea, we assayed 13 loci from 25 families from five sites. We determined the relative magnitude of genetic variance within a hierarchy that included families, patches, and total sample. For P. helianthi, slightly more genetic variation existed within families from the same patch than among families from different patches, but overall, variability was low (x̄ heterozygosity = 0.048). Paternity of individual egg masses was more than one in most cases. Chelymorpha cassidea was more variable (x̄ heterozygosity = 0.072) but showed less differentiation among patches. Multiple paternity was less frequent in this species than in P. helianthi. Differences in genetic structuring of these two species are likely because of their contrasting host-plant distributions and mating structure.

Characterization and Isolation of Five Microsatellite Loci in Northern Flying Squirrels, Glaucomys sabrinus (Sciuridae, Rodentia)

ABSTRACT. Northern flying squirrels (Glaucomys sabrinus) are found in boreal forests of northern and northwestern North America, but a small population is isolated to the Black Hills of South Dakota and Wyoming. Few micro satellite primers have been developed for this species, though they are needed to examine the genetic structure of these populations. Thus, we isolated and characterized 5 micro satellite loci in northern flying squirrels through a series of steps involving microbiology, molecular biology, and genetic techniques. Data analyses with these primers indicated that the northern flying squirrel population found in the Black Hills may have low heterozygosity and significant departure from Hardy-Weinberg equilibrium. The development of these primers not only provides additional data for analyzing a small, disjunct population but also serves as a mechanism for understanding population dynamics and assisting with overall management and conservation of unique populations.