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USGS2007

Genetic studies of freshwater turtle and tortoises: a review of the past 70 years

Powerful molecular techniques have been developed over many decades for resolving genetic relationships, population genetic structure, patterns of gene flow, mating systems, and the amount of genetic diversity in animals. Genetic studies of turtles were among the earliest and the rapid application of new genetic tools and analytical techniques is still apparent in the literature on turtles. At present, of the 198 freshwater turtles and tortoises that are listed as not extinct by the IUCN Red List, 69 species worldwide are listed as endangered or critically endangered, and an additional 56 species are listed as vulnerable. Of the ca. 300 species of the freshwater turtles and tortoises in the world, ca. 42% are considered to be facing a high risk extinction, and there is a need to focus intense conservation attention on these species. This includes a need to (i) assess our current state of knowledge regarding the application of genetics to studies of freshwater turtles and tortoises and (ii) determine future research directions. Here, we review all available published studies for the past 70 years that were written in English and used genetic markers (e.g. karyotypes, allozymes, DNA loci) to better understand the biology of freshwater turtles and tortoises. We review the types of studies conducted in relation to the species studied and quantify the countries where the studies were performed. We rack the changing use of different genetic markers through time and report on studies focused on aspects of molecular evolution within turtle genomes. We address the usefulness of particular genetic markers to answer phylogenetic questions and present data comparing population genetic structure and mating systems across species. We draw specific attention to whether authors have considered issues to turtle conservation in their research or provided new insights that have been translated into recommendations for conservation management.

Conference Paper↗

USGS2005

Rocky Mountain Center for Conservation Genetics and Systematics

The use of molecular genetic tools has become increasingly important in addressing conservation issues pertaining to plants and animals. Genetic information can be used to augment studies of population dynamics and population viability, investigate systematic, refine taxonomic definitions, investigate population structure and gene flow, and document genetic diversity in a variety of plant and animal species. Further, genetic techniques are being used to investigate mating systems through paternity analysis, and analyze ancient DNA samples from museum specimens, and estimate population size and survival rates using DNA as a unique marker. Such information is essential for the sound management of small, isolated populations of concern and is currently being used by universities, zoos, the U.S. Fish and Wildlife Service, and numerous state fish and wildlife agencies.

Fact Sheet↗

USGS2005

Bison PRNP genotyping and potential association with Brucella spp. seroprevalence

The implication that host cellular prion protein (PrPC) may function as a cell surface receptor and/or portal protein for Brucella abortus in mice prompted an evaluation of nucleotide and amino acid variation within exon 3 of the prion protein gene (PRNP) for six US bison populations. A non-synonymous single nucleotide polymorphism (T50C), resulting in the predicted amino acid replacement M17T (Met ??? Thr), was identified in each population. To date, no variation (T50: Met) has been detected at the corresponding exon 3 nucleotide and/or amino acid position for domestic cattle. Notably, 80% (20 of 25) of the Yellowstone National Park bison possessing the C/C genotype were Brucella spp. seropositive, representing a significant (P = 0.021) association between seropositivity and the C/C genotypic class. Moreover, significant differences in the distribution of PRNP exon 3 alleles and genotypes were detected between Yellowstone National Park bison and three bison populations that were either founded from seronegative stock or previously subjected to test-and-slaughter management to eradicate brucellosis. Unlike domestic cattle, no indel polymorphisms were detected within the corresponding regions of the putative bison PRNP promoter, intron 1, octapeptide repeat region or 3???-untranslated region for any population examined. This study provides the first evidence of a potential association between nucleotide variation within PRNP exon 3 and the presence of Brucella spp. antibodies in bison, implicating PrPC in the natural resistance of bison to brucellosis infection. ?? 2005 International Society for Animal Genetics.

Idaho, Montana, New Mexico, North Dakota, South Da↗

USGS2009

Kappa-casein polymorphisms among cattle breeds and bison herds

We identified the Hind III restriction site polymorphism of K-casein in cattle reported by Pinder et al. ( Animal Genetics 22, 11, 1991) and found an additonal polymorphism ( Rsa I) in cattle and bison. The Hin dIII and Rsa I restriction sites were mapped and three haplotypes (alleles) were identified. Preliminary screening of 39 cattle and 71 bison revealed one allele restricted to cattle, one restricted to bison, and one shared by the species. No fixed allelic differences were observed among cattle breeds or among bison herds or subspecies.

Animal Genetics↗

USGS2021

Macrogenetic studies must not ignore limitations of genetic markers and scale

Millette et al . (Ecology Letters, 2020, 23:55–67) reported no consistent worldwide anthropogenic effects on animal genetic diversity using repurposed mitochondrial DNA sequences. We reexamine data from this study, describe genetic marker and scale limitations which might lead to misinterpretations with conservation implications, and provide advice to improve future macrogenetic studies.

Ecology Letters↗

USGS2019

Within-group relatedness and patterns of reproductive sharing and cooperation in the tropical chestnut-crested yuhina

In cooperatively breeding animals, genetic relatedness among group members often determines the extent of reproductive sharing, cooperation and competition within a group. Studies of species for which cooperative behaviour is not entirely based on kinship are key for understanding the benefits favouring the evolution and maintenance of cooperative breeding among nonrelatives. In the cooperatively breeding chestnut-crested yuhina, Yuhina everetti , a songbird endemic to Borneo, we tested whether unrelated helpers are more likely to gain parentage than are related helpers consistent with the hypothesis that inbreeding risk constrains reproduction by related helpers. We also examined whether related or unrelated helpers provision broods more because of differences in their potential indirect or direct fitness benefits of helping. Kin structure of breeding groups (breeding pair and up to eight helpers of both sexes, median = 2 helpers, 96% of 57 pairs had helpers) based on genetic analysis was mixed; 48% of 76 breeder/helper dyads were first-order (26%) or second-order (22%) relatives of one or both members of the breeding pair, and 52% were nonrelatives. Only unrelated male and female helpers gained parentage, and helpers did not differ in their provisioning rate according to their relatedness to the broods. We documented quasi-parasitism or co-breeding by female helpers in 14% of 29 broods and extrapair paternity by male helpers in 21% of 47 broods. This rate of extrapair paternity is relatively high among the few tropical species examined but fit with predictions for mixed-kin groups where inbreeding is avoided. These findings support the emerging pattern for cooperative breeding in birds with mixed-kin groups, wherein unrelated helpers are more likely to gain parentage than are related helpers and helping effort is not necessarily predicted by kinship.

Animal Behaviour↗

USGS2015

Evaluation of fisher (Pekania pennanti) restoration in Olympic National Park and the Olympic Recovery Area: 2014 annual progress report

With the translocation and release of 90 fishers ( Pekania pennanti ) from British Columbia to Olympic National Park during 2008–2010, the National Park Service and Washington Department of Fish and Wildlife accomplished the first phase of fisher restoration in Washington State. Beginning in 2013, we initiated a new research project to determine the current status of fishers on Washington’s Olympic Peninsula 3–5 years after the releases and evaluate the short-term success of the restoration program. Objectives of the study are to determine the current distribution of fishers and proportion of the recovery area that is currently occupied by fishers, determine several genetic characteristics of the reintroduced population, and determine reproductive success of the founding animals through genetic studies. During 2014, we continued working with a broad coalition of cooperating agencies, tribes, and nongovernmental organizations (NGO) to collect data on fisher distribution and genetics using noninvasive sampling methods. The primary sampling frame consisted of 157 24-square-kilometer hexagons (hexes) distributed across all major land ownerships within the Olympic Peninsula target survey area. In 2014 we expanded the study by adding 58 more hexes to an expanded study area in response to incidental fisher observations outside of the target area obtained in 2013; 49 hexes were added south and 9 to the east of the target area. During 2014, federal, state, tribal and NGO biologists and volunteers established three baited motion-sensing camera stations, paired with hair snaring devices, in 80 hexes; 69 in the targeted area 11 in the expansion areas. Each paired camera/hair station was left in place for approximately 6 weeks, with three checks on 2-week intervals. We documented fisher presence in 5 of the 80 hexagons, and identified 5 different fishers through a combination of microsatellite DNA analyses and camera detections. All fisher detections were in the target area. These 5 individuals included 2 of the original founding population of 90, 1 of the 2 rescued and rehabilitated kits that were released in 2010, and 1 new recruit to the population (1 individual was not identified). Additionally, we identified more than 40 other species of wildlife at the baited camera stations. We also obtained eight incidental fisher observations through photographs and carcass retrieval. During 2015, we plan to sample 75 hexagons in the target area and 12 in the expansion area. We plan to sample all unsampled accessible hexes in the target area (26 hexes), and re-sample accessible hexes sampled in 2013 (49 hexes).

Washington↗

USGS2016

Evaluation of fisher (Pekania pennanti) restoration in Olympic National Park and the Olympic Recovery Area: 2015 final annual progress report

With the translocation and release of 90 fishers (Pekania pennanti) from British Columbia to Olympic National Park during 2008–2010, the National Park Service (NPS) and Washington Department of Fish and Wildlife (WDFW) accomplished the first phase of fisher restoration in Washington State. Beginning in 2013, we initiated a new research project to determine the current status of fishers on Washington’s Olympic Peninsula 3–8 years after the releases and evaluate the short-term success of the restoration program. Objectives of the study are to determine the current distribution of fishers and proportion of the recovery area that is currently occupied by fishers, determine several genetic characteristics of the reintroduced population, and determine reproductive success of the founding animals through genetic studies. During 2015, we continued working with a broad coalition of cooperating agencies, tribes, and nongovernmental organizations (NGO) to collect data on fisher distribution and genetics using noninvasive sampling methods. The primary sampling frame consisted of 157 24-km2 hexagons (hexes) distributed across all major land ownerships within the Olympic Peninsula target survey area. In 2014 we expanded the study by adding 58 more hexes to an expanded study area in response to incidental fisher observations outside of the target area obtained in 2013; 49 hexes were added south and 9 to the east of the target area. During 2015, Federal, State, Tribal and NGO biologists and volunteers established three Distributioned motion-sensing camera stations, paired with hair snaring devices, in 87 hexes; 75 in the targeted area and 12 in the expansion areas. Each paired camera/hair station was left in place for approximately 6 weeks, with three checks on 2-week intervals. We documented fisher presence in 7 of the 87 hexagons. Four fishers were identified through microsatellite DNA analyses. The 4 identified fishers included 1 of the original founding population of 90 and 3 new recruits to the population. Three additional fishers were detected with cameras but not DNA, consequently their identities were unknown. All fisher detections were in the target area. Additionally, we identified 46 other species of wildlife at the baited camera stations. We also obtained 4 additional confirmed records of fishers in the study area through photographs provided by the public and incidental live capture. During 2016, we plan to resample 69 hexagons sampled in the target area in 2014 and 12 new hexes in the expansion area. In addition, we plan to sample non-selected hexes in-between hexes where we had a cluster of fishers in 2014, to provide better understanding of occupancy patterns and minimum number of individuals in an area where fishers appear to be concentrating.

Washington↗

USGS2003

Dinucleotide repeat polymorphisms in waterfowl (family Anatidae): Characterization of a sex-linked (Z-specific) and 14 autosomal loci

Canada goose ( Branta Canadensis ) and harlequin duck ( Histrionicus histrionicus ) DNAs were digested with Sau 3AI, and size selected (300-700 bp) fragments were ligated into Bam HI-digested pBluscriptII KS + . The enrichment protocol of Ostrander et al . 1 was followed. The resulting libraries were screened using a [ƴ- 32 P]ATP end-labelled (CA) 20 oligonucleotides as a hybridization probe. Positive clones were sequenced using cycle-sequencing protocols (Epicentre Technologies, Madison, WI) and primers flanking the inserts. PCR primers were designed to amplify the repeat and yield amplification products of ≈100-200 bp. DNA samples were screened for variation at these loci using [ƴ- 32 P]ATP end-labelled primers. The products were resolved using 6% denaturing polyacrylamide gels and autoradiography.

Animal Genetics↗

USGS1987

Genetic management of endangered species at the Patuxent Wildlife Research Center

Summary: The Patuxent Wildlife Research Center conducts one of the world's largest and best-known research programs for captive propagation of endangered wildlife. In order to be effective and to ensure the long-term survival of species, researchers at Patuxent attempt to manage captive populations according to the principles of population genetics. This includes the use of estimated inbreeding levels for mate selections in Masked Bobwhites and biochemical analyses to measure extant genetic material and determine relationships among Whooping Cranes. As added insurance against catastrophic losses, or even random losses of key individuals representing unique lineages, cryopreservation of semen has been studied and used for some species. Artificial insemination, using either stored or fresh semen, is used to improve fertility rates, thereby increasing the chances for survival of unique genetic lines. Finally, a periodic influx of unrelated stock occurs, when feasible, in order to enhance the genetic base of captive populations. The application of these techniques will ensure that future releases utilize genetically viable animals, thereby improving the potential for successful reintroductions into the wild.

Book chapter↗

USGS1995

Biodiversity: a new challenge

Resource managers at many state and federal agencies are in the middle of a fundamental change in the practice and objectives of conservation. Traditional management has been directed toward maintaining, usually for harvest purposes, populations of individual species such as ducks, deer, or salmon. Increasingly, however, resource managers are recognizing the critical important of conserving biological diversity, or biodiversity. In its simplest terms, biological diversity is the variety of life at all levels: it includes the array of plants and animals; the genetic differences among individuals; the communities, ecosystems, and landscapes in which they occur; and the variety of processes on which they depend. Conserving biological diversity poses dramatic new problems for comprehensive inventory and monitoring: what should be measured or monitored?

Book chapter↗

USGS2018

Isolation by distance versus landscape resistance: Understanding dominant patterns of genetic structure in Northern Spotted Owls (Strix occidentalis caurina)

Landscape genetics investigations examine how the availability and configuration of habitat influence genetic structure of plants and animals. We used landscape genetics to evaluate the role that forest connectivity plays in determining genetic structure of the federally-threatened Northern Spotted Owl ( Strix occidentalis caurina ) using genotypes of 339 Northern Spotted Owls obtained for 10 microsatellite loci. Spatial clustering analyses identified a distinct genetic cluster at the southern extent of the region examined. This cluster could not be linked to landscape connectivity patterns and suggested that post-Pleistocene processes were involved with its development rather than contemporary landscape configuration. We also compared matrices of pairwise inter-individual genetic distances with resistance distances derived from a circuit-theory based framework. Resistance distances were obtained for an idealized raster map that reflected continuous unimpeded dispersal habitat across the landscape along with five empirically-derived raster maps reflecting the 1870’s, 1940’s, 1986, 1994, and 2012. Resistance distances from the idealized map served as surrogates for linear geographic distances. Relative to idealized conditions, resistance distances were ~250% higher in the 1940’s and ~200% higher from 1986 onward. Resistance distances from the 1870’s were ~40% higher than idealized conditions. Inter-individual genetic distances were most highly correlated with resistance distances from the idealized map rather than any of the empirical maps. Two hypotheses explain our results. First, our results may reflect temporal lags between the onset of large-scale habitat alterations and their novel effects on genetic structure in long-lived species such as Northern Spotted Owls. Second, because Northern Spotted Owls disperse over long distances, our results may indicate that forest habitat has never been sufficiently fragmented to the point where connectivity was disrupted. The second hypothesis could indicate that forest management practices mandated by the Northwest Forest Plan succeeded with one of its primary goals. However, our results do not represent a complete portrayal of the status of Northern Spotted Owls given detection of significant population declines and bottlenecks in other studies. Future investigations based on computer simulations may help distinguish between hypotheses.

California, Oregon, Washington↗

USGS2012

The genetic structure of a relict population of wood frogs

Habitat fragmentation and the associated reduction in connectivity between habitat patches are commonly cited causes of genetic differentiation and reduced genetic variation in animal populations. We used eight microsatellite markers to investigate genetic structure and levels of genetic diversity in a relict population of wood frogs ( Lithobates sylvatica ) in Rocky Mountain National Park, Colorado, where recent disturbances have altered hydrologic processes and fragmented amphibian habitat. We also estimated migration rates among subpopulations, tested for a pattern of isolation-by-distance, and looked for evidence of a recent population bottleneck. The results from the clustering algorithm in Program STRUCTURE indicated the population is partitioned into two genetic clusters (subpopulations), and this result was further supported by factorial component analysis. In addition, an estimate of FST (FST = 0.0675, P value \0.0001) supported the genetic differentiation of the two clusters. Estimates of migration rates among the two subpopulations were low, as were estimates of genetic variability. Conservation of the population of wood frogs may be improved by increasing the spatial distribution of the population and improving gene flow between the subpopulations. Construction or restoration of wetlands in the landscape between the clusters has the potential to address each of these objectives.

Colorado↗

USGS2005

Influence of barriers to movement on within-watershed genetic variation of coastal cutthroat trout

Because human land use activities often result in increased fragmentation of aquatic and terrestrial habitats, a better understanding of the effects of fragmentation on the genetic heterogeneity of animal populations may be useful for effective management. We used eight microsatellites to examine the genetic structure of coastal cutthroat trout (Oncorhynchus clarki clarki) in Camp Creek, an isolated headwater stream in western Oregon. Our objectives were to determine if coastal cutthroat trout were genetically structured within streams and to assess the effects of natural and anthropogenic barriers on coastal cutthroat trout genetic variation. Fish sampling occurred at 10 locations, and allele frequencies differed significantly among all sampling sections. Dispersal barriers strongly influenced coastal cutthroat trout genetic structure and were associated with reduced genetic diversity and increased genetic differentiation. Results indicate that Camp Creek coastal cutthroat trout exist as many small, partially independent populations that are strongly affected by genetic drift. In headwater streams, barriers to movement can result in genetic and demographic isolation leading to reduced coastal cutthroat trout genetic diversity, and potentially compromising long-term population persistence. When habitat fragmentation eliminates gene flow among small populations, similar results may occur in other species.

Oregon↗

USGS2017

Using high-throughput DNA sequencing, genetic fingerprinting, and quantitative PCR as tools for monitoring bloom-forming and toxigenic cyanobacteria in Upper Klamath Lake, Oregon, 2013 and 2014

Monitoring the community structure and metabolic activities of cyanobacterial blooms in Upper Klamath Lake, Oregon, is critical to lake management because these blooms degrade water quality and produce toxic microcystins that are harmful to humans, domestic animals, and wildlife. Genetic tools, such as DNA fingerprinting by terminal restriction fragment length polymorphism (T-RFLP) analysis, high-throughput DNA sequencing (HTS), and real-time, quantitative polymerase chain reaction (qPCR), provide more sensitive and rapid assessments of bloom ecology than traditional techniques. The objectives of this study were (1) to characterize the microbial community at one site in Upper Klamath Lake and determine changes in the cyanobacterial community through time using T-RFLP and HTS in comparison with traditional light microscopy; (2) to determine relative abundances and changes in abundance over time of toxigenic Microcystis using qPCR; and (3) to determine relative abundances and changes in abundance over time of Aphanizomenon , Microcystis , and total cyanobacteria using qPCR. T-RFLP analysis of total cyanobacteria showed a dominance of only one or two distinct genotypes in samples from 2013, but results of HTS in 2013 and 2014 showed more variations in the bloom cycle that fit with the previous understanding of bloom dynamics in Upper Klamath Lake and indicated that potentially toxigenic Microcystis was more prevalent in 2014 than in years prior. The qPCR-estimated copy numbers of all target genes were higher in 2014 than in 2013, when microcystin concentrations also were higher. Total Microcystis density was shown with qPCR to be a better predictor of late-season increases in microcystin concentrations than the relative proportions of potentially toxigenic cells. In addition, qPCR targeting Aphanizomenon at one site in Upper Klamath Lake indicated a moderate bloom of this species (corresponding to chlorophyll a concentrations between approximately 75 and 200 micrograms per liter) from mid-June to mid-August, 2014. After August 18, the Aphanizomenon bloom was overtaken by Microcystis late in the season as microcystin concentrations peaked. Overall, results of this study showed how DNA-based, genetic methods may provide rapid and sensitive diagnoses for the presence of toxigenic cyanobacteria and that they are useful for general monitoring or ecological studies and identification of cyanobacterial community members in complex aquatic habitats. These same methods can also be used to simultaneously address spatial (horizontal and vertical) and temporal variation and under different conditions. Additionally, with some modifications, the same techniques can be applied to different sample types, including water, sediment, and tissue.

Oregon↗

USGS2005

Comparison of Bacteroides-Prevotella 16S rRNA genetic markers for fecal samples from different animal species

To effectively manage surface and ground waters it is necessary to improve our ability to detect and identify sources of fecal contamination. We evaluated the use of the anaerobic bacterial group Bacteroides-Prevotella as a potential fecal indicator. Terminal restriction length polymorphism (T-RFLP) of the 16S rRNA genes from this group was used to determine differences in populations and to identify any unique populations in chickens, cows, deer, dogs, geese, horses, humans, pigs, and seagulls. The group appears to be a good potential fecal indicator in all groups tested except for avians. Cluster analysis of Bacteroides-Prevotella community T-RFLP profiles indicates that Bacteroides-Prevotella populations from samples of the same host species are much more similar to each other than to samples from different source species. We were unable to identify unique peaks that were exclusive to any source species; however, for most host species, at least one T-RFLP peak was identified to be more commonly found in that species, and a combination of peaks could be used to identify the source. T-RFLP profiles obtained from water spiked with known-source feces contained the expected diagnostic peaks from the source. These results indicate that the approach of identifying Bacteroides-Prevotella molecular markers associated with host species might be useful in identifying sources of fecal contamination in the environment.

Applied and Environmental Microbiology↗

USGS2022

Scat as a source of DNA for population monitoring

Sampling fecal droppings (scat) to genetically identify individual animals is an established method for monitoring mammal populations and could be highly useful for monitoring reptile populations. Whereas existing protocols for obtaining DNA from reptile scat focus on analyses of whole, fresh scat deposited during animal handling, the collection of scat naturally deposited by reptiles in situ, as required for non-invasive population monitoring, requires protocols to extract highly degraded DNA. Using surface swabs from such scats can reduce PCR inhibition, ecological impacts of removing scat, and zoonotic risks. We report on three related but independently designed studies of DNA analyses from scat swabs of herbivorous reptiles under natural desert conditions: two free-ranging desert tortoise species (Agassiz's desert tortoise, Gopherus agassizii , California, US, and Morafka's desert tortoise, G. morafkai , Arizona, US) and the common chuckwalla (Sauromalus atar ) (Arizona, US, and Sonora, MX). We analyzed samples from both tortoise species with the same set of 16 microsatellites and chuckwalla samples with four mtDNA markers; studies also varied in swab preservation medium and DNA extraction method. Microsatellite amplification success per sample, defined as ≥9 loci with amplification, varied between studies, with 15% for Agassiz's desert tortoise and 42% Morafka's desert tortoise. For chuckwallas, we successfully amplified and sequenced 50% of samples. We recovered fragments up to 400 bp for tortoises and 980 bp for chuckwallas from scat swab samples. This study demonstrates that genotypes can successfully be obtained from swabs of herbivorous reptile scat collected in the field under natural environmental conditions and emphasizes that repeat amplifications are necessary for genetic identification of individuals from non-invasive samples.

Arizona, California↗

USGS2006

Population structure, persistence, and seasonality of autochthonous Escherichia coli in temperate, coastal forest soil from a Great Lakes watershed

The common occurrence of Escherichia coli in temperate soils has previously been reported, however, there are few studies to date to characterize its source, distribution, persistent capability and genetic diversity. In this study, undisturbed, forest soils within six randomly selected 0.5 m2 exclosure plots (covered by netting of 2.3 mm2 mesh size) were monitored from March to October 2003 for E. coli in order to describe its numerical and population characteristics. Culturable E. coli occurred in 88% of the samples collected, with overall mean counts of 16 MPN g-1, ranging from <1 to 1657 (n = 66). Escherichia coli counts did not correlate with substrate moisture content, air, or soil temperatures, suggesting that seasonality were not a strong factor in population density control. Mean E. coli counts in soil samples (n = 60) were significantly higher inside than immediately outside the exclosures; E. coli distribution within the exclosures was patchy. Repetitive extragenic palindromic polymerase chain reaction (Rep-PCR) demonstrated genetic heterogeneity of E. coli within and among exclosure sites, and the soil strains were genetically distinct from animal (E. coli) strains tested (i.e. gulls, terns, deer and most geese). These results suggest that E. coli can occur and persist for extended periods in undisturbed temperate forest soils independent of recent allochthonous input and season, and that the soil E. coli populations formed a cohesive phylogenetic group in comparison to the set of fecal strains with which they were compared. Thus, in assessing E. coli sources within a stream, it is important to differentiate background soil loadings from inputs derived from animal and human fecal contamination. ?? 2005 Society for Applied Microbiology and Blackwell Publishing Ltd.

Environmental Microbiology↗