Conservation Genetics

A carnivores' oasis? An isolated fisher ( Pekania pennanti ) population provides insight on persistence of a metapopulation Abstract Landscape level species assessments are rarely available to inform conservation planning. Recent advances in multi-taxa survey techniques, however, have made them more obtainable by improving efficiency of collecting species occurrence data and genetic samples. Here, we used a multi-taxa survey technique to provide a landscape level portrait of fisher (Pekania pennanti) population status in a portion of the northern Rockies where the species' history has been heavily influenced by humans. From 2010 to 2014 we deployed 497 winter forest carnivore bait stations across a 23,000 km2 study area centered on northern Idaho, United States. The stations collected remote camera images and hair samples for DNA analysis. We used 12 microsatellite loci to identify 58 individual fisher in the study area. We identified two fisher populations for which we estimated effective population size (Ne) and genetic neighborhood size (NS). We calculated local genetic diversity measures [observed (Ho) and expected (He) heterozygosity, allelic richness (Ar), fixation index (FST), and number of migrants per generation (Nm)] and mapped continuous gradients of genetic diversity [Ar and inbreeding coefficient (FIS)] across the study area. We identified a fisher population in the West Cabinet Mountains [Ne of 26 (15.3–55.5), NS of 50–60] which is effectively isolated from the small population we detected in the Saint Joe/Coeur d'Alene Mountains [Ne of 6.5 (2.7–15), NS of 10–25]. We determined fisher have been effectively extirpated from the Selkirk and Purcell mountains and the lack of genetic connectivity between the two small remaining populations casts doubt on long term viability. Our study suggests northern Rockies fisher populations are vulnerable to landscape barriers and successful recovery will depend on future management and augmentation. We recommend these factors be considered when determining how, and if, species recovery should proceed.

Lack of spatial and temporal genetic structure of Japanese eel ( Anguilla japonica ) populations Abstract Japanese eel (Anguilla japonica) is an important food source in East Asia whose population has dramatically declined since the 1970s. Despite past analysis with DNA sequencing, microsatellite and isozyme methods, management decisions remain hampered by contradictory findings. For example, it remains unresolved whether Japanese eels are a single panmictic population or whether they harbor significant substructure. Accurate assessment of population genetic substructure, both spatial and temporal, is essential for determining the relevant number of distinct management units appropriate for this species. In the present study, we assayed genetic variation genome-wide using Restriction Site Associated DNA Sequencing (RAD-seq) technology to analyze the population genetic structure of Japanese eels. For analysis of temporal isolation, five "cohort" samples were collected yearly from 2005 to 2009 in the Yangtze River Estuary. For analysis of spatial structure, five "arrival wave" samples were collected in China in 2009, and two arrival wave samples were collected in Japan in 2001. In each cohort of each arrival wave, five individuals were collected for a total of 55 eels sampled. In total, 214,210 loci were identified from these individuals, 106,652 of which satisfied quality checks and were retained for further analysis. There was relatively little population differentiation between arrival waves and cohorts collected either at different locations during the same year (Fst = 0.077) or at the same location collected over subsequent years (Fst = 0.082), and locations displayed no consistent isolation-by-distance.

Assessing the contribution of aquaculture and restoration to wild oyster populations in a Rhode Island coastal lagoon Abstract The decline of the eastern oyster (Crassostrea virginica) has prompted various restoration and aquaculture efforts. Recent field surveys in Rhode Island suggest that wild populations are increasing, yet the factors contributing to expansion are unknown. We used a population genetic approach to characterize genetic differences between wild and cultured oyster populations and explore the extent of connectivity and admixture between groups. Individual oysters from four wild, three farmed, and two restored populations were collected within or just outside Ninigret Pond, a coastal lagoon highly influenced by human activity, and genotyped at 13 microsatellite loci. Results from the multi-locus genotype data showed that wild populations were more genetically diverse than the cultured populations. We also observed significant genetic differentiation between paired wild and cultured populations but not between pairs of wild populations. A cluster analysis detected substantial admixture between wild and cultured groups. As oyster aquaculture and restoration activities are forecasted to increase in the future, this study highlights the potential degree of genetic introgression between remnant wild populations and less diverse, hatchery-reared stocks. Those tasked with preserving our living natural resources should carefully consider how the juxtaposition of aquaculture, restored, and wild populations at small spatial scales will impact the genetic composition and evolutionary trajectories of species in decline for generations to come.

Distribution, fine-scale subdivision, and population size of San Joaquin kit foxes in the Ciervo-Panoche Natural Area, California Abstract The endangered San Joaquin kit fox (SJKF; Vulpes macrotis mutica) is strongly linked ecologically to xeric areas with high abundance of kangaroo rats. Endemic to the San Joaquin Desert of central California, the elusive nature of SJKF, coupled with steady habitat loss and lack of comprehensive surveys, has precluded efforts to quantify the species' population size and distribution, especially in the central and northern parts of its range. Because the Ciervo-Panoche Natural Area contains the largest area of high-quality habitat in this central/northern region, we conducted systematic transect surveys for SJKF scats with professionally trained dog-handler teams throughout the area during 2009–2011. We collected almost 600 scats over 473 km of transects, documenting the freshness and location of each scat. Using molecular methods, we identified 93 SJKF individuals (56 males and 37 females) from 332 samples. Half of the individuals carried a mtDNA haplotype with a 16 bp deletion that had not been previously detected in other areas surveyed. Four individuals were recaptured in 2010 and five in 2011, including one female that was captured every year. We documented a unique mtDNA haplotype and more individuals across a wider area of the Ciervo-Panoche Natural Area than expected. Population analyses revealed two distinct subpopulations, with low connectivity between foxes in the Panoche Valley that are separated by hills with unsuitable habitat from those on the adjacent valley floor next to a major Interstate highway (I-5). While individuals detected within 6 km of each other were closely related, overall relatedness within each subpopulation approached zero. Genetic population models indicated a conservative population estimate of 90 kit foxes in total, with 60–90 individuals in the Panoche Valley and 17–27 individuals in the I-5 area. These results will help to inform management of the SJKF and identify areas that may be important to maintaining connectivity between populations.

Genetic diversity and population demography of Geranium soboliferum var. kiusianum : a glacial relict plant in the wetlands of Japan Abstract Geranium soboliferum var. kiusianum and var. hakusanense (Geraniaceae) are believed to be relict plants that migrated southward from the Eurasian continent to the Japanese archipelago during the last glacial period. G. soboliferum var. kiusianum individuals and populations, which occur in wetlands in the Aso-Kuju region of Kyushu Island, are declining as a result of the deteriorating quality of the microhabitat, a process that is associated with the decrease in semi-natural grasslands. Using 10 polymorphic microsatellite markers, we evaluated the genetic diversity and population structure of the G. soboliferum complex, with the aim of understanding the species' genetic characteristics to facilitate efficient conservation. Genetic demographic analysis suggested that Japanese G. soboliferum populations branched off from the Eurasian continental G. soboliferum var. soboliferum during the last glacial period and that, thereafter, the Japanese populations of the plant diverged into two lineages at the beginning of the Holocene epoch. The genetic diversity of G. soboliferum var. kiusianumwas significantly lower than that of var. hakusanense, which is more widely distributed in central Honshu Island (mean Ar = 3.288 vs. 3.830 and He = 0.366 vs. 0.546). Genetic differentiation among populations was significantly higher (mean FST = 0.368 vs. 0.184) in var. kiusianum populations and, notably, exhibited non-significant patterns of isolation by distance, indicating that the populations underwent strong genetic drift independent of adjacent populations. Our findings suggested that the population's isolation, and its associated habitat specialization and limited inter-population gene flow, would have accelerated genetic differentiation in var. kiusianum. Overall, G. soboliferum var. kiusianum, which is genetically unique to wetland habitats in the Aso-Kuju region, merits conservation, and appropriate human intervention and management are critical to maintaining its remaining habitats in semi-natural grasslands.

Conservation genetic analysis of a Central-European range-margin population of the yellow-bellied toad ( Bombina v. variegata ) Abstract The aim of this study was to investigate the genetic structure and diversity of populations of the yellow-bellied toad in Lower Saxony, Germany. These populations provide a good example of a fragmented distribution of amphibian pioneer species on the outer rim of their occurrence. In total, 150 individuals from 11 sites were genotyped using ten highly polymorphic microsatellites. The genetic diversity, as measured by allelic richness and heterozygosity, was lower than that of other European amphibian populations; the individuals could be assigned to five genetic clusters. The average genetic differentiation between populations was very high (mean FST = 0.27) and no migration was detected between the two major populations, which were about 50 km apart. Nonetheless, natural migration as well as translocations between some subpopulations could be detected. In one population, which was introduced in the 1970s, we were able to verify the potential source population of the introduced toads. The results are discussed in light of conservation management strategies applied to this highly endangered species.

Genetic diversity and population structure of Kichulchoia multifasciata in South Korea Abstract Kichulchoia multifasciata, a small-sized loach (Family Cobitidae) inhabiting only the Nakdong River on the Korean Peninsula, is generally found around well-oxygenated minor tributaries. Despite the scarcity, this species is not yet a legally protected species in South Korea, and the spatial pattern of genetic diversity, which is essential to create management strategies, has never been studied. In this study, three mitochondrial loci and ten microsatellites were used to analyze the genetic diversity and the structure among eight K. multifasciata populations collected from different tributaries. K. multifasciata populations showed a high level of mitochondrial variability, with above 0.9 of average haplotype diversity in all three loci analyzed. A substantial microsatellite polymorphism was also found. Weak genetic structuring among populations and isolation by distance were clearly revealed in both mitochondrial and microsatellite data, indicating the high levels of gene flow among the tributaries. However, a few of the populations exhibited a genetic signature that they have experienced the historical size decline. Our data suggest that unequal gene flow might be one of the causes; nevertheless, the possibility of effective population size reduction by other factors could not be completely excluded. Our genetic evidence can be the critical information needed to protect the populations from the genetic erosion in the Nakdong River that has been destroyed and fragmented with increasing anthropogenic interferences.

Combining molecular and incomplete observational data to inform management of southern white rhinoceros ( Ceratotherium simum simum ) Abstract Conservation efforts have preserved the southern white rhinoceros (SWR) in protected areas and have resulted in substantial overall growth in population size, but in small, fragmented populations in which inbreeding is an important risk. However, field observation of breeding often lacks sufficient accuracy to inform translocation strategies that are intended to increase genetic variation. The purpose of this study was to integrate microsatellite genotypes with an incomplete, field-observed pedigree to make inferences about mean kinship and basic demographic data that could be used to inform translocation programmes for SWR in a confined population in Botswana. Using this approach, we identified parents for 29 out of 45 offspring born in the reserve between 1993 and 2013 and detected eight non-breeding bulls with high mean kinship as candidates for translocation. The method also allowed inferences about demographic parameters that could influence the effectiveness of intervention strategies, such as age and timing of reproduction, and natal sex ratios. Importantly, the reproductive dominance of the bulls was not as skewed as expected after the original dominant bull was removed from the population, suggesting that closed populations can maintain multiple, simultaneously breeding males. The genetic data also confirmed that the accuracy of field-based parentage assignment was increased after implementation of an ear-notching programme. This study demonstrates the value of combining genetic information with ongoing surveillance to inform management of threatened populations, and of using mean kinship to inform metapopulation management by identifying candidates for translocation.

Population genetics, speciation, and hybridization in Dicerandra (Lamiaceae), a North American Coastal Plain endemic, and implications for conservation Abstract Understanding patterns of speciation and subsequent gene flow can clarify the evolutionary origins and history of species endemic to a specific geographic region and reveal genetic patterns important for conservation and management of rare species. We chose Dicerandra from the North American Coastal Plain biodiversity hotspot as a model to explore these concepts because of its endemism and the threatened status of most of its species. Using microsatellite-based population-level analyses of 32 populations from four of the annual species (D. linearifolia var. linearifolia, D. linearifolia var. robustior, D. fumella, D. odoratissima, and D. radfordiana), we addressed questions of genetic diversity, population structure, and hybridization. Strong support was found for the species-level recognition of the recently described D. fumella from the Florida panhandle. Dicerandra linearifolia var. linearifolia showed some regional cohesion of populations, but there was no consistent geographic pattern to the clustering of populations. Dicerandra radfordiana showed consistent clustering with proximate populations of D. odoratissima. Given that D. radfordiana is found at the southeastern extreme of the range of D. odoratissima, these populations may represent the early stages of speciation by isolation. While there are morphological and bioclimatic niche distinctions between D. odoratissima and D. radfordiana, there is no molecular support for a distinct D. radfordiana. Overall, there is modest genetic diversity found at the population level for all Dicerandra annuals. Microsatellite data support previously proposed hypotheses of hybridization between D. linearifolia var. linearifolia and D. odoratissima, but do not support such hypotheses for D. fumella and D. linearifolia var. robustior.

Rangewide tidewater goby occupancy survey using environmental DNA Abstract Rangewide monitoring of fish species is critical for determining status and trends in distribution; however, implementations of large-scale distribution surveys have generally been constrained by time and cost. This study used environmental DNA (eDNA) to monitor the presence or absence of two endangered tidewater goby species, the northern tidewater goby (Eucyclogobius newberryi) and the southern tidewater goby (Eucyclogobius kristinae), across their combined geographic range that encompasses the entire California coast (1350 km). A total of 197 estuary sites were surveyed in coastal California from Del Norte to San Diego counties between May and September 2016. Among the 197 sites, a total of 430 water samples were collected (one to six per site), filtered, and tested for the presence/absence of northern and southern tidewater goby, using species-specific quantitative PCR assays. Northern tidewater goby were detected at 81 out of 175 sites and southern tidewater goby were detected at 4 out of 22 sites, resulting in a combined naïve occupancy of 0.43. In contrast, application of a multi-scale occupancy model that accounted for imperfect detection estimated site occupancy at 0.55 (95% CRI 0.46–0.64), indicating that tidewater goby were present but not detected at 23 additional sites. This study illustrates that eDNA methods represent a reliable and efficient tool for aquatic species monitoring, but highlight the importance of accounting for imperfect detection by use of occupancy models in eDNA surveys. The tidewater goby eDNA distributional snapshot represents a baseline for evaluation of future trends in site occupancy that will inform conservation and management of this endangered species.