Genetic Assignment Tests Research Articles

Population Genetic Assignment of Confiscated Gopher Tortoises

Abstract As gopher tortoises (Gopherus polyphemus) increasingly become threatened throughout their range in Florida, USA, the need for management and conservation will intensify. Here we evaluate the forensic applicability of genetic assignment tests based on microsatellite genotypic data to 1) accurately assign individuals in our genetic database to the sample location or population of origin and 2) determine the origin of 6 confiscated tortoises. Overall, we could correctly assign 90% of the individuals in the database to their population of origin, but we were unable to determine the source of the confiscated tortoises. However, these individuals are unlikely to have come from any of our sampled sites and all 6 may have come from the same population. This approach can be used by law enforcement personnel to identify the origin of confiscated tortoises as well as by developers and wildlife managers to determine the genetic appropriateness of potential recipient populations when it is necessary to relocate individuals.

Patterns of genetic diversity in Great Lakes bloaters (Coregonus hoyi) with a view to future reintroduction in Lake Ontario

The originally diverse ciscoe fish fauna of the Laurentian Great Lakes has suffered many extinctions and local extirpations. Bloaters (Coregonus hoyi) are presumed extirpated from Lake Ontario and the reintroduction of this deepwater fish is under consideration. Given the demographic fluctuations of this species in the other Great Lakes and its recent intralacustrine origin, we sought to identify a genetically diverse and similar source of C. hoyi via an analysis of genetic diversity and population structure using 10 microsatellite loci. Despite well-documented demographic declines, we found no genetic evidence of bottlenecks in 12 C. hoyi samples from the four potential donor lakes (Huron, Michigan, Superior and Nipigon). By contrast, evidence of bottlenecks in historical samples of C. artedi from Lake Ontario suggested that standard genetic methods frequently used to identify population bottlenecks can only detect very severe and long-lasting demographic declines in naturally large populations. Patterns of genetic differentiation and assignment tests indicated that C. hoyi from Lake Huron and Lake Michigan, which are not differentiated, are genetically most similar to Lake Ontario ciscoes. The small available sample of deepwater ciscoes recently caught in Lake Ontario did not allow determining if these represent a small undetected C. hoyi population or a recent invasion of the deep section by C. artedi. On the basis of genetic criteria, we conclude that C. hoyi from any location within Lake Huron or Lake Michigan would be judicious sources of breeders for reintroducing C. hoyi in Lake Ontario.

Evaluation of genetic diversity of eight grouper species Epinephelus spp. based on microsatellite variations

Genetic diversity among eight species of grouper, Epinephelus bleekeri, E. coioides, E. malabaricus, E. ongus, E. akaara, E. maculatus, E. merra and E. fuscoguttatus, was studied using six microsatellite loci, Em-01⋆, Em-03⋆, Em-07⋆, Em-08⋆, Em-10⋆, and CA-07⋆, with the aim of exploring the feasibility of using microsatellite data for species identification. The results showed high levels of genetic differentiation among species (Fst=0.4403 and Rst=0.4954). Species identification based on fixed allelic differences was possible between E. coioides, E. ongus, and E. fuscoguttatus at Em-01⋆ and between E. fuscoguttatus and E. ongus at Em-08⋆. Private alleles were found in all species, except for E. ongus. Pairwise Fst ranged 0.238–0.578 (P<0.008 Bonferroni correction), and Nei’s genetic distance ranged 0.433–2.710. Size homoplasy was observed at Em-03⋆157 allele, which was characterized by a T-C transition at the 119th nucleotide site of PCR products. The genetic assignment test unambiguously assigned each individual to the correct species. Thus, this test can be used for species identification of unknown individuals when the multilocus genotypes of the six microsatellite loci are available. The phylogenetic (neighbor-joining) tree, which was constructed based on the genetic distance matrix, separated the eight grouper species into two main groups.

Density‐dependent dispersal suggests a genetic measure of habitat suitability

Recent research shows that density dependence should result in predictable movements between habitats of different suitability, depending on whether population densities are increasing or decreasing. When population densities are increasing, habitats become filled in order of their suitability, resulting in a net flow from high suitability to low suitability. When populations decrease in density, the reverse can happen. These patterns suggest that genetic information can be used to infer habitat suitability since individual‐based genetic assignment tests permit high resolution assessments of migration. We used replicated landscapes to study fishers (Martes pennanti) during a population increase and predicted that there should be a net flow of individuals from areas of shallow to deep snow, since snow depth has previously been linked to fisher fitness. A total of 769 fishers were sampled from 35 different landscapes and profiled at 16 microsatellite loci. From assignment tests, we inferred five genetic populations. By assigning each of the 35 landscapes to one of these five populations, we were able to determine the proportion of immigrants to each. Consistent with our prediction, there was a positive relationship between the proportion of immigrants and snow depth. The best model of fisher habitat suitability was one with both snow depth and the proportion of coniferous forest in landscapes. Our findings suggest that where population trend is known, genetic information can be used to measure habitat suitability.

Do dams increase genetic diversity in brown trout (Salmo trutta)? Microgeographic differentiation in a fragmented river

Abstract – Local genetic differentiation may potentially arise in recently fragmented populations. Brown trout is a polytypic species exhibiting substantial genetic differentiation, which may evolve in few generations. Movement (semi‐)barriers in rivers may cause fragmentation, isolation and genetic differentiation in fish. In the Måna River (28 km) flowing from the alpine Lake Møsvatn to the boreal Lake Tinnsjø, construction of four hydropower dams during the period 1906–1957 have fragmented the previously (since last Ice Age) continuous wild resident brown trout population. Samples from the two lakes (N = 40) and six sites in the river (N = 30) isolated at different times were analysed at nine microsatellite loci. All populations showed substantial genetic variation (mean number of alleles per locus 5.3–8.9, observed heterozygosity 0.57–0.65 per population, overall Fst = 0.032). Pairwise multilocus Fst estimates indicated no significant differentiation between populations in the two lakes, and no or little differentiation in the lower river (Fst = 0.0035–0.0091). The microgeographic differentiation among wild resident trout at these sites was less than expected based on similar previous studies. However, results from the upper river, in particular the site immediately below the Lake Møsvatn outlet and dam, indicated isolation (Fst &gt; 0.035). Calculation of genetic distances and assignment tests corroborated these results, as did a significant correlation between years of isolation (since dam construction) and Fst. The population structuring is most likely a result of fragmentation by dams, which has increased overall genetic diversity. This increased local differentiation may be caused by natural selection, but more likely by genetic drift in small, recently fragmented populations. Increased local genetic diversity by genetic drift does not justify conservation measures aiming at preserving genetic diversity.

Genetic structure and assignment tests demonstrate illegal translocation of red deer (Cervus elaphus) into a continuous population

Molecular forensic methods are being increasingly used to help enforce wildlife conservation laws. Using multilocus genotyping, illegal translocation of an animal can be demonstrated by excluding all potential source populations as an individual's population of origin. Here, we illustrate how this approach can be applied to a large continuous population by defining the population genetic structure and excluding suspect animals from each identified cluster. We aimed to test the hypothesis that recreational hunters had illegally introduced a group of red deer into a hunting area in Luxembourg. Reference samples were collected over a large area in order to test the possibility that the suspect individuals might be recent immigrants. Due to isolation-by-distance relationships in the data set, inferring the number of genetic clusters using Bayesian methods was not straightforward. Biologically meaningful clusters were only obtained by simultaneously analysing spatial and genetic information using the program baps 4.1. We inferred the presence of three genetic clusters in the study region. Using partial Mantel tests, we detected barriers to gene flow other than distance, probably created by a combination of urban areas, motorways and a river valley used for viticulture. The four focal animals could be excluded with a high certainty from the three genetic subpopulations and it was therefore likely that they had been released illegally.

Microsatellite‐based diversity analysis and genetic relationships of three Indian sheep breeds

The genetic structure of three Indian sheep breeds from two different geographical locations (Nali, Chokla from north-western arid and semi-arid region; Garole from eastern saline marshy region) of India was investigated by means of 11 ovine-specific microsatellite markers as proposed in FAOs MoDAD programme. Microsatellite analysis revealed high allelic and gene diversity in all the three breeds. Nali sheep showed higher mean number of alleles and gene diversity (6.27 and 0.65) than Chokla (5.63 and 0.64) and Garole (5.63 and 0.59). High within population inbreeding estimates observed in the three breeds (FIS, Chokla = 0.286, Nali = 0.284, Garole = 0.227) reflected deficit of heterozygotes. The overall estimates for F-statistics were significantly (p < 0.05) different from zero. High values of FST (0.183) across all the loci revealed substantial degree of breed differentiation. Based on pair wise FST and Nm between different breeds, Nali and Chokla (FST = 6.62% and Nm = 4.80) were observed to be the closest followed by Garole and Nali (FST = 20.9% and Nm = 1.80), and Garole and Chokla (FST = 21.4% and Nm = 1.71). In addition, genetic distance estimates, phylogeny analysis and individual assignment test used to evaluate interbreed genetic proximity and population structure also revealed substantial genetic differentiation between Garole and the other two Rajasthani (Nali and Chokla) sheep. This divergent status of Garole sheep indicated genetic uniqueness of this breed suggesting higher priority for its conservation.

A genetic test for recruitment enhancement in Chesapeake Bay oysters, Crassostrea virginica, after population supplementation with a disease tolerant strain

Many of the methods currently employed to restore Chesapeake Bay populations of the eastern oyster, Crassostrea virginica, assume closed recruitment in certain sub-estuaries despite planktonic larval durations of 2–3 weeks. In addition, to combat parasitic disease, artificially selected disease tolerant oyster strains are being used for population supplementation. It has been impossible to fully evaluate these unconventional tactics because offspring from wild and selected broodstock are phenotypically indistinguishable. This study provides the first direct measurement of oyster recruitment enhancement by using genetic assignment tests to discriminate locally produced progeny of a selected oyster strain from progeny of wild parents. Artificially selected oysters (DEBY strain) were planted on a single reef in each of two Chesapeake Bay tributaries in 2002, but only in the Great Wicomico River (GWR) were they large enough to potentially reproduce the same year. Assignment tests based on eight microsatellite loci and mitochondrial DNA markers were applied to 1579 juvenile oysters collected throughout the GWR during the summer of 2002. Only one juvenile oyster was positively identified as an offspring of the 0.75 million DEBY oysters that were planted in the GWR, but 153 individuals (9.7%) had DEBY ×wild F1 multilocus genotypes. Because oyster recruitment was high across the region in 2002, the proportionately low enhancement measured in the GWR would not otherwise have been recognized. Possible causes for low enhancement success are discussed, each bearing on untested assumptions underlying the restoration methods, and all arguing for more intensive evaluation of each component of the restoration strategy.

Gene-flow between populations of cotton bollworm Helicoverpa armigera (Lepidoptera: Noctuidae) is highly variable between years

Both large and small scale migrations of Helicoverpa armigera Hübner in Australia were investigated using AMOVA analysis and genetic assignment tests. Five microsatellite loci were screened across 3142 individuals from 16 localities in eight major cotton and grain growing regions within Australia, over a 38-month period (November 1999 to January 2003). From November 1999 to March 2001 relatively low levels of migration were characterized between growing regions. Substantially higher than average gene-flow rates and limited differentiation between cropping regions characterized the period from April 2001 to March 2002. A reduced migration rate in the year from April 2002 to March 2003 resulted in significant genetic structuring between cropping regions. This differentiation was established within two or three generations. Genetic drift alone is unlikely to drive genetic differentiation over such a small number of generations, unless it is accompanied by extreme bottlenecks and/or selection. Helicoverpa armigera in Australia demonstrated isolation by distance, so immigration into cropping regions is more likely to come from nearby regions than from afar. This effect was most pronounced in years with limited migration. However, there is evidence of long distance dispersal events in periods of high migration (April 2001-March 2002). The implications of highly variable migration patterns for resistance management are considered.

Population structure and colonization history of the olive fly, Bactrocera oleae (Diptera, Tephritidae)

The olive fly, Bactrocera oleae, is the major pest of olives in most commercial olive-growing regions worldwide. The species is abundant in the Mediterranean basin and has been introduced recently into California and Mexico, creating problems for quarantine protection and international trade. Here, we use nuclear microsatellite markers and mitochondrial sequences to examine the history of olive fly range expansion and colonization. Sampled populations span the current distribution of the olive fly worldwide, including South and Central Africa, Pakistan, Mediterranean Europe and Middle East, California, and Mexico. The Pakistani populations appear to be genetically well differentiated from the remaining populations, though rooting the origins of the species is problematic. Genetic similarity and assignment tests cluster the remaining populations into two genetic groups--Africa and a group including the Mediterranean basin and the American region. That Africa, and not the Mediterranean, is the origin of flies infesting cultivated olive is supported by the significantly greater genetic diversity at microsatellite loci in Africa relative to the Mediterranean area. The results also indicate that the recent invasion of olive flies in the American region most likely originated from the Mediterranean area.

Effect of Vegetation Matrix on Animal Dispersal: Genetic Evidence from a Study of Endangered Skinks

Abstract: Maintaining connectivity in fragmented landscapes is a key principle of biological conservation. Although corridors are a widely accepted approach to connecting populations, their merits are still debated, and they may be impractical in many situations. A focus on management of the vegetation matrix between populations has been advocated as an alternative way to deal with habitat fragmentation and has theoretical support. We combined microsatellite DNA and demographic data to provide an empirical account of how two forms of agricultural land use affect the connectivity of insular populations of an endangered skink in southern New Zealand. The grand skink (Oligosoma grande) lives in small populations (approximately 20 individuals) on rock outcrops separated from one another by 50–150 m of inhospitable matrix vegetation (either native tussock grassland or exotic pasture). Skinks typically dispersed short distances, and the nature of the matrix both quantitatively and qualitatively affected dispersal dynamics. Skink populations in pasture were significantly more genetically structured and had less genetic variation than similar populations in tussock, implying less dispersal between populations in pasture than tussock. Furthermore, although female‐biased dispersal was a feature of populations in tussock, no sex bias was evident in pasture. In addition, Bayesian individual‐based genetic assignment tests that incorporated prior mark‐recapture information revealed that some populations produced many emigrants but received few immigrants, whereas other populations were relatively insular. Patterns of dispersal and response to matrix vegetation were complex, and the causes of these patterns deserve attention in future studies of habitat fragmentation. Managing the vegetation matrix may be a practical way to connect animal populations in some situations.

Genetic diversity and assignment tests among seven French cattle breeds based on microsatellite DNA analysis.

Genetic variability and relationships among six native French cattle breeds (Abondance, Tarentaise, Villard de Lans, Montbéliarde, Limousin, and Charolais) and one foreign breed (Holstein) were investigated using 23 microsatellite markers. These breeds were also compared with four Swiss breeds genotyped in a previously published study. Interestingly, the French alpine breeds have smaller population sizes but showed higher genetic variability than the larger Holstein breed. Neighbor-joining trees and PCA (principal components analysis) showed that alpine breeds tend to cluster together. Abondance and Tarentaise breeds were closely related, whereas the Holstein was highly differentiated from all breeds analyzed. Two different assignment tests for determining the breed of origin of individuals were compared: "direct" and "exclusion-simulation" approaches. The exclusion-simulation significance test correctly assigns fewer individuals than the direct approach but provides a confidence level (e.g., P < 0.01) for each individual being assigned. Accurate assignment with high statistical confidence is required for animal traceability. Unfortunately, the accuracy of assignment greatly decreases as the threshold level of confidence of assignment increases (e.g., from P < 0.05 to P < 0.001). Assignment accuracy also greatly declines as the level of population differentiation decreases below the level often found between related breeds (e.g., F(ST) < 0.1).

Female dispersal and relatedness structure in common wombats (Vombatus ursinus)

AbstractDispersal patterns were investigated in the common wombat Vombatus ursinus using microsatellite genetic data collected from three study sites. Two of these were within large, high‐density populations in Victoria, south‐eastern Australia, while the third was a small, isolated population in South Australia. Genetic assignment tests revealed that in the continuous populations, females were more likely to be immigrants into the sampling areas than were males, who were more likely to have been born locally. No such difference between the sexes was detected for the isolated population. At all three sites, pairwise relatedness estimates for males were negatively correlated with geographic distance, suggesting related males were clustered. This pattern was not observed among females, other than in the isolated population, where pairs of females showed a weak decline of relatedness with increasing geographic distance over a large spatial scale. Taken together, these data provide strong evidence that dispersal is predominantly by females. Furthermore, the observed spatial structure of relatedness suggests female breeding dispersal, a rare finding among mammalian species. Genetic variation within the isolated population was significantly reduced compared to the sites in Victoria. Our data are not sufficient to distinguish between two possible causes for this: (1) long‐term small size and isolation of populations at the edge of the species' range where resources may be more patchily distributed; (2) a genetic bottleneck effect of recent isolation resulting from agricultural development.

Dispersal patterns and population structure in a small marsupial, Antechinus agilis, from two forests analysed using microsatellite markers

Juvenile male agile antechinus (Antechinus agilis) are thought to disperse immediately after they leave the nest, whereas females are philopatric. However, no genetic data are available to support a hypothesis of male-biased dispersal, and the possible effects of habitat characteristics on dispersal are not known. We used five microsatellite loci to describe the dispersal patterns and population structure of A. agilis at two different forested sites (Mt Donna Buang and Mt Disappointment in central Victoria, south-eastern Australia). Within each site, DNA samples were collected from individuals from different trapping grids (four grids in Mt Donna Buang, collected over two years, and seven grids at Mt Disappointment, collected over one year). To assess levels of genetic structuring among the populations, Fst values (the proportion of genetic variance among populations) were calculated for each site and sex separately, and a test for isolation by distance was performed. Overall, Fst values were low, and did not increase with distance, and this may be the result of high levels of gene flow for both sexes at both sites, at least on the scale of less than 10 km. To investigate microsatellite patterns at a finer scale, we calculated pair-wise relatedness values separately for sites, years and sexes. By comparing these values for animals within and between grids, evidence was found for male-biased dispersal. A genetic assignment test provided further evidence for this: males were more likely to be assigned to grids other than their capture site. Most females had high assignment values for their capture site, although some of the females may have been immigrants. This result could be due to inaccuracies in the assignment test, or due to a higher frequency of dispersal of females than previously thought. The sex-biased dispersal was less pronounced at Mt Disappointment than at Mt Donna Buang. This may be related to the level of disturbance: logging patches, roads and fire-breaks (grass strips) disrupt the forest at Mt Disappointment more than at Mt Donna Buang.

Admixture analysis and stocking impact assessment in brown trout (Salmo trutta), estimated with incomplete baseline data

Studies of genetic interactions between wild and domesticated fish are often hampered by unavailability of samples from wild populations prior to population admixture. We assessed the utility of a new Bayesian method, which can estimate individual admixture coefficients even with data missing from the populations contributing to admixture. We applied the method to analyse the genetic contribution of domesticated brown trout (Salmo trutta) in samples of anadromous trout from two stocked populations with no genetic data available before stocking. Further, we estimated population level admixture proportions by the mean of individual admixture coefficients. This method proved more informative than a multidimensional scaling analysis of individual-based genetic distances and assignment tests. The results showed almost complete absence of stocked, domesticated trout in samples of trout from the rivers. Consequently, stocking had little effect on improving fisheries. In one population, the genetic contribution by domesticated trout was small, whereas in the other population, some genetic impact was suggested. Admixture in this sample of anadromous trout despite absence of stocked domesticated trout could be because of introgression by domesticated trout adopting a resident life history.