SambaR: An R package for fast, easy and reproducible population-genetic analyses of biallelic SNP data sets.

SNP data sets can be used to infer a wealth of information about natural populations, including information about their structure, genetic diversity, and the presence of loci under selection. However, SNP data analysis can be a time-consuming and challenging process, not in the least because at present many different software packages are needed to execute and depict the wide variety of mainstream population-genetic analyses. Here, we present SambaR, an integrative and user-friendly R package which automates and simplifies quality control and population-genetic analyses of biallelic SNP data sets. SambaR allows users to perform mainstream population-genetic analyses and to generate a wide variety of ready to publish graphs with a minimum number of commands (less than 10). These wrapper commands call functions of existing packages (including adegenet, ape, LEA, poppr, pcadapt and StAMPP) as well as new tools uniquely implemented in SambaR. We tested SambaR on online available SNP data sets and found that SambaR can process data sets of over 100,000 SNPs and hundreds of individuals within hours, given sufficient computing power. Newly developed tools implemented in SambaR facilitate optimization of filter settings, objective interpretation of ordination analyses, enhance comparability of diversity estimates from reduced representation library SNP data sets, and generate reduced SNP panels and structure-like plots with Bayesian population assignment probabilities. SambaR facilitates rapid population genetic analyses on biallelic SNP data sets by removing three major time sinks: file handling, software learning, and data plotting. In addition, SambaR provides a convenient platform for SNP data storage and management, as well as several new utilities, including guidance in setting appropriate data filters. The SambaR source script, manual and example data set are distributed through GitHub: https://github.com/mennodejong1986/SambaR.

Tracking invasions of a destructive defoliator, the gypsy moth (Erebidae: Lymantria dispar): Population structure, origin of intercepted specimens, and Asian introgression into North America.

Genetic data can help elucidate the dynamics of biological invasions, which are fueled by the constant expansion of international trade. The introduction of European gypsy moth (Lymantria dispar dispar) into North America is a classic example of human-aided invasion that has caused tremendous damage to North American temperate forests. Recently, the even more destructive Asian gypsy moth (mainly L. d. asiatica and L. d. japonica) has been intercepted in North America, mostly transported by cargo ships. To track invasion pathways, we developed a diagnostic panel of 60 DNA loci (55 nuclear and 5 mitochondrial) to characterize worldwide genetic differentiation within L. dispar and its sister species L. umbrosa. Hierarchical analyses supported strong differentiation and recovered five geographic groups that correspond to (1) North America, (2) Europe plus North Africa and Middle East, (3) the Urals, Central Asia, and Russian Siberia, (4) continental East Asia, and (5) the Japanese islands. Interestingly, L. umbrosa was grouped with L. d. japonica, and the introduced North American population exhibits remarkable distinctiveness from contemporary European counterparts. Each geographic group, except for North America, shows additional lower-level structures when analyzed individually, which provided the basis for inference of the origin of invasive specimens. Two assignment approaches consistently identified a coastal area of continental East Asia as the major source for Asian invasion during 2014-2015, with Japan being another source. By analyzing simulation and laboratory crosses, we further provided evidence for the occurrence of natural Asian-North American hybrids in the Pacific Northwest, raising concerns for introgression of Asian alleles that may accelerate range expansion of gypsy moth in North America. Our study demonstrates how genetic data contribute to bio-surveillance of invasive species with results that can inform regulatory management and reduce the frequency of trade-associated invasions.

Genetic Characterization of the Local Pirenaica Cattle for Parentage and Traceability Purposes.

Pirenaica is the most important autochthonous cattle breed within the Protected Geographic Indication (PGI) beef quality label in the Basque region, in northern Spain. The short tandem repeats (STRs) are powerful markers to elucidate forensic cases and traceability across the agri-food sector. The main objective of the present work was to study the phylogenetic relationships of Pirenaica cattle and other breeds typically raised in the region and provide the minimum number of STR markers for parentage and traceability purposes. The 30-STR panel recommended by the International Society of Animal Genetics-Food and Agriculture Organization of the United Nations (ISAG-FAO) was compared against other commercial STR panels. The 30-STR panel showed a combined matching probability of 1.89 × 10-25 and a power of exclusion for duos of 0.99998. However, commercial STR panels showed a limited efficiency for a reliable parentage analysis in Pirenaica, and at least a 21-STR panel is needed to reach a power of exclusion of 0.9999. Machine-learning analysis also demonstrated a 95% accuracy in assignments selecting the markers with the highest FST in Pirenaica individuals. Overall, the present study shows the genetic characterization of Pirenaica and its phylogeny compared with other breeds typically raised in the Basque region. Finally, a 21-STR panel with the highest FST markers is proposed for a confident parentage analysis and high traceability.

Development and characterization of 19 polymorphic microsatellite loci from the Red-cowled Cardinal (Paroaria dominicana, Passeriformes, Aves) using next-generation sequencing.

The Red-cowled Cardinal (Paroaria dominicana) is an endemic passerine of the Caatinga biome in Brazil, and is one of the most traded passerines in the country. Illegal trade can have serious impacts on wild populations, such as reduced population sizes, the introduction of the species to areas outside their historical range or mixing individuals from different populations. Microsatellites constitute an important tool for population genetics and forensics studies, and hold great potential to help authorities manage illegal trafficking and inspect commercial breeders. We developed new microsatellite loci using massive parallel sequencing and characterized them in 23 seized Red-cowled Cardinals with unknown geographic origin. The DNA sequencing generated 2,068,684 paired-reads of which we identified 10,322 tri- to hexanucleotide loci. We selected 30 loci for amplification and polymorphism tests, of which 21 successfully amplified and 19 were polymorphic. The number of alleles ranged from 7 to 18 and the mean expected heterozygosity was 0.863. Six loci deviated from Hardy-Weinberg equilibrium probably due to null alleles and/or the Wahlund effect. Polymorphic loci in Hardy-Weinberg equilibrium showed low identity probability and high paternity exclusion probability. Our results indicate that this new set of microsatellite loci constitutes an important tool for both population genetic and forensic studies, with ultimate potential for assisting authorities in managing animal victims of illegal trafficking and the inspection of commercial breeders of the Red-cowled Cardinal.

Dog breed affiliation with a forensically validated canine STR set.

We tested a panel of 13 highly polymorphic canine short tandem repeat (STR) markers for dog breed assignment using 392 dog samples from the 23 most popular breeds in Austria, Germany, and Switzerland. This STR panel had originally been selected for canine identification. The dog breeds sampled in this study featured a population frequency ≥1% and accounted for nearly 57% of the entire pedigree dog population in these three countries. Breed selection was based on a survey comprising records for nearly 1.9 million purebred dogs belonging to more than 500 different breeds. To derive breed membership from STR genotypes, a range of algorithms were used. These methods included discriminant analysis of principal components (DAPC), STRUCTURE, GeneClass2, and the adegenet package for R. STRUCTURE analyses suggested 21 distinct genetic clusters. Differentiation between most breeds was clearly discernable. Fourteen of 23 breeds (61%) exhibited maximum mean cluster membership proportions of more than 0.70 with a highest value of 0.90 found for Cavalier King Charles Spaniels. Dogs of only 6 breeds (26%) failed to consistently show only one major cluster. The DAPC method yielded the best assignment results in all 23 declared breeds with 97.5% assignment success. The frequency-based assignment test also provided a high success rate of 87%. These results indicate the potential viability of dog breed prediction using a well-established and sensitive set of 13 canine STR markers intended for forensic routine use.

Population differentiation or species formation across the Indian and the Pacific Oceans? An example from the brooding marine hydrozoan Macrorhynchia phoenicea.

Assessing population connectivity is necessary to construct effective marine protected areas. This connectivity depends, among other parameters, inherently on species dispersal capacities. Isolation by distance (IBD) is one of the main modes of differentiation in marine species, above all in species presenting low dispersal abilities. This study reports the genetic structuring in the tropical hydrozoan Macrorhynchia phoenicea α (sensu Postaire et al., 2016a), a brooding species, from 30 sampling sites in the Western Indian Ocean and the Tropical Southwestern Pacific, using 15 microsatellite loci. At the local scale, genet dispersal relied on asexual propagation at short distance, which was not found at larger scales. Considering one representative per clone, significant positive FIS values (from -0.327*** to 0.411***) were found within almost all sites. Gene flow was extremely low at all spatial scales, among sites within islands (<10 km distance) and among islands (100 to >11,000 km distance), with significant pairwise FST values (from 0.035*** to 0.645***). A general pattern of IBD was found at the Indo-Pacific scale, but also within ecoregions in the Western Indian Ocean province. Clustering and network analyses identified each island as a potential independent population, while analysis of molecular variance indicated that population genetic differentiation was significant at small (within island) and intermediate (among islands within province) spatial scales. As shown by this species, a brooding life cycle might be corollary of the high population differentiation found in some coastal marine species, thwarting regular dispersal at distances more than a few kilometers and probably leading to high cryptic diversity, each island housing independent evolutionary lineages.

Genetic rescue, the greater prairie chicken and the problem of conservation reliance in the Anthropocene.

A central question in conservation is how best to manage biodiversity, despite human domination of global processes (= Anthropocene). Common responses (i.e. translocations, genetic rescue) forestall potential extirpations, yet have an uncertain duration. A textbook example is the greater prairie chicken (GRPC: Tympanuchus cupido pinnatus), where translocations (1992-1998) seemingly rescued genetically depauperate Illinois populations. We re-evaluated this situation after two decades by genotyping 21 microsatellite loci from 1831 shed feathers across six leks in two counties over 4 years (2010-2013). Low migration rates (less than 1%) established each county as demographically independent, but with declining-population estimates (4 year average N = 79). Leks were genetically similar and significantly bottlenecked, with low effective population sizes (average Ne = 13.1; 4 year Ne/N = 0.166). Genetic structure was defined by 12 significantly different family groups, with relatedness r = 0.31 > half-sib r = 0.25. Average heterozygosity, indicating short-term survival, did not differ among contemporary, pre- and post-translocated populations, whereas allelic diversity did. Our results, the natural history of GRPC (i.e. few leks, male dominance hierarchies) and its controlled immigration suggest demographic expansion rather than genetic rescue. Legal protection under the endangered species act (ESA) may enhance recovery, but could exacerbate political-economic concerns on how best to manage 'conservation-reliant' species, for which GRPC is now an exemplar.

RAD genotyping reveals fine-scale genetic structuring and provides powerful population assignment in a widely distributed marine species, the American lobster (Homarus americanus).

Deciphering genetic structure and inferring connectivity in marine species have been challenging due to weak genetic differentiation and limited resolution offered by traditional genotypic methods. The main goal of this study was to assess how a population genomics framework could help delineate the genetic structure of the American lobster (Homarus americanus) throughout much of the species' range and increase the assignment success of individuals to their location of origin. We genotyped 10 156 filtered SNPs using RAD sequencing to delineate genetic structure and perform population assignment for 586 American lobsters collected in 17 locations distributed across a large portion of the species' natural distribution range. Our results revealed the existence of a hierarchical genetic structure, first separating lobsters from the northern and southern part of the range (FCT  = 0.0011; P-value = 0.0002) and then revealing a total of 11 genetically distinguishable populations (mean FST  = 0.00185; CI: 0.0007-0.0021, P-value < 0.0002), providing strong evidence for weak, albeit fine-scale population structuring within each region. A resampling procedure showed that assignment success was highest with a subset of 3000 SNPs having the highest FST . Applying Anderson's (Molecular Ecology Resources, 2010, 10, 701) method to avoid 'high-grading bias', 94.2% and 80.8% of individuals were correctly assigned to their region and location of origin, respectively. Lastly, we showed that assignment success was positively associated with sample size. These results demonstrate that using a large number of SNPs improves fine-scale population structure delineation and population assignment success in a context of weak genetic structure. We discuss the implications of these findings for the conservation and management of highly connected marine species, particularly regarding the geographic scale of demographic independence.

Empirical Selection of Informative Microsatellite Markers within Co-ancestry Pig Populations Is Required for Improving the Individual Assignment Efficiency.

The Lanyu is a miniature pig breed indigenous to Lanyu Island, Taiwan. It is distantly related to Asian and European pig breeds. It has been inbred to generate two breeds and crossed with Landrace and Duroc to produce two hybrids for laboratory use. Selecting sets of informative genetic markers to track the genetic qualities of laboratory animals and stud stock is an important function of genetic databases. For more than two decades, Lanyu derived breeds of common ancestry and crossbreeds have been used to examine the effectiveness of genetic marker selection and optimal approaches for individual assignment. In this paper, these pigs and the following breeds: Berkshire, Duroc, Landrace and Yorkshire, Meishan and Taoyuan, TLRI Black Pig No. 1, and Kaohsiung Animal Propagation Station Black pig are studied to build a genetic reference database. Nineteen microsatellite markers (loci) provide information on genetic variation and differentiation among studied breeds. High differentiation index (FST) and Cavalli-Sforza chord distances give genetic differentiation among breeds, including Lanyu's inbred populations. Inbreeding values (FIS) show that Lanyu and its derived inbred breeds have significant loss of heterozygosity. Individual assignment testing of 352 animals was done with different numbers of microsatellite markers in this study. The testing assigned 99% of the animals successfully into their correct reference populations based on 9 to 14 markers ranking D-scores, allelic number, expected heterozygosity (HE) or FST, respectively. All miss-assigned individuals came from close lineage Lanyu breeds. To improve individual assignment among close lineage breeds, microsatellite markers selected from Lanyu populations with high polymorphic, heterozygosity, FST and D-scores were used. Only 6 to 8 markers ranking HE, FST or allelic number were required to obtain 99% assignment accuracy. This result suggests empirical examination of assignment-error rates is required if discernible levels of co-ancestry exist. In the reference group, optimum assignment accuracy was achievable achieved through a combination of different markers by ranking the heterozygosity, FST and allelic number of close lineage populations.

Molecular identification of the hybrid between the catfish species Pseudoplatystoma corruscans and Pseudoplatystoma reticulatum using a set of eight microsatellite markers.

This study distinguished hybrids of surubim or pintado Pseudoplatystoma corruscans and cachara Pseudoplatystoma reticulatum from pure strains using a set of eight microsatellite markers and population assignment methods. Applications of this molecular tool range from certification of hybrid-free breeders in restocking conservation programmes to the identification of fish products lacking traditional morphological characteristics.

Can interbreeding of wild and artificially propagated animals be prevented by using broodstock selected for a divergent life history?

TWO STRATEGIES HAVE BEEN PROPOSED TO AVOID NEGATIVE GENETIC EFFECTS OF ARTIFICIALLY PROPAGATED INDIVIDUALS ON WILD POPULATIONS: (i) integration of wild and captive populations to minimize domestication selection and (ii) segregation of released individuals from the wild population to minimize interbreeding. We tested the efficacy of the strategy of segregation by divergent life history in a steelhead trout, Oncorhynchus mykiss, system, where hatchery fish were selected to spawn months earlier than the indigenous wild population. The proportion of wild ancestry smolts and adults declined by 10-20% over the three generations since the hatchery program began. Up to 80% of the naturally produced steelhead in any given year were hatchery/wild hybrids. Regression model selection analysis showed that the proportion of hatchery ancestry smolts was lower in years when stream discharge was high, suggesting a negative effect of flow on reproductive success of early-spawning hatchery fish. Furthermore, proportions of hybrid smolts and adults were higher in years when the number of naturally spawning hatchery-produced adults was higher. Divergent life history failed to prevent interbreeding when physical isolation was ineffective, an inadequacy that is likely to prevail in many other situations.