Penalized classification for optimal statistical selection of markers from high-throughput genotyping: application in sheep breeds

The identification of individuals’ breed of origin has several practical applications in livestock and is useful in different biological contexts such as conservation genetics, breeding and authentication of animal products. In this paper, penalized multinomial regression was applied to identify the minimum number of single nucleotide polymorphisms (SNPs) from high-throughput genotyping data for individual assignment to dairy sheep breeds reared in Sicily. The combined use of penalized multinomial regression and stability selection reduced the number of SNPs required to 48. A final validation step on an independent population was carried out obtaining 100% correctly classified individuals. The results using independent analysis, such as admixture, F_st, principal component analysis and random forest, confirmed the ability of these methods in selecting distinctive markers. The identified SNPs may constitute a starting point for the development of a SNP based identification test as a tool for breed assignment and traceability of animal products.     

The selection of the ‘wild’: A combined molecular approach for the identification of pure indigenous fish from hybridised populations

The identification of pure indigenous fish from hybridised populations represents a key issue in fisheries management and conservation biology. In the present study an approach for selection of purebred marble trout (Salmo trutta marmoratus C.) individuals out of admixed populations was set up and assessed. In a first step, baseline data sets of pure marble trout and pure brown trout specimens based on twelve microsatellite loci were used to simulate five consecutive generations of admixture. The baseline and the resulting simulation data sets were then combined with data of a ‘real’ hybridised marble trout population to perform a single individual assignment test as implemented in STRUCTURE. By this procedure the assignment approach was calibrated and it was possible to compare admixture coefficients obtained for individuals from different populations. The ranking of individual admixture coefficients on a plot and comparison with simulated data revealed that the test population was composed of pure marble trout individuals, first generation hybrids between marble trout and brown trout, and hybrid backcross specimens between both groups. However, by defining a critical q-value of 0.1 and additionally integrating individual sequence data of the mtDNA control region, it was possible to indicate individuals, which could be selected for the establishment of a pure marble trout strain.     

Methods Used to Assess Non-target Effects of Invertebrate Biological Control Agents of Arthropod Pests

The number of concerns regarding potential non-target effects of invertebrate biological control agents of arthropods has risen over the last decade and an increasing number of studies have since dealt with this topic. Despite some recent international initiatives aimed at providing guidance for risk assessment of biological control agents, detailed methods on how tests should be designed and conducted to assess for potential non-target effects still need to be provided. It is believed that this review comes at an ideal time, giving an overview of methods currently applied in the study of non-target effects in biological control of arthropod pests. It provides the first step towards the ultimate goal of devising guidelines for the appropriate methods that should be universally applied for the assessment and minimisation of potential non-target effects. The main topics that are reviewed here include host specificity (including field surveys, selection of non-target test species and testing protocols), post-release studies, competition, overwintering and dispersal. Finally, a number of conclusions that have emerged from this comprehensive compilation of studies are drawn, addressing potential non-target effects in arthropod biological control.     

A comparison of four methods for detecting weak genetic structure from marker data

Genetic structure is ubiquitous in wild populations and is the result of the processes of natural selection, genetic drift, mutation, and gene flow. Genetic drift and divergent selection promotes the generation of genetic structure, while gene flow homogenizes the subpopulations. The ability to detect genetic structure from marker data diminishes rapidly with a decreasing level of differentiation among subpopulations. Weak genetic structure may be unimportant over evolutionary time scales but could have important implications in ecology and conservation biology. In this paper we examine methods for detecting and quantifying weak genetic structures using simulated data. We simulated populations consisting of two putative subpopulations evolving for up to 50 generations with varying degrees of gene flow (migration), and varying amounts of information (allelic diversity). There are a number of techniques available to detect and quantify genetic structure but here we concentrate on four methods: F(ST), population assignment, relatedness, and sibship assignment. Under the simple mating system simulated here, the four methods produce qualitatively similar results. However, the assignment method performed relatively poorly when genetic structure was weak and we therefore caution against using this method when the analytical aim is to detect fine-scale patterns. Further work should examine situations with different mating systems, for example where a few individuals dominate reproductive output of the population. This study will help workers to design their experiments (e.g., sample sizes of markers and individuals), and to decide which methods are likely to be most appropriate for their particular data.     

Source population of dispersing rock-wallabies (Petrogale lateralis) identified by assignment tests on multilocus genotypic data

The ability to confidently identify or exclude a population as the source of an individual has numerous powerful applications in molecular ecology. Several alternative assignment methods have recently been developed and are yet to be fully evaluated with empirical data. In this study we tested the efficacy of different assignment methods by using a translocated rock-wallaby (Petrogale lateralis) population, of known provenance. Specimens from the translocated population (n = 43), its known source population (n = 30) and four other nearby populations (n = 19-32) were genotyped for 11 polymorphic microsatellite loci. The results identified Bayesian clustering, frequency and Bayesian methods as the most consistent and accurate, correctly assigning 93-100% of individuals up to a significance threshold of P = 0.01. Performance was variable among the distance-based methods, with the Cavalli-Sforza and Edwards chord distance performing best, whereas Goldstein et al.'s (deltamu)2 consistently performed poorly. Using Bayesian clustering, frequency and Bayesian methods we then attempted to determine the source of rock-wallabies which have recently recolonized an outcrop (Gardners) 8 km from the nearest rock-wallaby population. Results indicate that the population at Gardners originated via a recent dispersal event from the eastern end of Mt. Caroline. This is only the second published record of dispersal by rock-wallabies between habitat patches and is the longest movement recorded to date. Molecular techniques and methods of analysis are now available to allow detailed studies of dispersal in rock-wallabies and should also be possible for many other taxa.     

Variation of cats under domestication: genetic assignment of domestic cats to breeds and worldwide random‐bred populations

Both cat breeders and the lay public have interests in the origins of their pets, not only in the genetic identity of the purebred individuals, but also in the historical origins of common household cats. The cat fancy is a relatively new institution with over 85% of its 40–50 breeds arising only in the past 75 years, primarily through selection on single‐gene aesthetic traits. The short, yet intense cat breed history poses a significant challenge to the development of a genetic marker–based breed identification strategy. Using different breed assignment strategies and methods, 477 cats representing 29 fancy breeds were analysed with 38 short tandem repeats, 148 intergenic and five phenotypic single nucleotide polymorphisms. Results suggest the frequentist method of Paetkau (single nucleotide polymorphisms = 0.78, short tandem repeats = 0.88) surpasses the Bayesian method of Rannala and Mountain (single nucleotide polymorphisms = 0.56, short tandem repeats = 0.83) for accurate assignment of individuals to the correct breed. Additionally, a post‐assignment verification step with the five phenotypic single nucleotide polymorphisms accurately identified between 0.31 and 0.58 of the misassigned individuals raising the sensitivity of assignment with the frequentist method to 0.89 and 0.92 for single nucleotide polymorphisms and short tandem repeats respectively. This study provides a novel multistep assignment strategy and suggests that, despite their short breed history and breed family groupings, a majority of cats can be assigned to their proper breed or population of origin, that is, race.     

Genetic diversity and evolutionary patterns of Taraxacum kok‐saghyz Rodin

Taraxacum kok‐saghyz Rodin (TKS) is an important potential alternative source of natural inulin and rubber production, which has great significance for the production of industrial products. In this study, we sequenced 58 wild TKS individuals collected from four different geography regions worldwide to elucidate the population structure, genetic diversity, and the patterns of evolution. Also, the first flowering time, crown diameter, morphological characteristics of leaf, and scape of all TKS individuals were measured and evaluated statistically. Phylogenetic analysis based on SNPs and cluster analysis based on agronomic traits showed that all 58 TKS individuals could be roughly divided into three distinct groups: (a) Zhaosu County in Xinjiang (population AB, including a few individuals from population C and D); (b) Tekes County in Xinjiang (population C); and (c) Tuzkol lake in Kazakhstan (population D). Population D exhibited a closer genetic relationship with population C compared with population AB. Genetic diversity analysis further revealed that population expansion from C and D to AB occurred, as well as gene flow between them. Additionally, some natural selection regions were identified in AB population. Function annotation of candidate genes identified in these regions revealed that they mainly participated in biological regulation processes, such as transporter activity, structural molecule activity, and molecular function regulator. We speculated that the genes identified in selective sweep regions may contribute to TKS adaptation to the Yili River Valley of Xinjiang. In general, this study provides new insights in clarifying population structure and genetic diversity analysis of TKS using SNP molecular markers and agronomic traits.     

"Reverse ecology" and the power of population genomics

Rapid and inexpensive sequencing technologies are making it possible to collect whole genome sequence data on multiple individuals from a population. This type of data can be used to quickly identify genes that control important ecological and evolutionary phenotypes by finding the targets of adaptive natural selection, and we therefore refer to such approaches as "reverse ecology." To quantify the power gained in detecting positive selection using population genomic data, we compare three statistical methods for identifying targets of selection: the McDonald-Kreitman test, the mkprf method, and a likelihood implementation for detecting d(N)/d(S) > 1. Because the first two methods use polymorphism data we expect them to have more power to detect selection. However, when applied to population genomic datasets from human, fly, and yeast, the tests using polymorphism data were actually weaker in two of the three datasets. We explore reasons why the simpler comparative method has identified more genes under selection, and suggest that the different methods may really be detecting different signals from the same sequence data. Finally, we find several statistical anomalies associated with the mkprf method, including an almost linear dependence between the number of positively selected genes identified and the prior distributions used. We conclude that interpreting the results produced by this method should be done with some caution.     

Inference of population structure under a Dirichlet process model

Inferring population structure from genetic data sampled from some number of individuals is a formidable statistical problem. One widely used approach considers the number of populations to be fixed and calculates the posterior probability of assigning individuals to each population. More recently, the assignment of individuals to populations and the number of populations have both been considered random variables that follow a Dirichlet process prior. We examined the statistical behavior of assignment of individuals to populations under a Dirichlet process prior. First, we examined a best-case scenario, in which all of the assumptions of the Dirichlet process prior were satisfied, by generating data under a Dirichlet process prior. Second, we examined the performance of the method when the genetic data were generated under a population genetics model with symmetric migration between populations. We examined the accuracy of population assignment using a distance on partitions. The method can be quite accurate with a moderate number of loci. As expected, inferences on the number of populations are more accurate when theta = 4N(e)u is large and when the migration rate (4N(e)m) is low. We also examined the sensitivity of inferences of population structure to choice of the parameter of the Dirichlet process model. Although inferences could be sensitive to the choice of the prior on the number of populations, this sensitivity occurred when the number of loci sampled was small; inferences are more robust to the prior on the number of populations when the number of sampled loci is large. Finally, we discuss several methods for summarizing the results of a Bayesian Markov chain Monte Carlo (MCMC) analysis of population structure. We develop the notion of the mean population partition, which is the partition of individuals to populations that minimizes the squared partition distance to the partitions sampled by the MCMC algorithm.     

Individual organisms as units of analysis: Bayesian-clustering alternatives in population genetics

Population genetic analyses traditionally focus on the frequencies of alleles or genotypes in 'populations' that are delimited a priori. However, there are potential drawbacks of amalgamating genetic data into such composite attributes of assemblages of specimens: genetic information on individual specimens is lost or submerged as an inherent part of the analysis. A potential also exists for circular reasoning when a population's initial identification and subsequent genetic characterization are coupled. In principle, these problems are circumvented by some newer methods of population identification and individual assignment based on statistical clustering of specimen genotypes. Here we evaluate a recent method in this genre--Bayesian clustering--using four genotypic data sets involving different types of molecular markers in non-model organisms from nature. As expected, measures of population genetic structure (F(ST) and phiST) tended to be significantly greater in Bayesian a posteriori data treatments than in analyses where populations were delimited a priori. In the four biological contexts examined, which involved both geographic population structures and hybrid zones, Bayesian clustering was able to recover differentiated populations, and Bayesian assignments were able to identify likely population sources of specific individuals.     

The ideal free distribution: a review and synthesis of the game-theoretic perspective

The Ideal Free Distribution (IFD), introduced by Fretwell and Lucas in [Fretwell, D.S., Lucas, H.L., 1970. On territorial behavior and other factors influencing habitat distribution in birds. Acta Biotheoretica 19, 16-32] to predict how a single species will distribute itself among several patches, is often cited as an example of an evolutionarily stable strategy (ESS). By defining the strategies and payoffs for habitat selection, this article puts the IFD concept in a more general game-theoretic setting of the “habitat selection game”. Within this game-theoretic framework, the article focuses on recent progress in the following directions: (1) studying evolutionarily stable dispersal rates and corresponding dispersal dynamics; (2) extending the concept when population numbers are not fixed but undergo population dynamics; (3) generalizing the IFD to multiple species.For a single species, the article briefly reviews existing results. It also develops a new perspective for Parker’s matching principle, showing that this can be viewed as the IFD of the habitat selection game that models consumer behavior in several resource patches and analyzing complications involved when the model includes resource dynamics as well. For two species, the article first demonstrates that the connection between IFD and ESS is now more delicate by pointing out pitfalls that arise when applying several existing game-theoretic approaches to these habitat selection games. However, by providing a new detailed analysis of dispersal dynamics for predator-prey or competitive interactions in two habitats, it also pinpoints one approach that shows much promise in this general setting, the so-called “two-species ESS”. The consequences of this concept are shown to be related to recent studies of population dynamics combined with individual dispersal and are explored for more species or more patches.     

Genetic diversity, population structure, effective population size and demographic history of the Finnish wolf population

The Finnish wolf population (Canis lupus) was sampled during three different periods (1996-1998, 1999-2001 and 2002-2004), and 118 individuals were genotyped with 10 microsatellite markers. Large genetic variation was found in the population despite a recent demographic bottleneck. No spatial population subdivision was found even though a significant negative relationship between genetic relatedness and geographic distance suggested isolation by distance. Very few individuals did not belong to the local wolf population as determined by assignment analyses, suggesting a low level of immigration in the population. We used the temporal approach and several statistical methods to estimate the variance effective size of the population. All methods gave similar estimates of effective population size, approximately 40 wolves. These estimates were slightly larger than the estimated census size of breeding individuals. A Bayesian model based on Markov chain Monte Carlo simulations indicated strong evidence for a long-term population decline. These results suggest that the contemporary wolf population size is roughly 8% of its historical size, and that the population decline dates back to late 19th century or early 20th century. Despite an increase of over 50% in the census size of the population during the whole study period, there was only weak evidence that the effective population size during the last period was higher than during the first. This may be caused by increased inbreeding, diminished dispersal within the population, and decreased immigration to the population during the last study period.     

Management of genetic diversity in small farm animal populations

Many local breeds of farm animals have small populations and, consequently, are highly endangered. The correct genetic management of such populations is crucial for their survival. Managing an animal population involves two steps: first, the individuals who will be permitted to leave descendants are to be chosen and the number offspring they will be permitted to produce has to be determined; second, the mating scheme has to be identified. Strategies dealing with the first step are directed towards the maximisation of effective population size and, therefore, act jointly on the reduction in the loss of genetic variation and in the increase of inbreeding. In this paper, the most relevant methods are summarised, including the so-called 'Optimum Contribution' methodology (contributions are proportional to the coancestry of each individual with the rest), which has been shown to be the best. Typically, this method is applied to pedigree information, but molecular marker data can be used to complete or replace the genealogy. When the population is subjected to explicit selection on any trait, the above methodology can be used by balancing the response to selection and the increase in coancestry/inbreeding. Different mating strategies also exist. Some of the mating schemes try to reduce the level of inbreeding in the short term by preventing mating between relatives. Others involve regular (circular) schemes that imply higher levels of inbreeding within populations in the short term, but demonstrate better performance in the long term. In addition, other tools such as cryopreservation and reproductive techniques aid in the management of small populations. In the future, genomic marker panels may replace the pedigree information in measuring the coancestry. The paper also includes the results of several experiments and field studies on the effectiveness and on the consequences of the use of the different strategies.     

Characterizing dispersal patterns in a threatened seabird with limited genetic structure

Genetic assignment methods provide an appealing approach for characterizing dispersal patterns on ecological time scales, but require sufficient genetic differentiation to accurately identify migrants and a large enough sample size of migrants to, for example, compare dispersal between sexes or age classes. We demonstrate that assignment methods can be rigorously used to characterize dispersal patterns in a marbled murrelet (Brachyramphus marmoratus) population from central California that numbers approximately 600 individuals and is only moderately differentiated (F_ST～ 0.03) from larger populations to the north. We used coalescent simulations to select a significance level that resulted in a low and approximately equal expected number of type I and II errors and then used this significance level to identify a population of origin for 589 individuals genotyped at 13 microsatellite loci. The proportion of migrants in central California was greatest during winter when 83% of individuals were classified as migrants compared to lower proportions during the breeding (6%) and post‐breeding (8%) seasons. Dispersal was also biased toward young and female individuals, as is typical in birds. Migrants were rarely members of parent‐offspring pairs, suggesting that they contributed few young to the central California population. A greater number of migrants than expected under equilibrium conditions, a lack of individuals with mixed ancestry, and a small number of potential source populations (two), likely allowed us to use assignment methods to rigorously characterize dispersal patterns for a population that was larger and less differentiated than typically thought required for the identification of migrants.     

Comparative study of the population structure and population assignment of sockeye salmon Oncorhynchus nerka from West Kamchatka based on RAPD-PCR and microsatellite polymorphism

Using two types of molecular markers, a comparative analysis of the population structure of sockeye salmon from West Kamchatka as well as population assignment of each individual fish were carried out. The values of a RAPD-PCR-based population assignment test (94-100%) were somewhat higher than those based on microsatellite data (74-84%). However, these results seem quite satisfactory because of high polymorphism of the microsatellite loci examined. The UPGMA dendrograms of genetic similarity of three largest spawning populations, constructed using each of the methods, were highly reliable, which was demonstrated by high bootstrap indices (100% in the case of RAPD-PCR; 84 and 100%, in the case of microsatellite analysis), though the resultant trees differed from one another. The different topology of the trees, in our view, is explained by the fact that the employed methods explored different parts of the genome; hence, the obtained results, albeit valid, may not correlate. Thus, to enhance reliability of the results, several methods of analysis should be used concurrently.     

Comparative study of the population structure and population assignment of sockeye salmon Oncorhynchus nerka from West Kamchatka based on RAPD-PCR and microsatellite polymorphism

Using two types of molecular markers, a comparative analysis of the population structure of sockeye salmon from West Kamchatka as well as population assignment of each individual fish were carried out. The values of a RAPD-PCR-based population assignment test (94–100%) were somewhat higher than those based on microsatellite data (74–84%). However, these results seem quite satisfactory because of high polymorphism of the microsatellite loci examined. The UPGMA dendrograms of genetic similarity of three largest spawning populations, constructed using each of the methods, were highly reliable, which was demonstrated by high bootstrap indices (100% in the case of RAPD-PCR; 84 and 100%, in the case of microsatellite analysis), though the resultant trees differed from one another. The different topology of the trees, in our view, is explained by the fact that the employed methods explored different parts of the genome; hence, the obtained results, albeit valid, may not correlate. Thus, to enhance reliability of the results, several methods of analysis should be used concurrently.     

Analysis of Human Mitochondrial DNA Polymorphisms in the Japanese Population

The highly polymorphic nature and high amplification efficiency of mitochondrial DNA (mtDNA) is valuable for the analysis of biological evidence in forensic casework, such as the identification of individuals and assignment of race/ethnicity. To be useful, a mtDNA polymorphism database for the Japanese population requires an understanding of the range of haplotype variation and phylogenies of mtDNA sequences. To extend current knowledge on the haplotypes in the Japanese population, this study defines new lineages and provides more detail about some of those previously described. We compared the hypervariable regions (HVRs) of 270 healthy, unrelated Japanese individuals and demonstrated 192 haplotypes. Combining HVR1 and HVR2, the genetic diversity was 0.9935, thus providing a high level of identification capability. Haplogroup status was defined for 160 individuals using HVR1, HVR2, and particular coding region polymorphisms; these individuals belonged to 94 haplotypes, four of which were new lineages. The complete mtDNA sequence was also determined from seven individuals.     

Interplay between ecological, behavioural and historical factors in shaping the genetic structure of sympatric walleye populations (Sander vitreus)

Disentangling ecological, behavioural and evolutionary factors responsible for the presence of stable population structure within wild populations has long been challenging to population geneticists. This study primarily aimed at decoding population structure of wild walleye (Sander vitreus) populations of Mistassini Lake (Québec, Canada) in order to define source populations to be used for the study of spatial partitioning using individual-based multilocus assignment methods, and decipher the dynamics of individual dispersal and resulting patterns of spatial resource partitioning and connectivity among populations. A second objective was to elucidate the relationships between biological characteristics (sex, size, age and population of origin) and an individual's probability to migrate and/or disperse. To do so, a total of 780 spawning individuals caught on five distinct spawning sites, and 1165 postspawning individuals, captured over two sampling seasons (2002-2003) were analysed by means of eight microsatellite loci. Four temporally stable walleye populations associated with distinct reproductive grounds were detected. These populations were differentially distributed among lake sectors during their feeding migration and their spatial distribution was stable over the two sampling seasons. Dispersing individuals were identified (n=61); these revealed asymmetrical patterns of dispersal between populations, which was also confirmed by divergent admixture proportions. Regression models underlined population of origin as the only factor explaining differential dispersal of individuals among populations. An analysis of covariance (ancova) indicated that larger individuals tended to migrate from their river of origin further away in the lake relative to smaller fish. In summary, this study underlined the relevance of using individual-based assignment methods for deciphering dynamics of connectivity among wild populations, especially regarding behavioural mechanisms such as differential spatial partitioning and dispersal responsible for the maintenance of genetic population structure.