AB048. X-chromosomal SNPs variation in populations of Russia

Proportioning Whole-Genome Single-Nucleotide–Polymorphism Diversity for the Identification of Geographic Population Structure and Genetic Ancestry

Background: Global trade of plant products is a major driving force for the unintended spread of economically harmful insect pests. This PhD thesis aimed at (i) developing and implementing molecular tools for the on-site identification of invasive insect pests at points of entry (POEs) for plant import products as a prevention measure; and (ii) investigating the invasion history of the mosaic leafhopper Orientus ishidae, a potential vector of grapevine Flavescence doree phytoplasma. Methods: To achieve the first goal, loop-mediated isothermal amplification (LAMP)-based genetic assays for the rapid on-site identification of Bemisia tabaci, Thrips palmi and several invasive fruit flies of the genera Bactrocera and Zeugodacus were developed. Using publicly available DNA sequences, LAMP primers were designed to specifically target a fragment of the mitochondrial cytochrome c oxidase subunit 1 gene. To address the second goal of this PhD thesis, the invasion genetics of O. ishidae was studied, an invasive insect species that spread from its native range from in East Asia to North America in the first half of the 20th century and only recently colonised Europe. Possible source populations and invasion pathways were investigated by assessing the genetic structure of 41 O. ishidae populations from Asia, Europe, and North America based on a mitochondrial marker and 641 single nucleotide polymorphisms (SNPs) generated by double digest restriction-site associated DNA (ddRAD) sequencing. Results: Validation performed under laboratory and on-site conditions demonstrated the robustness and reliability of the developed LAMP identification assays. Analysing 319 insect specimens, the overall diagnostic test efficiency was 98% and the overall diagnostic test specificity was 100%. The small number of false-negative results (2%) originated either from previously unknown biotypes, not included in the initial primer design, or from handling errors during LAMP preparation. The results from the molecular genetic analyses of O. ishidae revealed a clear genetic separation between a native population from Asia and the non-native populations from Europe and North America. Among the non-native populations, only faint signals of spatial genetic structuring were found. However, when comparing non-native populations from Europe and North America, elevated levels of admixture of genetically distant mitochondrial haplotypes were observed for European populations. Conclusion: Characterised by high analysis speed (<1 h) and simplicity in use (only 1 pipetting step), the validated LAMP assays were found to be suitable identification tools for on-site application by plant health inspectors. Since completion of the validation phase, the developed identification assays are routinely deployed in the phytosanitary import control process of Switzerland. The considerable genetic separation between native and non-native populations of O. ishidae together with the strikingly high genetic similarity of European and North American populations suggest an invasion scenario in which North American populations served as source for the European invasion. A slightly reduced genetic structure combined with increased admixture of genetically distant mitochondrial haplotypes furthermore indicate that the European colonisation history was shaped by multiple introductions from North America, complemented by frequent intra-European gene flow. Taken together, it is hypothesised that the overall genetic complexity of non-native populations was strongly driven by frequent international trade of plants infested by O. ishidae.

Although ability to digest lactose generally declines after weaning in all mammals, in some human populations it persists also in adult individuals, a condition named lactase persistence (LP). Studies on the prevalence of the LP phenotype in worldwide human populations have shown that the frequency of this trait is highly variable in different ethnic groups, appearing to be positively correlated with the importance of milk in the diet. In particular, several single-nucleotide polymorphisms (SNPs) in the proximity of the LCT gene have been proved to be associated with LP. Nevertheless, few studies have till now analyzed genetic variation underlying LP in a wide set of Eurasian populations and, especially, in the Italian one. In the present study, we thus typed 40 SNPs surrounding the LCT gene in more than 1,000 samples from Italian and Arabic peninsulas to investigate patterns of LP-related genetic diversity in two regions which have played a pivotal role in the recent human evolutionary history according to their geographical position and historical/archaeological records. Our results underline a high and complex variability of the explored genomic region in both studied populations. In particular, a clear diversification of Northern Italian groups from the rest of the peninsula, was observed, with the formers being genetically more similar to Northern European populations than to Southern Italians. These observation are consistent with known decreasing pattern of LP from Northern to Southern Italy and suggest the possibility of an independent evolution of LP-associated genotypes in Northern Italy. A similar scenario was observed in the Arabian peninsula, with Dhofari Arabs from Southern Oman and Yemeni clustering together with respect to Arabs from Northern Oman and the subgroup of Omanis of Asian origin which appeared instead to be genetically closer to Europeans than to the rest of Arabic groups.

AimHow historical and contemporary eco‐evolutionary processes shape the patterns of genetic diversity and differentiation across species’ distribution range remains an open question with strong conservation implications. Focusing on the orange stony coral, Astroides calycularis, we (a) characterized the pattern of neutral genetic diversity across the distribution range; (b) gave insights into the underlying processes; and (c) discussed conservation implications with emphasis on a national park located on a hotspot of genetic diversity.LocationSouth Mediterranean Sea and Zembra National Park.MethodsWe combined new data from 12 microsatellites in 13 populations located in the Centre and in the Western Periphery of the distribution range with a published dataset including 16 populations from the Western and Eastern Peripheries. We analysed the relationship among parameters of genetic diversity (He, Ar(g)) and structure (population‐specific FST) and two measures of geographic peripherality. We compared two estimators of pairwise genetic structure (GST, DEST) across the distribution range. The evolutionary and demographic history of the populations following the Last Glacial Maximum was reconstructed using approximate Bayesian computations and maximum‐likelihood analyses. We inferred the contemporary connectivity among populations from Zembra National Park and with the neighbouring area of Cap Bon.ResultsWe demonstrate a decrease in genetic diversity and an increase in genetic differentiation from the Centre to the Eastern and Western Peripheries of the distribution range. Populations from Zembra show the highest genetic diversity reported in the species. We identified a spillover effect towards Cap Bon.Main conclusionsThe patterns of genetic diversity and differentiation are most likely explained by “the postglacial range expansion hypothesis” rather than the “central–peripheral hypothesis.” Enforcement of conservation measures should be considered to protect this genetic diversity pattern, in particular when considering the low effective population size inferred at many sites.

The history of livestock started with the domestication of their wild ancestors: a restricted number of species allowed to be tamed and entered a symbiotic relationship with humans. In exchange for food, shelter and protection, they provided us with meat, eggs, hides, wool and draught power, thus contributing considerably to our economic and cultural development. Depending on the species, domestication took place in different areas and periods. After domestication, livestock spread over all inhabited regions of the earth, accompanying human migrations and becoming also trade objects. This required an adaptation to different climates and varying styles of husbandry and resulted in an enormous phenotypic diversity. Approximately 200 years ago, the situation started to change with the rise of the concept of breed. Animals were selected for the same visible characteristics, and crossing with different phenotypes was reduced. This resulted in the formation of different breeds, mostly genetically isolated from other populations. A few decades ago, selection pressure was increased again with intensive production focusing on a limited range of types and a subsequent loss of genetic diversity. For short-term economic reasons, farmers have abandoned traditional breeds. As a consequence, during the 20th century, at least 28% of farm animal breeds became extinct, rare or endangered. The situation is alarming in developing countries, where native breeds adapted to local environments and diseases are being replaced by industrial breeds. In the most marginal areas, farm animals are considered to be essential for viable land use and, in the developing world, a major pathway out of poverty. Historic documentation from the period before the breed formation is scarce. Thus, reconstruction of the history of livestock populations depends on archaeological, archeo-zoological and DNA analysis of extant populations. Scientific research into genetic diversity takes advantage of the rapid advances in molecular genetics. Studies of mitochondrial DNA, microsatellite DNA profiling and Y-chromosomes have revealed details on the process of domestication, on the diversity retained by breeds and on relationships between breeds. However, we only see a small part of the genetic information and the advent of new technologies is most timely in order to answer many essential questions. High-throughput single-nucleotide polymorphism genotyping is about to be available for all major farm animal species. The recent development of sequencing techniques calls for new methods of data management and analysis and for new ideas for the extraction of information. To make sense of this information in practical conditions, integration of geo-environmental and socio-economic data are key elements. The study and management of farm animal genomic resources (FAnGR) is indeed a major multidisciplinary issue. The goal of the present Research Topic was to collect contributions of high scientific quality relevant to biodiversity management, and applying new methods to either new genomic and bioinformatics approaches for characterization of FAnGR, to the development of FAnGR conservation methods applied ex-situ and in-situ, to socio-economic aspects of FAnGR conservation, to transfer of lessons between wildlife and livestock biodiversity conservation, and to the contribution of FAnGR to a transition in agriculture (FAnGR and agro-ecology).

Camellia oleifera is a subtropical evergreen plant. Cultivated C. oleifera is the most important woody oil crop in China. Wild C. oleifera is an essential genetic resource for breeding. The patterns of genetic differentiation among altitudes/latitudes in wild C. oleifera are still unknown. Camellia oleifera may be predominantly hexaploid. The characteristics of polyploidy may lead to considerable biases in estimates of genetic diversity and differentiation. Our study used C. oleifera as a case study for analysing genetic diversity, structure and differentiation in polyploid plants using simple sequence repeats (SSRs). Wild C. oleifera samples were collected at different altitudes on the Jinggang and Lu mountains of China. The ploidy levels were determined with flow cytometry analysis. Eight highly polymorphic SSRs were used to genotype the samples. Genetic diversity and structure were analysed. Various estimates of genetic differentiation were compared. The flow cytometry results indicated that wild C. oleifera samples were all hexaploid at various altitudes of the Jinggang and Lu mountains. High levels of genetic diversity were found on both the Jinggang and Lu mountains. Genetic structure analyses indicated clear genetic differentiation between the Jinggang and Lu mountains and lower genetic differentiation among altitudes within each mountain. Classical genetic differentiation estimates of Fst failed to discriminate genetic differentiation between and within mountains. The Rho statistic showed a moderate level of genetic differentiation between mountains and lower levels of genetic differentiation within each mountain. Our study demonstrates that Rho is the statistic of choice for estimating genetic differentiation in polyploids.

Patchy distribution and stressful conditions of environment can induce the emergence of locally adapted phenotypes. Evolutionary theory supports that local adaptation is drown by strength of divergent selection to favor the genotype that better performs in a specific habitat. Nevertheless, adaptation could occur also via phenotypic plasticity that allows individuals to rapidly change their phenotypic response to environment and this ability may even slow down the effect of adaptive genetic divergence. Plants from serpentine represent a typical model for studying local adaptation to soil type as selection in this environment is very intense and leads to the evolution of locally adapted populations, a phenomenon known as “serpentine syndrome”. Dianthus sylvestris Wulfen (Caryophyllaceae) is frequently found both on serpentine and limestone bedrocks along Apennine chain. Here we investigated populations of D. sylvestris in North-Center of Italy in order to clarify if phenotypic variation among populations of D. sylvestris, on both serpentine and limestone soils, could be defined as an example of local adaptation or is due to strong phenotypic plasticity. We used a molecular approach based on EST SSR marker to infer on genetic diversity and populations structure. Moreover, to verify if serpentine populations are locally adapteted we use an ecologiac approach based on transplanting field experiment and morphological and physiological measurements. Population genetic analyses showed a high percentage of polymorphic loci (ranging from 71% to 100%) and the distribution of allele frequencies showed no significant differences among populations from the two soil types. Similarly, allele richness was comparable among populations of serpentine and limestone. Both ANOVA and the low values of differentiation among populations (mean Fst= 0.119, among populations) confirmed the low overall genetic differentiation. Bayesian (STRUCTURE) and multivariate approach (PCoA) ruled out that populations from limestone and serpentine soils cluster in two genetically differentiated groups, even if according to Mantel test, subdivision was on geographic distribution more than on edaphic base. Thus, no evident genetic differentiation among D. sylvestris populations from serpentine and limestone was found with neutral markers. To determine the contribution of selective factors and/or phenotypic plasticity to local adaptation of D. sylvestris to serpentine, from populations already examined in the genetic analysis, we estimated the metal content in plant aerial parts, collected data on morphological traits, and performed field reciprocal transplantations. High metal content (Ni, Cr) in plants aerial part confirmed, as in previous studies, the bioaccumulation of heavy metals in D. sylvestris plants from serpentine soils. In these plants, several morphological traits were found statistically decreased when compared to plants from limestone so highlighting that serpentine is a less permissive habitat than limestone. However, most of the morphological differences disappeared in transplanted individuals suggesting a large contribution of phenotypic plasticity in determining the observed morphological divergences. Nevertheless, in transplanted plants from serpentine soil to limestone, a two-way ANOVA resulted in a significant difference in biomass with an effect of the original soil on the transplanting soil. Significant differences were also found in flowering time, as plants from serpentine, when transplanted on limestone, flowered before than resident limestone plants. These differences, persisting independently from the original soil type, should have genetic bases. Thus genetic differentiation of populations of D. sylvestris is occurred at least in a few selected loci determining different affinity for the two habitats. This divergence is maintained among populations from different soil types even in the face of extensive gene flow as observed at neutral loci.

Distribution margins constitute areas particularly prone to random and/or adaptive intraspecific differentiation in plants. This trend may be particularly marked in species discontinuously distributed across mountain ranges, where sharp geographic isolation gradients and habitat boundaries will enhance genetic isolation among populations. In this study, we analysed the level of neutral genetic differentiation among populations of the long-lived shrub Daphne laureola (Thymelaeaceae) across the Baetic Ranges, a glacial refugium and biodiversity hotspot in the western Mediterranean Basin. Within this area, core and marginal populations of D. laureola were compared with regard to their spatial isolation, size, genetic diversity and differentiation. A spatially explicit analysis conducted on the vast majority of the species' known populations in the study area (N = 111) showed that marginal populations (western and eastern) present larger spatial isolation than core populations, but are not smaller. We compared genetic diversity and differentiation between core and marginal populations using a subsample of 15 populations and 225 amplified fragment length polymorphism (AFLP) markers. Core and marginal populations did not differ in genetic diversity, probably because of the occurrence of large populations on the local margins. Western populations were strongly differentiated from the other populations. In addition, spatial and genetic differentiation among populations was larger on the western margin. Eastern populations constituted a genetically homogeneous group closely related to core populations, despite their greater spatial isolation. Results suggest that studies on phenotypic differentiation between core and marginal populations of D. laureola, and presumably other species having discontinuous distributions across the Baetic ranges, should take into account geographical differences in levels of genetic differentiation between the different distribution borders.

This research was undertaken with the aim to identify the robustness of available genetic markers (chloroplast DNA sequences and nuclear microsatellites) to differentiate G. parviflora subsp. parviflora (Gpp) from five closely related taxa within the Linearifolia Group of grevilleas; particularly G. humilis subsp. humilis (Ghh) in the Lake Macquarie area. One chloroplast region had a consistent character that separated the northern populations of Gpp (northern: Hunter area) from populations in the south of its range (southern: Cumberland Plains area) and all other taxa. This character was an 11 base pair sequence inversion. An assessment of over 50 microsatellite loci led to the identification of nine primer pairs that consistently amplified variable product in the target taxon. Genetic profiling using the microsatellites from a minimum of 10 individuals per taxon demonstrated a number of important issues: (1) some individuals amplify three alleles at one or more loci, suggesting possible aneuploidy or triploidy; (2) some individuals from the same population have identical, or near identical, genotypes which may be indicative of vegetative propagation; (3) genetic profiles among all samples were complex and the microsatellite loci provided moderate support for some of the taxa while other samples suggest a likely history of admixture, hybridisation, or the mis-identification of plant material. These possibilities need to be assessed along with morphological studies being undertaken by Mr Bob Makinson as a complementary component to this research. Particular consideration should be given to the southern populations of the focal taxon which, based on cpDNA and microsatellites, were genetically more similar to the geographically closer G. parviflora subsp. supplicans. Overall, the genetics tools (cpDNA and microsatellites) show great promise for delineating northern populations of G. parviflora subsp. parviflora and for deeper investigations into population genetic structure including the number of unique genotypes within and among populations, the physical spread of clonal genotypes within populations, the genetic differentiation and the genetic health (as assessed by heterozygosity and inbreeding) of populations of this threatened taxon.

Salt sensitivity is an independent cardiovascular disease and mortality risk factor, present in both hypertensive and normotensive population. It is genetically determined and it may affect the relationship between salt taste perception and salt intake. The aim of this study was to explore the genetic predisposition to salt sensitivity in young and middle-aged adult population and its effects on salt taste perception and salt intake. Salt sensitivity was investigated in 20 normotensive subjects and defined as the change in blood pressure (BP) after 7 days of low-salt (51.3 mmol sodium/day) and 7 days of high-salt diet (307.8 mmol sodium/day). Salt taste perception was identified using British Standards Institution sensory analysis method (BS ISO 3972:2011). Salt intake was assessed with a validated food frequency questionnaire. DNA was genotyped for single nucleotide polymorphisms in the SLC4A5, SCNN1B and TRPV1 genes coding for sodium and ion channels and transporters. The subjects with the rs7571842 (SLC4A5) AA genotype exhibited the highest increase in BP (systolic BP (∆SBP)=7.75 mmHg, p=0.002; diastolic BP (∆DBP)=6.25 mmHg, p=0.044; mean arterial pressure (∆MAP)=6.5 mmHg, p=0.014). There was an association between salt taste perception and salt sensitivity (rs=0.551, p=0.041) with salt intake being unaffected. There is a genetic predisposition to salt sensitivity and it is associated with salt taste perception. Unaffected salt intake suggests that factors other than taste sensitivity and/or other genetic variants may have more pronounced effects in healthy adults. Moreover, experiments are conducted in our laboratory to increase the understanding of the SLC4A5 effects on salt-sensitive changes in BP. Urine samples have been collected from subjects phenotyped for salt sensitivity to measure the expression of this co-transporter. If differently expressed in salt-sensitive and salt-resistant subjects, together with its genotype, it may serve as a personalised salt sensitivity biomarker.

The first part of this PhD thesis is devoted to the application of molecular genetics to describe demographic trends, geographic distribution patterns and genetic status of the Alpine population 15 years after the reintroduction program. In order to achieve the first objective we a) increased the number of STR markers (from 10 to 15 loci) in the Alpine population to raise the informativity content for population genetics studies, and possibly resolve uncertain parentage assignments; b) presented an annual overview of the demographic status of the Alpine population for a long-term monitoring program; c) measured the genetic diversity over generations, highlighting possible trends d) identified parentage relationships and provided a pedigree reconstruction, showing if there is an increase of inbreeding events over generations; e) estimated the effective population size; f) verified whether or not a connection between the reintroduced population in the central Alps and the Dinaric population was established. We furthermore provided considerations for conservation and management of this species in the Alps, taking into account the emerged demographic, spatial and genetic aspects. The second part is methodological and is about developing a new set of SNP markers to enhance the resolution power for population genetics analysis of the Alpine and Apennine populations. In order to identify reliable and informative SNPs we a) tested the effectiveness of an existing SNP panel, developed for the Scandinavian brown bear populations, on the two Italian brown bear subspecies, identifying a set of SNPs that has potential for a SNP-based individual and sex identification system. I took into consideration the ascertainment bias that arises when transferring SNP markers across populations; and b) tested the selected subset of SNPs for parentage assignments in the Alpine population, comparing its resolution power with that derived from STRs.

My PhD project was focused on Atlantic bluefin tuna, Thunnus thynnus, a fishery resource overexploited in the last decades. For a better management of stocks, it was necessary to improve scientific knowledge of this species and to develop novel tools to avoid collapse of this important commercial resource. To do this, we used new high throughput sequencing technologies, as Next Generation Sequencing (NGS), and markers linked to expressed genes, as SNPs (Single Nucleotide Polymorphisms). In this work we applied a combined approach: transcriptomic resources were used to build cDNA libreries from mRNA isolated by muscle, and genomic resources allowed to create a reference backbone for this species lacking of reference genome. All cDNA reads, obtained from mRNA, were mapped against this genome and, employing several bioinformatics tools and different restricted parameters, we achieved a set of contigs to detect SNPs. Once a final panel of 384 SNPs was developed, following the selection criteria, it was genotyped in 960 individuals of Atlantic bluefin tuna, including all size/age classes, from larvae to adults, collected from the entire range of the species. The analysis of obtained data was aimed to evaluate the genetic diversity and the population structure of Thunnus thynnus. We detect a low but significant signal of genetic differentiation among spawning samples, that can suggest the presence of three genetically separate reproduction areas. The adult samples resulted instead genetically undifferentiated between them and from the spawning populations, indicating a presence of panmictic population of adult bluefin tuna in the Mediterranean Sea, without different meta populations.

Understanding the factors that contribute to loss of genetic diversity in fragmented populations is crucial for conservation measurements. Land-bridge archipelagoes offer ideal model systems for identifying the long-term effects of these factors on genetic variations in wild populations. In this study, we used nine microsatellite markers to quantify genetic diversity and differentiation of 810 pond frogs (Pelophylax nigromaculatus) from 24 islands of the Zhoushan Archipelago and three sites on nearby mainland China and estimated the effects of the island area, population size, time since island isolation, distance to the mainland and distance to the nearest larger island on reduced genetic diversity of insular populations. The mainland populations displayed higher genetic diversity than insular populations. Genetic differentiations and no obvious gene flow were detected among the frog populations on the islands. Hierarchical partitioning analysis showed that only time since island isolation (square-root-transformed) and population size (log-transformed) significantly contributed to insular genetic diversity. These results suggest that decreased genetic diversity and genetic differentiations among insular populations may have been caused by random genetic drift following isolation by rising sea levels during the Holocene. The results provide strong evidence for a relationship between retained genetic diversity and population size and time since island isolation for pond frogs on the islands, consistent with the prediction of the neutral theory for finite populations. Our study highlights the importance of the size and estimated isolation time of populations in understanding the mechanisms of genetic diversity loss and differentiation in fragmented wild populations.

Life finds a way: the recovery of frog populations from a chytridiomycosis outbreak

This thesis examined if signals of selection are present in the bottlenecked and reintroduced populations of the Alpine ibex (Capra ibex). By utilizing single nucleotide polymorphism (SNP) data, I identified weak signals of purifying selection and positive selection. Furthermore, I quantified the detection accuracy achievable for studies using genetic outliers or associations among allele frequencies and environmental variables to detect positive selection after a bottleneck. Additionally, I discussed the biases present in a high-throughput sequencing dataset due to batch effects that can arise in long-term studies where sequencing data is added incrementally. Finally, in this thesis I discussed the importance of considering genetics when planning the reintroduction of a species. Population bottlenecks can have profound and long-lasting genetic consequences. Due to their reduced effective population size, bottlenecked species experience strong genetic drift, loss of genetic variation, and increased inbreeding. They are therefore at risk of maladaptation and extinction. Despite these risks examples of thriving bottlenecked populations are known. Outside of conservation biology, population bottlenecks are important evolutionary forces because they can affect the rate and direction of adaptation. As a result, identifying examples of selection after a bottleneck is of importance to evolutionary and conservation biology. In this thesis, I utilised genome-wide SNP data for 27 populations of Alpine ibex, including the remnant population in the Gran Paradiso National Park. This data was generated with restriction site associated DNA sequencing (RADseq). RADseq is a high-throughput sequencing method that sequences genomic DNA around a restriction enzyme site. I identified over 6000 SNPs in the Alpine ibex genome. With this data, I identified putative signals of purifying selection by comparing exonic SNPs, which are likely under selection, with intronic SNPs and SNPs in intergenic regions, that are expected to be largely neutral. Furthermore I examined the ratio of non-synonymous to synonymous sites. The heterozygosity of exonic SNPs was significantly below that of introns and of intergenic SNPs. In addition, the ratio of non-synonymous to synonymous sites was below one. While this suggests purifyingii selection, due to marker and test limitations, these results are not conclusive and the presence of purifying selection should be viewed with caution. I then searched for signals of positive selection by scanning for large differences in allele frequencies among populations and for correlations between allele frequencies and an environmental variable. The high rates of genetic drift in bottlenecked populations can create false signals of positive selection when using such methods. Therefore, I used a population genetic (forward-time) simulation approach that followed Alpine ibex demography, to generate a simulated set of SNPs including neutral loci and loci that were under selection. I then used these loci to quantify the accuracy of three selection detection methods. To this end, I examined the number of false positive neutral SNPs identified by each method, as well as the number of true positive and false negative simulated selected SNPs. I found that a true discovery rate of over 70% can be achieved by combining three selection detection methods to identify “triple positive” SNPs, and an environmental correlation detection approach. When I applied the selection detection methods to the Alpine ibex empirical RADseq dataset no triple positive SNPs were identified by the triple positive environmental correlation approach. Thus there are no SNPs I confidently identified as under selection, though weak candidates were found by the lower accuracy methods (30- 50% true discovery rate) that may be suitable for further examination. High-throughput sequencing is maturing and an increasing number of studies have used data obtained over time or generated by different investigators. In this thesis, I also discuss the biases and errors, so-called ‘batch effects’, that can be introduced into a study if subsets of data differ in how they were obtained and contain different technical artefacts. I present a case study in the Alpine ibex where batch effects lead to a misleading biological conclusion. Finally in an additional co-authored publication, the importance of considering the long-term genetics of a population during a reintroduction was presented and discussed.