The spatial distribution of genetic variants is jointly determined by geography, past demographic processes, natural selection, and its interplay with environmental variation. A fraction of these genetic variants are “causal alleles” that affect the manifestation of a complex trait. The effect exerted by these causal alleles on complex traits can be independent or dependent on the environment. Understanding the evolutionary processes that shape the spatial structure of causal alleles is key to comprehend the spatial distribution of complex traits. Natural selection, past population size changes, range expansions, consanguinity, assortative mating, archaic introgression, admixture, and the environment can alter the frequencies, effect sizes, and heterozygosities of causal alleles. This provides a genetic axis along which complex traits can vary. However, complex traits also vary along biogeographical and sociocultural axes which are often correlated with genetic axes in complex ways. The purpose of this review is to consider these genetic and environmental axes in concert and examine the ways they can help us decipher the variation in complex traits that is visible in humans today. This initiative necessarily implies a discussion of populations, traits, the ability to infer and interpret “genetic” components of complex traits, and how these have been impacted by adaptive events. In this review, we provide a history-aware discussion on these topics using both the recent and more distant past of our academic discipline and its relevant contexts. SignificanceComplex traits such as height, educational attainment, or diseases like hypertension vary across the globe due to the interplay of genetics and the environment. In this review, we describe the evolutionary processes that could influence complex trait variation and highlight the challenges of separating the environmental and genetic contribution to a complex trait. We discuss these topics in light of the advantages and limitations of current approaches for studying the genetic basis of complex traits.One of the main questions of interest in human genetics is to understand the genetic factors driving the phenotypic diversity observed among individuals. This is a difficult enterprise because the majority of the phenotypes of interest are complex traits that are jointly determined by many genetic loci and environmental variables. Genome-wide association studies (GWAS) have allowed us to make progress in identifying the genetic loci that influence the phenotypic diversity of a complex trait in a particular study cohort and environment. This has allowed us to start identifying similarities and differences in the genetic architecture of a complex trait across human diversity. Here, the genetic architecture is defined by the number, effects, frequencies, and heterozygosity of the genetic variants changing the value of the trait in a particular environment, taking into account the interactions of the variants with each other and the environment . The unrResearchers typically try to make sense of biological variation by grouping individuals based on shared observed characteristics. This is a complicated task that can be approached differently depending on the purpose of the definition that we seek. Focusing on natural populations, Waples and Gaggioti (2006) categorize two main approaches to define populations, the ecological approach and the evolutionary approach. The first one proposes that individuals of a species belong to the same population based on interactions within the same space and time but not necessarily requiring reproduction, whereas the latter requires the potential for reproduction which implies being in close enough proximity to mate . HoweverDespite the problems of the definition of populations in space, the concise definition of populations in theoretical models has been useful to advance our understanding of the spatial patterns of genetic diversity. As an example, models of populations with predefined sizes and migration rates that have a clearly delimited space have helped us elucidate important concepts such as the patterns of isolation by distance (IBD). IBD is a very robust prediction derived from a mathematical framework and posits that there will be more genetic differences between individuals that are farther away from each other geographically under a continuous spatial range . This ovIt is important to note here that idealized population definitions that aid the development of mathematical models do not generally present an accurate representation of present-day human diversity, hence one should be wary when mapping these constructs to empirical biological data. In practice, the fields of human genetics and genomics have grouped individuals into populations with a certain ambiguity, where populations have been used to refer to races, ethnic groups, individuals sharing genetic ancestry (see Appendix—Keywords), a nationality, a religion, or a geographic region, with multiple definitions used interchangeably . AlthougTo prevent such essentialization of discrete ancestry labels, it is reasonable to describe them as outputs of specific methods such as admixture with certain assumptions, and label them as ancestry from present-day Europe instead of European ancestry for example. Furthermore, when determining how to analyze individuals in genetic and genomic studies, it would be advisable to select grouping criteria that are most in alignment with the research questions being asked, as well as clearly stating such criteria and why they are important for the analysis. For example, how do you define and label groups and why? Should your grouping be defined by genetic ancestry (and at which time slice), self-identified race, or some other variable? As such, it is also advisable to group only when it is justified by the research question and not by default. Additionally, when sampling is limited or not uniform across a grouping criterion, it should be pointed out that there is insufficient data to generalize particular results to the larger group.The concept of population is commonly used to group individuals for further analyses including those of trait variation. Traits are often described as a phenotypic characteristic that can be either measured or directly observed in an organism. One particularly useful definition was put forward by Dobzhansky who defined a “visible trait” as “the outcome of certain developmental, physiological and ultimately physico-chemical processes in the organism”  noting tTraits such as height that show continuous variation are referred to as “quantitative” or “complex” traits. Common disorders that seem binary such as Type 2 Diabetes (T2D) are also quantitative traits . R.A, FiFinding statistically conclusive support for epistasis is a challenging task mainly due to it having small effects, and its contribution to complex traits remains to be accurately quantified . On the Beyond genetic background, complex traits can be heavily influenced by the contribution of the environmental background that alters the processes in the organism giving rise to the trait. These gene by environment interactions and environmental effects make it complicated to assess how much of the variation of a trait comes from genetics or the environment. In this sense, estimating heritability is always a local analysis , for eitGWAS are our current best tool to analyze the genetic basis of complex traits in humans . To do tTwo primary statistical models are used to estimate genetic effect sizes a. The fiGenetic effects can be overestimated if an unknown environmental factor is differentiated along the same axis as the studied trait . This caCorrelation among causal variants induced by assortative mating can confound their genetic effect estimates from GWAS, inflating the additive genetic variance beyond its value under random mating  fig. 4ccfig. 4c.Investigating the relationship between spatial genetic and trait structure often requires performing predictions of complex phenotypes using SNP effects recovered in GWAS e. These Past demographic history plays an important role in the prediction of phenotypes. Its impact is mainly due to a larger genetic divergence (as measured by Fst) between the training population used to obtain the effects of associated SNPs via a GWAS study and the target population where we perform the predictions. This impact is manifested in two main ways: differences in the frequency of causal alleles and varying patterns of LD . Allele Potential variation of SNP effect sizes in different human populations may be another factor affecting the accuracy of PGSs. A recent study examined this problem by looking at the cross-population genetic correlation between effect sizes in regions of the genome with a particular functional annotation . The autFTO locus  in a female prediction set. In turn, the PGS resulting from the male GWAS cohort was similarly accurate for both sexes. Similarly, BMI and years of schooling were also sensitive to the choice of GWAS cohort. When PGS was constructed from GWAS of the youngest group (BMI) and lowest SES (years of schooling), it had a higher prediction across all groups. Whether these effects are widespread across other traits or group classifications is unknown, but for these three traits, prediction accuracy of PGS can vary in individuals of a shared genetic ancestry based on sex, age, and SES. A broader conclusion from these results is that the prediction accuracy of PGS between individuals with a different ancestry will also be affected by factors, such as SES or age, when those factors change between groups.Some solutions have been proposed to improve the prediction of phenotypes using data from multiple studies. One potential problem is that phenotypic predictions in a particular target population can be inaccurate because they are based on GWAS data using a small sample size from that target population. Due to this small sample size, the effect sizes estimated are inaccurate and lead to poor phenotypic predictions. However, one solution that has been proposed to tackle this problem is to create a multiethnic PGS that includes a linear combination of two PGSs from: 1) A study cohort where accurate effect sizes can be obtained due to a large sample size available, and 2) a study cohort with less accurate effect sizes due to a smaller sample size available . This apR2 values compared with PGSs that only use effect sizes from one particular ancestry  by ancestry and have only shown modest improvement in polygenic prediction. Meanwhile, the PRS-CSx approach assumes a mostly shared genetic architecture across ancestries, and boosts power by leveraging summary statistics and LD information across populations while allowing for population-specific effects as well. Finally, the field is also attempting transancestry GWAS in an attempt to increase prediction power across groups . It is aGWAS are used to identify trait-associated alleles that are in LD with causal alleles. Those causal alleles, which change the value of a complex trait, define the genetic component of a complex trait. Understanding the evolution of those causal variants around the globe is of fundamental importance to understand the spatial distribution of complex traits. The global spatial distribution of these causal alleles across the globe, as of other genetic variants, is broadly determined by degrees of genetic differentiation that are correlated with geographical distances  and geogThe environmental conditions present in different regions can lead to selection of particular phenotypic trait values and lead to variation in complex traits across geographies that mirrors genetic variation. The relationship between the reproductive success (defined as fitness) of an individual and their trait value determines the type of selection acting on that trait. In humans, the reproductive success is a reflection of natural and biological factors along with cultural factors that also play a strong role. Conceptually, natural selection acting on a trait has been broadly classified as directional selection, stabilizing selection, and disruptive selection .N at each time and the differences in fitness between individuals that possess a particular genotype. Broadly, those changes in frequency are higher when N is larger. We illustrate the impact of population size changes on allele frequency changes using simulations , genetic architecture and the environment will compensate each other to maintain the trait at an intermediate value . As showSimilar to genetic variants, complex traits also present their own spatial patterns. Statistical and population genetics research has been concerned with identifying the genetic factors that may underlie such variation in trait values or disease incidence. How well do spatial genetic patterns mirror spatial trait patterns? What other correlated environmental patterns may be mirroring trait patterns? Complex traits can vary across genetic and environmental axes. Simply conceived, the environmental axis can be biogeographical or sociocultural. Biogeographical axes are structured by the conditions along which climatic and ecological variables may change . Sociocultural axes are arranged by differences in diet, income, access to education, access to healthcare, ethnicity, and experience of racism and discrimination. The genetic axes capture variation in allelic frequencies and in levels of genomic homozygosity. This genetic variation at a given time is shaped by ancestral demographic histories and is often captured using the concept of genetic ancestry. Biogeographical, sociocultural, and genetic axes are often correlated in complex ways, depending on the specific environmental and cultural history of a place and people. Therefore, disentangling them and their interactions presents a very difficult problem. In particular, gene–environmental correlations (e.g. between a specific ancestry and discrimination) can cause the environment to lead to trait variation that can then be mistakenly perceived as caused by genetics .The case of hypertension will provide an illustrative example. Hypertension is observed at a higher rate in the United States in African Americans compared with European Americans . One maySlavery, colonization, and decolonization along skin color lines are processes that have underscored global history in the last 500 years. To understand traits like hypertension, we have to understand how those dynamics impacted the individuals studied. One way to do so is using a comparative method to look across populations with different histories (e.g. African Americans vs Africans in Africa). Indeed, as a wider selection of people have been sampled, it has become evident that Africans in Africa, especially rural Africans, actually have some of the lowest levels of hypertension in the world  fig. 6)fig. 6). Another study highlighting the importance of sociocultural axes found that blood pressure in Puerto Rico showed an association with skin color as defined through social classification using the ethnographic method of cultural consensus analysis which estimates how respondents are perceived by others in social interactions but not skin color as measured through reflectance spectrophotometry . The autSeveral present-day societies have been structured along lines of color and race with respect to social standing and opportunity which can lead to observable variation in complex traits. Within the general phenomenon of cultural contact, diets have also gone through larger turnovers for certain people and places during colonization or due to westernization driven partly by imperialism. This look at the spatial patterns of hypertension and blood pressure in a global context illustrates that purely genetic explanations for complex trait variation should be avoided when other correlated sociocultural variables, or an interaction between genetic and sociocultural variables, may be more likely to explain the patterns of variation within a single country with a specific history where discrimination has been correlated with skin color  or where big environmental shifts such as those in diet have taken place for certain groups.The joint action of genetics and the environment determine the final trait value in a given individual or group. Due to this, it is important to consider the role of the environment when performing phenotypic predictions based on the genetic information from PGS. To illustrate this point, we will study the relationship between PGS and trait values in groups distributed across space. First, we will consider two populations in Japan studied with respect to their trait distributions and their PGS distributions. A recent study looked at a number of different complex traits in individuals from mainland Japan and nonmainland Japan . The autPlasmodium vivax and Plasmodiumfalciparum. Predicted genetic risk for malaria infections is lower in Choco, whereas the malaria prevalence is higher in Choco compared with Antioquia because both P. vivax and P. falciparum are more prevalent in Choco. Overall, these results point to the importance of environmental factors such as diet, lifestyle, and the abundance of pathogens as elements that can play a role in causing discrepancies between polygenic phenotypic predictions and trait values.For two populations in Colombia, Antioquia (Mestizo) and Choco (Afro-Columbian), Next, we will consider how a widely studied trait, educational attainment (EA), and its PGS varies by birth year in the UK Biobank. Abdellaoui et al. used summary statistics from a meta-analysis performed on a large number of cohorts of European descent ancestry to compute PGS for EA. Principal components computed on their prediction cohort (UK Biobank) were later regressed out of the PGSs to correct for stratification effects. Across the country, EA in Great Britain shows a positive correlation with birth year, whereas the PGS for EA shows a negative correlation with birth year . That isGiven the detailed prior discussion of problems of biased estimation of genetic effects and of variable powers of prediction, the reader is justified in wondering if the results presented in the section may be due to the same. This is certainly possible and remains an area of active research. Nevertheless, we think that these examples are instructive to make the point that a trait’s value results from a combination of genetic and environmental processes. An often underlooked cause for the discrepancy between a PGS prediction and an actual trait value is the role of environmental factors. It is in this sense that a PGS can never be considered determinative of a trait value, as it is only one aspect of trait/disease manifestation . As suchAs described before, the spatial environmental structure is an important factor to take into account when describing the evolution of complex traits. The environment can change the distribution of a trait in a particular location due to the effect of natural selection. In this section, we will discuss the main findings of studies conceptualizing and elucidating the roles of stabilizing, disruptive, and directional natural selection acting on complex traits.The contributions of stabilizing and disruptive selection have been recently studied in a set of complex traits using data from the UK Biobank . Sanjak Populations can adapt to new environmental conditions through directional selection on complex traits. This evolutionary force has been hypothesized to drive phenotypic differences between different populations and, therefore, has motivated a vigorous line of research during the past decade. However, the detection of polygenic adaptation is challenging because the signal depends on frequency shifts at many alleles. There will not be a LD signal around the alleles that is indicative of the action of natural selection if those allele frequency changes are small. This particular issue has been investigated using analytical theory  and simuResults from GWAS have allowed the development of new statistical tests to find evidence of directional selection by incorporating information from trait-associated alleles and their effect sizes. Broadly, these tests use information from coordinated allele frequency shifts that are common when there is a large background mutation rate. One of the pioneer studies to incorporate GWAS information to detect directional selection found that height increasing trait-associated alleles have a significantly higher frequency in Northern Europeans compared with Southern Europeans . FurtherDespite the statistical soundness of tests to detect directional selection events, there have been major concerns regarding the validity of the current approaches available to detect signatures of directional natural selection due to population stratification, the transferability of effect sizes between populations, and ascertainment biases on the genotyping chip . AdditioA clear understanding of the factors that lead to phenotypic differences requires a careful assessment of how sociocultural, biogeographical, and genetic factors drive those changes. As an example, we can think of the impact that different variables associated with these factors have on human height. SES has a direct impact on net nutrition, the most important nongenetic factor affecting height . ExposurGWAS of height have been performed on cohorts of individuals from single countries such as United Kingdom , Japan , Peru A, and SarDirectional selection on complex traits has been hypothesized to be an important factor driving phenotypic differences in height among and between populations. There have been suggestions that the height differences between Northern and Southern European populations are driven by polygenic adaptation. This problem was first approached using GWAS summary statistics from the GIANT consortium, a meta-analysis containing samples from European populations , where tA look to the past can help contextualize the research trajectory so far, and provide a guide for research in the future. Populations as used in evolutionary genetics today can be directly linked to a racialized worldview that was inherited by the field of genetics from earlier academic and public discourse alike . During History shows us that science and scientists can be operating under extrascientific forces, and as such, the process of science has a fundamentally subjective aspect to it. This is a lesson for us today. As we familiarize ourselves with this history, we can understand the role it played in motivating and being generated by the development of concepts and ideas in genetics, statistics, and related disciplines. This will encourage us to revisit assumptions or ambiguities regarding the existence of populations , and theAlthough biology plays a role in shaping complex trait variation, it should not be assumed that it follows some natural hierarchy across all traits along color lines and can, therefore, be used to justify discrimination and racism. Indeed, methods that assume population-specific genetic effects and biology show a marginal improvement to predict complex traits and diseases across human diversity over methods that assume largely shared genetic effects. These results argue in favor of largely shared biological functions among individuals living in different geographies, with room for population-specific effects and interactions with the environment. As such, complex trait studies that include individuals living in different regions and environments will be crucial to get an accurate picture of the factors shaping complex trait diversity across the globe, helping identify shared and private aspects of genetic architecture and interactions with the environment. Such diverse sampling and methodological and conceptual advances will take us further in understanding genetics as mediated by the biogeographical and sociocultural environment to generate the mosaic picture of complex trait variation we see today.


We found that most ecotype studies involve fish, mammals and plants with a focus on habitat use, which at 60% was the most common criterion used for categorization of ecotypes. Only 53% of the studies incorporated genetic analyses, and major discrepancies in available genomic resources among taxa could have contributed to confusion about the role of genetic structure in delineating ecotypes. Our results show that the rapid advances in genetic methods, also for nonmodel organisms, can help clarify the spatiotemporal distribution of adaptive and neutral genetic variation and their relevance to ecotype designations. Genetic analyses can offer empirical support for the ecotype concept and provide a timely measure of evolutionary potential, especially in changing environmental conditions. Genetic variation that is often difficult to detect, including polygenic traits influenced by small contributions from several genes, can be vital for adaptation to rapidly changing environments. Emerging ecotypes may signal speciation in progress, and findings from genome‐enabled organisms can help clarify important selective factors driving ecotype development and persistence, and thereby improve preservation of interspecific genetic diversity. Incorporation of genetic analyses in ecotype studies will help connect evolutionary biology and applied conservation, including that of problematic groups such as natural hybrid organisms and urban or anthropogenic ecotypes.The concept of ecotypes is complex, partly because of its interdisciplinary nature, but the idea is intrinsically valuable for evolutionary biology and applied conservation. The complex nature of ecotypes has spurred some confusion and inconsistencies in the literature, thereby limiting broader theoretical development and practical application. We provide suggestions for how incorporating genetic analyses can ease confusion and help define ecotypes. We approach this by systematically reviewing 112 publications across taxa that simultaneously mention the terms  Specifically, we (1) review publications from a Web of Science search to determine the present characteristics used to define ecotypes in the context of conservation and management, and 2) discuss factors that could limit or confound integration of the ecotype concept with analyses in landscape, evolutionary and conservation genomics. Finally, we (3) offer recommendations for investigating and integrating genetic structure relevant to ecotypes, including problematic groups such as natural hybrid organisms and putative anthropogenic or urban ecotypes, where parallel selective pressures such as tolerance to higher temperatures may promote important adaptations to global climate change. By expanding the focus from specific genes or genome regions towards environmental processes and variables  populations in the outer regions of the Pacific Coast of North America have a closer ecological relationship with salmon and other marine resources than their conspecifics farther inland, although populations occur well within dispersal distance  and black bears (U. americanus). The latter comprises the local white form known as the Kermode or Spirit bear, a recessive homozygote morph where a recent study found the relevant allele occurring in a broader area, but with a lower landscape‐level frequency (up to 26%), than had previously been estimated  and a Kermode or Spirit bear capturing salmon (photo: Paul C. Paquet).2https://www.webofscience.com/wos/woscc/basic‐search) that included records published until 23 January 2020 with the terms ‘ecotype AND conservation AND management’ in the topic field, which searches title, abstract and keywords. Although our criteria thereby narrowed the focus of the literature search, it facilitated our central aim of evaluating how consistently the ecotype concept has been used in applied research directly relevant to conservation and management. Moreover, our search was done without bias towards specific taxonomic groups or organisms with more available genomic resources. We obtained n = 118 records. Further reading revealed six records where ‘ecotype’ referred to features or entities not directly applicable for genetic analyses (e.g. habitat types such as sand dunes). We excluded these records but retained three simulation/review studies, resulting in n = 112 publications . We divided taxa into seven groups , and only two of these listed ecotypes linked to genetic differentiation, whereas seven reported habitat use. Caribou are a well‐known species of conservation concern for which there have been different and sometimes confounding results on the relationship between genetic structure and ecotypes , and fish ecotypes most frequently being defined by genetic differentiation, whereas mammals were more often categorized by habitat use. The ecotype concept may thus have appeared more relevant for certain taxa in the context of conservation and management. In contrast, we found substantially fewer records for invertebrates, birds and reptiles, and none for amphibians. The variability might indicate that less ecotype‐related research has been done within certain fields, or that one or more of the search terms were not included, and the use of alternate names for conservation units . These guidelines recognize evolutionary significance as phylogenetic divergence, ecological (and likely adaptive) differentiation, a remnant population representing a natural occurrence, or a population whose loss would cause extensive range disjunction. The United States Endangered Species Act also permits assignment of Distinct Population Segments for vertebrate populations . Although genetic data are typically included, such groupings could thus encompass units that are linked to specific habitat types without considerations of genetic criteria, and this uncertainty complicates the delineation and monitoring of evolutionary and conservation units, including ecotypes. Genetic structure plays an important role in defining management units for wild species, including endangered , where logistical difficulties have limited data collection during winter . In marine and terrestrial environments, some populations, that is, genetic clusters, occur in sympatry yet exhibit distinct spatiotemporal patterns  of the studies we reviewed included analyses of genetic structure in their categorization of ecotypes. Therefore, asking if genetic measures were not considered necessary or informative seems relevant. Alternatively, few or no genetic markers may have been available for the organism of interest. Our temporal analysis of studies that included genetic and genomic analyses suggests that this factor could have played a role, especially for nonmodel species Figure . We use 13 of the4.14.1.1The listing of subspecies or distinct population segments can help focus management on vulnerable areas, allowing protection of vital evolutionary potential while reducing impacts on landowners or other special interests relative to that of listing the entire species across its range  found on the north Pacific Coast of Canada. This morph is linked to a recessive mutation at the melanocortin 1 receptor gene (Mc1r), and for this reason, isolation and limited gene flow may be essential for its continued persistence , for example, has colonized areas where it was never previously recorded and is now successfully reproducing in those ranges  habitats, rapid environmental changes in urban and other anthropogenic environments also appear relevant for ecotype identification and conservation management. Recent reports have indicated differential selective pressures in urban and nonurban environments that influence morphology, physiology and behaviour  with a long history in human settlements  at present, the role of spatial genetic structure in delineating ecotypes ranges from all important to unspecified, and this lack of consistent definition limits broader application, and 2) current discrepancies between genetic structure and ecotype may reflect lack of genomic resources for many taxonomic groups and therefore limited investigation of adaptive vs. neutral differentiation. Although existing conservation units variously include ecological distinctness and the presence of adaptive genetic diversity, the application of an ecotype concept centred on adaptive diversity can offer additional direction. Preservation of genome‐wide variation is critical for evolution and conservation (Kardos et al.,  current The authors declare no conflict of interest.Supplementary MaterialClick here for additional data file.


Sus scrofa) is a widely distributed ungulate whose success can be attributed to a variety of ecological features. The genetic variation and population structure of Lithuania’s wild boar population have not yet been thoroughly studied. The purposes of this study were to investigate the genetic diversity of S. scrofa and assess the effects of habitat fragmentation on the population structure of wild boar in Lithuania. A total of 96 S. scrofa individuals collected from different regions of Lithuania were genotyped using fifteen microsatellite loci.Wild boar (st values and AMOVA, which indicated no differentiation between the four sampling areas. The results of the Mantel test revealed a weak isolation by distance and geographic diversity gradients that persisted between locations. Motorway fencing and heavy traffic were not an effective barrier to wild boar movement.The microsatellite analysis of the wild boars indicated high levels of genetic diversity within the population. Microsatellite markers showed evidence of a single panmictic wild boar population in Lithuania according to STRUCTURE’s highest average likelihood, which was K = 1. This was supported by pairwise FThere was limited evidence of population genetic structure among the wild boar, supporting the presence of a single population across the study area and indicating that there may be no barriers hindering wild boar dispersal across the landscape. The widespread wild boar population in Lithuania, the high level of genetic variation observed within subpopulations, and the low level of variation identified between subpopulations suggest migration and gene flow between locations. The results of this study should provide valuable information in future for understanding and comparing the detailed structure of wild boar population in Lithuania following the outbreak of African swine fever.The online version contains supplementary material available at 10.1186/s12863-021-01008-8. Sus scrofa) is among the most widespread large mammals, with a natural range extending from western Europe and the Mediterranean Basin to the eastern Russian Federation and Japan, and throughout southeast Asia [The wild boar . Hunting statistics reveal that 176,722 wild boars were hunted in the last 5 years (2014–2019). Hunting serves a population-regulation function and could affect the dispersal behaviour and population structure of this species.Central European wild boar subspecies are also abundantly distributed in Lithuania . The sizhttps://lakd.lrv.lt/en/). In Lithuania, Balčiauskas [Habitat fragmentation caused by transport infrastructures could have an influence on the wild boar population. Roads and traffic volumes act as barriers for individuals, can hinder migration, and strongly influence road mortality rates and gene flow between populations , 11. Thečiauskas  recordedčiauskas , 14.Numerous studies have focused on the ecological impact of wild boar, but little research has been performed on their genetic diversity and population structure in Lithuania. Molecular techniques can serve as valuable tools for improving understanding of genetic changes in populations, population structuring and genetic differentiation .S. scrofa and assess the effects of habitat fragmentation on the population structure of wild boar in Lithuania.The main purposes of this study were to investigate the genetic diversity of A) ranged from 2 to 13, with an average over all loci and all sample sites of 6.817  revealed that 11 of the 15 markers were at equilibrium, while the remainder showed a significant departure from HWE in the wild population due to heterozygote deficiency  and Fst analysis indicated low or no genetic differentiation between all pairs of the subpopulations  and analysis of molecular variance revealed no significant genetic differentiation between the wild boar subpopulations (Table is = 0.145 (p < 0.001) and Fit = 0.148 (p < 0.001). These data indicated that a greater genetic variability in S. scrofa was mainly distributed within individuals  revealed that 86% of the variance was found to be between individuals, while 14% came from differences between individuals within population, and 0% was observed among subpopulations Table . Statistns Table . Other Fls Table .Table 32 = 0.1658, P = 0.009). A relatively high regression coefficient and significant association between genetic and geographical distances were obtained among individuals from subpopulations II and III , and a weak but significant  correlation was found among individuals from subpopulations I and III. However, there was no significant association between genetic and geographical distances for individuals in the combined subpopulations III and IV , I and II   mean observed heterozygosity values were detected in this study compared with those for wild boar populations in Bulgaria (Ho = 0.63) [o = 0.63), Hungary (Ho = 0.75) [o = 0.627) [o was slightly higher in the Lithuanian population than in the Polish population (Ho = 0.51) [o = 0.63) has been reported in wild boar populations inhabiting East Asia [o) were lower than the expected heterozygosities (HE) in four subpopulations   , Italy ( = 0.75)  and Port= 0.627) . Furtherast Asia . The dif Ho = 0.5 [o = 0.6Analysis of molecular variance (AMOVA) revealed a high intra-population genetic variation in the wild boar population in Lithuania Table . A similst values were effectively zero and none was significant, suggesting that the grouping of Lithuanian wild boar individuals due to traffic barriers had no tangible effect on population structure. Fst analyses found no evidence of genetic differentiation between subpopulations living on opposite sides of the motorway, but a significant positive correlation between genetic and geographical distance suggested isolation by distance. In contrast, the higher genetic differentiation (Fst = 0.0816) obtained from Bulgarian populations occurs due to geographical barriers such as mountain ridges and human impact [Population pairwise Fn impact . One posn impact .Analysis using the Mantel test did not show significant correlation between different sampling areas, except between western and eastern subpopulations . These results suggested that weak differentiation could occur due to habitat fragmentation by the main motorways: the E67 connecting Vilnius and Klaipėda and the E85 “Via Baltica” motorway connecting Lithuania and neighbouring Poland. The E67 and E85 are the busiest roads in Lithuania (6873 and 9523 vehicles per day respectively). Barriers created due to anthropogenic activity, such as the busiest motorways with high volumes of traffic, fencing and contiguous urban areas, could reduce gene flow and affect the formation of population structure. Conversely, the results of the Mantel test for all subpopulations together demonstrated weak isolation by distance, indicating that geographic distance weakly contributed to the genetic differentiation in the wild boar population.The multi-locus microsatellite data presented here suggested that the most likely explanation for the lack of genetic structure was that wild boar breeding areas in Lithuania comprise a single panmictic unit. Dispersal of successfully reproducing animals among breeding areas exhibited high genetic connectivity between sampling areas. Bayesian analysis in STRUCTURE showed that all wild boar individuals could probably be assigned to one genetic cluster. This was supported by pairwise FST calculations that demonstrated little (ranging from - 0.000 to 0.007) or no significant differentiation between locations. Nikolov et al. 2009  identifiThere was limited evidence of population genetic structure among the wild boar, which therefore supported the presence of a single population across the study area. It indicated that there may be no barriers hindering wild boar dispersal across the landscape. The widespread wild boar population in Lithuania, the high level of genetic variation observed within subpopulations, and the low level of variation identified between subpopulations suggested migration and gene flow between locations. The results of this study should provide valuable information in future for understanding and comparing the detailed structure of wild boar population in Lithuania following the outbreak of African swine fever.S. scrofa individuals legally harvested by licensed hunters in different parts of Lithuania were investigated. A decision was taken to focus on the single population of Lithuania and a sample size that would be sufficient to characterise population-level genetic diversity when using microsatellites.Tissue samples of wild boar were collected over a five-year period 2009–2013) from 35 sampling sites across Lithuania  sampling area.The main habitats favoured by boars vary from semi-arid environments to marshes, forests and alpine grasslands . In LithFresh muscle, spleen and blood were sampled from wild boars and either stored in plastic tubes (5–30 ml) filled with 96% alcohol or kept frozen at a temperature of − 20 °C. All the samples were legally collected and deposited at the State Food and Veterinary Service of the Republic of Lithuania (SFVS). The study did not involve the collection of samples from live animals. An ethics statement was not required. The hunters collected samples in accordance with national regulations on wild boar management.In this research, samples were extracted using the “DNeasy Blood and Tissue Kit”  following the manufacturer’s instructions. The concentration and purity of the isolated DNA were determined using Nanodrop 2000 Spectrophotometer . Samples were used immediately for amplification or stored at − 20 °C for later use.A set of 15 microsatellite markers were selected from the list of microsatellite markers recommended by the International Society of Animal Genetics (ISAG) – Food and Agriculture Organization (FAO) . The marPCR reactions were carried out in a total volume of 25 μL, containing 1 μL of DNA template, fluorescent forward primer (2 μM) and non-fluorescent reverse primer (2 μM), and 2x QuantiTect Multiplex PCR NoROX Master Mix . PCR reactions were carried out in the following steps: 10 min an initial denaturation at 95 °C, 30 or 35 cycles at 95 °C for 30 s depending on the primer set used, annealing at an optimal temperature ranging from 57 to 58 °C, extension at 72 °C for 1 min, then a final extension at 72 °C for 30 min. The ABI 3100  DNA Analyzer was used to genotype alleles with a GeneScanTM-500 ROX size standard (Applied Biosystems). Gene Mapper 3.7 (Applied Biosystems) software was used to estimate the size of the alleles.A), observed heterozygosity (Ho) and expected heterozygosity (HE) under Hardy-Weinberg assumptions were obtained in GenAlEx v6.1 [P values for HWE were corrected for multiple comparisons by applying a sequential Bonferroni correction, with an initial probability of p = 0.05 [st, Fis, and Fit). FST values were estimated according to Weir and Cockerham’s [In order to estimate the population genetic structure of wild boars in Lithuania, the number of alleles per locus  to estimate the optimal value of K using the Delta (K) method [http://clumpak.tau.ac.il [The determination of the most probable number of clusters for Lithuania’s wild boar population  was assessed by the STRUCTURE program version 2.3.4 . The pro) method  and allo) method . Based oau.ac.il ).Additional file 1: Table S1. Genetic diversity of wild boar in Lithuania estimated based on polymorphisms in 15 microsatellite loci.Additional file 2: Figure S1. Mantel tests of the relationships among genetic differentiation  and geographical distance (km) in different sampling areas. A Mantel test between Fst values and the geographical distance in I and II sampling areas. B Mantel test between Fst values and the geographical distance in I and III sampling areas. C Mantel test between Fst values and the geographical distance in II and III sampling areas. D Mantel test between Fst values and the geographical distance in III and IV sampling areas.


The word “bamboo” reminds us of “a hollow stick,” but it is filled with a plethora of health benefits. The tribals of northeastern India ferment these beneficial bamboo shoots for the goodness of mankind. Fermentation is an important age-old biotechnological procedure used for the preservation of food products. Fermented bamboo shoots form the niche for many microorganisms, and this confers positive effects and advantages in many ways. These magical shoots have tremendous health benefits like anti-cancer, anti-oxidant, anti-aging, cardioprotective, weight loss, probiotics, to name a few. Apart from health benefits, fermented bamboo shoots form important functional foods and have industrial and economical values. Though these are commonly found and started in the tribal area, and local markets, today, they are valuable all around the world, as popular as gold. Hence, fermented bamboo shoots are referred as “green gold” of India. This review briefs about various health benefits, advantages, disadvantages, future scope, and finally the economic values of fermented bamboo shoots, the “green gold” of the twenty-first century. Aspergillus spp., and Saccharomyces spp., respectively. Globally, fermented foods and beverages ranges from 20 to 40% of the food supply. The fermented foods such as bread, wine, cheese, yogurt, idli, and dosa [Fermentation may be defined as a method of generation of a product, by culture of mass of microorganisms . People The table shows the fermented foods from different parts of the world Table .Table 1F. These fermented bamboo shoot food products, along with their nutritional value, mode of preparation, and the presence of microorganisms have been described here. This review is to account for the benefits of fermented bamboo shoot-based foods of northeast India. Bamboo shoots are fermented in northeast India by the tribal people, due to their plethora of health benefits with high medicinal value and immense food value. They are a storehouse of many important microorganisms especially the lactic acid bacteria (LAB) and yeast strains. They give the color, aroma, flavor, taste, and texture to the food product. They can be used as functional foods as they are rich in probiotics. Many fermented bamboo shoot-based food products are produced by the local tribes of northeastern India, of which few are readily available in the local market, like soidon, soibum, mesu, ekung, lung-siej, hirring, and eupAs this review lists the various advantages of fermented bamboo shoots, especially the health benefits, it is very much required to prove why fermented bamboo shoots are important and thus, how they increase the economic value and that is why it is called the “green gold” in India in this twenty-first century. Bamboo being green in color and is as valuable as gold due to the plethora of benefits, it bestows on mankind.Lactobacillus sp., Bacillus sp., Candida sp., and Saccharomyces cerevisiae are present especially in Kinema, Hawaijar, Tungrumbai, Bekang, Peruyyan, Soibum, Soidon, Mesu, Soijim, Ekung, Hirring, Ngari, Hentak, Tungtap, Gnuchi, Gundruk, Sinki, Ziang-sang, Goyang, Khalpi, Ipoh, Atingba, Kodo ko Jaanr, and Zutho [Northeast India consists of eight states, namely, Arunachal Pradesh, Assam, Manipur, Meghalaya, Mizoram, Nagaland, Sikkim, and Tripura. It consists of a variety of people having diverse ethnic backgrounds. Most people are tribal about 75% of the population. These tribes have great knowledge about plants, forests, and edible food products from plants. They use fermentation techniques for improving food processing, preservation, and taste enhancement. In the fermented foods, microbes such as nd Zutho .Fermentation is used to preserve food products in Manipur . There aBamboo shoots are edible. Fermented bamboo shoots are important to human life due to their high nutritional and medicinal value. The bamboo shoots also form an integral part of the tribal diet. They are high in dietary fiber and mineral content, low in fat, and are economically important. Traditional medicine systems like Ayurveda uses bamboo shoots as medicines for many diseases .They are rich in xylan or xylooligosaccharides. Some of the medicinal applications and health benefits are as follows: anti-oxidant, anti-cancer, anti-aging, anti-free radical, weight loss, prevents cardiovascular diseases, improves digestion, anti-microbial activity due to the presence of different glycosides, flavones, and also decreases blood pressure .Lactobacillus sp. [Fermented bamboo shoot products like soidon, soibum, soijim, lung-siej, and bamboo shoot pickle samples undergo natural fermentation by llus sp. .. These bacteria add flavor, aroma, and sour taste to the fermented foods. Soidon mainly has lactobacillus sp. such as L. curvatus and Lactococcus lactis.These foods act as a storehouse of a group of gram-positive bacteria namely, lactobacillus (LAB) species . These bLeuconostoc fallax, L. mesenteroides, Lactobacillus brevis, L. curvatus, and Lactococcus lactis [Lung-siej mainly has s lactis .The list of fermented bamboo shoot-based products and the associated microorganisms is shown in Table Some examples of northeast Indian fermented bamboo shoots have been discussed below:Meiteis, the inhabitants of Manipur [Bambusa tulda (Utang), B.balcona (Ching saniebi), Wanap, Unap, Dendrocalamus hamiltonii, Pecha, Melacona bambusoide (Moubi/Muli), Bambusa tulda (Utang), and B.balcona (Ching saniebi), by natural fermentation. The outer casings of young shoots are removed, and the inner part is cut into pieces and washed and fermented for 3–12 months. The Meitei women make this in Manipur. It is eaten with steam rice as a normal dish by the Meitei tribe. Soibum is commonly sold as a vegetable in the market by the Meitei women in Manipur. It is rich in microorganisms such as Lactobacillus plantatum, L.brevis, L. coryniformis, L. delbrueckii Leuconostic fallax, L. mesenteroides Lactococcus lactis, Streptococcus lactis, Enterococcus durans, Bacillus subtilis, and yeasts like Candida, and Saccharomyces, to name a few [Manipur is a center for different types of fermented food products, especially prepared by  Manipur . Soibum me a few , 13.The nutritional value of soibum includes, moisture: 92%, acidity: 0.98%, pH: 3.9, fat: 3.2% DM, carbohydrate: 47.2% DM, protein: 36.3% DM, food value: 362.8 kcal/100 g DM, Ca: 16.0 mg/100 g, K: 212.1 mg/100 g, and Na: 2.9 mg/100 g .The preparation of soibum has been represented as a flow chart in Fig. Lactobacillus plantarum, confers antimicrobial property and increases the shelf- life of soibum [As shown in Fig. f soibum .karati bans (Bambusa tulda), choya bans , and bhalu bans  are used. The shoots are defoliated and chopped and pressed tightly, into a hollow bamboo stem. The tip of the vessel is covered tightly with leaves and allowed to ferment naturally under anaerobic conditions for 7–15 days. Mesu is consumed as a pickle. Usually, microorganisms found in Mesu are Lactobacillus brevis, L. plantarum, L. curvatus Leuconostoc citreum, and Pediococcus pentosaceus [Limboo women of Nepal. Mesu in green bamboo vessel is sold in rainy season in the local markets of Sikkim and Darjeeling hills by the limbo women [Mesu is a common fermented bamboo shoot that is made and eaten by Gorkha tribe of Sikkim. To prepare this, edible bamboo shoots tosaceus . Mesu habo women .The nutritional value of mesu includes moisture: 89.9%, acidity: 0.88%, pH: 3.9, ash: 15.0% DM, protein: 27.0% DM, fat:2.6% DM, carbohydrate: 55.6% DM, Ca: 7.9 mg/100 g, K: 282.6 mg/100 g, Na: 2.8 mg/100 g, food value: 352.4 kcal/100 g DM .The method of preparation of mesu has been illustrated in Fig. Mechanism of action of different microorganisms occurs at the stage where the chopped bamboo shoots are kept in air-tight bamboo vessels and for fermentation for 7–12 days, gives the texture and color to the mesu food product  is used. In other words, the apical meristem of Teinostachya wightii (Nath) - bamboo shoots are taken. The lower portions and the outer casings are removed. In an earthen pot of water, the whole tips are submerged. The soijim or the sour liquid of the previous batch is added as a starter in 1:1 dilution and fermented for 3-7 days at room temperature. To increase the flavor of soidon, leaves of Garcinia pedunculata Roxb., known as heibung, locally, may be added to the fermenting vessel. Soidon can be kept for a year in a closed container, at room temperature. It is used as a curry or as a pickle [Soidon denotes fermented bamboo shoot tips sold by Meitei women and forms a diet of inhabitants of Manipur. It is prepared in the following ways: the tip of the matured bamboo shoot  are collected and the leaves are removed, and the bamboo shoots are cut into small pieces. In the forest, near a water body, a pit is dug, and the bamboo shoot pieces are washed. In the pit, the chopped bamboo pieces are laid in a bamboo basket and are covered with leaves and sealed. Heavy stones are kept for draining the water down and ferment the bamboo shoots for 1–3 months. This can be kept for a year in an air-tight container. Ekung can be cooked with meat, vegetables, or fish and is also sold in the local markets [Ekung is a fermented bamboo shoot food product, ethnic to Arunachal Pradesh, produced by Nishi. It is called by different names by different dialects like  markets .Lactobacillus plantarum, L.casei, L. brevis, and Tetragenococcus halophilus.The microorganisms which are mainly present in ekung are The nutritional value of ekung consists of moisture: 94.7%, acidity: 0.94%, pH: 3.9, ash: 14.0% DM, food value: 363.0 kcal/100 g DM, protein: 30.1% DM, carbohydrate: 52.1% DM, fat: 3.8% DM, Ca: 35.4 mg/100 g, K: 168.6 mg/100 g, and Na: 10.9 mg/100 g .The method of preparation of ekung has been illustrated in Fig. Lactobacillus plantarum in ekung, shows mechanism of action by conferring antifungal and antimicrobial properties, and also increases storage life [age life .ipe, and Apatani call them hi. In the case of eup, bamboo shoots are cut into small pieces and fermented-like ekung, in 1–3 months. Eup is a dry food product, and bamboo shoots are chopped into pieces and dried in sun for 5–10 days till the color changes from white to chocolate brown. Eup is consumed as a curry with vegetables, fish, or meat and can be stored up to 2 years [Eup is a dry fermented bamboo shoot of Arunachal Pradesh. The word is derived from the Nishi dialect. Eup has synonyms like Khampti call them nogom; Adi call them  2 years .Lactobacillus fermentum and L. plantarum.The microorganisms which are mainly found in eup are The nutritional value of eup consists of moisture: 36.8%, acidity: 0.80%, pH:4.1, ash: 18.2% DM, fat: 3.1% DM, protein: 33.6% DM, carbohydrate: 45.1% DM, food value: 342.7 kcal/100 g DM, Na: 3.4 mg/100 g, Ca: 76.9 mg/100 g, and K: 181.5 mg/100 g .The preparation of eup has been illustrated in Fig. Apatani tribe. The Nishi call them hitak or hitch. In the production of hirring, bamboo shoots are cut longitudinally into 2–3 pieces or the shoots are flattened by crushing and are put into bamboo baskets with leaves. The baskets are placed into a pit, covered with leaves and sealed, and fermented for 1–3 months. Hirring can be consumed as a curry and is generally sold in local markets [Hirring is a fermented bamboo shoot product in Arunachal Pradesh made by the  markets .Lactobacillus plantarum and Lactococcus lactis [The microorganisms commonly present in hirring are s lactis .The nutritional value of hirring includes moisture: 88.8%, pH: 4.0, acidity: 0.81%, protein: 33.0% DM, carbohydrate: 49.3% DM, fat: 2.7% DM, food value: 353.5 kcal/100 g DM, Ca: 19.3 mg/100 g, K: 272.4 mg/100 g, and Na: 3.4 mg/100 g .The method of preparation of hirring is illustrated in Fig. Bamboo shoots are fermented and consumed by tribal people of northeast India and form an integral part of their basic diet. They are rich in nutrition. They consist of a plethora of health benefits such as anti-free radicals, help in reducing cholesterol, anti-cancer, anti-oxidant, act as immune-booster, anti-aging, prevent cardiovascular diseases by protecting the heart, improve digestion, weight loss, decrease blood pressure, rich in flavones, glycosides, and it is anti-microbial, and rich in  probiotics . AccordiNot only in India but also in the Philippines, Korea, and Tibet, bamboo shoots find its applications Table .Table 3ABambusa blumeana (Kawayangtinik), the anti-fatigue property was reported [Bambusaarundinacea species is considered as a beneficial tonic for respiratory disorders [In the Philippines, reported . Even inisorders .Apart from food and medicine, bamboo shoots are also used in the cosmetics industry. Korea has released bamboo sea salt, which is used as a cleansing agent called bamboo bath salts .Bamboo shoots have industrial importance too. They are used for the production of bioethanol. They are also used as a source of natural products like potassium, dietary fibers, carbohydrates, and vitamins. Bamboo shoots may be used to produce functional xylooligosaccharides which find application in food, biodegradable plastics or nanoparticles, and pharmaceutical industries .Lactobacillus plantarum most frequently found in most fermented bamboo shoots, have potential probiotic effects along with cholesterol-lowering feature [L. brevis, they exhibit high hydrophobicity which indicates the ability of bacterial culture to adhere to the epithelial cell layer of the digestive tract for good colonization [Fermented bamboo shoots like soidon, lung-siej, ekung, soibum, and mesu are rich in microorganisms which have probiotic property. Microbes like LAB, species like  feature . With L.nization , 27.The bamboo-based fermented foods require a large water body for its preparation and fermentation process. A long time is required for fermentation. Large volumes of bamboo shoots are chopped and cut in the forest area. This leads to deforestation.Sometimes, the consumption of fermented bamboo shoots may lead to toxicity. The presence of cyanogenic glycosides known as taxiphyllin in bamboo shoots may lead to cyanide poisoning. This has been reported due to inhalation of hydrogen cyanide gas (HCN), which was produced from pickled bamboo shoots .But, if we compare the benefits of fermented bamboo shoots with the disadvantages, it outweighs the latter and is a rich source of food value, functional food, and medicinal value.Fermented bamboo shoots have many advantages that can be used in the future. Not only do they have innumerable health benefits, but are applied in many other areas also, which are quite lucrative in the future. Consequently, the future scope of fermented bamboo shoots is very high. It can be done in the following areas:Fermented bamboo shoots can be used in different industrial applications. It can be used in food, pharmaceuticals, biofuels, to name a few. In the food industry, it has high growth potential. It can be used as functional and healthy foods, as medicines, and as a source of bioactive compounds. It is a niche for various lactic acid bacteria which act as probiotics. Though the preparation of fermented bamboo shoots is local and prepared by tribal people of Northeast India, they have great scope and value among the food source of plant origin in the Asian countries. It may be just “supporting sticks” for poor people, and also a tasty cuisine. In the international market, fermented bamboo shoots form an important aspect in terms of nutrition, health, and medicinal property due to probiotic microflora .In the biofuel industry, it can be used to produce ethanol or methane. It has been used to produce bioethanol, which is a source of carbohydrates, potassium, vitamins, and dietary fibers. Bio-methane can be produced from bio-ethanol which has high holocellulose content and biomass yield .Meiteis. However, due to numerous food and health benefits, the medicinal values, it can be sold out in the international market also, to other countries [Fermented bamboo shoots have high economic importance . Usuallyountries –26. HencIn our opinion, fermented bamboo shoots are very healthy and have high future scope in the fields of food, pharmaceutical, and other industries, due to their plethora of health benefits, medicinal properties, and probiotic nature. It should be publicized in the national and international markets, and not be hidden only in the local tribal markets. More care should be taken in the area of preservation of fermented bamboo shoots for a longer time so that the shelf- life can be increased.Fermented bamboo shoots are a storehouse of microorganisms, many of them being probiotics in nature. When consumed, they produce a lot of health benefits, such as anti-oxidant, anti-cancer, reduces blood pressure, prevents cardiovascular diseases, weight loss, to name a few. Besides these, they can also be applied in the industries, especially in the food, pharmaceutical, and biofuel industries. They have economic value and the benefits are far more than the disadvantages, that is why, they are called the “green gold,” of India.


Salvelinus confluentus) extent and abundance are in decline due to historic overharvest, invasive non‐native species, and habitat loss. In Alberta's Eastern Slope region, populations at the range margin have progressively been lost, motivating us to better understand the amount and distribution of genetic variation in headwater habitats and some downstream sites where they continue to persist. Across this region, we sampled 431 Bull Trout from 20 sites in the Athabasca and Saskatchewan River basins and assayed 10 microsatellite loci to characterize within‐ and among‐population genetic variation. The Saskatchewan and Athabasca River basins contained similar levels of heterozygosity but were differentiated from one another. Within the Athabasca River basin, five genetically differentiated clusters were found. Despite the evidence for genetic differentiation, we did not observe significant isolation‐by‐distance patterns among these sites. Our findings of ample genetic diversity and no evidence for hybridization with non‐native Brook Trout in headwater habitats provide motivation to ameliorate downstream habitats and remove anthropogenic barriers to connectivity towards the goal of long‐term persistence of this species.Freshwater ecosystems are negatively impacted by a variety of anthropogenic stressors, with concomitant elevated rates of population decline for freshwater aquatic vertebrates. Because reductions in population size and extent can negatively impact genetic diversity and gene flow, which are vital for sustained local adaptation, it is important to measure these characteristics in threatened species that may yet be rescued from extinction. Across its native range, Bull Trout ( Principal component analysis for individuals sampled in the Athabasca River basin, with 95% confidence ellipses for the five population groups identified by STRUCTURE analyses. For this dataset, differentiation among groups was best explained when using these five distinct clusters that were detected compared to either the tributary or larger regional area that the sampling site was located in. Estimates suggest that they support >10% of all described species, including ~30% of all vertebrate species and 40% of all fish species  is an apex predator endemic to northwestern North America. Across this vast range, spanning different watersheds, ecoregions, and management areas, the species is declining in population size and spatial extent due to overharvest, hybridization with invasive non‐native species, reduced habitat connectivity, and habitat degradation  is part of Canada's strategy for protecting and managing threatened species. One important component of SARA's framework are Assessment and Status Reports, which are generated by the independent Committee on the Status of Endangered Wildlife in Canada (COSEWIC). Where there are a priori expectations for requiring management “below” the species level, genetic information can inform the establishment of Designatable Units (DUs), which represent evolutionary significant within‐species diversity COSEWIC, . These uHere, we assess the population genetic structure of Bull Trout, a globally recognized species as risk COSEWIC, , in the 22.1We sampled 14 sites to characterize the genetic diversity of Bull Trout populations within the Athabasca River Basin (Western Arctic), with six sites from the North Saskatchewan and Bow River Basins (Saskatchewan‐Nelson) sampled for comparison , were multiplexed in four optimized groups Table . These lPCR products were run on a 1.5% w/v agarose gel using a standard gel electrophoresis protocol and visualized using a DigiDOC IT electrophoresis gel imager (UVP Inc.). Samples showing clearly defined bands representing the DNA fragment PCR products of the specified microsatellite loci were then analyzed. Fragment lengths were determined using a standard protocol for microsatellite fragment analysis with an ABI 3500XL Capillary Electrophoresis Genetic Analyzer  and scaled against GeneScan‐500LIZ size standard . For each sample, electropherograms were produced, which were scored using GENEMAPPER v.4.1 .Raw genotype scores were assessed by MICROCHECKER v.2.2.3 . Subsequent STRUCTURE analyses were performed on Bull Trout captured within the Athabasca River basin to determine sub‐basin structure using 10 independent runs at K‐values of 1–15. For all K‐values in STRUCTURE, 10 independent replicates were run. Initial STRUCTURE runs were conducted at 100,000 burn‐in lengths and 100,000 Markov chain Monte Carlo (MCMC) iterations  was deemed the most likely correct K value. The hierarchical partitioning of genetic variation among river basins and drainages using Analysis of Molecular Variance (AMOVA) in GenAlEx version 6.5  and inbreeding , heterozygote excess and deficiency, and linkage disequilibrium (LD) between pairs of loci were determined using Markov‐chain methods and the following parameters in Genepop version 4.2 Rousset, : MCP – 1on Rice, .FST) and geographic distance (km) with 1,000 randomizations in each scenario  to 51 (Sco109). Genotypic frequencies differed from HWE in a number of locations and loci. Heterozygote excess was rare and found only in Little Elbow River (Sco102) and Storm Creek (Omm1128). Evidence for heterozygote deficiency was found in the populations from Colt Creek , Elk Creek (Sfo18), Kerkeslin Lake , Miette Lake (Sfo18), Moon Creek , and Storm Creek (Sco109). One pair of loci  was observed at one of 450 total locus pairs across sites to be in linkage disequilibrium after Bonferroni correction, with no linkage trends between any two loci across all sites and loci. Infrequent evidence for null alleles was seen throughout the dataset.The two loci used to differentiate Bull Trout from Brook Trout and their hybrids, Sco216 and Sfo18, all scored alleles outside the range indicative of Brook Trout DNA, 172–192 bp and 145–165 bp, respectively . In the Athabasca River basin, expected heterozygosity values ranged from 0.473 (Gregg River) to 0.736 (Miette Lake). Allelic richness values averaged 5.52 and ranged from 2.8 (Unnamed Lake) to 8.6 (Athabasca River). Inbreeding coefficients (FIS) ranged from −0.122 (Ranger Creek) to 0.152 (Moon Creek). Of the 41 private alleles detected in this study, 20 of them were found in the Athabasca River basin, with values ranging from zero (Little Berland River and Ranger Creek) to four (Kerkeslin Creek and Unnamed Lake). Sites within the Saskatchewan River basin had similar expected heterozygosity, ranging from 0.468 (Elbow River) to 0.690 (Colt Creek). Allelic richness averaged 6.5 and ranged from 3.7 (Elbow River) to 8.1 (Colt Creek). The Saskatchewan River basin contained the remaining 21 private alleles detected, with values ranging from zero  to nine (Little Elbow River). All calculated expected heterozygosity, private alleles, allelic richness and FIS are presented in Table Genetic diversity was not significantly different between the Athabasca River basin and Saskatchewan River basin  to 0.49 (between McLeod River and Kerkeslin Lake). The mean FST in the Saskatchewan River basin was similar (0.24) to the Athabasca river basin samples, with FST values ranging from 0.15 (between Storm Creek and Little Elbow River) to 0.33 (between Elbow River and Pinto Lake)  primarily reflecting differentiation of Miette individuals from all others but for a cluster of Kerkeslin‐Osprey individuals, which were themselves differentiated from nearly all others, including the majority of Kerkeslin‐Osprey individuals Figure . On this3.4FST values were variable at all geographic distances. Mantel tests performed on Athabasca River basin Bull Trout revealed no significant isolation‐by‐distance pattern between genetic distance and geographic distance using either waterway distance  or linear distance .In the Athabasca River basin, pairwise 44.1FST, FIS, and private alleles) and genetic diversity . The majority of genetic variation was explained within each population with less variation explained among populations, and even less variation explained by the separate river basins.Population genetics is an important conservation tool for understanding the genetic health of vulnerable populations , STRUCTURE results showed that the two river basins are differentiated from one another, exemplified by populations being clustered based on grouping fish to their origin in the Western Arctic or Saskatchewan‐Nelson DU. In both river basins, high heterozygosity values and low FIS were observed, alluding to large genetic differences between populations, which is common for Bull Trout systems  exhibits moderate isolation‐by‐distance patterns, which is thought to be due to a combination of their large geographic ranges and population sizes , also a Salmonid, tends to exhibit weak isolation‐by‐distance trends, although this has been attributed to large population sizes and high levels of gene flow, which prevent differentiation  values and AMOVA results indicate that genetic variance largely resides within the population level. Thus, although there is differentiation between groups, there currently appears to be sufficient genetic diversity that can be drawn upon, acting as evolutionary potential to allow adaptation to changing habitat conditions and long‐term persistence.Because the range of Bull Trout is declining, this elevates the importance of conserving genetic diversity in remaining extant populations as a means to withstand stochastic events ; Data curation ; Formal analysis ; Funding acquisition (supporting); Investigation (lead); Methodology (lead); Project administration ; Writing‐original draft (lead); Writing‐review & editing . Steven M. Vamosi: Conceptualization ; Data curation ; Formal analysis ; Funding acquisition (lead); Investigation (supporting); Methodology (supporting); Project administration (supporting); Writing‐original draft (supporting); Writing‐review & editing .Figure S1aClick here for additional data file.Figure S1bClick here for additional data file.Figure S2aClick here for additional data file.Figure S2bClick here for additional data file.Figure S3Click here for additional data file.Figure S4Click here for additional data file.Figure S5Click here for additional data file.Figure S1‐5Click here for additional data file.Table S1Click here for additional data file.Table S2Click here for additional data file.


Solanum lycopersicum L.) is considered one of the most valuable and versatile vegetable crops globally and also serves as a significant model species for fruit developmental biology. Despite its significance, a severe genetic bottleneck and intense selection of genotypes with specific qualitative traits have resulted in the prevalence of a restricted number of (geno)types, also causing a lack of diversity across widespread cultivated types. As a result, the re-emergence of landraces as well as traditional and heirloom varieties is largely acknowledged as a countermeasure to restore phenotypic, phytochemical and genetic diversity while enriching the aroma/taste tomato palette. On those grounds, the Cypriot tomato germplasm was assessed and characterized. Ten landrace accessions were evaluated under greenhouse conditions and data were collected for 24 IPGRI discrete phenotypic traits. Grouping of accessions largely reflected the fruit shape and size; four different fruit types were recorded across accessions . Moreover, a single run panel consisting of ten SSRs was developed and applied in order to genetically characterize 190 Cypriot genotypes and foreign heirloom varieties. Based on genetic indexes it was established that tomato landraces have a rather low level of heterogeneity and genetic variation. Finally, mineral and phytochemical analyses were conducted in order to estimate biochemical attributes  across genotypes; thus, ascertaining that the Cypriot panel has a high nutritional value. Due to the thermo-drought adaptation and tolerance of these genotypes, the current study serves as a roadmap for future breeding efforts in order to incorporate desirable traits or develop novel tomato lines combining resilience and alimentary value.Tomato ( Solanum lycopersicum L.) is one of the most highly appraised and versatile vegetable crops globally and is widely cultivated for fresh-fruit consumption, grown as an industrial/cash crop or even for processed  products [Tomato  are adapted to an assortment of edaphoclimatic environments spanning from the sea level up to the highlands of the Andean sierra, and thrive in arid, brackish water or rainy conditions . DespiteFor that matter, early introduced genotypes (up to the nineteenth century) were mostly open-pollinated and on-farm propagated at small local scales. As a result, novel varieties emerged via spontaneous mutations and/or recombination of pre-existing genetic diversity . The bulS. lycopersicum gene pool [Nowadays, the fruit weight across modern tomato varieties range from about 10 g in acorn and cherry type tomatoes to more than 450 g in some beefsteak fresh tomatoes . To thisene pool ,17,18. ACurrently, a significant turn towards qualitative nutriment attributes is occurring. Moreover, food science and technology have been shifting away from merely the notion of adequate calorie intake, towards an optimal and efficient nutrition scheme. Thus, currently, the breeding aim is to promote the consumption of bio-functional foods and contribute to improved physical health, therefore preventing the risk of diseases . TomatoeTomato landraces epitomize the most basic form of existing cultivars and are largely regarded as intermediates of the breeding process . OpposedWhen aiming to harness the phenotypic and phytochemical variation of cultivated tomato, and in order to engage this assortment in applied and basic breeding schemes, it is vital to quantify and count such traits in a precise and factual manner. Moreover, phenotypic and biochemical analyses of genetic resources are essential in order to appraise the genetic basis that connects these characters and their overall yields . HoweverIn view of these remarks and due to the shortage of comprehensive information regarding the Cypriot tomato genetic recourses, the objectives of the current study were: (i) to perform a morphological description using scores and descriptors according to worldwide standard norms  descriptors); (ii) to use microsatellite genetic markers and append landraces of other origins to define the genetic structure and intra-relations of Cypriot tomato varieties; (iii) to assess the variation in phytochemicals and other traits of tomato fruits from plants grown under uniform greenhouse conditions and (iv) to investigate the relations between them. The overall goal was to increase the information regarding the local tomato germplasm, which has the potential for exploitation in modern cultivation schemes and direct usage in marketplaces, breeding schemes and for future genomic surveys.The plant material used in the current study consisted of 19 discrete landraces. Ten were Cypriot landraces, namely AR100731, ARI00732, ARI00733, ARI00734, ARI00735, ARI00736, ARI00737, ARI00872, ARI00905 and ARI00906  reservedTuta absoluta was conducted via the repeated application of insecticides  when needed, before anthesis.All Cypriot tomato genotypes were seeded at a nursery (using peat as a substrate) and transplanted at a two-leaves-stage to a greenhouse (March 2018), according to a randomized complete block design, and within row distance of 50 cm. Three discrete blocks were employed and within each plot five plants per accession were studied . For fruit morphological traits, at least ten randomly selected fruits from the second and third truss were analyzed as indicated (IPGRI descriptors for Tomato). Irrigation was provided via drippers at a frequency of two days (for 30 min). Five intervals of fertilization were conducted using an all-purpose water-soluble commercial fertilizer (20-20-20), while pest management against whiteflies and On a single tomato plant basis, 24 discrete agro-morphological traits were scored as detailed in For nucleic acid extraction, tips from young tomato leaves were collected and kept among moist towel paper on ice, till storage at −70 °C. DNA extraction was conducted using the Dneasy Plant Mini Kit  according to the manufacturer’s instructions. The purity and concentration of DNA were estimated by means of nanodrop spectrophotometry. Ten plants per accession were randomly sampled.TM, JOETM ROXTM, or TAMRATM). This permitted a one-tube, single-reaction nested PCR, as previously described [Fifteen simple sequence repeat (SSR) markers were initially screened for the preliminary analysis of the Cypriot tomato germplasm. Based on the efficiency, reproducibility and allele size generated, ten loci  were selescribed . Based o2 in a 12 μL final reaction volume. Conditions for the PCR amplification were: 94 °C , followed by 39 cycles at 94/56/72 °C (60 s), and a final extension at 72 °C for 30 min.For PCR reactions, each panel master mix contained 50 ng of template DNA, 10 pmole of the labeled M13 tailed forward primer, 10 pmole of the reverse and 2.5 pmole of the forward primer, 0.2 mM dNTPs, 0.5 U KAPA Taq DNA Polymerase  and a 2.5 mM final concentration of MgCl2O. Diluted PCR products across panels were all mixed in one tube (per accession). One μL of the mixture was added to 10 μL deionized formamide and 0.2 μL of DNA size standard , before denaturing at 95 °C (5 min). Allele fragments were separated by capillary electrophoresis using an Applied Biosystems 3130® Genetic Analyzer .Amplification products were verified using a standard 2% agarose electrophoresis and diluted at a 1:40 ratio with dd H^2); I = Shannon’s Information Index = −1* Sum (pi * Ln (pi)); Ho = Observed Heterozygosity = No. of Hets/N; He = Expected Heterozygosity = 1 − Sum pi^2; uHe = Unbiased Expected Heterozygosity = (2N/(2N − 1)) * He; F = Fixation Index = (He − Ho)/He = 1 − (Ho/He), where pi is the frequency of the ith allele for the population and sum pi^2 is the sum of the squared population allele frequencies. Moreover, the proportion of polymorphic loci (PIC) and discrimination power (Dp) of each locus was determined.Allele fragments were identified (de-multiplexed) by means of fluorochrome colour and/or bin size, and tandem software was utilized to verify/correct bins . Microsahttps://irscope.shinyapps.io/iMEC/ (accessed on 1 August 2021)).The same dataset was similarly used to test for linkage disequilibrium and Hardy–Weinberg equilibrium (HWE) in the tomato accessions. Genetic relationships between individuals (MLGs) were assessed using the ‘dissimilarity’ distance algorithm and visualized as a minimum spanning network (MSN) and a discriminant analysis of principal components (DAPC). All of the above-mentioned statistics/analyses were performed using the Poppr (V. 2.8.5) package  and the A phylogenetic tree was also constructed using the binary template  using the R package polysat . An apprhttp://clumpak.tau.ac.il/).A Bayesian statistic-employing method for estimating genetic kinship was performed using Structure 2.3.4 . The admIn order to examine the possible correlation of mean morphological traits with the genetic distance of landraces, a mantel test (999 permutations) was computed using the distances matrix of morphological descriptors and the genetic distances matrix calculated via the GENALEX v. 6.501 software .2O, dried with absorbent paper and immediately placed at −80 °C to stop possible metabolic processes. For the phytochemical analyses, fruits were cut in quarters and at least five discrete fruits were bulked in order to create one sample . Bulked samples were blended and juice was lyophilized. In total, three discrete biological repetitions were estimated across phytochemical analyses.Harvesting (approximately 30 to 40 days after anthesis) was performed at the same ripeness level  across all tomato fruits from the second and third trusses. Tomatoes were washed with dH2CO3 were further added, incubated at 40 °C for 30 min, agitated for one min, and absorbance was estimated at a 750 nm wavelength . For the standard curve, serial dilutions of gallic acid (GA) were employed, and results were expressed as mg GA equivalents/100 g FW. All trials were performed in biological triplicates.Total phenols were estimated following the Folin–Ciocalteu reagent methodology revised for a microplate reader, as previously reported . To attaDetermination of the vitamin C content followed the reduction of the 2,6-dichloroindophenol sodium salt (DCIP) method revised for a microplate reader according to Ochoa-Velasco and co-workers . In geneβ-carotene was conducted according to a method previously described [v/v) acetone-hexane mixture. Samples were vigorously vortexed and incubated in dark for 20 min at 4 °C. A five min centrifugation followed and 200 μL of extracts were transferred to a 96-well polypropylene microplate (resistant to organic solvents). Absorbance readings were performed at the following wavelengths: 663 nm, 645 nm, 505 nm and 453 nm, and pigments determination was calculated using the referred equations:Quantification of lycopene and escribed . All pigAll trials were performed in biological triplicates.The juice from three biological replicates was used to estimate the total soluble solids (TSS) via a portable digital refractometer , and results were expressed in Brix. pH was measured with a benchtop pH-meter . Titratable acidity (TA) was assessed employing the potentiometric titration of 0.1 mol/L NaOH up to pH 8.1, using five mL of undiluted juice. Estimations were performed on a DL22 Mettler Toledo titrator  and were expressed as citric acid g in 1 L of tomato juice.Tissues were ground into a fine powder and passed through a 30-mesh screen. Each sample (0.5 g) was dry-ashed in a muffle furnace at 515 °C for 5 h. Then, the ash was digested in 3 mL of 6 N HCl and diluted with double-distilled water up to 50 mL. The concentrations of P, K, Ca, Mg, Mn, Zn and Cu were determined by ICP . Nitrogen was determined by the Kjeldahl method .http://factominer.free.fr/).In order to combine morphological, genetic, mineral and phytochemical analyses, we employed a Generalized Procrustes Analysis (GPA) employing all quantitative traits. The FactoMineR, an R package dedicated to multivariate Exploratory Data Analysis was used under the ‘Commandeur’ algorithm  to p < 2.2 × 10−16 (number of locules). The number of cavities containing seeds was largely correlated to fruit size and ranged from 5.80 ± 1.03 locules for accession ARI00734 to 13.73 ± 2.46 locules for landrace ARI00906 , while the number of locules was also correlated to fruit width  and weight .All quantitative fruit traits had statistically significant differences across landraces and a probability ranging from ARI00906 . AdditioARI00906 . Across ARI00906 . Fruit wA principal component analysis (PCA) was plotted  aiming t−2 across populations. All loci were found to be polymorphic across landraces, still, the percentage of polymorphic loci within the landraces and heirloom varieties was significantly reduced. Since tomatoes are highly autogamous plants and have been present for a short period in the Old World, a restricted amount of genetic and allelic diversity is expected within landraces. Across loci, two to four different alleles were detected having a mean value of 3.2 (The Cypriot tomato germplasm collection (ten accessions) was genotyped alongside nine foreign heirloom tomato cultivars. After preliminary experiments where fifteen primer sets were screened, ten loci were selected in terms of fragment size bins, allele variability and reproducibility of proliferation . The gene of 3.2 .p < 0.5), except loci LEMDDNA, LELEUZIP and EST258529, where a significant (p = 0.05) value of disequilibrium was established. This suggests that sexual propagation has also occurred in the lineage of Cypriot tomato varieties, even though at a reduced rate.Moreover, the observed and expected heterozygosity indexes were also restricted in range and in all cases were below the 0.5 threshold; showing a restricted allelic variation among and within landraces. Cypriot landraces were found to be virtually entirely homogeneous, since all individuals within a landrace (except ARI00735 and ARI00736) presented a single genotype. Nonetheless, across loci and genotypes a moderate amount of genetic diversity was evident based on several diversity indexes ; Table 3st values (p < 0.001 level and ranged from 0 (among Cypriot landraces ARI00733 and ARI00906 that were grouped as one cluster) up to 0.864 (among the ARI00733/ARI00906 group and the Greek heirloom variety ‘Stithos Aphroditis’).The analysis of molecular variance (AMOVA) revealed that 58% of the total variation was attributed to the genetic diversity among varieties, while the remaining 42% was attributed to the genetic variability within landraces and heirloom varieties. Ft values  across gp < 0.045). It was established that within the tomato germplasm, complex genetic relationships could be identified; but all landraces and heirloom varieties were clearly demarcated. Moreover, bootstrap values further confirmed that these varieties have significant discrepancies that reflect a discrete genetic markup. However, Cypriot accessions ARI00733 and ARI00906 (both having the same fruit morphotype) were found identical and thus could signify a remarkably close genetic ancestry. Accessions ARI00735 and ARI00736  were also found highly affiliated but presented a clear cut-off value; hence stand as discrete varieties or clones. Grouping of genotypes did not clearly reflect the country of origin nor did it absolutely reflect morphotypes, signifying that the markers selected are not strongly associated to agro-morphological traits but are rather unrelated.Genetic distances across genotypes were used to construct a dendrogram depicting linear phylogenetic relationships and relevant bootstrap support values . AdditioA minimum spanning network (MSN) was also constructed A to deteA Bayesian-based method was additionally employed in order to survey the distribution of the genetic diversity and the population structure of the tomato germplasm . The optp < 0.01) exist across the tomato genotypes . Interestingly, a mild negative correlation  among the pH and titratable acidity  was estimated, indicating that several organic acids  make up the acidic profile of Cypriot tomato landraces and that the total titratable acidity is a much more complex trait than pH. Moreover, pH discrepancies across genotypes were smaller—in absolute numbers—comparative to TA; an element that also reflects the vast difference in the ANOVA probability value across groups , total soluble solids (Brix), ascorbic acid (vitamin C), total phenolics, lycopene and e levels . StatistSignificant differences in sugar levels were also recorded via the estimation of the refractive index, the total soluble content (TSS) solids concentration and the TSS/TA ratio. On average, approximately 4.11 °Brix were estimated for Cypriot landraces, making the Cypriot germplasm neither sweet nor sour. Specifically, landrace ARI00735 had the lowest TSS content (3.2 °Brix), while ARI00737 was found to have an increased TSS/TA ratio and almost twice the sugar content (5.07 °Brix).Cypriot tomato landraces were also found to have substantial levels of ascorbic acid (AA). The analysis showed that the vitamin C capacity of Cypriot tomatoes ranged from a minimum of 24.37 ± 1.04 mg AA/100 g FW for ARI00906, to a maximum of 48.02 ± 0.94 mg AA/100 g FW for landrace ARI00735. Overall, the mean value of AA was above 30 mg AA/100 g FW, suggesting that Cypriot varieties are a rich source of vitamin C.Additionally, tomato fruits were found to hold significant amounts of phenolic substances at relatively elevated levels as determined using the Folin–Ciocalteu assay (about 7 mg of GAE/100 g FW were estimated on average). Interestingly, the ARI00734 accession (the only pomodoro type in the Cypriot collection) was distinctively disassociated from the remaining cluster of genotypes and presented significantly lower levels of phenolics (4.50 ± 0.16 mg of GAE/100 g FW).β-carotene levels were also evaluated for landraces, and it was proven that significant discrepancies exist across genotypes . Lycopene values ranged from 1.42 ± 0.05 mg/100 g FW (ARI00734) to 5.85 ± 0.04 mg/100 g FW (ARI00905). Surprisingly, landraces ARI00734, ARI00735 and ARI00736 were considered within the same ANOVA group, all presenting low levels of pigments. In the case of β-carotene, even though there were significant deviations among the landraces, nonetheless, differences were less broad (β-carotene concentration (0.92 ± 0.02 mg/100 g FW), while ARI00733 (0.42 ± 0.07 mg/100 g FW) alongside to ARI00906 (0.49 ± 0.11 mg/100 g FW) had the lowest.Lycopene and ss broad . ARI0073It appeared that phytochemical grouping did not necessarily correlate to fruit types or morphological traits. Nonetheless, landraces ARI00735 and ARI00736, as well as landraces ARI00733 and ARI00906 were clustered within the same or adjacent ANOVA groups; hence a metabolic affiliation was proven alongside agro-morphological and genetic affinity. To identify the hierarchical phytochemical proximity among the landraces, values of quality traits were standardized, and the transformed matrix was depicted as a heat map . Two majIn order to combine all types of diverse analyses  into a single analysis, a generalized Procrustes analysis (GPA) was employed . Accessip < 0.05 as analyzed by one-way ANOVA and the Duncan Multiple Range Test. Coefficient of variation (CV) was calculated as the ratio of the standard deviation to mean values.Mean values and relative standard deviations across phytochemical analyses. Different letters (a–g) refer to statistically significant differences at Cyprus, the largest island in the eastern Mediterranean basin is located at the crossroad point of three continents  and historically it has been on the map of millenia-old trade routes. Many discrete civilizations have shaped the Cypriot agricultural tradition throughout the centuries, each contributing to the introduction of non-indigenous species; such as tomato. Moreover, Cyprus has a complex edaphoclimatic background that is divided into four discrete geological zones: (a) the Pentadaktylos (Keryneia) zone, (b) the Troodos Ophiolite, (c) the Mamonia zone and (d) the zone of the autochthonous sedimentary rocks . ComplimNowadays, a gradual detachment of consumers from established tomato cultivars towards locally-grown traditional cultivars is ongoing. This attitude is revitalizing the interest in heirloom and rustic tomato landraces and is leading to the rediscovery of local agricultural traditions . Hence, All landraces presented an intermediate growth trait and had dense foliage as frequently reported across Mediterranean genotypes ,47. As aThe present collection included genotypes producing fruits mostly appropriate for local trading and self-consumption, since tomato seeds were initially collected directly from farmers who grew tomatoes for such purposes, rather than having long shelf life for wider distribution, thus, the predominant fruit shapes and colour hues mostly reflected local preferences. We also found a considerable level of variation for fruit size and weight, while in several cases, the fruits exceeded the 500 g threshold, having an extremely large size . Parisi The majority of accessions had also distinguished greenback (green shoulders). The shape of the pistil scar was also a significant breeding trait that showed variation across genotypes. In general, flattened/heart-shaped tomato fruits presented an irregular shape of pistil scar  with the2) may cause a geographical bias that further restricts landrace diversity and narrows the genetic basis.All morphological traits were converted to eigenvalues and a PCA was constructed . The PCASeveral molecular marker systems have been employed to survey the genetic diversity of tomato collections such as amplified fragment length polymorphisms (AFLPs), randomly amplified polymorphic DNA (RAPD), sequence-characterized amplified regions (SCARs) and single nucleotide polymorphisms (SNPs) . NonetheBoth expected and observed heterozygosity indexes  varied bAll genetic data analyses  indicated that there is neither a geographical structure nor does clustering correspond with fruit type. This is accordance with several studies that describe a genetic grouping uncorrelated to several morphological traits and fruit characteristics ,54. HencOverall, the current germplasm seems to be genetically distinct and thus could serve as a valuable addition for breeding schemes targeting the genetic enhancement of modern cultivars. Moreover, the employment of heirloom varieties and tomato landraces could reverse the loss of genetic diversity, the farming of allochthonous varieties and ameliorate the ongoing genetic erosion. Moreover, since these landraces have evolved under low input agricultural systems in the semi-arid Cypriot environment, these genotypes might serve as valuable germplasm in sustainable farming systems.β-carotene); β-carotene). In that respect, rustic tomato Tuscan varieties were reported to have a three-fold increase in antioxidant phytochemicals, compared to the commercial counterparts [Taking into consideration a typical proteinic, lipidic and sugar content in order to describe the nutritional value , it seems that tomato fruits do not offer a high nutritional value . Still, terparts . All Cypterparts ,59 and Fterparts  reportedterparts . MoreoveBacillus licheniformis) on a commercial cultivar [Cypriot tomato fruits also contained significant amounts of phenolic compounds (averaging 7 mg GAE/100 g FW). It has been reported that phenolics are a highly desirable feature since these moieties encompass the major contribution of antioxidant activities in tomato fruits . Flattencultivar . Nonethecultivar .A critical element of consumers’ demand for tomato fruits is the TSS and acidity characteristics. The pH parameter ranged from 4.55 (ARI00732) to 4.68 (ARI00735); hence Cypriot tomatoes do not present an elevated acidity. Scarano and colleagues analyzed the phytochemical composition of an Italian landrace tomato germplasm collection under an elevated temperature and established a very narrow pH range (4.3 to 4.6) that corresponds to the present study . NonetheThe present study provides the first attempt to characterize and evaluate the traditional Cypriot tomato landraces. For that purpose, a multidisciplinary combined experimental approach was adopted. Twenty-four IPGRI descriptors were used in order to identify morphological and agronomical affinities, and it was determined that morphotypes were largely grouped according to fruit-type . Cypriot and foreign tomato germplasm was also genotyped by means of a single-tube microsatellite analysis and it was established that a rather limited amount of intra-variability exists within landraces. Nonetheless, it was ascertained that complex interchange of genetic material must have taken place in the tomato lineages. Finally, mineral levels and bioactive phytochemicals were determined in order to acquire the nutrient profile of these traditional tomato types and their eligibility in future breeding efforts.