Tracheliastes polycolpus. We collected parasites according to a hierarchical sampling design, with the sampling of all parasites from all host individuals captured in eight sites spread along an upstream–downstream river gradient. Individual multilocus genotypes were obtained from 14 microsatellite markers, and used to assign parasites to full‐sib families and to investigate the genetic structure of T. polycolpus among both hosts and sampling sites. The distribution of full‐sibs obtained among the sampling sites was used to estimate individual dispersal distances within families. Our results showed that T. polycolpus sibs tend to be aggregated within sites but not within host individuals. We detected important upstream‐to‐downstream dispersal events of T. polycolpus between sites , becoming scarcer as the geographic distance from their family core location increases. Such a dispersal pattern likely contributes to the strong isolation‐by‐distance observed at the river scale. We also detected some downstream‐to‐upstream dispersal events  that likely result from movements of infected hosts. Within each site, the dispersal of free‐living infective larvae among hosts likely contributes to increasing genetic diversity on hosts, possibly fostering the evolutionary potential of T. polycolpus.Inferring parameters related to the aggregation pattern of parasites and to their dispersal propensity are important for predicting their ecological consequences and evolutionary potential. Nonetheless, it is notoriously difficult to infer these parameters from wildlife parasites given the difficulty in tracking these organisms. Molecular‐based inferences constitute a promising approach that has yet rarely been applied in the wild. Here, we combined several population genetic analyses including sibship reconstruction to document the genetic structure, patterns of sibship aggregation, and the dispersal dynamics of a non‐native parasite of fish, the freshwater copepod ectoparasite  Tracheliastes polycolpus and its principal fish host Leuciscus burdigalensis in a French river network. Using genetic assignment approaches, we found that dispersal in T. polycolpus at the metapopulation level is mostly driven by waterflow during its free‐living infectious stage (downstream movements of ~25.4 km) and, to a lesser extent, by movements of its hosts during its adult parasitic stage (upstream movements of ~2.6 km). Furthermore, T. polycolpus sibs do not tend to aggregate on the same fish, thus fostering genetic variation within hosts.Knowledge about parasite dispersal is still scarce, although it may greatly influence key ecological and evolutionary processes such as virulence and local adaptation. We here focused on the freshwater ectoparasite  We analyzed the distribution of full‐sib families and the genetic structure of T. polycolpus at different scales, by hierarchically sampling all parasites from all hosts captured within eight sites along the upstream–downstream gradient of the Viaur River. Based on the ecological information available for T. polycolpus and its host , we built several nonmutually exclusive predictions. After hatching, the free‐living larvae of T. polycolpus released into the water column almost instantaneously develop into an infectious stage  over “large” distances until encountering a new host. We thus expected that parasites from the same clutch would drift, infecting hosts from downstream non‐natal populations. Finally, because daces are relatively sedentary and their dispersal particularly constrained by several artificial weirs and dams in the Viaur River  and, to a lower extent, several other cyprinid fish species , with a high prevalence (10%–90%) when compared to the average prevalence on alternative hosts . The attached parasites to each fin, if any, were counted before being collected using forceps and stored in ethanol for subsequent genetic analyses. All host individuals were then returned alive to their original sampling site.We focused our study on the Viaur River, a 169 km‐long river located in the Adour‐Garonne drainage basin in southwestern France Figure . Eight s® Multiplex PCR Kit (Qiagen). The two PCR were carried out in a 10 μl final volume containing 5–20 ng of genomic DNA, 5 μl of 2 × QIAGEN Multiplex PCR Master Mix, and locus‐specific optimized combination of primers (Appendix ®) under the following conditions: 15 min at 95°C followed by 30 cycles of 30 s at 94°C, 90 s at 56°C, and 60 s at 72°C and finally followed by a 45 min final elongation step at 60°C. The resulting PCR products were separated by electrophoresis on an ABI3730 at the GenoToul (Toulouse France). Allele scoring was performed using GENMAPPER version 4.0.Individual DNA extractions were performed on parasite trunks to avoid any contamination with genetic material from eggs, following a standard salt protocol  using Microchecker v2.2  using GENETIX  was estimated using NeEstimator v.2.1  within hosts, (b) among hosts within sites, and (c) among sites.We then explored how the genetic diversity of 2.5T. polycolpus were reconstructed using the full‐likelihood approach implemented in COLONY 2.0  the membership status of each pair of individuals to the same family  and (b) the membership status of each pair of individuals to the same site . Based on our observed dataset, we computed the proportion of full‐sib pairs sharing the same site  given the total number of full‐sib pairs over the river . This observed proportion was compared to a series of expected proportions under the null hypothesis of a random distribution of full‐sib pairs among sites, using 10,000 random permutations of the site matrix to compute the probability of correctly rejecting the null hypothesis  the membership status of each pair of individuals to the same family (sibship matrix) and (b) the membership status of each pair of individuals to the same host . We then computed the proportion of full‐sib pairs sharing the same host  given the total number of full‐sib pairs within the considered site and compared this observed proportion to a series of expected proportions under the null hypothesis of a random distribution of full‐sib pairs among hosts, using 10,000 random permutations of the host matrix.2.7T. polycolpus along the Viaur River, we focused on a subset of full‐sib families including at least five full‐sibs (N = 94 families). For each of these 94 families, we first determined a “family core location” as the mode of the kernel distribution of the geographical distance of each family member to the river source using the R‐package stats  with a parasite load of 13.4 ± 13.2 (mean ± SD). A total of 1,207 parasites were sampled from infected hosts. Over all sampling sites, He was 0.52 ± 0.01 (mean ± SD), Ar ranged from 3.43 to 4.05, and FIS ranged from −0.02 to 0.01 . The mean genetic differentiation estimated overall sites, and overall loci was φST = 0.08, and pairwise φST values between sites ranged from 0 to 0. 21, suggesting weak to moderate genetic structure in the Viaur River. We found however a strong and significant correlation between pairwise φST and pairwise riparian distances between sites  as expected under an isolation‐by‐distance pattern . Moreover, the proportion of full‐sib pairs belonging to the same site and infecting the same host differed slightly (but significantly) between sites, and ranged from 5.7% to 23.6%  and ranged from 0 to 78.6 km  fraction of the overall reconstructed full‐sib pairs was found to be “aggregated” within sites . This pattern of within site “aggregation” strongly suggests that some recently hatched T. polycolpus were at migration‐drift equilibrium. This isolation‐by‐distance pattern also conforms to the result obtained from the AMOVA, which indicates that a significant fraction of the overall genetic variability of T. polycolpus along the river occurs among sites. With dispersal among hosts facilitating random mating and dispersal among sites resulting in gene flow, the hierarchical dispersal strategy of T. polycolpus probably contributes to maintaining high genetic diversity  that is mostly found on L. burdigalensis in the studied system and the latter showing both small population sizes and spatially limited movements in the studied system ; Investigation ; Methodology ; Writing‐original draft ; Writing‐review & editing . Keoni Saint‐Pé: Formal analysis ; Investigation ; Methodology ; Writing‐original draft ; Writing‐review & editing . Simon Blanchet: Conceptualization (supporting); Methodology (supporting); Project administration (supporting); Supervision ; Writing‐review & editing . Geraldine Loot: Conceptualization ; Funding acquisition (lead); Methodology (supporting); Project administration (supporting); Supervision ; Writing‐review & editing . Olivier Rey: Conceptualization ; Funding acquisition (supporting); Investigation ; Methodology ; Project administration (lead); Supervision ; Writing‐original draft ; Writing‐review & editing .Fieldwork was conducted with adequate administrative permits for electrofishing , and fish were processed in accordance with the French Law .Appendix S1‐S5Click here for additional data file.


Xa23) and rice blast (Pid3), and identified unique haplotypes that were different from those of the six rice ecotypes in Guizhou. This study shows that KSR is an excellent rice germplasm resource, provides important information for the improvement and utilization of rice landraces, and serves as a reference for formulating effective rice conservation measures.Kam Sweet Rice (KSR) is a special kind of rice landrace that has been domesticated for thousands of years by the local Dong people in southeast Guizhou province, China. KSR has many distinguishing characteristics including strong fragrance; high resistance to diseases, pests, and adverse abiotic conditions; difficulty of threshing; and glutinous texture. There is a lack of systematic research on its genetic diversity. In this study, we analyzed the levels and patterns of genetic diversity and nucleotide variation in 1,481 rice germplasm using simple sequence repeat (SSR) markers and single nucleotide polymorphism (SNP) haplotype analysis of six unlinked nuclear loci. The accessions included 315 KSR resources from southeast Guizhou, 578 rice landraces from six rice-growing ecological zones in Guizhou, 546 rice landraces from nine provinces around Guizhou, and 42 wild rice sources. Genetic diversity and heterozygosity of KSR were both low, and thus KSR might be close to a pure rice line. Population structure analysis showed that KSR was isolated into a single type of rice, which had a large genetic distance and a unique genetic background compared to the local varieties in Guizhou province, indicating that KSR is a special rice ecotype. Haplotype analysis of the target genes showed that the population of KSR was rich in haplotypes for resistance to bacterial blight ( Oryza sativa L.), one of the three major global food crops, has a long history of domestication and is now the staple food of nearly half of the world’s population. Rice germplasm resources are the material basis supporting original innovation of agricultural science and breeding  (japonica varieties in China (16.72%) (P < 0.01). The low amylose content is one of the key factors for the stickiness and good taste of KSR, and indica (2.71%) and japonica (2.85) screened in China   and the (16.72%)  , rice landraces of neighboring provinces (NP), and wild rice (WR). Among them, 315 KSR were collected ; for GZ, 578 rice landraces distributed in 72 counties and six ecological rice-growing areas from other areas of Guizhou except southwest were selected; 546 NP accessions were selected from Jiangsu, Zhejiang, Anhui, Jiangxi, Guangdong, Guangxi, Hunan, Hubei, and Yunnan; and 42 WR accessions were selected from Guangdong, Guangxi, and Hunan. Furthermore, the 1,481 materials included 537 core germplasms from the primary core germplasm of cultivated rice in China, to represent rice grown throughout China. Rice landraces in Guizhou covered all ecological and climatic types in the province. Rice landraces from all of the surrounding provinces of Guizhou were also taken, and counties in these provinces were sampled as much as possible to achieve comprehensive coverage. The sources and quantities of 1,481 accessions are shown in 2O and finally preserved at −20°C.Total genomic DNA was extracted manually from fresh young leaves using a modified CTAB procedure . Fresh lA set of 36 SSRs , evenly Sorting the peak values obtained by capillary electrophoresis. Method of molecular marker data conversion, statistical analysis, Nei’s genetic distance calculation, analysis of molecular variance (AMOVA) and population differentiation coefficient calculation refer to SKC1, SAP8, Pid3, Xa23, GS5, and Ehd1). The structure of the gene is showed in According to the characteristic of KSR described by the Dong farmers in local research and existing research results , KSR is SKC1, SAP8, Pid3, Xa23, GS5, and Ehd1) were obtained from the NCBI database, and oligonucleotide primers were designed using primer5. PCR amplifications are the same as The DNA sequences of parts of six rice genes , at 6.01 and 93.99%, respectively, indicating that the genetic differences mainly came from within populations.The assessment of genetic diversity in the four groups of rice accessions is shown in K = 2, the statistical value ΔK showed a peak, suggesting that the number of subpopulations should be two, namely, japonica (Group I) and indica (Group II) , indicating that KSR were mostly japonica. II-2 had 268 rice accessions that were indica-NP and WR.To infer population structure, we performed model-based simulations for the 1,481 accessions using 36 SSRs. When roup II) . Furtherindica and japonica subpopulations and wild rice subpopulations. The indica population was further divided into two groups and the japonica population into three groups. The results of N-J clustering and the structure analysis based on the model were consistent, indicating that most KSR germplasm was japonica, which was different from other rice resources in Guizhou, though all of them originated from the same province. Rice resources from other parts of Guizhou are mixed and clustered together, but all KSR accessions were clustered together, highlighting its unique genetic background, and identifying KSR as a special ecotypic rice species.To further study the population structure of the 1,481 rice varieties, we constructed an N-J cluster diagram based on Nei’s genetic distance . The 1,4P < 0.001), indicating that the genetic differences mainly came from within subpopulations, and the average coefficient of genetic differentiation between subpopulations was 0.2201 (P < 0.001), with a range of 0.0985–0.3198. We calculated the genetic distance and differentiation coefficients of five subpopulations based on Nei’s minimum distance (1972) using Power Marker3.25 and Arlequin 3.5.2.2, respectively  showed the same result, indicating that KSR has a large genetic differentiation (0.15 < 0.2308 < 0.25) from GZ, marking it as a special ecotypic rice variety different from the local rice varieties in Guizhou province. On the contrary, KSR had small genetic differentiation and a close relationship with NP, suggesting that there may be a certain evolutionary relationship between them. The result of N-J clustering dendrogram based on Nei’s genetic distance is the same  of mean nucleotide variation among KSR , GZ , NP , and WR   on these genes, which control salt tolerance, grain size, rice blast resistance, and cold tolerance in rice. This may be due to negative selection or strong positive selection during the long-term domestication by the Dong people.Six unlinked gene regions– 0.0026) . The neuch locus . There wGS5, Ehd1, Pid3, SAP8, SKC1, and Xa23 gene loci, respectively. Among the six genes sequenced, KSR, GZ, NP, and WR shared dominant haplotypes H_1 and H_2, which appeared most frequently, and other haplotypes were radiating or staggered around them. The haplotype frequencies were as follows: GZ (39.0%) > NP (36.8%) > KSR (21.3%) > WR (2.6%)  were common (frequency > 25%) in KSR. Further, three haplotypes  were dominant in KSR, with proportions of 56.8, 70.5, and 75.0%, respectively. This indicated that genes for resistance to blast and bacterial blight were abundant in KSR, consistent with the nucleotide diversity and the neutral test of the six genes. Subsequently, we performed haplotype cluster analysis of Pid3 and Xa23 genes  and the dominant NP haplotypes  clustered into one group. These results suggested that KSR might have a close evolutionary relationship with NP, consistent with the clustering results based on SSR molecular marker data.We investigated the network of major haplotypes at six loci , with alR (2.6%) . NP had 23 genes , which sAMOVA  showed tPid3 . For Pid3 , the domd3  , 40 riceIn terms of KSR diversity, Cluster analysis and population structure analysis are effective means to explore the genetic diversity and background of crop germplasm resources, and are prerequisites for the protection and utilization of crop germplasm resources . Many scindica and japonica subgroups based on Structure analysis and the N-J cluster map. The indica and japonica populations were further divided into two and three groups, respectively, similar to the study of population structure division of 537 rice varieties in Guizhou Province by japonica subgroup, according to the calculation of Nei’s genetic distance and the FST value; the genetic distance between KSR and GZ (FST = 0.2308) was higher than that between KSR and NP (FST = 0.1141). The 1,481 accessions were divided into GS5; H_10 of Pid3; H_7, H_8, and H_9 of SKC1), which are essential in the breeding of superior varieties.Polymorphic variation in gene sequences results in the genetic diversity of species. As the environment changes, some mutation sites that are more adaptable to the environment are retained. The most common sequence variation is the insertion and/or deletion of single nucleotide polymorphisms (SNPs) and InDel fragments . SNPs arex situ conservation, and should be encouraged and strengthened, especially in the genetic diversity centers of crops in southwest China.Therefore, on-farm conservation of farmers’ traditional tillage system is an important way to conserve germplasm diversity, which supplements Eleusine coracana L.), wheat (Triticum turgidum L.) (Hordeum vulgare L.)  around the world . Understidum L.) , and higgare L.)  is to meThe Dong people are the descendants of the ancient Yue people, who lived on glutinous rice as their staple food, and as such, the Dong people have a long history of eating glutinous rice. As the most distinctive rice landrace in Guizhou province, KSR not only has good resistance to diseases and insects, but is also used in the traditional diet, festival celebrations, sacrificial activities, and other cultural customs of the Dong people, and thus has been planted and used by the Dong people for thousands of years. In addition, KSR adopts the organic ecological planting mode of “rice-fish-duck” co-existence completely , which hThe datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/CL performed statistical analysis and wrote the manuscript. CL, YW, DC, and BH conceived and designed the experiments. YW, AJ, XM, HC, RR, and XL conducted field surveys and collected germplasm resources. YW, AJ, and LH sorted out the collected germplasm resources and incorporated them into the National Medium term Gene bank. YW performed the experiments. LH supervised the thesis. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


We did semi-structured interviews, free listings, and tours with 11 agave managers. We analyzed morphology and studied genetic diversity and structure using nuclear microsatellites. We found wild-collected, tolerated, transplanted, and cultivated varieties of agave. This comprised 19 traditional varieties of pulque agave, 12 of them in production during the study, which corresponded to the species Agave americana, A. salmiana y A. mapisaga and five intraspecific entities. The varieties were grouped morphologically according to a management gradient; the wild-collected varieties were the smallest, with more lateral teeth and a larger terminal spine. The cultivated varieties clearly exhibited domestication syndrome, with larger plants and smaller dentition. The expected heterozygosity (He) of the varieties ranged from 0.204 to 0.721. Bayesian clustering suggested the existence of three genetic groups, both at the level of traditional varieties of pulque agaves and for management categories, a result that matches multivariate clustering. Pulque producers in the studied localities maintain high agrobiodiversity. The cultivated varieties exhibit domestication syndrome, as has been reported for other species of the genus with the same selection purposes. Our results support the hypothesis of a decrease in genetic diversity in crops compared to wild-growing agaves, which seems to be due to vegetative propagation, among other factors.The agaves are plants of cultural importance which have been used by humans for about 10,000 years and about 40 specific uses. The most culturally and economically important of those uses are for the production of fermented (pulque) and distilled beverages . Pulque continues to be produced in nearly all of Mexico, and the agaves used for this purpose have shown domestication syndrome. We carry out an ethnobotanical, morphological, and genetic analysis of the traditional varieties of pulque agave used in the production of  Agave genus is the richest taxon within this subfamily, with 210 species distributed from the southern United States to Colombia and Venezuela and the Caribbean Islands [Agaves are a group of monocotyledonous, rosetophilic, succulent and monocarpic plants. They were reclassified to the Asparagaceae family, but the Agavoideae subfamily continues to be recognized. The  Islands . In Mexi Islands ,4.Agave and Littaea. The greatest diversity of Agave is found in the Tehuacan-Cuicatlan Valley, shared between the states of Oaxaca and Puebla (approximately 20 species), while the area with the greatest diversity of Littaea (eight species) is in the Barranca de Metztitlán, in the state of Hidalgo [According to the morphology of their inflorescences, Gentry  classifi Hidalgo ,7.Agave taxa have been reported, including intraspecific varieties [Agave americana, A. lechuguilla, A. mapisaga¸ and A. salmiana—have been widely used for several purposes in the region. For example, A. lechuguilla is used to obtain fibers, make articles for cleaning and for bags. The other three species are mainly used for culinary purposes; for example, flowers are collected to prepare different dishes ; the leaves (penca) and cuticle (mixiote) are used as utensils and ingredients for cooking food; drinks are prepared from the sap extracted from the central corm, both fresh as aguamiel and fermented as pulque; and preparing syrup sap (jarabe de aguamiel).In the Hidalgo State, 17 arieties . Of thesArchaeological records suggest that the use of agaves dates back at least 9,000 years, evidenced by findings of remains of chewed agave in different sedimentary floors in caves of the Tehuacan-Cuicatlan Valley –10. TherThe ancient and constant interaction between humans and agaves has generated different evolutionary pressures on the plants, such as artificial selection by several cultural groups that have selected plants with characteristics to satisfy their necessities. This type of selection has exacerbated or reduced the frequency of certain characteristics of the plants, which favor the abundance or reproduction of individuals with these attributes. Plants under the same artificial selective pressures converge on a set of traits that differentiate managed individuals from their ancestors and/or wild relatives, a process known as domestication syndrome –16.It has been suggested that for the agave species whose main use is the production of fresh or fermented beverages, the domestication syndrome is aimed at obtaining a greater quantity and quality of sap, which leads to plants presenting: 1) gigantism—taller overall height and leaf length and width—since larger individuals produce greater volume of sap, 2) sap with desirable organoleptic properties like higher sugar content, neutral pH, reduction in irritating compounds and structures such as saponins and raphides, and 3) reduced “thorniness—size, number, and closeness of lateral teeth and size of the terminal spine, all relative to leaf length—to avoid injury to sap collectors from these plant defense mechanisms –21. ThisAgave mapisaga is the largest species and has the smallest lateral teeth; its varieties have only been recorded in anthropic environments, with no records in natural ecosystems. A. salmiana is a species with high morphological diversity, several varied of forms to the exploitation and had a gradient of domestication. On the other hand, A. macroculmis and some wild varieties such as A. salmiana ssp. crassispina, do not show characteristics associated with domestication syndrome [The morphological variation and domestication syndromes of agaves used for pulque production have been studied by several authors. The results indicate that syndrome –26.ST = 0.68), and low gene flow (Nm = 0.24). The second study was of the wild relative of the pulque agave, A. salmiana ssp. crassispina in San Luis Potosí. In that study, AFLPs were used as the molecular marker, and the authors found high genetic diversity (He = 0.403), no population structure (FST = 0) and high gene flow (tending toward infinity) [A. hookeri species in the P´urhepecha highlands in Michoacán, Mexico; nuclear microsatellites showed low population genetic diversity (He = 0.485) compared to the most likely wild ancestor (A. inaequidens He = 0.704–0.733), strong population structure (FST = 0.28) and fixation of heterozygotes, a genetic trait associated with the selection of favorable genotypes in crops [The levels of genetic diversity of the agaves used for pulque production has only been addressed in five studies. The first was by Alfaro-Rojas and collaborators , who stunfinity) . The thiin crops ,20. Finain crops , studiedin crops ,29.Agave salmiana var. salmiana and Agave mapisaga are the most widely used species for the extraction of sap, both in extensive crops and within the plots of small producers [Hidalgo is the state with the largest cultivable area of pulque agave in Mexico. This area of approximately 4,905 ha represents 60% of the cultivation in the country for the year 2019, followed by Mexico State with 19% and Puebla with 12%. Hidalgo is also the top nationwide producer of pulque, producing 117,432,130 liters per year  [roducers ,23.in situ or cultivated, 5) they are found to a greater extent in traditional production systems, although they can also be found in intensified systems, 6) there is a wealth of traditional knowledge and techniques associated with the management and use of each variety. These traditional varieties are of cultural and economic importance, are part of the identity of cultural groups and satisfy multiple needs—especially the nutritional needs—since the derived products are used both for self-consumption and for commercialization, generating monetary income for the families.“Traditional varieties” refers to useful plants that are recognized, named, managed, propagated, and preserved by the producers ,31,32. TIn localities of the Mezquital Valley, Hidalgo, 25 traditional varieties of agaves are reported. Richness is higher in indigenous than mestizo communities, but in both cases, agaves are pillars of food self-sufficiency and family economy ,23,35,36Analyzing the characteristics and status of these plant genetic resources is essential to ensure the maintenance of the biological and cultural diversity of Hidalgo and Mexico, generate information that allows decision-making and the design of strategies for sustainable resource management, and improve the livelihood that communities will be able to maintain, manage, and continue to use these resources.This research was carried out in two localities in the state of Hidalgo with a tradition of using agaves for producing pulque and other purposes. Our objectives were: 1) to carry out an ethnobotanical characterization of the traditional agave varieties, determine their management categories and describe the productive system, 2) to evaluate the morphological characteristics of the different varieties and to analyze their correspondence with domestication syndrome, and 3) to quantify the genetic variability of the different species and traditional varieties of agave. This will allow us to discern the consequences of management and the degree of domestication of the traditional varieties of agave whose main uses are sap extraction and pulque production in this region of Mexico. We hypothesize that agave varieties with a more intense degree of management will have morphological characteristics associated with the domestication syndrome that have been previously described for pulque agaves, while varieties that are less intensely managed will present the characteristics of the syndrome to a lesser degree. On the other hand, we expected cultivated plants to have lower genetic diversity and stronger population structure than wild-growing plants.The institutions to which the authors belong do not have internal ethics committees that endorse the ethnobiological methodology applied in this research, however it has been reviewed and approved by peers of our institutions, in addition this research was carried out following the statutes of the Code of Ethics for research, action research, and ethno-scientific collaboration in Latin America of the Latin American Society of Ethnobiology (SOLAE). At the beginning of the research, we established contact with civil authorities and people in the communities and owners of agave crops. We presented the study project, its aims, and methods and asked their consent to collaborate with us. All information obtained  were acquired with previous express permission from the participants.2, which represents 1.1% of the total area of Mexico [El Cubo, in the Cardonal municipality of the Mezquital valley, has 63 inhabitants, ten of which speak the Hñähñu indigenous language [Grutas de Tolantongo caves, located 11 km away. There, we studied the agave plantations of two producers  of forward and reverse primers, 10 mM (0.3 μL) of dNTP mix, 2.3 μL of buffer (5x), 0.5 μL of DNAse-free water and 1.5U (0.3 μL) GoTaq® Flexi DNA Polymerase (Promega). The PCR products were resolved on a 15% polyacrylamide gel stained with ethidium bromide (10 mg/mL) run for 90 min at 90 V.In total, 127 samples of agave tissue were collected from healthy leaves  were scored using GelAnalyzer 2010a . We chooO and He), and genetic diversity—were calculated using the R package “Poppr” [The descriptive diversity statistics—the percentage of polymorphic loci (PPL), mean allele diversity (A), effective alleles (Ae), observed and expected heterozygosity  was calculated, correcting for null alleles, with the INEst program [3 bootstrap replications in Poptree2 software [K), were performed [https://lmme.qdio.ac.cn/StructureSelector/ [Salmianae group were included, and we excluded varieties that were represented by a single individual.The global and pairwise Fetitions . The inb program  using th program  using 10software  at the lsoftware . Ten inderformed . The apperformed  generateelector/  at the lelector/ ,60. For El Cubo, the producers have small plots that have been used for producers for at least two generations. The agaves are cultivated in hileras (rows) interspersed with other crops or as living fences within producers’ properties. The CCUB1 plot is a production system measuring 3.529 ha, where agaves are cultivated in rows within fields of seasonal crops, such as maize and beans. The CCUB2.1 plot is a backyard orchard with fruit trees and ornamental plants, occupying an area of 0.35 ha. In the CCUB2.2 and CCUB2.3 plots, agaves are cultivated in rows among crops, such as maize and beans, and occupy a space of 1.309 and 0.781 ha, respectively, belonging to a single producer of Hñähñu origin. The agaves are propagated by transplanting 50- to 100 cm-tall suckers from the agaves from the plot into rows. Agaves purchased from other producers in the area are also occasionally incorporated into the plots.In barbacoyeros  for 100 Mexican pesos (4 USD) per plant. In this locality we recorded eight traditional varieties of agave used for pulque production. There was one wild-growing variety, known as 1) Corriente, or Bronco or Verde de monte, and seven cultivated varieties: 2) Chino, 3) Xaminí, 4) Penca larga, 5) Mutá, 6) Hoc´uadá, 7) Poblano, and 8) Guanté  (Corriente), but that they currently use cultivated agaves.Agave suckers are classified for sale according to their vigor into first-rate plants, sold for about 50 Mexican pesos (2 USD) each, and second- and third-rate plants that sell for about 30 Mexican pesos (1.2 USD). Some families in the area sell their plants to pulque producers and  blanco) , Fig 2. acocote, essentially a large straw. One end is inserted into the cavity of the agave, and suction is generated by mouth at the other end, drawing the sap into the acocote which is then emptied into a bucket. Acocotes were traditionally made with a long, dried Cucurbitaceae fruit (Lagenaria siceraria), however due to the low availability of this material and its fragility, it has been replaced by plastic materials. Currently, acocotes are made by attaching a plastic hose (the end inserted into the plant) to a plastic bottle, which is drilled at the base . Pulque is produced with a combination of different sap varieties. The producers mix the fresh sap with old pulque, produced the day before. The fresh sap and pulque are used for self-consumption and sold locally at 10 Mexican pesos (0.4 USD) per liter, or to tourists for 25 Mexican Pesos (1 USD) per liter.In this locality, each producer uses six to 20 plants simultaneously. The sap is collected with the elongated tool known as an La Coyotera has been cultivating agaves for around 30 years. A total of 44 hectares are dedicated to agave production, where the plants are found at different densities and are managed in three ways: living fences, jilas, and nursery. For living fences, used to delimit spaces and paths, the Corriente variety is preferred, since it has more teeth , longer spines , is fast growing, and has deep roots. Jilas refers to rows of agaves that are interspersed with native vegetation and are planted perpendicular to the slope of the land. Juvenile agaves are cultivated and kept in nurseries to protect them from goat browsing and allow them to develop and are then transplanted into the jilas when they reach 100 cm in height.Corriente, 2) Corriente cenizo, 3) Corriente colorado, 4) Corriente espina china 5) Corriente penca ancha, 6) Corriente penca larga, 7) Manso de zoqui, 8) Manso del altiplano, 9) Xaminí, 10) Púa larga, 11) Verde, 12) Penca larga 13) Blanco and 14) Sabililla  abililla , Fig 2. In both localities there is a gradient of management practices that can be grouped into the following categories:in situ and used for various purposes. According to the people interviewed, some 70 years ago there were no crops, and these agaves were used by wild collection.Wild collected (WCO). Agaves that inhabit the surrounding areas of native vegetation. These plants are collected Tolerated (T). Agaves that were already growing wild in an area of native vegetation that was later converted to a productive system and have been maintained by the managers within the new system. These plants receive some management, such as removing dry leaves.Transplanted (TR). Agaves removed from their original place, generally from nearby ecosystems, and placed within productive systems, forming part of the rows and living fences.Cultivated (C): Agaves cultivated within productive systems. This category generally contains the most used and valued traditional varieties. These are given greater care, including pruning and more intense propagation. These are the varieties that are bought or exchanged between localities.cerro and monte agaves (wild-collected) from cultivated ones. These characteristics are the size of the plant and its leaves, the shape and length of the spines, and the number of lateral teeth. Another important characteristic for managers is the color of the plant, which ranges from different shades of green to glaucous. Characteristics such as the time to reproductive maturity, the sap quantity, and qualities (such as sweetness or viscosity), and the duration of sap production are also considered . The other subgroup was composed exclusively of cultivated agaves, highlighting the presence of individuals of the variety Penca larga (A. mapisaga), which represent the most marked domestication syndrome. It should be noted that it is a species whose wild ancestor is unknown, and it is a species that is exclusively under cultivation.The heatmap showed that the agaves were grouped according to the management category. Two large groups were observed, the first one in the upper part composed mainly of cultivated agaves, and the second group mostly containing wild-collected agaves. The attributes that distinguished them were related to domestication syndrome (gigantism and thorniness). The cultivated agaves (top in the heatmap) had larger size and were less thorny, regardless of their variety. They had smaller lateral teeth and the teeth were smaller proportionally to the leaf. This first group of cultivated agaves was divided into two subgroups: the first one made up of C and some WCO and T individuals that, although they belong to other management categories, present some morphological characters with tendencies towards domestication syndrome and represent those agaves that are progressively selected for possessing these desirable characteristics, such as larger size (Guanté (A. americana), which represents a species It distinguishes the Salmianae group and there it is located with greater morphological similarity with the WCO agaves, since it is not a species of size as large as Agave salmiana var. salmiana or A. mapisaga. The group at the bottom of the heatmap contained the agaves with the highest values of variables related to thorniness, such as high number of teeth in relation to the size of the leaf  was the smallest in terms of GPL, SL, D, and LL. The cultivated variety Penca larga (A. mapisaga) was the largest in terms of GPL, D and LL, while the cultivated variety Poblano  had the highest average SL. In terms of thorniness, Penca larga had more teeth and more closely spaced teeth than the other varieties, but the size of the teeth was smaller than the wild varieties. The variety Manso de zoqui also had a smaller tooth size. This confirms the domestication syndrome of lesser dentition for easier manipulation  was made up of individuals cultivated from traditional varieties Penca larga, Manso de zoqui, Mutá and Poblano. The third group (bottom center) corresponded to tolerated and transplanted individuals of different varieties (their centroids did not differ), even wild and cultivated individuals are distinguished. Wilk’s Lambda had a value close to zero , indicating that the information provided by the variable was statistically significant, allowing the discrimination of groups whose centroids are not same (wild collected and cultivated) and have little overlap. Only 7% of the individuals were not correctly classified into the category management assigned a priori. 90% of the individuals in the management category WCO were classified correctly; 7.5% were classified T and 2.5% to TR. 85% of the individuals in the T category were classified correctly and 15% classified as TR. All individuals in the TR category were correctly classified. In the case of management category C, 96% were classified correctly and 4% to category T.The DFA explained 96.03% of the variance . Three large groups were differentiated; one contained the wild collected varieties (left side of p < 0.05). LD was observed between four pairs of loci  in traditional varieties ranged from 43.75% (Guanté) to 100% . In the management categories, this was 100% for wild collected (WCO) and cultivated (C) and 93.75% for tolerated (T) and transplanted (TR). The allele richness (A) in the traditional varieties ranged from 1.563 (Guanté) to 6 (Corriente), and the effective number of alleles (Ae) varied from 1.459 to 4.076 in the same traditional varieties  to 0.214 (Poblano) and the expected heterozygosity (He) varied from 0.219 (Guanté) to 0.730 (Corriente) (O) ranged from 0.135 (TR) to 0.204 (T) and the expected heterozygosity (He) varied from 0.495 (T) to 0.701 , for A. salmiana ssp. crassispina , for Agave salmiana var. salmiana ; for A. mapisaga  and for A. americana . FST with ENA correction was 0.127, indicating a moderate genetic differentiation among management categories. FIS ranged from 0.194 (Guanté) to 0.720 (Corriente), indicating strong inbreeding. The neighbor joining tree based on Nei’s distances showed three groups. The first included the traditional varieties: WCO Corriente, WCO Corriente cenizo and WCO Corriente penca ancha. The second group contained TR Corriente, C Manso de zoqui, TR Corriente penca larga, T Corriente Penca larga and T Corriente, and the third contained the C Poblano, C Mutá, C Penca larga varieties .Null alleles were found at 16 loci, suggesting homozygote excess. Only one locus (BYU4463) did not exhibit null alleles. The estimated null allele frequencies over traditional varieties and managed category varied from -0.0738 (7%) at BYU4463 (this suggest heterozygote excess) to 1 (monomorphic loci) at APAR3-11, APARLC28, APARLC34 and BYU 4012. Sixteen loci showed departures from HWE (rriente) . By mana0.701 W, . Averagearieties . The Bayarieties . The bluarieties  matched A rich diversity of traditional varieties of pulque agave are used in the localities studied in the state of Hidalgo, which are maintained in a management gradient in traditional agroforestry systems. The cultivated plants clearly exhibited domestication syndrome related to gigantism and reduced thorniness. We also found support for the hypothesis of decreased genetic diversity and moderate population structure among cultivated pulque agaves compared to wild populations.Agave americana, A. salmiana and A. mapisaga. The assignment of taxonomic identities in agaves is a complex task. Some of the species descriptions are based on ex situ specimens, and agaves are a group of plants where hybridization is common, even more so in cultivated species and varieties. In our case, the plants managed in crops are difficult to identify at the species level because reproductive structures are frequently removed as part of management, so these structures are not available to aid in identification.In Mexico, about 117 agave taxa have been reported to be used for various purposes, which have been assigned about 570 common names and represent varieties of agronomic interest . We founA. applanata among their crops, which was widely used in the past. In the state of Tlaxcala, nine varieties of pulque agaves have recently been identified, with the novelty that the Ayoteco variety, which had been located taxonomically as A. salmiana, was revealed by molecular analyses to be more closely related to varieties of A. mapisaga [Xaminí.In the southern highlands, Mora-López et al.  recordedmapisaga . In the mapisaga  found simapisaga . AccordiAgave is highly variable, with high plasticity. This is due to the large number of ecotypes and clinotypes, as well as to the impact of factors such as arid environments and life history characteristics like pollination on Agave evolution [Several authors agree that the genus volution –65. In tLa Coyotera and El Cubo corresponded to Metepantles, from nahuatl metl: agave, pantli: rows, a traditional agroforestry system that consists of rows of agave interspersed in plots of different plants, which maintain and conserve a high diversity of crops and native vegetation, such as Corriente or Cimarron agaves. The management of agaves has led to maintaining higher densities of traditional varieties that yield greater productivity in terms of quantity and quality of sap, as is the case of the Manso de zoqui variety. However, producers maintain and protect other varieties even when they are not so profitable, as part of their interest in safeguarding part of their cultural heritage. The continuous selection of some of these agave varieties has led to individuals presenting a larger rosette size and decreased thorniness, as has been recorded in other works exploring domestication syndrome in pulque and mezcal agaves [In Mexico there is a wide diversity of systems and intensity of management of pulque agaves . In thisl agaves ,19–23,67l agaves . ProduceAgave are polyploid [Littaea and 64.3% of the subgenus Agave [A. americana, diploid (2x), tetraploid (4x), and hexaploid (6x) plants have been found. A. mapisaga presents sterile pentaploids (5x) [A. salmiana tetraploid and hexaploid plants (6x) have been recorded [It has been proposed that producers traditionally select vigorous, vegetatively propagated suckers, and that these plants can be polyploid, which are maintained for generations through clones ,64. Polyolyploid ; 40% in ids (5x) ,69, and recorded . This maFor future analyzes of agave populations, we consider it important to include, in addition to the taxonomic classification, the classification of traditional varieties used by producers in each specific region, since this classification system allows analyzing the artificial selection processes to which agave populations are subject, both domesticated species in crops, and wild species that are being managed.ST, FIS and decrease genetic diversity [For the varieties and species in this study, the low levels of observed heterozygosity compared to expected heterozygosity and the high levels of inbreeding may be related to the presence of null alleles with frequencies greater than 8%, which may increase the parameters of population differentiation such as Fiversity . On the S = 0.190, FST = 0.150) to monocot angiosperms in isoenzymes  to our values, although their values for the species A. americana were higher than ours  and RAP = 0.31; ). The di = 0.31;  evaluatean ours  presented lower levels of genetic diversity than its wild ancestor, A. inaequidens (He = 0.704). Similarly, Alfaro-Rojas et al. [A. salmiana var. salmiana  and A. mapisaga  in northeastern Mexico state using RAPD’s, which compared to the high levels of diversity recorded in wild populations of A. salmiana ssp. crassispina in San Luis Potosí, using AFLP’s (Hs = 0.408). The low genetic diversity in pulque agaves may be due to intrinsic characteristics of the plant, as well as the fact that the crops have mostly been propagated vegetatively, possibly for the past few thousand years, according to archaeological evidence. In addition, the fact that the management of plants eliminates the possibility of sexual reproduction, also prevents genetic recombination.The hypothesis of higher genetic diversity in wild than cultivated populations of pulque agaves has been supported by several authors previously. Figueredo et al.  found ths et al.  found thIn the case of agaves used to produce traditional mescal, patterns of genetic diversity have been found to be the opposite of those reported for pulque agaves. This high diversity among cultivated varieties is due to the fact that there is a wide range of management systems, where traditional agroforestry systems (TAFS) are a reservoir for a high number of traditional varieties, morphological and genetic diversity, due to the frequent and constant introduction of plants from other sources in these systems, effective management practices such as the formation of seedbeds and the implementation of management plans to promote and maintain sexual reproduction. In addition, there is historical information that in some species of agaves the production of mezcal began approximately 400 years ago, with cultivation being an even more recent phenomenon (approximately 30 years) .Metepantles and backyard gardens. The varieties studied presented a great diversity in their form, which is a product of the varied environments where these agaves grow as well as the diverse cultural motivations of pulque producers to maintain diversity in their crops and management practices. As such, these systems should be considered productive spaces that maintain diversity, practices that should be maintained in other productive systems in the country. The species with the greatest diversity of forms and traditional varieties was Agave salmiana, while the species that most clearly exhibited the domestication syndrome was A. mapisaga. The wild-collected agave species showed high levels of genetic diversity, while the cultivated varieties exhibited low levels of diversity, probably due to vegetative propagation. Varieties such as Corrientes or Cimarrones  are found in the natural ecosystems, which eventually enrich the gene pool of the crops through their occasional introduction into crops (Tolerated and Transplanted). Similarly, some cultivated agaves "escape" , which in turn has maintained some diversity in the agave populations.We found a high agrobiodiversity of agaves, which are maintained in traditional agroforestry systems such as S1 File(DOCX)Click here for additional data file.


Purpose: Informed consent for genetic testing has historically been acquired during pretest genetic counseling, without specific guidance defining which core concepts are required. Methods: The Clinical Genome Resource (ClinGen) Consent and Disclosure Recommendations Workgroup (CADRe) used an expert consensus process to identify the core concepts essential to consent for clinical genetic testing. A literature review identified 77 concepts that are included in informed consent for genetic tests. Twenty-five experts  completed two rounds of surveys ranking concepts’ importance to informed consent. Results: The most highly ranked concepts included: (1) genetic testing is voluntary; (2) why is the test recommended and what does it test for?; (3) what results will be returned and to whom?; (4) are there other types of potential results, and what choices exist?; (5) how will the prognosis and management be impacted by results?; (6) what is the potential family impact?; (7) what are the test limitations and next steps?; and (8) potential risk of genetic discrimination and legal protections. Conclusion: Defining the core concepts necessary for informed consent for genetic testing provides a foundation for quality patient care across a variety of healthcare providers and clinical indications. While they are focused on the entire genetic counseling process, which includes educational topics and psychosocial assessment and support beyond what may be needed for informed consent, some examples of professional guidelines specific to the genetic testing and counseling process include the National Society of Genetic Counseling (NSGC) Practice Guidelines  genetic testing. The Consent and Disclosure Recommendations (CADRe) workgroup of the NIH funded Clinical Genome Resource project  has been focused on proposing and standardizing approaches to communication about genetic testing beyond the traditional genetic counseling model. The CADRe framework proposes that there are many clinical indications where a targeted discussion or brief communication may be an appropriate pretest communication approach ,24. The This study used an expert consensus process to define the critical concepts required for informed consent for genetic testing in a clinical setting. We focused on clinical genetic testing primarily because the research on the genetic testing consent process may have additional considerations related to study protocols and IRB requirements. The Stanford University Institutional Review Board (IRB) reviewed this study and declared it exempt.We modeled the consensus study process after the approach of a modified Delphi method . SpecifiWe completed a literature review to develop a comprehensive list of the concepts involved in the consent process for genetic testing. The review identified publications listed in PubMed before June 2019 using the keywords “informed consent”, “consent”, “consent form”, AND “genetic or genomic” AND “counseling.” We then limited the papers reviewed to those that covered germline genetic testing in clinical and research settings and focused only on those that described the content of either consent forms or of genetic counseling that was part of the informed consent process. We also reviewed the relevant practice guidelines, consent forms from clinical laboratories offering exome testing, and direct-to-consumer genetic testing company websites’ educational materials. The concepts were extracted from the results of the literature review, combined with concepts extracted from prior focus groups with both patients and providers that were conducted by the Clinical Genome project’s CADRe workgroup  and discIn our study we focused on experts (defined below), because they brought content knowledge to the prioritization. We also utilized an anonymous process in order to allow equal voice to each participant, so that the more vocal or more esteemed experts were not given more weight than other participants. A critical feature of any consensus process is clearly defining the expert participant population, in this case, individuals who were considered experts in the topic of informed consent for genetic testing. In this study, experts were defined as genetics clinicians  or bioethicists with at least 5 years of experience in obtaining informed consent and results disclosure of genetic testing results, or research experience in the ethical, legal, and social implications (ELSI) of genetic testing who practiced in the United States. Experts were identified through their involvement in one of three areas: (1) ethics-related committees; (2) leadership participation in various clinical special-interest groups within genetics organizations including the National Society of Genetic Counselors (NSGC), the American College of Medical Genetics and Genomics (ACMG), the American Society of Human Genetics (ASHG), and Clinical Genome Resource (ClinGen); (3) genetics special-interest groups within non-genetics organizations ). Additional ELSI thought leaders in this area were identified as those who had authored multiple publications included in the literature review. The study participants were provided with USD 75 for each completed round of the surveys, for a maximum of USD 150 each.Between November 2019 and February 2020, we conducted two rounds of surveys using Qualtrics online survey administration software; the surveys were linked using a codename created by each participant at the end of Survey 1. We initially selected a two-round approach based on data that suggest that two to three rounds of surveys are adequate to obtain consensus .In Survey 1 , particiBefore completing Survey 2, approximately 2 months later, the participants were given their individual results from Survey 1 and a summary of the group results. Participants were asked to rerate all 77 concepts in light of reviewing this data and rated an additional 12 concepts that were suggested by participants during Survey 1. Participants were asked to comment on a list of the 10 most highly rated concepts, considering whether they thought the concepts included “the minimum necessary and critical information to aid an individual in making an informed decision/consent,” and, if not, which concepts they felt were missing or could be removed. They were subsequently shown a list of 18 ‘second-tier’ concepts and asked if they felt that any of these issues warranted inclusion on a core concept list and why. Lastly, participants piloted and provided open-ended comments on four consent scenarios that were intended to be used for a future national survey that would follow up on the results of the present study. Comments from these scenarios were used to inform discussions as the workgroup developed the ‘final concept list’ in this study.In order to define which concepts were most highly rated by the expert participants of Survey 1, we used SPSS version 26.0  to calculate the means, standard deviations, and modes for each concept. In Survey 1, we also tallied the number of times a concept was ranked in the top five within a category. For the “potential risks” category, since there were only five items, we tracked the top three items. The open-ended comments from both surveys were reviewed and discussed by the entire CADRe workgroup in order to determine which concepts were most highly endorsed for inclusion in the subsequent survey.We created a list of the top concepts from Survey 1 based on the items that scored a mean >4.0 (out of 5.0) and were also frequently ranked in the top-five lists. To ensure that we did not miss any critical concepts with these cutoffs, we created a second-tier of concepts that met one but not both of the criteria above; for example, concepts that either scored between 3.75–3.99 but were frequently ranked in the top five, or concepts that had a score >4.0 but were not frequently ranked in the top five.To identify the final consensus list of the core concepts that would allow “the minimum necessary and critical information to aid an individual in making an informed decision/consent”, the CADRe workgroup examined the mean responses for two rounds of survey results, using a combination of the criteria described above and looking for consistency across both surveys, as well as the open-ended comments from participants. The workgroup combined some closely related items to avoid repetition. One hundred and thirty-two potentially eligible experts were identified. After excluding 36 individuals who were members of the ClinGen Steering Committee or who were genetics laboratory directors without patient-facing roles, and 8 without publicly available emails, 88 potential participants were emailed in November 2019. Three emails were undeliverable, and 32 confirmed their interest through an eligibility survey , one of whom was ineligible based on years of experience. Ultimately, 25 individuals  completed Survey 1  wrote comments regarding the list in general. Ten participants were supportive of the proposed list.“I agree that it covers the concepts that most patients want to know about genetic testing.”“I think the list is helpful and focused. It should help improve the efficiency of obtaining consent and will probably improve the understanding of the people getting testing by simplifying the process.”“The list should allow for an informed decision. Some patients/families will have additional needs/interests which can be addressed during time allotted for questions.”Two added that they had no further comments, and one highlighted that the survey and their responses “underlines the challenges in streamlining consent”. Two participants highlighted items that they still felt were missing from the proposed concept list: the manner in which the sample will be obtained (and its respective invasiveness) and how potential results will be interpreted.Finally, when participants reviewed the list of the 18 second-tier concepts, the most highly endorsed potential concepts were: test limitations , variant interpretation will change over time , and the potential for genetic discrimination or stigma . Of note, six people, at least some of whom overlapped with those who endorsed including “genetic discrimination or stigma”, also endorsed “GINA (Genetic Information Nondiscrimination Act) and relevant state laws provide some protection”. Most of the comments regarding GINA/discrimination issues stressed that there are important issues that people may not know about, and may actively wonder or worry about, that should be discussed prior to testing.“I think this [GINA] can be wrapped into ‘potential risks for genetic discrimination and stigma, but patients should understand what is/isn’t protected legally before proceeding with testing.”Eight concepts  met the Over the past several years, the CADRe workgroup has taken up the question of how to “operationalize new approaches to provide patients with the education and support that each needs and desires when undergoing genetic testing” . Prior sThis study highlighted the following concepts as critical : the volWe propose that this list is tractable for both patients and clinicians, and provides all clinicians, whether genetics specialists or not, a focal point to begin the discussion about genetic testing with a patient. This concept list does not imply that only these concepts are important in order for a patient to be able to make an informed decision about whether to undergo genetic testing, but rather it prioritizes a ‘most critical’ list as a starting place for discussion, so that clinicians can avoid overwhelming patients with information that is potentially superfluous to their decision-making process. It may well be that some concepts, particularly the ‘second-tier’ concepts, are highly critical for a minority of patients in the pretest consent process, or that they are more uniformly important in the post-test genetic counseling process when the results are available. Furthermore, the core-concept list recognizes that the traditional genetic counseling process may go beyond the concepts required to allow patients to make informed decisions about genetic testing since these sessions often include information regarding the logistics of sample collection and the return of results. Given that our prior work suggests that, for most predictive genetic testing, an abbreviated (‘targeted discussion’) approach would be feasible and appropriate for many testing indications , this lihttps://www.federalregister.gov/documents/2020/05/01/2020-07419/21st-century-cures-act-interoperability-information-blocking-and-the-onc-health-it-certification, accessed on 26 June 2021), along with laws that allow for the direct release of genetic testing results to patients, may heighten this concern if they lead to limits in post-test counseling, potentially making some providers worry that it is even more critical to provide as much information as possible during the pretest consent conversation. Rather, we would emphasize that tailoring a pretest genetic testing discussion to these points allows a patient either to make an informed decision or to request further information, and that information beyond these points may be best suited to a post-test conversation when providers can contextualize the results and their implications in a focused manner. By shortening the pretest communication process to the discrete set of concepts that this study proposes, we expect that patients will still be able to make informed decisions about genetic testing, providers will feel more comfortable that they are covering the key concepts, and access may be improved through both the broadening pool of providers who are able to facilitate the genetic testing process as well as the shorter time frame required, allowing more patients to be seen in relevant clinics. This hoped-for improvement to access to quality care remains to be tested.We anticipate that there will be some genetics providers who feel uncomfortable with any concept list that attempts to prioritize concepts for informed consent. The recent US-based 21st Century Cures Act Information Blocking Rule  with more than 5 years of experience obtaining consent for genetic testing, as well as bioethics researchers who research and write about the topic. While our demographics suggest that we obtained a range of opinions from people who practice in different clinical areas, different geographic parts of the USA, and with different amounts of experience, it is possible that the views of these experts do not represent the broader professional attitudes (particularly those of non-genetics providers), or even the attitudes that patients and families would have towards the same concepts.Our methods had several limitations that are worth considering. First, we purposefully started with a long list of potential concepts that were extracted from the literature about both clinical- and research-based genetic testing consents. Both the fact that we had a long and somewhat granular list, and the fact that it included some concepts that were more focused on research genetic testing may have distracted the expert respondents from some concepts that may otherwise have ‘risen to the top’ as highly important. Given that we did not provide specific definitions for each concept, it is also possible that our experts interpreted concepts (for example “test limitations”) in different ways. Finally, we opted to take the final list of concepts back to the CADRe workgroup for a consensus discussion after two rounds of the survey process. It is possible that our determinations would have been different had we held the final consensus discussions with the experts who engaged in the survey process.Our study showed that it is possible to reach expert consensus on a minimal list of concepts that are critical to facilitate informed consent in patients considering genetic testing, and we anticipate the medical genetics community will robustly debate whether these concepts are the ‘right’ concepts. Future research directions could include considering whether this concept list could also be applied to somatic genetic testing, pharmacogenomic testing, and direct-to-consumer testing, and also if it could be applied in a research genetic testing context.Our next steps include determining if a broader sample of medical geneticists and genetic counselors agree that these concepts would allow patients to make informed decisions in a variety of clinical scenarios and indications, and assessing whether these views of critical concepts are supported by patients and families who have previously undergone genetic testing across a broad range of conditions.


Scorpidium cossonii was higher and the genetic variation among sites was smaller in disturbed than late‐successional rich fens. Sex ratio was balanced in S. cossonii and Campylium stellatum in disturbed fens and skewed in C. stellatum in late‐successional fens, thus supporting H I. In line with our prediction, sex expression incidence was higher in, and sporophytes were confined to, late‐succession compared to disturbed rich fens. Late‐successional S. cossonii sites had more within‐site patches with two or more genotypes, and both species displayed higher sex expression levels in late‐successional than in disturbed sites. We conclude that diverse genotypes and both sexes disperse efficiently to, and successfully colonize new sites, while patterns of genetic variation and sexual reproduction in late‐successional rich fens are gradually shaped by local conditions and interactions over extended time periods.To understand colonization processes, it is critical to fully assess the role of dispersal in shaping biogeographical patterns at the gene, individual, population, and community levels. We test two alternative hypotheses (H I and H II) for the colonization of disturbed sites by clonal plants, by analyzing intraspecific genetic variation in one and reproductive traits in two typical fen mosses with separate sexes and intermittent spore dispersal, comparing disturbed, early‐succession (limed) fens and late‐successional rich fens. H I suggests initial colonization of disturbed sites by diverse genotypes of which fewer remain in late‐successional fens and an initially balanced sex ratio that develops into a possibly skewed population sex ratio. H II suggests initial colonization by few genotypes and gradual accumulation of additional genotypes and an initially skewed sex ratio that alters into the species‐specific sex ratio, during succession. Under both scenarios, we expect enhanced sexual reproduction in late‐successional fens due to resource gains and decreased intermate distances when clones expand. We show that the intraspecific genetic diversity, assessed by two molecular markers, in  We exploit a unique system of recently and large‐scale anthropogenically disturbed habitats and natural rich fens to test two alternative models for recruitment of clonal plants in disturbed sites. Patterns of intraspecific genetic diversity in Scorpidium cossonii (A) and reproductive features in this species and Campylium stellatum (B) show that initial colonization of disturbed sites is by diverse genotypes and a few attain dominance in late‐successional fens and that an initially balanced sex ratio develops into a possibly skewed population sex ratio. We conclude that the genetic variation and sexual reproduction in late‐successional rich fens are gradually shaped by local conditions and interactions over extended time periods. We selected S. cossonii, and sex expression and sex ratio in S. cossonii and C. stellatum, in disturbed limed and natural rich fens, representing early‐ and late‐successional environments. We ask whether within‐ and between‐site patterns of intraspecific genetic variation, sex ratios, and frequencies of sex expression in the study species agree with either of the hypotheses for recruitment of clonal plants in disturbed sites.We test two alternative models for recruitment of clonal plants in disturbed sites  and exhibits a balanced phenotypic sex ratio in natural vegetation . Since the proportion of populations with reproductive structures is higher in C. stellatum than in S. cossonii, it likely produces sporophytes more frequently than S. cossonii.Hedenäs, , 2019. BHedenäs, . They eiHedenäs, . BryophyHedenäs, . NeverthHedenäs, . In this2.2Sphagnum spp. and transfers the mires, at least temporarily, into new artificial habitats that resemble natural rich fens in terms of water chemistry and wetness, thereby creating substrates suitable for colonization by rich fen species. In a postglacial perspective, these anthropogenically disturbed habitats became available for colonization by rich fen species very recently. Natural rich fens are sparse in the immediate areas of liming because base‐rich rocks and soil are rare in the region  belonged to Bryum pseudotriquetrum (Hedw.) G. Gaertn., B. Mey. & Scherb. s.l. .We randomly picked ten individual shoots from the material from each patch to score the presence of male or female sexual structures (sex expression). Out of the 10 2.4rpl16 sequences for S. cossonii as described by Hedenäs  and the plastid  Hedenäs , except http://www.phyde.de/index.html; accessed 23 March 2020). We aligned the assembled sequences manually in PhyDE®. We identified regions of partially incomplete data in the beginning and end of the sequences and excluded them from subsequent analyses. Gaps were coded using the simple indel coding of Simmons and Ochoterena , effective number of haplotypes (Ne), haplotype diversity (H)  , and values of 1 indicate only unique haplotypes and lack of clonality  and was normally distributed by checking that no outliers were present among the residuals in a preliminary ANOVA. We calculated product–moment correlations among the four diversity parameters. Further, we examined whether haplotype numbers per patch (within the sites), as a proxy for the number of clones, differed between fen types with a nonparametric Mann–Whitney U test; patch‐level data did not meet the criterion of homogeneity of variance. Patches were pooled across sites per fen type.For the analyses of genetic diversity and variation, we consider “site” as sampling unit, from where 15 shoots each were analyzed. The null hypothesis in the following analyses is that there exist no genetic differences between the moss populations from different study sites. We revealed the existing haplotypes based on ITS plus D test of selective neutrality using Arlequin , where the moss population showed signs of possible increase. For limed fens, it indicates that the initial colonization phase is completed, that is, the population is not further expanding, while populations in the natural rich fens are currently in equilibrium with their habitat.To be able to correctly interpret potential differences between the two fen types, we investigated whether the fen moss populations are currently stable or not with Tajima's PT . Genetic variation between populations from different sites based on pairwise ΦPT values and average numbers of pairwise nucleotide differences were visualized with principal coordinate analyses . To explore potential isolation‐by‐distance effects, which may obscure habitat effects, we evaluated the relationship between interpopulation ΦPT, based on ITS + rpl16 haplotypes, and interpopulation distance by Mantel tests (GENALEX).We investigated patterns of haplotype variation, that is, by partitioning the total variation into the relative amounts within sites, between sites, and between fen types, by analyses of molecular variance . We run AMOVAs for the limed and natural rich fen data separately and for the total data set. We computed average number of pairwise nucleotide differences between populations  for sexual structures and categorized those bearing perigonia as phenotypically male (M), those with perichaetia, and if fertilized with sporophytes, as phenotypically female (F), and shoots without sex structures as nonreproductive (NR). Perigonia and perichaetia in pleurocarpous mosses are reduced branches with specialized leaves where male and female gametangia, respectively, are formed and which eventually produce spermatozoids and egg cells. F plus M are termed reproductive shoots. We calculated sex expression (SE) as the proportion of shoots with sexual structures  out of the 10 shoots per patch. We calculated sex ratio (SR) as the number of male shoots divided with total number of shoots with sex organs, that is, a SR > 0.5 indicates male dominance, while SR < 0.5 denotes a female bias. We summed up the occurrence of spore capsules for each species per fen type. We computed mean SE ± standard error and mean SR ± standard error for both natural rich and limed fens for each species across sites, after pooling shoot number per reproductive category  from five patches per site. We tested whether mean SR per species and fen typed differed from an unbiased SR (0.5) with Pearson's chi‐squared tests. To examine differences in SE and SR between fen type, we used generalized mixed effects models with binomial error distributions and logit link functions. We included fen type as a fixed factor and site as random factor. Lastly, we tested whether number of patches without reproductive shoots, number of unisexual patches, and number of bisexual patches differed between fen type (rich fen or limed fen) for both species using Pearson's chi‐squared tests. Generalized linear models were conducted in the R 4.0.2 environment , was higher in limed than in rich fens  or both fen types, and 16 were restricted to a single fen  was found within sites and little was referable to between fen types . In natural rich fens, S. cossonii exhibited an even sex ratio , while it was skewed towards males in C. stellatum  . However, it was only borderline so (p = .066) when applying a generalized mixed effect model that takes within‐site variation into account . Despite considerable variation in sex expression within sites, the number of patches without, relative to patches with, sex expression was higher in the limed than in the natural rich fens in both species . Moreover, the number of patches carrying both sexes was higher in natural rich than in limed habitats in C. stellatum . Sample patches carrying sporophytes were rare in both species, and they were all found in natural habitats .Sex expression at the site level spans from 0% to 54% (limed) and 0% to 100%  in i Figure . Sex expl Figure . Despit4S. cossonii at the site level, other than haplotype numbers, were higher in limed than in natural rich fens. The limed sites were more similar to each other in genetic composition than were natural rich fens. Sex expression levels in both S. cossonii and C. stellatum were higher in the natural rich fens than in the limed fens; sex ratios were even in both species at the limed sites and skewed in C. stellatum in the natural rich fens. We infer that site‐level genetic diversity and possibly the frequency of one sex decrease with progressing succession, while genetic differentiation among sites increases and more plants reach sexual maturity.We compared genetic diversity and reproductive traits of wetland mosses in a unique system of anthropogenic early‐successional rich fen habitats, recently created by liming, with those in late‐successional natural rich fens. Our findings support hypothesis I for recruitment of clonal plants . We therefore infer that the population expansion in the disturbed fens was terminated after at most 30 years since the initial liming events. Further, we suggest that the development between the limed fen environments and the late‐successional natural rich fens, that are several 1000 years old ; Data curation (lead); Formal analysis ; Funding acquisition ; Investigation ; Methodology ; Visualization ; Writing‐original draft . Kristoffer Hylander: Conceptualization ; Formal analysis (supporting); Funding acquisition ; Investigation (supporting); Methodology ; Writing‐original draft (supporting). Niklas Lönnell: Conceptualization (supporting); Investigation (supporting); Methodology (supporting); Resources (lead); Writing‐original draft (supporting). Irene Bisang: Conceptualization ; Formal analysis ; Funding acquisition (supporting); Investigation ; Methodology ; Visualization ; Writing‐original draft .Supplementary MaterialClick here for additional data file.


Phaseolus vulgaris L.) is an economically important food legume for direct human consumption worldwide, common bean production in Croatia is based almost exclusively on landraces and there is no common bean breeding program. Information on phaseolin type and results of population structure and genetic diversity obtained by analysis of SSR and SNP markers, in combination with the morphological characterization of 174 accessions of 10 common bean landraces (morphotypes), enabled thorough classification of accessions. The accessions were classified into phaseolin type H1 (“S”) of Mesoamerican origin and phaseolin types H2 (“H” or “C”) and H3 (“T”) of Andean origin. By applying distance- and model-based clustering methods to SSR markers, the accessions were classified into two clusters at K = 2 separating the accessions according to the centres of origin, while at K = 3, the accessions of Andean origin were further classified into two clusters of accessions that differed in phaseolin type (H2 and H3). Using SNP markers, model-based analysis of population structure was performed, the results of which were consistent with those of SSR markers. In addition, 122 accessions were assigned to 14 newly formed true-type morphogenetic groups derived from three different domestication events: (1) Mesoamerican (H1A) , (2) Andean—indeterminate type (H2B1)  and (3) Andean—determinate type (H3B2) . The rest of the accessions could represent putative hybrids between morphogenetic groups. The differences between the true-type groups of accessions were further analysed based on nine quantitative traits, and the subsets of traits that best distinguish among centres of origin  and genetic groups  were proposed.Landraces represent valuable genetic resources for breeding programmes to produce high-yielding varieties adapted to stressful environmental conditions. Although the common bean ( Landraces are characterized by specific adaptation to the environmental conditions of the area of cultivation ,2 and caPhaseolus vulgaris L.) is an economically significant vegetable crop and food legume for direct human consumption worldwide . Analysis of variance (ANOVA) was conducted based on nine quantitative morphological traits between two groups of accessions classified according to centres of origin (A: Mesoamerican group and B: Andean group) as well as among three genetic groups: H1A  and H2B1  and H2B3  . Six outp < 0.001). PC2 primarily separated the true-type accessions of H1A  from the true-type accessions of groups H2B1 and H3B2, based on the traits seed width (T4), seed height (T5), and 100 seed weight (T6). The off-type accessions  were positioned without any clear distribution pattern among the true-types.Principal component analysis (PCA) A based oThe same plot showing the classification of the true-type accessions into 14 morphogenetic groups B revealeFour out of nine quantitative morphological traits  were selected as the best discriminating factors between genetic groups A (Mesoamerican) and B (Andean) by stepwise discriminant analysis. The discriminant function based on four quantitative traits showed a classification success of 95.90% after cross-validation indicating its usefulness in genetic groups discrimination.Using stepwise discriminant analysis, seven quantitative morphological traits  were selected as the best discriminant factors between the Andean genetic groups B1 and B2. The discriminant function based on these quantitative traits showed a classification success of 92.62% after cross-validation, indicating its usefulness in discrimination of genetic groups.The ordination diagram  showed cAll analysed common bean accessions showed a unique multilocus genotype using 26 SSR markers. Analysis of genetic diversity using SSR markers shows that Croatian common bean landraces have a high degree of genetic diversity with 135 alleles detected in 174 accessions with an average of 5.192 alleles per locus.After analysis of the genetic structure using both SSRs and SNPs and phaseolin type, the accessions were almost completely grouped into two main groups, Mesoamerican (A) and Andean (B), and further analysis separated the Andean into two subgroups (B1 and B2). These results confirm that Croatian common bean landraces originate from both centres of origin, Mesoamerican, and Andean, which agrees with numerous studies done in Europe ,33,34,35It was found that 24.71% of the accessions are of Mesoamerican origin  and 45.40% are of Andean origin , while 29.89% of the accessions were classified as off-types and can be considered as hybrids between different groups. Moreover, the Andean group is divided into two subgroups and it was found that 10.92% of the accessions belong to phaseolin type H2 (“H” or “C” type of phaseolin), while most of the accessions 34.48%) belong to phaseolin type H3 (“T” phaseolin type). The results are in agreement with other studies that found that common bean of Andean origin is 66–76% predominant in Europe ,44,45,46.48% beloAnalysis of the genetic diversity of the two groups using both SSR and SNP markers revealed that the allelic richness was higher in the Andean (B) group than in the Mesoamerican (A) group, but when the Andean group was further divided into two groups (H2B1 and H3B2), the Mesoamerican group (H1A) had the highest allelic richness. Similar results were obtained by Leitão et al.  based onThe results show that the expected heterozygosity is larger in the Andean group when SSRs are used and in the Mesoamerican group when SNPs are used. The contrasting results could be due to the nature of the markers used as SSRs are usually more polymorphic between populations that have recently diverged in comparison to SNPs . In thisWhen accessions were collected for this study, the vernacular names of the landraces were recorded as assigned by farmers and the most frequent was chosen to identify them. However, we have observed that the landraces often included accessions with slightly different seed coat colour and pattern but similar enough to be considered as belonging to the same landrace. After genetic analysis based on SSR and SNP markers and phaseolin type determination, 122 accessions (70.12%) with cluster membership higher than 75% (Q >75%) in both SSR and SNP analysis and phaseolin type agreement were categorized as true-type accessions . The rest of the accessions were classified as off-type , which can be considered as hybrids between different true-type groups.Based on genetic analysis of SSR and SNP markers and phaseolin type determination, 174 accessions initially grouped into 10 morphotypes (landraces), were classified into 14 morphogenetic groups. Landraces “Trešnjevac”, “Puter” and “Dan noć” were further separated based on genetic data and growth habit. That is, Andean group B was further separated into H2B1  and H3B2 , from which it can be seen that the genetic groups correspond to growth habit. Based on genetic analysis of SSR and SNP markers and determination of phaseolin type, 122 accessions were assigned to 14 newly formed true-type morphogenetic groups derived from three different domestication events: (1) Mesoamerican (H1A) , (2) Andean—indeterminate type (H2B1)  and (3) Andean—determinate type (H3B2) .The genetic and morphological data collected can therefore contribute to an accurate classification that will allow cultivars to be registered as ‘conservation varieties’ in the List of Varieties of the Republic of Croatia . That caAn analysis of Croatian common bean landraces based on quantitative traits revealed a high degree of morphological diversity of morphogenetic groups. That is, principal component analysis and discriminant analysis divided genetic groups H1A , H2B1 , and H3B2  based on quantitative traits.In congruence with results of SSRs, SNPs, and phaseolin type the PCA using quantitative traits confirmed the existence of three distinct groups. The true-type accessions of the H2B1 group  are separated from the true-type accessions of the H3B2 group  based on the traits seed length, seed flatness, and flatness index. The true-type accessions of H1A  were separated from the true-type accessions of groups H2B1 and H3B2, based on the traits seed width, seed height, and 100 seed weight. These results are in agreement with previous studies that found that seeds of Mesoamerican origin differ morphologically from those of Andean origin, i.e., landraces of Andean origin have larger seeds than those of Mesoamerican origin . It was Four of nine quantitative morphological traits  were revealed as the best discriminating factors between genetic groups A (Mesoamerican) and B (Andean) using discriminant analysis. Discriminant analysis was ultimately applied to the whole dataset and showed a clear separation between the groups of true-type accessions. Seed height, seed width, and seed elongation were the three morphological traits of five in an analysis of Portuguese common bean landraces , which wIn contradiction to the general conclusion that the Mesoamerican centre of origin is represented by smaller seeds than the Andean centre of origin , in the It was hypothesized that among the three Mesoamerican races , the small-seeded beans of the “Mesoamerica“ race were either not brought to Europe or were not preferred by farmers . MoreoveThe results show that the Croatian common bean landraces can be classified into morphogenetic groups according to their origin and growth habit: H1A , H2B1 , H3B2 . The almost complete correspondence of classifications based on phaseolin analysis and cluster membership based on SSR and SNP markers, as well as the strong association between group membership and growth habit, could be explained by a series of bottlenecks during the introduction of the common bean into Portugal and Spain, and eastward expansion throughout Europe .The divergence between the Mesoamerican and Andean genetic resources of common bean preceded domestication, which occurred independently in the two geographic regions . The genZea mays L.) and common bean with indeterminate habit were domesticated in different regions of Mesoamerica and later reunited in a single system as a combined crop with pumpkin (Cucurbita spp.) (milpa cropping system) which have formed the basis of traditional agriculture of Central America [Wild common bean has an indeterminate growth habit, and it was thought that the first types, domesticated in the Mesoamerican centre of origin, were also indeterminate ,61. It wIn the Andean region, the determinate types of common beans were domesticated first as no natural support for the growth of indeterminate common beans was available being root crops, lupins and quinoa domesticated earlier . HoweverAlthough most accessions of the determinate habit originate from the Andean centre of origin, the determinate habit is also present in the Mesoamerican region. Kwak et al.  have shoThe research was carried out on 174 accessions of common bean landraces originating from diverse geographical regions of Croatia. The accessions belonging to 10 morphotypes (landraces) which are known by their traditional names based mostly on seed coat colour and pattern: M01 (“Kukuruzar”), M02 (“Tetovac”), M03 (“Biser”), M04 (“Sivi”), M05 (“Sivi prošarani”), M06 (“Trešnjevac”), M07 (“Puter”), M08 (“Dan noć”) M09 (“Zelenčec”), and M10 (“Bijeli”).Amplification of phaseolin sequences  was usedTwenty-six primers developed by Blair et al.  Córdoba https://www.diversityarrays.com/, accessed on 20 March 2021). The quality of DArTseq-derived SNP markers was determined by several parameters, including the percentage of technical replicate pairs scored identically for a given marker (reproducibility), the percentage of samples scored for a given marker , minor allele frequency (MAF), and the percentage of heterozygotes (heterozygosity). Alignment of the marker sequences against the reference genome of Phaseolus vulgaris [DArTseq analysis was performed by Diversity Arrays Technology Pty Ltd., Bruce, Australia  [Data on the morphological and agronomic traits of Croatian common bean landraces were obtained based on nine quantitative morphological characteristics based on the list of descriptors for ch 2021) . QuantitaN) and polymorphic information content (PIC) in Cervus v3.0 [OH), and gene diversity  in GENEPOP v4.0 [For each microsatellite locus we calculated the average number of alleles per locus  were performed on the Isabella computer cluster at the University Computing Centre (SRCE), University of Zagreb, Croatia consisting of a burn-in period of 20,000 steps and followed by 1,000,000 MCMC replicates assuming an admixture model and correlated allele frequencies. We calculated ΔK values [K to choose the most likely number of clusters (K) as implemented in STRUCTURE HARVESTER v0.6.94 [The genetic structure was inferred using STRUCTURE v2.3.4 . Thirty K values  for each v0.6.94 . Runs we v0.6.94 .OH), and gene diversity  in GENEPOP. Population structure (using STRUCTURE) was analysed in the same manner as in the case of the SSR data.SNP diversity was assessed by calculating observed heterozygosity (Data on landrace (morphotype) membership, growth habit, phaseolin type, and cluster membership based on both SSR and SNP markers of each accession were used to classify the accessions into three true-type genetic groups  and three off-type groups . True-type accessions had a percentage of cluster membership (Q in STRUCTURE) higher than 75% in both SSR and SNP analyses and phaseolin type matching the cluster membership. Off-type accessions were categorized as: (1) X1—admixed: Q < 75% in either SSR or SNP analysis, (2) X2—genetically non-corresponding: no correspondence between phaseolin type and SSR/SNP cluster membership, and (3) X3—non-corresponding: no correspondence between morphotype membership and genetic cluster membership . Further analyses were performed exclusively on true-type accessions classified according to (1) centres of origin , (2) genetic groups , and (3) morphogenetic groups.aN), allelic richness (arN), number of private alleles (prN), and private allelic richness (parN) in HP-RARE v1.0 [OH), and gene diversity (EH). The values of allelic richness (Nar), observed heterozygosity (OH), and gene diversity (EH) were compared between Mesoamerican (H1A) and Andean (H2B1 and H3B2) groups of accession as well as among the three genetic groups (H1A vs. H2B1 vs. H3B2) by the analysis of variance carried out using PROC GLM in SAS in SAS v9.3 [Microsatellite diversity of accessions was assessed by calculating the average number of alleles   using ARTo graphically represent genetic relationships among individual genotypes, a factorial correspondence analysis (FCA) was carried out using GENETIX v4.05 .The analysis of genetic diversity, AMOVA, and FCA based on SNP marker data were carried out as previously described for SSR markers.The analysis of variance (ANOVA) was conducted to test the mean differences between Mesoamerican (A) and Andean (B) groups of accession as well as among the three genetic groups (H1A vs. H2B1 vs. H3B2) in nine morphological traits using PROC GLM in SAS, followed by Tukey’s studentized range test.Pearson’s correlation coefficients were calculated among the seven morphological traits using PROC CORR and principal component analysis (PCA) was carried using PROC PRINCOMP from SAS.Discriminant analysis (DA) was performed to determine which of the seven morphological traits were the most useful for maximum discrimination between Mesoamerican (H1A) and Andean (H2B1 and H3B2) groups of accession as well as among the three genetic groups (H1A vs. H2B1 vs. H3B2) using PROC STEPDISC, DISCRIM, and CANDISC in SAS. Thus, obtained discriminant function was finally applied to the total data set (including off-type accessions).


We review some of the current insights derived from the analyses of new large-scale, genome-wide autosomal variation data studies incorporating Ethiopians. Consistent with their substantial degree of cultural and linguistic diversity, genetic diversity among Ethiopians is higher than that seen across much larger geographic regions worldwide. This genetic variation is associated in part with ethnic identity, geography and linguistic classification. Numerous and varied admixture events have been inferred in Ethiopian groups, for example, involving sources related to present-day groups in West Eurasia and North Africa, with inferred dates spanning a few hundred to more than 4500 years ago. These disparate inferred ancestry patterns are correlated in part with groups’ broad linguistic classifications, though with some notable exceptions. While deciphering these complex genetic signals remains challenging with available data, these studies and other projects focused on resolving competing hypotheses on the origins of specific ethnolinguistic groups demonstrate how genetic analyses can complement findings from anthropological and linguistic studies on Ethiopians. Australopithecus afarensis specimen named Lucy, with 40% of her osteological remains recovered  data. We provide a current snapshot of the complex Ethiopian genetic landscape, including a description of features associated with this genetic structure, signatures of admixture and the ancestral histories of various groups. We also highlight a few of the many possible future directions in genetic studies involving Ethiopians.Genetic data from several recent studies using both whole-genome sequencing (WGS) and genotyping arrays now allow joint analyses of genome-wide SNP data from >1000 Ethiopians representing 68 ethnolinguistic groups . Here, wConsistent with their cultural and linguistic diversity, Ethiopians display a high degree of genetic heterogeneity relative to non-Africans. For example, we used hap-ibd , with deFST, calculated here using HIERFSTAT  and Nilo-Saharan (NS) . Severalm SNPs targeted to represent genetic diversity in multiple African populations  genome published from an African was that of a ≈4500-year-old individual found in Mota Cave in the Gamo highlands of southwest Ethiopia . This ‘Met al.  infer thet al. . They noet al. .These complex, disparate admixture histories can at least partially explain the differences in ancestry patterns observed among Ethiopian ethnic groups. As an illustration, we used CHROMOPAINTER  and SOURet al.  people that practice farming often have limited interactions with Ari who practice artisanal activities such as blacksmithing, pottery and tanning ,38. In gFST . While the Chabu appear genetically distinguishable from other neighboring Ethiopian groups, three separate studies using different data and applying various methods, including those mentioned in the previous section, have shown that this is in part owing to a relatively high degree of recent isolation , who are genetically more similar to certain AA-speaking groups than to NS speakers ; the National Institute for Health Research University College London Hospitals Biomedical Research Centre; the Natural Environment Research Council (NE/L002485/1 to N.B.).