Mental imagery has long been part of cognitive behavioural therapies. More recently, a resurgence of interest has emerged for prospective mental imagery, i.e. future-directed imagery-based thought, and its relation to reward processing, motivation and behaviour in the context of depression.We conducted a selective review on the role of prospective mental imagery and its impact on reward processing and reward-motivated behaviour in depression.Based on the current literature, we propose a conceptual mechanistic model of prospective mental imagery. Prospective mental imagery of engaging in positive activities can increase reward anticipation and reward motivation, which can transfer to increased engagement in reward-motivated behaviour and more experiences of reward, thereby decreasing depressive symptoms. We suggest directions for future research using multimodal assessments to measure the impact of prospective mental imagery from its basic functioning in the lab to real-world and clinical implementation.Prospective mental imagery has the potential to improve treatment for depression where the aim is to increase reward-motivated behaviours. Future research should investigate how exactly and for whom prospective mental imagery works. This review provides a selected update of the literature on prospective mental imagery.Prospective mental imagery might decrease depression via reward processing and reward-motivated behaviours.Suggestions for future research to investigate these hypotheses are provided. According to Beck’s cognitive model, individuals with depression hold negative views about the self, others and the future . In addiMajor Depressive Disorder (MDD) is characterized by low mood and/or the loss of interest in previously rewarding or enjoyable activities as well as a number of other emotional, cognitive and physical symptoms . MDD is By drawing on shared brain structures and functions , vivid mRecently, a number of studies have emerged that tested the impact of prospective mental imagery of positive events or activities on reward anticipation and reward-motivated behaviour. These studies have the common aim of investigating new ways of promoting positive experiences, in line with recent calls for treatment innovation in depression to focus on positive affect systems . StudiesSeveral studies have investigated the impact of mental imagery on reward anticipation. In a case-series, Other studies have focussed on the effects of mental imagery on approach motivation. For example, Finally, a number of studies have investigated the transfer of the motivating effect of mental imagery interventions to self-reported activity levels outside the lab. One study conducted a secondary analysis of a randomized controlled trial  to test The studies reviewed here suggest that positive prospective mental imagery of activities can facilitate reward anticipation, reward motivation and reward-motivated behaviour. This is clinically relevant given that reward anticipation deficits are not adequately addressed in current treatments of depression . Drawingmotivational amplifier effect by facilitating reward anticipation, reward motivation and reward-motivated behaviour. Given the power of mental imagery to amplify emotions (emotions , it has emotions . Anticipemotions . This tremotions . Actual motivational amplifier boosting behavioural activation and thereby alleviating depressive symptoms (The conceptual model has clinical potential insofar as it illustrates how positive prospective mental imagery could be used to promote behavioural activation in depression. The central assumption here is that reduced reward anticipation in depression contributes to a downward-spiral of reduced reward-motivated behaviours due to a loss of interest in previously rewarding activities that reduces the experience of rewards in daily life and, consequently, worsens depressive symptoms such as low mood . Based osymptoms .In summary, the recent literature reviewed above supports the idea that positive prospective mental imagery of activities can facilitate reward anticipation, reward motivation and reward-motivated behaviour. However, the reviewed studies primarily relied on self-report and more work is needed to investigate how the transfer of imagery to behaviour beyond the laboratory can be facilitated and how prospective mental imagery might benefit clinical practice.Recent literature has emphasised the importance of conducting multimodal research to understand and thereby improve clinical interventions . A futurAnother important question in experimental psychopathology research is how lab-based findings hold up under everyday circumstances. Recent research in the broader field of clinical psychology demonstrated the added value of combining laboratory experiments with Ecological Momentary Assessment (EMA; e.g. Moreover, individuals may differ in the extent to which they benefit from prospective mental imagery interventions. Studies already pointed to individual variation in processes related to prospective mental imagery, such as anticipatory pleasure  and the Imagery based interventions have been used as stand-alone interventions as well as part of regular CBT for depression . So far,In this review, we provided a selected update of the recent scientific literature on prospective mental imagery and its impact on reward processing and reward-motivated behaviour in depression. Overall, the studies presented here suggest that prospective mental imagery simulations of activities can increase reward processing related to these activities as well as reward motivation and reward-motivated behaviors. Thus, these initial studies suggest that prospective mental imagery is a promising experimental intervention in the context of depression, where the aim is to increase engagement in potentially rewarding activities. Future directions for research in this area may focus on multimodal assessments of prospective mental imagery effects to gain a better understanding of the processes involved, from basic mechanisms to everyday situations and its clinical applications.


Jasus edwardsii) recruits, known as pueruli, along a latitudinal and temporal gradient on the east coast of Tasmania, Australia. We further explored whether natural selection could be driving morphological variation. We used double digest restriction site‐associated DNA sequencing (ddRADseq) to assess differences in the genetic structure of recently settled recruits on the east coast of Tasmania over 3 months of peak settlement during 2012 (August–October). Phenotypic differences in pueruli between sites and months of settlement were observed, with significantly smaller individuals found at the northernmost site. Also, there was a lack of overall genetic divergence; however, significant differences in pairwise FST values between settlement months were observed at the southernmost study site, located at an area of confluence of ocean currents. Specifically, individuals settling into the southernmost earlier in the season were genetically different from those settling later. The lack of overall genetic divergence in the presence of phenotypic variation indicates that larval environmental history during the dispersal of J. edwardsii could be a possible driver of the resulting phenotype of settlers.Environmental conditions experienced during the larval dispersal of marine organisms can determine the size‐at‐settlement of recruits. It is, therefore, not uncommon that larvae undergoing different dispersal histories would exhibit phenotypic variability at recruitment. Here, we investigated morphological differences in recently settled southern rock lobster ( Jasus edwardsii settlers exhibit phenotypic variability across a geographical and temporal gradient; however, this is not correlated with genetic divergence. Given the protracted pelagic larval duration of the species, we hypothesize that dispersal history drives phenotypic variability in this species.We found that  The present study explored patterns of morphological and genetic differences in recently settled 22.1Lobster pueruli were caught in crevice collectors . To determine whether pueruli weight varied with settlement month, site, and the interaction of both, a two‐way analysis of variance (ANOVA) was performed using the function “aov” implemented within the R package stats v4.2.1  following the manufacturer's instructions. DNA concentration was quantified on a Qubit® 2.0 Fluorometer (Life Technologies). DNA integrity was determined through gel electrophoresis to verify that high molecular weight DNA was obtained in all samples (>1000 base pairs [bp]).Four ddRADseq libraries were prepared using a modified version of Peterson et al.  ddRAD pr2.4“process_radtags” protocol from Stacks v1.29  to obtain a catalog of reference loci detailed within Villacorta‐Rath et al. (https://github.com/torognes/vsearch). Initially, all reads were pooled and clusters with a depth of at least 193 were retained, assuming that there would be at least one read per sample represented in each cluster. Subsequently, all clusters that were comprised of reads with more than a 4% mismatch (3 bp) were discarded. Assuming that each member of a cluster was an allele, only clusters that had between two and 16 members were retained. Finally, only samples with a minimum of 10,000 alleles and 30% missing data were retained for subsequent read alignment and SNP calling.Filtered reads were processed through a custom‐built “rad‐loci” pipeline , to remove SNPs that were potentially in linkage disequilibrium (−‐min‐r2 0.2), to discard SNPs with a minor allele frequency (MAF) of less than 5% (−‐maf 0.05), and to ensure that the minimum SNP depth was 5 (−‐minDP 5). The maximum amount of missing data for each locus was set to 25% (−‐max‐missing 0.75) and individuals with more than 25% missing data were removed from subsequent analyses. Finally, only one SNP per locus (−‐thin 75) was retained.A total of 22 technical replicates remained after the removal of individuals with more than the missing data threshold. A Principal Component Analysis (PCA) was performed to visualize the spatial distribution of replicated samples using the function "dudi.pca" of the R package ade4 v1.7‐19 , the minimum heterozygosity required before including calculations from a locus was set to 0.1 (Hmin = 0.1), and the false discovery rate was 0.1 (qthreshold = 0.1). Only SNPs characterized as being under putative positive selection by both software packages were retained for downstream analyses.Two outlier identification methods were used in order to minimize false positives in the panel of SNPs under putative positive selection. The first SNP characterization was performed in the software Lositan   using BLAST+ v2.2.29. Queries with statistically significant e‐values (E < 10−6) and more than 84% identity were considered as valid alignments. Transcriptome sequences that provided significant alignments were annotated using the Trinotate pipeline (https://trinotate.github.io/) to determine whether they aligned with any known protein domain.To determine whether the loci containing SNPs putatively under positive selection were contained in protein‐coding regions, sequences were BLASTed against the complete 2.7ST values and confidence intervals for each SNP panel were estimated using the R package mmod v1.3.2  . This metric represents the proportion of heterozygous loci over the mean heterozygosity across all markers, so that heterozygosity of all individuals is measured on the same scale  was estimated using Mantel tests between temporal and genetic distance using 9999 permutations within the R package ade4 v1.7‐19 . Pairwise FST values and AMOVA were not calculated for this SNP panel, since it was only comprised of two loci. Transcriptome sequences from the eyestalk, optical nerve, green gland, and hepatopancreas of J. edwardsii exhibited significant hits with one locus containing SNPs under putative positive selection. However, the transcriptome sequences could not be successfully annotated to any gene, possibly due to the short fragment size . Additionally, no significant hits between the panel of SNPs under putative positive selection and the H. americanus transcriptome were found.On the other hand, the SNPs under putative positive selection exhibited a stronger overall level of genetic differentiation , indicating the existence of a very low correlation. Pueruli settling into Recherche Bay during the same month was genetically more similar to each other than individuals settling one or two months apart, as shown by the slight but significant regression  was genetically divergent from the other sites and also exhibited temporal genetic divergence across settling cohorts.4.1Palaemon serratus) larvae exposed to different combinations of temperature, food, and salinity treatments exhibited a longer duration of development (increased number of instars) under suboptimal conditions  different cohorts of phyllosoma metamorphosing into pueruli and settling simultaneously, and (2) to pueruli originating from different parental stocks  varies across months within each sampling site, suggesting that settlers could have undergone different environmental histories. Oceanographic modeling would allow determining whether collective dispersal, larval cohesiveness close to settlement sites, or a combination of both could be driving the observed phenotypic, but no genetic differences were observed in the present study.While larval cohesiveness could be a plausible mechanism of dispersal in Cancer magister  sourcing pueruli during the winter months. The slight differences in individual weights between months found and size‐at‐settlement could be a signature of each of these populations. However, the small sample size of the present study precludes the possibility of proving that the observed genetic divergence at Recherche Bay is due to chaotic genetic patchiness or IBT.5J. edwardsii maintaining cohesiveness and experiencing the same environmental conditions is a likely explanation for the observed phenotypic differences between settlement months and sites in the lack of overall neutral population differentiation. The lack of neutral population differentiation suggests that neither the time nor place of reproduction acts as a biological barrier to the dispersal of J. edwardsii populations on the east coast of Tasmania. A larger panel of markers under putative positive selection and a larger sample size could help determine whether observed phenotypic differences across sites and months were driven by selection in response to environmental factors.The observed variation in size‐at‐settlement of pueruli observed in the present study is likely due to phenotypic variability during dispersal, rather than the genetic signature of the parental stock. The hypothesis of cohorts of larval Cecilia Villacorta‐Rath: Conceptualization ; data curation (lead); formal analysis (lead); investigation (lead); methodology (lead); writing – original draft (lead). Bridget S Green: Conceptualization (lead); funding acquisition (lead); methodology (lead); project administration (lead); resources (lead); writing – review and editing . Caleb Gardner: Conceptualization (supporting); funding acquisition (lead); investigation (supporting); methodology (supporting); project administration (lead). Nicholas P Murphy: Formal analysis (supporting); investigation (supporting); resources ; writing – review and editing . Carla A. Souza: Data curation ; formal analysis (supporting); investigation (supporting). Jan Strugnell: Conceptualization ; formal analysis ; investigation ; methodology ; project administration (lead); resources (lead); writing – review and editing .The authors declare that they have no competing interests.


The Guzerá breed evolved from the introduction of breeds from India, mainly the Kankrej breed, into Brazilian livestock at the end of the 19th century. Guzerá adapted well to the climatic conditions of Brazil, where it is considered a dual-purpose breed and has been used for pasture-based beef, milk or dual-purpose production systems with the use of low-medium inputs. The importance of this genetic resource for milk production in tropical regions moved breeders to implement the National Breeding Program for the Improvement of Guzerá in 1994, based on both progeny testing and MOET nucleus schemes. We sought to evaluate the role of the MOET nucleus scheme in the phenotypic and genetic progress for milk traits in this breed. The initial database used in the present study consisted of 6,513 cows, daughters of 761 bulls. We performed genetic evaluations with different datasets using a linear mixed model in a single trait analysis, including the relationship matrix, in order to estimate breeding values. Inbreeding coefficients were also calculated using the relationship of descent between two parents. Annual phenotypic, genetic and inbreeding trends were obtained for each dataset, considering the genetic pathways of both the bull and the cow. The low genetic progress found for milk yield in the whole population (5.27 ± 0.30 kg/year) partially accounted for the dual-purpose selection goal, despite the higher genetic progress in the MOET nucleus (9.39 ± 0.79 kg/year). The inbreeding coefficient was minimized at the beginning of the breeding program based on the use of new lineages. Posteriorly, it started increasing again from 0.002 in 1991 to 0.008 in 2019. The results provided evidence of the significant contribution of the MOET nucleus scheme for the phenotypic and genetic progress of Guzerá breed for milk traits, as well as of the impact of the breeding program on the inbreeding coefficient rate in the early years. New strategies need to be designed for the Guzerá breed, to allow for greater improvement of milk traits and minimizing the rate of the inbreeding coefficient. Nucleus-breeding schemes present an interesting approach to dairy cattle improvement. These schemes may be effective in organizing the breeding program, particularly when large-scale recording is lacking, for comparing animals under standard conditions . Many auMultiple ovulation and embryo transfer (MOET) allow an increase in family sizes, mainly in collateral relatives, enabling improvement of the nucleus-breeding scheme . The useThe use of MOET also allowed reduction of the generation interval, by making it possible for the mothers of the bulls to produce a greater number of progeny in a shorter period than it would normally take during the animal’s life . FurtherFactors of genetic progress variation in nucleus schemes such as herd size, mating system, sibling family type, selection age, number of sires selected per full-sib group, have also been studied . NicholaSome authors have proposed a nucleus that is continuously open to the introduction of individuals from other herds of known genetic merit . This woZebu dairy herds in Brazil have a long generation interval as a result of both a late age at first calving and a large calving interval related to genetic and environmental circumstances , limitinTropical dairy systems demand specific selection objectives and management geared to regional reality, focusing on adaption to the harsh environment, in order to become sustainable . Regardi2), in partnership with Embrapa-Dairy Cattle, implemented the National Program for the Improvement of Guzerá for Dairy Purposes (PNMGuL). This program is based on both the progeny test and MOET nucleus schemes  or above 37°C in the rainy period (October to March); precipitation between 20 and 209 mm; and, humidity varying between 1 and 86%.in vitro embryo production biotechnology (IVP), the MOET biotechnology was replaced and IVP became the technique of choice for embryo generation, mainly after the success of in vitro fertilization (IVF) in Zebu breeds. However, the original denomination MOET Nucleus was kept.MOET was conducted in a commercial corporation. Following the development of Initially, to constitute the families in each annual battery, bulls and cows with the potential to maximize milk production were chosen based only on their phenotypes for milk and beef production. These phenotypes comprised the following data: lactation records from ancestors, collaterals, descendants and, in the case of cows, the lactation record itself. Subsequently, in the posterior batteries, the choice of parents for the next generation (later batteries) was based mainly on the following criteria: EBVs from the most recent genetic evaluation as well as on genealogy, inbreeding coefficient, and average relationship. Since the beginning, a commission formed by breeders and the Embrapa research team annually evaluates the proposed progeny test (PT) bulls and MOET families, and ranks those to be included in subsequent batteries.Selected donors were sent for assisted reproduction at a private center. There, after an adaptation period, they were synchronized and super-ovulated. Embryos of satisfactory quality were then transferred to recipients (Holstein-Zebu crossbreeds). Initially, after the 2-months positive pregnancy diagnosis, these recipients were sent to the MOET Nucleus (Fazenda Taboquinha), 400 km distant from the embryo-transfer center, to give birth and where calves were raised. Males were sent to the MOET family’s owner after 15 months. Females were inseminated and kept at the Nucleus until the end of their first lactation. Subsequently, there was a change in the logistics at the Nucleus. The recipients started to be sent to the farms of the MOET family’s owners, and the calves were sent to the Nucleus after weaning. There, they were inseminated and kept until the end of their first lactation.®, Sigma-Aldrich, United States). These samples were immediately refrigerated and sent to the Milk Analysis Laboratory at Embrapa Dairy Cattle for milk composition analysis. Data from test-day and milk composition were sent to the database team for computational processing. After processing, all these data were analyzed together with data from database of the Brazilian Association of Zebu Cattle. Joint genetic evaluations are carried out using the animal model methodology for BLUP evaluation, connecting data from all those sources. This procedure makes a significant contribution to the accuracy of breeding value for the animals born at the MOET Nucleus. Some proven bulls from this scheme were submitted to the progeny test in order to obtain additional accuracy. In sire summary, only bulls with minimum accuracy of 0.50 are listed, and few bulls have stayed below 0.60 in the last summary.Milk production recording and individual milk sampling were performed monthly at the MOET Nucleus and in the PT herds. Individual milk samples were collected after each milking during the test-day using tubes containing Bronopol  and bulls for forming MOET families at each battery were variable, depending on the logistic and financial circumstances, oscillating from 4 to 16 donors and 4–13 bulls at each battery. Eventually, some bulls were mated to more than one cow.1- Cows to maximize milk (minimum 1/3 of the vacancies)—cows with PTA (predicted transmitting ability) > 300, choosing among them, preferentially, the 1st within each lineage, followed by those with the highest number of ancestral selection and lastly, those with good conformation and type. If there are no candidates with a PTA >300 available, use the ones with the highest PTA.2- Lineage option cows—low risk—cows with PTA >100 were chosen, prioritizing rarer genetics within the program, followed by milk PTA cows with good conformation and type.3- Risk cows (maximum 1/3 of the vacancies)—cows with innovative, and other, genetic attributes that are important for the program, as determined by the technical committee and breeders. Except in exceptional cases, the cow must have high production measured in a lactation.1- Bulls to maximize milk —bulls chosen from the highest PTA, not yet used in the program, preferably one within each lineage (maximum one within each MOET family), with the highest number of generations of selection, and of better conformation and type of breed.2- Bulls lineage and/or risk option —bulls prioritized for keeping the inbreeding coefficient (inbreeding) at minimum levels and adding favorable aspects in other traits, such as those morphological and growth. Chosen for new genetics or distant parentage, based on the best dairy information available. They must have breed and/or conformational attributes that are important to the breed.Data for the present study were obtained from the National Program for the Improvement of Guzerá Cattle (PNMGuL) database. The complete dataset, including data from MOET Nucleus, was formed: from 6,513 first-lactation records from Guzerá cows, daughters of 761 bulls; the reduced dataset  from 4,482 Guzerá cows; and, 2,031 cows came from MOET Nucleus truncated to 305-days. Therefore, the traits evaluated were the 305-days first-lactation cumulative yields (kg) of milk (MY305), fat (FY305), and protein (PY305). The number of test-day records used for obtaining the cumulative yields were 7.6 ± 1.9 (milk yield), 5.8 ± 2.1 (fat yield), and 5.6 ± 1.9 (protein yield).The complete relationship matrix comprised data from 120,655 animals. The reduced relationship matrix contained 75,378 animals , being 2,144 sires and 18,560 dams from 13 overlapping generations. The MOET animals came from 182 full sib families formed over the years.b and a to The genetic evaluation was performed using the complete data and the reduced data. The following mixed linear model single trait was assumed:Contemporary groups (CGs) were formed by combining herd, year, and season of calving (defined as dry [April–September] or rainy [October to March]). There were 58, 24, and 16 herds for respectively MY305, FY305 and PY305, and 604, 212, and 123 CGs were formed for MY305, FY305, and PY305. Data that belonged to the CG with fewer than three records and with only one sire as a parent were excluded. The age at calving was considered a linear covariate.The genetic evaluation was performed using the complete data and the reduced data using BLUPF90 family programs .Inbreeding coefficients (F) were calculated using the relationship of descent between two parents X and Y; that is, the kinship coefficient  by birth year, considering the different datasets. In the same way, the averages of the inbreeding coefficients were regressed by birth year. Furthermore, EBV averages for cows and bulls were regressed separately by birth year, in order to evaluate the genetic contribution of each pathway for the improvement of the Guzerá breed for milk traits. In all cases, linear regressions were performed.The descriptive statistics are shown in The annual phenotypic and genetic trends for MY305 considering complete data, reduced data and only data from the MOET Nucleus are shown in p-values < 0.001), respectively, for the whole population, the reduced population, and the MOET nucleus. The genetic increases for milk production for the whole population , and the population excluding MOET records , were notably lower than the genetic rate for the MOET Nucleus population . The increases in the female and male pathways for the whole population were similar. However, for the MOET scheme, the female pathway contributed more for genetic progress than did the male pathway. Excluding data from the MOET nucleus, though, the male pathway contributed slightly more to the rate of genetic progress.The annual phenotypic increases for milk yield reveal favorable improvements of 41 ± 11 kg/year , 17 ± 7 p-value < 0.001) and in the population excluding MOET records  were notably lower than the genetic rate in the MOET Nucleus population . The female and male pathways behaved the same as for milk production, with the female pathway higher than the male pathway in the MOET nucleus population.The annual genetic progress for fat production in the whole population  and in the population excluding MOET records  were notably lower than the genetic rate in the MOET Nucleus population  and, in some cases, were even negative. The genetic progress for protein in the population excluding MOET records can be considered null. The female and male pathways behaved the same as for milk production, with the female pathway higher (36%) than the male pathway in the MOET nucleus population.The annual genetic progress for protein production in the whole population , with MOET animals (Fwith_MOET) and without MOET animals (Fwithout_MOET) for (A) all animals, and for (B) males and females, can be seen in Annual inbreeding trends for selected Guzerá animals, considering data of only MOET animals , 7.7% presented an inbreeding coefficient less than 10%, 0.9% of animals presented F between 10 and 20%, and 0.2% presented an inbreeding coefficient greater than 20%. Considering reduced data, 93.7% of animals presented F equal to zero (not shown), 5.2% presented F less than 10%, 0.9% of animals presented F between 10 and 20%, and 0.2% presented F greater than 20%. Considering data containing only MOET animals, 87.2% of animals presented an inbreeding coefficient equal to zero (not shown), 11.8% presented an inbreeding coefficient less than 10%, 0.8% of animals presented an inbreeding coefficient between 10 and 20%, and 0.2% presented an inbreeding coefficient greater than 20%.Although there are many theoretical studies, to our knowledge, there are no practical results on the performance of any other selection nucleus in the literature. This is, therefore, the first study to present such results.As shown in Results from Bos indicus breed, in a low-input and pasture-based production system and under direct selection for milk production in Brazil from 1987 to 2012. This study, although conducted for only one herd, revealed higher genetic and lower phenotypic progress rates as those obtained for the Guzerá breed, and had the same low selection response. However, to our knowledge, there is no recent study about Gir cattle trends for milk traits. As this breed has long been selected for milk yield, these results probably do not reflect the actual phenotypic and genetic progress for the whole population of this breed.Studies conducted in developed countries for other breeds, just to access the genetic possibilities for improvement, have found the highest genetic progress for the milk purpose to be due to directional selection. For the Holstein population in the United States, e.g., higher annual phenotypic (≅+273 kg) and genetic (≅+168 kg) progress in milk production was observed from 1957 to 2020. A continuous and intense selection process focused on milk traits combined with the constant adoption of modern methodologies during this period support this result . For GuzFurthermore, in an attempt to control inbreeding, the influence that the strategy of introducing animals from different lineages or lineages still not used, as well as risk animals about which little was known could have contributed to the timid results for genetic gain for milk production. Joint to the mating planning implemented since the beginning of the nucleus scheme, inbreeding has been an important and constant concern.The results for the fat yield revealed low or even negative annual phenotypic progress, despite the positive genetic progress in the population, both with  and without  data from the MOET nucleus. For this trait, the progress in the MOET scheme had increased phenotypically and genetically by 380 g and 220 g of fat/year, respectively. It seems that these increases could not have contributed in any meaningful way to the improvement of this trait in the population. Two aspects should be taken into account. First, the genetic correlation between milk and fat yield (0.96 ± 0.02) in Guzerá , althougFor the protein yield, the phenotypic (+320 g) and genetic (+70 g) progress was positive in the whole population. Considering the negative annual phenotypic (−160 g) and genetic (−10 g) progress for the population without data for the MOET animals, and considering the notable annual phenotypic (+960 g) and genetic (+160 g) progress in the MOET nucleus, the progress in the whole Guzerá population can be also attributed to the contribution of the indirect selection for protein production in this scheme. This was because the genetic correlation between milk and protein production (0.97) is high . TherefoUnfortunately, other traits important for dairy systems, such as those related to reproduction, growth and development, morphology, and disease occurrence, were not recorded in the MOET nucleus due to financial and logistic circumstances what would have allowed to evaluate the impact on other important traits. Information on the beef traits of the Guzerá breed, for instance, is processed in specific genetic evaluations for beef cattle, carried out by beef improvement programs. However, as the database is genetically connected to the database of the milk program, the joint availability of genetic values for dual-purpose bulls has been allowed . There aIt is important to clarify in this publication that Guzerá breeders who opted for the dual-purpose systems have no interest in specializing their herds, so the model of maximizing genetic gains in specific traits is not their objective.The genetic progress achieved for milk yield, from the selection of female and male pathways in the whole population, was similar. However, in the MOET nucleus, the genetic progress from the female pathway (+11 kg) contributed more than that from male pathway (+6.5 kg) to the genetic progress in this scheme. Excluding data of the MOET nucleus from the whole population, the male pathway contributed slightly more (+1.31 kg) to the rate of genetic progress, although it was quite similar to the female pathway contribution (+1.15 kg). Probably, progeny from test-proven bulls also contributed to this slight difference. In a study with the Gir breed in Brazil, Balieiro et al. (2000) found positive, higher phenotypic and genetic progress in both pathways. The Gir genetic progress was higher in the male pathway, which was the main contribution responsible for the improvement of milk traits in the population. The breeding program with the Gir breed, however, was initiated in 1985, 10 years before the PNMGuL, and since then has focused only on milk production traits.Observing the genetic progress of Holstein or Red and White female and male pathways in USA, it can be noticed that genetic progress came mainly from the progeny of test-proven bulls and recently from the inclusion of genomic data in genetic evaluations, although the contribution of the females is also high and follows the same trend . To our From It is important to consider that the inclusion of risk animal/lineage options in the PT and in the MOET nucleus, which was used as an attempt to minimize the inbreeding coefficient progress in the population, also contributed to the low phenotypic and genetic progress found in the present study. However, it was an important approach, which together with the mating planning carried out in the population as well as in the MOET nucleus scheme allowed control for the increase of F in the first years, thus maintaining this parameter at acceptable levels in the population. Concern about inbreeding led the PNMGuL to continuous monitoring of the genetic contribution of bulls by calculating annually the average relationship coefficient and making available proven bulls of different origins. Breeders are guided breeders to diversify the use of bulls in their herds. The Optimum Contribution Selection is an important strategy to avoid unbalanced use of bulls and decrease the rate of inbreeding .It is also important to stress that the Guzerá breed went through many bottlenecks during its evolution in Brazil, mainly as a result of its intense use in crossbreeding and the formation of synthetic breeds . TherefoIn 2000, the first Guzerá sire summary containing the results of accurate genetic evaluations was made available and producers felt confident in using bulls from other origins, which also contributed to keeping low F levels. The positive impact of the breeding program on the F of the population became evident from 2003 to 2012, when the progeny of proven sires were born. After this period, it started to increase again, maybe due to the use of superior bulls from the same family that will require new control efforts. The intense use of MOET proven bulls, the top of the sire-summary ranking, probably contributed to the increase of F in the population after 2012, as shown in In an interesting review, In conclusion, we can say that the MOET Nucleus scheme has allowed the genetic improvement of Guzerá cattle, genetically lifting the dual-purpose population for milk production in realistic, tropical production systems, and contributing to the economic return of the activity. The low phenotypic progress of the Guzerá population is partially due to the low-medium input production systems. These production systems impair the expression of the genetic potential and limit the possible phenotypic trends, as well as raise concerns about losses of genetic variability due to the increasing inbreeding coefficient. The selection for milk production in MOET nucleus allowed correlated response in fat and protein production, albeit small. The PNMGuL should start phenotyping for other important traits as soon as possible, in view of the prospects of future demands, focusing on the sustainability of the program. The increase in F in recent years will demand an implementation of a mating strategy to minimize the consequences of family selection based on the scheme of PT bulls. The MOET nucleus scheme represents an important tool for the rapid and easy change of production traits, requiring a well-organized partnership.


Labeo ariza), a newly potential aquaculture species in Bangladesh. The improved performances of this crossbreed were observed phenotypically (growth performances) and genetically (variation at selected microsatellite loci) and manifested through myofiber histology. Captive breeding of this species in carp hatcheries of Bangladesh could be initiated and facilitated through maintaining the gene pool of the L. ariza of Kangsha and the Atrai River. Conservation could also be considered for advancing the better performer crossbred (G4K♀A♂) as the reference diallel crossbred. The simple yet efficient breeding technique and the manifestation of growth performances using histological and phenotypical observations will encourage the hatchery operators of Bangladesh and elsewhere to initiate the induced breeding of L ariza in large scale aquaculture.Diallel crosses are breeding schemes used to estimate combining ability and the effects of reciprocal crosses among strains or varieties. Using this diallel cross scheme, we produced a crossbred population of Ariza labeo (Labeo ariza) originating from three geographically separated rivers  in Bangladesh. Intra  and inter  stocks were produced following diallel cross . Reproductive and growth performance, muscle cellularity and genetic variation following genotyping of eight microsatellite markers  and analysis of all crossbreeds was performed. The fertilization (95% ± 2.11%), hatching (88% ± 1.03%), and survival rates (82% ± 1.88%) of G4K♀A♂ were higher compared to other groups. With respect to length and weight gains (2.67 ± 0.4 cm and 3.39 ± 0.2 g), SGR (3.23% ± 0.20%), and heterosis (8.87% and 24.74%) G4K♀A♂ was the superior group. A higher number of hyperplastic muscle fibers, mean number of alleles (2.75) and mean observed heterozygosity (0.417) from G4K♀A♂ could be interpreted to mean that G4K♀A♂ comprise better performance efficiency compared to others and are considered for continuing the L. ariza stock improvement program.The objective of the present study was to evaluate the growth performance and genetic variation in diallel crosses of Ariza labeo ( Labeo ariza)  is considered one of the most commercially important indigenous species (belongs to minor carp group) in the Asian subcontinent including Bangladesh, and it inhabits small rivers, floodplains, and creeks throughout the Indo-Pacific region [L. ariza is an omnivore and column feeder that grows to a maximum size of 50 cm in length [Labeo rohita; Catla: Catla catla; Black Rohu: Labeo calbasu; Mrigal: Cirrhinus cirrhosus), and high market demand [L. ariza has been listed as a vulnerable fish species in Bangladesh [Dawkinsia rohani (Rohan’s barb) [Osteobrama belangeri (Pengba) [Labeo gonius (Minor carp) [Ariza labeo (c region . L. arizn length  and attan length . This fingladesh . The devor carp) . Howeveror carp) ,11,12. Tor carp) , the comor carp)  and the or carp) .L. ariza, where the success of breeding schemes is countered by the complexities of negative selection  and inbreeding, a diallel mating design could help improve the genetic and phenotypic gains and may offer additional management tools to develop the base population of a breeding program [Diallel crossing is a breeding scheme that has been extensively applied to estimate the combining ability as well as the effect of reciprocal crosses in plants and animals ,17,18. T program ,21. Heal program ,23,24. M program ,26.Muscle growth in fish develops through the generation of new muscle fibers (hyperplasia—a condition that augments cell numbers of a particular organ and/or tissue) and/or the expansion of existing muscle fibers (hypertrophy—An increase in the size of cells (or tissues) in response to various stimuli) ,27,28. RL. ariza in the wild and the long history of poor outcomes of the breeding schemes in the carp hatcheries of Bangladesh [L. ariza and comprehensively studied the performance of the intraspecific crossbred populations based on physical growth and myofiber histology. We also assessed the molecular genetic variation in crossbreeds using microsatellite DNA markers. Therefore, the aim of the present study was to evaluate the growth performance and genetic variation in diallel crosses of the Ariza labeo (L. ariza) originating from three geographically separated rivers in Bangladesh. We also aimed to conserve the best group as a founder stock that will facilitate future research aiming to promote sustainable genetic and phenotypic gains in L. ariza populations.Considering the threatened status of ngladesh , we implThis study included sampling from the wild, breeding in captivity, and fin clipping of the animals without euthanizing them, among other activities. Animal scientific procedures were rigorously followed during these activities with prior approval by the Animal Welfare and Ethics Committee of the Bangladesh Agricultural University (BAU) (ref. no. BAURES/ESRC/FISH-30/22).L. ariza (n ≥ 900) were collected from each of the three river populations of Bangladesh, i.e., the Atrai (Dinajpur District), Kangsha (Mymensingh District), and Jamuna Rivers (Sirajganj District) (Advanced fry of istrict) . The aveL. ariza (late juvenile stage with 1:1 sex ratio) from each stock  were reared for six months in nine separate grow-out ponds (18 × 14 × 1.3 each). A vitamin E supplement was incorporated into the protein-rich feed (35%) during the rearing period and fed at 5% of the body weight. Consequently, after 12 months of culturing (including six months of nursery rearing and six months of grow-out culture), the final length, weight, and survival rates of the fish were determined.One hundred and eighty The GSI, a metric representing the relative weight of the gonad to the fish body weight, was determined over a period of 12 months for mature fish. Year-round (between October 2017–September 2018) GSI data was collected following the sacrifice of three males and three females  in every−1 bw and 8 mg·kg−1 bw) at 6-h intervals, while the males  were treated with a single dose (4 mg·kg−1 bw) at times when the females received their second dose.A total 48 broods (16 from each stock source from three riverine system following a sex ratio of 1:1) were selected for a 3 × 3 diallel crossing, within and among stocks of the Kangsha, Jamuna, and Atrai Rivers. The pituitary extract of carp was used for effective induced breeding from selected brood samples. Female broods (having an average length 16.34 ± 0.44 cm and weight 90.13 ± 0.23 g) were treated with two doses (4 mg·kgWe designated the intraspecies inter- (and intra) stock crosses to produce the F1 generation and the groups were acronymed as G1K♀K♂, G2J♀J♂, G3A♀A♂, G4K♀A♂, G5K♀J♂, G6A♀K♂, G7A♀J♂, G8J♀K♂, and G9J♀A♂ (For details see Twenty-one-day-old fry were released in nine different earthen ponds (9 × 7 × 1.3 m each) with the same facilities as those described in 2), these measurements were further converted to the values of equivalent diameter, in accordance with Weatherley and Gill [The muscle growth patterns of early juveniles (82 days post fertilization) of both the intra- and inter-stock crossbreds were analyzed using trunk muscle histology counterstained with hematoxylin ,39. Brieand Gill . The groMeasured muscle fibers were categorized into small (<20 µm), medium (21–30 µm), and large (>30 µm), in conformity with Johnston , and forLr1, Lr2, Lr3, Lr22, Lr24, Lr27, Lr28, and Lr29)  using the GeneJET Genomic DNA Purification Kit . The microsatellite loci were amplified using PCR in a 10-μL reaction volume following the standard PCR protocol [Eight microsatellite markers (nd Lr29)  develope. rohita  were useprotocol . For fraa), the effective number of alleles (Ne), the frequency of alleles, observed heterozygosity (Ho), expected heterozygosity (He), fixation index (F = 1−Ho/He), Nei’s genetic distance, and gene flow (Nm).Significant difference of growth indices and muscle fibers mean’ number of different F1 generation groups was calculated by one-way analysis of variance. A multiple range test of Duncan’s at a 5% significance level was performed to specify the difference between groups. These growth and muscle fiber data were analyzed using SPSS . Alpha Ease FC 4.0 was used to estimate markers and allelic length. The GenAlEx program version 6.51b2 proposed by Peakall and Smouse  was usedL. ariza originating from three riverine sources after a domestication period of 12 months are shown in p < 0.05; 16.78 ± 0.68 cm and 95.92 ± 2.24 g, respectively) than those of the Kangsha  and the Jamuna  river stocks. The survival rate of Kangsha fish was significantly higher (91% ± 0.3%) than those of the Atrai (86% ± 0.12%) and Jamuna (84% ± 0.15%) fish.The observed growth performance and survival rates of Naturally, the GSI values of females becomes more compared to males for wight variation between the ovary and testis. In this experiment, the GSI value of males varies between 0.55 ± 0.26 and 4.95 ± 0.35. As opposed to females, the lower and upper bounds of GSI was 1.15 ± 0.21 and 11.5 ± 0.37, respectively. Coincidentally, August was the peak GSI-recorded month for both sexes. In contrast, the GSI indices of both sexes were the lowest in November. The GSI of males plateaued in April, with a steady increase until May; it then increased gradually for the next three months, peaked in August, and then declined moderately . A similp < 0.05) than those of the other groups during seed production. Comparatively lower fertilization and hatching rates were found in G6A♀K♂ and G2J♀J♂ , with G2J♀J♂ exhibiting the lowest survival rate among all groups (The fertilization (95% ± 2.11%), hatching (88% ± 1.03%), and survival rates (82% ± 1.88%) of G4K♀A♂ were higher (l groups .n = 20 in each group) (p < 0.05) in G4K♀A♂ (3.23% ± 0.20%) and lower in G2J♀J♂ (2.98% ± 0.20%). Heterosis, for both length and weight, was higher in G4K♀A♂  than in the other crossbred groups  after 60 days of rearing (h group) . The lowd groups .A mosaic pattern distribution of fibers (with fibers of various diameters) was observed in transverse sectional trunk skeletal muscle histology . Small, Lr3, Lr24, and Lr27 were identified. Loci Lr24 and Lr27 were polymorphic, while loci Lr1, Lr2, Lr22, Lr28, and Lr29 were monomorphic in all groups; Lr3 was polymorphic in five groups. The average Na (2.75) and average Ho (0.417) were the highest in G4K♀A♂. The lowest Na (2.000) and Ho (0.167) were observed in G6A♀K♂ and G1K♀K♂, respectively. Negative average fixation indices over all loci were observed in G4K♀A♂ (−0.116), G6A♀K♂ (−0.452), and G7A♀J♂ (−0.144) (m (7.277) was observed between G3A♀A♂ (A♀ × A♂) and G6A♀K♂ (A♀ × K♂), while the lowest Nm (0.838) was observed between G1K♀K♂ and G3A♀A♂ (The microsatellite profiles of loci (−0.144) . The higd G3A♀A♂ . The higd G3A♀A♂ .L. ariza, a newly introduced but popular aquaculture species in Bangladesh. This present study aimed to observe and evaluate the outcomes of a 3 × 3 diallel crossing of riverine populations of L. ariza to assess the performance of the progeny in terms of i) the phenotypic gain (or loss) of economically important traits , ii) muscle fiber morphometry, and iii) genetic variation observed among the crossbred groups. Moreover, following the genetic principles of fisheries management, the superior group attained from this study (if any) aimed to be conserved as a founder population and also aimed to retain or increase the variability of this population.Sufficient genetic variation in the base population of a selective breeding program is a prerequisite for its success. In the present study, we performed a complete diallel cross to increase the genetic variation and improve the growth performance of L. ariza from three different rivers were collected and reared in on-farm ponds. After 12 months of on-farm rearing, in terms of different growth indices, populations from different riverine systems exhibited the best performance. For length gain and weight gain, the population from the Atrai River was superior, with the population from the Kangsha River having the highest survival rate. The growth trial of the present study was conducted in nine uniform earthen ponds located side by side under similar environmental conditions. All of the major water quality parameters observed during the experiment were within suitable ranges for aquaculture. Therefore, the growth performance variation was likely due to heterozygosity. Bangladesh is a small country with little climatic and environmental variation in its different regions. Samples were collected from the north (Atrai River), central (Jamuna River), and eastern (Kangsha River) regions. Over hundreds of thousands of years, an independent genetic structure evolved in these rivers. Therefore, we conclude that the better performance shown by the Atrai River stock was because of genetic effects. We also found better growth performance in G4K♀A♂. Corroborating our results, higher length and weight gains in L. ariza from the Atrai River were also reported by Asaduzzaman [In this study, the advanced fry of L. ariza can be considered a synchronous spawner, unlike other carps  and some minor carps and minnows . The results of the present study suggest that L. ariza has only one breeding season (between July and September), and it is short. It reaches its peak in August  that could facilitate the planning and conducting of induced breeding. GSI has also recently been considered as a reference point against any changes in sexual development and/or reproductive success, both in the wild and in captivity . In thismolitrix  and Hypo nobilis ), but lin August , in cont October ,50 with  October .C. reba  seed production, the application of Ovaprim as an inducing agent was conducive to the fertilization rate (86.4%) and hatching rate (91.6%) at a satisfactory level [L. rohita) (Hatchery♂ × Padma♀). Positive heterosis in growth was also reported in another major Indian carp, Cirrhinus cirrhosus, by Sayeed [Macrobrachium rosenbergii, was assessed by Suburamanian [Macrobrachium rosenbergii stock shown by Suburamanian [The higher fertilization, hatching, and survival rates in F1 progeny from G4K♀A♂ might be attributable to the combination of better alleles and the dominant genetic variance. However, the dominant genetic variance is not predictable. In the case of ry level . The intry level . An incrry level . One of ry level  et al. oy Sayeed , with a ramanian . Weight ramanian  have hadL. ariza resulted in an uncounted number of fibers with small-diameter which were further encompassed by large fibers. Hyperplasia-derived new fibers remained small during its’ first generation and the size tended to increase as it grew. As skeletal muscle is the main contributor to body weight in most fish, it is probable that the size of a fish is limited by the growth of this tissue [Dicentrarchus labrax [Dentex dentex [Gadus morhua [Sparus aurata [Solea senegalensis [In the early juvenile stage, the trunk skeletal myofiber morphometry of distinct groups of s tissue . Muscle s labrax , Dentex x dentex , Gadus ms morhua , Sparus s aurata , Solea sgalensis , and mangalensis ).Sparus aurata, was studied by Rowlerson [Hyperplasia, hypertrophy, and elongation are the subsequent events involved in muscle formation and muscle fiber enlargement of teleost fish. Myogenesis consists of serial complex events involving the specification, proliferation, differentiation, migration, and fusion of precursor cells to form multinucleated muscle fibers. The myogenesis rate, the subcellular organelles’ architecture, gene expression patterns, and muscle fibers size and numbers of an organism are all determined by the environment where they belong . The posowlerson , who aimowlerson . In contowlerson  and Johnowlerson  substantL. ariza was the main phage where muscle growth eventuated by way of hyperplasia—ascertained from the appearance of mosaic fiber lines. Among all nine groups, the highest muscle hyperplasia eventuated in G4K♀A♂ , of which two (Lr24 and Lr27) were polymorphic in all groups. Previously, these markers were used in population genetic studies of major Indian carp, where polymorphisms were detected at Lr3 in C. cirrhosus [Lr1, Lr2, Lr3, Lr22, Lr24, Lr28, and Lr29 in L. rohita [a, indicated that this crossbred group was superior to the other groups. In contrast, the lowest average Na (2.000) was observed in G6A♀K♂, which indicates the allelic deficiency of that group. Overall, low levels of variation in terms of allelic richness and heterozygosity were observed in the three riverine stocks of L. ariza, which indicates that the populations might have suffered from a reduction in their sizes, causing bottleneck effects. The loss of alleles and heterozygosity may increase with bottlenecking and inbreeding over generations in hatchery stocks [L. ariza samples used in the present study were collected from wild sources. The reduction in allelic variation may also be explained by the bottleneck effect and genetic drift in the population, resulting from a sudden decrease in population size (Ne), which might have occurred in the case of L. ariza.Genotyping using appropriate molecular markers is used to assess genetic variations in fish populations. In the present study, we created nine groups by applying a 3 × 3 diallel crossing design in three different wild stocks of irrhosus  and at L. rohita ,68,69. Ty stocks . HoweverL. ariza through crossbreeding. Moreover, we observed higher growth in terms of length and weight gain in four of the six crossbreeds compared to that of the original riverine stocks. Zeng [Siniperca chuatsi), which supports the findings of the present study. In crossbred stocks of Macrobrachium rosenbergii from the diallel program, genetic variation increased in an analogy of their cultured stock [L. ariza among three river populations, including Atrai, Jamuna, and Kangsha, one of the parental populations of G4K♀A♂. The Kangsha and Atrai Rivers are geographically distantly located, and there is little chance of genetic interchange between the two stocks. The crossing between the G4K♀A♂ stocks in the present study resulted in genetic intermixing, which could have caused the high genetic variation in G4K♀A♂. The high genetic diversity observed in a population could be explained by the overlap of generations, population mixing from different geographical locations, natural selection favoring heterozygosity, or subdivision accompanied by genetic drift.Populations with less genetic variation have little chance of improvement through selective breeding. In that case, crossbreeding is the most promising alternative that can improve genetic variation and the potential for the genetic improvement of the population. Accordingly, as hypothesized, we observed higher heterozygosities in four  of the six crossbred groups compared to their parental groups. Therefore, the results of the present study have created an opportunity to improve the genetic quality of existing stocks of ks. Zeng  also obsed stock . Asaduzzed stock  found hiL. ariza stock of the Atrai River is genetically distinct from that of the Kangsha River. Along with the stock of Atrai River, populations of Kangsha and Jamuna expressed phenotypic variations [The lowest gene flow (0.838) and the highest genetic distance (0.335) were observed in G1K♀K♂ and G3A♀A♂, respectively. These results could be attributed to the geographic isolation of the Atrai and Kangsha Rivers, which minimizes the chances of population exchanges between the two rivers. Thus, the riations  and generiations  found a L. ariza populations, venturing to develop an effective stock improvement method and generate high-quality L. ariza seeds. We analyzed 30 individuals from each of the nine groups of L. ariza using eight microsatellite primers. This small number of samples is a limitation of the study to elucidate an extensive epilogue on genetic variation of L. ariza in Bangladesh. However, based on the results of the present study, it can be inferred that the diallel cross increases the genetic variation in the crossbreeds compared to the purebred L. ariza. These results also highlight the natural resources that require scientific management for the preservation of pure breeds  in the case of poor broodstock performances of the crossbred populations owing to allelic segregation. This G4K♀A♂ line could be further advanced, improving genetic and phenotypic variation in their subsequent generations following pertinent selection as well as breeding.In this study, we examined for the first time the muscle growth and genetic variation in diallel crosses of


According to the Culture Code Theory of Design, posters can be divided into strategy aspect, meaning aspect, and technical aspect. This study explored the impact of visual imagery on poster communication and effect focus on the meaning and technical aspects which directly related to the audiences. The visual imagery preference of ten public poster samples is collected from 40 participants by online questionnaire with the semantic differential method. The results show that the audience’s visual imagery at the technical aspect is mainly related to the richness of colors, graphics, and texts. When receiving the information from the meaning aspect of the poster correctly, the visual imagery will be influenced by the type of theme. Most importantly, the same or similar visual imagery at the technical and meaning aspect encourages audiences to explore the deep information of posters. Poster works that keep visual imagery coherent and smooth at the two aspects can achieve a better effect. Imagery, also known as psychological image, is the psychological feeling based on physical image and personal perception. Imagery can be both produced in the original consciousness field of the information producer, and extracted and formed after the audience receives the information . AccordiThe poster is a functional visual design, which plays an important role in information communication and owns a significant relationship with visual imagery . The visCurrent pieces of research mainly focus on the visual attraction and semantic understanding of posters. Most scholars believe that a good poster design needs to attract the audience’s attention at first. For example, colors and graphics with strong contrast will leave a deep visual impression on the audience, stimulate them to decode the information expressed by designers, and re-code it into personal cognition . They apAs a culture system theory, the Culture Code Theory of Design was first proposed by Yufu Yang based on semiotic theory . The CulPoster design is an art design that requires both visual attraction and readability. Applying the Culture Code Theory of Design, we can completely study the visual imagery of posters from the technical aspect  and the meaning aspect (theme) to provide designers with design and creation strategies to improve the effect of poster design.1. Every year, the 25 best posters will be awarded at a public exhibition with thousands of international posters. It has been held for four sessions since 2017 and involves four public themes: Woman, Tolerance, House and Living, and Water. A total of 100 winning works from the four sessions were selected as the initial sample set. An expert group composed of two design teachers and two design students coded 100 posters from the meaning aspect (theme) and the technical aspect  of the poster as shown in The poster sample for this research comes from the German ANFACHEN Poster Design Award2 is used for word segmentation and word frequency statistics. A total of 54 adjectives are obtained. The expert group selects ten adjectives as the final evaluation basis for the poster work. Then, ten pairs of representative opposite words are chosen, including Soft-Intense, Popular-Individualized, Plain-Fashionable, Traditional-Modern, Elegant-Heroic, Serious-Lively, Cold-Enthusiastic, Static-Dynamic, Empty-Plump, Harmonious-Conflicting.The poster evaluation text is collected from relevant literature, magazines, and websites. The Weiciyun websiteForty participants are recruited in total. Half of the participants come from design majors, and the other half come from non-design majors, with 50% male and 50% female to eliminate the potential influence of gender and specialty on experiment results. All participants are informed of the experiment procedure and sign informed consent. This experiment has been approved by the Ethics Review Committee of the South China University of Technology.Using the 7-level Likert scale and regarding the two words of each adjective pair as two extreme cases, the participants use the scale to choose the degree of personal feeling between each adjective pair. The poster samples appear completely on the computer screen or mobile screen at a specific time. All participants need to fill in the questionnaire twice. In the first questionnaire, the participants are reminded to complete the questionnaire for each poster sample based on first impressions. The first appearance time of the poster is 7 s according to the research on the first impression of online web pages . In the p < 0.001), and the interpretation rate of the total variance is 69.939%.First, the questionnaire with the same score of ten dimensions is considered invalid and is eliminated. There is an overall analysis of the visual imagery of the poster sample at the technical aspect and the meaning aspect with the factor analysis of SPSS . The factors with large loads in multiple factors are deleted  and three principal factors are extracted from the seven factors. The final factor analysis result is shown in The evaluation criteria of the word pairs Traditional-Modern, Plain-Fashionable, and Popular-Individualized in Factor 1 is related to the social trend, so Factor 1 is named Sense of Contemporary. Factor 2 including Cold-Enthusiastic, Serious-Lively mainly describes the temperature of the poster, so Factor 2 is named Sense of Humanity. Factor 3 contains Elegant-Heroic and Soft-Intense that focus on the impact of the poster, so it is named Sense of Power. Take Factor 1, Factor 2, and Factor 3 as x, y, z coordinates, respectively, and the average of all samples in the three factors as the origin, the poster samples’ visual imagery maps of every two aspects are drawn according to the difference . Figure T Test of SPSS (IBM) was used to explore the differences between gender and profession (n = 40). It showed that there is no significant difference in the questionnaire results between different genders. Only in the meaning aspect, the professional group compared with the non-professional group consider that sample 5 was more fashionable . Therefore, this research will ignore the influence of gender and specialty on visual imagery. Using the paired-samples T Test of SPSS (IBM), the results show that the meaning aspect makes sample 1 more dynamic and enthusiastic, sample 2 more modern and intense, sample 5 more plump, sample 6 more personalized and fashionable, and sample 8 more dynamic. The visual imagery of the other samples doesn’t change significantly.The independent-samples The visual imagery at technical and meaning aspects of the poster is the emotional tendencies created by different colors, graphics, text, compositions, and themes. Current research shows that emphasizing emotion in design can help enhance the audience’s acceptance and understanding of design works . The expIn the detailed comparative analysis of the technical aspect and meaning aspect, there is no significant difference in the visual imagery of samples 4, 7, 9, and 10. Considering the higher understanding degree of samples 4 and 7 than that of samples 9 and 10, there may be two reasons to explain this situation. One reason is that the audience did not grasp the deep meaning of the poster sample so the meaning aspect does not affect the final visual imagery. The second reason is that the visual imagery of the meaning aspect and technical aspect is consistent, so the emotion would not change. The comparison between samples 7 and 9 can verify the two reasons. In sample 7, the IKEA banners are entwined to form a monster roaring which covers the full poster, and the graphics are abstract with blue and yellow color. The audience’s first feeling about the poster elements is conflict and strong which is consistent with the poster’s meaning of housing tension and urban crowding. Even if some audiences do not notice the hidden IKEA logo and the vague monster image in the picture, they can still accurately receive the expected message of the poster. In contrast, sample 9 also expresses the theme Housing and Living where the text “We are all migrants” assists the semi-abstract graphics like a human-shaped airplane to express the housing crisis and housing instability. The visual imagery of the technical aspect with blue, white, and black color and the central composition conflicts with the strong visual imagery of the meaning layer about human existence. Even with the help of text, the understanding of the meaning of sample 9 is not better than that of sample 7.It can be seen that only when the audience receives the information correctly, the meaning aspect affects the visual imagery of the technical aspect. And the poster with unified visual imagery at both aspects has a higher understanding. The consistency between the first impression and deep thinking, from the technical aspect to the meaning aspect plays a vital role in the persuasive effect for poster design.This research found that different visual imagery will be produced by different colors, compositions, and themes of the poster. The audience will have a higher understanding of the poster that owns consistent visual imagery in the viewing process. The result suggests that the posters with the theme of women and children will awaken the softness of the audience, which can be assisted by softer colors and stable composition. The strong and conflicting visual effect will be more conducive to the expression of social themes such as environmental protection and social competition.In conclusion, the same or similar visual imagery at the technical aspect and the meaning aspect encourage audiences to explore and think further. The posters that keep the visual imagery coherent can achieve a better effect.The original contributions presented in the study are included in the article/The studies involving human participants were reviewed and approved by Ethics Review Committee of South China University of Technology. The patients/participants provided their written informed consent to participate in this study.LW contributed to the conception of the study. WJ performed the experiment and the data analyses and wrote the manuscript. WC contributed significantly to manuscript preparation. SL helped perform the analysis with constructive discussions. 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.


Xanthophyllomyces dendrorhous is an established platform for genetic pathway modification. A genetic tool box is available and can be used extensively to select from for different engineering strategies. Due to the diploid nature of X. dendrorhous, genetic transformation typically results in heterozygous lines. They are genetically unstable and lose their phenotypes caused by mitotic recombination. In addition, targeted integration for inactivation of genes of the carotenoid pathway resulted in an intermediary phenotype of incomplete pathway disruption. This issue is the main scope of this review. It is illustrated by using genetic modification of the carotenoid pathway of X. dendrorhous as a model system with a focus on the demonstration of how to solve these problems by generation of homozygous lines. They can be selected from heterozygous transformants after spontaneous mitotic recombination and selection or after induced meiotic recombination. Corresponding methods of how to proceed including the initiation of a sexual cycle are described. The selected segregated lines are stable in fermenter cultures without the need of selection pressure. This is an essential requirement for any industrial application.The nonconventional yeast • Genetic interventions of diploid yeasts result in heterozygous transformants that are unstable without selection pressure.• This is due to mitotic recombination leading to the elimination of inserted DNA.• Stable homozygous lines can be obtained and selected after either meiotic or mitotic recombination. Xanthophyllomyces dendrorhous is a heterobasidiomycetous budding yeast belonging to the order Cystofilobasidiales  contains not only colourless phytoene in contrast to the wild type but also astaxanthin, the end-product of the pathway, as indicated by its yellowish pigmentation  and crtI with the inserted resistance gene  (Liu et al. Candida utilis (Liang and Bennett X. dendrorhous are already of interest for genetic engineering of the carotenoid biosynthesis pathway (Shimada et al. Kluyveromyces marxianus (Ortiz-Merino et al. Yarrowia lipolytica (Schwartz et al. X. dendrorhous has been successful (Hong et al. X. dendrorhous and other diploid yeasts.Genetic pathway engineering of so far unnoticed nonconventional yeasts including diploid species offers promising opportunities for new industrial production processes. The described methods for obtaining homozygous transformants are applicable in combination with any genetic intervention of diploid yeasts. They may be specifically useful for diploid strains of


Loligo forbesii Steenstrup, 1856, occurs at the European Shelf areas including the Azores and represents a valuable resource for the European commercial fishery in the North East Atlantic. However, very little is known about its population structure and phylogeography. This lack of knowledge also impedes the development of sustainable fishery management for this species. The present study combined the use of two types of markers that retrieve patterns of gene flow in different time spans; the analysis of 16 nuclear microsatellites and sequencing of the mitochondrial cytochrome oxidase subunit I (COI). Whereas the high mutation rate of microsatellites allows the description of recent patterns of connectivity in species, the lower mutation rate of COI provides phylogeographic patterns on a longer timescale. A total of 347 individuals of L. forbesii were investigated from nearly the entire distribution range of the species, including the North East Atlantic Shelf, the Azores and the Mediterranean. Individuals from the Western and Eastern Mediterranean Sea have never been included in a genetic study before. We were able to analyse COI sequences from all 12 sampling areas and define three clades of L. forbesii. Due to our large sampling area, we are presenting 13 COI-haplotypes that were previously unknown. The microsatellite analysis does not include the Azores but three main clades could be identified at the remaining 11 sampling sites. Low FST values indicate gene flow over large geographical distances. However, the genetically significant differences and an additional slight grouping in the microsatellite structure reveal that geographical barriers seem to influence the population structure and reduce gene flow. Furthermore, both markers provide strong evidence that the observed phylogeographic pattern reflects the geographical history of the Azores and the Mediterranean Sea.The veined squid,  Especially in the Atlantic Ocean, the commercial importance of the loliginids has grown3. Loliginids landings reached about 12,000 t3 in 2017 showing a fivefold increase between 2000 and 2017 in the entire Northeast Atlantic, which illustrates the growing socio-economic importance. Among the loliginids, the veined squid Loligo forbesii Steenstrup, 1856 is one of the most important species for the European fishery3. This species is neritic, associated to the shelf and equally distributed on the lower shelf (80–200 m) and the upper slope (200–500 m) in the northern Mediterranean4. In the North East Atlantic it occurs from the North Sea and UK waters to the Canary Islands and Azores2. This opportunistic predator is characterized by high growth rates. During its 12–16 months of life span, it can reach a dorsal mantle length of up to 900 mm6. As a semelparous species L. forbesii spawns only once in its lifetime2.Within the last decades biomass of various cephalopod populations and commercial catches increased world-wideL. forbesii is unregulated fished as bycatch, but European target fisheries exist in the English Channel, Scottish and Portuguese waters3 and recreational fishery develops in UK and Norway. First prediction models for L. forbesii have been developed7, but up to now it has been difficult to develop a sustainable fishery management for this valuable resource due to knowledge gaps regarding population structure8, life cycle and spawning areas9. Here, we will focus on the first aspect and will shed new light on the genetic population structure of L. forbesii.In most regions 10. However, to elucidate the complex population structure of L. forbesii, allozymes and mitochondrial DNA sequences were found apparently unsuitable because of their extremely low levels of genetic variability12. In such cases, a more appropriate approach is the analysis of microsatellites. Microsatellites are short repetitive sequences in the nuclear genome showing high mutation rates and multiple alleles13, allowing a powerful analysis of recent patterns of population connectivity. Shaw et al.12 found a higher genetic variability by using microsatellites for L. forbesii compared with previous studies using allozyme and mitochondrial DNA (mtDNA) markers, and were able to identify subtle genetic differentiation between species occurring at the European shelf seas (Scotland to Northern Spain) and offshore population including the Azores, Rockall and Faroes. They suggest that water depth and isolating current regimes are responsible for the genetic differences between offshore and onshore areas.The mitochondrial cytochrome-c-oxidase I (COI) gene is one of the most commonly used markers for molecular systematics and is often used to infer phylogeographic patternsL. forbesii we performed a mitochondrial COI and nuclear microsatellite analysis using the same dataset. In comparison to Shaw et al.12 we were able to geographically expand the study area to almost the entire distribution range of L. forbesii including the North-East Atlantic  and the Mediterranean Sea . The new findings in population structure, gene flow and habitat connectivity of L. forbesii might have a substantial influence on the management of this species’ fisheries, as they provide new information regarding the identification of different appropriate management units, so called “stocks”14.To shed new light on the genetic structure and phylogeography of Loligo forbesii was sampled at 12 European areas, including shelf and upper slope areas and the Azores in 2019 , were sent to the TI-OF, alcohol was changed once more and stored until the beginning of the genetic analysis at University of Rostock, where DNA was extracted using “innuPrepMini Kit”  following the manufacturer’s standard protocol.Loligo forbesii is not protected under any legislation and not considered threatened or endangered. Atlantic samples were collected as bycatch during ICES coordinated international fishing trawl surveys  and CEFAS Otter Trawl Survey (Celtic Sea) or at the fish market (English Channel). L. forbesii samples from the Mediterranean were bycatch from the International bottom trawl survey in the Mediterranean (MEDITS) except samples from the Balearic Sea which were collected from a commercial vessel. Azorean samples have been collected by the artisanal jigging fishery. All individuals were already dead when they were handed over to collect the tissue samples for our study and therefore handled in accordance with relevant guidelines and regulations. Fishing licenses for all surveys and trips were available.L. forbesii16. Specimens from the Azores were not included in this analysis, because (1) we received the samples only 1 year after completion of the microsatellite analysis and (2) Shaw et al.12 already showed the uniqueness of L. forbesii in this area based on microsatellites. All primers were marked with a fluorescent dye. The master mix for the PCR included 0.5 µL of both primers (10 µM), 1.0 µL dNTPs , 1.0 µL 10 × Buffer, 0.085 µL Taq-Polymerase (5 units/µL) and between 0.8 and 1.2 µL MgCL2 (25 mM), referring to the primer. This was filled up to a volume of 9 µL with water. For the PCR reaction 1 µL DNA isolate was added. The reaction conditions followed the description of Shaw et al.12. Next, the concentration of DNA was estimated using calibrated gel electrophoresis and diluted accordingly. The diluted PCR product was added to 9.8 µL HiDi™ formamide and 0.02 µL size standard GeneScanTM LIZ™ . The samples were denatured at 96 °C for 4 min and fragments were size-separated in a capillary sequencer . If the signal in the electropherogram was too weak, the whole procedure was repeated up to three times.Population genetics of 300 individuals from eleven sampling areas were performed via microsatellite analysis at the University of Rostock using 16 specific primers (Lfor1–Lfor16) for 17. Thereafter, the software performed the allele binning and the length of the different alleles was determined.The recorded electropherograms were evaluated using “Geneious Prime” . The evaluation followed the guidelines of Butler18 calculated the number of alleles per locus as well as the expected heterozygosity19 and the observed heterozygosity20. Furthermore, statistics on linkage disequilibrium and Hardy–Weinberg Principle21 were performed with “GenePop”23. “HP RARE”24 was used to determine allelic richness and private allelic richness, based on 15 individuals. The calculation and the pairwise comparison of the FST values were performed in “Arlequin Ver 3.5”25. Cluster analyses based on the microsatellite data were conducted with “STRUCTURE” version 2.3.4426. All settings were selected according to the standard settings for microsatellite data as recommended by Gilbert et al.27 and Porras-Hurtado et al.28. For a K-value from 1 to 10 a total of 25 iterations were calculated. Next, the best K-value was calculated with the ΔK method29, using the online software “Clumpak” (http://clumpak.tau.ac.il/)30.For the statistical evaluation several programs were applied. The “Microsatellite Toolkit” by ParkL. forbesii, 218 individuals from all 12 sampling areas were analysed. The primers LCO1490 und HCO219831 were used to amplify a 648 bp long part of the mitochondrial cytochrome oxidase I (COI) gene. For samples with poor DNA quality, internal primers (see Supplementary Table 2 (25 mM), 0.21 µL Taq-Polymerase (5 units/µL) and 9.99 µL water. The PCR was conducted in 38 cycles: 30 s at 94 °C, 30 s at 55 °C, 60 s at 72 °C and finally 300 s at 72 °C.To describe the phylogeography of 32) of which 28 were retrieved from GenBank40. The phylogenetic analyses  was performed using “MEGA” version 641 with 1000 bootstraps. We determined the best fitted model of the ML method with “MEGA 6” on the basis of the Akaike Information Criterion (AIC), TN93 + G + I  was used. Cycle sequencing products were analysed by using capillary separation on an ABI Genetic Analyzer 3130 xl (Applied Biosystems/Hitachi). All products were sequenced in both directions. The sequences obtained were analysed with the software “CEQ8000 Genetic Analysis System”  and an alignment of 249 sequences was created (“BioEdit” version 7.2.5.42 was used for the Bayesian Inference (BI) method. Two independent runs with four chains were performed for 2 million generations using the Markov Chain Monte Carlo (MCMC) method. Calculations of the consensus tree, including clade posterior probability (PP), were conducted based on the trees sampled after the chains converged using “Tracer 1.7”43. The first 25% were discarded as burn-in. We determined the best fitted model of the BI method with “Modeltest”, implemented in “MEGA 6”, on the basis of the Bayesian information criterion (BIC), T92 + G.“MrBayes 3.2.7”44) using “Network 5.0.1.0” and its standard settings . Additionally, a second haplotype network was calculated containing 313 individuals with shorter sequences (432 bp), because some individuals yielded incomplete or shorter sequences  to 49 (Lfor12). The test of linkage disequilibrium showed that all loci are inherited independently. Some deviations of expected and observed heterozygosity were found to be significant. On average across all loci for each sampled area, only non-significant deviations were determined, meaning the Hardy–Weinberg equilibrium could not be rejected.The average private allelic richness Table  across aST values between the sampled populations (areas) were low but after Bonferroni-correction still some significant differences (p < 0.0009) were found  belong exclusively to the “Mediterranean” clade, while individuals from the eastern Atlantic, with few exceptions, are members of the “East Atlantic” clade. The Gulf of Cadiz and the west and east coast of Sardinia represent a transition zone between the North Atlantic and the Mediterranean, with both clades occurring in these areas Fig. .Figure 4The shorter and longer sequences show similar haplotype networks, although some mutations are omitted in the network performed with the shorter sequences length  with high gene flow between populations within and, partially, between the genetic clusters. Due to our substantially expanded sampling area and increased number of microsatellite loci in comparison to previous studies, we were able to identify a higher number of alleles giving us a more robust dataset for gene flow inferences48.Our results based on 47. This inconsistency between different studies may be due to different methods of allele scoring. Furthermore, loci Lfor14 and Lfor15 showed additional polymorphic amplification products. As unspecific priming and chain-termination can be excluded, we consider that these amplifications may eventually represent repeated regions in the genome. Not every locus delivers suitable results for every sample. This is not unusual as microsatellites are sensitive and the quality of the tissue sample plays an important role. Our dataset has 4.92% missing data, which is slightly higher than the 3.75% in Shaw and Boyle47. Not every locus was affected equally by missing data. The differences in the present study are most likely due to low DNA quality. This was especially the case for some samples from the Bay of Biscay and Adriatic Sea for which the ethanol may have not been changed adequately.Locus Lfor7 showed up to eight peaks per individual, which was never described beforeST values are indicative of a considerable gene flow across the entire range of this species. Only sampling areas with a substantial geographic distance show low significant differences. Our STRUCTURE results show clear differences between Atlantic and Mediterranean L. forbesii individuals. The genetic composition of individuals from the Gulf of Cadiz to the western Mediterranean assigning to two or three genetic clusters is an evidence for recent gene flow, or otherwise a short period of genetic isolation. It is well known that the relatively shallow area of the Strait of Gibraltar has the potential to be a barrier for some species50. But it is not surprising that the area represents a transition zone for a shelf-living species such as L. forbesii. In addition, admixture within the Atlantic and the Mediterranean sampling sites seems to occur, represented by high gene flow rates  and Doryteuthis gahi (formerly known as Loligo gahi) show poor genetic differentiation as well52. Our results from northern Atlantic sample sites correspond in general to the findings of Shaw et al.12 and Brierly et al.11. Hence, although we were not able to incorporate the Azorean samples in our microsatellite analysis, we anticipate that the Azores would represent a fourth microsatellite cluster as Shaw et al.12 uncovered significant genetic differences between L. forbesii from the archipelago and the North East Atlantic Shelf area.The observed low values of private allelic richness, as well as the low pairwise FL. forbesii, whereof three have been reported before40 (H7 and H10, “East Atlantic” clade; H25, Azorean clade, see Fig. 53 and were caught at a known spawning area during spawning time54. The elektropherogramms revealed that a sibling or half-sibling relationship can be excluded for these three individuals, additionally indicating multiple events of gene flow from the Mediterranean into the Atlantic. It is known that Loligo reynaudii, a closely related species, performs migrations over large geographical distances during spawning time and does not exhibit homing behavior55. When ready to spawn, individuals of this species show an average migration speed of 3 km per day, and exceptional migration events were also reported, as a movement of 207 km in 18 days55. While this information is not available for L. forbesii, it may be hypothesised that the spawning behaviour and the ability to move over long distances of L. reynaudii may be also present in L. forbesii.We found a total of 16 COI-haplotypes for 56 (Supplement Fig. . Our phylogeographic results are concordant with COI-data from Sepia officinalis, a species with a similar distribution to L. forbesii. Perez-Losada et al.57 inferred that S. officinalis expanded from Northwestern Europe coast into the Mediterranean, and observed that Aegean and Ionian populations of S. officinalis are genetically clearly separated from the remaining Mediterranean. We also found this differentiation between Eastern and Western Mediterranean but for L. forbesii mitochondrial haplotypes are generally more admixed between the sampling sites. This result may be due to gene flow following secondary contact between these clusters. Differences between local patterns of L. forbesii and S. officinalis may correspond to the different dispersal ability of the two species, with L. forbesii being a way more mobile species. Both the geological history of the Mediterranean and physical barriers between eastern and western Mediterranean Sea seem to have had a huge impact on the phylogeographical patterns of both cephalopod species.The three mitochondrial clades could also be differentiated from the phylograms Fig. . The booL. forbesii. Comparing the same geographical range for both markers, we found two mitochondrial clades and three microsatellite clusters, which expectably express the different mutation rates of these markers. As microsatellites are known to better describe recent patterns of connectivity between populations13, our results reveal a recent differentiation into three clades within the European Shelf population.The microsatellite and mitochondrial COI analyses show concurring results in population structure of 12 described the influence of geographic barriers, like deep sea basins, on the migration patterns of L. forbesii. The Italian peninsula as well as the Hellenic trench are potential migration barriers in the Mediterranean Sea. The offshore location of the Azores is very likely the reason for the genetic isolation between L. forbesii from the archipelago and the remaining North East Atlantic shelf area12, as the squids prefer the upper shelf slope4. Active migrations over large distances also seem to be important58 regarding the exchange between the Atlantic and Mediterranean over the Strait of Gibraltar. The two-layer water exchange in the Strait of Gibraltar described by Izquierdo et al.59 may influence the distribution of L. forbesii. Less dense Atlantic water forms a current in the upper water layer which flows into the Mediterranean, while the denser Mediterranean water forms the Mediterranean outflow, an underwater current which brings Mediterranean water masses into the Atlantic. This water exchange may affect differently adult L. forbesii, which spend the daytime close to the ground and ascend to surface waters at night for feeding2. This way, adults are directly influenced by the water flow in both directions, which supports the exchange of individuals between Atlantic and Mediterranean triggering the genetic admixture that can be seen in the present results. It is also expectable that the transport of paralarvae is influenced by currents.Shaw et al.L. forbesii. We found a subtle genetic differentiation between the Atlantic and the Mediterranean, but statoliths of Loligo, indistinguishable from those of current L. forbesii and L. vulgaris, were collected in early Miocene deposits at southern French shores60. This means that an ancestral of the current L. forbesii population existed in the Mediterranean sea, but we anticipate that the final establishment of the current Mediterranean population took place only after the Messinian Salinity Event 5.5 million years ago62, as the survival of the species during this crisis is very unlikely. We thus assume a repetitive colonialization of the Mediterranean by L. forbesii from the Atlantic across the Strait of Gibraltar. A geographical separation of the species took apparently place during the last ice age in the Late Pleistocene63, and the East Atlantic and Mediterranean lineage could evolve in allopatry. Today, the two previously evolved lineages have come into contact, supported by the combination of gene flow rates and the distribution of clade-specific haplotypes.With our new findings of the COI-gene analysis, we are able to describe uncovered phylogeographic patterns for L. forbesii that inhabited both the Atlantic and Mediterranean, got isolated in the Mediterranean due to sea level change eventually during the Late Pleistocene glaciations, becoming later the common ancestor of the three clades observed today, the “East Atlantic” clade, the “Mediterranean clade “ and the endemic clade in the Azores.Under the assumption of the above mentioned scenario, the Sardinian cluster may have established following a second wave of immigration into the Mediterranean Sea. This cluster is represented in mitochondrial COI diversity by the “East Atlantic” haplotypes 5 and 6, which were only found around Sardinia. However, an alternative explanation for the genetic patterns observed is that a subpopulation of an ancestral population of L. forbesii. Even knowing that the structure analysis illustrates admixture in some regions, a more accurate classification into smaller management units seems not possible using the current genetic markers. Therefore, understanding the complex population structure of L. forbesii and differentiate stocks for a more precise management needs further research and more alternative methods. Some studies indicate that statolith shape analysis and statolith elemental analysis or the combination of both might be potential methods to identify smaller management units64.Based on our results, we recommend that future fishery management should consider at least three different genetic groups of Supplementary Information.