Lucilia eximia   is a blowfly with medical and forensic importance that shows genetic and color variation, however, these variations have not justified the description of new species. But in forensic entomology an accurate identification of species and subpopulations is crucial. We explored the genetic variation of L. eximia from eight localities, in five natural regions in Colombia using two mitochondrial fragments, including the standard locus for insect identification COI and the Cytb-tRNA-Ser-ND1 region. We found significant differentiation at COI and Cytb-tRNA-Ser-ND1 level, characterizing two lineages and revealing a deep and significant genetic split. High values of FST and genetic distances supported the two lineages. The origin of the divergence of L. eximia remains to discover. Examining whether the lineages have diverse ecological and biological behaviors could be a significant impact on the use of L. eximia in forensic and medical science. Our results could have relevant implications for the use of post-mortem interval estimation based on insect evidence, as well as our sequences improve the database used in DNA-based methods for identifying forensically important flies. Lucilia eximia   as an annoying fly, despite its valuable and unexpected uses. Applying controlled myiasis, their larvae remove necrotic tissue of chronic wounds , suggesting the presence of two species within—also called cryptic species  is also used to study intraspecific variability in Diptera (L. eximia genetic variability Cytb and the NADH dehydrogenase genes (Cytb-tRNA-Ser-ND1). However, this study showed a moderate level of variability, conversely to the high level of variability revealed by COI sequences in Colombian specimens , including highly disturbed areas, semi-rural areas, and secondary forests ( forests .Lucilia (We used Van Someren-Rydon traps (DeVries 1987) using a mixture of rotten fish heads and chicken viscera as bait. In each locality, we placed four traps at 1.5 m above the ground for 96 h and separated the traps 200 m from each other. Every 48 h, we replaced the baits and collected the specimens and identified the samples using taxonomic keys for the Neotropical species of Lucilia , 2014.COI gene was amplified using primers developed by Cyb-tRNA-Ser-ND1 region, using primers developed by L. sericata (NC_009733) in Geneious v.8.0.4.From one specimen leg of the 61 collected specimens, we extracted DNA using the DNeasy Blood & Tissue Handbook (Qiagen) kit. A 650 pb fragment from the We calculated haplotype diversity (Hd), haplotype number (n), nucleotide diversity (π), and genetic diversity parameters using DNASP v. 6.0  and calcCOI and the Cytb-tRNA-Ser-ND1 genes, we built a tree using Bayesian Inference Analysis and applied the evolutionary model GTR + T based on the Akaike criterion  and 61 sequences, representing 8 haplotypes of the Cytb-tRNA-Ser-ND1 region (H1b to H8bCytb-tRNA-Ser-ND1 haplotypes). COI sequences were deposited in GenBank under accession number KT160173–KT160218 and the Cytb-tRNA-Ser-ND1 sequences under KU665416–KU665476  had the highest frequency (28.3%) followed by H2 (17.4%) both shared among Copacabana, Caldas, Pajarito, and Cola del Zorro, the four closest localities in Antioquia  was the most frequent haplotype (60.6%) and was found in Cola del Zorro, Copacabana, Caldas, Pajarito, and Leticia—the first four localities are close to each other but far from the last Leticia. The second most frequent haplotype H6b (21.3%) was distributed in Playa Huina, Puerto Gaitán, and Santa Marta.We found a total of 8 o Gaitán . H1b  and “lineage II” , separated by a significant number of mutations. The haplotype network based on COI showed 25 mutational steps and the haplotype network based on Cytb-tRNA-Ser-ND1 showed 11 mutational steps  and the Mantel test indicated the absence of correlation between FST and geographic distances . Whereas genetic distance ranged from 0.2% to 6.2% between the two lineages. Lineage I showed higher genetic diversity than the lineage II . Specimens from Playa Huina showed the lowest COI and Cytb-tRNA-Ser-ND1 diversity, showing one haplotype. For lineage II, all localities of the department of Antioquia shared the two most frequent COI haplotypes, but the diversity varied per locality. Copacabana showed the highest COI  followed by Caldas , whereas Leticia showed the lowest COI and Cytb-tRNA-Ser-ND1 haplotype diversity with a unique haplotype.L. eximia in Colombia using two mitochondrial genes. Lucilia eximia showed significant levels of genetic variability with two mitochondrial lineages, revealing a deep and significant genetic split. The mutational steps observed in the haplotype networks supported the idea of L. eximia in Colombia could be a cryptic species complex. This is the first approach to the genetic variability of COI barcode region for delimiting Calliphoridae showed intraspecific distance values between 0.0%–1.487% in Colombia  from the forest and urban areas showed genetic differences attributed to environmental factors  showed a substantial level of variation in mitochondrial genes but a low variation in nuclear genes and non-morphological variation. This study concluded that the variation between P. regina populations was related to intraspecific mitochondrial divergence rather than species-level differentiation (We detected high levels of genetic variability of ntiation .L. eximia in forensic and other medical fields. In other species, fertility, life cycle, emergence, and adult longevity varied between populations and this variation was related to noteworthy molecular variation (P. regina shows developmental rate variation, which was explained by mitochondrial divergence between populations (L. eximia may differ biologically, studying the lineages of L. eximia in more detail is necessary. Differences in the developmental rate between lineages in L. eximia would affect the use of L. eximia in forensic entomology applications.Our results could have important implications for the use of ariation . For insulations . These fL. eximia exhibits considerable genetic variability and two mitochondrial lineages in Colombia. It remains to elucidate the origin of this divergence and explore if the lineages exhibit differences in ecological/biological behavior, which may have important implications for the use of L. eximia in the forensic field.We found significant results when analyzing a small sample of the population, we suggest that future research include more specimens, as well as nuclear genes. In conclusion, tjad031_suppl_Supplementary_Table_S1Click here for additional data file.


In promoting genomic medicine, genomic literacy, which is the public’s ability to acquire and utilize the knowledge and skills related to genetics and genomes, requires urgent improvement. This study determined how the three components of genomic literacy  are associated with risk and benefit perception for genetic testing and decision-making related to genetic testing. Using an online questionnaire, we surveyed genetic/genomic knowledge, health numeracy, interactive and critical health literacy, risk and benefit perception of genetic testing, and intention toward genetic testing of 857 Japanese citizens (aged 20–39). A vignette was created to measure the intention toward genetic testing, and respondents were asked about their willingness to undergo genetic testing and to share the results with their partners and children. The path analysis, which was done by creating a path diagram revealing the relationship of the three components with risk and benefit perception, revealed that genetic and genomic knowledge and interactive and critical health literacy affected the respondents’ intention to undergo genetic testing via recognition of the benefits of such testing. Further, it was suggested that health numeracy affects benefit perception through risk perception toward genetic testing. However, the goodness of fit of this model for the intention of conveying positive results to partners and children was relatively low, thus indicating that the hypothetical model needs to be reexamined. In recent years, expectations for genomic medicine have rapidly increased with advancements in genetics and genomics. For example, in the field of cancer and intractable diseases, the practical application of genomic medicine, such as more effective and efficient diagnosis and treatment of diseases suitable for an individual’s constitution and medical condition, has begun . MeanwhiHowever, it is difficult for the general public to fully understand genomic medicine due to its complexity and the high level of technological expertise required; as such, the general public’s knowledge of heredity and genomics is insufficient , 3. FurtIt has been noted that knowledge about genetics and genomics alone does not lead to appropriate action. Based on the concept of health literacy, the American Public Health Association  states tTypical health behavioral theories, such as the Health Belief Model and Protection Motivation Theory, demonstrate that the perception of risks and benefits is an important determinant of decision-making . Health According to a study of the processes by which risk and benefit perceptions influence decision-making, the following association has been found in the risk perception process: general tendency → situation recognition factor → option recognition factor → behavior. This suggests that the decision-making process in which each individual’s abstract way of thinking and recognizing influences the recognition of a concrete object, and as a result, determines their behavior, exists securely . FurtherAlthough genomic medicine is becoming widespread in Japan, there are still few opportunities for the general public to make decisions, because related genetic tests and treatments are not yet commonly known. In Japan, decision-making research on genetic testing is limited to the perception of patients undergoing genetic testing and their decision support , 21. TheBased on the above understanding, this study considers genetic/genomic knowledge, health numeracy, and interactive and critical health literacy as components of genomic literacy. It examines the impact of risk and benefit perception on decision-making relating to genetic testing.The survey covered 2,000 respondents  between the ages of 20 and 59, and 857 respondents  between the ages of 20 and 39 were included in the analysis of this survey. Since it has been noted that online surveys can bias respondents based on factors such as gender and age, the number of respondents for this survey was determined after evenly stratifying the gender and age groups according to the population distribution and percentages in the 2019 national survey. This study was approved by the Research Ethics Review Board of the Tokyo Gakugei University with which the authors are affiliated .The purpose of the study and the handling of personal information were explained on the webpage, and responses were considered to be the acquisition of consent.The respondents were asked about their sex, age, marital status, children, occupation, level of education, whether or not they had taken biology during high school, whether or not they had learned about genetics, and whether or not they or someone in their family had a genetic disorder.It has been pointed out that the existing genetic and genomic knowledge scales are biased toward assessing professional and academic knowledge . TherefoHealth numeracy was measured using the validated scale “the health numeracy test” by Miyoshi et al. . The queJapan is a country with high literacy, and the Communicative and Critical Health Literacy (CCHL) scale , developBased on the “Guidelines for Genetic Testing/Diagnosis in Medicine” , we creaThe content of questions on risk perception for genetic testing consists of five items such as “if I discover a genetic mutation (genetic change) associated with a disease, I will continue to be anxious about developing it in the future,” and “if I discover a genetic mutation (genetic change) associated with a disease, it will put a strain on my family and future children.”The content of the questions on benefit perception for genetic testing consists of five items such as “if I discover a disease-related gene mutation (genetic change), it will help me prevent, detect early, and choose treatment for my own disease,” and “if I discover a genetic mutation (genetic change) related to a disease, it will help prevent, diagnose early, and treat the disease in my family and future children.” The response to each question is based on a five-point scale with “strongly agree” (5 points) as the highest and “strongly disagree” (1 point) as the lowest value.Although genomic medicine is becoming widespread in Japan, there are still few situations in which the general public makes decisions of their own, because genetic tests and treatments are not commonplace. Therefore, in measuring behavioral intention toward genetic testing, we used a method of assessing behavioral intention by presenting a situation regarding genetic testing in the form of a vignette; a vignette assumes family medical history and genetic risk.The vignettes were created based on the advice of seven genomic medicine doctors and researchers. In this study, behavioral intention was regarded as decision-making in a narrow sense, and the term “behavioral intention” was used for measuring decision-making.“You are in your twenties and engaged. Your mother got sick at the age of 40. Your grandmother had also developed the same illness. You are in good health now. It is said that in 5–10% of cases, this disease is hereditary. If a genetic test is performed and a positive diagnosis is made, the probability of developing the disease in your lifetime is said to be 25–85%.”Regarding the question items on behavioral intentions, the question on “the respondent’s genetic testing intention” is as follows: “Do you plan on taking a genetic test for this disease?” The response to this question is based on a five-point scale with “I will take the test” (5 points) as the highest and “I will not take the test” (1 point) as the lowest. The question on “intention to convey positive results to my partner” is as follows: “The genetic test revealed that you were positive for a specific genetic mutation. Would you share this result with your partner?” The question on “intention to convey positive results to your children” is as follows: “The genetic test revealed that you were positive for a specific genetic mutation. Would you tell this result to your future children?” The responses to both questions are based on a five-point scale with “I will tell them” (5 points) as the highest and “I will not tell them” (1 point) as the lowest.2. Next, a correlation analysis was conducted to examine the relationship between the three genomic literacy variables, risk perception score, benefit perception score, and behavioral intentions toward genetic testing. To investigate the effects of genomic literacy components on behavioral intention toward genetic testing, those who had a genetic disorder or a family member with a genetic disorder were excluded and a multiple regression analysis was performed, with behavioral intention towards genetic testing as the dependent variable and the three components of genomic literacy as the independent variables. Next, we created a path diagram demonstrating the relationship between the three components of genomic literacy centered on behavioral intention toward genetic testing and the risk and benefit perception of genetic testing and performed path analysis. The values of GFI, AGFI, CFI, and RMSEA were calculated as indicators of the goodness of fit of the model. The significance probability of all analyses was set to the 5% level (both sides), and IBM SPSS Statistics 25 and IBM SPSS Amos 22 were used for the statistical analyses.In the comparison of the average scores of the three components of genomic literacy, risk perception, benefit perception, and behavioral intention toward genetic testing by attribute, the effect size d was obtained by conducting a t-test on sex, age group, marital status, children, occupation, whether the respondent took biology in high school, whether the respondent had an educational background in genetics, and whether the respondent or a family member had a genetic disease. For any level of education, a one-way ANOVA was performed to obtain the effect size ηThe research company that conducted this survey has obtained the Privacy Mark and ISO/IEC27001 (ISMS) certifications from the Japan Information Processing Development Corporation (JIPDEC), and is an organization that gives due consideration to the protection of registered users’ personal information, and has obtained the Privacy Mark and ISO/IEC27001 (ISMS) certifications. The personal information of the monitors is managed by the research company, and no more personal information than necessary will be obtained by the researcher. This study has been approved by the University’s Research Ethics Committee . The purpose of the study and the handling of personal information were explained on the webpage, and responses were considered to be the acquisition of consent.At the time of the survey, the responses were collected after excluding those who responded to the straight line; thus, a total of 857 respondents were included in the analysis. The attributes of the subjects are shown in The percentages of correct answers for genetic and genomic knowledge are shown in We confirmed the ceiling effect (mean score + standard deviation ≧5) and floor effect (mean score—standard deviation ≦1) for five items on risk perception and five items on benefit perception for genetic testing. As a result, no ceiling effect or floor effect was observed for any of the five items on risk perception or the five items on benefit perception.Next, we performed a principal component analysis on five items for risk perception and five items on benefit perception for genetic testing. As a result, across the five risk perception items, the load of the first component was .734 to .789, and the contribution rate was 59.8%. In the five benefit perception items, the loading amount of the first component was 0.80 to 0.82, and the contribution rate was 65.1%. The total score was calculated for each of the five items on risk perception and the five items on benefit perception. Hereinafter, the total scores are referred to as the “risk perception score” and “benefit perception score.” The score range is 5 to 25 points in each case.To investigate the relationships between the variables of the three components of genomic literacy, risk perception scores, benefit perception scores, and behavioral intention toward genetic testing, a correlation analysis was performed . The parTo investigate the effects of genomic literacy components on behavioral intention toward genetic testing, those who had a genetic disorder or a family member with a genetic disorder were excluded and a multiple regression analysis was performed, with behavioral intention toward genetic testing as the dependent variable and the three components of genomic literacy as the independent variables . ConsequWe set up a hypothetical model where components of genomic literacy  influenced behavioral intentions toward genetic testing through risk and benefit perception, created a path diagram, and performed path analysis after excluding those who had a genetic disease or had a family member with a genetic disease.The effect on “one’s intention toward genetic testing” is as shown in For “one’s intention toward genetic testing,” the goodness-of-fit indicators showing the fit of the path diagram and data in the analysis were GFI = .994, AGFI = .960, CFI = .987, and RMSEA = .067, indicating that the model has high goodness of fit. However, the goodness-of-fit indicators of “intention to convey positive results to my partner” were GFI = .974, AGFI = .820, CFI = .925, and RMSEA = .162. The goodness-of-fit indicators of “intention to convey positive results to children” in respondents were GFI = .983, AGFI = .882, CFI = .950, and RMSEA = .128. Thus, the goodness of fit of the model was lower than that of “one’s intention toward genetic testing.”In previous studies investigating genetic/genomic knowledge and intention toward genetic testing, those with high genetic/genomic knowledge tend to want to undergo genetic testing, and it has been shown that genetic/genomic knowledge influences decision-making –33. The It is not only genetic/genomic knowledge, but also health numeracy and interactive and critical health literacy that influences behavioral intention toward genetic testing. Enhancing genetic/genomic knowledge leads to appropriate decision-making, and the genomic literacy scales , 34, 35 In addition, the genetic and genomic knowledge required for decision-making may change with rapidly advancing genomic medicine. However, health numeracy and interactive and critical health literacy are universal abilities that are not situation-dependent. From the results of this study, it can be said that health numeracy and interactive and critical health literacy are important decision-making components of genomic literacy.To clarify the process by which genomic literacy influences decision-making, a model was assumed in the present study in which the behavioral intention toward genetic testing was influenced through risk and benefit perception based on previous studies. In other words, we created a path model of intention toward genetic testing from the components of genomic literacy through the cognitive factors of risk and benefit perception and conducted path analysis to examine it. Thus, the association of the genomic literacy component → risk and benefit perception → intention toward genetic testing was found, and the model revealed high goodness of fit. This association supported the results of previous studies regarding the decision-making process , 18, 37.In this study, we reveal the path from risk perception to benefit perception to the influence on one’s intention toward genetic testing. In previous studies –40, therIn previous studies on numeracy, those with high numeracy made appropriate decisions based on the numbers themselves. However, people with low numeracy are susceptible to non-numerical information (emotions and moods) and cognitive bias , 44. In Many previous studies on health literacy have shown that those with high health literacy have a high intention to undergo cancer screening –49. AccoIn this light, it was revealed that a model in which the three components of genomic literacy influence behavioral intention through risk and benefit perception is effective for one’s own intention toward genetic testing.We believe that our study makes a significant contribution to the literature because we clarify the three components of genomic literacy  which, through the recognition of the benefits and risks of genetic testing affect the respondents’ intention to undergo such testing.We created a path model of “intention to convey positive results to partners” and “intention to convey positive results to children” from the components of genomic literacy through cognitive factors of risk and benefit perception and performed path analysis. However, the goodness of fit of the model was lower than that for “one’s intention toward genetic testing,” indicating that the hypothetical model needs to be re-examined. The reason for this is considered to be the peculiarity of the genetic information handled in genetic testing. The peculiarities of genetic information include the fact that it is information unique to a person and does not change or cannot be changed in their lifetime (invariance), and further that there is commonality regarding such information with other individuals in a genetically-linked family , 51. The 8 Feb 2023
 PONE-D-22-23128

 Impact of genomic literacy components on genetic testing decision-making in the general Japanese population in the 20s and 30s

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 After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands and requires a few minor revisions. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.Thank you for submitting your manuscript to PLOS ONE. I apologize for the delay in the review process.

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Sinapis alba L. The comprehensive collection of genetic resources represented 187 registered varieties, landraces, and breeding materials. Microsatellites generated 44 polymorphic alleles in 15 loci. Eleven of them were medium to highly polymorphic, and the high levels of observed heterozygosity (0.12–0.83) and Nei’s gene diversity index (0.11–0.68) indicated a high level of polymorphism. Based on PCoA and neighbor joining analyses, the genetic resources were divided into two groups. The range of genetic dissimilarity in the analysed collection was in the range of 0.00–1.00. The high level of dissimilarity between the accessions was documented by the high WAM value (33.82%). Bayesian clustering algorithms were performed in the STRUCTURE 2.3.4 software. The number of clusters was estimated at K = 2. The accessions were classified according to Q1/Q2 values. The low average values of the parameters Fst_1 (0.3482), Fst_2 (0.1916), and parameter alpha (0.0602) indicated substantial mating barriers between varieties and reproductive isolation due to the limited exchange of genetic resources between breeders. These results demonstrated the importance of extensive collections of genetic resources for the maintenance of genetic diversity and indicated considerable genetic differentiation among accessions.Microsatellite markers were used for the assessment of genetic diversity and genetic structure in a germplasm collection of yellow mustard,  Sinapis alba L.) is a vital crop for Czech agriculture’s food and condiment sectors. It originated in the Middle East and Mediterranean regions. Still, this crop has been produced worldwide for millennia as a condiment crop. It is extensively planted as a prominent speciality crop [S. alba exposed to elements such as metals from the thallium, arsenic, and platinum groups were collected, ensuring crop application for phytoremediation in contaminated areas [Yellow mustard  was represented by 198 unique accessions . Microsatellites generated 113 alleles and, of the total alleles, 44 were polymorphic and detected in 15 loci, with an average of 2.93 alleles per locus and a maximum of six alleles in D3_A, P7, and P35_A loci. Five loci  were classified as uninformative loci with MAF < 0.01. Only one allele was detected in these loci of one to three individuals/accessions. BoREM1b_B was successfully amplified in genotype 9  and BoREM1b_C in genotypes 51 , 167, and 177 . BolAB19TF was amplified in genotype 130 , P30_C in genotypes 169 and 186 , and P35_B in genotype 174 . This study is part of a project focused on yellow and coloured mustard, and only the markers successfully amplified in yellow mustard are presented in The wide collection of genetic resources for yellow mustard . Thus, there are genetic differences between types of genetic resources, as well as between individual accessions of yellow mustard within the analysed set of genetic resources.The results of the AMOVA  showed that the highest proportion of diversity was found within the analysed set of genetic resources (87.13%) and that the proportion of diversity and differentiation between types of genetic resources (4.6%) or between genotypes within types of genetic resources (8.27%), although significant, was always much lower . LikewisBrassica napus [The genetic diversity of the yellow mustard germplasms was calculated using DARwin 6 software. The dendrogram in ca napus , we obseca napus , probablBased on microsatellite analysis, it was possible to identify a large proportion of the analysed accessions unambiguously, but even when a relatively high number of markers were evaluated, several accessions of different origins had precisely the same patterns of microsatellite markers. There were two main groups of accessions with the same pattern of microsatellites. The first group included genotypes from Germany, Denmark, Netherlands, and Poland, but also genotypes from Australia and an old landrace from the Czech Republic/former Czechoslovakia . The second block of genotypes with the same pattern of microsatellites mainly included Czech varieties (‘Andromeda’ and ‘Severka’), as well as varieties from Germany, Austria, and the Netherlands . There were also other groups or pairs of indistinguishable accessions, such as ‘OP-2408′ and ‘OP-2415′, or ‘Olga’ and ‘OP-2291′, originating in one breeding station, but also cases have been reported where landraces have the same microsatellite profile as modern varieties, e.g., ‘Přerovská bílá’ vs. ‘Ludique’ and ‘Budakalászi sárga’, or ‘Dr. Schneider’ vs. ‘Concerta’ and ‘Esprit’. A relatively narrow genetic base should explain these results, and unrelated genotypes with an identical pattern of microsatellites are used in different breeding programmes. These findings also reveal the need for better management of breeding populations based on genetic analysis, and further improvement is also increasingly dependent on the evaluation of genetic diversity resources and breeding populations. Breeding materials from the TILLING population showed significantly higher variability and distinct differences from other genetic resources. These materials were created precisely with the goal of extending genetic variability and their further utilization in breeding programmes  belonged to the same group with closely similar Q1 values. They were bred at one breeding station, and their genetic structure reflects the significant similarity of the genetic resources used in breeding programmes. In the case of Czech varieties, both landraces , the older variety ‘Zlata’, and modern varieties from the breeding stations Opava and Krukanice belong to the same group. The second cluster comprises current modern varieties from breeding programmes in Selton and BOR (‘Agent’ and ‘Veronika’). In these cases, there is a strong influence of a limited set of genetic resources, and molecular marker methods and the evaluation of genetic diversity can help to appropriate the management of genetic resources and to spread genetic variability and shifts in genetic structures. In this study, some landraces and older varieties were not significantly different from modern varieties. However, it is still evident that the origin of the variety, country of origin, or the breeding programme significantly influences the genetic structure. Similar results were presented in oil seed brassicas  or in B.. juncea  and B. ccarinata  genetic ultivars . The lar. juncea .An essential problem in current breeding is the insufficient range of genetic diversity and the related limited availability of interesting genetic resources, such as donors of required traits and characteristics . The depPlant material and DNA extraction. For the evaluation of genetic diversity, a pilot set of 198 high-value accessions was chosen from the collection of yellow mustard kept at the OSEVA Development and Research in Opava, Czech Republic. These accessions were selected based on their value for current breeding programmes, including modern and old registered varieties, landraces, breeding material, and accessions from the TILLING population. The accessions came from various countries, representing a wide range of spring-type cultivars with a wide range of seed colours. The list of genetic resources, their accession numbers in the gene bank, and their country of origin are given in ® Mixer Mill MM400  for 30 s at a maximal frequency (30 Hz). The DNA was extracted by a modified CTAB PVP method [P method , which iMicrosatellite amplification. A total of 15 microsatellite markers were used: BoREM1b_A, BoREM1b_B, BoREM1b_C, BolAB19TF, BoPC34 [4)2 SO4, 0.01% Tween 20, 2.5 mM MgCl2, 200 µM dNTPs), 10 pmol of each primer, 0.5 U of Taq Purple DNA polymerase PPP Master Mix  or 2× Master Mix with standard buffer , 2× BSA, and 50 ng of template DNA. The amplification was carried out on a Biometra TProfessional thermocycler  in the following temperature profile: initial denaturation of 5 min at 94 °C, 31 cycles of 60 s at 95 °C, 60 s at 52–59 °C (depending on the primer used), and 60 s at 72 °C/68 °C, with a final elongation of 15 min at 72 °C/68 °C., BoPC34 , P381, D, BoPC34 . Three p, BoPC34 . The PCRData Analysis. The results of the PCR analysis were evaluated by fragment analysis on an ABI 3500 genetic analyser . Molecular data were analysed using GeneMapper version 3.0 software. The weighted arithmetic mean of genetic distances (WAM) was calculated based on frequency values in the appropriate class of genetic diversity [CT = 0), between genotypes within types of genetic resources (ΦSC = 0), or between genotypes within collection of genetic resources (ΦST = 0) [iversity . Diversiiversity ,54. Theyiversity  were periversity . The nulΦST = 0) . The datΦST = 0)  and KoláΦST = 0) . STRUCTUΦST = 0) .The issue of genetic diversity, its conservation, protection, and exploitation are the key points in plant breeding. In this study, we focused on the detailed description and molecular analysis of the genetic variability of a large collection of yellow mustard genotypes. High values of observed heterozygosity, diversity index, and WAM index indicated a high level of polymorphism between individual accessions. Also, outputs from AMOVA supported these findings and showed a low proportion of variance (4.6%) between types of genetic resources  and a high proportion of variance between genotypes within the analysed set of genetic resources (87.13%). In addition to the standard descriptive characteristics, two other procedures were applied, for an overall assessment of genetic variability and the degree of diversity of the genetic resources: PCoA analysis and Bayesian clustering algorithms. The results show a more valuable interpretation and highlight another way of processing the genetic diversity data and indicated mating barriers between varieties and reproductive isolation, due to the limited exchange of genetic resources between breeders. The range of genetic diversity was much higher than reported in other studies, concerning the large and variable collection of genetic resources. These results showed the importance of the proper management of genetic resources for the preservation of genetic diversity. Analysis of genetic structure may lead (as shown in our example) to a clarification of the grouping of accessions (genetic resources and varieties) and recommendations for expanding the genetic background in breeding programmes. The choice of parental components based on phenotypic characteristics and genetic diversity facilitates the selection of genetic resources for breeding programmes and can contribute to the success of breeding.


Urbanization and inequalities are two of the major policy themes of our time, intersecting in large cities where social and economic inequalities are particularly pronounced. Large scale street-level images are a source of city-wide visual information and allow for comparative analyses of multiple cities. Computer vision methods based on deep learning applied to street images have been shown to successfully measure inequalities in socioeconomic and environmental features, yet existing work has been within specific geographies and have not looked at how visual environments compare across different cities and countries. In this study, we aim to apply existing methods to understand whether, and to what extent, poor and wealthy groups live in visually similar neighborhoods across cities and countries. We present novel insights on similarity of neighborhoods using street-level images and deep learning methods. We analyzed 7.2 million images from 12 cities in five high-income countries, home to more than 85 million people: Auckland , Sydney , Toronto and Vancouver (Canada), Atlanta, Boston, Chicago, Los Angeles, New York, San Francisco, and Washington D.C. (United States of America), and London (United Kingdom). Visual features associated with neighborhood disadvantage are more distinct and unique to each city than those associated with affluence. For example, from what is visible from street images, high density poor neighborhoods located near the city center  are visually distinct from poor suburban neighborhoods characterized by lower density and lower accessibility . This suggests that differences between two cities is also driven by historical factors, policies, and local geography. Our results also have implications for image-based measures of inequality in cities especially when trained on data from cities that are visually distinct from target cities. We showed that these are more prone to errors for disadvantaged areas especially when transferring across cities, suggesting more attention needs to be paid to improving methods for capturing heterogeneity in poor environment across cities around the world.The online version contains supplementary material available at 10.1140/epjds/s13688-023-00394-6. Inequalities are on the rise globally and increasingly recognized as a social, political, and ethical problem –3. CitieVisual information from streetscapes have been used to study urban inequalities at least as far back as Charles Booth’s ‘Inquiry into the Life and Labour of the People in London’ in the 1880s . Urban gBooth and much of the early work on visual features of cities relied on in-person site visits which restricted the areas of the city that they could study. Large scale street-level images are a source of city-wide visual information and allow for comparative analyses of multiple cities . ComputeWe studied similarities of visual features and distinctness of urban environments of the most and least advantaged communities across 12 cities in five countries, home to more than 85 million people: Auckland , Sydney , Toronto and Vancouver (Canada), Atlanta, Boston, Chicago, Los Angeles, New York, San Francisco, and Washington D.C. (United States of America), and London (United Kingdom) . We evaluated similarity in visual features between cities by measuring how well a model trained in one city performed when making income decile predictions using images from other cities . We use We used a deep learning method to automatically recognize, i.e. learn, visual features from street images that are associated with different levels of income , 26–31. Countries differ in how city boundaries are defined based on population size and density, as well as city history, urban function, and administrative and political organization. We used definitions by national statistical offices in each of the respective countries as follows . We first created a 50 m grid for each of the cities using city boundary shape files. We then used the API to retrieve the unique panorama ids (‘panoid’) of images near each grid point acquired by Google for the years 2007-2019. A fixed number of panoids for each census tract were used prioritizing images taken in recent years. Census tracts are defined to have a similar population size for each country, yet their sizes vary across countries depending on how they are defined by statistical agencies in respective countries. We wanted our sampled number of images to be roughly proportional to the census tracts’ population. Therefore, we used 20 images from census tracts with smaller population sizes  and 30 images from census tracts from all other cities that have larger census tract populations  city were used for training. We measured how well the trained network uses images to predict outcomes in all other target  cities that were not used in training . We evalFor evaluating how well models performed when making predictions for communities within the same city, i.e., single city experiments, we used five-fold cross validation. The census tracts of a city were randomly allocated to five equally sized groups of 20%. In each fold, 80% of census tracts  were used for training the network and the remaining 20% were withheld. We then measured how well the trained network uses images to predict outcomes for census tracts that were not used in training. We repeated this process five times for each of the cities holding out a different 20% of census tracts each time.For a comparative measure of population density, we computed the number of people per km2 for each census tract. For distances to CBDs, we used coordinates for the CBD for each city from OpenStreetMaps and computed distances between the centroid point of each census tract and the CBD in kilometers.Distributions of the wealthiest and poorest census tracts over space were substantially different for Canadian cities and most American cities compared to London, Sydney, and Auckland. Figure S5 and Figure S6 show comparisons in population density and distances to the CBD in each city between the highest and lowest income neighborhoods. North American cities are characterized by highest income groups living in car-dependent low-density suburbs whereas the lower income groups live near the center with high population density. The poor tend to live close to the city center as highlighted by the right-skewed distributions in Figure S5. Most of the highest income groups live farther away from the center as they have access to cars and ability to afford costs of private transport, and hence can afford to live farther from the center. Population density distributions  for high income groups are much narrower, which is intuitive as they have more opportunities to choose their neighborhoods to live in their preferred density. This is consistent with the suburbanization and car-oriented development history of American and Canadian cities, where high income people who can afford cars chose to move outside the city to consume more land  alongsidThese patterns, which will also have visual signs captured by street-level images, are reversed for London, Sydney, and Auckland where it is mostly the high-income groups that live closer to CBDs and the lower income groups father away from the center. Land near CBDs is valuable, and it is mostly the rich who can afford to live there as affordable housing in the center of these cities are scarce. The distributions of population density for the best-off 10% and worst-off 10%, as shown in Figure S6, are mostly overlapping where both income segments live in similar areas with respect to their density. In these cities, especially for London, dense, walkable, transit-oriented neighborhoods are also attractive for high-income segments of the population. San Francisco and New York had somewhat different patterns from other US cities, and similar to London, where instead many high-income neighborhoods are located near the center, populated by people who prefer to live near well-paying jobs and urban amenities.We tested whether within city differences in population density and distance to city center as experienced by the wealthiest and poorest deciles would explain poorer transferability performance between city pairs. To do this, we needed a quantitative metric to summarize ‘the difference in differences’, comparing the difference in density and distance to CBD between the richest and poorest deciles for each city. We used the test-statistic of a two-sample t-test for each city and each metric; large positive or negative values of the test-statistic indicate strong evidence of a difference between richest and poorest deciles in that city, smaller values indicate weak or no evidence for a difference.s and t are source and target cities respectively. CSC is the city specific constant to be estimated, β are parameters to be estimated. ddensity is the difference between source and target city t-statistics computed for population density. ddistance is the difference between source and target city t-statistics computed for distance to CBD. We then carried out a regression analysis (Eq. ) using tTolstoy famously said “Happy families are all alike; every unhappy family is unhappy in its own way .” In thePoor areas in all cities display signs of disrepair and poor condition of pavements, roads, homes, and gardens, but differ in other features such as building design. As examples, Fig. Wealthier residents can afford to live in neighborhoods with better amenities , as a rePoor and rich communities looked more similar to their counterparts in other cities in the same country, than to those in cities in different countries. Models trained on data from an US city made better predictions in other US cities than in non-US cities, as was the case for Canadian cities. The same pattern was observed between Sydney and Auckland, which are in the pacific region although not in the same country. These cities share similarities in their climates, cultures, social and political history, and architecture. We also hypothesized that neighborhoods in big cities with populations of more than 5 million each  would be more similar. However, we did not observe this from our results.In addition, poor prediction performances for Auckland, Sydney, London, and Atlanta when transferring from other source cities  are consistent with observed differences in spatial patterns described in Sect. We considered differences across cities between the highest and lowest income deciles in two well-defined and quantifiable features described above: their population density and distances to the city’s central business district (CBD), as predictors of inter-city similarity. Both features have direct visual signs, such as more buildings, cars, and less sky view, that are captured by street images. To summarize and quantify the role of differences and similarities observed in Figure S5 and Figure S6, we conducted regression analyses Fig. . Indeed,Between city MAEs were regressed against an indicator for whether the predicted and training cities were in the same country, and differences in population density and distances to city center between the corresponding two cities (Eq. ). The teRegression results revealed that a moderate share of variation in transferability errors  varied across cities ranging from r = 0.17 for Vancouver to r = 0.81 for London and Atlanta . In Canada, many highly educated immigrants cannot get high paying jobs as their credentials are not recognized . Similar yet less pronounced dynamics are in play for both Sydney and Auckland. When we repeated the same set of analyses with street images using educational attainment instead of income, we found even higher visual similarity between areas with highest educational attainment. Lowest educational attainment tracts were even more different from each other  have more of an influence on high-skill workers’ decisions of where they live compared to low-income workers where the priority is on rents and wages , 38. TraPlato’s statement from more than 2300 years ago, that “any city, however small, is in fact divided into two, one the city of the poor, the other of the rich,” still holds true today as neighborhood experiences of the wealthiest and poorest residents differ widely even when they live in the same city , 41. To In previous work, visual elements in street images, including window and roof styles, number and types of cars, store signs, trees, fire escapes, and road widths, were found to be correlated with socioeconomic variables in city or country specific studies , 20, 21.We find that, only some visual elements associated with poverty or wealth in one city only generalize to others. Similarities as well as differences across cities are driven by historical factors, policies, and local geography. Car-oriented development often results in lower density especially when geography allows for city expansion. Poor suburban neighborhoods often have green space visible from images. However, in such cities, poor neighborhoods often lack access to sufficient levels of urban amenities, affordable and sustainable transport options. On the other hand, cities that have higher shares of public transit use are often higher in density. From what is visible from street-level images, high-density poor neighborhoods lack trees or green space in proximity. In addition, residents in such neighborhoods may be more exposed to hazards related to traffic such as higher levels of pollution, and road collisions. They may also suffer from poorer municipal services including waste collection. Further, residents of such neighborhoods may enjoy higher levels of access to urban amenities, but the type and quality of amenities may be poor and inadequate for the population density . While we used the full dataset containing all deciles from all cities during training, we used the best-off and worst-off deciles for our evaluations relating to the wealthiest and poorest areas in each city in line with the focus of the paper. Similarities or differences in other deciles and their transferability across cities and countries, which are arguably harder to interpret, are not part of our analysis.Cities are dynamic and their visuals change with poor and rich residents moving in and out of neighborhoods, city investments and regulations. History of migration and gentrification also affect visual similarities, where new social groups settle into existing neighborhoods changing their social fabric and environmental quality. For example, land ownership changes over time where the wealthy migrate according to changing opportunities , sometimes pushing out the poor . FeatureMethodologically, we found that transferability errors were higher for poverty compared to that of wealth. This finding raises the need for caution in using imagery for measurements of socioeconomic status for data-poor geographies . SpecifiIn high-income countries, inequalities are increasingly concentrated in urban areas , 45. WhiSupplementary information (PDF 8.0 MB)Below is the link to the electronic supplementary material.


New plant breeding techniques may play an important role in improving food quality, global food security and sustainability. Previous breeding techniques have, however, met with substantial resistance from society. This study examined the role of associations and deliberation in the evaluation of breeding techniques. Breeding techniques studied included conventional breeding, gene-editing, genetic modification (cisgenesis and transgenesis), marker-assisted breeding and synthetic biology. By using focus group discussions that included individual tasks, we found that when participants relied on their spontaneous associations, gene-editing was evaluated similarly as genetic modification. However, after information provision and group discussion, gene-editing was preferred over genetic modification. Perceived naturalness was found to be the main reason for obtaining different levels of acceptance, not only between gene-editing and genetic modification but across all breeding techniques examined. These findings highlight the importance of associations and show that beliefs about naturalness remain crucial in understanding how consumers evaluate breeding techniques. Global food security is under substantial pressure . It is eConfédération paysanne and Others v Premier ministre and Ministre de l’agriculture, de l’agroalimentaire et de la forêt Case C528/16 that gene-editing is a form of genetic modification. This implies that the use of gene-editing for food production falls under the same strict regulations as genetically modified foods. Hence, according to the European Court of Justice, the European legislation does not differentiate between these different types of gene-based breeding, even though there are technical distinctions that separate gene-editing from earlier genetic modification techniques. It remains however unclear, to what extent the opinion of the European Court of Justice reflects the opinion of the public. That is, it remains unclear whether the public also perceives these techniques as similar or whether they perceive these techniques as distinctively different.The distinction between genetic modification and gene-editing is particularly relevant since the An important distinction between gene-editing and genetic modification is that certain subtypes of gene-editing can block specific genes without inserting exogenous DNA . The insPrior research suggests that consumers have limited knowledge of breeding techniques and therefore often rely on quick responses that determine their attitude towards these techniques . Quick rThe aim of this study is to explore (1) whether new breeding techniques have different levels of acceptance compared to the current breeding techniques, (2) the role of associations in the evaluation of breeding techniques and (3) how these associative responses compare to deliberative responses. Studied techniques range from conventional breeding and gene-based selection techniques to genetic modification, gene-editing and synthetic biology. Specifically, we focus on how genetic modification compares to a (sub)type of gene-editing that solely blocks genes (site-directed nuclease 1 generations). To keep the distinction between gene-editing and genetic modification clear to participants, this study does not focus on other (sub)types of gene-editing that can insert exogenous genes such as site-directed nuclease 3 . GainingConsumers often rely on associative processes when evaluating a product, person or situation . Associagene-editing appears to be both conceptually and semantically related to genetic modification. Semantically, the similarity between ‘genetic’ from genetic modification and ‘gene’ from gene-editing will likely activate similar associations. The words ‘modification’ and ‘editing’ are not semantically similar; however, both words represent a concept of (human) intervention that may activate conceptually similar associations.Associations can be semantically and conceptually related to the presented topic. Semantic associations are related to representation of the topic in words, signs and symbols . ConceptAssociations are highly context-dependent . DiffereWhile initial associations may play an important role in shaping judgement, they are not necessarily decisive in the final evaluation. The extent to which associations are activated and influence the final judgement depends on the amount of deliberation people use when making a judgement . DeliberIn summary, associations are expected to be important in the evaluation of new breeding techniques due to semantic and conceptual similarity between associations retrieved from memory. Similar associations may lead to similar evaluations. Associations are context-dependent and therefore the food product type may influence acceptance. Finally, deliberative reasoning may lead to different evaluations compared to association-based evaluations.n = 6–8 each) were conducted, two each in three European countries , to gain insights in the range of associations with and deliberations on breeding techniques. The countries represented the North-Western, Mediterranean and Central European regions. Each focus group was conducted in the local language and moderated by a native speaker. Focus groups were audio recorded after consent of all participants. The study received ethical approval from the Social Science Ethics Committee of Wageningen University & Research.Six face-to-face focus groups ), Italy ), and Czech Republic ). To prevent participants from searching for information before the start of the focus group, they were told that the topic would be food-related but were not informed that the discussion would be about breeding techniques used for food/crop production. For each focus group, quotas were set for gender, age and education to create a diverse group that represents a broad set of views. We limited the number of participants with food allergies and/or dietary preferences to a maximum of two participants per focus group session, to prevent personal concerns becoming a majority view in the discussions. Participants were rewarded according to the agreements made with the marketing recruitment agencies in the different countries.A total of 45 participants were recruited by marketing recruitment agencies across three European countries: the Netherlands  genetic modification, (2) cross-pollination, (3) gene-editing and (4) synthetic biology. As we did not want to provide information at this stage, the distinction between cis and trans-genetic modification techniques was omitted. A booklet was prepared in which each individual participant wrote down the words that spontaneously came to mind when thinking of each breeding technique, with a maximum of 10 words per technique. Subsequently, participants selected one of the reported words which best reflected their concept of each breeding technique. Participants evaluated each word on how that word made them feel on a 5-point scale ranging from ‘very negative’ to ‘very positive’. Finally, participants provided an (associative) overall evaluation of the breeding technique on another 5-point scale. Together, two associative evaluation scores were obtained: (1) the evaluation of each (association-based) word mentioned and (2) the (associative) overall evaluation of each breeding technique. This process was repeated 4 times such that each breeding technique was evaluated. Because we were interested in spontaneous associations that would directly come to mind, participants were given a time limit of 15 minutes in total. Pre-test revealed that booklets were understandable but that many people had no associations with the term Supplemental materials for all information provided). The final leaflet contained a ranking form which had to be used for the ranking task that followed the reading task. The following six breeding techniques were described on the leaflets: (1) cisgenesis (variant of genetic modification), (2) transgenesis (variant of genetic modification), (3) conventional breeding, (4) CRISPR-Cas9 (subtype of gene-editing), (5) synthetic biology  and (6) marker-assisted breeding (non-invasive breeding tool). We specified cisgenesis  and transgenesis (where foreign genes from other species are used) as variants of genetic modification given these techniques substantially differ from each other and are regularly compared genetic modification techniques in social sciences  explore the range of perceptions, deliberations and preferences towards different breeding techniques through group discussion; (2) raise group discussion about the likes and dislikes of different breeding techniques and (3) have a group discussion on the implementation of different breeding techniques in food products to explore the practical relevance and, given the contextual dependency of associations, whether acceptance is in part product-specific.The final evaluation task aimed to gain insights into the stability of the associative evaluation and to what extent evaluations changed due to the information provided and the discussion raised. The final evaluation task was an individual task that consisted of an evaluation form to evaluate the six breeding techniques discussed. On the evaluation form, participants had to evaluate each technique one final time on a 5-point scale ranging from ‘very negative’ to ‘very positive’.online Supplemental materials.All materials were prepared in English and translated into local languages by a native speaker. Pre-tests confirmed all materials  to be understandable. The English version of the booklet, the information leaflets, the ranking task form and the final evaluation form can be found in the Participants were welcomed to the focus group and offered refreshments. The moderator and participants introduced themselves by mentioning their name and, as icebreaker, their favourite dish. Thereafter, the moderator discussed the ground rules and provided participants an informed consent form. The informed consent form included consent for participation, recording and anonymised data collection and was to be signed if agreed. Participants were then handed out the booklet and asked to complete the associative evaluation task. This was followed by a short break after which participants were provided with the information leaflets about the six breeding techniques. When finished reading the information leaflets, participants did the ranking task. Thereafter, the focus group discussion started (stage 2), which lasted approximately 1 hour. After the discussion, participants were asked to make a final evaluation of the breeding techniques discussed (stage 3). Finally, participants were thanked for their participation, debriefed and booklets were collected. The entire procedure lasted approximately 1.5 hour.All associations mentioned in the individual booklets were translated into English and summarised in two ways. First, frequency tables were created for the words that were literally written down with each breeding technique. A cut off point of at least three words was used as indication of generally occurring associations, given fewer words would capitalise too much on individual opinions cf. . Second,https://doi.org/10.17026/dans-xbg-kkbr. Finally, means were calculated and reported as illustration of the final evaluation. Given the small sample size, no formal statistical analyses were deemed relevant.For the rankings, cross tabulations were calculated with the breeding techniques as rows and the rankings as columns. Audio recordings of the group discussions were transcribed verbatim and translated into English. Transcripts were coded into emergent codes using Atlas.Ti 9 (version 9.1.6.0). Full (anonymised) data can be accessed at artificial, which might have been due to a carry-over effect. Synthetic biology was associated with both the concepts of artificiality  and nature . When comparing the most frequently written down words between the three genetic breeding techniques , all genetic breeding techniques appear to be associated with similar concepts. Six out of eight most mentioned words with gene-editing were similar to the most mentioned words with genetic modification. The majority of (most) mentioned words with genetic breeding techniques were related to the concepts of genetics, artificial, modification or research. In contrast, cross-pollination was most frequently associated with concepts related to nature. None of the most mentioned words with cross-pollination were related to the top mentioned words of gene-editing and genetic modification, except for the word When categorising all words written down, it appears that from the genetic breeding techniques, genetic modification was associated more frequently with food/agriculture compared to gene-editing and synthetic biology. In contrast to genetic modification and gene-editing, synthetic biology was less associated with the concept of modification. Furthermore, gene-editing elicited substantially less associations in total compared to genetic modification and raised more associations with complexity and with DNA/genes. In contrast to the genetic breeding techniques, cross-pollination was highly associated with concepts related to nature. In addition, participants hardly associated cross-pollination with DNA/Genes, science, or with the concept of modification. Categorised associations with synthetic biology once more showed that synthetic biology was associated with both artificiality and nature. An overview of all categories and their frequency of being associated with each breeding technique is shown in When comparing the means of the overall associative evaluations, cross-pollination was evaluated most positively. Furthermore, results show a negligible difference in evaluation between genetic modification and gene-editing (mean genetic modification of 3.24 vs mean gene-editing of 3.26). A similar pattern is shown when comparing the average evaluation of all the words written down per breeding technique (mean genetic modification of 3.48 vs mean gene-editing of 3.49). Synthetic biology received the lowest associative evaluation. An overview of the mean associative evaluation and the mean score of all words per breeding technique is shown in Results from the ranking task show patterns of different levels of acceptance across breeding techniques, with most participants preferring conventional breeding over the genetic breeding techniques .I had it [conventional breeding] on top because it is the most natural, without any manipulation. Actually, ah well, just as it always was. Across all participants, 58% ranked conventional breeding as most preferred breeding technique. The reasoning behind this was that in almost every case, conventional breeding was perceived as the most natural, or least invasive, method:It [conventional breeding] is less invasive of nature also possibly . . . that is . . . the words [conventional breeding] are closer to us. The other words [genetic breeding techniques], as terms, are distant and even [conventional breeding] gives more the idea of pollination by bees, as to say like flowers in history. Additionally, to this we also use the word conventional , it helps to assimilate the concept. Associative responses were also observed when participants explained their reasoning for preferring conventional breeding. Some participants explained how the words associated with conventional breeding influenced their perception and therefore their preference:By exclusion, I would have put [conventional breeding] as an average, but the others inspired me more. It is nothing new, nothing advanced, all the others are new techniques. But on the other side, from a breeder perspective, it is a problem that conventional breeding takes so much time. When participants deviated from the majority and preferred genetic breeding techniques over conventional breeding, their preferences were based on multiple reasons. Some participants perceived greater opportunities with genetic breeding techniques compared to conventional breeding, whereas others focused on perceived disadvantages of conventional breeding:In my opinion it [marker-assisted breeding] is a great technology, but it seemed to me to have little to do with the others where there is talk of an alteration of plant genes, while this is a technology that could be used together with the others, but which in itself does not change anything. A breeding technique that stood out in terms of ranking was marker-assisted breeding. In general, participants evaluated marker-assisted breeding positively; however, it was rarely selected as the most preferred breeding technique. Perceptions of marker-assisted breeding were related to research and controlling risks, which was perceived beneficial. Moreover, participants highly valued marker-assisted breeding being non-invasive. Yet, marker-assisted breeding was clearly perceived as an additional tool rather than a breeding technique, which was the primary reason for not ranking it as the most preferred technique:It [synthetic biology] is too invasive for the plant and for us. Regarding the least preferred breeding techniques, participants were clear. Approximately half the participants ranked synthetic biology as least preferred breeding technique and 83% of participants ranked synthetic biology in the lower bracket. The reasoning behind this was mostly similar to the reasoning for ranking conventional breeding as most preferred. That is, synthetic biology was perceived as the most invasive and least natural breeding technique:synthetic and biology represent opposite concepts. Specifically, focussing on each word in isolation, the associations participants had with ‘biology’ were related to nature, whereas the associations with ‘synthetic’ were related to artificiality. Responses by participants indicated that combining these concepts creates tension, leading to a negative evaluation:It [synthetic biology] seems like a contrast: biology is the study of life and synthetic is an opposite juxtaposition. Participants’ perceptions were often strengthened by their conflicting associations with the semantics of synthetic biology. According to participants, the words I think this [CRISPR-Cas9] is the least tampering, the easiest modification in the way that they are trying to erase the bad genes that can affect that the food will spoil easily, as my colleague here said and so on. Most participants preferred gene-editing over genetic modification, whereas approximately one-third preferred genetic modification over gene-editing or had no clear preference. The reasoning for preferring gene-editing techniques was similar to the reasoning for preferring conventional breeding methods. That is, gene-editing was perceived less invasive, less artificial and it is not expected to lead to the development of new species as it was described in terms of gene blocking:blocking:And somehow, uhm, is CRISPR also inside my head applied to people. That they use it [CRISPR-Cas9] on babies to remove things. That I believe is also CRISPR-Cas I think, so I have a really negative association with this. Most of the times when genetic modification was preferred over gene-editing, it was not due to a heuristic decision rule such as naturalness, but to negative associations with gene-editing. For some participants, the associations they had with gene-editing were related to scariness. Other participants had negative associations with the word I feel like it [cisgenesis] is less difficult than CRISPR-Cas, where you have to basically eliminate all of the bad qualities that you do not want. Because, like, how many of them would you have to delete in order to produce a good result? Additional reasons for preferring genetic modification over gene-editing appeared to be similar to preferring genetic breeding techniques over conventional breeding. That is, a minority of participants preferred genetic modification over gene-editing due to opportunities, effectiveness and/or efficiency benefits:When asking participants about the implementation of different breeding techniques in food products, the level of acceptance varied depending on the type of food product. The exception holds for synthetic biology, which was perceived unacceptable regardless of the type of food product. Food categories that were mentioned to affect participants’ level of acceptance were (un)natural/processed, (un)healthy and/or hedonic food products. The application of genetic breeding techniques on products that were perceived as natural was deemed less acceptable compared to processed products. Natural products that were mentioned were products such as vegetables, fruits and eggs. For processed hedonic products that were perceived unhealthy participants showed greater acceptance of genetic breeding techniques: Hedonic products that were mentioned were products such as chips, pizza and cola.‘Are there any food products where it matters more to know that they were made like this [genetic breeding techniques]? . . .’ Everyone: ‘yes’ . . . Participant: ‘Fruit, vegetables and dairy products’. This thing [CRISPR-Cas9] of artificially correcting the defects that nature imposes, is a remarkable thing and will also bring a quality product that will have a balanced taste at the table. Although there was general agreement on most issues, some differences between the countries emerged. In line with Italian food culture, Italians raised taste as an important aspect:I always try to look where the product comes from. If there’s a possibility to buy a product from the Czech Republic, I will do that to support local products. Participants from the Czech Republic attributed great value to the country of origin of the products they buy:A lot of testing etc. . . And it probably can go wrong on the long term, you do not know that either. Just like with, yes with the Corona vaccine, for example. You do not know that either. So that will take time. . . I guess? Finally, the Dutch showed associations with the COVID-19 vaccine in both focus groups, whereas the Italians and Czechs did not mention anything related to COVID. This could have been influenced by the timing of the focus group , or due to the associations that Dutch participants have with genetic modification and the COVID-19 vaccine, such as long-term risk effects:When comparing the final evaluation with the associative evaluation, we observe different evaluations. Both conventional breeding and gene-editing were evaluated more positively. Cisgenesis was evaluated slightly more positively compared to the primary associative evaluation of genetic modification, whereas transgenesis and synthetic biology were evaluated less positively than their associative response. This study shows different levels of acceptance across a spectrum of breeding techniques. Perceived naturalness appears to be the main differentiation when evaluating different breeding techniques. This is in line with previous research that suggests naturalness to be an important heuristic when evaluating breeding techniques . In thisA technique that stood out was marker-assisted breeding. Responses to marker-assisted breeding show that using DNA technology to select breeds based on genetic composition is not perceived negatively, even if the technology to do so is distinctly unnatural. We argue that the distinction lies in the fact that marker-assisted breeding is not invasive in the natural process of breeding itself. Hence, we argue that naturalness as a heuristic  is used Prior research has shown that associations are of great importance in the evaluation of genetic modification . This stActivation and evaluation of conflicting associations influence opinions on breeding techniques. Breeding is related to crops, which in turn are associated with nature . While cThe perception of naturalness is more than a shallow association. Naturalness is a deeply rooted belief , that isThe difference between associative evaluations and final evaluations shows the instability of initial judgements or attitudes when expressed without a substantial amount of knowledge . FollowiOpinions on gene-editing were different. They started with the same negative associations with artificiality as genetic modification, yet gene-editing was ultimately perceived as more positive. The deliberative line of reasoning that gene-editing is less invasive compared to genetic modification could be an explanation for this change from the initial attitude. Due to the information provided and the discussion raised, participants perceived gene-editing as less invasive compared to genetic modification. Given their initial limited knowledge of gene-editing, the new knowledge may have changed their relatively unstable concept of gene-editing, which appeared to be quite similar to genetic modification beforehand. Hence, deliberating on information and discussion caused participants to differentiate between techniques, which may have led to the accommodation of knowledge . In contOur study also sheds some light on how consumers in general deliberate on topics of which they have limited knowledge. During deliberation, we observed that additional associations became active, which were then used to justify initial associative responses. These additional associations were only indirectly related to the information provided. Deliberation appeared to trigger a deeper exploration of the associative network until participants were sufficiently confident about their opinion based on these associations. That is, associations activated additional associations, which in turn triggered larger networks that include heuristics and beliefs learned from prior experience. As a result, more information was retrieved from memory, leading to greater attitudinal certainty. Hence, we argue that the deliberation process operated more as a meta-cognitive process steering towards certainty in associative response, rather than a separate, logic-based reasoning process on the topic in hand.On a practical level, our findings show differences in acceptance across a spectrum of breeding techniques ordered on perceived naturalness and invasiveness. When using these techniques for societally relevant purposes such as climate change and global food security, it may be useful to consider using a less invasive technique to gain greater acceptance and less resistance. Furthermore, especially when advocating gene-editing, it is important to provide consumers with information on the technique. Due to the retrieval of semantically similar associations derived from the ‘gene’ word element and conceptually similar associations from the ‘editing’ word element, gene-editing and genetic modification are likely to be perceived similar when no information about these techniques is provided. This also stresses the importance of the naming of breeding techniques, as ‘unlucky’ name choice may set the public against these techniques . Our finFor this study, we conducted six focus groups across three European countries. Although the sample size was limited, the reduced additional insights with each consecutive group suggest theoretical saturation regarding the range of perceptions. In all countries, we observed the same pattern in which a majority of participants mentioned naturalness as the most important driver for preferring conventional breeding over genetic breeding techniques. In addition, supporters of genetic breeding techniques also showed similar patterns in reasoning across countries. That is, supporters of genetic breeding techniques focused on either abstract benefits described as ‘opportunities’ or disadvantages of conventional breeding such as ‘time-consuming’. We found only minor differences in reasoning between countries such as the importance of taste , the country of origin (Czech Republic) and COVID associations (the Netherlands).Providing information about multiple techniques and asking participants to rank these techniques may have influenced opinions and is thus a limitation of this study. In this study, such comparison was crucial to foster an appropriate discussion about the breeding techniques. In reality, consumers often do not have the information at hand and will often only consider one technique at a time. In addition, the exact contents of the information may have influenced the evaluation, although in this study the extent and direction in which the evaluation changed did not necessarily follow the information provided. To disentangle these explanations, in future research the effects of deliberation and information should be studied in isolation. Another limitation of the information provided is the number of variants used. We limited the discussion to six techniques, and for gene-editing we only included CRISPR-Cas9 with a focus on blocking genes. There are, however, many more techniques particularly in the emerging field of gene-editing, several of which can also insert exogenous DNA (e.g. site-directed nuclease 3 generations). We decided to exclude these options as we wanted to present the differences between a limited set of breeding techniques in such way that it would be understandable and manageable in the context of a focus group discussion without causing confusion between techniques. It is conceivable that including the insertion of genes as possibility for gene-editing would lead to different comparison with, in particular, genetic modification. Further study is needed to establish to what extent our findings reflect opinions on gene-editing in general, or that the difference between blocking and insertion within gene-editing options matters.Consumers show different levels of acceptance across a spectrum of breeding techniques. Associations and deliberation on information play an important role in both the evaluation of breeding techniques and in the extent to which consumers develop different levels of acceptance. When relying on associations, gene-editing and genetic modification are at first similar, while only with information provision and group discussion, consumers differ in acceptance of these techniques. After information and discussion, conventional breeding remains the most preferred option, followed by gene-editing, genetic modification and, finally, synthetic biology. These preferences rely on the perception that invasive techniques are unnatural and therefore disliked.Click here for additional data file.Supplemental material, sj-pdf-1-pus-10.1177_09636625231168087 for Breeding by intervening: Exploring the role of associations and deliberation in consumer acceptance of different breeding techniques by Paul Nales and Arnout R.H. Fischer in Public Understanding of Science