Different control of resistance to two Colletotrichum orbiculare pathogenic races 0 and 1 in cucumber (Cucumis sativus L.).

KEY MESSAGE: Quantitative trait locus analysis identified independent novel loci in cucumbers responsible for resistance to races 0 and 1 of the anthracnose fungal pathogen Colletotrichum orbiculare. Cucumbers have been reported to be vulnerable to Colletotrichum orbiculare, causing anthracnose disease with significant yield loss under favorable conditions. The deployment of a single recessive Cssgr gene in cucumber breeding for anthracnose resistance was effective until a recent report on high-virulent strains infecting cucumbers in Japan conquering the resistance. QTL mapping was conducted to identify the resistance loci in the cucumber accession Ban Kyuri (G100) against C. orbiculare strains 104-T and CcM-1 of pathogenic races 0 and 1, respectively. A single dominant locus An5 was detected in the disease resistance hotspot on chromosome 5 for resistance to 104-T. Resistance to CcM-1 was governed by three loci with additive effects located on chromosomes 2 (An2) and 1 (An1.1 and An1.2). Molecular markers were developed based on variant calling between the corresponding QTL regions in the de novo assembly of the G100 genome and the publicly available cucumber genomes. Multiple backcrossed populations were deployed to fine-map An5 locus and narrow the region to approximately 222 kbp. Accumulation of An2 and An1.1 alleles displayed an adequate resistance to CcM-1 strain. This study provides functional molecular markers for pyramiding resistance loci that confer sufficient resistance against anthracnose in cucumbers.

Evaluation of the Influence of Genetic and Environmental Factors on the Ephedrine Alkaloids Content of Ephedra sinica.

Ephedra herb, a dried terrestrial stem of Ephedra sinica, is used in traditional Japanese medicine (Kampo) and Chinese medicine to treat the common cold, headaches, bronchial asthma, and nasal inflammation. E. sinica predominantly contains two ephedrine alkaloids-(-)-ephedrine and (+)-pseudoephedrine-which are crucial for its medicinal effects. This study aimed to reveal the influence of genetic and environmental factors on ephedrine alkaloids content using statistical genetic analyses. To evaluate the influence of genetic factors on ephedrine alkaloids content, 25 clonal lines were cultivated in Ibaraki and the broad-sense heritability of the traits was estimated. The heritabilities of (-)-ephedrine, (+)-pseudoephedrine, and "total alkaloids" (TA) content were 0.871, 0.969, and 0.865, respectively. The heritabilities of ephedrine alkaloids content were high. To evaluate the influence of environmental factors on ephedrine alkaloids content, four clonal lines which have different genotypes were cultivated in three locations (Ibaraki, Shizuoka, and Yamanashi prefectures). The effects of genotype (G), location (L), and genotype by environment (G × E) interactions on ephedrine alkaloids content were found to be significant (p < 0.05) by two-way ANOVA, and, in particular, the genotypic effects were found to be the largest. Our results indicate that the ephedrine alkaloids content in E. sinica is under relatively strong genetic control and remains stable under various environments. These findings suggest that E. sinica with a higher and stable ephedrine alkaloids content could be cultivated in different locations through selective breeding.

Identification of novel sex determination loci in Japanese weedy melon.

KEY MESSAGE: Japanese weedy melon exhibits unique sex expression with interactions between previously reported sex determination genes and two novel loci. Sex expression contributes to fruit quality and yield in the Cucurbitaceae. In melon, orchestrated regulation by sex determination genes explains the mechanism of sex expression, resulting in a great variety of sexual morphologies. In this study, we examined the Japanese weedy melon UT1, which does not follow the reported model of sex expression. We conducted QTL analysis using F2 plants for flower sex on the main stem and the lateral branch and mapped "occurrence of pistil-bearing flower on the main stem" locus on Chr. 3 (Opbf3.1) and "type of pistil-bearing flower" (female or bisexual) loci on Chr. 2 (tpbf2.1) and Chr. 8 (tpbf8.1). The Opbf3.1 included the known sex determination gene CmACS11. Sequence comparison of CmACS11 between parental lines revealed three nonsynonymous SNPs. A CAPS marker developed from one of the SNPs was closely linked to the occurrence of pistil-bearing flowers on the main stem in two F2 populations with different genetic backgrounds. The UT1 allele on Opbf3.1 was dominant in F1 lines from crosses between UT1 and diverse cultivars and breeding lines. This study suggests that Opbf3.1 and tpbf8.1 may promote the development of pistil and stamen primordia by inhibiting CmWIP1 and CmACS-7 functions, respectively, making the UT1 plants hermaphrodite. The results of this study provide new insights into the molecular mechanisms of sex determination in melons and considerations for the application of femaleness in melon breeding.

Prospects and potentials of underutilized leafy Amaranths as vegetable use for health-promotion.

Climate change causes environmental variation worldwide, which is one of the most serious threats to global food security. In addition, more than 2 billion people in the world are reported to suffer from serious malnutrition, referred to as 'hidden hunger.' Dependence on only a few crops could lead to the loss of genetic diversity and high fragility of crop breeding in systems adapting to global scale climate change. The exploitation of underutilized species and genetic resources, referred to as orphan crops, could be a useful approach for resolving the issue of adaptability to environmental alteration, biodiversity preservation, and improvement of nutrient quality and quantity to ensure food security. Moreover, the use of these alternative crops will help to increase the human health benefits and the income of farmers in developing countries. In this review, we highlight the potential of orphan crops, especially amaranths, for use as vegetables and health-promoting nutritional components. This review highlights promising diversified sources of amaranth germplasms, their tolerance to abiotic stresses, and their nutritional, phytochemical, and antioxidant values for vegetable purposes. Betalains (betacyanins and betaxanthins), unique antioxidant components in amaranth vegetables, are also highlighted regarding their chemodiversity across amaranth germplasms and their stability and degradation. In addition, we discuss the physiological functions, antioxidant, antilipidemic, anticancer, and antimicrobial activities, as well as the biosynthesis pathway, molecular, biochemical, genetics, and genomic mechanisms of betalains in detail.

Colletotrichum orbiculare strains distributed in Japan: race identification and evaluation of virulence to cucurbits.

The pathogen Colletotrichum orbiculare is causal fungus of cucurbit anthracnose. Multiple races have been identified in the United States, suggesting that it is necessary to cultivate suitable resistant cultivars and breed new cultivars with the most suitable resistance gene. This study examined the pathogenicity and virulence of 20 strains in Japan to clarify the existence of races and virulence differences. Based on the symptoms on inoculated cotyledons and true leaves of watermelon, we could evaluate the compatibility of each strain to each host cultivar. Our analysis based on the reaction to the host cultivar harboring the resistance gene Ar-1 (Cla001017) revealed the existence of three races in Japan. An alarming result was that a race that overcame Ar-1, which is a target gene in current watermelon breeding in Japan, is present in Japan. The cucumber and melon host cultivars showed diverse symptoms, whereas a squash cultivar was resistant to all strains. Three strains caused severe damage even to the most resistant cucumber cultivar 'Ban Kyuri' and resistant cultivars harboring Cssgr, a well-known gene conferring loss-of-susceptibility resistance. Screening genetic resources for novel resistance genes using strains with high virulence is of vital importance for watermelon, cucumber, and melon production.

Molecular Analysis of Genetic Diversity and Structure of the Lablab (Lablab purpureus (L.) Sweet) Gene Pool Reveals Two Independent Routes of Domestication.

In this study, genetic diversity and structure of 474 cultivated and 19 wild lablab (Lablab purpureus) accessions. were determined using 15 nuclear and 6 chloroplast SSR markers. The overall gene diversity was relatively low (0.3441). Gene diversity in the wild accessions (0.6059) was about two-folds greater than that in the cultivated accessions. In the wild accessions, gene diversity was greatest in the southern Africa, followed by East Africa. In the cultivated accessions, gene diversity was highest in the eastern Africa. The results suggested that South Africa is the center of origin and East Africa is the center of domestication of lablab. Different cluster analyses showed that 2-seeded-pod cultivated accessions (ssp. uncinatus) were clustered with wild accessions and that 4-(6)-seeded-pod cultivated accessions (ssp. purpureus and bengalensis) were intermingled. UPGMA tree suggested that ssp. purpureus and bengalensis were domesticated from 4-seeded-pod wild accessions of southern Africa. Haplotype network analysis based on nuclear SSRs revealed two domestication routes; the ssp. uncinatus is domesticated from 2-seeded-pod wild lablab (wild spp. uncinatus) from East Africa (Ethiopia), while the ssp. purpureus and bengalensis are domesticated from 4-seeded-pod wild lablab from Central Africa (Rwanda). These results are useful for understanding domestication and revising classification of lablab.

Stability and Reproducibility of Ephedra sinica Ephedrine Alkaloid Content and Terrestrial Stem Dry Weight.

Dried terrestrial stems of Ephedra sinica are known as 'Ephedra herb.' The pharmacological effects are mainly related to two major ingredients, (-)-ephedrine and (+)-pseudoephedrine (total alkaloids which are defined in Japanese Pharmacopoeia, TA). In this study, in order to aid in cultivation and breeding, the stability of TA content and stem dry weight of 46 E. sinica genets was evaluated from the first year of transplantation to the sixth year. TA content and composition ratio of these genets were stable after the second year, and dry weight was stable after the fourth year. These traits showed high inter-genet variability but low annual variability for each genet. Additionally, rank correlation coefficients of each trait among the genets were high. There was no significant correlation between these traits. Furthermore, to assess the reproducibility of these traits in clones, we evaluated TA content and dry weight of three clonal lines with high TA contents. TA content and composition ratio of the clonal lines were also stable after the second year of transplantation, and dry weight of the clonal lines was also stable after the fourth year. Moreover, TA content and composition ratio in each clonal line were comparable with those of each original genet after the second year. These results suggested that ephedrine alkaloids content and dry weight of E. sinica plants are stable, and that these traits are highly reproducible in clones. Therefore, selection breeding of E. sinica using vegetative propagation can be effective for high and stable quality of Ephedra herb.

Identification of quantitative trait loci for powdery mildew resistance in highly resistant cucumber (Cucumis sativus L.) using ddRAD-seq analysis.

Powdery mildew, caused by Podosphaera xanthii (syn. Sphaerotheca fuliginea ex Fr. Poll.), is one of the most economically important foliar diseases in cucumber (Cucumis sativus L.). Cucumber parental line 'Kyuri Chukanbohon Nou 5 Go', developed from weedy cucumber line CS-PMR1, is highly resistant to powdery mildew and is promising breeding material. We performed quantitative trait locus (QTL) analysis using double-digest restriction-site-associated DNA sequencing (ddRAD-Seq) in a population from a cross between 'Kyuri Chukanbohon Nou 5 Go' and the Japanese native cultivar 'Kaga-aonaga-fushinari', which is susceptible to powdery mildew. The resistance of the population and its parents was evaluated using leaf disc assays and image analysis. We detected one major QTL on Chr. 5 that was effective at both 20°C and 25°C and one minor QTL on Chr. 1 effective at 20°C. We detected two additional QTLs in subpopulation: one on Chr. 3 effective at 20°C and one on Chr. 5 effective at both 20°C and 25°C in a position different from the major QTL. The resistance alleles at all four QTLs were contributed by 'Kyuri Chukanbohon Nou 5 Go'. The results of this study can be used to develop practical DNA markers tightly linked to genes for powdery mildew resistance.

Validity of Selection Breeding for High Ephedrine-Alkaloid Content in E. sinica under Different Environments in Japan.

Dried terrestrial stems of Ephedra sinica are called 'Ephedra herb,' whose pharmacological effects are due mainly to two major ingredients, (-)-ephedrine and (+)-pseudoephedrine (total alkaloids which are defined in Japanese Pharmacopoeia (TA)). Ephedra herb is an important crude drug in Japan. However, E. sinica is widely distributed in arid areas of northeastern China and Mongolia. Recently, E. sinica has started to be cultivated in Japan. This study aimed to assess the validity of selection breeding on TA content of E. sinica in several locations in Japan. In this experiment, we grew approximately 350 seedlings and divided them randomly into seven groups. Nearly fifty plants were cultivated at each of seven locations. In Ibaraki, Yamanashi, and Shizuoka, average TA content of whole samples satisfied the criteria for Ephedra herb defined in Japanese Pharmacopoeia (7.0 mg/g of dry weight (DW)). Plants with high and intermediate TA content at four locations were selected and transplanted to Ibaraki. There were significant differences in TA content between selected plants with high and intermediate TA content before and after transplanting (p < 0.05). TA content of high-TA plants was significantly higher than that of control plants cultivated continuously at Ibaraki (p < 0.05). These results suggest that the selection on content of ephedrine alkaloids in E. sinica under various locations in Japan is valid, and high- TA E. sinica plants can be selected at various locations.

Genetic diversity and population structure of feral rapeseed (Brassica napus L.) in Japan.

Rapeseed (Brassica napus L.) is one of the most economically important oilseed crops worldwide. In Japan, it has been cultivated for more than a century and has formed many feral populations. The aim of this study was to elucidate the genetic diversity of feral rapeseeds by genotyping 537 individuals (among which 130 were determined to be genetically modified) sampled from various regions in Japan. Analysis of 30 microsatellite markers amplified 334 alleles and indicated moderate genetic diversity and high inbreeding (expected heterozygosity, 0.50; observed heterozygosity, 0.16; inbreeding coefficient within individuals, 0.68) within the feral populations. The Mantel test showed only an insignificant weak positive correlation between geographic distance and genetic distance. Analysis of molecular variance showed a greater genetic diversity among individuals than between populations. These results are in accordance with population structure assessed by using principal coordinate analysis and the program STRUCTURE, which showed that the 537 individuals could be assigned to 8 genetic clusters with very large genetic differences among individuals within the same geographic population, and that among feral individuals, many are closely related to rapeseed accessions in the NARO Genebank but some have unknown origins. These unique feral rapeseeds are likely to be affected by strong selection pressure. The results for genetically modified individuals also suggest that they have two different sources and have a considerable degree of diversity, which might be explained by hybridization with nearby individuals and separation of hybrid cultivars. The information obtained in this study could help improve the management of feral rapeseed plants in Japan.

Genetic diversity of leafy amaranth (Amaranthus tricolor L.) resources in Vietnam.

Leafy amaranths, which are consumed as traditional food in Asia and Africa, are now considered among the most promising vegetables. In Vietnam, leafy amaranths, particularly Amaranthus tricolor L., are important summer vegetables due to their excellent nutritional values and high tolerance to biotic and abiotic stresses. However, this species has not been subjected to systematic breeding. Here we describe species identification and evaluation of the genetic diversity of Vietnamese amaranth collection by using matK and simple sequence repeats (SSR) markers. Our phylogenetic analysis based on the matK marker classified the species of 68% of the accessions, of which 120 belonged to A. tricolor. We developed 21 SSR markers, which amplified a total of 153 alleles in 294 A. tricolor accessions originating from Vietnam and overseas, with a mean allelic richness of 7.29 per marker, observed heterozygosity of 0.14, expected heterozygosity of 0.38, and polymorphic information content of 0.35. The STRUCTURE and FST analysis indicated a positive relationship between geographic distance and genetic differentiation among most of the overseas groups and the Vietnamese collection, but not among geographic groups within the Vietnamese collection. Vietnamese amaranths could be divided into two major types, one common in East Asia and the other one unique to Vietnam.

Analysis of genetic diversity of rapeseed genetic resources in Japan and core collection construction.

Diversity analysis of rapeseed accessions preserved in the Japanese Genebank can provide valuable information for breeding programs. In this study, 582 accessions were genotyped with 30 SSR markers covering all 19 rapeseed chromosomes. These markers amplified 311 alleles (10.37 alleles per marker; range, 3-39). The genetic diversity of Japanese accessions was lower than that of overseas accessions. Analysis of molecular variance indicated significant genetic differentiation between Japanese and overseas accessions. Small but significant differences were found among geographical groups in Japan, and genetic differentiation tended to increase with geographical distance. STRUCTURE analysis indicated the presence of two main genetic clusters in the NARO rapeseed collection. With the membership probabilities threshold, 227 accessions mostly originating from overseas were assigned to one subgroup, and 276 accessions mostly originating from Japan were assigned to the other subgroup. The remaining 79 accessions are assigned to admixed group. The core collection constructed comprises 96 accessions of diverse origin. It represents the whole collection well and thus it may be useful for rapeseed genetic research and breeding programs. The core collection improves the efficiency of management, evaluation, and utilization of genetic resources.

Gene pyramiding enhances durable blast disease resistance in rice.

Effective control of blast, a devastating fungal disease of rice, would increase and stabilize worldwide food production. Resistance mediated by quantitative trait loci (QTLs), which usually have smaller individual effects than R-genes but confer broad-spectrum or non-race-specific resistance, is a promising alternative to less durable race-specific resistance for crop improvement, yet evidence that validates the impact of QTL combinations (pyramids) on the durability of plant disease resistance has been lacking. Here, we developed near-isogenic experimental lines representing all possible combinations of four QTL alleles from a durably resistant cultivar. These lines enabled us to evaluate the QTLs singly and in combination in a homogeneous genetic background. We present evidence that pyramiding QTL alleles, each controlling a different response to M. oryzae, confers strong, non-race-specific, environmentally stable resistance to blast disease. Our results suggest that this robust defence system provides durable resistance, thus avoiding an evolutionary "arms race" between a crop and its pathogen.

On plant detection of intact tomato fruits using image analysis and machine learning methods.

Fully automated yield estimation of intact fruits prior to harvesting provides various benefits to farmers. Until now, several studies have been conducted to estimate fruit yield using image-processing technologies. However, most of these techniques require thresholds for features such as color, shape and size. In addition, their performance strongly depends on the thresholds used, although optimal thresholds tend to vary with images. Furthermore, most of these techniques have attempted to detect only mature and immature fruits, although the number of young fruits is more important for the prediction of long-term fluctuations in yield. In this study, we aimed to develop a method to accurately detect individual intact tomato fruits including mature, immature and young fruits on a plant using a conventional RGB digital camera in conjunction with machine learning approaches. The developed method did not require an adjustment of threshold values for fruit detection from each image because image segmentation was conducted based on classification models generated in accordance with the color, shape, texture and size of the images. The results of fruit detection in the test images showed that the developed method achieved a recall of 0.80, while the precision was 0.88. The recall values of mature, immature and young fruits were 1.00, 0.80 and 0.78, respectively.

Mutation of FVS1, encoding a protein with a sterile alpha motif domain, affects asexual reproduction in the fungal plant pathogen Fusarium oxysporum.

Fusarium oxysporum produces three kinds of asexual spores: microconidia, macroconidia and chlamydospores. We previously analysed expressed sequence tags during vegetative growth and conidiation in F. oxysporum and found 42 genes that were markedly upregulated during conidiation compared to vegetative growth. One of the genes, FVS1, encodes a protein with a sterile alpha motif (SAM) domain, which functions in protein-protein interactions that are involved in transcriptional or post-transcriptional regulation and signal transduction. Here, we made FVS1-disrupted mutants from the melon wilt pathogen F. oxysporum f. sp. melonis. Although the mutants produced all three kinds of asexual spores with normal morphology, they formed markedly fewer microconidia and macroconidia than the wild type. The mutants appeared to have a defect in the development of the conidiogenesis cells, conidiophores and phialides, required for the formation of microconidia and macroconidia. In contrast, chlamydospore formation was dramatically promoted in the mutants. The growth rates of the mutants on media were slightly reduced, indicating that FVS1 is also involved in, but not essential for, vegetative growth. We also observed that mutation of FVS1 caused defects in conidial germination and virulence, suggesting that the Fvs1 has pleiotropic functions in F. oxysporum.

Use of image analysis to estimate anthocyanin and UV-excited fluorescent phenolic compound levels in strawberry fruit.

Strawberry is rich in anthocyanins, which are responsible for the red color, and contains several colorless phenolic compounds. Among the colorless phenolic compounds, some, such as hydroxycinammic acid derivatives, emit blue-green fluorescence when excited with ultraviolet (UV) light. Here, we investigated the effectiveness of image analyses for estimating the levels of anthocyanins and UV-excited fluorescent phenolic compounds in fruit. The fruit skin and cut surface of 12 cultivars were photographed under visible and UV light conditions; colors were evaluated based on the color components of images. The levels of anthocyanins and UV-excited fluorescent compounds in each fruit were also evaluated by spectrophotometric and high performance liquid chromatography (HPLC) analyses, respectively and relationships between these levels and the image data were investigated. Red depth of the fruits differed greatly among the cultivars and anthocyanin content was well estimated based on the color values of the cut surface images. Strong UV-excited fluorescence was observed on the cut surfaces of several cultivars, and the grayscale values of the UV-excited fluorescence images were markedly correlated with the levels of those fluorescent compounds as evaluated by HPLC analysis. These results indicate that image analyses can select promising genotypes rich in anthocyanins and fluorescent phenolic compounds.

Ability of bumblebees to discriminate differences in the shape of artificial flowers of Primula sieboldii (Primulaceae).

BACKGROUND AND AIMS: Flower shapes are important visual cues for pollinators. However, the ability of pollinators to discriminate between flower shapes under natural conditions is poorly understood. This study focused on the diversity of flower shape in Primula sieboldii and investigated the ability of bumblebees to discriminate between flowers by combining computer graphics with a traditional behavioural experiment. METHODS: Elliptic Fourier descriptors described shapes by transforming coordinate information for the contours into coefficients, and principal components analysis summarized these coefficients. Using these methods, artificial flowers were created based on the natural diversity of petal shape in P. sieboldii. Dual-choice tests were then performed to investigate the ability of the bumblebees to detect differences in the aspect ratio of petals and the depth of their head notch. KEY RESULTS: The insects showed no significant ability to detect differences in the aspect ratio of the petals under natural conditions unless the morphological distance increased to an unrealistic level. These results suggest the existence of a perception threshold for distances in this parameter. The bumblebees showed a significant preference for narrow petals even after training using flowers with wide petals. The bumblebees showed a significant ability to discriminate based on the depth of the petal head notch after training using artificial flowers with a deep head notch. However, they showed no discrimination in tests with training using extreme distances between flowers in this parameter. CONCLUSIONS: A new type of behavioural experiment was demonstrated using real variation in flower corolla shape in P. sieboldii. If the range in aspect ratios of petals expands much further, bumblebees may learn to exhibit selective behaviour. However, because discrimination by bumblebees under natural conditions was low, there may be no strong selective behaviour based on innate or learned preferences under natural conditions.

Bayesian association mapping of multiple quantitative trait loci and its application to the analysis of genetic variation among Oryza sativa L. germplasms.

One way to use a crop germplasm collection directly to map QTLs without using line-crossing experiments is the whole genome association mapping. A major problem with association mapping is the presence of population structure, which can lead to both false positives and failure to detect genuine associations (i.e., false negatives). Particularly in highly selfing species such as Asian cultivated rice, high levels of population structure are expected and therefore the efficiency of association mapping remains almost unknown. Here, we propose an approach that combines a Bayesian method for mapping multiple QTLs with a regression method that directly incorporates estimates of population structure. That is, the effects due to both multiple QTLs and population structure were included in our statistical model. We evaluated the efficiency of our approach in simulated- and real-trait analyses of a rice germplasm collection. Simulation analyses based on real marker data showed that our model could suppress both false-positive and false-negative rates and the error of estimation of genetic effects over single QTL models, indicating that our model has statistically desirable attributes over single QTL models. As real traits, we analyzed the size and shape of milled rice grains and found significant markers that may be linked to QTLs reported previously. Association mapping should have good prospects in highly selfing species such as rice if proper methods are adopted. Our approach will be useful for the whole genome association mapping of various selfing crop species.

Analysis of petal shape variation of Primula sieboldii by elliptic fourier descriptors and principal component analysis.

BACKGROUND AND AIMS: Petals are important for Primula sieboldii because of the commercial value of its flowers, and their form is a target characteristic for breeding. An appropriate understanding of petal form in terms of genetic mechanisms and environmental effects is necessary for improvement of this species. The aim of this study was to establish a quantitative evaluation method of petal shape by elliptic Fourier descriptors and principal component analysis (EF-PCA), and thus to investigate genotypic and environmental effects on petal morphology. METHODS: EF-PCA describes an overall shape mathematically by transforming coordinate information concerning its contours into elliptic Fourier descriptors (EFDs) and summarizing the EFDs by principal component analysis. To examine varietal effects on principal component (PC) scores and petal area among commercial varieties, nested ANOVAs were performed (since the samples had a hierarchical structure with four sources, i.e. variety, plant, flower and petal). KEY RESULTS: Petal shape variation could be evaluated successfully and the symmetrical and asymmetrical elements of the overall shape variation could be detected. The proportions of the variance component due to varietal differences were more than 70 % in the first five PCs of the symmetrical elements and petal area. By contrast, the proportions due to varietal effects of all PCs of the asymmetrical elements were less than 20 %, and the proportions of the variation within a flower were more than 75 %. It was also demonstrated that the yearly variance of petal shape was small, and that of petal area was large. CONCLUSIONS: Within a flower the major source of the symmetrical elements is genotypic and the asymmetrical elements are strongly affected by the environment. With respect to petal area, the contribution of genotypes is also large; it is, however, affected by the macro-environment more notably than is petal shape.