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International audience; The 'pollen test' and 'fruit set test' following controlled crossing combinations of parents are the most commonly used methods for pollination incompatibility studies in Olea europaea L. Self-incompatibility (SI), with diagnoses based on the pollen test and pollen germination, indicating self-compatibility, is not always followed by fruit set in this species. To solve this dispute, we have reconciled all observations into a new model. Mismatches between field and laboratory data and between methods are resolved by the dual-successive-screen model (DSSM) supposing two different loci for the expression of the two SI mechanisms. Pollen/stigma is controlled by diallelic SI, or DSI, inferring two G1 and G2 compatibility/incompatibility (C/I) groups for varieties, whereas pollen tubes in ovaries are controlled by poly-allelic SI or PASI with twenty C/I groups. To explain the selfing of varieties, we have suggested that some determinants in the pollen tube and stigma are unstable and degrade (DS-D for degradation of S-determinant) after three to five days, enabling some pollen tubes to avoid being rejected, hence reaching ovules. DSI and PASI in the DSSM and DS-D mechanisms, plus the andromonoecy of the olive tree, complexify SI studies. Inferences from DSSM and DS-D mechanisms in olive orchard practice are detailed.

AGAP : équipe DAVEM; International audience; Olive fruit production is dependent on wind pollination, and recently sporophytic self-incompatibility has been advocated to function with six S-alleles. The aim here is to show fruit setting data gathered from literature can be interpreted and their standardization help to release firm conclusions on the S-allele pair carried by most varieties. Under a bag, fruit settings for mates do not allow to infer inter-compatibility of pairs of mates. Each variety displays a different number of flowers per inflorescence, and for a given proportion of hermaphrodite flowers, fruit sets per hermaphrodite flowers were calculated. Thus, broad fruit set data were standardized per 100 hermaphrodite flowers. The ratio fruit sets from self-pollination and fruit sets from crosses are commonly referred to fruit sets after open pollination, as the index of self-incompatibility (ISI) and the index for cross inter-compatibility (ICOI), respectively. Open pollination rate (OP-R) is considered to be the best rate of fruit sets for a variety. ICOI values were classified with the threshold at 0.6 to rank a pair of varieties as inter-incompatible and inter-compatible. Before, we have proceeded by trials and errors. Here we showed that rationale reasoning is efficient to analyze fruit settings. Computations on ICOI showed that matings that succeeded are more confident than matings that failed, because pollen efficiency appeared insufficient. Computations on ISI revealed high correlations between SI and co-dominant S-allele pairs as for SC and S-allele pairs sharing dominance relationships. Thus, the sporophytic self-incompatibility (SSI) model will help geneticists to unravel the S-allele series and to predict suitable pollinizers in olive orchards. Moreover, compensation for OP-R deficiency in orchards could be achieved by supplementing suitable pollinizers computed in silico.

International audience; The new self-incompatibility system (SI) was presented by Saumitou-Laprade, Vernet, Vekemans et al. (2017). Evolutionary Applications based on 89 crosses between varieties in the olive tree. Four main points are not clear. We are examining here as follows: (i) the assertion that the self-incompatibility system is sporophytic was not sustained by pollen germination data; (ii) surprisingly, the new model does not explain that about one-third of pairwise combinations of olive varieties leads to asymmetric fruit setting; (iii) DNA preparation from one seed may contain two embryos, and thus, embryos should be separated before seed extraction; (iv) although effective self-fertility in olive varieties was reported by many studies, the DSI model fails to explain self-fertility in some olive varieties. Moreover, we cannot discuss result data, as science cannot be verified because variety names were encoded, this does not allow comparison of data with previous works. The DSI model on olive self-incompatibility should explain more features than the model based on four dominance levels shared by six S-alleles. Perspectives for orchard management based on this model may face serious limitations. An olive variety does not have a fifty percent chance of cross-incompatibility, but surely fewer, and thus, the sporophytic system limits fruit production. Evolutionary perspectives of self-incompatibility in Oleaceae should include data from the Jasmineae tribe that displays heterostyly SI.

Fruit setting after self-pollination, crosses and free-pollination appears to be erratic in the cultivated olive tree [Olea europaea subsp europaea L. (O. e. europaea L.)] because of a lack of a suitable model to enable prediction of rates. The same lack of prediction also applies to the wild taxon Olea europaea subsp cuspidata (O. e. cuspidata). Because of their close phylogenetic relationships, we hypothesize that O. e. cuspidata and cultivated olive share the same self-incompatibility system. We used data recently published in a wide study involving four O. e. cuspidata accessions and four olive cultivars. Because the olive varieties have been deciphered for their S-allele pair, that infer determinants present in the stigma and pistil, and that coat the pollen, we deciphered the S-alleles carried by three of the O. e. cuspidata accessions. Data are too scarce and the number of accessions too small to speculate on the O. e. cuspidata genetic population structure. The working hypothesis is confirmed. This study and data from the Italian team will enable us to embark on a large-scale hybridization program between the two subsp. to obtain a wide range of progenies for screening for responses to cold, diseases and pests.

International audience; Self-fertility is largely decreased and even prevented by various mechanisms because, broadly, it causes inbreeding depression, although some species have retained self-reproduction regimes. Species of plants that display the self-incompatible sporophytic type of self-incompatibility may rarely self-pollinate. It is only possible in the absence of foreign compatible pollen. In the olive tree with a sporophytic mechanism, we will show that three co-dominant S-alleles R1, R3 and R5 do not lead to the same level of self-fertility. All varieties that carry R1 are less self-fertile than those that carry R5, whatever the other S-alleles, while those carrying R3 are intermediate. S-allele pair-wise combinations that differ by two or three levels of dominance, and not the other combinations allow self-fertility, and moreover each S-allele R1, R3 and R5 decreases, maintains and enhances the self-fertility rate, respectively.

Contact: berville@supagro.inra.fr; The genetic base of sunflower elite lines is very narrow, due to many years of selection and breeding. To broaden the genetic diversity of the cultivated sunflower, in 1995 73 wild sunflower populations were crossed with 3 cultivated lines (Testers), and 219 hybrid offspring's were evaluated in the field. GCA and SCA effects were computed suggesting for all traits a genetic potential for improvement through selection. Study of the hybrids revealed that the wild accessions bear different genetic abilities to combine with the testers for traits of morphological architecture, phenology and yield (seed weight and seed oil). The variance due to GCA and SCA showed that gene action was additive for days to flowering, branching and plant height. Genotypes derived from the same geographic origin may have either good or poor general combing ability. The correlation between GCA and per se genotype performance was positive for all traits except for seed oil content. This was the first attempt to evaluate wild-cultivated hybrids in sunflower on a large scale and will be the starting point for the management of hybrid Helianthus annuus populations for breeding. GCA and SCA estimations will facilitate the definition of strategies to manage and exploit the natural diversity for this crop

Sunflower (2n=17) belongs to the Helianthus genus (Asteraceae). Wild Helianthus species display morphological variation for branching and stem number, for architecture and seed size, and for resistance to abiotic and biotic stresses due to which they thrive in different environments in North America. The genus is divided into botanical sections, two for annual as sunflower, and two for perennial species as Jerusalem artichoke that produces rhizomes (tubers). We explain the difficulties and successes obtained by crossing sunflower with these species to improve the agronomic traits of the sunflower crop. It is easier to cross the annual species than the perennials’ with sunflower. Several traits such as Cytoplasmic male sterility and restorer Rf-PET1 genes, Downy mildew resistance, Phomopsis resistance, Sclerotinia resistance, Rust resistance, and Orobanche resistance have already been introduced from annual species into sunflower crop, but the complex genomic organization of these species compared to sunflower limits their important potential. Perennial species are much more diverse, and their genomes display 2n, 4n, or 6n chromosomes for n 17. The realities of inter-specific hybridization are relatively disappointing due to the introgression lines that have low oil and low seed yield. We report here several attempts to introgress agronomic traits from these species to sunflower, and we present as a case study, an introgressed progenies from H. mollis, a diploid species with sessile small leaves. We constructed a preliminary genetic map with AFLP markers in 21 BC1 plants, and we then showed that some progenies display 6 to 44% of introgression from H. mollis. Although this study is promising due to the novel compact architecture of the progenies, we cannot estimate the transferability from H. mollis to other perennial Helianthus to improve sunflower.

The olive is widely cropped in Tunisia where also oleaster trees thrive all around orchards and in natural sites. Little is known on the genetic relationships between the olive crop and oleaster trees in Tunisia. Fifty-two oleaster trees and fifteen cultivars were sampled from Tunisia. SSR genotyping was performed in polyacrylamide gels after fluorescent labeling. We used seven nuclear and two chloroplast SSR markers. AFC analyses showed close genetic relationships between cultivated and oleaster trees. Genetic relationships were also displayed in a dendrogram based on Unweighted Pair Group Method (UPGMA). Five clusters were defined mixing cultivar and oleaster trees suggesting close relationship between some cultivar and some oleaster trees. One oleaster is single in a cluster. The chlorotype SSR markers show probably three olive origins. Some cultivars have the CE chlorotype originates from the East of the Mediterranean basin, the CCK haplotype originates from Maghreb and the COM chlorotype originates from West Mediterranean. The cultivars were 1) introduced from the East; 2) selected in the West; 3) or selected in the North Africa region. The Tunisian oleaster trees carry eastern and western Mediterranean chlorotype CCK, COM and CE.

To determine whether oil accumulation pattern is parallel to drupe olive (Olea europaea L) growth and if common climatic parameters may influence oil content we conducted an experiment in rainfed orchards with four olive cultivars, Amygdalolia, Arbequina, Lucques, and Oliviere, differing by fruit size at maturity. Fruits were harvested weekly from July to November. They were counted and weighted before being crushed. Fat content was determined on dry matter using a Minispec RMN. Common climatic parameters were recorded. Variance analyses showed stage effects highly significant. Results showed three different patterns for fruit growth. Dry matter accumulated broadly similarly and the weekly rates were positively correlated with fruit size. Oil accumulation is mostly independent of cli- matic variation and probably depends on genetic programmes for each cultivar. We defined the main steps and events for olive fruit ripening according to recent knowledge on fruit development.

E-mail Addresse: andre.berville@supagro.inra.fr Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; The olive (Olea europaea) and the wattles (mimosas or Acacia spp.) are typical tree species of the Mediterranean and Australia, respectively. Both trees have been subject to trans-oceanic movements, including the introduction of Mediterranean olives to Australia, and the introduction of Australian wattles to the Mediterranean. In each case, the trees have naturalised and become problematic invasives. Based on case studies of such movements, we compare and contrast the processes of transfer, dispersal, and genetic changes of these trees, with particular attention to crop-feral-wild complexes in both their home ranges and in introduced areas. Contrasting ecological, social, and economic landscapes shape the possibilities for crop-feral-wild interactions and for invasiveness

SUMMARY The Pervenets mutation raises the oleic acid content (OAC) from about 25% (LO linoleic sunflower) to over 75% (HO sunflower) in sunflower seed oil whatever the genetic backgrounds of the genotypes: fixed mutant lines, hybrid between fixed mutant lines and, surprisingly, both direction cross hybrids between high oleic × linoleic and linoleic × high oleic lines. Here we develop the methodology to point out the coincidence between the mutation and the oleate-desaturase locus. The mutation function is modeled to explain the phenotype. A series of sunflower genotypes developing embryos between 12 and 24 days after pollination without or carrying the Pervenets mutation was used to compare oleate-desaturase transcript accumulation. This revealed the presence and absence of the transcript, in normal LO and Pervenets HO embryos, respectively, whereas stearate-desaturase transcript accumulation was equivalent in both types. The RFLP approach using oleate-desaturase cDNA as a probe has revealed that in comparison with the linoleic sunflower, those carrying Pervenets mutation displayed, besides a common 5.8 kb EcoRI fragment, an extra 8 kb EcoRI fragment and, with HindIII, the shift from 8 kb to 16 kb of a HindIII fragment. The common 5.8 kb EcoRI fragment to Pervenets and normal sunflower carries an oleate-desaturase gene. The insertion unique to the sunflower carrying Pervenets mutation has to carry oleate-desaturase sequences. The genetic studies of this mutation have lead to contradictory results in the inheritance pattern of the HO trait due to variable mutation expression and several genetic factors affecting the HO level. Thus, it appeared important to study the inheritance in a set of recombinant inbred lines. Moreover, we pointed out that the presence of the mutation is not sufficient to induce the HO phenotype since a suppressor of the mutation “supole” may mask the HO trait. The supole allele is revealed in half of the HO RI lines, and consequently its accurate mapping is difficult. Thus conjunction of the molecular and genetic methods and of material construction was required to solve this problem and to provide efficient tools to breed HO sunflower.

A genetic map was constructed with specific PCRs, DALPs and AFLPs using F8-generation sunflower recombinant inbred lines. RI lines generated from a F2 population of one cross between the two cultivated inbred lines HA89 (maintainer for Pet1 CMS) and LR4 (restorer for Pet1 CMS) were used. A total of 305 markers were located using seven sPCR, 64 DALP and 301 AFLP loci. They were generated with one, seven and 14 primer pairs, respectively. The map construction consisted of a two-step strategy using 6 and 3.1 LOD scores revealed by a simulation file. Mapped markers were assembled into 18 linkage groups covering 2,168.6 cM with an average of 6.1 cM. The distribution of DALPs and AFLPs revealed that both markers tagged different regions to enable covering most of the sunflower genome. This leads to the longest map published so far for sunflower.

SUMMARY The effect of osmotic and water stresses on various morphological traits was studied in eight experimental varieties of sunflower bred for Moroccan conditions or issued from interspecific hybridization with H.argophyllus or H.debilis. Seed yield of the same genotypes was also evaluated under rainfed and irrigated field conditions. Shoot and root growth were significantly reduced by osmotic stress (-0.6 and -1.0 MPa) induced with polyethylene glycol (PEG 6000). Considerable genotype variation was expressed for all traits. Significant genotype by stress interaction was observed for root dry weight and volume as well as seed yield. The morphological traits were not related to seed yield in the correlation and principal component analyses. Four genotypes issued from introgression with H.argophyllus were grouped and distinct from other genotypes according to hierarchical ascending classification. One genotype issued from introgression with H.debilis and three non-introgressed sunflower varieties were found to be very distinct from each other and from those introgressed with H.argophyllus.

Previous papers have dealt with olive chloroplastic DNA (cpDNA) variation revealed using several methods (RFLPs, PCR-RFLPs and microsatellites) and have led to different conclusions. This paper aims to reconsider these divergences. A Southern approach was applied to reveal polymorphism. We used chloroplast DNA of Phillyrea media as a probe. Based on these data, only four chlorotypes were identified in the olive complex. The number of detected lineages was lower than reported in the literature using a direct cpDNA RFLP approach, and was insufficient to distinguish the North African subspecies europaea, maroccana, guanchica and laperrinei. Furthermore, one individual considered belonging to the subspecies laperrinei was questionable. Using other cpDNA and mitochondrial DNA (mtDNA) polymorphisms - based on PCR and RFLP methods, respectively - we showed that this individual displays the cytoplasmic lineage CE1-ME1 characteristic of most Eastern mediterranean cultivars and of Olea europaea subsp. laperrinei from Hoggar. However, based on RAPDs, this individual appeared as mislabelled and probably corresponded to a Mediterranean cultivar or a feral form. In addition, we checked O. e. subsp. laperrinei herbarium samples using two cpDNA microsatellites, which revealed polymorphisms. These also supported that both populations from Niger and Algeria displayed a chlorotype related to CE1. Consequently, based on cpDNA, the relationships of O. e. subsp. laperrinei from Hoggar with a Mediterranean lineage appeared well supported, whereas the South West Moroccan and Macaronesian olives appeared in a different clade using both mtDNA and cpDNA polymorphisms. We conclude that methods based on PCR reveal more polymorphisms in the cpDNA and lead to more-reliable results that the classical RFLP method.

Apart from the wild species P. orientalis and P. occidentalis, the cultivated plane trees constitute a wide and heterogeneous group, with uncertain genetic status and largely debated names. The recent canker stain problem in Europe makes it necessary at the present time to consider the genetic resources and to determine the genetic bases of all these trees. To attain this objective, a genetic molecular approach was used to analyze 60 trees of P. orientalis and P. occidentalis,different London planes (P. hispanicaand P. densicoma), a few controlledP. occidentalis × P. orientalishybrids and particular trees from arboreta and old parks. Molecular analysis involved thirty RAPD fragments generated with nine primers, PCR-RFLP in the 5S RNA genes and mitochondrial polymorphisms revealed by RFLP method. Clones were recognized amongP. hispanica and P. densicomatrees. A Correspondence Analysis and a dendrogram constructed according to the genetic distances confirmed the supposed hybrid origin of P. hispanica and P. densicoma between P. occidentalisand P. orientalis. Contribution ofP. orientalis to their constitution seems more important than that of P. occidentalis. Mitochondrial DNA polymorphisms indicated that crosses occurred in both directions. Moreover, P. occidentalis as female parent led toP. densicoma whereas P. orientalis as female parent led to P. hispanica. Low prevalence of pure species individuals and confusion risks with hybrid trees even for old trees are highlighted.

SUMMARY This work aims to clone the genomic region of the sunflower genome that may carry the Pervenets mutation causing high oleic content of seed oil. We previously reported that the oleHOS allele at the oleate-desaturase locus (oleHL) carries or is genetically closely linked to Pervenets mutation. We pointed out that the fragments that border the 5.8 kb EcoRI fragments carry the Pervenets mutation. The oleHOS reference physical map was constructed. It displays a common part made of the 5.8 kb EcoRI fragment, also present in oleLOR allele, and a specific region with the 7.9 kb EcoRI fragment, also carrying oleate-desaturase sequences. A genomic library was constructed in λfixII with an average insert size of 15 kb. Two millions clones were screened enabling to isolate clones from Group I, which were entirely sequenced and revealed carrying a gene for an oleate probably located in the RE. This corresponds to the invariant part of the oleHL locus. Two clones of Group II (11.1, 27.1) are overlapping but 11.1 is characterized by new specific restriction fragments and instability leading to a smear when probed with the oleate cDNA. Consequently, 11.1 sequence probably carries oleate repeated sequences that cause instability of the clone. The clone 11.1 is a good candidate to carry a part of the specific oleHOS allele, but due to its organization in repeats it is not yet sequenced. This organization leads to speculate on the mechanisms that could disturb oleate function in Pervenets [HOAC] genotypes.

The self-incompatibility type is of key importance to understanding pollination in orchards, because most olive cultivars are partially self-incompatible and thus require pollinizers to ensure fruit set. The gametophytic model has been advocated to function in the olive, but no allele pair has been attributed to any variety. The GSI model failed in most combinations to explain fruit set. Olive growers must screen experimentally and empirically to look for inter-compatible pair-wise combinations of varieties for optimum pollination. The sporophytic model, with given dominance relationships for six S-alleles matches 98 % of the experimental data of the two sets investigated. We propose a method to analyze data from controlled crosses between olive cultivars applied to two experiments for varieties crossed in a diallel design. Furthermore, the dominance between the S-allele pair allows rational prediction of olive variety self-incompatibility levels. The S-allele pairs were unraveled for more than 60 cultivars. To go further, crosses between reference varieties—those in which the S-allele pair was unraveled—and varieties under experimentation (VarE) with an unknown S-allele pair will enable an increase in knowledge and the choice of the best pollinizers in silico. Nevertheless, we pose outstanding questions in orchards where open-pollination efficiency with varieties harboring the R2R3, R1R3, R1R5, or R3R5 pairs. These S-allele pairs require pollen grains without R2 or R3 , R1 or R3, and R3 or R5 determinants. Such pollinizer varieties are not abundant in France and Italy, and this questions whether their spread is sufficient for optimal pollination of main varieties.

SUMMARYAn experiment has been carried out in order to study the behaviour under mineral deficiency of three sunflower genotypes, a population variety (Oro 9) and two hybrids (Mirasol and Albena). Sunflower seedlings were submitted to five treatments: N deficiency (N0), P deficiency (P0), K deficiency (K0), N and K deficiency (N0K0) and a control. Plants were harvested when they reached 3-4 true pairs of leaves. Growth parameters measured (height, total leaf area, root length, root and shoot dry mater) were all significantly reduced by mineral deficiency. Leaf area was most reduced by N0 (-61%) and P0 (-56%). Total dry matter was most affected by N0 (-63%) and by N0K0 (-66%). Genotype comparisons showed that Oro 9 had the highest shoot dry matter while Albena had the lowest root dry matter. Effect of mineral deficiency on content and partitioning of N, P, K, Ca and Na was significant and varied according to treatments and among plant parts. Shoot dry weight was significantly correlated with root N content (r2=0.81) and root K content (r2=-0.61) for N0 and K0.

SUMMARYIn arid and semi arid areas, water is the most limiting factor of crop production. Germination is susceptible to water shortage. Experimentation was undertaken to study the effect of water stress on germination of seeds of three sunflower genotypes (Oro 9, Mirasol and Albena). Six PEG 6000 osmotic constraints were tested (0, -0.4, -0.6, -1, -1.2 and -1.6 MPa). Results showed no significant difference between germination percent of the control and -0.4 MPa treatment and between -0.8, -1 and -1.2 MPa treatments. Germination percent was most reduced at -1.6 MPa (-65%). Mean germination duration was also affected and the highest value was obtained for -1.6 MPa (4.95 days, related to an increase of 185%). Differences between genotypes were significant in the treatment at -1 MPa.

Pervenets is a sunflower mutant with a seed oil oleic acid content greater than 65%. It was obtained after mutagenesis treatment on VNIIMK 8931. Several commercial varieties derived from Pervenets and breeding materials with a high oleic acid content have been marketed. However, the genetics of this trait are still not fully understood by breeders. To characterize the Pervenets mutation, we studied RFLP in relation to high oleic acid content. We performed diversity analyses on 239 genotypes with cDNA sequences coding for Δ9- and Δ12-desaturases as probes. The Δ12 RFLPs enabled us to identify at least two independent loci. One Δ12 RFLP allele (Δ12HOS) was strictly correlated to high oleic acid content, whereas no correlation was found between Δ9-desaturase polymorphism and high oleic acid content. These results enabled to us estimate the genetic distance between the marker and the Pervenets mutation loci. An F2 segregating population of 107 plants confirmed the correlation between high oleic acid content and Δ12HOS, indicating tight genetic linkage. The nature of the Pervenets dominant mutation and the complexity of the high oleic acid content trait are discussed.

A method based upon targetting of introgressed markers in a Phomopsis-resistant line (R) of cultivated sunflower, issuing from a H. argophyllus cross was used to mark the Phomopsis resistance regions. Our study was based upon 203 F(3) families derived from a cross between an inbred line susceptible to Phomopsis (S1) and the introgressed resistant line (R). Families were checked for Phomopsis resistance level in a design with replicated plots and natural infection was re-inforced by pieces of contaminated stems. Thirty four primers were employed for RAPD analysis. Out of 102 polymorphic fragments between (S1) and H. argophyllus, seven were still present in (R) suggesting that they marked introgressions of H. argophyllus into (R). The F(2) plants were scored for the presence or absence of 19 fragments obtained from five primers, and the relationships between the presence/absence of fragments in F(2) plants and Phomopsis resistance/susceptiblity in the F(3) progenies was determined by using an analysis of variance. We found that at least two introgressed regions, as well as favourable factors from sunflower, contributed to the level of Phomopsis resistance in cultivated sunflower.

Summary Trees or seedlings representing the species Platanus occidentalis, P. orientalis, P. acerifolia and P. densicoma completed by three P occidentalis x P. orientalis hybrids and two plants of P wrightii and P. racemosa were analysed. DNA from leaves were used for RAPD amplification with thirteen primers. Factorial correspondence analysis and Sokal and Michener genetic distances were computed. They confirmed the hybrid origin of P. acerifolia between P. occidentalis and P orientalis, and revealed that P. densicoma resulted from an equivalent cross. So, the latter has to be assimilated into P. pyramidalis from Rivers. P. wrightii and P racemosa individuals appeared much more closer to P occidentalis than to P. orientalis.

Although the tree development directs the olive tree yield we focus here to describe the main phases, stages and key steps from blossoming to harvest. The olive tree produces much more flowers than all other trees, however, fruit set and final harvest are limited by several parameters. Most articles in literature deal with the physiological aspects of the transformation of the olive flower into the drupe. Little attention has been given to genetic involvements that underlay the physiology and the biochemistry. A comprehensive survey of the literature as much as we can is given with genetic commentaries based upon recent genetic progresses made in the field. The olive tree derived from the oleaster and the oleaster belongs to an intertropical (Asian, African) genus and a major difference between northern species (Fraxinus, Syringa, Ligus‐ trum) is the bare axillary buds at the basis of leaves. This explains why the olive is more susceptible to freeze than other Oleaceae as Fraxinus. Olive flower buds appear on one-yearold twig in terminal position or at the axillary of the leaves.

Seven wildPetunia species including 2n = 18 species (P. parviflora Jussieu,P. linearis Hook.) and those with 2n = 14 (P. parodii Steene,P. axillaris Lam.,P. integrifolia Hook.,P. inflata R.E. Fries,P. violacea Lindl.) and tenPetunia hybrida horticultural lines were compared for polymorphisms in rDNA genes using the four restriction enzymesEcoRI,BamHI,HindIII andXhoI. All the unit types found in the lines pre-existed in the wild forms. There are two different sizes of either 11.45 or 11.6 kb./The 2n = 18 species are closely related to the 2n = 14 species, thus making thePetunia genus homogeneous. Moreover, it is likely thatP. hybrida lines originated in several kinds of crosses between these species. We constructed a dendrogram for all the 15 rDNA unit types found. Two main branches of the tree result from the presence or the absence ofHindIII sites. The main branch is divided according to variability at theEcoRI andBamHI sites. Taking into account the existence of several loci which carry one unit type only, we consider whether or not exchanges might occur between loci. Lines carrying two unit types and lines carrying three unit types support such a hypothesis.XhoI andBamHI fragments enable us to distinguish two types of 5S DNA corresponding to 2n = 18 and 2n = 14 species, respectively.P. hybrida lines and each 2n = 14 wild species carry one of the types only, that corresponds to one 5S DNA locus. The most parsimonious phylogenetic trees whatever the species chosen as the outgroup, do not fit with our knowledge ofPetunia and with taxonomy. This is likely because only few loci formed the basis of these phylogenetic constructions.

Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; Most olive varieties are not strictly self-incompatible, nevertheless, they request foreign pollen to enhance fruit yield, and consequently orchards should contain pollinisers to ensure fruit set of the main variety. The best way to choose pollinisers is to experiment numerous crosses in a diallel design. Here, the genetic mode of inheritance of SI in the olive is deciphered and it does not correspond to the GSI type, but to the SSI type. It leaves S-allele dominance relationship expression in the male (pollen and pollen tube), but not in the female (stigma and style). Thus, a pair-wise combination of varieties may be inter-compatible in one direction (male to female, or female to male) and inter-incompatible in the other direction. Dominance relationships also explain different levels of self-pollination observed in varieties. Little efficient pollinisers were found and predicted in varieties; nevertheless, some new efficient pair-wise allele combinations are predicted and could be created. This model enables one to forecast compatibility without waiting for several years of yield records and to choose pollinisers in silico.

To begin, the methodology followed to reconstruct the origin and history of the olive is presented. The genetic structure (density of alleles across the geographic distribution of individuals) based on allele frequencies of the present oleaster tree in the Mediterranean Basin computed with different methods of comparison with the genetic structure of cultivars grouped based on their geographic and genetic origins infers several possible scenarios for the transition from the oleaster to the olive. To screen among the scenarios requires solid dating in oleaster presence, diffusion and physical remains (from oleaster and cultivar trees) from different sites. Consequently, reconstructing the origin of the olive is based upon data from diverse disciplines and integrating them appears fruitful. Genetic data show that an event (such as bottleneck, migration, differentiation, adaptation) has occurred, but it cannot be dated. Thus it requires crossing genetic data with data gathered from different biological disciplines to make a strong case for this history. We examine successively

Common sunflower (i.e., Helianthus annuus) is occasionally crossed with other Helianthus species to gain favourable agronomic traits. An interspecific cross between H. annuus and Helianthus mollis was used to examine events that occurred during the five first generations. First generation hybrid plants were verified using GISH, and F1 hybrid (#17) was back-crossed to common sunflower to obtain BC1 (#4) and BC2 (#21). Some BC2 progeny possessed new traits, i.e., compact (Compact plant architecture is due to short inter-node length, short petiole length and short leaf lamina length) architecture without and with corrugated leaves, which were not observed in the parents. These traits were selected for and fixed after the BC2-S3 (#18) to produce a H. mollis introgression line (namely HM374). Although the size of the population was small, we tentatively construct a genetic map using data from the BC2 generation. Two linkage groups were identified carrying H. mollis specific AFLP markers that were associated with compact plant architecture, and one unlinked marker was shown to explain variation for the corrugated leaves. Introgression line HM374 was used to test the inheritance of compact plant architecture in a cross with an elite high oleic line, OL75HO. Microsatellite (SSR) markers were used to genotype F1 and F2 offspring’s. Our data suggested that the line HM374 was still heterozygous for introgression after eight generations. HM374 displays new architecture traits (i.e. compact architecture with or without corrugated leaves) and carries H. mollis fragments anchored on LG1 and LG11 of sunflower public map.

Because they usually correspond to unique sequences, RFLP markers are used in genomic mapping, variety distinction, variety homogeneity and in reconstruction of phylogenies. Nevertheless, they are not the most suitable markers for all these applications. Their use in taxonomy is not always straightforward, since these sequences may be highly conserved as in the genus Beta L. The analysis of genetic variability allows calculation of genetic distances and enables the construction of phenograms, but not phylogenetic trees. Phylogenetic trees are constructed using other methods based on parsimony analysis and, hence, they do not always fit within taxonomic groups. Therefore, we have turned to other molecular markers which have already been found in sections Beta, Corollinae and Procumbentes, and, therefore, fit well into the main taxonomic divisions. RFLPs with repeated sequences are generally not used, although they offer a wide range of potential applications. Some have already been used and others appear suitable for use as markers. Here we present results with moderately and highly repeated sequences which display RFLP or are genome specific when used as probes. The main advantages and disadvantages of both RFLP types are discussed.

Dans 6 lignees de Petunia hybrida dont l'ADN a ete hydrolyse par EcoRI, BamHI, HindIII, KpnI, SacI ou XhoI, l'ADN ribosomique est apparu tres polymorphe. Chaque lignee porte plussieurs types d'unites; ainsi 13 types differents sont reveles dans les lignees, ce qui est surprenant. Nous avons caracterise les differents types et determine le nombre de loci. Deux types d'unites avec et sans sites HindIII sont reveles. Pour la lignee St43 les fragments EcoRI et BamHI permettent de construire une unite de 11,4 kb. En outre un fragment EcoRI de 2,6 kb ne peut etre place dans l'unite de 11,4 kb. Il est en effet equivalent au fragment de 2,4 kb portant la sequence codante du gene 25 S. Il est donc place dans une unite de 11,7 kb. Dans la lignee Tlh1 les fragments correspondants ne permettent de construire que des unites de 8,8 kb, 9,2 kb, et 10,8 kb, donc considerees comme une partie d'unites de 11,4 kb. Les autres lignees montrent une combinaison des fragments precedents. L'heredite du polymorphisme dans 50 descendants du couple de lignees (St43 et Tlh1) et les 2 retrocroisements possibles [(St43 × Tlh1I) × St43] et [(St43 × Tlh1) × Tlh1] indique au moins 2 loci. La presence dans Sk176 des fragments EcoRI 6,2, 5,7 et 5,4 suggere 3 loci. Parmi les plantes haploides obtenues de l'hybride F1 (St43 × Tlh1), un descendant porte les 3 types d'unite presents dans l'hybride, ce qui ne peut s'expliquer que s'ils sont repartis sur 3 loci. De plus la segregation dans les retrocroisements correspond a 2 loci pour St43, il porte donc un nulliallele. La confirmation des hypotheses est apportee par hybridation in situ avec une sonde ribosomique biotinylee. Les 2 lignees dihaploides Tlh1 et T1h7 montrent le plus d'unites et par consequent le polymorphisme le plus eleve

Contact: berville@supagro.inra.fr; International audience; Seventy-seven wild sunflower accessions from USA were compared in France (Mauguio) for 13 quantitative characters using multivariate methods, including clustering, principal component and canonical discriminate analyzes to assess the patterns of morphological and climatic variation. Geographic and climatic data from their sampled sites such as latitude, longitude, elevation, rain fall, temperature, number of rain fall days, temperature range, and sunniness were also analyzed and we sought for correlations of climatic variables and morphological traits. Climatic data clustered the accessions into four groups. Principal component analysis was used to summarize the characteristic habits of annual wild sunflower sampled sites in terms of geography and climates, thus we characterized the ecogeographical profiles of wild sunflowers. As we studied the traits in a common environment we speculated whether local adaptation patterns are sustained by our data. Indeed traits shared by accessions that belong to climatic clusters are suggested to be due to local adaptation

Contact: berville@supagro.inra.fr Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; The present diversity of the olive (crop) and oleaster (wild) tree was investigated with nuclear and cytoplasm markers. Patterns of diversity of the wild form inferred eleven ancestral populations in the East and the West of the Mediterranean basin. Patterns of diversity for cultivars are less clear, but we showed that cultivars admixed to nine groups that corresponded to oleaster ancestral populations. We inferred that nine domestication events took place in the olive, but these origins were blurred by gene flow from oleaster and by human displacements. These origins of domestication probably reflected different reasons and uses to domesticate the oleaster. To cite this article: C Breton et al., C R. Biologies 332 (2009). (C) 2009 Academie des sciences; La diversité actuelle de l’olivier (cultivé) et de l’oléastre (sauvage) a été étudiée avec des marqueurs nucléaires et cytoplasmiques. Les patrons de diversité de la forme sauvage conduisent onze populations ancestrales à l’Est et l’Ouest du bassin méditerranéen. Pour les cultivars, les patrons de diversité sont moins tranchés, mais nous avons montré qu’ils se rattachent à neuf groupes qui correspondent à neuf des onze populations ancestrales d’oléastre. Nous en avons déduit que neuf évènements de domestication différents se sont produits pour conduire à l’olivier actuel, mais les origines sont rendues diffuses du fait de flux de gènes avec l’oléastre et des déplacements des cultivars par l’homme. Ces origines de domestication reflètent probablement diverses motivations et utilisations de l’olivier et de ses produits. Pour citer cet article : C. Breton et al., C. R. Biologies 332 (2009)

International audience; Classical sunflower varieties display a high linoleic acid content in their seeds [low oleic (LO) varieties] whereas genotypes carrying the Pervenets mutation display an increased oleic acid content of above 83% [high oleic (HO) varieties]. Despite the advantage in health terms of oleic acid, the nature of the mutation was still unknown. Previous work reported that HO genotypes carried a specific oleate desaturase (OD) allele. This enzyme catalyses the desaturation of oleic acid into linoleic acid. The present work demonstrates that this allele is organised in two parts: the first section present in both HO and LO genotypes carries a normal OD gene, the second section is specific to HO genotypes and carries OD duplications. The study of mRNA accumulation in LO and HO seeds revealed that the mutation is dominant and induces an OD mRNA down-regulation. Furthermore, OD small interfering RNA, characteristic of gene silencing, accumulated specifically in HO seeds. Considered together, these observations show that the mutation is associated with OD duplications leading to gene silencing of the OD gene and consequently, to oleic acid accumulation. This finding allowed the development of molecular markers characterising the mutation that can be used in breeding programmes to facilitate the selection of HO genotypes.

Contact: berville@supagro.inra.fr Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; The olive belongs to Olea, a complex genus and to a species with six subspecies. Subsp. europaea includes both the cultivated olive and the oleaster, the wild ancestor of the olive. Little is known on the phenotype of fruits from subsp. cuspidata. We aimed to compare europaea and cuspidata trees for oil content and composition and to verify whether natural hybrids may exist between the two subspecies. Specimens were from Kenya and putative natural hybrids between cuspidata and europaea were from Stellenbosch (South Africa). Cultivar and oleaster trees were sampled in France (continental and Corsica), Italy (Continental and Sardinia) and Tunisia. We have examined the fruit (drupe) structure between subsp. cuspidata and europaea and extracted oil from the pulp, seed and total drupe. Comparison for oil content was made between the cultivated olive and the oleaster and some cuspidata trees from Kenya. A few of cuspidata individuals from Botanical gardens do not enable phenotyping for oil content and composition. Oil composition analyses were performed for the main fatty acids to compare the three taxa. We used microsatellite markers at 11 loci to compute genetic distances between cuspidata, oleaster and cultivar trees and to reveal eventual hybrids. The SSR polymorphisms were huge between the two subspecies and they revealed that putative hybrids were true hybrids sampled around olive orchards. The whole comparison of oil content shows that olive cultivars display higher oil content than the subsp. cuspidata and that the oleaster trees are intermediate, whereas for oil composition of the drupe, cuspidata from Kenya shows less oleic acid than europaea. However, the cuspidata trees show seed oil composition similarities with the other two taxa. The discussion deals with possible reasons to explain the differences and of the possible uses of these taxa for breeding both the olive and brown olive

email: berville@supagro.inra.fr Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; Olive cultivars are diversified but nothing is known on their origins and if they are local or introduced in any regions. The study aims to determine which traits may help to identify native from introduced cultivars and oleaster trees. We compared cultivars and oleasters from North Tunisia to determine their relationships based on morphological traits, oil composition and SSR genotyping at seven loci. We used those parameters to examine 32 cultivar trees from 17 denominations and 70 oleaster trees sampled. We used multivariate analysis, enabling to retain the best variables, to establish relationships among trees based on morphological and pomological parameters. Gas chromatography was used to determine fatty acid composition of 30 cultivar trees and 13 oleaster trees. We determined for one cultivar Gerboui the steady drupe, pit morphological and oil composition variation ranges in six different contrasted agro-systems. SSR genotyping was performed in polyacrylamide gels after fluorescent labelling. Based on morphology, oleaster trees from agro-ecosystems clustered broadly in an intermediate position between cultivars and oleasters from natural ecosystems. SSR revealed that the feral and genuine oleasters plus cultivars are always overlapping. Relationships between cultivars are displayed in two dendrograms. They revealed six and three main clusters based on Unweighted Pair Group Method (UPGMA) and Ward algorithm, respectively. They mix olive cultivar and oleaster trees suggesting kinship relationships between some cultivar and some oleaster trees. In contrast, on PCA, some morphological parameters split our sample approximately between olive and oleaster trees. Oil composition was similar between cultivar and oleaster trees. Kinship relationships between cultivar and oleaster trees based on molecular polymorphisms suggested that olive cultivars may have origin in local oleasters. Oil composition as fruit descriptors and drupe size appeared inefficient to discriminate between olive and oleaster trees, in comparison to SSR. Our results suggested several domestication events for the olive. It is important to know which cultivars have local origin to promote and sale products from Tunisia as from all around the Mediterranean basin

Correspondance: andre.berville@supagro.inra.fr Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; Olive cultivar origins are unknown. We attempted to use multivariate analyses and based Bayesian methods to look for sites of olive cultivar geographic origins. In this study, to establish the method, forty cultivars were chosen with different chlorotypes that represent the olive RAPD-based diversity in the Mediterranean basin. We constructed a dendrogram, as reference, using molecular SSR data. Cultivar origins in the oleaster were examined by using Bayesian clustering methods. It was calculated whether some cultivars may be grouped by admixture analysis to these GRPs and how much olive cultivars admixed to the previously defined oleaster GRPs: two in the east and four in the west of the Mediterranean basin. The comparison between dendrogram and Bayesian analyses showed that when the dendrogram suggests relationships, the Bayesian method quantifies them by probabilities and proportions. The method used is applicable to microsatellite data from other crop/wild complexes

Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; The study aims to determine which traits may help to differentiate cultivars and oleaster trees based on drupe and pit morphological traits, oil content and composition and nuclear DNA markers at seven SSR loci. We determined for one cultivar (Gerboui) the steady drupe, pit morphological and oil composition variation ranges in six different contrasted agro-systems. We then investigated 33 cultivars and 40 oleaster trees. PCA analysis on morphological traits was examined. Gas chromatography was used to determine fatty acid composition of 20 cultivars and 13 oleaster trees, and the Soxhlet method was used to determine fruit oil content. SSR genotyping was performed in poly-acrylamide gels. Five clusters based on UPGMA mixed olive cultivars and oleaster trees suggesting kinship relationships between some cultivars and some oleasters; and thus cultivars could derive from domestication of oleaster trees.; L'étude a pour objet de déterminer les caractères permettant de différencier les oléastres et les cultivars d'olivier en se basant sur la morphologie du fruit et du noyau, le contenu et la composition de l'huile, et de la variation génétique à 7 locus SSR. Nous avons au préalable déterminé l'amplitude de la variation morphologique et biochimique induite par l'environnement pour le cultivar Gerboui dans 6 agro-écosystèmes différents. Nous avons ensuite caractérisé 33 cultivars et 40 d'oléastres. La variation morphologique est analysée par ACP, la composition en acides gras par chromatographie en phase gazeuse. Nous avons utilisé la méthode de Soxhlet pour quantifier la teneur en huile de 20 cultivars et 13 oléastres. Les SSR ont été séparés en gel de poly-acrylamide. Cinq groupes d'individus construits selon la méthode UPGMA basés sur les données génétiques suggèrent des parentés étroites entre oléastres et cultivars qui découleraient de domestication locale ou de flux de gènes entre les deux compartiments.

The isolation of mitochondria from sugar beet and sunflower has been carried outina buffered medium of high ionic strength (I=1.40 M). Them tDNAs obtained were enriched for supercoiled molecules of low molecular weight. Chloroplast and mitochondrial DNA were successively prepared and used from a single leaf preparation for further analysis of restriction fragment length polymorphism. The few available wild relatives of sugar beet or sunflower have also been analyzed by comparison of the restriction patterns of their chloroplast and mitochondrial DNAs.

Correspondance: berville@supagro.inra.fr Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; Aim The oleaster is believed to have originated in the eastern Mediterranean, implying that those in the western Mediterranean basin could be feral. Several studies with different molecular markers (isozymes, random amplified polymorphic DNA, amplified fragment length polymorphism) have shown a cline between the eastern and the western populations, which supports this hypothesis. To reconstruct the post-glacial colonization history and establish a relationship between olive and oleaster populations in the Mediterranean basin, analyses were carried out on the genetic variation of chloroplast DNA (chlorotype) and at 12 unlinked simple sequence repeat (SSR) loci, sampling a total of 20 oleaster groves. Location: This is the first known large-scale molecular study of SSR loci based on samples of both oleasters and cultivars from the entire Mediterranean basin.Methods Samples were taken from 166 oleasters in 20 groves of modern populations, and 40 cultivars to represent molecular diversity in the cultivated olive. The Bayesian method and admixture analysis were used to construct the ancestral populations (RPOP; reconstructed panmictic oleaster populations) andto estimate the proportion of each RPOP in each tree. If one tree can be assigned to two or more RPOPs, it can be regarded as a product of hybridization between trees from different populations (i.e. admix origin).Results: On this first examination of the SSR genetic diversity in the olive and oleaster, it was found to be structured in seven RPOPs in both eastern and western populations. Based on different population genetic methods, it was shown that: (1) oleasters are equally present in the eastern and the western Mediterranean, (2) are native, and (3) are not derived from cultivars. Chlorotypes (one and three in the eastern and western Mediterranean, respectively) revealed fruit displacement for the oleasters.Main conclusions: Oleaster genetic diversity is divided into seven regions that could overlay glacial refuges. The gradient, or cline, of genetic diversity revealed by chloroplast and SSR molecular markers was explained by oleaster recolonization of the Mediterranean basin from refuges after the last glacial event, located in both eastern and western regions. It is likely that gene flow has occurred in oleasters mediated by cultivars spread by human migration or through trade. Animals may have helped spread oleasters locally, but humans have probably transported olives but not oleaster fruits over long distances. We found that cultivars may have originated in several RPOPs, and thus, some may have a more complex origin than expected initially.

The other chapters in this book demonstrate the vast array of systems that plants utilize to evolve endoferality and exoferality, and mixtures between the two. This chapter presents short case histories of other crop species that extend this biodiverse array and accentuates other issues and implications that may derive from ferality. Clearly, other cases could be added to this, as many as there are additional other crops, but the idea is not to be exhaustive. Our intent is to supply a feeling of further issues and to provide parameters that must be discerned when analyzing the potential problems, if any, that may arise when a given crop becomes a persistent volunteer or introgresses genes from a wild relative.

Virgin olive oil is made from diverse cultivars either mixed or single. Those ensure different tastes and typicity, and these may be also enhanced by the region of production of cultivars. The different olive oil labels correspond to their chemical composition and acidity. Labels also may correspond to a protected origin indication, and thus, such oils contain a given composition in cultivars. To verify the main cultivars used at the source of an olive oil sample, our method is based on DNA technology. DNA is present in all olive oil samples and even in refined oil, but the quantity may depend on the oil processing technology and oil conservation conditions. Thus, several supports were used to retain DNA checking different techniques (silica extraction, hydroxyapatite, magnetic beads, and spun column) to prepare DNA from variable amounts of oil. At this stage, it was usable for amplification through PCR technology and especially with the magnetic beads, and further purification processes were checked. Finally, the final method used magnetic beads. DNA is released from beads in a buffer. Once purified, we showed that it did not contain compounds inhibiting PCR amplification using SSR primers. Aliquot dilution fractions of this solution were successfully routinely used through PCR with different SSR primer sets. This enables confident detection of eventual alien alleles in oil samples. First applied to virgin oil samples of known composition, either single cultivars or mixtures of them, the method was verified working on commercial virgin oil samples using bottles bought in supermarkets. Last, we defined a protocol starting from 2 x 40 mL virgin olive oil, and DNA was prepared routinely in about 5 h. It was convenient to genotype together several loci per sample to check whether alleles were in accordance with those of expected cultivars. Thus, forensic applications of our method are expected. However, the method needs further improvement to work on all oil samples.

With more than 100 accessions, the CBNMP olive collection includes a major part of the French germplasm. We used molecular markers to characterise all accessions and to study genetic relationships between cultivars. Firstly, 497 olive trees were genotyped using 32 RAPD markers. We identified 114 RAPD profiles and detected several cases of mislabelling, synonymy and homonymy. Secondly, for each RAPD profile, one tree was analysed using mtDNA RFLPs to determine the cytoplasmic lineage of each cultivar and using five nuclear SSR loci. French germplasm displayed ME1, MOM and MCK mitotypes with ME1 prevailing (84%). Based on SSR markers, we revealed a slight differentiation between French cultivars growing in the West and the East side of the Rhône Valley. This study allowed us to construct a molecular data-base for the reference collection and to analyse genetic diversity for further prospecting, and for introducing new olive accessions.

To obtain introgressed sunflower lines with improved disease resistance, interspecific crosses were performed with foreign perennial species. We report on several unusual features displayed by these hybrid plants. The methods used to produce the kernels affected yield and genotypes of progeny. Phenotypic traits and DNA markers were investigated in 97 plants derived from cross-pollination between annual diploid cultivated sunflower (Helianthus annuus) and the perennial diploid species H. mollis or H. orgyalis, and the reverse reciprocal crosses. The level of hybridization in progeny was determined using RAPD and RFLP markers. Hybridization was performed by leaving embryos to develop normally on the head (classical crossing) or using embryo rescue. All observed plants derived from H. mollis were diploid (2n = 34). Phenotypes were predominantly similar to the female when cultivated sunflower was the female parent. Progeny from crosses using a wild species as the female parent resembled that parent. Thus, reciprocal crosses led to different progeny. F1 sister progeny shared different sets of molecular markers representing a few of those of the wild species used as the pollen donor. Our results indicate mechanisms leading to the unusual event of partial hybridization. Possible mechanisms behind these unusual events and their possible impact on evolution are discussed.