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commonly known as ‘Esca’, is a detrimental fungus for many tree species and grapevine and is considered to be one of the main causal agents of wood decay of olive plantations in the Mediterranean...

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.

Some imperative developed genotypes of almond (‘Tardy Nonpareil’, ‘Nonpareil’, ‘Sahand’, ‘Rabie’, ‘Azar’, ‘Shekofeh’, ‘Sefied’ and ‘Mamaei’) were utilized in two experiments for 3 years to elucidate the genotype response of almond pollen germination and tube development at various temperature levels. Pollen was subjected to pre-incubation at 10, 20, 30 or 40∘C for 24 h to reproduce temperature stress during pollen dispersal; and then in vitro cultured. In the second examination, pollen was exposed at 15, 20, 25 or 30 °C for 24 h in vitro to assess pollen reaction in state of water and supplements accessibility and to determine the ideal pollen germination and tube development temperatures for every genotype. The most elevated pre- incubation temperature treatment (40 °C) completely prevented pollen germination in “Mamaei” and ‘Sefied’, while even in the less influenced cultivars ('Shekofeh’, ‘Azar’, ‘Rabie’, ‘Sahand’, ‘Tardy Nonpareil’ and ‘Nonpareil’) germination rates were as low as 2–3%. Pre-incubation at 30 ∘C or 40 °C negatively affected pollen germination and pollen tube length. In the second experiment, in vitro pollen germination increased after incubation at 25 °C for ‘Mamaei’ (+2%), ‘Sefied’ (+3%), ‘Shekofeh’ (+3%), ‘Azar’ (+3%), ‘Rabie’ (+4%), ‘Sahand’ (+5%), ‘Nonpareil’ (+5%) and ‘Tardy Nonpareil’ (+5%) compared to the control (20∘C). At 30 ∘C, germination percent for ‘Mamaei’, ‘Sefied’, ‘Shekofeh’, ‘Azar’, ‘Rabie’, ‘Sahand’, ‘Nonpareil’ and ‘Tardy Nonpareil’ was 35, 35, 10, 15, 25, 15, 35, and 20% lower, respectively, compared to the control (20 ∘C). Pollen tube length additionally increased with incubation temperature for all of the considered genotypes. In view of the cumulative stress response index (CSRI) that was calculated for high temperature stress the developed genotypes were arranged: ‘Shekofeh’ and ‘Tardy Nonpareil’ as tolerant and ‘Nonpareil’, ‘Rabie’ and ‘Sahand’ as intermediate at 40 ∘C, while ‘Azar’, ‘Rabie’, Sahand’ and ‘Tardy Nonpareil’ as tolerant, ‘Shekofeh’ and ‘Nonpareil’ as intermediate and ‘Mamaei’ and ‘Sefied’ as sensitive at 30 °C. The observed strong genotype differentiated reaction in high and low temperature stress could be utilized by plant breeders towards producing new tolerant almond genotypes.

In the present study, four Greek (Agiorgitiko, Asyrtiko, Roditis and Xinomavro) and one international (Soultanina) grapevine cultivars (Vitis vinifera L.) were screened for their resistance to Phaeomoniella chlamydospora. Artificial inoculation was carried out by drilling a hole into the trunk and injecting a concentrated conidial suspension into the vessels. Disease reactions were evaluated in an 87-day assessment period, on the basis of external symptoms (disease incidence, disease severity and mortality) and by calculating the relative areas under disease progress curves (relative AUDPC). The extension of vascular browning as well as the isolation ratio along the inoculated vine trunks were also taken into account as additional parameters for evaluating resistance. The results indicated that the resistance of grapevine cultivars to P. chlamydospora varied significantly. ‘Agiorgitiko’ and ‘Soultanina’ were susceptible, whereas ‘Asyrtiko’ and ‘Xinomavro’ were resistant; ‘Roditis’ showed an intermediate level of resistance. Cultivars’ resistance was mostly distinguished in terms of the extension of vascular browning and pathogen isolation ratio. On the contrary, the disease incidence, final disease severity, mortality and relative AUDPC provided less distinctive efficiency in resistance evaluation. The robust methodology presented here could be useful in rapid evaluation experiments for future screening programs to search and recognize natural resistant sources within grapevine genotypes against P. chlamydospora.

A two-year survey was conducted to identify fungi associated with wood decay in a range of tree species and grapevine. Fifty-eight fungal strains isolated from plants of 18 species showing typical wood decay symptoms were characterized by morphological, physiological, and molecular analyses. By 5.8S rRNA gene-ITS sequencing analysis, these isolates were classified into 25 distinct operational taxonomic units, including important phytopathogenic species of the phyla Pezizomycotina and Agaricomycotina, such as Fomitiporia, Inonotus, Phellinus, Inocutis, Fuscoporia, Trametes, Fusarium, Eutypa, Phaeomoniella, Phaeoacremonium, and Pleurostomophora spp. The white rot basidiomycetes Fomitiporia mediterranea (20 isolates, 34.5%) and Inonotus hispidus (6 isolates, 10.3%) were the most prevalent. Pathogenicity tests revealed for the first time that certain fungal species of the genera Fomitiporia, Inonotus, Phellinus, Pleurostomophora, and Fusarium caused wood infection of various tree species in Greece and worldwide. To the best of our knowledge, this is the first report of F. mediterranea as the causal agent of wood decay in pear, pomegranate, kumquat, and silk tree. This is also the first record of Inonotus hispidus, Phellinus pomaceus, Pleurostomophora richardsiae, and Fusarium solani in apple, almond, avocado, and mulberry tree, respectively, whereas P. richardsiae was associated with wood infection of olive tree for the first time in Greece. Cross pathogenicity tests with F. mediterranea strains originated from grapevine applied on other woody hosts and from olive on grapevine demonstrated partial host specificity of the fungus. The potential of F. mediterranea to transinfect hosts other than those originated, along with the host range extension of the fungus, is discussed.

Barbary fig ( Opuntia ficus indica ) could be an economically important species as it could be an alternative crop extremely tolerant to dry condition and water deficiency. Moreover, it could be used in alternative sustainable cultivation systems and landscape conservation. In this work, we report the analysis of the genetic diversity of Greek Barbary fig genotypes using ISSR molecular markers. Six primers were screened to assess their ability to detect polymorphisms within twenty-two Barbary fig accessions and generated 57 markers (bands), with an average of 9.5 markers per primer. The percentage of polymorphic bands (50.21%) and the resolving power (R P ) (28.85) showed the efficiency of the used primers. Mean values for GD (gene diversity) and I (Shannon index) were found as 0.215 and 0.355, respectively. The revealed ISSR markers allow distinguishing all accessions analyzed except for one case. UPGMA dendrogram and PCoA (Principal Coordinate Analysis) were performed to access patterns of diversity among genotypes. The high genetic diversity existing in the Greek germplasm suggests that it would be beneficial to utilize this pool in Barbary fig breeding programs and germplasm management activities.

The present study attempts to determine how some physiological and reproductive functions of olive tree (Olea europaea L., cv. Koroneiki) respond to enhanced UV-B radiation or heat. Enhanced UV-B radiation was applied to (1) three-year-old potted plants in an open nursery (corresponded to ca. 16% ozone depletion), and (2) in vitro cultured pollen samples (220 μmol m−2 s−1, PAR = 400−700 nm + UV-B at 7.5, 15.0, or 22.5 kJ m−2 d−1). Potted olive plants were also subjected to high temperature (38 ± 4°C) for 28 h to mimic heat levels regularly measured in olive growing areas. A significant effect of UV-B on photosynthetic rate was observed. However, enhanced UV-B radiation did affect neither chlorophyll nor carotenoid content, supporting previous reports on hardiness of the photosynthetic apparatus in olive. Increased superoxide dismutase activity was observed in UV-B-treated olive plants (+ 225%), whereas no effect was found in the plants under heat stress. Neither UV-B and nor heat did affect H2O2 accumulation in the plant tissues. However, the same treatments resulted in enhanced lipid peroxidation (+ 18% for UV-B and + 15% for heat), which is likely linked to other reactive oxygen species. The increased guaiacol peroxidase activity observed in both treatments (+ 32% for UV-B and + 49% for heat) is related to the defense against oxidative membrane damage. The observed reduction in pollen germination (20–39%) and tube length (11–44%) could have serious implications on olive yields, especially for low fruit-setting cultivars or in years and environments with additional unfavorable conditions. UV-B and heat effects described here support the hypothesis that plant response to a given stressor is affected by the overall context and that a holistic approach is necessary to determine plant strategies for climate change adaptation.

Olea europaea L. has been cultivated in the Mediterranean region for thousands of years and is of major economic importance. The origin of olive cultivars remains as complex to trace as their identification. Thus, their molecular characterization and discrimination will enable olive germplasm management. In addition, it would be a useful tool for authentication of olive products. High-resolution melting (HRM) analysis, coupled with five microsatellite markers, was integrated to facilitate molecular identification and characterization of main O. europaea cultivars collected from the National Olive Tree Germplasm Collection established in Chania, Greece. The five microsatellite loci used were highly informative and generated a unique melting curve profile for each of the 47 cultivars and for each microsatellite tested. In particular, three microsatellite markers (DCA03, DCA09 and DCA17), which generated 29 HRM profiles, were sufficient to genotype all the olive cultivars studied, highlighting their potential use for cultivar identification. Furthermore, this assay provided a flexible, cost-effective and closed-tube microsatellite genotyping method well suited for molecular characterization of olive cultivars.

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.

The genetic relationships between and within some traditionally grown cultivars of olive tree (Olea europaea L.) in Greece (island of Crete) and in Turkey were investigated. Cultivars from Crete included ‘Koroneiki’, ‘Throubolia’ and ‘Mastoidis’, while those from Turkey included ‘Samanli’ and ‘Gemlik’. Cultivars were represented by multiple genotypes of aged trees collected from the field, each one complying with established descriptors. Representative genotypes of wild olive trees from Crete were also employed. A total of 112 genotypes were analysed, employing seven microsatellite (SSR) loci yielding a total of 81 alleles, and reaching a cumulative probability of identity of 6.73 × 10−09 with a mean observed heterozygosity of 0.852. Analysis of molecular variance significantly partitioned genetic diversity between and within cultivars, albeit with no appreciable difference between the two levels of diversity. All cultivar genotypes aggregated along single, cultivar-specific clusters, pointing to human-driven selection. The two Turkish cultivars ‘Samanli’ and ‘Gemlik’ were grouped together. The Cretan cultivar ‘Throubolia’ grouped together with the two Turkish cultivars, indicating germplasm movement across the Aegean Sea during historical times. Some gene flow was observed between the Cretan cultivars and the native wild populations (likely feral forms). SSR alleles were ranked for their efficiency in discriminating the examined materials, thus establishing a molecular key for cultivar identification. An identification process is proposed including a classification binary tree and provided a method for sorting any new unknown material purportedly originating from any of the analysed cultivars.

The floral biology of ‘Koroneiki’, ‘Kalamata’ and ‘Mastoidis’ was studied for three consecutive years to elucidate the potential physiological and genetic controls of inflorescence architecture and phenology and to identify potential genotype-distinguishing characters that could be employed for morphological cultivar discrimination. The first open flowers were mainly observed at position I for ‘Koroneiki’ (83%) while they were mainly located at position II for ‘Mastoidis’ (40%) and ‘Kalamata’ (63%). The last open flowers were principally located at the apical position for ‘Koroneiki’ (46%) and ‘Kalamata’ (68%) and at position II for ‘Mastoidis’ (75%). Most of the first open flowers were perfect for ‘Koroneiki’ (89%) and ‘Mastoidis’ (97%) while lower percentages were observed in the last open flowers (67% and 80%, respectively). Higher percentages of perfect flowers were observed in ‘Mastoidis’ compared to ‘Koroneiki’ in both cases. The average flowering shoots of ‘Kalamata’ produced significantly more flowers (1523/m) compared to ‘Koroneiki’ (1139/m) and ‘Mastoidis’ (1044/m). The half of the flowers in ‘Mastoidis’ were located at position I while the 38% was located at position II and only the 12% at the apex. By contrast, approximately the 80% of flowers in ‘Koroneiki’ and ‘Kalamata’ panicles was equally distributed at positions I and II, while the 18% and 19%, respectively, was located at the apex. A marked relationship was also observed between flower position in the panicle and flower gender.

Olive varieties ‘Koroneiki’, ‘Kalamata’, ‘Mastoidis’ and ‘Amigdalolia’ were employed in two experiments for 3 years to assess the effect of temperature on olive pollen germination and tube growth in relation to relative humidity and genotype. Pollen samples were subjected to pre-incubation at 10, 20, 30 or 40 °C in combination with decreased air relative humidity – 80, 40, 30 or 20%, respectively – for 24 h to simulate temperature stress that is observed during pollen dispersal; and subsequently in vitro cultured. In the second experiment, pollen was exposed at 15, 20, 25 and 30 °C for 24 h in vitro to evaluate pollen response in conditions of water and nutrients availability and to determine the optimum pollen germination and tube growth temperatures for each cultivar. The highest pre-incubation temperature treatment (40 °C) prevented pollen germination in ‘Koroneiki’ and ‘Mastoidis’, with the less affected varieties (‘Amigdalolia’ and ‘Kalamata’) having average germination percentages of only 7.6 and 2%, respectively. Pre-incubation at 30 °C had a negative impact on pollen germination in ‘Koroneiki’ (−65%), ‘Kalamata’ (−20%) and ‘Amigdalolia’ (−72%) compared to the control (20 °C). Pollen pre-incubation at 40 °C decreased significantly the pollen tube length in ‘Kalamata’ (−50%) and ‘Amigdalolia’ (−52%). In the second experiment, in vitro pollen germination increased after incubation at 25 °C for ‘Koroneiki’ (+6%), ‘Mastoidis’ (+52%), ‘Kalamata’ (+10%) and ‘Amigdalolia’ (+10%) compared to the control (20 °C). At 30 °C germination percentages for ‘Mastoidis’, ‘Kalamata’ and ‘Amigdalolia’ were 8, 6 and 14% higher, respectively, compared to the control (20 °C). Pollen tube length also increased with incubation temperature for all of the studied cultivars. Based on the cumulative stress response index (CSRI) that was calculated for high temperature stress the varieties were classified: ‘Mastoidis’ and ‘Kalamata’ as tolerant and ‘Koroneiki’ and ‘Amigdalolia’ as intermediate at 30 °C while all studied cultivars were sensitive at 40 °C. The observed strong genotype-differentiated response in high and low temperature stress could be exploited by plant breeders towards producing new tolerant olive varieties.

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.

The effectiveness of in vitro thermotherapy (explants at 35–37°C for 20 days) in obtaining propagating material of apricot (Prunus armeniaca L.) cultivar Bebecou that is free of Plum pox virus (PPV) was compared to a conventional heat treatment technique (potted plants at 30–35°C for 8 weeks). An improved protocol for in vitro thermotherapy was combined with shoot tip culture. The protocol is 5.8 times more effective than conventional thermotherapy while taking half the time and is likely to have a high commercial impact for nurseries.

Factors affecting successful establishment in vitro, rapid proliferation and rooting of apricot cultivar ‘Bebecou’ were studied. Ethanol and NaOCl were applied in several combinations for disinfection; chilling, plant growth regulators BA, IAA and GA3, antibiotics, different culture vessels and systems of subculture were evaluated for the optimization of shoot proliferation and the auxins NAA and IBA were assessed for root induction. The highest number of new microshoots/explant (18.7) was obtained in a culture medium supplemented with 2.2 μM BA+0.57 μM IAA after 300 h of chilling. The effect of GA3 (11.4 μM) on shoot proliferation was positive in combination with 4.4 or 8.9 μM BA. Shoot length and productivity were highest at 2.2 μM BA+11.4 μM GA3+0.57 μM IAA and at 2.2 μM BA+0.57 μM IAA, respectively and decreased as cytokinin concentration increased. The antibiotic ‘Na-cefotaxime’ had a minimal impact on shoot growth when used at the lowest concentration (250 mg l−1). Subculture every 2 weeks in a medium supplemented with 2.2 μM BA and 0.57 μM IAA was more efficient for shoot induction than alternation of 20 days culture in a propagation medium supplemented with 2.2 μM BA and 10 days culture in an elongation medium supplemented with 1.1 μM BA and 5.71 μM IAA. The highest number of roots/shoot (8.1) was recorded at 19.6 μM IBA.

Variability of pollen morphology and ultrastructure of Olea europaea L. ( Oleaceae ) cultivars ‘Koroneiki’, ‘Mastoidis’ and ‘Kalamata’ was studied with scanning electron microscopy to identify genotype- distinguishing characters that could be employed for morphological cultivar discrimination. Pollen viability and germination was also measured for the three cultivars. Significant variation was observed in minimum diameter, maximum/minimum diameter (L/W) ratio and size index of pollen grain of ‘Mastoidis’ and the other two cultivars. Regarding the exine pattern, muri width of ‘Koroneiki’ and ‘Mastoidis’ pollen was similar and significantly lower than that of ‘Kalamata’. The highest values of maximum diameter were observed in ‘Kalamata’ and differed significantly with ‘Mastoidis’ but not with ‘Koroneiki’. No marked variation was noted in minimum diameter and L/W ratio. ‘Kalamata’ was characterized by higher size index than the other two cultivars, though statistical difference was observed only when compared with ‘Mastoidis’. Higher pollen viability and in vitro germination was recorded for ‘Kalamata’ and ‘Mastoidis’ compared to ‘Koroneiki’. Acquired information could contribute to the establishment of a database for future olive germplasm classification studies. Keywords: Floral biology, olive, pollen ultrastructure, scanning electron microscopy

Avocado (Persea americana) is an important crop for Chania, Crete, Greece, and is grown on more than 800 ha. In November 2013, 4-year-old trees in a new avocado grove of cv. Hass grafted onto the rootstock ‘Bacon,’ previously planted in citrus trees, showed symptoms of yellowing, leaf fall, twig and branch dieback and vascular tissue discoloration. Disease incidence was estimated at 2.3% (12 out of 530 trees affected). A fungus was consistently and readily isolated from symptomatic vascular tissue, previously surface-disinfested with 95% ethanol, on acidified potato dextrose agar (APDA). After 7 days, slow-growing colonies were transferred to PDA and the growth rate of the fungus was 2.9 mm/day at 24°C in the dark. Microscopic observations revealed hyaline hyphae with many irregular, dark microsclerotia measuring 40 to 200 × 30 to 75 μm (average 94.5 × 50.3 μm) developing after 21 days of growth. Hyaline, elliptical, single-celled conidia measuring 2.8 to 7.5 × 2.5 to 4.3 μm (average 4.8 × 3.1 μm) developed on verticillate conidiophores. For molecular characterization, Verticillium dahliae specific primer pair ITS1-F/ITS2-R that amplifies the rRNA internal transcribed spacer (ITS) region was used (2). Band of expected size was amplified, sequenced, and deposited in GenBank (Accession No. KJ818294). On the basis of morphological characteristics (3) and a BLAST search with 100% identity to the published ITS sequence of a V. dahliae isolate in GenBank (KC834733.1), the fungus was identified as V. dahliae. Five 1-year-old avocado plants of cv. Hass, grafted onto the rootstock ‘Bacon,’ were used for pathogenicity tests. Artificial inoculation was performed by making a 5.0 × 3.5 mm hole in the rootstock trunk, injecting approximately 40 μl of a 2.8 × 107 conidia/ml suspension into the vessels (spores were introduced passively), sealing with Vaseline, and covering with adhesive paper tape. Five control plants were mock inoculated with sterilized distilled water. Disease symptoms that appeared 18 days post artificial inoculation were similar to those observed under natural infection conditions. Thirty-five days post artificial inoculation, disease incidence was 80%, whereas the percentage of positive V. dahliae re-isolations from infected tissues was 95% (96.7 and 93.3% from rootstock and graft, respectively). The extent of vascular tissue discoloration from the point of inoculation ranged from 11 to 62 cm, whereas V. dahliae was successfully re-isolated even from the end of the graft (approximately 60 cm above the initial inoculation point), thus confirming Koch's postulates. Neither symptoms nor positive isolations were observed in control plants. The pathogenicity test was repeated twice with similar results. Verticillium wilt of avocado has been observed in several countries including Argentina, Chile, Ecuador, Israel, Mexico, Morocco, Spain, and the United States (1). To the best of our knowledge, this is the first report of Verticillium wilt on avocado in Greece. This disease could potentially be an increasing problem in areas where young avocado trees are established on land previously planted in vegetable crops. References: (1) J. C. Goud and J. A. Hiemstra. Chapter 3 in: A Compendium of Verticillium Wilt in Trees Species, 1998. (2) E. A. Markakis et al. Eur. J. Plant Pathol. 124:603, 2009. (3) G. F. Pegg and B. L. Brady. Verticillium Wilts. CABI Publishing, Wallingford, UK, 2002.