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Pintos B.,Complutense University of Madrid | Sanchez N.,Biotecnologia Forestal | Bueno Ma.A.,Biotecnologia Forestal | Navarro R.Ma.,ETSIAM | And 3 more authors.
Silvae Genetica | Year: 2013

This paper describes a method to obtain haploid and doubled-haploid (DH) embryos using anther cultures of holm oak (Quercus ilex L.). The production of haploids and DH through gametic embryogenesis provides an attractive biotechnological tool for developing homozygous lines from heterozygous parents, which is important in breeding programs, as well as in genetic studies. As a consequence, protocols to produce homozygous plants have a significant impact on forest tree improvement. Anthers were subjected to different temperature treatments for embryo induction: a cold pre-treatment (4°C) from 3 to 7 days was carried out at the beginning, followed by a heat shock (33°C) from 2 to 5 days. Most anthers responding to these stress treatments contained vacuolated microspores, indicating that this developmental stage is responsive to embryogenesis induction in holm-oak microspores. In all cases, embryos grew from the interior of the anthers, breaking through the degenerating anther walls. Under these conditions, embryo formation occurred in 31 anthers between 46 and 95 days after culture initiation. Embryo analysis performed with flow-cytometry and DNA-microsatellite markers showed haploid profiles and/or spontaneous doubling of the chromosomes during early regeneration stages. This is, to our knowledge, the first published report on gametic embryogenesis in holm oak.

Drake F.,University of Chile | Martin M.A.,ETSIAM | Alvarez A.,ETSIAM | Molina J.R.,University of Cordoba, Spain | And 3 more authors.
Spanish Journal of Agricultural Research | Year: 2012

Seed storage proteins have been used as genetic marker in forest species to evaluate genetic variability, demonstrating its effectiveness both in conifers and broad-leaved. In conifers, megagametophyte storage proteins are particularly useful because of their haploid nature. The aim of this study was to determine whether these proteins could be used as a new marker of genetic diversity in Araucaria araucana, one of the oldest conifers of South America and a representative symbol of Chilean forest biodiversity. For this, megagametophytes from two A. araucana populations were assessed to identify polymorphic bands and to obtain a preliminary estimation of the genetic diversity. The results revealed that globulin is the best fraction for measuring the variability in the species, due to their high level of variation (20 identified bands, 11 of them polymorphic). Both populations showed high genetic diversity, with more than 92% of the variation within populations. The study highlighted that these proteins can be used to measure the genetic diversity in A. araucana, providing good information to ensure the preservation of the species genetic resources.

Jimenez-Fernandez D.,University of Cordoba, Spain | Navas-Cortes J.A.,CSIC - Institute for Sustainable Agriculture | Montes-Borrego M.,CSIC - Institute for Sustainable Agriculture | Jimenez-Diaz R.M.,ETSIAM | Landa B.B.,CSIC - Institute for Sustainable Agriculture
Plant Disease | Year: 2011

The association of Fusarium redolens with wilting-like symptoms in chickpea in Lebanon, Morocco, Pakistan, and Spain is reported for the first time, together with the molecular and pathogenic characterization of isolates of the pathogen from chickpea of diverse geographic origin. Maximum parsimony analysis of sequences of the translation elongation factor 1α (TEF-1α) gene grouped all F. redolens isolates from chickpea in the same main clade. Pathogenicity assays using three chickpea cultivars and isolates from different geographic origins indicated that F. redolens is mildly virulent on chickpea. Moreover, infection of chickpea by F. redolens induces a disease syndrome similar to that caused by the yellowing pathotype of F. oxysporum f. sp. ciceris, including leaf yellowing and necrosis that develop upward from the stem base, and premature senescence of the plant. In contrast, F. redolens does not cause discoloration of the vascular tissues in chickpea but does cause brown necrotic lesions in the tap root and necrosis of lateral roots. F. redolens is not easily differentiated from F. oxysporum f. sp. ciceris using morphology-based diagnosis, and the two species cause similar symptoms on chickpea; therefore, the use of molecular protocols should help to avoid misdiagnoses of Fusarium yellows in chickpea. © 2011 The American Phytopathological Society.

Quesada-Moraga E.,ETSIAM | Valverde-Garcia P.,ETSIAM | Garrido-Jurado I.,ETSIAM
Environmental Entomology | Year: 2012

Laboratory studies were conducted to assess the effect of soil moisture and temperature on the development of the preimaginal stages of Ceratitis capitata (Wiedemann) (Diptera: Tephritidae). The number of days required for the immature flies to complete their development and reach the adult stage (development time) were studied at five temperatures (15, 20, 25, 30, and 35°C) and under five soil moisture regimes (-2.14 Mpa [mega Pascal] [1% (wt:wt)], -0.5 MPa [5.0% (wt:wt)], -0.47 MPa [9.0% (wt:wt)], -0.28 MPa [13.0% (wt:wt)], and -0.23 MPa [17.0% (wt:wt)]). A parametric survival model describing the effect of linear and quadratic combinations of temperature and soil moisture and their interaction on the probability of completing the development over the experimental period was used. The lognormal parametric survival model was significant for C. capitata preimaginal development, with DT 50 (the development time for 50% of the preimaginal C. capitata to reach the adult stage) ranging from 12.8 to 32.4 d. The highest percentages of adult emergence at 30 d were obtained at 24.8°C and intermediate soil moistures of 5.0 to 13.0% wt:wt. The average development time of the medfly preimaginals reaching the adult stage was inversely related to temperature and ranged from 7.4 to 26.1 d. This model could allow the monitoring of medfly preimaginal natural mortality in the soil, the prediction of adult emergence under field conditions, and therefore, the identification of suitable application times in the medfly lifecycle to achieve the maximum degree of adult and preimaginal control. © 2012 Entomological Society of America.

Moral J.,ETSIAM | Jurado-Bello J.,ETSIAM | Sanchez M.I.,ETSIAM | Oliveira R.D.,Agostinho Neto University | Trapero A.,ETSIAM
Phytopathology | Year: 2012

The influence of temperature, wetness duration, and planting density on infection of olive fruit by Colletotrichum acutatum and C. simmondsii was examined in laboratory and field experiments. Detached olive fruit of 'Arbequina', 'Hojiblanca', and 'Picual' were inoculated with conidia of several isolates of the pathogen and kept at constant temperatures of 5 to 35°C in humid chambers. Similarly, potted plants and stem cuttings with fruit were inoculated and subjected to wetness periods of 0 to 48 h. Infection occurred at 10 to 25°C, and disease severity was greater and the mean latent period was shorter at 17 to 20°C. Overall, C. acutatum was more virulent than C. simmondsii at temperatures <25°C. When temperature was not a limiting factor, disease severity increased with the wetness period from 0 to 48 h. Disease severity was modeled as a function of temperature and wetness duration; two critical fruit incidence thresholds were defined as 5 and 20%, with wetness durations of 1.0 and 12.2 h at the optimum temperature. In the field, anthracnose epidemics progressed faster in a super-high-density planting (1,904 olive trees/ha) than in the high-density plantings (204 to 816 olive trees/ha) and caused severe epidemics in the super-high-density planting even with the moderately resistant Arbequina. Data in this study will be useful for the development of a forecasting system for olive anthracnose epidemics. © 2012 The American Phytopathological Society.

Anthracnose, caused by Colletotrichum spp., is a destructive disease of olive fruit worldwide. The objective of this study was to investigate the influence of agronomical and weather factors on inoculum production using detached olive fruit and on the development of epidemics in the field. The pathogen produced very large numbers of conidia on rotted (>1.87 × 108 conidia/fruit) or mummified (>2.16 × 104 conidia/fruit) fruit under optimal conditions. On mummified fruit, conidial production was highest on mummies incubated at 20 to 25°C and 96 h of wetness. Repeated washings of mummies reduced conidial production until it was very low after five washings. When mummies were placed in the tree canopy, conidial production was not reduced after 6 months (May to October); but, when they were held on the soil or buried in the soil, conidial production comparatively decreased up to 10,000 times. Anthracnose epidemics on susceptible 'Hojiblanca' and 'Picudo' during three seasons (2005-08) were influenced by rainfall, temperature, and fruit ripening, and had three main phases: the latent period (May to October); the onset of the epidemic, which coincided with the beginning of fruit ripening (early November); and disease development, which was predicted by the Weibull model (November to March). No epidemics developed on the susceptible cultivars during the driest season (2007-08) or on the resistant 'Picual' olive during any of the three seasons. These results provide the basis for a forecasting system of olive anthracnose which could greatly improve the management of this disease. © 2012 The American Phytopathological Society.

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