Allozyme variation of oleaster populations (wild olive tree) (Olea europaea L.) in the Mediterranean Basin

As a result of the early domestication and extensive cultivation of the olive tree throughout the Mediterranean Basin, the wild-looking forms of olive (oleasters) presently observed constitute a complex, potentially ranging from wild to feral forms. Allozyme variation was analysed at 10 loci in 31 large and 44 small oleaster populations distributed in various habitats of the Mediterranean Basin and in two populations of the wild subspecies Olea europaea subsp (ssp) guanchica, endemic to the Canary islands and closely related to oleasters. At eight polymorphic loci, 25 alleles were identified. Genetic evidence that nondomesticated oleasters still survive locally was provided by the occurrence of four and one alleles shared exclusively by the eight western and two eastern oleaster populations, respectively, which were collected in forests potentially containing genuinely wild forms according to environmental, historical and demographic criteria. As reported previously from cytoplasmic and RAPDs analysis, substantial genetic differentiation was observed between the eastern oleaster populations genetically close to most olive clones cultivated in the Mediterranean Basin, and the western populations that are related to the wild Canarian populations. In addition, the occurrence of significantly lower heterozygosity in cultivated olive than in oleasters, whatever their origin, suggests that intensive selection involving inbreeding has taken place under cultivation to obtain particular characteristics in the olive cultivars.     

Quantitative and qualitative variability of epiphytic algae on one Apiaceae (<Emphasis Type="Italic">Apium nodiflorum</Emphasis> L.) in a karstic river (Southeast of France)

Spatial distribution of the firmly attached epiphytic algae was investigated directly on the leaves of one Apiaceae (Apium nodiflorum L.) commonly found in chalky Mediterranean rivers, at two sampling dates (summer and autumn). The composition and structure of the epiphytic communities of the samples were determined at different levels: along the leaves, on both their sides and within each leaflet (n = 13). Forty-eight leaflets from four randomly selected plants were cut along the longitudinal and vertical axes so as to define 16 microhabitats located on the edge, the centre and the vein. Only a few species of algae were found to be constantly present whatever the sampling date, and dominated all microzones (n = 768). Their presence seemed to depend, rather, on the architectural structure of the plant. The other algae represented the major part of the epiphytic community and were only occasionally present. Those were more subject to temporal and environmental variations and less specific of the substrate. The epiphytes showed considerable spatial heterogeneity within their microhabitats, with an evident preference for the centre and the edge of the leaves, whereas two inhospitable spaces were the vein and apex. On the other hand, when the study was performed at a larger scale (along the leaf), the epiphytic populations of the youngest leaves (near the surface) and the eldest ones (close to the sediments) were homogenous, hence suggesting that other factors, independent from the physiology of the plant, may have an impact on epiphytic distribution.     

Effects of browsing in relation to vegetation cover on common yew (<Emphasis Type="Italic">Taxus baccata</Emphasis> L.) recruitment in Mediterranean environments

Common yew (Taxus baccata L.) stands are recognized as prioritary habitats for biodiversity conservation within the European Union. The effects of browsing on the regeneration capacity and spatial dispersal of T. baccata recruits at the European southern limit of the species in the Mediterranean Basin have been herein studied. The efficacy of T. baccata recruitment has been evaluated at six localities in the Northern Sardinia mountains, which have similar altitude, climate, soil, and vegetation but have different types of uses (three were grazed by livestock and three were not). At each site, five habitats have been identified for T. baccata seed dispersal: reproductive female T. baccata canopy, reproductive female Ilex aquifolium canopy, non-fleshy-fruited tree canopy, fleshy-fruited shrubs, and open areas. The density of seedlings was found to be greater under fleshy-fruited trees (reproductive female T. baccata and I. aquifolium) than under shrubs, whereas the sapling density was higher in shrubby habitats, especially at grazed sites due to the mechanical protection afforded by the spiny shrubs against herbivores. Land use (LU) has been found to be the most important factor in determining the spatial distribution of seedlings and saplings in relation to forest habitats. Although browsers had an ephemeral but positive effect on seed germination through their trampling and the resultant scarification, this process eventually became ineffective as was shown by the occurrence of the lowest density of saplings in those habitats where the density of seedlings was the highest. The ultimate and most important effect of browsing was the sharp decrease in the density of saplings, and their almost complete extinction, in non-shrubby habitats. This study highlights the result that, in Mediterranean ecosystems, browsing constitutes the main negative factor on T. baccata seedling-sapling transition and furthermore confirms the necessity to preserve shrubby patches in the vicinity of reproductive female T. baccata and I. aquifolium to permit the regeneration of T. baccata in the presence of livestock. Moreover, at ungrazed sites, T. baccata is able to colonize non-shrubby shady habitats. The application of different management strategies to ungrazed and grazed sites should therefore be the main direction in the management and preservation of T. baccata stands in the Mediterranean region.     

Oncopsis flavicollis (L.) associated with tree birches (Betula): a complex of biological species or a host plant utilization polymorphism?

The closely related species of leafhoppers, Oncopsis flavicollis (L.) and 0. subangulata (Sahl.), are restricted to birches, Betula pendula Roth and B. pubescens Ehrh., as host plants. Morphometric discriminant analyses of adult insects from S Wales showed O. flavicollis populations from the two Betula species to be significantly different. The best discrimination was provided by characters of the male dorsal abdominal apodemes. Analyses of such apodeme morphology in populations more widely from S Britain showed three distinct types: in western localities type 1 dominantly on B. pubescens , type 2 dominantly on B. pendula and type 3 absent; in eastern localities type 3 only on B. pendula , type 2 on both species of Betula and type 1 only on B. pubescens. Acoustic calling and courtship signals of males showed clear differences between the three apodeme types of 0. flavicollis. It is concluded that the three 0. flavicollis types, together with 0. subangulata , are distinct but very closely related biological species. Contrary to earlier suggestions, this example provides no evidence for host plant utilization polymorphisms.     

Studies on genetic transformation of olive (<Emphasis Type="Italic">Olea europaea</Emphasis> L.) somatic embryos: I. Evaluation of different aminoglycoside antibiotics for <Emphasis Type="Italic">nptII</Emphasis> selection; II. Transient transformation via particle bombardment

As a first step to the establishment of a genetic transformation protocol for olive somatic embryos obtained from the seeds of cv. ‘Picual’, the efficiencies of different aminoglycoside antibiotics as selective agents to be used with the nptII marker gene, and the particle bombardment technique for transient transformation have been evaluated. Among the three antibiotics tested, paromomycin and kanamycin showed a similar inhibitory effect and, at 200 mg l⁻¹, both of them impaired callus growth after 8 weeks of culture. However, when isolated embryos were cultured in the presence of these antibiotics, a 20% of the embryos still remained viable at 400 mg l⁻¹. Neomycin was discarded as a selective agent since it showed only a moderate toxic effect. Contrary to solid medium, when olive callus was cultured in liquid medium supplemented with different paromomycin concentrations for 3 weeks, the callus growth was impaired at the lowest antibiotic concentration, 3 mg l⁻¹. Best conditions for transient transformation of olive callus using PDS-1000/He system were a 6 cm target distance and a 900 psi bombardment pressure. pCGU∆1 plasmid, containing the gus gene under the control of sunflower ubiquitin promoter yielded a significantly higher number of gus expression areas per bombarded explant than pGUSINT or pJGUS5 plasmids, where the gus gene is driven by CaMV35S promoter or CaMV35S with enhancer, respectively. Almost 45% of bombarded explants showed gus expression 12 weeks after bombardment.     

Effects of Dredging Activities on Population Dynamics of Posidonia oceanica (L.) Delile in the Mediterranean Sea: The Case Study of Capo Feto (SW Sicily, Italy)

Between 1981 and 1993 a methane pipeline was deployed between Sicily (Italy) and Tunisia. This involved the construction of a pipeline trench, which damaged the Posidonia oceanica (L.) Delile meadow at Capo Feto (SW Sicily, Italy) and disturbed the surrounding meadow. Seagrass growth and population dynamics were examined at different depth ranges and at increasing distances from the construction site outer limit (5, 15, 30, 50 m). Results showed significant differences between the shallow (10±3.3 m) and the deep (20±4.6 m) meadow as well as differences among distances. The age structure of P. oceanica varied along the distance gradient and with depth. The mortality rate decreased with distance from the trench at all depth ranges, showing that the plants close to the excavation suffered a higher level of disturbance. Turnover and annual gross shoot recruitment rate (R_gross) were higher in the shallow portion of the meadow than in the deep range. Forecast of future meadow development (R_net) close to the trench indicates that, if present conditions are maintained, shoot density will be reduced by 50% over the next 6 to 17 yrs.     

Understanding biological conservation strategies: a molecular-genetic approach to the case of myrtle (<Emphasis Type="Italic">Myrtus communis</Emphasis> L.) in two Italian regions: Sardinia and Calabria

Myrtle (Myrtus communis L.), a shrub widespread in the Mediterranean area, is the only species belonging to the Myrtaceae family growing in Europe. The pharmacological and aromatic properties of myrtle have caused a growing interest in this plant. The use of myrtle as an aromatic plant is traditionally established in the Italian regions of Sardinia and Calabria, where it is subjected to an ecological stress due to the large biomass removal for liquor production and environmental damage. The reduction in size and number of wild populations is often related to a loss of genetic variation and reproductive potential. In this study fluorescent amplified fragment length polymorphisms (fAFLPs) were employed to assess the genetic variation within and among natural populations of myrtle from Sardinia and Calabria to gain new insights into their fitness and survival potential. AMOVA analysis indicated that genetic variation was greater within populations (51.86%) than among populations (16.99%), as previously reported for outcrossing species. A significant amount of variation (31.15%) was attributed to variation between Sardinia and Calabria population groups, suggesting a genotypic differentiation between the myrtle populations of these two regions. Intra-population genetic variation, assessed by estimating expected unbiased heterozygosity (H _E), ranged from 0.0595 to 0.2595. These values resulted correlated with population extension (r = 0.918; P < 0.01) and with two reproductive parameters: seed germinability (r = 0.793; P < 0.01) and number of seeds per fruit (r = 0.631; P < 0.05). A moderate gene flow among Sardinia myrtle populations and Calabria populations (1.2719 and 1.0478, respectively) counteracts the low level of genetic variation observed in some␣populations and avoids their differentiation and isolation.     

Pollen limitation and fruit abortion in a declining rare tree,the Eurasian yew (Taxus baccata L.): A reproductive cost of ecological marginality

Populations of rare tree species such as the dioecious and anemophilous yew (Taxus baccata) are especially prone to extinction if they occupy marginal habitats. This is the case of yew populations growing in Mediterranean mountains under dry climate and a severe anthropogenic disturbance regime, which show insufficient regeneration. We examined two of the likely mechanisms driving this population decline. The study was conducted in a central Spanish yew population in 2005 and 2006. On one hand, we tested for the effect of the amount of pollen received by female flowers by means of a pollen addition experiment. On the other hand, we tested for the effect of initial fruit set on abortion and mature fruit production. Finally, we evaluated the effect of the spatial arrangement of male and female yew trees on realized fecundity. The success of fertilization and fruiting in the experimental population was pollen-limited. Female trees aborted ca. 70% of the fruit set in both years and treatments. Fruit set was significantly influenced by the females' neighbourhood, with fecundity decreasing exponentially with distance to the nearest male in both years. Overall, our results indicate that factors associated with ecological marginality have a strong influence on yew performance.     

Free-living nematodes of the genus <Emphasis Type="Italic">Alaimella</Emphasis> Cobb 1920 (Nematoda: Leptolaimidae): A description of <Emphasis Type="Italic">A. macramphis</Emphasis> sp. n. from the White Sea and a revision of the genus

Alaimella cincta Cobb 1920 and Alaimella macramphis sp. n. are described and illustrated. Both the species were first recorded for the White Sea in northern Russia. A. cincta occurs in the shells of the agglutinated foraminiferan Reophax curtus, as well as freely in bottom sediments. A. macramphis sp. n. is described from a single male sampled from bottom sediments. A. macramphis sp. n. is distinguished from two previously known species of Alaimella (A. cincta Cobb 1920 and A. truncate Cobb 1920) by having a longer body, longer cephalic setae, and a wide amphid equal to the respective body diameter. The new species also differs from A. truncate by the distinct striation of the cuticular annulations. The Alaimella species are additionally characterized by having a posterior glandular widening of the esophagus. An emended diagnosis of the genus Alaimella Cobb 1920 and a key for species identification are provided.     

A CAPS marker associated with the partial restoration of cytoplasmic male sterility in chili pepper (<Emphasis Type="Italic">Capsicum annuum</Emphasis> L.)

Cytoplasmic male sterility (CMS), an economically important trait for hybrid seed production in many crops, is a maternally inherited trait in which a plant fails to produce functional anthers, pollen grains, or male gametes. It has long been reported that the restoration of CMS in chili pepper is controlled by a major nuclear gene termed restorer-of-fertility (Rf), along with several modifiers and some environmental factors. In this study, we identified the partial restoration (pr) locus related to the fertility restoration of CMS, demonstrated the inheritance of the trait, and developed a CAPS marker closely linked to the locus. The partially restored plant had normal anthers that produced a mix of normal and aborted pollen grains that stuck tightly to the anther wall, even after dehiscence. This trait was expressed only when the pepper plant had the sterile (S) cytoplasm and homozygous recessive pr alleles. A total of 768 AFLP primer combinations were screened, and bulked segregant analysis (BSA) was performed by preparing two pools of eight Pr/Pr (fully fertile) and eight pr/pr (partially fertile) plants, respectively, selected from the 87 individuals of the F₂ segregating population. Of the eight Pr-linked AFLP markers that were identified, E-AGC/M-GCA₁₂₂ and E-TCT/M-CCG₁₁₆ were the closest to the locus, estimated at about 1.8 cM in genetic distance. E-AGC/M-GCA₁₂₂ was converted into a CAPS marker, PR-CAPS, based on the sequences of the internal and flanking regions of the AFLP fragment. This PR-CAPS marker could be useful in selecting fully fertile lines (Pr/Pr) and eliminating partially fertile (pr/pr) and potential (Pr/pr) lines in segregant populations during the development of new inbred restorer lines.     

A review of white rust (Puccinia horiana Henn.) disease on chrysanthemum and the potential for its biological control with Verticillium lecanii (Zimm.) Viégas

The biology, aetiology and epidemiology of Puccinia horiana, the cause of white rust disease of chrysanthemum (Chrysanthemum spp.) is reviewed in relation to current environmental, cultural and chemical methods for its control. Importantly, basidiospore release, germination and infection can take as little as 5 h at optimum r.h. (96%) and temperature (between 17–24°C). Recent developments using the fungus Verticillium lecanii for the control of insects on glasshouse-grown all-year-round chrysanthemums rely upon the maintenance of r.h. during night periods in excess of 95%, thus predisposing plants to white rust attack. However, V. lecanii is unusual in that it can also parasitise spores and fruiting structures of a range of rust fungi including P. horiana. This mycoparasitic ability is also reviewed, and against this background, the potential for an integrated insect and white rust control programme on all-year-round chrysanthemums is assessed.     

A molecular phylogeny of the wild onions (Allium; Alliaceae) with a focus on the western North American center of diversity

Nuclear ribosomal DNA (ITS and ETS) sequences from 39 native Californian (USA) Allium species and congeners were combined with 154 ITS sequences available on GenBank to develop a global Allium phylogeny with the simultaneous goals of investigating the evolutionary history (monophyly) of Allium in the Californian center of diversity and exploring patterns of adaptation to serpentine soils. Phylogenies constructed with ITS alone or ITS in combination with ETS provided sufficient resolution for investigating evolutionary relationships among species. The ITS region alone was sufficient to resolve the deeper relationships in North American species. Addition of a second marker (ETS) further supports the phylogenetic placements of the North American species and adds resolution within subgenus Amerallium, a clade containing many Californian endemics. Within the global phylogeny, the native North American species were found to be monophyletic, with the exception of Allium tricoccum and Allium schoenoprasum. All native Californian species included in the analysis fell into a monophyletic subgenus Amerallium section Lophioprason, although endemic Californian species were not monophyletic due to the inclusion of species with ranges extending beyond the California Floristic Province. The molecular phylogeny strongly supports previous morphology-based taxonomic groupings. Based on our results, serpentine adaptation appears to have occurred multiple times within section Lophioprason, while the ancestor of the Californian center of diversity may not have been serpentine-adapted.     

Impact of suspended mussels (Mytilus edulis L.) on plankton communities in a Magdalen Islands lagoon (Québec, Canada): A mesocosm approach

The Grande-Entrée Lagoon (Magdalen Islands, Canada) has supported mussel (Mytilus edulis) cultivation for the last 25 years. Algal biomass in this lagoon is relatively low while heterotrophic plankton biomass is high. Although often considered herbivorous, it is known that filter-feeding bivalves can consume various types of food, from bacteria to zooplankton. We hypothesize that along with phytoplankton, heterotrophs constitute an important food resource for the Grande-Entrée mussels. In situ mesocosm experiments were undertaken at different seasons using short socks filled with mussels from the same cohort taken from an aquaculture farm, in order to determine the impact of cultured mussels on local plankton communities and assess the role of heterotrophs. Filtration activity by the mussels and associated epibionts present in the socks was expressed as clearance rates (CR). The average CR over all taxa was lowest in June and highest in October. Diatoms, dinoflagellates and heterotrophic protists constituted the bulk of planktonic carbon removed by mussels. While smaller-sized taxa contributed little (< 5%) to mussel carbon intake, large-sized heterotrophs (namely ciliates) contributed 69 to 88%. Taxon-marker pigment analyses generally confirmed these observations for groups containing phototrophic pigments. The high heterotrophic biomass retained by mussels indicates they are a major food source for mussels in this environment and should be considered both in the evaluation of mussel feeding and in assessing the influence of cultured mussels on local plankton ecosystems.     

Development of sequence-specific PCR markers associated with a polygenic controlled trait for marker-assisted selection using a modified selective genotyping strategy: a case study on anthracnose disease resistance in white lupin (<Emphasis Type="Italic">Lupinus albus</Emphasis> L.)

Selection for anthracnose disease resistance is one of the top priorities in white lupin (Lupinus albus) breeding programs. A cross was made between a landrace P27174 (resistant to anthracnose) and a cultivar Kiev Mutant (susceptible). The progeny was advanced to F₈ recombinant inbred lines (RILs). Disease tests on the RIL population from field trials over 2 years indicated that the disease resistance in P27174 was polygenic controlled. A modified selective genotyping strategy was applied in the development of molecular markers linked to quantitative loci conferring anthracnose diseases resistance. Eight individual plants representing high level of anthracnose resistance (HR), eight plants representing susceptibility (S), together with eight lines representing medium level of anthracnose resistance (MR), were subjected to DNA fingerprinting by Microsatellite-anchored Fragment Length Polymorphisms (MFLP). Six MFLP polymorphisms, which had the banding pattern matching the HR plants and the S plants, were identified as candidate markers linked to quantitative loci conferring anthracnose resistance. The six candidate MFLP markers were delineated into three groups based on their banding variation on the eight MR plants. One candidate MFLP marker each from the three groups was selected, cloned, sequenced, and converted into co-dominant, sequence-specific PCR markers. These three markers, designated as WANR1, WANR2 and WANR3, were tested on a segregating population containing 189 F₈ RILs. The disease phenotyping data and the marker genotyping data on the F₈ RILs were merged and analysed by the JMP software using the ‘fit-model’ function, which revealed that 71% of the phenotypic variation was controlled by genetic factors, while the other 29% of the phenotypic variation was due to environmental factors and environment × genotype interactions. On individual marker basis, marker WANR1 conditioned 39% of phenotypic variations of anthracnose resistance, followed by marker WANR2 with 8%, and WANR3 with 12%. Further analysis showed that WANR2 and WANR3 were on the same linkage group with a genetic distance of 15.3 cM. The combination of the two markers WANR1 and WANR3 explained 51% out from the 71% of the genetic controlled variations for disease resistance, indicating that the two QTLs working additively for anthracnose disease resistance. A simulation of marker-assisted selection on the F₈ RIL population using the two markers WANR1 and WANR3 identified 42 out of the 189 RILs being homozygous for resistance-allele bands for both markers, and 41 of them showed disease severity below 3.0 on the 1 (highly resistant) to 5 (susceptible) scale. The two markers WANR1 and WANR3 have now been implemented for marker-assisted selection for anthracnose resistance in the L. albus breeding program in Australia.     

Quantifying two-dimensional dichromatic patterns using a photographic technique: case study on the shore crab (<Emphasis Type="Italic">Carcinus maenas</Emphasis> L.)

Contrasting patterns of pigmentation, such as those associated with crypsis and aposematism, are common in many taxa. In order to determine why patterning varies among individuals or populations, it is important to quantify how these patches of pigment are arranged. Here we present a simple technique for measuring areas of pigmentation as well as their spatial distribution, and demonstrate its application to the study of substrate-associated patterning in shore crabs (Carcinus maenas L.). The results, based on a virtual grid laid over digital images of crab carapaces, allow for correlations to be made among sample populations. The technique, or variations of it, can be applied to any situation where two-dimensional dichromatic patterns need to be quantified.