Enhanced seed defenses potentially relax selection by seed predators against serotiny in lodgepole pine

Serotiny, the retention of seeds in a canopy seed bank until high temperatures cause seeds to be released, is an important life history trait for many woody plants in fire‐prone habitats. Serotiny provides a competitive advantage after fire but increases vulnerability to predispersal seed predation, due to the seeds being retained in clusters in predictable locations for extended periods. This creates opposing selection pressures. Serotiny is favored in areas of high fire frequency, but is selected against by predispersal seed predators. However, predation also selects for cone traits associated with seed defense that could reduce predation on serotinous cones and thereby relax selection against serotiny. This helps explain the elevated defenses in highly serotinous species. However, whether such interactions drive variation in seed defenses within variably serotinous populations has been studied rarely. We investigated the effects of phenotypic selection exerted by red squirrel (Tamiasciurus hudsonicus) predation on Rocky Mountain lodgepole pine (Pinus contorta latifolia) seeds. Squirrels preferentially harvested cones with more and larger seeds, indicating a preference for a higher food reward. We found evidence for stronger selection on trees with serotinous cones, which presumably accounts for the elevated defenses of and lower predation on serotinous compared to non‐serotinous cones. Lower levels of predation on serotinous cones in turn lessen selection against serotiny by squirrels. This has important implications because the frequency of serotiny in lodgepole pine has profound consequences for post‐fire communities and ecosystems widespread in the Rocky Mountains.     

Variation in serotiny of three Banksia species along a climatic gradient

The degree of serotiny (i.e. the proportion of follicles remaining closed in each year's crop of cones since the last fire) was measured in Bank-sia attenuata, B. menziesii and B. prionotes at five sites along a climatic gradient extending 500 km north of Perth, Western Australia. The decrease in annual rainfall and increase in average temperature along the gradient paralleled a decrease in plant height and an increase in the degree of serotiny of all species. Extreme serotiny was recorded in the scrub-heath at the xeric end of the gradient whereas two species were essentially non-serotinous in the low woodland at the most mesic site. It is concluded that degree of serotiny is related to the fire characteristics of the site which depend on plant height. In xeric scrub-heath, the entire canopies of the Banksia spp. are consumed by fire which promotes massive release of seed. This facilitates recruitment in an otherwise unpredictable and unreliable seedbed. In mesic woodland, where cones rarely come into contact With flames, seeds are released spontaneously and site conditions are more conducive to recruitment in the inter-fire period.     

Non‐fire induced seed release ina weakly serotinous pine: climatic factors,maintenance costs or both?

The advantages of canopy seed retention (serotiny) for plants inhabiting fire‐prone ecosystems are well documented. However, very few species are completely serotinous and non‐fire induced opening of serotinous fruits is commonly observed (weak serotiny). Two non‐mutually exclusive causes are envisaged to contribute to this process: mechanical changes in serotinous fruits mediated by climatic conditions (e.g. drought) or the costs of maintenance for the plant of these long‐lasting structures. However, their relative contribution to the spontaneous opening of serotinous fruits remains elusive as well as the consequences for the build‐up of the canopy seed bank and inter‐individual differences in serotiny. In this study we monitored the dynamics of cone production and cone opening in the weakly serotinous Pinus halepensis for five years (2004–2008), including two severe drought episodes (2005, 2006). Drought decreased the production of conelets, increased the abortion of immature cones, reduced the seed quality in the cohorts of cones produced during these years, and increased the opening of serotinous cones. During the first drought episode, a higher proportion of serotinous cones opened in those pines bearing a larger crop of younger cones. This suggests that not only passive changes induced by drought but also competition among cones for resources (e.g. water) might be involved in this process. The opening of serotinous cones in pines bearing more cones made inter‐individual differences in the size of the canopy cone bank to narrow or even to reverse from 2004 to 2008. These results may help to understand the decrease in serotiny when pines grow and accumulate more cones and the large inter‐individual variability in the degree of serotiny observed in P. halepensis forests. In addition, the negative effects of drought episodes for the size of the canopy cone bank and the seeds contained can be an unexplored cause of post‐fire regeneration constraint.     

Fruit-seed relations in Hakea: serotinous species invest more dry matter in predispersal seed protection

Abstract The ability to maintain a canopy stored seedbank (serotiny) is characteristic of many woody genera inhabiting fire-prone environments. The relationship between level of serotiny for 94 Hakea (Proteaceae) species in southwestern Australia and follicle mass, density, three wall thicknesses and seed mass was investigated. Two species were non-serotinous (fruits open at maturity), 12 were weakly serotinous (majority of fruits open at maturity), 9 were moderately serotinous (fruits open within five years of maturity) and 71 were strongly serotinous (fruits still closed at least five years after reaching maturity). A positive relationship existed between the level of serotiny and follicle morphology. Strongly serotinous species were more likely to have heavier, woodier and thicker-walled follicles than non- and weakly serotinous species. Moderately/strongly serotinous species invested more energy (six times higher follicle:seed mass ratio) than non weakly serotinous species, consistent with increased protection of the seeds from granivores, pathogens, desiccation and/or heat. Recent work has shown that thicker fruit walls (strongly serotinous species) provide better insulation to seeds from heat, although the need to survive fire is just as critical for thin-walled, weakly serotinous species. Greater protection from granivores may provide a better explanation for the adaptive significance of dense, thick-walled serotinous follicles, as the opportunities for predispersal granivore damage are low among weakly serotinous species.     

Canopy seed bank structure in relation to: fire,tree size and density

To assess the canopy seed bank structure of Pinus halepensis, we measured the level of serotiny and the seed bank size and density of trees in unburned stands and post-fire regenerated stands in Israel. We analysed the effects of tree size, tree density and fire history on the level of serotiny. The level of serotiny decreased with an increase in tree height. The high level of serotiny in short trees could be explained by selection to increase regeneration chances after burning at pre-mature age. Also, limitation of long-distance seed dispersal opportunities in short trees may favour high serotiny levels. The level of serotiny was higher in post-fire stands than in unburned stands, suggesting a fast selection for serotiny by fire. Unburned stands had a higher total stand seed density than post-fire regenerated stands, but the proportion of seeds in serotinous cones of the total stand seed density was higher in post-fire regenerated stands. The fact that P. halepensis bears simultaneously serotinous and non-serotinous cones reflects its dual strategy as both a post-fire obligate seeder, mainly from serotinous cones and an early coloniser during fire-free periods, mainly from non-serotinous cones. The relative investment in these strategies is dependent on fire history and varies with tree height. Furthermore, mature brown cones can contribute to post-fire regeneration in case of spring fires, and serotinous cones are known to open partially also in dry spell events. Thus, post-fire regeneration and invasion are strategies, which seem to complement each other.     

The invasive potential of Australian banksias in South African fynbos: A comparison of the reproductive potential of Banksia ericifolia and Leucadendron laureolum

Abstract Several taxa of the Australian Proteaceae have invaded South African fynbos and require costly management programmes to bring under control. Banksia spp. have been introduced only recently to fynbos regions. The invasive potential of Banksia ericifolia (Proteaceae) was investigated by comparing its recruitment potential with that of an indigenous proteaceous shrub, Leucadendron laureolum. Both species are overstorey shrubs that are killed by fire and rely on canopy-stored seeds (serotiny) for recruitment. Eight year old B. ericifolia shrubs produced an average of 16 500 seeds per plant, which is thirty times more than the average of 570 seeds produced by 10 year old L. laureolum shrubs. The seed bank of B. ericifolia was not only larger than that of L. laureolum (1098 vs 525 viable seeds m^?2 projected canopy cover), but also considerably larger than that described for the species in its native environment (200–330 seeds m^?2 in a 9 year old stand north of Sydney). Leucadendron laureoleum released most of its seed a few days after the cones were burnt, whereas seed release in B. ericifolia was spread over 12 weeks. The seeds of B. ericifolia had lower wingloading and fall rates than L. laureolum and were dispersed over greater distances. The relative seedling growth rates of the two species were very similar (0.03 g per day), but below-ground biomass was greater and proteoid roots were more developed in B. ericifolia seedlings than in L. laureolum after 100 days. Four year old B. ericifolia plants growing in the field had attained over twice the height of indigenous pro-teoids and accumulated up to 10 times the fresh biomass of L. xanthoconus, a species which is ecologically similar to L. laureolum. The Bioclimatic Prediction System (BIOCLIM) was used to create a bioclimatic profile of B. ericifolia and identify climatically suitable areas in the Cape Province. Results show that its potential distribution covers most fynbos areas in the southwestern Cape. It is concluded that B. ericifolia has the potential to be highly invasive in fynbos.     

Expression of gibberellin mutations in fruits of Pisum sativum L.

The gibberellin (GA) economy of young pea (Pisum sativum L.) fruits was investigated using a range of mutants with altered GA biosynthesis or deactivation. The synthesis mutation lh-2 substantially reduced the content of both GA₄ and GA₁ in young seeds. Among the other synthesis mutations, ls-1, le-1 and le-3, the largest reduction in seed GA₁ content was only 1.7-fold (le-1), while GA₄ was not reduced in these mutants, and in fact accumulated in some experiments (compared with the wild type). Mutation sln appeared to block the step GA₂₀ to GA₂₉ in young pods and seeds, but not as strongly as in older seeds. Mutations ls-1, le-1 and le-3 markedly reduced pod GA₁ levels, but pod elongation was not affected. After feeds of [¹³C,³H]GA₂₀ to leaves, the pods contained ¹³C,³H-labelled GA₂₀, GA₁, GA₂₉ and GA₈₁, and the seeds, [¹³C,³H]GA₂₀ and [¹³C,³H]GA₂₉. These findings are discussed in relation to recent suggestions regarding the role and origin of GA₁ in pea fruits. Received: 6 June 1997 / Accepted: 15 July 1997     

Evolution of serotiny in maritime pine (Pinus pinaster) in the light of increasing frequency of fires

Wildfire frequency and intensity in the Mediterranean region are predicted to increase with climate and anthropogenic changes in the following decades. Pines species often posses fire-embracing and fire-avoiding strategies that increase the probability of persistence and performance in fire-prone habitats. One such strategy is serotiny, i.e., the capacity to retain seeds in long-closed cones within the plant canopy; serotinous cones release seeds only when either a fire or a heat shock occurs. In this work, we used a simulation approach and Pinus pinaster populations as a model system to investigate how (i) an increased frequency of fire, (ii) genetic characteristics of serotiny, and (iii) observed differences in life histories interact to determine (a) risk of local population extinction and (b) temporal changes in the prevalence of serotiny in the modeled population. In addition, we tested whether the contemporary evolution of serotiny in the face of increased probability of occurrence of fires increased the probability of population persistence with respect to a scenario in which serotiny was not allowed to evolve. Our simulations showed that over the 300 years of simulated time, the evolution of serotiny did not substantially contribute to the persistence of populations. Extinction risk increased with the increasing probability of occurrence of fire and slightly decreased with (i) higher gene flow from outside the modeled population, and (ii) higher prevalence of serotiny at the beginning of the simulation. The prevalence of serotiny at the end of simulation time was difficult to predict and mostly driven by stochasticity.     

Phosphorus accumulation in Proteaceae seeds: a synthesis

The family Proteaceae dominates the nutrient-poor, Mediterranean-climate floristic regions of southwestern Australia (SWA) and the Cape of South Africa. It is well-recognised that mediterranean Proteaceae have comparatively large seeds that are enriched with phosphorus (P), stored mainly as salts of phytic acid in protein globoids. Seed P can contribute up to 48% of the total aboveground P, with the fraction allocated depending on the species fire response. For SWA species, 70–80% of P allocated to fruiting structures is invested in seeds, compared with 30–75% for Cape species, with SWA species storing on average 4.7 times more P per seed at twice the concentration. When soil P is less limiting for growth, seed P reserves may be less important for seedling establishment, and hence plants there tend to produce smaller seeds with less P. For Australian Hakea and Grevillea species the translocation of P from the fruit wall to the seed occurs in the days/weeks before final fruit dry mass is reached, and accounts for 4–36% of seed P. Seed P content increases with the level of serotiny, though it decreases marginally as a fraction of the total reproductive structure. The greater occurrence of serotiny and higher seed P content within the Proteaceae in SWA supports the notion that SWA soils are more P-impoverished than those of the Cape.     

Elevated CO2 decreases seed germination in Arabidopsis thaliana

The impact of elevated [CO₂] on seed germination was studied in different genotypes of Arabidopsis thaliana from natural populations. Two generations of seeds were studied: the maternal generation was produced in the greenhouse (present-day conditions), the offspring generation was produced in two chambers where the CO₂ concentration was either the present atmospheric concentration (about 350 ppm) or elevated (700 ppm). The seeds were tested for proportion of germinated seeds and mean germination time in both chambers to study the impact of elevated [CO₂] during seed production and germination. Elevated [CO₂] during maturation of seeds on the mother-plants decreased the proportion of germinated seeds, while elevated [CO₂] during germination had no effect on the proportion of germinated seeds. However, when seeds were both produced and germinated under elevated [CO₂] (situation expected by the end of next century), germination was slow and low. Moreover, the effect of the [CO₂] treatment differs among genotypes of Arabidopsis: there is a strong treatment × genotype interaction. This means that there is ample genetic variance for a selective response modiying the effects of high levels of [CO₂] in natural populations of Arabidopsis thaliana. The outcome at the community level will depend on what seeds are available, when they germinate and the resulting competition following germination.     

Heritability and quantitative genetic divergence of serotiny,a fire-persistence plant trait

Background and Aims

Although it is well known that fire acts as a selective pressure shaping plant phenotypes, there are no quantitative estimates of the heritability of any trait related to plant persistence under recurrent fires, such as serotiny. In this study, the heritability of serotiny in Pinus halepensis is calculated, and an evaluation is made as to whether fire has left a selection signature on the level of serotiny among populations by comparing the genetic divergence of serotiny with the expected divergence of neutral molecular markers (Q_ST–F_ST comparison).

Methods

A common garden of P. halepensis was used, located in inland Spain and composed of 145 open-pollinated families from 29 provenances covering the entire natural range of P. halepensis in the Iberian Peninsula and Balearic Islands. Narrow-sense heritability (h²) and quantitative genetic differentiation among populations for serotiny (Q_ST) were estimated by means of an ‘animal model’ fitted by Bayesian inference. In order to determine whether genetic differentiation for serotiny is the result of differential natural selection, Q_ST estimates for serotiny were compared with F_ST estimates obtained from allozyme data. Finally, a test was made of whether levels of serotiny in the different provenances were related to different fire regimes, using summer rainfall as a proxy for fire regime in each provenance.

Key Results

Serotiny showed a significant narrow-sense heritability (h²) of 0·20 (credible interval 0·09–0·40). Quantitative genetic differentiation among provenances for serotiny (Q_ST = 0·44) was significantly higher than expected under a neutral process (F_ST = 0·12), suggesting adaptive differentiation. A significant negative relationship was found between the serotiny level of trees in the common garden and summer rainfall of their provenance sites.

Conclusions

Serotiny is a heritable trait in P. halepensis, and selection acts on it, giving rise to contrasting serotiny levels among populations depending on the fire regime, and supporting the role of fire in generating genetic divergence for adaptive traits.     

Sunflower seed deterioration as related to moisture content during ageing, energy metabolism and active oxygen species scavenging

The objectives of the present work were to investigate whether loss of sunflower (Helianthus annuus L.) seed viability was affected by the embryo moisture content (MC) during seed pretreatment at 35°C, and was related to changes in energy metabolism and in the antioxidant defence system. Non‐dormant seeds were equilibrated at MC of the embryonic axis ranging from 0.037 to 0.605 g H₂O g^?1 dry matter (DM) for 1 day at 15°C, and they were then placed at 35°C for various durations up to 14 days before the germination assays at 15°C. As expected, the higher the MC, the faster was seed deterioration. There existed a negative linear relationship between the time taken for germination to drop to 50% (P₅₀) and the embryonic axis MC ranging from 0.108 and 0.438 g H₂O g^?1 DM. In dry seeds, adenosine triphosphate (ATP), adenosine diphosphate (ADP) and adenosine monophosphate represented 6.3, 14.8 and 70.9% of the adenylate pool, respectively, and the energy charge (EC) was very low (0.14). ATP and ADP levels and EC increased sharply during the first day of equilibrium of seeds at a MC above 0.158 g H₂O g^?1 DM. Subsequent controlled deterioration at 35°C resulted in a decrease in the adenylate pool, and consequently in ATP level. The higher the energy metabolism during ageing, the lower was seed viability. Loss of seed viability was associated with an accumulation of H₂O₂, and then of malondialdehyde (MDA) suggesting that lipid peroxidation was not the only cause of seed deterioration. When there was a sublinear relationship between H₂O₂ content in the embryonic axis and seed viability, MDA accumulation only occurred when 50% of the seed population died within 7 days, i.e. when MC was higher than 0.248 g H₂O g^?1 DM. Ageing was associated with a decrease in the activity of superoxide dismutase, catalase and glutathione reductase, the main enzymes involved in cell detoxification. The involvement of seed MC, as key factor of ageing is discussed with regards to energy metabolism and the regulation of active oxygen species accumulation.     

Reproductive traits of two closely related species-pairs on adjacent,different soil types in South African Fynbos

High levels of edaphic endemism and soil-related beta-diversity in Agulhas Plain fynbos communities suggest that reproductive traits of plants growing on different fynbos soils would be related to differences in soil regime. We investigated reproductive traits in two closely related Proteaceae species-pairs growing on adjacent soil types: Protea obtusifolia and Leucadendron meridianum occurring in shallow pockets of limestone-derived soils, and P. susannae and L. coniferum on the adjacent, uniformly deep colluvial sands.We found that species growing on the limestone soil comprised smaller plants, with fewer cones and seeds per plant, than species on the colluvial sands. These differences suggest that the small soil pockets of limestone soil limit plant size, in turn limiting the number of reproductive structures. Annual variation in cones and seed was the same in all species. There were no consistent trends in degree of serotiny, or sex allocation across soil types. The higher cone and seed predation levels of both limestone species than the colluvial sands species were ascribed to the higher plant densities of the former leading to lower insect search times. It was concluded that there were no overall patterns in reproductive traits that could be ascribed to differences in soil regime, other than through size-related effects. Fire regime is likely to have played a more important role in determining reproductive traits.Department of Statistical Sciences, University of Cape Towm     

Reversal of ethylene action on cocklebur seed germination in relation to duration of pre-treatment soaking and temperature

Abstract At 23°C, both C₂H₄ and CO₂ stimulated the germination of freshly imbibed upper cocklebur (Xanthium pennsylvanicum Wallr.) seeds, but C₂H₄, unlike CO₂, changed to an inhibitor of germination under some soaking conditions. However, when seeds were pre-soaked for more than several hours at 23 °C prior to treatment, C₂H₄ strongly inhibited their germination at 33 °C, the degree of inhibition increasing with the duration of pre-soaking. Maximum inhibition occurred at 1–3 cm³ m^?3 C₂H₄ when seeds were pre-soaked for 1 week; further increases of C₂H₄ concentration and pre-soaking period decreased the inhibitory effect. C₂H₄ was synergistic with CO₂ when C₂H₄ promoted germination, whereas it was antagonistic when inhibitory. Such a transition of the C₂H₄ action occurred at ca. 27 °C. Also 1-andnocyclopropane-1-carboxylic acid, a C₂H₄ precursor, inhibited the germination of pre-soaked seeds at 33 °C, although it promoted the germination at 23 °C. When pre-soaked seeds were prepared for germination by chilling at 8 °C for 3 d, the inhibitory effect of C₂H₄ on the subsequent germination was manifested even at 23 °C. The reversal of the C₂H₄ action from promotion to inhibition in cocklebur seed germination is discussed in relation to the engagement of two respiratory pathways in the imbibed seeds.     

Fire, soil fertility and delayed seed release: a community analysis of the degree of serotiny

Delayed seed release (serotiny) is a convergent plant trait in fire-prone regions of the world but explaining the degree of serotiny has remained elusive because of the paucity of community data. Selective forces involving seed predators, fire and soil nutrients have been suggested as factors influencing serotiny. We tested whether protection of seeds and/or synchronized dispersal were associated with different levels of serotiny and if resprouting ability influences selection for strong serotiny. We compared the numbers and abundance of 146 woody species with delayed dispersal among five community types varying in combinations of fire severity, fire frequency, soil fertility and seed predators. The strength of the relationship between levels of serotiny and environmental factors was tested among community types ranging from rainforests to heathlands. Highest levels of serotiny were recorded in low nutrient shrublands with intermediate fire return intervals that burn at high severity, while the lowest were recorded in high nutrient, low flammability forests. Both protection of seeds and synchronized seed release were related to fire effects in nutrient-limited environments. Strong serotiny is prominent in species killed by fire whereas weak serotiny is more common in resprouting species. Recruitment failure in the inter-fire interval appears to drive selection for strong maternal care of seeds and synchronized seed dispersal in fire-prone environments. Weak serotiny is proposed as a bet-hedging strategy that relies on resprouting after fire for population persistence and higher probability of inter-fire recruitment. The spectrum of serotiny (weak to strong) in these communities is proposed to be driven by the interactive effect of both fire and soil nutrients on the selection for delayed seed dispersal.     

Deuterium effect on ionization and fragmentation patterns of monosaccharides ionized by atmospheric pressure chemical ionization

Glucose, galactose, and mannose in H₂O and D₂O were ionized by an atmospheric pressure chemical ionization (APCI) method. Isotope effects on fragmentation patterns of the monosaccharides were examined by deuterium replacement of the -OH groups to distinguish the isomers with a single mass spectrometer. The most abundant ions were the [M+H₂O]⁺ and [M_D5+D+D₂O]⁺ for using H₂O and D₂O as solvent and eluent, respectively. Major fragment ions were the [M−OH]⁺ and [M−OH−H₂O]⁺ in H₂O, while those in D₂O were the [M_D5+D−D₂O]⁺ and [M_D5+D−2D₂O]⁺. The differences in the product ions generated in H₂O and D₂O were due to enhancement of the strength of hydrogen bonding by the deuterium replacement. Variations of the ion intensity ratios of the [M−OH]⁺/[M−OH−H₂O]⁺ and [M_D5−OD]⁺/[M_D5−OD−D₂O]⁺ with the fragmentor voltage showed different trends depending on the kind of monosaccharides. By comparing the ion intensity ratios of the [M+H₂O]⁺/M⁺, [M_D5+D+D₂O]⁺/[M_D5+D]⁺, [M−OH]⁺/[M−OH−H₂O]⁺, and [M_D5+D−D₂O]⁺/[M_D5+D−2D₂O]⁺, it was possible to distinguish the isomers of monosaccharides.     

Diurnal water balance of the cowpea fruit

The vascular network of the cowpea (Vigna unguiculata [L.] Walp.) fruit exhibits the anatomical potential for reversible xylem flow between seeds, pod, and parent plant. Feeding of cut shoots with the apoplast marker acid fuchsin showed that fruits imported regularly via xylem at night, less frequently in early morning, and only rarely in the afternoon. The dye never entered seeds or inner dorsal pod strands connecting directly to seeds. Root feeding (early morning) of intact plants with ³²PO₄ or ³H₂O rapidly (20 min) labeled pod walls but not seeds, consistent with uptake through xylem. Weak subsequent (4 hours) labeling of seeds suggested slow secondary exchange of label with the phloem stream to the fruit. Vein flap feeding of subtending leaves with [¹⁴C]sucrose, ³H₂O, and ³²PO₄ labeled pod and seed intensely, indicating mass flow in phloem to the fruit. Over 90% of the ¹⁴C and ³H of fruit cryopuncture phloem sap was as sucrose and water, respectively. Specific ³H activities of transpired water collected from fruits and peduncles were assayed over 4 days after feeding ³H₂O to roots, via leaf flaps, or directly to fruits. The data indicated that fruits transpired relatively less xylem-derived (apoplastic) water than did peduncles, that fruit and peduncle relied more heavily on phloem-derived (symplastic) water for transpiration in the day than at night, and that water diffusing back from the fruit was utilized in peduncle transpiration, especially during the day. The data collectively support the hypothesis of a diurnally reversing xylem flow between developing fruit and plant.     

Sexual reproduction of Lolium perenne L. and Trifolium repens L. under free air CO2 enrichment (FACE) at two levels of nitrogen application

Field experiments in managed grassland have shown that the response of vegetative growth to elevated CO₂ is nitrogen‐dependent in grasses, but independent in N₂‐fixing legumes. In the present study, we tested whether this is also true for reproduction. We evaluated reproductive growth, flowering phenology, seed development, reproductive success and seed germination in the grass Lolium perenne L. and the legume Trifolium repens L., growing in monocultures in a free air carbon dioxide enrichment (FACE) system at ambient (35 Pa) and elevated (60 Pa) partial pressure of CO₂ and two levels of nitrogen fertilization (14 and 56 g N m^?2 a^?1). In both species, elevated CO₂ had no significant effect on sexual reproduction. In L. perenne, reproduction was mainly nitrogen‐dependent. The weak interactions between CO₂ and mineral N supply (13% more flowers and 8% more grains per spike at high N, 7% less flowers and 8% less grains at low N) were not significant. Under elevated CO₂, grain maturation was slightly enhanced and grain weight tended to decrease. No influence could be ascertained in the date of anthesis, the temporal pattern of grain growth, the rate of grain abortion and germination. Trifolium repens, grown under CO₂ enrichment at both levels of N fertilization, flowered 10 d earlier, tended to form more inflorescences per ground area and more flowers (8–12%) and seeds (>18%) per inflorescence than at ambient CO₂. The temporal pattern of seed growth was about the same in all treatments; embryo development, however, was accelerated in fumigated plants. The number of aborted seeds per pod, seed size, thousand‐seed weight and germinability did not show any influence of CO₂. Fumigated plants at high N were attacked slightly more frequently by seed‐eating weevils, which lowered the seed output per pod. In summary, the reproductive response of L. perenne and T. repens to CO₂ enrichment on the flower and inflorescence level was far weaker than expected from the results on vegetative growth.     

Anatomic basis and insulation of serotinous cones in Pinus halepensis Mill

Pinus halepensis Mill., a widespread, low elevation conifer common in Mediterranean Basin, shows a dual reproductive strategy: post-fire obligate seeder (from serotinous cones) and an early coloniser (from non-serotinous cones). Release of seeds encased in serotinous cones is induced either by fire (pyriscence, serotiny or bradychory) or by drying (xeriscence). Morphological differences in serotinous and non-serotinous cones in natural populations of P. halepensis in Southeastern Italy were analyzed. Relationships between tree size (diameter class) and serotiny were checked by counting and sampling serotinous and non-serotinous cones. The macro and microscopic characteristics that could affect cones’ opening were measured in sampled cones. Protection against high temperatures offered by wood scales was also evaluated by applying different temperatures and time exposures, and following the inner thermal raise. Results showed that non-serotinous cones had bigger resin ducts and more separate scales. Also it was highlighted that ovuliferous scales of serotinous cones were bigger and thicker. These scales had more lamellated (multilayered) sclereid cells, and were significantly thinner with a shorter lumen diameter. Continuous temperature-monitoring heat tests inside cones showed that temperatures close to the cone axis were rather low, so seed germination was not influenced. Results confirm that serotinous cones are more compact, rigid and consistent than non-serotinous cones. These characteristics explain the lower insulation, seed protection and the ease opening of non-serotinous cones as well. In conclusion, opening mechanism of pinecone scales under the effect of fire or dry conditions seem related to anatomic differences and it provides seeds with an efficient protection against heat.     

No contribution of the pentose phosphate pathway in dormancy-breaking of cocklebur seeds

The role of the oxidative pentose phosphate (PP) pathway in the dormancy-breaking of cocklebur (Xanthium pennsylvanicum Wallr.) seeds was investigated. D-[1-¹⁴C]-glucose or D-[6-¹⁴C]-glucose was fed to dormant and non-dormant lower seeds or to their axial or cotyledonary segments which were imbibed for different durations, and C₆/C₁ ratios of respired ¹⁴CO₂ as an index of the PP pathway activity were calculated. Contrary to expectation, there was no significant difference in the C₆/C₁ ratios between the dormant and non-dormant seeds or segments during a water imbition period of 24 h, although the PP pathway actually operated already in an early stage of water imbibition. Also concerning the activities of G6PDH and 6PGDH, the key enzymes of this pathway, no difference between the dormant and non-dormant seeds was found. It was thus concluded that, unlike other seeds, there is no contribution of the PP pathway to the regulation of dormancy of the cocklebur seed.