Reproductive modes in the epixylic hepatic Anastrophyllum hellerianum

The frequency and dynamics of sexual and asexual reproduction were investigated in a dioecious epixylic hepatic, Anastrophyllum hellerianum, which has declined in recent decades in Finland as a consequence of forestry practices. In our investigation asexual reproduction by gemmae was the dominant mode of reproduction and specialised gemmiparous shoots were present in all colonies studied. The proportions of dead shoots were considerably higher among sex-expressing than among non-sexexpressing shoots. Our results suggest that lower reproductive investment is required for asexual than for sexual reproduction. For instance, no trade-off is detected between asexual reproduction and survival of the gemmiparous shoots in A. hellerianum. Sexual reproduction occurred only in 12% of the colonies and it was promoted by the following factors: medium shoot density, high proportion of sex-expressing shoots, an even sex ratio and very short distances between individuals representing opposite sexes. The ratio of dead males to dead females was significantly female-biased, which suggests higher mortality among female shoots. At the level of individual shoots, more spores than gemmae were produced. However, as a consequence of the low frequency of sporophyte-bearing shoots, gemma production highly exceeded spore production at the colony level. Furthermore, cultivation tests of the propagules showed that gemmae germinate faster than spores.     

Plant regeneration from leaf petioles in <Emphasis Type="Italic">Camptotheca acuminata</Emphasis>

Summary Camptothecin, produced by Camptotheca acuminata, is a pharmaceutically important compound. Transgenic technology has potential uses for the enhancement of camptothecin production; however, an efficient plant regeneration protocol for C. acuminata is not currently available. Factors that affected successful seedling germination were evaluated. The regeneration potential of various parts of seedlings was tested. Camptothecin production in regenerated plants was compared to its production in calluses and the original seedlings. Dark incubation and seed coat removal led to a higher germination rate and a higher survival rate after germination. The best shoot induction medium was found to be Gamborg's B5 medium+8.9 μM benzyladenine. Among the calluses induced from various parts of seedlings, leaf petiole calluses, leaf dise calluses, and cotyledon calluses regenerated shoots, but internode calluses did not. Furthermore, leaf petiole calluses and leaf dise calluses regenerated normal shoots, while cotyledon calluses regenerated hyperhydric shoots. Moreover, leaf petiole calluses had a higher shoot regeneration rate, 50% versus 9%, and a higher shoot number, 6.2±0.5 versus 2.0±0.3, than did leaf dise calluses on the best shoot induction medium. It took 4–6 wk to regenerate shoots after transfer into shoot induction media. Camptothecin concentration in the regenerated plants was significantly higher than that in the calluses and similar to that in the original seedlings. In conclusion, leaf petioles provide efficient plant regeneration of C. acuminata.     

Vegetative regeneration in invasive Reynoutria (Polygonaceae) taxa: the determinant of invasibility at the genotype level

Vegetative regeneration of individual genotypes of Asian Reynoutria taxa, which are invasive in the Czech Republic, was studied in R. sachalinensis (five genotypes), R. japonica (a single genotype present in the country), and their hybrid R. ×bohemica (nine genotypes). Identity of genotypes was confirmed by isozyme analysis. Ten rhizome segments of each genotype were planted in a randomized block design. After 30 d, the regeneration rate of each genotype was measured as the proportion of rhizomes that produced shoots. Emergence time and final mass of each shoot were recorded. The regeneration rate and final shoot mass were significantly affected by genotype in R. ×bohemica but not in R. sachalinensis. In R. ×bohemica, easily regenerating genotypes grew faster. Regeneration characteristics that crucially contribute to the fitness of these vegetatively spreading plants are closely related to each other. In genotypes with a low regeneration rate, early-emerging shoots produced more biomass, while in those with a high regeneration rate, shoot mass was independent of emergence time. Mean clone size recorded in the field was marginally significantly related to emergence time during regeneration; regeneration characteristics might thus affect the extent of R. ×bohemica invasion at a regional scale. Hybrids genetically intermediate between the parents regenerated better than those closely related to parents. Novel hybrid invasive genotypes may be produced by rare sexual reproduction, fixed by clonal growth, and present a previously unknown threat to native vegetation.     

Two new species of Distichophyllum from Malaya

Abstract

The germination and establishment abilities of different types of propagules under natural conditions is an area in bryophyte reproductive biology that is largely unknown. In some species gametophyte fragments, rather than spores, may be primarily responsible for establishment of new colonies in nature, and the presence of other mosses may have an effect on establishment processes. These ideas were tested experimentally using four species of Tortula grown in a glasshouse under semi-natural conditions. The relative germination and growth abilities of spores and gametophytic fragments were tested under each combination of the following treatments: (1) continuous hydration (using a mist system), or periodic drying; (2) substrate of bare sand, a clump of Dicranum scoparium, or (for spores only) the clump of Tortula from which sporophytes were taken. Fragments produced new shoots much more quickly than spores under either hydration condition but spores germinated at a higher rate. Germination rates of both spores and fragments were greatly reduced (usually to zero) in clumps of Tortula or Dicranum under either hydration condition. Some differences between species were noted. In a further experiment filter-sterilized water extracts of each species were applied to spores on agar. Again, strong effects on spore germination and protonemal growth were noted. Patterns of inhibition differed between species and reflected those found in the previous experiment, indicating that the inhibition effect may be due to a water soluble chemical rather than the physical properties of the moss clumps. These studies present the first experimental evidence for the chemical inhibition of spores by mature plants. When taken together with previous reports of chemical interactions among protonemal systems, considerable ecological significance of these results is suggested.     

In vitro multiple shoot regeneration and plant production in Alysicarpus rugosus DC. var. heyneanus Baker

A protocol for in vitro multiple shoot regeneration and plant production through seedling (shoot tip) culture was established for Alysicarpus rugosus DC. var. heyneanus Baker. Maximum number of adventitious shoots (14.4) per shoot tip explant were initiated after two subcultures on MS solid medium supplemented with IAA (2.85 microM) plus BAP (2.22 microM) after 4 weeks. Shoot elongation (3.0-3.5 cm) was achieved on MS medium without any hormones. Stunted shoots elongated on half MS medium without growth hormones. Rooting occurred in MS medium containing IAA (1.14 - 2.85 microM) alone or in combination with IBA (0.89 - 2.46 microM) and or NAA (1.07 - 2.69 microM). Maximum rooting was established in MS medium supplemented with IAA (2.85 microM). The plants were acclimatized successfully with 55% survival in pot containing cocoa peat and sand (1:1). After a month, hardened plants were transferred to pots with manure, garden soil and sand (1:2:1) for further growth and finally planted in field.     

Cultivation of Gigartina skottsbergii (Gigartinales, Rhodophyta): Recent advances and challenges for the future

This study integrates landings statistics and biological studies of the red algaGigartina skottsbergii Setchell & Gardner. The analysis of the landings and carrageenan production in Chile suggeststhat this resource will suffer a strong harvesting pressure during the nextyears. Biological results on sporulation, germination, sporeling growth and survivorship in laboratory,indoor tanks and field conditions, indicated that cultivation of this species istechnically feasible, as spores can be seeded on ropes and other substrata. Vegetative propagation of this species through tissue fragmentationis also possible. Vegetative fragments of this carrageenophyte have 20 to30% higher growth rates than whole fronds in suspended culture systems. Protoplast production can be also explored for bypassing restrictions inspore availability. Major advantages that encourage the cultivation of G. skottsbergii include its gel quantity and quality, its pathogen-freecondition, a high reproduction potential and its regeneration capacity. Onthe other hand, the major constraints are related to its relatively slowgrowth as compared to other carrageenophytes, limited availability ofspores and high mortality during juvenile stages.     

Sexual reproduction in the phyllodioicous bryophyte Homalothecium lutescens (Hedw.) H.Rob. in relation to habitat age,growth conditions and genetic variation

Sporophyte production and female fertility were investigated in seventeen calcareous grassland demes of the moss Homalothecium lutescens (Hedw.) H.Rob. on the Baltic island of Öland, with the aim of understanding the relationships between sexual reproduction, habitat age, genetic variation and factors related to growth conditions. The overall proportion of fertile female shoots (with perichaetia) was 35%. Fertility status at the level of individual shoots was positively associated with shoot length and density, while within deme fertility was positively associated with bush cover. There was no association between female fertility and habitat age, genetic diversity (H_S) or allelic richness. Out of 1344 investigated shoots, only two were normal-sized fertile males. Dwarf males were also extremely rare, and found almost exclusively on shoots with sporophytes. Few sporophytes were observed (in the two demes with highest fertility and bush cover). No relationship between genetic variation and the frequency of sporophytes and males was found. The lack of a relationship between sexual reproduction and genetic variation suggests that sexual reproduction may not occur in the same grassland fragments as the recruitment of new clones (from spores or vegetative fragments). The majority of the dry, open grassland habitats, where H. lutescens is typically found in the study area, appear to be suboptimal for both dwarf males and fertilization. Sexual reproduction is more likely to occur in shaded (although grazed) grassland patches, where moisture levels are likely to be higher and the moss colonies are generally more vigorous.     

Acorn production is linked to secondary growth but not to declining carbohydrate concentrations in current-year shoots of two oak species

In trees, reproduction constitutes an important resource investment which may compete with growth for resources. However, detailed analyses on how growth and fruit production interact at the shoot level are scarce. Primary canopy growth depends on the development of current-year shoots and their secondary growth might also influence the number and size of fruits supported by them. We hypothesise that an enhanced thickening of current-year shoots is linked positively to acorn production in oaks. We analysed the effect of acorn production on shoot growth of two co-occurring Mediterranean oak species with contrasting leaf habit (Quercus ilex, Quercus faginea). Length and cross-sectional area of current-year shoots, apical bud mass, number of leaves and acorns, xylem and conductive area, number of vessels of acorn-bearing and non-bearing shoots were measured in summer and autumn. Nitrogen and carbohydrates analyses were also performed in stems and leaves of both shoot types. Stem cross-sectional area increased in acorn-bearing shoots when compared with non-bearing shoots for both species and such surplus secondary growth was observed since summer. In bearing shoots, the total transversal area occupied by vessels decreased significantly from basal to apical positions along the stem as did the xylem area and the number of vessels. Leaves of bearing shoots showed lower nitrogen concentration than those of non-bearing shoots. Carbohydrate concentrations did not differ in stems and leaves as a function of the presence of acorns. Such results suggest that carbohydrates may preferentially be allocated towards reproductive shoots, possibly through enhanced secondary growth, satisfying all their carbon demands for growth and reproduction. Our findings indicate that acorn production in the two studied oaks depends on shoot secondary growth.     

Dark pretreatment improves adventitious shoot organogenesis from cotyledons of diploid watermelon

The influence of light incubation during embryo germination on shoot organogenesis from cotyledons of four diploid watermelon [Citrullus lanatus (Thumb.) Matsum. & Nakai cultivars was examined. Germinating embryos in darkness significantly improved the number of explants that produced harvestable shoots during the 6 week incubation period on shoot regeneration medium under a 16-h photoperiod. The percentage of explants with shoots more than doubled for `Crimson Sweet' and was about 1.5-fold greater for `Sweet Gem' and `Yellow Doll' when embryos were germinated in darkness. The percentage of explants with shoots was not significantly improved for `Minilee' by pretreating seedlings in darkness. This study demonstrates that optimal shoot regeneration can be obtained by germinating embryos in darkness before preparing cotyledon explants. This revised version was published online in June 2006 with corrections to the Cover Date.     

Vegetative reproduction capacities of floodplain willows--cutting response to competition and biomass loss

While several studies on regeneration in Salicaceae have focused on seedling recruitment, little is known about factors controlling their vegetative reproduction. In two greenhouse experiments, we studied the response of floodplain willows (Salix fragilis, S. viminalis, S. triandra) to competition with Poa trivialis, and to shoot and root removal when planted as vegetative cuttings. In the first experiment, growth performance variables were analysed in relation to full competition, shoot competition, root competition and control, taking into account two different water levels. After 9 weeks, shoots were removed and the resprouting capacity of the bare cuttings was recorded. In the second experiment, the cutting performance of the three floodplain and an additional two fen willow species (S. cinerea, S. aurita) was compared when grown in three different soil compositions and with two different water levels. After 9 weeks, shoot and root biomass was removed and the bare cuttings were replanted to test their ability to resprout. Cutting performance and secondary resprouting were negatively affected by full and shoot competition while root competition had no or weak effects. The floodplain species performed better than the fen species in all soil types and water levels. Secondary resprouting capacity was also higher in the floodplain species, which showed an additional strong positive response to the previous waterlogging treatment. The results contribute to understanding of the vegetative regeneration ecology of floodplain willows, and suggest that the use of vegetative plantings in restoration plantings could be an effective strategy for recovering floodplain forests.     

Establishment and in vitro regeneration studies of the potential oil crop species Camelina sativa

Camelina sativa was successfully established in vitro and systems for the regeneration of shoots from leaf explants developed. Methods for the surface-sterilisation of seeds were used which gave 95% germination, though the in vitro grown seedlings failed to develop beyond 28 days culture. In a micropropagation system, the rooting response of nodal explants was increased from a control level of 26.4% to 46.7% by the addition of 5.4 μM NAA. Leaf explants were more efficient for the regeneration of root and shoots than hypocotyls. For regeneration from leaf tissue the use of auxin (NAA) alone in the medium above a level of 0.54 μM resulted in root or callus growth. Cytokinin, in the form of BA alone failed to induce regeneration, but a combination of 4.44 μM BA and 0.54 μM NAA induced shoot regeneration at rates over 10.0 shoots per explant. Regenerated shoots were successfully transplanted to soil and flowered and set seed normally. This revised version was published online in June 2006 with corrections to the Cover Date.     

High-frequency direct shoot regeneration and continuous production of rapid-cycling Brassica oleracea in vitro

Summary An in vitro method was developed for high-frequency shool regeneration and continuous production of rapid-cycling B. oleracea in large numbers. The high regenerative capacity was tissue-dependent. Developmental polarity (apical and basal ends) of the explants appeared to play a regulatory role in shoot morphogenesis in this system. High-frequency shoot regeneration was obtained with N⁶-benzyladenine or thidiazuron-supplemented media. Delayed and reduced regenerative ability of cultures in air-tight vessels and the dramatic suppression of shoot regeneration in internodal explants by the ethylene precursor 1-aminocyclopropane-1-carboxylic acid implicate a possible involvement of ethylene in shoot morphogenesis in this species. Rotting of regenerated shoots of B. oleracea occurred readily on α-naphthaleneacetic acid-supplemented media. Rooted plantlets were successfully established in soil and developed normal fertile flowers and viable seeds.     

Shoot apical meristem: In vitro regeneration and morphogenesis in wheat (Triticum aestivum L.)

Summary We report a less genotype-dependent in vitro regeneration system capable of producing multiple shoot clumps and whole plants in four different wheat genotypes. Shool apical meristems from 7-d-old-seedlings produced axillary and adventitious shoots and somatic embryos on media containing N⁶-benzyladenine (BA) and 2,4-dichlorophenoxyacetic acid (2,4-D). All four genotypes responded positively to shoot multiplication depending upon media composition. Scanning electron microscopies of cultures showed a proliferating budding state that gave rise to adventitious shoots and somatic embryos on further multiplication. The percentage of relative shoot apical meristem multiplication was 80–90%, and the average number of shoot meristems per multiplied shoot was 40–50 in all genotypes. Among different concentrations of phytohormones, 2 and 4 mgl⁻¹ BA (8.8 and 17.7 μM) in combination with 0.5 mg l⁻¹ 2,4-D (2.26 μM) gave the best results. Actively multiplying shoot clumps were recovered with high frequency among 3-mo.-old cultures. These shoot clumps regenerated normally and produced fertile plants containing viable seeds. This in vitro system might prove useful for the production of transgenic plants of wheat in a relatively genotype-independent manner.     

Formation of shoots from leaf axils of Alstroemeria: The effect of the position on the stem

Direct shoot regeneration was induced from leaf explants of Alstroemeria. The explants contained a leaf blade and a small portion of stem node, which were cut from the erect shoots of in vitro multiplicated plantlets. The shoot regeneration capacity of the excised leaf explants was significantly related to the position of the explant on the stem. The youngest explant which was located closest to the shoot apex gave the highest response. A gradient response toward the shoot apex was observed in percentage of shoot regeneration and in the number of shoots per regenerating explant. Histological studies revealed that the shoots were initiated at the leaf axils. The origin of the adventitious buds was located at the epidermal layer of stem peripheral cells. This revised version was published online in June 2006 with corrections to the Cover Date.     

Trade-offs between vegetative growth and acorn production in Quercus lobata during a mast year: the relevance of crop size and hierarchical level within the canopy

The concept of trade-offs between reproduction and other fitness traits is a fundamental principle of life history theory. For many plant species, the cost of sexual reproduction affects vegetative growth in years of high seed production through the allocation of resources to reproduction at different hierarchical levels of canopy organization. We have examined these tradeoffs at the shoot and branch level in an endemic California oak, Quercus lobata, during a mast year. To determine whether acorn production caused a reduction in vegetative growth, we studied trees that were high and low acorn producers, respectively. We observed that in both low and high acorn producers, shoots without acorns located adjacent to reproductive shoots showed reduced vegetative growth but that reduced branch-level growth on acorn-bearing branches occurred only in low acorn producers. The availability of local resources, measured as previous year growth, was the main factor determining acorn biomass. These findings show that the costs of reproduction varied among hierarchical levels, suggesting some degree of physiological autonomy of shoots in terms of acorn production. Costs also differed among trees with different acorn crops, suggesting that trees with large acorn crops had more available resources to allocate for growth and acorn production and to compensate for immediate local costs of seed production. These findings provide new insight into the proximate mechanisms for mast-seeding as a reproductive strategy.     

Efficient in vitro regeneration systems for Vaccinium species

Efficient protocols for shoot regeneration from leaf explants suitable for micropropagation as well as for the development of transgenic plants were developed for blueberry (Vaccinium corymbosum) and lingonberry (Vaccinium vitis-idaea) cultivars. Nodal segments were used to initiate in vitro shoot cultures of lingonberry cultivar ‘Red Pearl’ and southern highbush blueberry cultivar ‘Ozarkblue’. In order to develop an optimized regeneration procedure, different types and concentrations of plant growth regulators were tested to induce adventitious shoot regeneration on excised leaves from micropropagated shoots of both cultivars. The effect on percentage regeneration and number of shoots per explant was investigated. Results indicated that zeatin was superior to TDZ and meta-topolin in promoting adventitious shoot formation. A concentration of 20 μM zeatin was most effective in promoting shoot regeneration in both cultivars, in case of ‘Red Pearl’ along with 1 μM NAA. Shoots were either allowed to root in vitro on medium containing IBA or NAA or ex vitro in a fog tunnel. IBA was superior to NAA for induction of root development in vitro in both Vaccinium cultivars. Ex vitro rooting under high humidity was tested with cuttings from mature field-grown plants, from acclimatized tissue culture derived plants and with unrooted in vitro proliferated shoots planted directly. It was found that in vitro shoots rooted better under fog than cuttings from the other plant sources and rooting was equivalent to that achieved in vitro.     

Barriers to Forest Regeneration in an Abandoned Pasture in Puerto Rico

Sources of forest regeneration (soil seed bank, seed rain) and barriers to seedling establishment were examined in a recently abandoned pasture in eastern Puerto Rico. Few woody species were found in the soil seed bank or in the seed rain. The number of seeds and species in the seed rain and soil seed bank declined with distance from the adjacent secondary forest. Nine species naturally dispersed and colonized plots during the study, with the wind‐dispersed tree Tabebuia heterophylla being the predominant colonizer (91% of all seedlings). Barriers to seedling establishment were determined using a blocked field experiment with eleven woody species representative of a variety of life histories. Each species was planted under the pasture vegetation (control) or in areas where all vegetation was removed (removal). Germination was enhanced for four species in the control treatment, five species were not affected, and two species did not germinate under either treatment. Survival to 6 months was higher in the removal treatment for two species. Seedling biomass was greater in the removal treatment at 12 months for one species. Seed mass was a good predictor of germination success and final shoot biomass, but not survival. This study demonstrates that seeding recently abandoned pastures with a mix of known pioneer species may accelerate the rate of secondary succession, but some species will have to be planted in later successional stages in order to overcome strong barriers to establishment.     

Seed, cutting and air-layering reproductive inefficiency of noxious woody vine Merremia biosiana and its implications for management strategy

Merremia boisiana (Gagnep) van Ooststr. is a noxious fast growing woody vine and is able to grow overtop other plants, causing the death of plants underneath and forming monospecies stands. To formulate management responses, we assessed its seed and vegetative reproduction efficacy through indoor and field experiments. The number of flowers counted from bagged infructescences in Guangzhou ranged from 25 to 172, with an average of 80.80. Counting the seeds of bagged infructescences had shown that there were only 1.58 hard testa seeds in each infructescence. Seed vitality tests using red ink indicated that only 68.6% of hard testa and filled seeds were viable. The emergence rate of scarified hard testa seeds in the sand bed was 31.96%. Under imitated natural conditions, 8% of hard testa seeds could germinate, and 9% still retained their germination potential in one year. Thus, seedlings should be monitored and removed in a timely fashion after any attempt of clearing. Moving soil or transplanting plant from infested patches should be strictly prohibited at least for several years. All considered, an infructescence contributed 0.3 seedlings. The investigation in the field found no seedlings either inside or at the perimeter of the patch, suggesting scant expansion by means of seed dispersal. Therefore, the elimination effort could be focused on a relatively restricted scale of patches. Of 630 cuttings of young shoots, old shoots and old lying shoots with or without growth regulators, there were only four (or 0.63%) established individuals. Air-layering shoots all died in two months. Poor cuttings and air-layering reproduction indicated that regeneration from fragments of removed stems or accidentally dropped ones was quite unlikely, and thus mechanical removal was safe.     

Seed, cutting and air-layering reproductive inefficiency of noxious woody vine Merremia biosiana and its implications for management strategy

Merremia boisiana (Gagnep) van Ooststr. is a noxious fast growing woody vine and is able to grow overtop other plants, causing the death of plants under-neath and forming monospecies stands. To formulate management responses, we assessed its seed and vegeta-tive reproduction efficacy through indoor and field experiments. The number of flowers counted from bagged infructescences in Guangzhou ranged from 25 to 172, with an average of 80.80. Counting the seeds of bagged infructescences had shown that there were only 1.58 hard testa seeds in each infructescence. Seed vitality tests using red ink indicated that only 68.6% of hard testa and filled seeds were viable. The emergence rate of scarified hard testa seeds in the sand bed was 31.96%. Under imitated natural conditions, 8% of hard testa seeds could germinate, and 9% still retained their germination potential in one year. Thus, seedlings should be monitored and removed in a timely fashion after any attempt of clearing. Moving soil or transplanting plant from infested patches should be strictly prohibited at least for several years. All considered, an infructescence contributed 0.3 seedlings. The investiga-tion in the field found no seedlings either inside or at the perimeter of the patch, suggesting scant expansion by means of seed dispersal. Therefore, the elimination effort could be focused on a relatively restricted scale of patches. Of 630 cuttings of young shoots, old shoots and old lying shoots with or without growth regulators, there were only four (or 0.63%) established individuals. Air-layering shoots all died in two months. Poor cuttings and air-layering reproduction indicated that regeneration from fragments of removed stems or accidentally dropped ones was quite unlikely, and thus mechanical removal was safe.     

人工油松种群生长与生殖分配关系的研究

植物种群营养生长和生殖生长的关系是植物生殖生态学研究的重要内容之一,自本世纪70年代,国外已有大量报道,但国内的研究报道很少。本文在种群水平上研究了人工油松种群一年生雌性枝条和雄性枝条在枝长、直径、叶数、叶生物量、枝生物量及新生枝条在大小孢子球生物量、雌雄枝条长度和生物量等方面的差异,结果表明,一年生枝条除在西方向上雌雄枝条直径差异显著和在东、西方向上雌雄枝条生物量差异显著外,其它处理条件下雌雄枝条在长度、直径、叶数、叶生物量和枝生物量等方面差异均不显著;而对新生枝条的取样分析表明同方向上雌雄枝条在大小孢子球生物量、雌雄枝条长度、雌雄枝条生物量等方面差异均显著或极显著。无论是一年生枝条,还是新生枝条,在东、西、南、北四个方向上其各项指标差异显著或极显著。