A review of progress towards the conservation of the Killarney Fern (Trichomanes speciosum Willd.) in the British Isles

Summary

The Killarney Fern (Trichomanes speciosum) was considered amongst the rarest and most vulnerable of Europe's plants. Research which fed into the Biodiversity Action Plan and developed alongside it, has forced us to reconsider the level of threat to this species. A hitherto overlooked stage of the life-cycle, the gametophyte, has been shown to be comparatively abundant, genetically diverse, largely threat-free and with the potential over time to regenerate the familiar sporophyte stage. This research also promoted wider public interest which itself generated additional records, such that even the sporophyte is now known at more British sites than at any time previously and the species is accordingly considered ‘recovered’. However, continued legal protection and the need for further research are still recommended.     

Population structure and conservation biology of the endangered fern Trichomanes speciosum Willd. (Hymenophyllaceae) at its northern distributional limit

Genetic diversity in the Killarney fern, Trichomanes speciosum Willd. has been investigated in south-western Scotland, the northern-most limit of the distribution of the sporophyte. T. speciosum is unique amongst European pteridophytes in that both phases of the life cycle are perennial and capable of vegetative propagation. Within sites no variation was revealed by allozyme electrophoresis, even where both generations were growing together. In contrast, diversity was observed among sites, with seven different multilocus phenotypes (MLPs) present in the area. Two of these MLPs covered large areas while the others were restricted to one, or few localities. Asexual reproduction of the gametophyte via gemmae is assumed to be the main means of dispersal in recent times, allowing single clones to become widespread, while the overall genetic variability may be attributed to sexual reproduction and spore dispersal in historic times under more favourable climatic conditions. We suggest that it is not inbreeding, nor lack of genetic variation that limits sporophyte production, but rather the prevailing climatic conditions. The sporophyte is extremely rare and vulnerable. However, when the gametophyte is considered, the species is neither threatened with extinction, nor does it appear to face the danger of marked genetic erosion, because the long-lived gametophyte stage contains all of the genetic variability present in the area and can be regarded as a valuable 'seed-bank'.     

Negative phototropism in young gametophytes of the fern Schizaea pusilla

Schizaea pusilla is a rare fern that occurs in acidic bogs and is one of the few fern species that maintains a filamentous gametophyte throughout its development. To expand our knowledge of the physiology of this fern, phototropic responses were examined in young gametophytes. In contrast to germ filaments of other fern species, apical protonemata of young gametophytes are negatively phototropic in continuous white, red and blue light at all fluence rates tested. The expression of phototropic curvature is not limited by time since apical protonemata are also negatively phototropic when they are given brief exposures of light and then placed in the dark. In other lower plant groups such as mosses and some algae, the direction of phototropic curvature can change depending on light quality and intensity, but in young gametophytes of Schizaea, negative phototropic curvature was observed in all conditions studied. Blue light is the most effective in promoting the negative phototropic response in Schizaea.     

A global molecular phylogeny of the fern genus Trichomanes (Hymenophyllaceae) with special reference to stem anatomy

To establish a basis for future taxonomic revisions and to infer the evolutionary traits of Trichomanes s.l. , one of two large filmy fern genera, molecular phylogenetic analyses were conducted using chloroplast rbcL sequences. Sampling covered most of the global distribution range of this genus throughout the tropical and temperate zones, as well as all taxonomically significant species by the addition of 51 samples principally from Asia. The evolution of eight selected characters, which were adopted as taxonomic keys and/or putatively reflect morphological regression, was optimized on the retrieved most parsimonious rbcL trees, together with new observations on the stem anatomy of 45 species. The eight robust clades retrieved within Trichomanes in the rbcL phylogeny conflict with the existing classifications. Our results also suggest that the cup-shaped sorus, the primary diagnostic character of the genus, appears in parallel in the Hymenophyllum lineage, as demonstrated by Pleuromanes (typified by Trichomanes pallidum ) included in the lineage. The suggestion that the regressive character states are homoplastic apomorphies in the evolution of the frond size, stem thickness, root system, and stem anatomy (stele and cortex) probably illustrates a strong tendency towards adaptive evolutionary transition in Trichomanes . © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society , 2007, 155 , 1–27.     

The composition of lipids and fatty acids determined at various stages of haploid and diploid generations in the fern Adiantum capillus-veneris

The composition of lipids and fatty acids was determined at various developmental stages of gametophytes and sporophytes in the fern Adiantum capillus-veneris L. When the lipid content was expressed per g fresh weight, triacylglycerol was a major reserve lipid in spores and remained the dominant lipid in the gametophytes up to the 15th day after germination, but it was a minor one in the pinnae of sporophytes. In contrast, triterpenoids were abundant in sporophytes but not detected in gametophytes. When the content of fatty acids was expressed in percentage of the total fatty acids, the content of arachidonic acid in phosphatidylcholine and linolenic acid in diacyglyceryltrimethylhomoserine and phosphatidylcholine was significantly higher in both pinnae and petioles of sporophtes than in gametophytes. Significant differences in fatty acid composition of phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol and triacylglycerol were also noted between gametophytes and pinnae of sporophytes. All these differences were attributed to the difference of generation rather than tissue differences or age.
The content of monogalactosyldiacylglycerol, hexadecatrienoic acid in monogalactosyldiacylglycerol, and 3- trans hexadecenoic acid in phosphatidylglycerol was higher in the pinnae of sporophytes than in gametophytes or in young leaves of sporophytes. Electron microscopy of the ultrastructure of chloroplasts indicated that the high content of these components was consistent with the development of chloroplasts in pinnae.     

Solute analysis and water relations of gametophyte mutants tolerant to NaCl in the fern Ceratopteris richardii

The stl1 and stl2 mutations confer low and high levels of NaCl tolerance to gametophytes of the fern Ceratopteris richardii, respectively. As an initial characterization of these mutations, the levels of various organic solutes, tissue ion content and water relations were examined in the wild-type and mutant strains in the absence and presence of 60 mol m^-3 NaCl stress (a level which results in a 20, 15 and 0% reduction in gametophyte growth in the wild-type, stl1 mutant and stl2 mutant, respectively). All strains exhibited major changes in organic and inorganic solute levels and water relations in response to 60 mol m^-3 NaCl stress. Differences in organic solute levels and water relations between the wild-type and mutant strains in the absence and in response to 60 mol m^-3 NaCl stress were minimal. Analysis of tissue ion content showed that stl1 was associated with a slight reduction in Na⁺ accumulation during 60 mol m^-3 NaCl stress. stl2 was associated with (1) higher constitutive levels of K⁺ and (2) continued selective accumulation of K⁺ and reduced accumulation of Na⁺ during 60 mol m^-3 NaCl stress. A K⁺/Na⁺ ratio close to 1 was observed in the wild-type during 60 mol m^-3 NaCl stress, while higher ratios were detected in stl1 and stl2 (1·7 and 4·0, respectively). The findings of this study suggest that the tolerance imparted by stl1 and stl2 is associated with altered ion accumulation during NaCl stress, rather than an enhanced ability to accumulate organic solutes to be used for osmotic adjustment of the cytoplasm.     

Ecological and evolutionary consequences of desiccation tolerance in tropical fern gametophytes

Ferns have radiated into the same diverse environments as spermatophytes, and have done so with an independent gametophyte that is not protected by the parent plant. The degree and extent of desiccation tolerance (DT) in the gametophytes of tropical fern species was assessed to understand mechanisms that have allowed ferns to radiate into a diversity of habitats. Species from several functional groups were subjected to a series of desiccation events, including varying degrees of intensity and multiple desiccation cycles. Measurements of chlorophyll fluorescence were used to assess recovery ability and compared with species ecology and gametophyte morphology. It is shown that vegetative DT (rare in vascular plants) is widely exhibited in fern gametophytes and the degree of tolerance is linked to species habitat preference. It is proposed that gametophyte morphology influences water-holding capacity, a novel mechanism that may help to explain how ferns have radiated into drought-prone habitats. Fern gametophytes have often been portrayed as extreme mesophytes with little tolerance for desiccation. The discovery of DT in gametophytes holds potential for improving our understanding of both the controls on fern species distribution and their evolution. It also advances a new system with which to study the evolution of DT in vascular plants.     

遮荫对东北地区四种可食用蕨类植物生长和光合特征的影响

以东北4种蕨类植物——荚果蕨(Matteuccia struthiopteris(L.)Todaro)、猴腿蹄盖蕨(Athyrium multidentatum(Dol1.)Ching)、分株紫萁(Osmunda cinnamomea(L.)var.asiatica Fernald)和蕨(Pteridium aquilinum (L.) Kuhn var.latiusculum (Desy.) Underw.ex Heller)为研究材料,分析在35%、13%和8%全光照处理下4种蕨类植物的生长和光合特征.结果发现:随着遮荫程度的增加,4种蕨类植物的株高减小,比叶面积增加,叶片含水量增加.4种蕨类植物的净光合速率、气孔导度、蒸腾速率随着遮荫程度的增加而降低,胞间CO2浓度呈上升趋势.在3种遮荫处理下4种蕨类植物的PSⅡ最大光化学效率在0.7~0.8;非光化学猝灭系数随着遮荫程度的增加而降低.4种蕨类植物的叶绿素含量,尤其是叶绿素b含量随着遮荫程度的增加而上升,但在8%全光照下蕨的叶绿素含量呈下降趋势.相比其他蕨类植物而言,蕨具有较高的比叶面积、净光合速率和量子产率,相对较低的非光化学猝灭系数和叶绿素a./b.结果表明,4种蕨类植物最适合在35%全光照下生长,虽然对较低光照条件具有一定的适应性,但生长受到一定的抑制;4种蕨类植物中,蕨具有较强的耐荫性.本研究为东北地区4种可食用蕨类植物的栽培管理及利用提供科学依据.     

Mixed mating system in the fern Asplenium scolopendrium: implications for colonization potential

Background and Aims

Human-mediated environmental change is increasing selection pressure for the capacity in plants to colonize new areas. Habitat fragmentation combined with climate change, in general, forces species to colonize areas over longer distances. Mating systems and genetic load are important determinants of the establishment and long-term survival of new populations. Here, the mating system of Asplenium scolopendrium, a diploid homosporous fern species, is examined in relation to colonization processes.

Methods

A common environment experiment was conducted with 13 pairs of sporophytes, each from a different site. Together they constitute at least nine distinct genotypes, representing an estimated approx. 95 % of the non-private intraspecific genetic variation in Europe. Sporophyte production was recorded for gametophytes derived from each parent sporophyte. Gametophytes were grown in vitro in three different ways: (I) in isolation, (II) with a gametophyte from a different sporophyte within the same site or (III) with a partner from a different site.

Key Results

Sporophyte production was highest in among-site crosses (III), intermediate in within-site crosses (II) and was lowest in isolated gametophytes (I), strongly indicating inbreeding depression. However, intragametophytic selfing was observed in most of the genotypes tested (eight out of nine).

Conclusions

The results imply a mixed mating system in A. scolopendrium, with outcrossing when possible and occasional selfing when needed. Occasional intragametophytic selfing facilitates the successful colonization of new sites from a single spore. The resulting sporophyte, which will be completely homozygous, will shed large amounts of spores over time. Each year this creates a bed of gametophytes in the vicinity of the parent. Any unrelated spore which arrives is then selectively favoured to reproduce and contribute its genes to the new population. Thus, while selfing facilitates initial colonization success, inbreeding depression promotes genetically diverse populations through outcrossing. The results provide further evidence against the overly simple dichotomous distinction of fern species as either selfing or outcrossing.     

Efficient induction of apospory and apogamy in vitro in silver fern (Pityrogramma calomelanos L.)

Silver fern (Pityrogramma calomelanos L.) is a terrestrial or lithophytic herbaceous fern used for ornamental and medicinal purposes. In its farina it produces the cytotoxic and anticancer compound dihydrochalcone. In vitro induction of apospory and apogamy, and direct field establishment of aposporous gametophytes and subsequent sporophyte development has been accomplished. Half-strength Murashige and Skoog (MS) medium with 3.33 μM N⁶-benzyladenine (BA) and 2.32 μM kinetin (Kn) showed earlier development and produced higher numbers of aposporous gametophytes than half-strength MS basal medium. Crozier explants developed higher numbers (mean value 29.2) of gametophytes, but were slower than frond explants (mean value 23.2). The gametophytes originated from the epidermal hairs progressed from uniseriate filamentous to cordate through bi-, tri- and multiseriate and spatulate stage with the development of antheridia. Reduction in the nutrient and sucrose concentrations in the media favoured apogamy. Sucrose-free 1/10 strength MS medium and agar plates developed a mean of 30.4 and 29.9 sporophytes, respectively in the light. The greenhouse-established gametophytes developed sporophytes. The established sporophytes ex vitro showed 95% survival rate. Apogamous sporophytes and the source plant showed the same chromosome numbers (2n=116). The established protocol accomplishes apogamy and apospory in silver fern, and the aposporous gametophytes can be used for genetic transformation and development of transgenic silver fern.     

Methylation of somatic and sperm DNA in the homosporous fern Ceratopteris richardii

Plants, in general, have a high proportion of their CpG and CpNpG nucleotide motifs modified with 5-methylcytosine (5mC). Developmental changes in the proportion of 5mC are evident in mammals, particularly during gametogenesis and embryogenesis, but little information is available from flowering plants due to the intimate association of gametes with sporophytic tissues. In ferns, sperm are uninucleate and free-swimming and thus are easily isolated. We have examined 5mC in DNA isolated from fern sperm and other tissues with methylation-sensitive and -insensitive restriction enzyme isoschizomers, Southern blots probed with chloroplast and nuclear ribosomal RNA genes and end-labeled restriction fragments. We conclude that fern sperm DNA is methylated to a similar or greater degree than DNA isolated from either sporophytes or gametophytes.     

Photocontrol of spore germination in the fern Thelypteris kunthii

The light requirement for germination in spores of the fern Thelypteris kunthii (Desv.) Morton was fully satisfied by a long period of continuous red light or partially by intermittent, short periods of red light. Red light-potentiated spore germination was inhibited by brief far-red light irradiation, indicating phytochrome involvement. Repeated exposure of spores to prolonged red and short far-red irradiations, or exposure of red-potentiated spores to far-red light after an extended period in darkness, led to their escape from inhibition of germination by far-red light. Prolonged irradiation of spores with blue light before or after red light treatment partially antagonized the effect of red light.     

Isocitrate lyase in germinating spores of the fern Anemia phyllitidis

Isocitrate lyase was partially purified from germinating spores of the fern Anemia phyllitidis. The enzyme requires Mg²⁺ and thiol compounds for maximal activity and has a pH optimum between 6.5 and 7.5. The K_m of the enzyme for threo-Δ_s-isocitrate is 0.5 mM. Succinate inhibits the enzyme non-competitively (K_i. 1.8 mM). The increase of isocitrate lyase activity is closely correlated with the induction of the germination process. The fall of enzyme activity during germination is associated with the decline in triglyceride reserves.     

濒危植物银杉幼树对生长光强的季节性光合响应

银杉(Cathayaargyrophylla)是我国松科中特有的单种属植物,被认为处于濒危状态。在对银杉群落多年调查研究的基础上,针对银杉幼树生长过程对光强的需求特性,我们开展了银杉幼树对光的适应性研究。试验在人工培育的银杉苗圃地,采用遮荫的方法设置不同的光环境处理(100%、45%和3%自然光强),利用气体交换技术和叶绿素荧光技术测定了3种光强下银杉叶片光合生理指标的变化,探讨了不同光环境下银杉幼树光合能力在夏季和冬季的变化及其对生长光强的响应等。结果表明:在夏季银杉生长旺盛时期,遮荫导致叶片最大光合速率(Pnmax)、羧化效率(CE)下降,但不同叶龄叶片的下降幅度不同。随生长光强的下降,银杉幼树的光补偿点(LCP)和光饱和点(LSP)有所降低,但全晴天时,低光强(3%自然光强)条件下实际的光辐射量高于当年生叶片光补偿点的累积时间约6h,而且与光饱和的区域相差极大,造成全天碳同化量低,同化物累积少,严重影响了银杉幼树的正常生长。在不同处理中全光强条件下银杉幼树长势最好,45%光强条件下幼树生长减慢。冬季银杉最大光合速率(Pnmax)、羧化效率(CE)值均低于夏季,光补偿点(LCP)和光饱和点(LSP)也较夏季降低。全光照条件下无论是当年生叶片和一年生叶片,在冬季均出现了轻微光抑制现象,适度遮荫有利于银杉抵御冬季光抑制。无论在遮荫或不遮荫条件下,冬季银杉叶片将所吸收的相对过剩光能通过非辐射途径耗散出去,表现出一种光保护策略。     

Characterization and genetic analysis of antheridiogen-insensitive mutants in the fern Ceratopteris

WARNE, T. R., HICKOK, L. G. & SCOTT, R. S., 1988. Characterization and genetic analysis of antheridiogen-insensitive mutants in the fern Ceratopteris . The pheromone antheridiogen mediates the differentiation of male gametophytes in the fern Ceratopteris . Mutants insensitive to antheridiogen were isolated using an in vitro selection procedure. Antheridiogen-insensitive mutants exhibited partial or complete insensitivity to antheridiogen, but were normal in all other respects. Two mutants were completely insensitive to antheridiogen, whereas, another mutant was insensitive to supplemented antheridiogen, but produced male gametophytes in multispore cultures. Genetic analysis suggested a single gene basis for each mutant.     

Overwintering leaves of a forest-floor fern, Dryopteris crassirhizoma (Dryopteridaceae): a small contribution to the resource storage and photosynthetic carbon gain

BACKGROUND AND AIMS: Dryopteris crassirhizoma is a semi-evergreen fern growing on the floor of deciduous forests. The present study aimed to clarify the photosynthetic and storage functions of overwintering leaves in this species. METHODS: A 2-year experiment with defoliation and shading of overwintering leaves was conducted. Photosynthetic light response was measured in early spring (for overwintering leaves) and summer (for current-year leaves). KEY RESULTS: No nitrogen limitation of growth was detected in plants subjected to defoliation. The number of leaves, their size, reproductive activity (production of sori) and total leaf mass were not affected by the treatment. The defoliation of overwintering leaves significantly reduced the bulk density of rhizomes and the root weight. The carbohydrates consumed by the rhizomes were assumed to be translocated for leaf production. Photosynthetic products of overwintering leaves were estimated to be small. CONCLUSION: Overwintering leaves served very little as nutrient-storage and photosynthetic organs. They partly functioned as a carbon-storage organ but by contrast to previous studies, their physiological contribution to growth was found to be modest, probably because this species has a large rhizome system. The small contribution of overwintering leaves during the short-term period of this study may be explained by the significant storage ability of rhizomes in this long-living species. Other ecological functions of overwintering leaves, such as suppression of neighbouring plants in spring, are suggested.     

Desiccation tolerance in the vegetative tissues of the fern Mohria caffrorum is seasonally regulated

As there is limited information on the mechanisms of vegetative desiccation tolerance in pteridophytes, we undertook a comprehensive anatomical, ultrastructural, physiological and biochemical study on the fern Mohria caffrorum . Our data show that this species is desiccation-tolerant during the dry season, and desiccation-sensitive in the rainy season. This system allows the verification of protection mechanisms by comparison of tolerant and sensitive tissues of the same species at the same developmental age. Tolerant fronds acquire protection mechanisms during drying that are mostly similar to those reported for angiosperms. These include: (i) chlorophyll masking by abaxial scales and frond curling; (ii) increased antioxidant capacity that is maintained in dry tissues; (iii) mechanical stabilization of vacuoles in the dry state; (iv) de novo production of heat stable proteins (at least one identified as a putative chaperonin); (v) accummulation of protective carbohydrates (sucrose, raffinose family oligosaccharides and cyclitols). This study has implications for the biotechnological production of drought-tolerant crops, and allows speculation on the evolution of vegetative desiccation tolerance.