GENETIC VARIATION IN THE HOMOSPOROUS FERN PELLAEA ANDROMEDIFOLIA

Genetic variability was examined in nine sexual and three apogamous natural populations of the homosporous fern Pellaea andromedifolia by electrophoretic analysis of enzymes specified by eight loci. Genetic interpretations of heterozygous banding patterns were determined by segregational analysis of gametophytes. High levels of segregating heterozygosity characterized the sexually reproducing populations, and genotype frequencies at the five polymorphic loci were consistent with those expected under conditions of random intergametophytic mating. Multiple-banded patterns in the apogamously reproducing populations resembled those of heterozygous sexual individuals, but did not segregate. The results suggest that genetic variation in sexual homosporous vascular plants is produced by cross fertilization of genetically different gametes and may not result from pairing between homoeologous chromosomes carrying duplicated loci as previously thought.     

Evidence of genetic segregation in the apogamous fern species Cyrtomium fortunei (Dryopteridaceae)

In apogamous ferns, all offspring from a parent are expected to be clonal. However, apogamous 'species' frequently demonstrate a large amount of morphological and genetic variations. Cyrtomium fortunei composed of four varieties (C. fortunei var. fortunei, var. clivicola, var. intermedium, and var. atropunctatum), is all reported to be apogamous triploids, but demonstrates large and continuous morphological variation. In previous studies, we showed that considerable genetic diversity was observed in many local populations of the apogamous fern 'species'. We hypothesized that genetic segregation has occurred, because neither sexual type nor intraspecific polyploidy have been observed in C. fortunei in Japan. Of 732 progeny examined (250 gametophytes and 482 sporophytes), obtained from a parental sporophyte whose pgiC genotype was estimated as aab, 11 (4.4%) gametophytes and 8 (1.7%) sporophytes showed a different genotype (aaa) from that of the parent sporophyte. We showed that genetic segregation occurs in apogamous C. fortunei in relatively high frequency. Moreover, we could first show that the segregation frequency in gametophytes is significantly higher than that in sporophytes of the next generation (χ (2)?=?4.90, P?=?0.027). It may suggest the existence of deleterious genes, which are expressed during the morphogenesis and growth of sporophytes.     

Size dependency of gametophytes decay inAthyrium brevifrons Nakai during spring desiccation

Gametophyte populations inAthyrium brevifrons were analysed with respect to population size and surviving area (%) of individual thalli in a transplant garden at Sapporo during 5–26 April 1983, to study the safe-microsite for gametophyte establishment in nature. Spores dispersed in August 1982 germinated and grew into thalli of various widths (<10 mm); 10.3% of the thalli matured by early October 1982. Maturation was attained by gametophytes of width 4–7 mm. The number of gametophytes gradually decreased with increasing width. By April 1983, 20.5% of total gametophytes were mature with a mode of 5–6 mm in width. The relative number of gametophytes with surviving area of 2–20% increased and that of 85–100% decreased in accordance with collection days delayed until after snow-melt. Surviving area (%) on gametophyte of all widths decreased with decreasing soil moisture contents. In particular, immature gametophytes of 2–4 mm width showed a significant correlation (P<0.01) between soil moisture content and relative number of gametophytes with 0–20% surviving area and mean surviving area (%) of every width of thalli. The spring desiccation might be a factor that reduces or limits gametophyte populations in nature.     

Growth and gender in the gametophyte of Blechnum spicant L

The growth and gender in the gametophyte of Blechnum spicant L. were strongly affected by its origin, spore or mature homogenized gametophytes, and also by the addition of plant growth regulators to the culture medium. Spore-derived gametophytes cultured in Murashige and Skoog liquid medium were females and heart-shaped, whereas those derived from mature homogenized gametophytes were shorter and male or asexual (1:3) because of the release of antheridiogen to the culture medium. In the latter, maleness was especially increased by the addition of BA. This cytokinin is a strong inductor of maleness in homogenized cultures; however, even though BA influenced sexual organ formation in spore-derived gametophytes, it does not change the␣female sexual pattern that these gametophytes have. Separate male and female populations of gametophytes were obtained in this work.     

Genetic variation in the ‘Japanese apogamous form’ of the fernAsplenium unilaterale Lam.

Genetic variation was examined in the Japanese apogamous and sexual forms ofAsplenium unilaterale by electrophoretic analysis of eight enzyme systems. The apogamous form consisted of four biotypes (A, B, C, and D) which show different band patterns. Biotypes C and D were distinguished from biotype A by a difference at only one locus each:Mdh-1 andPgi-2, respectively. Biotypes A, C, and D were sufficiently differentiated from the sexual form (Nei's genetic distance (D)=0.50). This suggests that the apogamous form is not directly derived from the sexual form. However, biotype B expressed the combined band pattern of biotype C and the sexual form, leading to the conclusion that biotype B probably originated by hybridization between those two.     

WILD GAMETOPHYTES OF PHANEROSORUS MAJOR (MATONIACEAE)

The gametophytes of Phanerosorus major collected from wild populations in Seram Island are described. Young gametophytes are slender, ribbon-like and uniformly one cell thick. They develop to old gametophytes with a broad thick cushion and coarsely ruffled wings. The gametophytes are monoecious and protandrous, although archegonia-bearing thalli occasionally become antheridia-bearing. Massive antheridia form on the ventral surface of young and old gametophytes while archegonia are borne on the cushion of old ones. Both young and old gametophytes reproduce vegetatively. The gametophytic characters are compared with those of Matonia. The life cycle of P. major is discussed with reference to its ecology.     

THE INFLUENCE OF CULTURAL CONDITIONS ON THE INDUCTION OF APOGAMY IN PTERIDIUM GAMETOPHYTES

Apogamous sporophytes formed on Pteridium gametophytes in response to concentrations of certain sugars which supported gametophytic growth. High osmotic concentration of the medium inhibited apogamy, while variations in the basic medium were not stimulatory. Agar, autoclaving, the ammonium ion, and dry media were not required for apogamy. Renewing the medium during an experiment enhanced the apogamous response. Changing the medium at set intervals facilitated the separation of apogamous plant development into gametophytic, initiative, and developmental phases, thus enabling testing of various factors at each of these stages. Apogamy was light-initiated, while the actual development of apogamous sporophytes was caused by light, succinic acid or sugar.     

THE INITIATION OF SPOROPHYTES BY OBLIGATE APOGAMY IN CHEILANTHES CASTANEA

The initiation of apogamous sporophytes in Cheilanthes castanea was recorded by daily photography of individual gametophytes. Whereas an ordinary embryo arises from a zygote, apogamous embryos of C. castanea originate from one to three initial cells which occur just behind the apical region of the prothallus. The initial (or initials) produce cells with small chloroplasts behind the sinus of the gametophyte. The appearance of cells with smaller chloroplasts than those normally found in gametophytes is the first indication that apogamy is occurring. The cells with small plastids produce a group of densely-cytoplasmic meristematic cells. The size of the meristematic mass increases until shoot and root apices of the apogamous embryo are organized.     

The effect of cytokinins on growth and sexual organ development in the gametophyte of <Emphasis Type="Italic">Blechnum spicant</Emphasis> L.

In this study, we report the role of exogenous and endogenous cytokinins on growth and sexual organ development in the fern Blechnum spicant L. Spore-derived gametophytes (SG) were cultured in full-strength Murashige and Skoog (1962) liquid medium supplemented with (a) 4.44 μMN⁶-benzyladenine (BAP), (b) a crude extract from mature female gametophytes, and (c) 4.44 μM BAP in combination with the crude extract from mature gametophytes, respectively. Both BAP and the crude extract delayed the gametophyte development, and this effect was increased when they were added together. With respect to sexual organ development, BAP inhibited the sexual organ formation, while the crude extract favored antheridia formation; however, when added together, the percentage of antheridia decreased. The endogenous level of the cytokinins cis-zeatin (cZ), cis-zeatin-riboside (cZR), dihydrozeatin (DHZ), dihydrozeatin riboside (DHZR), isopentenyl adenine (iP), isopentenyl adenosine (iPR), isopentenyl-9-glucoside (iP9G), trans-zeatin (tZ), and trans-zeatin riboside (tZR) were analyzed in female and male gametophytes of B. spicant L. The endogenous levels of cytokinins tZ, cZ, DHZ, cZR, iP, and iPR were higher in female gametophytes than in male gametophytes, with the endogenous iP and iPR content being increased more than 300 and 400 times, respectively.     

Parthenogenesis is rare in the reproduction of a sexual field population of the isogamous brown alga Scytosiphon (Scytosiphonaceae,Ectocarpales)

Parthenogenetic development of unfused gametes is commonly observed in laboratory cultures among various brown algal taxa. There is, however, little information on the contribution of parthenogenesis to the reproduction of field populations. In this study, we investigated whether parthenogenesis is present in a sexual population of the isogamous brown alga Scytosiphon with a 1:1 sex ratio. In culture, both female and male gametes showed higher mortality and slower development compared to zygotes. More than 90% of surviving partheno‐germlings formed parthenosporophytes irrespective of the culture conditions tested. Therefore, if parthenogenesis occurs in the field, most unfused gametes are expected to form parthenosporophytes. Contrary to this expectation, parthenosporophytes were rare in the field population. We collected 126 sporophytic thalli and isolated and cultured a unilocular sporangium from each of them. We confirmed that cultures of 120 unilocular sporangia produced both female and male gametophytes by the observation of zygotes or amplification of PCR‐based sex markers indicating that these sporangia originated from zygotic sporophytes. Only females were detected in cultures from two sporangia and only males from four sporangia suggesting that these sporangia originated from parthenosporophytes. In the Scytosiphon population, although parthenogenesis is observable in culture, our results demonstrate that the contribution of parthenogenesis to reproduction is small (≤4.8%) compared to sexual reproduction. Unfused gametes may not survive to form mature parthenosporophytes in significant numbers in the field partly due to their higher mortality and slower development compared from zygotes.     

The life history of Porphyra endiviifolium from the South Shetland Islands, Antarctica

This paper describes the reproduction and life history of an intertidal species, Porphyra endiviifolium, from Antarctica. Field specimens were examined microscopically, prepared for electron microscopy and used to establish cultures. Wild populations comprised two kinds of leafy thalli, morphologically similar but distinguished by their mode of reproduction, either sexual or asexual. Carpospores from monoecious leafy gametophytes developed into conchocelis filaments in culture, and under “winter-spring” conditions these formed conchospores that germinated to produce leafy thalli. Monospores from asexual leafy thalli developed directly into two different forms of leafy thalli. Only one of the cultured morphotypes became fertile, reproducing asexually by monospores. We conclude that the phases of the life history of P. endiviifolium show different ecological strategies, the conchocelis phase reproducing in response to short days unlike the leafy thalli in which growth and reproduction respond primarily to irradiance. Received: 2 May 1997 / Accepted: 11 October 1997     

Temporal variations of vegetative features, sex ratios and reproductive phenology in a Dictyota dichotoma (Dictyotales, Phaeophyceae) population of Argentina

This paper addresses the phenology of a Dictyota dichotoma population from the North Patagonian coasts of Argentina. The morphology of the individuals was characterized, and analyses of the temporal variations of vegetative features, diploid and haploid life cycle generations and sex ratios are provided. Individuals, represented by growing sporophytes and gametophytes, occurred simultaneously throughout the year. Morphological variables showed temporal variation, except the width and height of medullary cells, which did not vary between seasons. All vegetative variables were significantly correlated with daylength. Besides, frond length, frond dry mass and apical and basal branching angles were significantly correlated with seawater temperatures. Vegetative thalli were less abundant than haploid and diploid thalli. Sporophytes were less abundant than male and female gametophytes. Male gametophytes dominated in May, August, October and January, and female gametophytes were more abundant in September, November, December, February and March. The formation of female gametangia showed a significant correlation with daylength, and the highest number of gametangia was registered in spring. In general, the male/female sex ratio varied between 1:2 and 1:1. Apical regions were more fertile than basal regions. Our data about frequency in the formation of reproductive structures and male/female ratios are the first recorded in the Dictyota genus and thus could not be compared with populations from other regions of the world. Significant morphological variation was observed in thalli of both life cycle generations, regarding length and dry mass, number of primary branches and branching basal angle. In general, all variables analyzed varied seasonally except cortical cell width.     

Marginal distribution and high heterozygosity of asexual Caloglossa vieillardii (Delesseriaceae,Rhodophyta) along the Australian coasts

In animals and land plants, many asexual species originate through inter‐ or intraspecific crosses, and such heterozygous asexuals frequently are more abundant than their sexual relatives in marginal habitats. Although asexual species have been reported in various macroalgal taxa, detailed information regarding their distribution, heterozygosity, and origin is limited. Because many asexual tetrasporophyte strains of Caloglossa vieillardii have been isolated from South Australia, far from their core tropical habitats, we re‐examined the distribution range of asexual C. vieillardii and genotyped these and other western Pacific strains using an actin gene marker. We confirmed the marginal distribution of the asexuals; however, a small patch of sexual thalli was newly discovered 450 km further west from asexual populations in South Australia. Three heterozygous genotypes and one homozygous genotypes were detected from nine asexual populations; 21 heterozygous strains were obligately asexual, but one homozygous strain suddenly produced sexual gametophytes after several years of culture. We hypothesized that the most abundant heterozygous genotype (defined as type 3/4) in asexual populations occurred by a cross between type 3 and type 4 allele gametophytes, both of which were isolated from the Australian coasts. In the crossing experiments, certain combinations between type 3 females and type 4 males produced tetrasporophytes, which recycled successive tetrasporophytes. In the culture experiments, whereas both sexual and asexual strains successfully produced tetraspores at 12°C, no sexual strains released carpospores below 14°C. However, it is uncertain whether this slight difference of maturation temperature was related to the marginal distribution of asexual C. vieillardii.     

Reproductive biology and population dynamics of wild gametophytes ofEquisetum

This is d study of the reproductive biology of wild gametophytes of Equisetum arvense L., E. fluviatile L., and E. palustre L., which form extensive populations during the summer months on previously inundated bare mud along the margins of reservoirs and lakes in Northern England and Wales. The garnetophytes have all the characteristics of pioneer species: they rapidly attain sexual maturity and are adversely affected by competition. Early in the summer mixtures of male and female gametophytes are produced. The former are smaller and their frequency (which is always less) diminishes throughout the season as a result of bryophytic competition. The rarity of bisexual, initially female, gametophytes, in wild populations is related to the absence of metabolite accumulation which mediates this change in rulture. Significantly different sex ratios between populations and species underlines the labile nature of the sex-determining mechanism in Equisetum. The high frequency of females bearing sporophytes indicates that intergarnetophytic fertilization is highly effective in nature. Absence of any correlation between the incidence of fertilized females and the proportions of males, in conjunction with a consideration of the male gamete dispersal distance, suggests that sporophyte formation is restricted by the availability of ripe archegonia. Sexual reproduction in Equisetum is probably limited by the narrow range of conditions under which gametophytes can become established rather than availability of water for fertilization. The majority of females, which bear one sporophyte, are smaller than unfertilized or polyembryonic females. Their small size results from nutrient demands and allelopathic compounds from the sporophytes which also probably prevent the establishment of gametophytes within mature stands of the parent species. Correlations between female diameters and frequencies of males suggest that gametophytes are more likely to produce archegonia under favourable conditions. The natural reproductive biology of Equisetum is in accord with predictions based on an understanding of the mechanism of sex determination in axenic culture. Several striking parallels between sexuality in Equisetum and dioecious angiosperms are revealed. The absence of winter flooding at two of the gametophyte sites led to the establishment of mature rtands of Equisetum, which produced cones after 3–4 years; two hybrids, E. fluviatile×E. arvense and E. fluviatile×E. palustre, were detected.     

Biomass variation and reproductive phenology of Gracilaria gracilis in a Patagonian natural bed (Chubut, Argentina)

The biomass variation and the reproduction of the natural Gracilaria gracilis bed in Bahía Bustamante (Patagonia, Argentina) were analyzed for 2 years, with the aim of determining the present situation of the population for an updated status overview; establishing the relevant features of the temporal variation in both biomass and reproductive states in relation to environmental factors, epiphytes and associated algae; and assessing carpospore availability for future spore-culture development. Field measurements and sampling were performed monthly between March 2006 and February 2008. In both years, G. gracilis biomass presented marked seasonal variations, with a minimum in winter and a maximum in late spring and in summer. During both years, coexistence of the three life-cycle phases was found, with dominance of tetrasporophytes. Two data sets from individuals originated from sexual reproduction (tetraspores and carpospores) and from asexual reproduction by thallus fragmentation were analyzed separately. In the fragmentation fraction, tetrasporophyte frequencies remained higher than those for gametophytes. However, in the spore-originated fraction, a generation ratio close to 0.5 was observed. Female gametophytes bearing cystocarps were always present, with a maximum in summer and autumn. Biological data were related to environmental factors by means of canonical correspondence analysis (CCA). The first year was characterized by higher biomass values of G. gracilis and Undaria pinnatifida, lower epiphytism, larger Gracilaria thalli and greater proportion of mature tetrasporophytes and gametophytes. The second year was characterized by a high proportion of Gracilaria vegetative thalli and high epiphyte density. The best time to obtain spores from cystocarpic thalli would be in summer and early autumn.     

Reproductive phenology and growth rates in two species of Gracilaria from Hawaii

Observations on wild populations of Gracilaria bursapastoris (Gmelin) Silva and G. coronopifolia J. Ag. showed significant differences in gametophyte: tetrasporophyte ratios from the expected 1: 1 ratio. As in many other perennial red algae, the proportion of tetrasporic individuals in a population of these two Gracilaria spp. dominates the combined male and female gametophytc stage. There were significantly more male than female thalli in the G. cornopifolia population whereas the gametophytes of G. bursapastoris occurred in the expected 1: 1 ratio. In addition, there are seasonal changes in the proportions of tetrasporic and gametophytic individuals within the populations. Tetrasporic thalli of G. coronopifolia evinced a biphasic seasonal pattern with high proportions in winter and summer. The tetrasporic phase of G. bursapastoris, on the other hand, showed a low proportion in winter. Maximum biomass does not necessarily correlate with maximum proportion of the tetrasporophyte generation. Seasonal patterns in the proportion of male and female gametophytes differed for each stage as well as for each species. The proportion of male thalli in G. bursapastoris and G. coronopifolia showed high peaks in winter and autumn, respectively. Cystocarpic thalli were most abundant in the former in late winter and summer and in the latter in winter and spring. In both species the female gametophytes grew significantly slower than did the male gametophytic or tetrasporophytic stages. Practical applications regarding seasonal cycles in the various reproductive stages and their differential growth rates are discussed.     

Sexuality and apogamy in the Cuban Asplenium auritum–monodon complex (Aspleniaceae)

Asplenium auritum Sw. is a sexual fern that produces 64 spores per sporangium, while A. monodon Liebm. is an apogamous species that produces 32 spores. The hybrid between them, A. × lellingerianum Sánchez & Regalado, also shows an apogamous life cycle, with mainly abortive spores. The scope of this work was to study the sexual and apogamous behavior in these taxa. We studied spore germination and gametophyte development of the three species, a process that followed the Adiantum type. Afterward, we observed morphological apogamous characters in A. monodon and A. × lellingerianum. Apogamous sporophytes arose from apical and basal regions of gametophytes, lacking feet and roots in the first instance, but developing other normal sporophytic structures, such as tracheids, stomata, glandular hairs, and scales. Finally, we studied the gametangia production in all three taxa, finding that the scarce production of antheridia in A. monodon is indicative of the Braithwaite apogamous life cycle scheme, and this scheme has probably been inherited by A. × lellingerianum.     

Electrophoretic evidence for homoeologous chromosome pairing in the apogamous fern species<Emphasis Type="Italic"> Dryopteris nipponensis</Emphasis> (Dryopteridaceae)

Segregation of genotypes through homoeologous chromosome pairing in the apogamous species Dryopteris nipponensis was tested by electrophoretic analysis. Of 284 progeny examined (250 gametophytes and 34 sporophytes), from the parental sporophyte with the Pgi-2 genotype abc, five showed different genotypes from that of the parent (three aac, one bbc and one bcc). This is the first evidence for genetic segregation in the progeny of apogamous fern species. Electronic Publication     

Electrophoretic variation of the apogamousDryopteris varia group (Dryopteridaceae)

A total of 35 electrophoretic variants were detected in four apogamous species of theDryopteris varia group: 14 variants inD. bissetiana triploids, 7 inD. sacrosancta triploids, 6 inD. varia triploids, 3 inD. pacifica triploids, and 5 in apogamous diploids. The variants differed from each other in various combinations of a few alleles which were common in some of the variants, suggesting recurrent origin.Dryopteris insularis var.insularis (2x) and var.chichisimensis (3x), other members of the group, were electrophoretically monomorphic, suggesting single origin. Banding patterns infer that triploids ofD. bissetiana andD. sacrosancta may have originated from hybrids whose parent is the sexualD. saxifraga, whereas this species was not involved in the origin of the other apogamous triploids and the apogamous diploids. The other parent was also discussed and we proposed a hypothesis that diploid offsprings fromD. pacifica triploids hybridized withD. saxifraga giving rise to triploids ofD. bissetiana andD. sacrosancta.     

Exploring Phenotypic Plasticity in the LichenRamalina capitata: Morphology, Water Relations and Chlorophyll Content in North- and South-facing Populations

The present work analyses the morphology, anatomy, water relationsand chlorophyll content of thalli of the lichenRamalina capitatavar.protectafromtwo different populations exposed to contrasting microclimaticconditions due to differences in the orientation of the rocksurface. The population on the north-facing rock surface (NFS)was exposed to lower photosynthetic photon flux densities (PPFD),remained at high relative humidities for longer periods of timeand was exposed to lower temperatures than the population onthe south-facing surface (SFS). We proposed the hypothesis thatthe shadier the habitat the greater the ecological advantagefor enhanced light harvesting. Thalli from the SFS had shorterand wider lacinia, thicker thalli, mostly due to increased medullathickness, a higher water-retention capacity, a higher percentageof thallus volume occupied by the algal cells and a higher chlorophyllcontent than thalli from NFS. The phenotypic plastic responseof the traits studied inR. capitatavar.protectawas not directlyrelated to differences in the light availability, at least forthe range of PPFD experienced by the two populations studied,since the population exposed to higher PPFD exhibited largeramounts of light harvesting pigments. Both populations exhibitedthe same intrathalline distribution of algal cells and chlorophylls,which were more abundant in the apical than in the basal zonesof all thalli studied. Periods of water-induced metabolic activitywere shorter in the SFS than in the NFS, and structural andchlorophyll data indicated that thalli from the SFS were betterprepared for the photosynthetic exploitation of these brieferperiods and for maintaining thallus hydration into dry periods.These results suggest that differences in selective pressurebetween the two populations ofR. capitatavar.protectastudiedinvolved maximization of the photosynthetic exploitation ofthe periods of metabolic activity when they are brief, as hasbeen described for certain vascular plants from xeric environments. Ramalina capitatavar.protecta; algal cells; chlorophylls; water relations; microclimate; morphology; intrathalline variation; lichen; phenotypic plasticity