Gametophyte contribution to sporophyte growth on the basis of carbon gain in the fern <Emphasis Type="Italic">Thelypteris palustris</Emphasis>: effect of gametophyte organic-matter production on sporophytes

At an early stage of growth gametophytes support the sporophytes of ferns. Young sporophytes become independent of gametophytes when the first leaves develop. Although large fern gametophytes produce multiple archegonia simultaneously, only one sporophyte is typically established on one gametophyte. The number of sporophytes is believed to be controlled in two possible directions, from gametophyte to sporophyte or from preceding sporophyte to another sporophyte. To investigate the effects of gametophytes on their sporophytes, we studied the relationship between organic matter production by gametophytes and the growth of young sporophytes of Thelypteris palustris. We cut gametophytes in half (CGs) to reduce the gametophytes’ production of matter. There was no significant difference between the growth of sporophytes on intact gametophytes (IGs) and that on CGs. According to our estimates, based on the rate of organic matter production, the large gametophyte was able to produce two or more sporophytes. The resources required for CGs to make similar-sized sporophytes was twice that for IGs. In polyembryony each of the multiple sporophytes was similar in size to the single sporophytes. Resource limitation does not seem to explain why fern gametophytes establish single sporophytes.     

Isolation of protoplasts from fern prothallia and their regeneration to gametophytes

Protoplasts were isolated enzymatically from prothallia ofLygodium japonicum. The protoplasts grown in a culture medium containing 0.6 M mannitol and 0.05 M sucrose began to divide within 8 days of culture, and after 30 days 10-cell clusters were present. When the cell-clusters were transferred into fresh media followed by sequential reduction of mannitol concentration, they developed rhizoids and protonemata. The reduction of mannitol concentration to 0.3 M resulted in the regeneration of a common gametophyte within 50 days of culture, and subsequently the regenerated gametophytes produced sporophytic leaves and roots.     

Iridescence: a useful criterion to sort gametophytes from sporophytes in the red algaChondrus crispus

Carrageenan analyses ofChondrus crispus Stackhouse from Brittany, France, showed that the iridescent fronds always were kappa-carrageenan dominant and that the non-iridescent ones were lambda-carrageenan dominant, whatever their sizes and their vegetative or reproductive stages. Evidence is presented that the blue iridescence is independent of both the morphology of the fronds and their ecological situation (shelter, exposure). It is suggested that iridescence might be used to sort rapidly the kappa-carrageenan containingChondrus crispus (gametophytes) from the lambda-carrageenan containing ones (tetrasporophytes).     

Lead tolerance and accumulation in the gametophytes of the fern Athyrium yokoscense

The fern Athyrium yokoscense is known to be highly tolerant to lead toxicity, and is a lead hyperaccumulator that can accumulate over 1,000 g g^–1 of lead in its dry matter. In this work, we examined whether the gametophytic generation of A. yokoscense also resists lead toxicity like the sporophytic generation. Spore germination in A. yokoscense was more tolerant to Pb²⁺, compared to that in other fern species, such as Pteridium aquilinum, Lygodium japonicum and Pteris vittata. In addition, the early gametophyte development of A. yokoscense was not much affected by 10 M Pb²⁺, as evaluated from the prothallial growth and rhizoid development. We also showed that Athyrium gametophytes could accumulate more than 10,000 g g^–1 of lead, and that the lead was localized in the cytosol and vacuole of rhizoidal cells, as determined by a transmission electron micrograph. These results indicate that Athyrium gametophytes have the ability to accumulate lead in the rhizoids. Furthermore, the gametophytes were found to include a large amount of proanthocyanidins (condensed tannins). Because proanthocyanidins have a latent ability to complex with lead ions, the possible roles of proanthocyanidins in the lead tolerance and accumulation of Athyrium gametophytes are discussed.     

Oleosins in the gametophytes of Pinus and Brassica and their phylogenetic relationship with those in the sporophytes of various species

Oleosins, which are structural proteins on the surface of intracellular oil bodies, have been found in the sporophytic seeds of angiosperms. Here, we report an oleosin from the female gametophyte of gymnosperm Pinus ponderosa Laws, seed and another oleosin from the male gametophyte of Brassica napus L. With the pine seed gametophyte, we identified two putative oleosins of 15 and 10 kDa, which are similar to the oleosins in angiosperm seeds in terms of their presence in the oil bodies in massive quantity. The complete sequence of the cDNA encoding the gametophytic 15-kDa oleosin was obtained, and it has a predicted amino-acid sequence similar to those of oleosins in angiosperm sporophytic seeds. A Brassica napus pollen cDNA sequence, which was reported earlier, would encode an amino-acid sequence somewhat similar to those of seed oleosins. We tested if the dissimilarity signifies a substantially different oleosin in the Brassica male gametophyte or an analytic error. By direct sequencing of a polymerase chain reaction (PCR)-amplified fragment of genomic DNA, we obtained evidence showing that this reported dissimilarity is likely to have arisen from a sequencing error. Our predicted sequence of the Brassica pollen oleosin has all the structural characteristics of seed oleosins. A phylogenic tree of 20 oleosins, including those from sporophytic and gametophytic tissues of angiosperm and gymnosperm, was constructed based on their amino-acid sequences. We discuss the evolution of oleosins, and conclude that oleosins are ancient proteins with multiple lineages whose root cannot be determined at this time.Abbreviations PCR polymerase chain reaction - TAG triacylglycerols This work was supported by USDA grant 91-01439 (AHCH). We thank Dr. Mike Lassner of Calgene, Inc., (Davis, Calif., USA) for providing us with the unpublished jojoba oleosin amino acid sequence.     

Introduction of a seedling production method using vegetative gametophytes to the commercial farming of Laminaria in China

Laminaria is the main seaweed species under commercial cultivation in China, which utilizes 80 x 10⁹ artificially cultured seedlings each year. This paper reports the large-scale application of seedling production using vegetative gametophytes in Laminaria cultivation. Key techniques, such as the attachment of gametophytes to the collectors and tank culture of seedlings in the hatchery, are examined. The problems of detachment and irregular dispersion of seedlings were also investigated. An efficient and practical seedling production technology was developed, which consists of seeding gametophytes on collectors, control of seawater flow in tanks, washing of collectors and regulation of ambient light, seawater temperature and nutrition supplement. After 2 months of culture, seedlings grown on woven palm rope collectors 0.5 cm in diameter measured up to the national technical standard of China (i.e., > 20 individuals cm⁻¹ at a length of > 1.5 cm, and > 100 individuals cm⁻¹ at a length of 0.2 cm–1.5 cm, desolation rate < 1% and detachment rate < 5%). Compared with spore-derived seedlings, no significant differences in sporophyte recruitment and dispersion pattern were observed. Large-scale production of seedlings for commercial cultivation of 4,000 ha from 2001 to 2006 was conducted successfully.     

Culture and growth of Lessonia trabeculata (Phaeophyta,Laminariales) juvenile sporophytes in La Herradura de Guayacan Bay,northern Chile

Lessonia trabeculata is one of the major kelps found along the northern coast of Chile. In addition to its ecological and economic importance, L. trabeculata may be severely affected by environmental disturbances such as El Níño, which during 1982–1983 cleared wide areas along the coast of Peru and Chile. The main goal of this work was to mass culture L. trabeculata and to observe the growth of sporophytes obtained in the laboratory and cultured in the sea. Juvenile sporophytes obtained in the laboratory were attached between 1 and 6 m in depth. The linear growth rate, as blade elongation, was recorded weekly for seven months. No significant differences (p < 0.05) were found in sporophyte blade linear growth at different depths. The best elongation growth rate was 7.5 ± 1.6 mm d^–1 at 3 m during March. This preliminary work suggests that L. trabeculata follows an annual growth cycle similar to that of other Laminariales with a high rate of blade elongation during the summer and decreasing towards autumn. This species can be considered a potential candidate for aquaculture to increase the availability of raw material and aid in repopulation of overexploited areas.     

Gene localization on the chloroplast DNA of the maiden hair fern;Adiantum capillus-veneris

Gene maps were constructed for the inverted repeat region and for the adjacent large single copy region of the chloroplast genome of the maiden hair fern,Adiantum capillus-veneris L. Gene order and organization was very different from the typical angiosperm chloroplast genome (e.g. tobacco). Elongation of inverted repeat and a minimum of two inversions must be postulated to account for the unusual genome structure.     

Absence of major differences between soluble proteins from haploid gametophytes and diploid sporophytes in the green alga Ulva mutabilis Føyn

Soluble proteins from haploid gametophytes and diploid sporophytes of the marine green alga Ulva mutabilis Føyn have been reexamined, using polyacrylamide gel electrophoresis and isoelectric focusing. A two-dimensional system resolved about 150 protein spots. In contrast to an earlier report (Hoxmark (1976) Planta 130, 327–332), no major differences could be detected between soluble proteins from the two generation types by any of the methods used.To whom correspondence should be addressed     

Effects of daylength,irradiance and settlement density on the growth and reproduction of <Emphasis Type="Italic">Undaria pinnatifida</Emphasis> gametophytes

The effects of daylength, irradiance and spore settlement density on the growth, maturation and sporophyte production of Undaria pinnatifida (Harvey) Suringar gametophytes were examined using a factorial experimental design in culture. The growth of Undaria gametophytes increased with increasing daylength (8, 12 and 16 h), but the maximum fertility occurred at a daylength of 12 h followed by 8 and 16 h. Gametophytes grew better at the 16 h daylength under the same mean daily irradiance (MDI) of 20 μmol photons m⁻² s⁻¹. However, the fertility was higher at the short daylength (8 h), indicating that the maturation of U. pinnatifida gametophytes is influenced by daylength rather than by the MDI. Vegetative growth and sporophyte production of gametophytes were better at 60 μmol photons m⁻² s⁻¹ than at 30 μmol photons m⁻² s⁻¹ under a 8:16 h LD (Light: Dark) cycle, and their growth and maturation were density-dependant in 16 and 12 h daylength, respectively. These results suggest that the U. pinnatifida gametophytes require a certain amount of light for the growth and reproduction, and intraspecific competition occurred under the optimal growth and maturation conditions. However, the sporophyte recruits per unit has been enhanced with increasing spore settlement density at 8 and 12 h daylengths indicating that high settlement density gives a benefit for maintaining population, even though the sporophyte production of each female plant is inhibited. In conclusion, the vegetative growth, reproduction and sporophyte production of U. pinnatifida gametophytes are retarded at a low irradiance above growth saturation and a high settlement density, and are determined by daylength.     

Effects of carbon concentration and carbon to nitrogen ratio on the growth and sporulation of several biocontrol fungi

Effects of carbon concentration and carbon to nitrogen (C:N) ratio on six biocontrol fungal strains are reported in this paper. All fungal strains had extensive growth on the media supplemented with 6–12 g l⁻¹ carbon and C:N ratios from 10:1 to 80:1, and differed in nutrient requirements for sporulation. Except for the two strains of Paecilomyces lilacinus, all selected fungi attained the highest spore yields at a C:N ratio of 160:1 when the carbon concentration was 12 g l⁻¹ for Metarhizium anisopliae SQZ-1-21, 6 g l⁻¹ for M. anisopliae RS-4-1 and Trichoderma viride TV-1, and 8 g l⁻¹ for Lecanicillium lecanii CA-1-G. The optimal conditions for P. lilacinus sporulation were 8 g l⁻¹ carbon with a C:N ratio of 10:1 for M-14 and 12 g l⁻¹ carbon with a C:N ratio of 20:1 for IPC-P, respectively. The results indicated that the influence of carbon concentration and C:N ratio on fungal growth and sporulation is strain dependent; therefore, consideration for the complexity of nutrient requirements is essential for improving yields of fungal biocontrol agents.     

The effect of environmental factors on floating fresh marsh end-of-season biomass

We used both stepwise and quantile regression to determine the sources of environmental variation that explained the observed inter-annual variation in end-of-season freshwater floating marsh aboveground biomass over an 18-year period. The vegetation at our study site had high species diversity with an average of 20 species recorded from 10 0.25 m⁻² plots. However, Panicum hemitomon was clearly the dominant contributing 74% of the total biomass. Only three other species (Solidago sempervirens, Vigna luteola, and Thelypteris palustris) were so common that they were sampled in all years. We expected that the most important factors controlling interannual variation in aboveground biomass are temperature and nitrogen availability. We also expected that nitrogen availability to the plants is affected by water movement through and under the mat driven by precipitation (lower N), evaporation (transportation of higher N waters to roots), and local runoff (higher N). Stepwise regression analysis indicated that P. hemitomon average biomass was negatively related to average water level and positively related to maximum water level and had a curvilinear response with TKN. Using quantile regression the best fit for P. hemitomon maximum-biomass with two parameters was obtained using hot days (positive relationship) and maximum water level (negative relationship). Both analytical methods showed maximum water level (negative relationship) and cold front passage (positive relationship) to be the environmental parameters that best explained interannual variation in S. sempervirens biomass. V. luteola biomass was positively related to temperature. Stepwise regression added chloride concentration as an additional positive parameter explaining V. luteola biomass, while quantile regression identified nitrogen as an important positive parameter. Both analytical methods identified pH, TKN, and water level as environmental parameters that were negatively correlated with T. palustris biomass. The overall negative effect of water level on all species was unexpected in this floating mat system. We initially assumed that higher water levels were due to higher runoff which should have a positive effect on biomass. However, higher water levels may also be related to a higher retention time in this fresh-water tidal system, which decreases water exchange and nutrient replenishment.     

Growth and survival rates of large-type sporophytes of Ecklonia cava transplanted to a growth environment with small-type sporophytes

Stipe lengths of sporophytes of Ecklonia cava Kjellman have been reported to be longer along the southeast than southwest coast of the Izu Peninsula, central Japan. Two bays in this region that have natural populations of E. cava, but with different stipe lengths, were chosen for transplant experiments to examine if stipe length was an environmentally controlled trait. Transplant experiments were carried out in order to determine whether large-type sporophytes of E. cava with long stipes growing in Nabeta Bay (southeast Izu Peninsula, Japan) would turn into small-type sporophytes with short stipes when transplanted to Nakagi Bay (southwest Izu Peninsula). Ten juvenile sporophytes of E. cava (stipe length < 5 cm) were collected from Nabeta Bay (large-type habitat) and transplanted to Nakagi Bay (short-type habitat) in December 1995. As a transplant control, ten juvenile sporophytes of E. cava growing in Nakagi Bay were also transplanted to the same artificial reefs. Growth and survival rates of the sporophytes were monitored monthly for 3 y until December 1998. The transplanted sporophytes showed an increase in their stipe length and diameter from winter to spring, whereas almost no increase was observed from summer to autumn. However, the elongation was greater in Nabeta sporophytes than in Nakagi sporophytes. The primary blade length increased mainly from winter to early spring and decreased largely in autumn. Average primary blade lengths were similar in both Nabeta and Nakagi sporophytes from the end of the first year of transplanting. Although ca. 70% of both Nabeta and Nakagi sporophytes survived during the first 2 y after transplantation, no Nakagi sporophytes and only two Nabeta sporophytes survived to the end of the 3 y study period. Despite transplantation to Nakagi Bay, where short sitpes are naturally present, the sporophytes from Nabeta Bay persisted in having longer stipes, which suggests that stipe length is genetically, rather than environmentally, controlled.     

红香芋试管球茎膨大过程中主要碳水化合物含量以及淀粉合成相关酶活性的动态研究

为了探讨芋(Colocasia esculenta(L.)Schott)试管球茎膨大期间糖类物质积累特点,以红香芋无菌试管苗为材料,研究了高浓度蔗糖诱导条件下,红香芋试管球茎形成及膨大过程中主要碳水化合物的变化规律,以及与相关酶活性的关系。结果表明:(1)在红香芋试管球茎膨大过程中,果糖、葡萄糖和总可溶性糖含量均呈先升高后降低的变化趋势,果糖含量在诱导至第27天时达到最大值,而总可溶性糖和葡萄糖含量均在第34天达到峰值;蔗糖含量呈现先上升、后下降、再上升的变化趋势,在培养第48天时积累量达到最大值。(2)红香芋试管球茎总淀粉含量、直链和支链淀粉含量均随培养时间的延长而增加,至膨大后期总淀粉含量达到最大值,淀粉总含量约占干重的76%,并以支链淀粉含量为主。(3)解剖学观察发现,随着试管球茎的形成与膨大,贮藏组织中淀粉粒密度不断增大,至球茎膨大后期,淀粉粒布满薄壁细胞,并且处于比较稳定的水平。(4)诱导培养至第41天时,试管球茎的ADPG焦磷酸化酶和Q-酶活性均达到最大值,分别为1.22和2.39μmol·g~(-1)·min~(-1)。相关性分析发现,从茎基部开始膨大(20d)至ADPG焦磷酸化酶和Q-酶活性达峰值(41d)时,ADPG焦磷酸化酶活性与总淀粉含量、Q-酶活性与支链淀粉含量的相关系数分别为0.819和0.738,二者均呈极显著正相关。研究认为,淀粉的积累以及可溶性糖类含量的变化与红香芋试管球茎的膨大发育密切相关,并受到相关酶的调控。     

Mimosine produced by the tree-legume Leucaena provides growth advantages to some Rhizobium strains that utilize it as a source of carbon and nitrogen

Growth of most Rhizobium strains is inhibited by mimosine, a toxin found in large quantities in the seeds, foliage and roots of plants of the genera Leucaena and Mimosa. Some Leucaena-nodulating strains of Rhizobium can degrade mimosine (Mid⁺) and are less inhibited by mimosine in the growth medium than the mimosine-nondegrading (Mid^-) strains. Ten Mid⁺ strains were identified that did not degrade 3-hydroxy-4-pyridone (HP), a toxic intermediate of mimosine degradation. However, mimosine was completely degraded by these strains and HP was not accumulated in the cells when these strains were grown in a medium containing mimosine as the sole source of carbon and nitrogen. The mimosine-degrading ability of rhizobia is not essential for nodulation of Leucaena species, but it provides growth advantages to Rhizobium strains that can utilize mimosine, and it suppresses the growth of other strains that are sensitive to this toxin.     

Effect of pre-incubation irradiance on survival of cryopreserved gametophytes of Undaria pinnatifida (Phaeophyta) and morphology of sporophytes formed from the gametophytes

Gametophyte strains originating from indigenous sporophytes of Undaria pinnatifida (Harvey) Suringar in Iwate Prefecture, Northeast Japan, were maintained for 9–10 months at 45 μmol photons m⁻² s⁻¹. Before cryopreservation in liquid nitrogen for more than 12 h (1–14 days) using a two-step cooling method with a mixture of cryoprotectants (10% l-proline and 10% glycerol), these were pre-incubated for 2, 4 and 8 months at 15 μmol photons m⁻² s⁻¹. After 1 week of thawing, no surviving gametophytes were detected in the strains without pre-incubation, but both the female and male gametophytes, pre-incubated for more than 4 months, showed high survival rates (43–60% for females and 64–100% for males). This revealed the induction of freezing tolerance by incubation at low irradiance. Thereafter, sporophytes derived from cryopreserved gametophytes and subcultured gametophytes, stored under pre-incubation conditions, were formed from the strain, and a morphological comparison was conducted with 10 characters (stipe length, stipe wet weight, blade length, blade wet weight, blade width, incision depth, blade thickness, sporophyll length, sporophyll wet weight, and sporophyll width). The morphology of the sporophytes formed from the cryopreserved gametophytes corresponded well with that of the subcultured gametophytes from the same strain. The results suggest that the cryopreservation method is applicable for preserving culture stocks of U. pinnatifida to be used in mariculture.     

Estimation of the effect of photoinhibition on the carbon gain in leaves of a willow canopy

The occurrence of photoinhibition of photosynthesis in leaves of a willow canopy was examined by measuring the chlorophyll-a fluorescence ratio of F _V/F _M (F_M is the maximum fluorescence level of the induction curve, and F_V is the variable fluorescence, F _V=F _M–F ₀, where F₀ is the minimal fluorescence). The majority of the leaves situated on the upper parts of peripheral shoots showed an afternoon inhibition of this ratio on clear days. This was the consequence of both a decrease in F _M and a rise in F _O. In the same leaves the diurnal variation in intercepted photosynthetic photon flux density (PPFD) was monitored using leaf-mounted sensors. Using the multivariate method, partial least squares in latent variables, it is shown that the dose of PPFD, integrated and linearly weighted over the last 6-h period, best predicts photoinhibition. Photoinhibition occurred even among leaves that did not intercept PPFDs above 1000 mol·m^–2·s^–1. Exposure of leaves to a standard photoinhibitory treatment demonstrated that the depression in the F _V/F _M ratio was paralleled by an equal depression in the maximal quantum yield of CO₂ uptake and a nearly equal depression in the rate of bending (convexity) of the light-response curve of CO₂ uptake. As a result, the rate of net photosynthesis is depressed over the whole natural range of PPFD. By simulating the daily course in the rate of net photosynthesis, it is estimated that in the order of one-tenth of the potential carbon gain of peripheral willow shoots is lost on clear days as a result of photoinhibition. This applies to conditions of optimal temperatures. Photoinhibition is even more pronounced at air temperatures below 23° C, as judged from measurements of the F_V/F_M ratio on clear days: the afternoon inhibition of this ratio increased in a curvilinear manner from 15% to 25% with a temperature decrease from 23° to 14° C.Abbreviations and Symbols F_O minimum fluorescence - F_V variable fluorescence - F_M maximum fluorescence - PLS partial least squares in latent variables - PPFD photosynthetic photon flux density - VPD water vapour-pressure deficit This study was supported by the Swedish Natural Science Research Council. We are indebted to Dr. Jerry Leverenz (Department of Plant Physiology, University of Umeå, Sweden) for guidance with the modelling of the photosynthesis data.     

The effects of light on sex determination in gametophytes of the fern <Emphasis Type="Italic">Ceratopteris richardii</Emphasis>

The sexuality of homosporous fern gametophytes is usually determined by antheridiogen, a pheromone that promotes maleness. In this work the effect of photomorphogenically active light on antheridiogen-induced male development was examined for gametophytes of Ceratopteris richardii. Although blue light did not affect sensitivity to Ceratopteris antheridiogen (A_Ce) in wild-type gametophytes, it was found that the gametophytes of the her1 mutant, which are insensitive to A_Ce, developed into males when grown under blue light in the presence of A_Ce. Thus, we conclude that another A_Ce-signal transduction pathway activated by blue light exists latently in the gametophytes of C. richardii. Red light, on the other hand, suppressed male development. Because simultaneous red and blue light-irradiation did not promote male development in the her1 gametophytes, the action of red light seems to dominate that of blue light. The results of experiments with a photomorphogenic mutant also suggested that phytochrome may be involved in the action of red light.     

Contribution to the orchid flora of South Central Africa

11 new taxa (8 new species, 3 new varieties) ofOrchidaceae are described from South Central Africa, including data on their habitat and distribution and discussions of their systematic affinities.