EFFECT OF BUFFERED SOLUTIONS AND SULFATE ON VEGETATIVE AND SEXUAL DEVELOPMENT IN GAMETOPHYTES OF PTERIDIUM AQUILINUM

Experiments were performed to determine effects of buffered solutions (0.01 m ) on vegetative development and fertilization of gametophytes of Pteridium aquilinum. Buffered solutions were used to simulate exposures to acidic precipitation up to 3.5 h. Flagellar movement of sperm was reduced at buffer pH levels below 5.8. Specifically, longevity of motility was reduced so that no movements were observed 8–10 and 5–7 min after exposure to pH 5.6 and 5.2, respectively. Addition of sulfate (86 μm ) depressed the percent motile sperm by 50%, 2–4 min after exposure at all pH values tested. Longevity of flagellar movement was most affected by pH and additions of sulfate. The effect on fertilization was assayed directly by determinations of sporophyte production. Although gametophyte survival and development were not affected by solutions of pH 5.8 to 2.2, fertilization was reduced after exposure to buffers below pH 4.2. Sporophyte production was not altered at pH 5.2 but was reduced 50% at pH levels of 4.2 and 3.2 compared with exposures to pH 5.8. Addition of sulfate (86 μm ) decreased fertilization at least 50% at all pH levels observed. The results suggest that only limited fertilization, and therefore limited genetic recombinations could occur in Pteridium aquilinum under conditions of acidic precipitation (pH and sulfate levels) that prevail in the northeastern United States.     

蕨受精作用的细胞学观察

为了阐明进化蕨类受精作用的特点和细胞学机制,该文采用透射电镜观察了蕨(Pteridium aquilinum var.latiusculum)受精作用的主要过程,观察结果显示:(1)蕨精子通过受精孔进入卵细胞,多数情况下,该精子的螺旋运动先在受精孔的下方产生一个受精腔,然后精子再与卵细胞质融合。(2)第一个精子的这种延迟的螺旋运动和因精子的钻入而引起的卵细胞固缩反应可能是阻止多精受精的重要因素。(3)卵发育时期产生的核外突在受精后仍能持续12 h,然后与核本体分离,逐渐在细胞质中消解。(4)合子通过其后方细胞质的液泡化而建立了水平极性,此后再进行细胞分裂。该研究观察到了进化蕨类受精作用过程中的一些新现象,包括产生受精腔、卵细胞固缩反应、核外突的命运以及合子极性建立等,这有助于理解蕨类植物的受精作用机制及有性生殖的演化。     

ACTINOMYCIN D-INDUCED CHANGES IN GROWTH AND RIBONUCLEIC ACID METABOLISM IN THE GAMETOPHYTES OF BRACKEN FERN

Actinomycin D affected the morphological type of growth in the gametophytes of Pteridium aquilinum and the distribution of RNA and protein in their particulate fractions. Increasing concentrations of the drug progressively inhibited two-dimensional growth at the end of a period during which controls had formed typical two-dimensional plants. RNA was lost maximally from the nuclei-rich and ribosome-rich fractions of plants growing in a concentration of actinomycin D which inhibited two-dimensional growth. The magnitude of changes in protein content of the plants was less striking. Presence of actinomycin D in the medium also suppressed incorporation of uridine-H³ into cytoplasmic fractions of gametophytes. The possibility that two-dimensional growth in the gametophytes is under control of a newly synthesized messenger RNA, which is sensitive to actinomycin D, is discussed.     

Growth and development of fern gametophytes in an airlift fermenter

Spores of the fernsPteridium aquilinum andAnemia phyllitidis were grown in an airlift fermenter and subsequent growth and development of gametophytes was monitored. Both species produced greater biomass than that generated in any other solid- or liquid-based culture system tested.Pteridium generated more tissue thanAnemia in every system. The morphology of airlift-grown gametophytes was similar to that of soil-grown plants; fewer gametophytes with perturbed development were observed in airlift cultures than in the other liquid-based systems. No attempt was made to optimise airlift conditions for the species and tissue employed, so it is concluded that airlift cultivation is a promising system for the bulk production of fern gametophytic tissue.     

RNA and protein metabolism in the particulate fractions of the gametophytes of bracken fern during growth in red and blue light

Summary The metabolism of RNA and protein in the gametophytes of bracken fern (Pteridium aquilinum) is affected by the quality of light in which they are grown. When sporelings were grown as two-dimensional gametophytes in blue light, particulate fractions separated from the sporelings exhibited greater incorporation of uridine-³H and leucine-³H into RNA and protein, respectively, than those from sporelings grown as one-dimensional protonema in red light. After various periods of exposure of gametophytes to red or blue light in the presence of uridine-³H, the nuclei-rich fraction showed the highest specific activity in RNA, and irrespective of incubation time, blue light was more effective than red light. The possibility that enhanced synthesis of RNA in the nucleus in response to blue light is significantly related to the morphological growth pattern of the gametophytes, is discussed.     

The role of ethylene in the induction of apogamous buds in Pteridium gametophytes

Summary The level of induced apogamy in gametophytic colonies of Pteridium aquilinum (L.) Kuhn is altered by varying the number of colonies per culture vessel or by including a constant number of colonies in culture vessels of different volumes. In either case, placing vials of mercuric perchlorate, an absorber of ethylene, within the closed culture vessels reduced the apogamous response to a very low level. Production of ethylene by the gametophytes was demonstrated by gas chromatography. Ethylene supplied in a continuous-flow system promoted the apogamous response above that of an air control.     

THE TRANSITION FROM FILAMENTOUS TO TWO-DIMENSIONAL GROWTH IN FERN GAMETOPHYTES. I. THE REQUIREMENT FOR PROTEIN SYNTHESIS IN GAMETOPHYTES OF PTERIDIUM AQUILINUM

Two-dimensional (2-D) development of gametophytes of Pteridium aquilinum was estimated by the percentage of plants with at least one cell wall at an oblique angle, relative to the long axis. Inhibition of cell division per se was assayed by counting the cell number per filamentous (1-D) gametophyte. Specific concentrations of 8-azaguanine, 5-fluorouracil, p-fluorophenylalanine, actinomycin D, streptomycin, and cycloheximide were found to selectively inhibit 2-D development. A reduction in the percentage of 2-D gametophytes was accompanied by a reduction in the relative protein content (protein per dry weight). A comparable association was found only during the initial stages of 2-D development; after 5 days the relative protein content decreased as the percentage of 2-D gametophytes increased. Different intensities of white light resulted in a 10-fold difference in the percentage of 2-D plants with no alteration of the relative protein content. These results demonstrate that no strict relationship between 2-D development and relative protein content occurred in these gametophytes.     

FERTILIZATION SUCCESS CAN DRIVE PATTERNS OF PHASE DOMINANCE IN COMPLEX LIFE HISTORIES1

Many algal life cycles alternate between two free‐living generations. Life histories in which the two generations look identical (isomorphic) are common, particularly in the Rhodophyta. Reports of natural populations dominated by one generation of the life history have sought explanation in terms of phase‐specific differences in mortality and reproductive output, yet in many cases identification of these adaptations has been elusive or inconsistent with predictions. We hypothesized that the gametophyte‐to‐sporophyte ratio of ecologically equivalent isomorphs could result from variation in fertilization rate. We developed two models to test this hypothesis: one representing a generalized isomorphic life history and the other specific to red algae with a Polysiphonia‐type life history. Fertilization rate affected the gametophyte‐to‐sporophyte ratio, especially at low fertilization rates. In the general model, gametophytes dominated the population regardless of fertilization rate unless egg production greatly exceeded meiospore production. In the red algal model, phase dominance depended on the combination of fertilization rate and the number of carpospores produced per fertilization. The generational composition of model multiphasic algal populations results from their inherent reproductive characteristics and the dynamic environment to which fertilization and mortality rates are tied.     

Do motile spermatozoids limit the effectiveness of sexual reproduction in bryophytes? Not in the liverwort Marchantia polymorpha

To test the premise that fertilization distances in bryophytes are limited by spermatozoid motility, we measured the distances between over 80 000 male and female thalli of Marchantia polymorpha on heathland over 2 successive years after a major fire and recorded the incidence of sporophytes. The first comparative data on spermatozoid numbers per antheridium across bryophytes and pteridophytes were calculated from antheridial sizes and spermatid diameters and cross‐checked with the likely numbers produced by successive antheridial mitoses. Individual antheridia of Marchantia produce over 200 000 spermatozoids. Extrapolating from individual antheridia to the numbers of mature antheridia per antheridiophore and then to individual male plants, we calculated that a single flooding event on a male thallus with 10–12 antheridiophores will result in the release of over 50 million spermatozoids. Assuming radial dispersal in surface water films by lipids released from the dehiscing antheridia, spermatozoids can reach distances exceeding 20 m from the parent plants, in line with our finding of 100% fertilization in female plants of Marchantia up to 19 m from the nearest males, far beyond published fertilization distances in dioicous bryophytes. We attribute this to (i) much greater spermatozoid production and numbers in Marchantia than in other bryophytes and (ii) highly effective sperm transport both within the antheridiophores and archegoniophores via their overlapping scales and grooves in the stalks and between male and female thalli via surface water films. These features, coupled with the massive production of small spores, explain the success of Marchantia as the primary colonist of open habitats.     

Effects of canopy gaps on the demography of the subcanopy tree Styrax obassia

Abstract. To evaluate the gap dependency of the subcanopy tree species Styrax obassia, we estimated the demographic parameters of this species in a permanent plot in the Ogawa Forest Reserve, Central Japan, a temperate deciduous forest. From the data collected over eight years, we constructed four transition matrix models of Styrax: a whole population; a shaded subpopulation; a gap-site subpopulation; a compound population of shaded and gap-site subpopulations. The whole-population model suggested that the population structure of Styrax is stable in its present condition. The asymptotic population growth rate of the shaded subpopulation was larger than 1, suggesting that this species can maintain its population even under shaded conditions. The elasticity analyses showed the relative importance of survival of adult individuals that stay mostly under a closed canopy. Since the subpopulation in gaps is very small in a mature forest, the contributions of high seedling survival and high fecundity of adult trees in gaps to the whole population growth were smaller than the survival of adult individuals in the shade. Although we showed that this species has the potential to increase its population in larger disturbances, it actually dominates in the study site where the disturbance scale is relatively small. Styrax should be classified as a typical shade-tolerant species, and it also takes good advantage of small scale disturbance. It can sufficiently maintain its population in a mature forest with small-scale disturbances and has the adaptive characteristics of species that live under a closed canopy.     

THE LIFE HISTORY OF CALLITHAMNION BYSSOIDES IN CULTURE

The life history of Callithamnion byssoides Arnott ex Harv. in Hook. has been shown to comprise a regular sequence of gametophytic, carposporophytic, and tetrasporophytic phases in unialgal culture using supplemented seawater media. Tetraspore germlings gave rise to gametophytes bearing either antheridia or carpogonia in 13 days. Fertilization, carposporophyte development, and carpospore release took place within 5 days. Carpospore germlings produced mature tetrasporophytes in 13 days. The life history thus required approximately 1 month for completion. No deviations from this pattern were observed.     

MORPHOLOGY AND REPRODUCTION OF THE RED ALGA ACROCHAETIUM PECTINATUM IN CULTURE12

The life history of the marine red alga Acrochaetium pectinatum (Kylin) Hamel was studied in unialgal culture using supplemented natural seawater media. The tetrasporophytes are larger than the gametophytes, have a compact filamentous basal system, and produce monosporangia and tetrasporangia. Mono-spores give rise to tetrasporophytes. Tetraspores develop into small gametophytes with unicellular bases. The gametophytes are heterothallic when small (usually less than 500 μ) but as some females become larger (2-3 mm) they produce spermatangia as well as carpogonia. Gametophytes may bear mono-sporangia in addition to carpogonia or spermatangia. These monospores give rise to gametophytes. Fertilization of the carpogonia has not been observed. The tetrasporophytes produce only monosporangia in day-lengths of 12-16 hr, but both tetrasporangia and monosporangia are formed in daylengths of 6–10 hr. Tetrasporangial production is reduced at 15 C compared with 10 C. Light intensity in the range of 5-200 ft-c (cool white fluorescent lighting) has no apparent influence on induction of tetrasporangia. Induction of tetrasporangia is not a photoperiodic response because their development is not inhibited by a brief light break in the middle of the dark period in short daylengths. Plastid morphology, origin, and frequency of sporangia and vegetative branching are variable during the ontogeny, and consequently are somewhat unreliable as taxonomic criteria. Differences in basal systems between gametophytes and tetrasporophytes also indicate that this feature, which is used to distinguish major subgeneric groupings in Acrochaetium, may not be as useful as previously thought.     

GAMETOPHYTES AND YOUNG SPOROPHYTES OF OPHIOGLOSSUM CROTALOPHOROIDES WALT.

The lens-shaped, dorsiventral gametophytes of Ophioglossum crotalophoroides are superficially different from the cylindrical gametophytes of other species of Ophioglossum. However, they have the same features as other Ophioglossum gametophytes (fundamentally axial organization, radially symmetrical apical meristem, radial distribution of gametangia) except that the cylindrical axis is reduced in length. Young sporophyte development is unique in the genus: all primary organs clearly arise from the embryo and develop simultaneously. The length of the life cycle of O. crotalophoroicles is considerably shorter than that of some other species in the Ophioglossaceae. A timetable for gametophyte and young sporophyte development is postulated. Spores germinate soon after they are released in the spring, and mature gametophytes develop by the next growing season. Fertilization occurs approximately one year after spore dispersal, and after two years, the photosynthetic first leaf of the young sporophyte emerges.     

Effect of Heavy Metals （Cd, Cu） on the Gametophytes of Laminaria japonica Aresch

Effects of various concentrations of two heavy metals, namely Cd and Cu, on gametophytes of Laminariajaponica Aresch were determined by recording morphological changes of gametophytes, determining pH values and the heavy metal content of the culture solution, calculating the germination rate of sporophytes, and observing heavy metal （Cd） distribution using a fluorescence microscope. The results showed that heavy metals damaged the gametophytes, and were even lethal, and that the higher the concentration of heavy metal ions, the greater the injury to gametophytes. Gametophytes could not survive in culture solutions containing more than 100 mg/L Cd and 50 mg/L Cu and were only able to survive in culture solution containing a mixture of Cd and Cu up to a concentration of 10 mg/L, which indicates that gametophytes have a higher tolerance to Cd than Cu and that multiple heavy metal ions in solution markedly aggravate the damage to gametophytes compared with individual heavy metal ions. With increases in the concentration of the heavy metal, the burgeoning rate of sporophytes decreased acutely, and solutions containing multiple heavy metal ions caused even more marked harm to sporophytes than solutions containing a single heavy metal ion, because most sporophytes died in mixed solutions. The pH value of the culture medium dropped immediately at the beginning （the first day） of treatment, increased over the following days, and then decreased again. The pH of culture media containing multiple heavy metal ions showed greater variation than media containing a single heavy metal ion, with the extent of the decrease in pH of culture media containing multiple ions being greatest during the last period of the experiment. With increases in the concentration of heavy metals, the capacity of gametophytes to accumulate these ions increased. The blue fluorescent light emitted by the Cd-and Cd-binding protein complex existing in gametophytes in media containing different concentrations of Cd showed clearly the distribution of the ion in gametophytes and the results obtained were consistent with distribution determined using other methods. All results of the present study showed that gametophytes of L. japonica play a remarkable role as heavy metal decontaminators, especially with regard to Cd.     

The synthesis and distribution of ribonucleic acid in developing archegonia of Pteridium aquilinum

Summay Feeding gametophytes of Pteridium aquilinum with tritiated uridine, followed by autoradiography, revealed two waves of incorporation of the nucleoside into nuclei during oognesis. The first, affecting the nucleus of the primary cell of the archegonium, is interpreted as indicating the activation of the genes initiating oogenesis. The second, seen in the nucleus of the maturing egg, is believed to be concerned with the differentiation of the egg itself.The results of the autoradiography, corroborated by enzyme digestion and fluorescence microscopy, also demonstrated a high concentration of ribonucleic acid towards the periphery of the mature egg, some possibly located in nucleolus-like bodies found only in this region of the cytoplasm. It is suggested that part of this cytoplasmic ribonucleic acid may represent genetic information remaining untranslated until after fertilization.No evidence was found of any asymmetry in the distribution of the ribonucleic acid in mature eggs which might account for the polarity of developing zygotes.     

DIFFERENTIAL RESPONSES OF PLANT FOLIAGE TO SIMULATED ACID RAIN

Experiments were performed to show the responses of foliage of several clones of Tradescantia sp., Pteridium aquilinum, Quercus palustris, and Glycine max to simulated acid rain. These experiments were performed to (a) predict the relative sensitivities of foliage of these plants to acid rain, and (b) identify leaf surface and anatomical alterations to simulated acid rain that may be used to diagnose acid rain injury. Plants were exposed to simulated rain at pH levels of 5.7, 3.4, 3.1, 2.9, 2.7, 2.5, and 2.3. Sporophyte leaves of bracken fern {P. aquilinum) were most sensitive to simulated acid rain among the species tested. About 10% of the surface area of older leaves of P. aquilinum was injured after exposure to 10 rainfalls at pH 2.5 (a single 20-min rainfall daily). Foliage of pin oak (Q. palustris) exhibited less than one-percent leaf area injury after exposure to simulated rain at pH 2.5 after 10 rainfalls (one 20-min rainfall daily). The responses of soybeans (G. max) and spiderwort (Tradescantia sp.) were intermediate between these two extremes. Histological observations show that lesion development results in collapsed leaf tissue in all four species. Gall formation that resulted from both cell hypertrophy and hyperplasia occurred in lesions of Tradescantia, and Q. palustris. Limited hyperplastic and hypertrophic reactions occurred in G. max foliage after exposure to simulated acid rain but no leaf galls resulted. Sporophyte foliage of P. aquilinum exhibited a “granular” type abnormality in the cytoplasm prior to the collapse of some epidermal cells injured by simulated rain. Injury occurred most frequently near vascular tissues and trichomes in all four species. In general, plant species that show cell hyperplasia and hypertrophy of leaf tissues after exposure to simulated acid rain are injured less than species that do not show these responses.     

Ionic factors affecting motility,respiration and fertilization rate of the sperm of the bivalvePecten maximus (L.)

Fertilization of the scallopPecten maximus occurs after gametes were naturally released in sea water by the bivalve which has undergone stimulation. The motility of the spermatozoa requires their dilution in sea water (1/40). Dilution triggers an immediate increase of oxygen consumption by sperm, reflecting an activation of a cyanide-sensitive respiration of a cellular origin. When scallops were stimulated by thermal shocks or by serotonin injection, sperm sampled at the urogenital pore output duct shows a respiration-motility activation after sea water dilution which is not seen in sperm scarified from the gonad. Dilution of kidney-sampled sperm into acidic (pH 5) or Na⁺-free artificial sea water reversibly inhibits both respiration and motility. In all cases fertilization rate of sperm is correlated to the increase of respiratory rate and motility measured after dilution in different media. Whether the scallop was stimulated or not, the pH of haemolymph and pericardic fluids were one pH unit below the value of sea water, the pH of the gonad and of the kidney tissues being more acidic (6.5 in average). Our results suggest that the acidic pH of the genital tract maintains the spermatozoa in a quiescent state and that capacitation occurs when male gametes move from the gonad to the kidney from where it is naturally released.Abbreviations ASW artificial sea water - SW sea water - TRIS trishydroxymethyl-aminomethane     

THE DEVELOPMENT AND REPRODUCTIVE MORPHOLOGY OF HALOSACCION AMERICANUM I. K. LEE (RHODOPHYTA,PALMARIALES)1

Halosaccion americanum, a member of Palmariaceae, was grown in culture from spores and the life history was critically examined by the use of scanning EM and light microscopy. A mature tetrasporangium of H. americanum produces four spores that germinate to form two male and two female gametophytes. The male gametophytes grow to maturity in approximately eight months and macroscopically resemble the tetrasporophyte. Following the first division of the tetraspore, the two-celled female gametophyte consists of a vegetative cell and a carpogonium with trichogynes. Fertilization is accomplished by spermatia from male plants of the preceding generation, as male plants of the same season are immature. Spermatia are formed in a continuous layer over the surface of the mature male gametophytes and, when released, are entrained in long strands of mucous. Spermatia adhere to and fuse with trichogynes and, nuclear fusions presumably follow. The carposporophyte is absent; the new tetrasporophyte develops directly from the fertilized carpogonium. Growth of the sporophyte eventually obliterates the female gametophyte, and development into a mature tetrasporophyte proceeds over a period of approximately eight months. The development of H. americanum, with its extremely abbreviated female gametophyte stage and direct development of the tetrasporophyte from the zygote, indicates that this rhodophyte has the same life history as reported for other members of the Palmariales.     

Gametogenesis and chromosome number in Postelsia palmaeformis (Laminariales, Phaeophyceae)

Gametophytes of the ‘sea palm’, the kelp Postelsia palmaeformis Ruprecht, produced gametes whether or not chelated iron was supplied in the culture medium, in contrast to the inhibition of gametogenesis seen with the absence of iron in many other kelps. As gametogenesis proceeded, every cell of the gametophytes was converted into a gamete so that the gametophytes did not continue to grow vegetatively. The portion of the life history from spore release through germination, gametophyte growth, gametogenesis, fertilization and growth of the young sporophyte was completed in 9–10 days under laboratory conditions. Chromosome counts showed that sporophytes had a diploid number of 26–34 chromosomes while sporangia and gametophytes had a haploid number of 14–17 chromosomes, indicating a typical haplodiplophasic life history as seen in other Laminariales.     

THE TRANSITION FROM FILAMENTOUS TO TWO-DIMENSIONAL GROWTH IN FERN GAMETOPHYTES. II. KINETIC STUDIES ON PTERIDIUM AQUILINUM

Gametophytes of the fern Pteridium aquilinum were incubated 3 days in red light and then transferred to white light. The sequence of events occurring after the transfer was as follows: a swelling of the apical region within 1 hr; a reduction in cell elongation after 5 hr; a series of one-dimensional cell divisions between 10-25 hr; and the initiation of two-dimensional gametophytes after 25-50 hr. The percentage of two-dimensional gametophytes was proportional to the logarithm of the intensity of white light. The rate of elongation after 5 hr was inversely proportional to the logarithm of the intensity. The rates of cell divisions for one- and two-dimensional gametophytes were proportional to intensity up to 240 and 120 ft-c, respectively.