Effects of Dimethyl Sulphoxide on the Gametophytic and Sporophytic phases of Pteridium aquilinum (L.) Kuhn

Dimethyl sulphoxide (DMSO) at concentrations of up to 10 mlper litre of growth medium was found to have no significantinfluence upon Pteridium gametophyte growth or morphology However,significant effects upon embryo development and sporophyte morphologywere shown The most striking of these was that more than oneembryo developed from each fertilized gametophyte grown on DMSOmedium Two to five embryos regularly developed on each gametophytecultured on media containing this solvent, control gametophytesbore single sporophytes The significance of these findings inrelation to theories concerning polyembryony are discussed Pteridium, dimethyl sulphoxide, polyembryony     

Changes in the Level of the Protein Nitrogen during Growth of the Gametophyte and the Initiation of the Sporophyte of Dryopteris borreri Newm.

Estimations of the amounts of organic nitrogen in spores anddeveloping gametophytes of Dryopteris borreri, a fern reproducingapogamously and lacking archegonia, have shown that the nitrogenin the gametophytes increases exponentially. The logarithmicrate of increase of the nitrogen remains unchanged during theinitiation and emergence of the sporophyte. Gametophytes aboutto produce sporophytes yielded only filamentous growths whentransferred to a medium containing 8-azaguanine, and the increasein their content of organic nitrogen was very small. The resultsare discussed in relation to similar estimations made by otherworkers upon the gametophytes of Dryopteris erythosora(Eat.)Schott, a fern reproducing sexually. The interpretation placedupon the results obtained for D. erytkrosora is questioned,and a new interpretation of these results, together with thosefor D. borreri, is related to the difference in morphology ofthe two generations in the life cycle of the Polypodiaceousferns.     

变异鳞毛蕨的孢子培养与配子体发育研究

应用无菌培养和常规泥土培养两种方法对变异鳞毛蕨(Dryopteris varia)孢子进行了比较研究,并在光学显微镜下观察了其配子体的发育过程.结果表明:蔗糖浓度为2%的1/2MS与MS培养基对孢子萌发时间和萌发率影响不大,但前者较适于孢子萌发,而后者则适于孢子体形成;在1/2MS培养基上,1%的蔗糖浓度比其它浓度更适宜于孢子的萌发.以菜园士为培养基质时,变异鳞毛蕨孢子的萌发时间短且萌发率高,但幼孢子体出现的时间明显晚于无菌培养.孢子萌发为书带蕨型,原叶体发育为三叉蕨型,符合鳞毛蕨属配子体发育的特征.     

Apical Structure of Actively Growing Fern Rhizoids Examined by DIC and Confocal Microscopy

The development and cytology of gametophyte primary rhizoidsof the fern Dryopteris affinis was examined using actively growingmaterial. During development an apical cytoplasmic ‘ accumulation’forms and is associated with active tip growth. This accumulationdeteriorates as terminal differentiation and cessation of growthapproaches. During early development the nucleus moves fromthe rhizoid cell base into the newly extending rhizoid. Later,during the active elongation phase, the nucleus takes up a relativelystable location approx. 100 µm behind the extending apex.Towards terminal differentiation the nucleus lags further behindthe tip. In actively growing rhizoids four distinct zones weredistinguished: a richly cytoplasmic ‘cap’; an apicalregion with tubular vacuolar intrusions; a region distinguishedby a peripheral sheath of cytoplasm and fine irregular cytoplasmicstrands connecting to the nucleus; and the main subapical vacuole.Confocal microscopy of gametophytes stained with fluorescentvital dyes, not previously used to examine fern rhizoid structure,confirmed that the tubular vacuolar system extends well intothe apical cytoplasm, and that the network of fine cytoplasmicstrands leads back from the apical cytoplasm to the nucleus.It also revealed that mitochondria are distributed throughoutthe rhizoid and are not excluded from the extreme apex. Membranestaining by FM 4-64 suggested a high density of membrane vesicleswithin the cytoplasm of the extreme apex. Uptake of this endocytosismarker into endomembranes also suggested rapid plasma membraneturnover in the rhizoid. This study highlights the similarityin the developmental stages and appearance of D. affinis rhizoidsto angiosperm root hairs and their much less distinct apicalzonation compared to pollen tubes. Copyright 2000 Annals ofBotany Company Rhizoid, root hair, confocal imaging, vital stains.     

Microsporogenesis, Pollination, Pollen Germination and Male Gametophyte Development in Taxus brevifolia

Taxus brevifolia(Nutt.), commonly known as Pacific or westernyew, is a conifer native to the Pacific northwest of North America.Contrary to other Taxus species, T. brevifolia staminate strobiliare usually located on 2-year-old foliage although they mayoccur on foliage from 1 to 5-years-old. This delayed staminatestrobilus development may be an adaptation to the low lightenvironment where T. brevifolia grows. Microsporogenesis occurredin the autumn preceding pollination. Successive divisions producedisobilateral tetrads visible as early as mid-October. Over-winteringstaminate strobili usually contained separate microspores. In1996 to 1999, pollination occurred in March and April in twonatural forest sites on southern Vancouver Island, British Columbia,Canada. The low amounts of airborne pollen and prolonged pollinationperiod indicated low pollination success within T. brevifolia.Female receptivity was measured by the presence of a pollinationdrop. Protandry up to 18 d was observed. In vitro pollen germinationwas moderate to good, ranging from 65 to 88% depending on thetree and year. DAPI fluorescence staining showed successfulmale gametophyte development in vitro. The microspore dividedforming a tube nucleus and generative cell within 3 d of culture.The generative cell then divided forming a sterile nucleus andspermatogenous nucleus after 17 d. The spermatogenous nucleusacquired a cell wall then divided forming two equal sperm after24 d. Copyright 2000 Annals of Botany Company Taxus brevifolia, Pacific yew, microsporogenesis, pollination, pollen germination, male gametophyte development     

The Effect of the Female Gametophyte on the Growth of Cultured Douglas-fir Embryos

Several clones of Douglas-fir, Pseudotsuga menziesii (Mirb.)Franco, embryos grown in aseptic culture in the presence orabsence of the female gametophyte or a crushed aqueous extractof it, have shown striking differences in overall length, sizeand substance(s) responsible for this colour of the cotyledonsand in root length. The growth-promoting effect is diffusiblein agar, heat-stable and has detectable activity at a concentrationof about 0·1 to 1·0 per cent (v/v) in a basalnutrient medium. Evidence is provided that extracts of gametophytemay have synergistic effects with coconut water in promotingmorphogenesis of Douglas-fir cells in suspension culture. Pseudotsuga menziesii, Douglas-fir, female gametophyte, einbryo culture     

Influence of tissue culture conditions on apogamy in Dryopteris affinis sp. affinis

The naturally-occurring apogamy of some ferns can be modified by culture conditions and growth regulators. Gametophytes of the apogamic fern Dryopteris affinis sp. affinis L., were cultured on Murashige and Skoog (MS) basal medium. Changes in concentration of MS medium components, sucrose, agar and different pH values were tested. The addition of benzyladenine (4.43 M) and naphthalene acetic acid (0.53 M) enhanced sporophyte proliferation on the gametophytes. After one month in culture, the gametophytes formed callus with a high morphogenic capacity. Culture of calli on medium without growth regulators yielded about 10,000 sporophytes per 1 g fresh weight of callus. This pattern of differentiation slowed with time to a point where only gametophyte regeneration was observed.Abbreviations BA benzyladenine - 2,4-d 2,4-dichlorophenoxyacetic acid - F.W. fresh weight - MS Murashige & Skoog medium - NAA 1-naphthalene acetic acid - SE standard error     

同形鳞毛蕨成精子囊素系统的初步研究

研究了同形鳞毛蕨成精子囊素对该种和水蕨孢子萌发和配子体发育的影响,结果表明：同形鳞毛蕨配子体能产生成精子囊素,该成精子囊素能抑制同种孢子的萌发,抑制作用随配子体成熟度的增加而增强;同形鳞毛蕨成精子囊素还可促进同种孢子发育为雄配子体;光照条件下,同形鳞毛蕨成精子囊素对水蕨孢子萌发和配子体发育影响不大,黑暗条件下,同形鳞毛蕨成精子囊素能显著的促使水蕨孢子提早萌发,但都不影响其孢子最终萌发率和配子体的性别分化,表明同形鳞毛蕨和水蕨的成精子囊素不属于同一系统。     

Distribution of Potassium, Calcium and Magnesium in the Gametophyte and Sporophyte Generations of Funaria hygrometrica Hedw

Analyses have been made of the K, Ca and Mg in separated sporophytes,spores and gametophytes of the moss Funaria hygrometrica duringsporophyte development. Degeneration of the gametophyte wasaccompanied by loss of K and a gain in Ca while the K contentof the developing sporophyte increased more rapidly than thatof Ca. The presence of an air–gap in the expanding capsulesignificantly influenced the observed cellular location of ions.Relative to the sporophyte the spores were shown to have a higherK and a lower Ca concentration. The behaviour of Mg was intermediatebetween K and Ca throughout. Funaria hygrometrica, potassium, calcium, magnesium, cation distribution, gametophyte, sporophyte     

Seed Development in Podocarpus henkelii: an Ultrastructural and Biochemical Study

Biochemical and fine-structural studies were undertaken on theembryo and female gametophyte of seeds of Podocarpus henkeliiduring the 24-week period of post-fertilization growth and development. Although the d. wt of the embryo and female gametophyte showeda steady increase during development, the levels of sugars,amino acids, proteins and lipids differed between seed partsand showed changes during the four sampling intervals. Onlystarch was seen to increase steadily throughout development.Lipid levels were high in the mature embryo and free amino acidsshowed a steady increase until the seeds were shed. At thisstage seeds were fully hydrated, possessed abundant reservesand all organelles appeared fully functional. This was interpretedas part of the development strategy of neotonous (recalcitrant)seeds, namely, the maintenance of full metabolic competencefor continued growth in the absence of development arrest whichfollows drying in orthodox seeds. Podocarpus henkelii, yellow wood, embryo and endosperm growth, neotonous seed, recalcitrance     

Growth and Maintenance of an Embryogenic Cell Culture of Daylily (Hemerocallis) on Hormone-free Medium

Callus cultures of the diploid daylily (Hemerocallis) clone‘Autumn Blaze’ were initiated and maintained inhormone-containing nutrient medium. At various times (from 6weeks to 1 year) after being initiated, hormone-derived cultureswere evaluated for their ability to be maintained and to multiplyon hormone-free medium at low pH (between pH 4 and 4.5). Cultureshad to be exposed to hormone-containing medium for at least12 weeks before they could be maintained on hormone-free mediumat low pH. The transition to maintainability on low pH hormone-freemedium included the production of many aberrant embryonal forms('neomorphs'). However, all hormone-derived cultures testedconsisted entirely of preglobular stage proembryos (PGSPs) after12–24 weeks on low pH hormone-free medium. PGSP cultureshave been maintained and multiplied as such for over 1 yearon low pH hormone-free medium. PGSPs continue their developmentinto various somatic embryo stages when cultured on hormone-freemedium buffered at pH 5.8. The production of well-formed somaticembryos was greatly enhanced when PGSPs were plated on activatedcharcoal impregnated filter papers that were placed on top ofthe agar surface. The gross morphology and histology of thePGSPs and stages of somatic embryo development are presented.The work shows that the ability of hormone-free medium at lowpH to permit PGSP multiplication without development into laterstages of embryo development is not restricted to carrot. Hemerocallis cv, ‘Autumn Blaze’, daylily, somatic embryogenesis, hormone-free medium, tissue culture     

Experimental and Analytical Studies of Pteridophytes: XL. Factors in the Formation and Distribution of Sori in Leptosporanglate Ferns

In further studies of soral morphogenesis in common Britishferns, consideration is given to environmental and other factorswhich may be causally involved. Accounts are given of the normalcourse of sporophyll development in Dryopteris austriaca andof the effects of more or less extensive defoliation on theinception of sori in several ferns. Some anomalous developmentsin sporophylls of Blechnum spicant are described and illustrated.Various general ideas regarding factors involved in the onsetof the reproductive phase are discussed. It is concluded that,on the evidence which is admittedly very incomplete, the onsetof the reproductive phase in the fern sporophyte is determinedby the interaction of a number of factors, rather than by thespecial action of some particular factor. Because the nature,functional sorus is a complex and highly organized structure,its further investigation is considered to be a task of veryconsiderable magnitude.     

Investigations on Growth and Metabolism of Plant Cells: IV. Evidence on the Role of the Coconut-Milk Factor in Development

By the carrot-assay method it has been shown that the wateryendosperm of coconut contains the growth-promoting coconut-milkfactor at all stages of development. Some activity is shownby the parts of the immature embryo but not by the solid endosperm. Sources of analogous activity are in the endosperm of Zea maysin the milk stage, the gelatinous content of immature fruitsof Fuglans regia, and the young gametophyte of Ginkgo biloba.The data for other cases examined suggest that the materialdevelops best in nutritive tissues associated with delayed embryodevelopment.     

Seed Germination in Podocarpus henkelii: An Ultrastructural and Biochemical Study

Biochemical and ultrastructural studies were undertaken on theembryo and female gametophyte of neotonous (recalcitrant) seedsof Podocarpus henkelii over a 9-d period following scarificationand incubation on a moist substrate. After 3 days incubationat 25 °C, cells of the root tip were characterized at theelectron microscope level by increased vacuolation, numerousamyloplasts and lipid mobilization. By d 6 measurable embryonicgrowth was noted and ultrastructural evidence of synthetic activitywas suggested by abundant endoplasmic reticulum (ER), ribosomesand dictyosomes. Fine-structural changes suggestive of reservemobilization were also observed in the female gametophyte. Biochemicalstudies indicated a gradual decline in lipid and protein inboth the embryo, and female gametophyte, over the 9 d of incubation.A decline in embryonic starch levels contrasted with the increaseseen in the female gametophyte at d 6. Small changes in thesugar levels of the female gametophyte contrasted with increasesin the embryonic tissues between d 3 and 6, a period coincidingwith the first records of germination. The maintenance of highmoisture contant and the evidence of metabolic activity obtainedfrom biochemical and ultrastructural observations suggest that,following scarification, the transition between maturation andgermination is characterized by a continuation of earlier syntheticactivity and reserve interconversions. Podocarpus henkelii Stapf, yellow-wood, embryo, endosperm, overgrown seed, neotonous, recalcitrant     

Flower-inducing Effect of Benzoic and Salicylic Acids in Various Strains of Lemna paucicostata and L. minor

The flower-inducing activities of benzoic and salicylic acidsadded to the medium differ with the species (Lemna paucicostataand L. minor), and even with the strains used. The type andpH of the medium used, full or 1/10 strength M medium at pH3.8, 4.4 or 5.1, or 1/2 or 1/20 strength NH₄⁺-free Hutner'smedium at pH 5.0, 6.0 or 7.0, also modify their activity. L.paucicostata, strain 151 is the most sensitive of the strainsused to both benzoic and salicylic acids followed by strain381. Such dramatic flowering responses were not obtained withthe other strains, but even strain 321, reportedly insensitiveto benzoic acid, could be induced to flower by adding benzoicacid to a modification of the medium. Benzoic acid is more effectivethan salicylic acid for all strains of L. paucicostata, butthe contrary is true for two L. minor strains tested. A higherpercentage of flowering is obtained in L. paucicostata in 1/2strength NH₄⁺-free Huter'sn medium than in M medium, exceptfor strain 151. When diluted, both media enhance flowering inall L. paucicostata strains. Generally, a lower concentrationof benzoic acid or salicylic acid is enough to induce floweringwhen the pH of the medium is lower. (Received March 30, 1981; Accepted May 16, 1981)     

Growth and reproductive characteristics in artificially formed clonal gametophytes ofDryopteris filix-mas (Dryopteridaceae)

Artificially formed clonal gametophytes ofDryopteris filix-mas (L.)Schott were able to grow and reproduce, although growth rates, proportions of hermaphrodites and reproductive efforts were low. Variable density and nutrient levels appeared to affect gametophytic life histories and they continued to influence the viability of the developing sporophyte generation. Differences between populations and sporophytes were discovered in life histories. Hermaphroditic gametophytes were considerably larger and possessed higher viability than did males. Clear differences between clones in the level of phenotypic plasticity in gametophyte size were observed. No neighbor effect on gametophyte sex expression was detected with the densities and nutrient levels used.     

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.     

Fungal Assciations: II. Cultural Studies on Rhizoctonia solani Kuhn, Fusariutn solani Snyder and Hansen, and Other Fungi, and Their Interactions

Several types of interaction between Rhizoctonia solani, Fusariumsolani, and Phoma foveata were found when these fungi were grownon potato-dextrose agar. After being used by Rhizoctonia a potatomash medium gave better growth of Rizoctonia and Fusarium thanit did when the medium was initially used by Fusarium; and thiswas so whether the reaction of the spent medium was readjustedor not. It is suggested that potato mash medium used by Fusariumcontains a thermostable factor(s) affecting the subsequent growthof Rhizoctonia or Fusarium. The range of pH values suitable for Rhizoctonia growth was narrowerthan that for Fusarium, optimum values being approximately 5•9for the former and 7•8 for the latter. In mixed culturesof the two fungi on potato-dextrose agar adjusted to differentpH values, the fungus for which the reaction of the medium wasmore suitable usually became visually predominant after sometime. A study of various carbon sources showed that poor growth ofRhizoctonia was obtained when pectin was used as the sole sourceof carbon. On a pectin-agar medium, the rate of growth of aRhizoctonia colony increased on the sector which lay towardsan adjacent Fusarium colony; also, after the two fungi camein contact, there was more rapid growth of Rhizoctonia roundthe Fusarium colony than elsewhere. On a synthetic liquid mediumwith pectin as the carbon source better Rhizoctonia growth wasobtained when Fusarium-spent medium was added to it than whenRMzoctoma-spent medium was added. Rhizoctonia showed partial deficiencies in thiamine, biotin,and inositol. Both the extract of Fusarium mycelium, grown onvitamin-free medium, and the Fusarium-spent medium, stimulatedthe growth of Rhizoctonia on vitamin-free medium.     

Importance of B4 Medium in Determining Organomineralization Potential of Bacterial Environmental Isolates

B4 precipitation medium has been used as the preferred medium for studying mineral precipitation using bacterial strains in vitro since pioneer studies were performed by Boquet and coworkers in 1973. Using this medium, several authors have demonstrated that some environmental isolates were able to precipitate minerals, yet others did not. The main goal of the current study is to understand whether pH and buffer conditions would have a significant effect on mineral precipitation results for environmental isolates grown on B4. For this study, a total of 49 strains isolated from natural environments from Puerto Rico were grown on B4 plates, and their CaCO₃ precipitation potential was investigated. Our findings revealed a strong correlation between a lack of CaCO₃ precipitation and the acidification of the B4 plates by the colonies. The ability to precipitate CaCO₃ could be restored by buffering the B4 medium to a pH of 8.2. Buffering capacity of the medium was proposed to be involved in CaCO₃ precipitation: acid-base titrations conducted on the individual ingredients of B4 showed that yeast extract has a poor buffering capacity between pH 6.5–7.5. This pH range corresponds to the pH of B4 plates [6.87 (±0.05)] prior to the inoculation. This might explain why B4 is such a good precipitation medium: a small variation in the H⁺/OH^? balance during microbial growth and precipitation produces rapid changes in the pH of the medium. Finally, an amorphous matrix was distributed within 90% of the examined crystals generated on B4 medium by the environmental strains. Supplemental materials are available for this article. Go to the publisher's online edition of Geomicrobiology Journal to view the free supplemental file.     

Effects of Temperature, pH, and NaCl on Growth and Pectinolytic Activity of Pseudomonas marginalis

The interaction of temperature (4, 10, 18, and 30°C), pH (6, 7, and 8), and NaCl (0, 2.5, and 5%) and their effects on specific growth rate, lag phase, and pectinolytic enzymes of Pseudomonas marginalis were evaluated. Response surface methodology was adapted to describe the response of growth parameters to environmental changes. To obtain good conditions of storage, the combined action of salt and temperature is necessary. At 4°C with an NaCl concentration of 5% and a pH of 7, the lag time was 8 days and no growth was observed at 4°C with 5% NaCl and a pH of 6. In the absence of salt, P. marginalis could grow regardless of temperature and pH. Pectate lyase and pectin lyase were produced by P. marginalis, while pectin methyl esterase activity was not observed in our culture conditions. The enzyme production depended on temperature, pH, and salt concentration but also on the age of the culture. Pectinolytic enzymes were abundantly excreted during the stationary phase, and even at 4°C, after 2 weeks of storage, enzyme activities in supernatant culture were sufficient to damage vegetables. Both bacterial growth and enzymatic production have to be taken into account in order to estimate correctly the shelf life of vegetables.