CHROMOSOME MUTATIONS IN A FERN POPULATION GROWING IN A POLLUTED ENVIRONMENT: A BIOASSAY FOR MUTAGENS IN AQUATIC ENVIRONMENTS

A population of royal ferns, Osmunda regalis, which is periodically submersed by the waters of the Millers River was found to have a very high frequency of chromosome mutations. The Millers River is located in western Massachusetts and is heavily polluted with industrial wastes. Approximately 43 % of meiotic samples collected in 1973 from the Millers River population were heterozygous for mutations such as paracentric inversions or reciprocal translocations, whereas less than 1 % of meiotic samples from nearby non-polluted control populations were such heterozygotes. Cytological analysis within royal fern clones indicate that practically all of the chromosome mutations were post-zygotically induced and that at least 64 % were induced since 1969. In the course of this study over 26,000 spore mother cells were analyzed for chromosome aberrations.     

GENETIC LOAD IN OSMUNDA REGALIS POPULATIONS

Osmunda regalis sporophytes form haploid spores which develop into functionally hermaphroditic gametophytes. The self-fertilization of such gametophytes results in zygotes which are completely homozygous. Spore samples collected from sporophytes in natural populations were used to establish gametophyte cultures. The majority of these gametophytes were unable to form viable embryos when only self-fertilization was possible. Controlled selfing and crossing experiments revealed that the inability of these homozygous embryos to develop normally is attributable to the presence of recessive lethals. To account for this genetic load, an hypothesis is proposed integrating the morphology and ecology of the gametophyte generation with the polyploid genetic system of the sporophyte generation.     

GAMETOPHYTE ONTOGENY,SEX EXPRESSION,AND GENETIC LOAD AS MEASURES OF POPULATION DIVERGENCE IN BLECHNUM SPICANT

Early gametophyte ontogeny was quantitatively distinct for Olympic Peninsula, Alaskan, and disjunct Idaho populations of the homosporous fern Blechnum spicant (L.) J. Sm. Although variable, gametophyte sex expression was shown to have a genetic component. Statistically different patterns of sex expression characterize each population. The Olympic Peninsula populations were distinct from each other but consistent in having a predominantly unisexual pattern. The disjunct Idaho population was predominantly bisexual at the time when comparable field collected gametophytes bear sporophytes. Preliminary experiments suggest that an antheridogen operates in this species. Increased sowing density favors maleness, and an extract from soil cultures of gametophytes shifts cultures to an exclusively male pattern after a dramatic suppression of growth. Mating experiments revealed that all populations are interfertile, although fertility was highest when the test Idaho population underwent intergametophytic-selfing. The Idaho population evidenced a low level of genetic load consistent with predictions based on its sex expression. Although Olympic Peninsula populations evidenced apparent high genetic load in some experiments, failure to produce abundant sporophytes in other experiments suggested that additional cultural factors operated to reduce sporophyte formation. Moderate density mating experiments produced single sporophytes that were comparable to field collections. Isolated gametophytes underwent polyembryony after a time delay and gametophyte proliferation. Cultural conditions which allow sporophyte formation on isolated gametophytes without this delay or proliferation must be sought before further genetic analysis is undertaken.     

REPRODUCTIVE LONGEVITY OF DRIFTING KELP MACROCYSTIS PYRIFERA (PHAEOPHYCEAE) IN MONTEREY BAY,USA1

Drifting Macrocystis pyrifera (L.) C. Agardh sporophytes have long been viewed as the primary long‐distance dispersal vector; yet, few data exist that support the ability of reproductive viable sporophytes to actually travel the presumed hundreds to thousands of kilometers. This study addressed the reproductive longevity of experimental and naturally occurring M. pyrifera drifters. Temporal variability in sporophyte size and reproduction was estimated for experimental drifting sporophytes that were tethered to surface buoys and compared with attached plants (controls). Reproductive viability was also studied for beach‐cast drifters (BCD), and naturally drifting sporophytes observed during field surveys in Monterey Bay. Detached drifting sporophytes were tracked with radio transmitters to follow drifter trajectories and to measure drifting speed. Experimental drifters (ED) experienced a 74% reduction in frond length after 35 days, a 76% reduction in average frond number after 70 days, and a reduction in average sorus area by 83% after 28 days. Although zoospore production was reduced following detachment, sporophytes remained fertile with high zoospore germination success as long as sori were present (125 days). Zoospore production and germination success for natural and BCD was similar to ED. The average displacement of radio‐tagged drifters was 7.12 km·day^?1, suggesting that a sporophyte adrift for 125 days disperses viable propagules (zoospores) over 890 km (±363). Dispersal of propagules is important for population restoration, distribution, and genetic diversity. Such dispersal distances are long enough to connect potentially all Northern Hemisphere Macrocystis populations across a generational timescale and may facilitate inter‐hemispheric gene flow.     

OBLIGATE OUTCROSSING IN A HOMOSPOROUS FERN: FIELD CONFIRMATION OF A LABORATORY PREDICTION

While homosporous ferns are potentially capable of producing totally homozygous sporophytes in one generation via selfing of their bisexual gametophytes, laboratory analyses indicate that a variety of mechanisms promote gametophytic outcrossing. The operation of these mechanisms in natural sporophyte populations, however, has not been previously demonstrated. Laboratory analyses of gametophyte ontogeny show that Bommeria hispida is obligately outcrossing. Electrophoretic data presented here indicate that individuals from natural sporophyte populations of this species are highly heterozygous. Electrophoretic data, therefore, corroborate evidence from the in vitro analysis of gametophyte development and demonstrate that sporophytes of B. hispida in nature typically are products of outcrossing between genetically different gametophytes. Extrapolations from the literature, together with our findings, indicate that outcrossing mechanisms may operate frequently in ferns, thereby maintaining genetic variability between individuals within populations. This evidence questions whether most ferns are highly inbred and therefore predominantly homozygous.     

MUTATION IN FERNS GROWING IN AN ENVIRONMENT CONTAMINATED WITH POLYCHLORINATED BIPHENYLS

Polychlorinated biphenyls (PCBs) are ubiquitous and persistent environmental contaminants. The Housatonic River in Massachusetts is heavily contaminated with PCBs in the vicinity of the city of Pittsfield. The ostrich fern, Matteuccia struthiopteris, forms extensive populations in the flood plain of this river. The frequency of somatic mutations was determined in these fern populations as well as control populations growing in uncontaminated environments in the Connecticut River and its tributaries. The frequency of somatic mutations in the Housatonic River populations was 5.26–6.14 times the controls. Whether similar genetic damage has occurred to other inhabitants of the Housatonic environment is unknown.     

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.     

Herberta borealis,a new species from Scotland and Norway

Abstract

The reproduction of Pohlia nutans (Hedw.) Lindb. and Pleurozium schreberi (Brid.) Mitt. were compared in the surroundings of copper smelters at Harjavalta, Finland. Samples of both species were collected from 16 sample plots, which were situated on a 12-km transect at intervals of half a kilometre from the smelters. As a result of pollution the production of gametangia decreased in P. schreberi, and near the smelters most of the shoots were sterile. The biomass of shoots seems to have an effect on the production of perichaetia; reproducing females were always the largest ones in the colonies. The reproductive strategy of Pohlia nutans also varied. While, in moderately clean areas, colonies reproduced sexually and formed sporophytes, in the vicinity of the smelters caducous shoot apices were produced. The numbers of sporophytes in colonies were highest in moderately polluted areas and decreased further away from the smelters. The numbers of spores per capsule in P. nutans decreased at polluted sites and the proportion of aborted spores increased.     

GROWTH INHIBITION IN GAMETOPHYTES AND OAT COLEOPTILES BY THELYPTERIN A AND B RELEASED FROM ROOTS OF THE FERN THELYPTERIS NORMALIS

Gametophytes, when grown in the immediate vicinity of a Thelypteris normalis sporophyte—in soil or in sterile culture on agar—showed a reduced number of cells and an altered gross morphology. This is attributed to the action of the thelypterins, which are inhibitors released from T. normalis sporophytes. Growth inhibition of the gametophytes was greatest when thelypterins were added during early stages of gametophyte development. Removal of thelypterin A permitted resumption of growth. Thelypterin A noncompetitively inhibited auxin-enhanced elongation of A vena coleoptiles. The growth of T. normalis root segments was not affected by thelypterins. The growth of young sporophytes of T. normalis was inhibited by mature sporophytes.     

A methodology for large-scale Athyrium sheareri gametophyte proliferation and sporophyte production using tissue culture

Tissue culture methods using gametophytes are considered the easiest ways to mass-produce fern sporophytes. The aim of this study was to develop a practical propagation method for the ornamental fern, Athyrium sheareri. The gametophytes obtained from in vitro spore germination were used as experimental materials. We used the chopping method to investigate the culturing conditions for proliferating gametophytes and the blending method for evaluating the mass production of sporophytes in mixed soil. Gametophyte proliferation was determined via Knop medium, various concentrations of Murashige and Skoog (MS) basal medium (1, 1/2, 1/4), and media components (sucrose, nitrogen source, and activated charcoal). The fresh weight of the gametophytes increased by more than 24-fold in 1/2 MS medium. In addition, 1 g of gametophyte could produce a maximum of 255.3 sporophytes in a mixed soil of 7.5 cm² area. Treating gametophytes with exogenous plant growth regulators promoted the formation and growth of sporophytes. The cultivated young sporophytes were acclimated and successfully grown in greenhouses. We developed a mass production protocol for A. sheareri sporophytes suitable for field application, which is expected to have commercial value.

     

STUDIES ON THE GENETIC CONTROL OF RESISTANT SPORANGIUM FORMATION IN ALLOMYCES

The diploid sporophyte of the phycomycetous fungus Allomyces arbuscula bears two types of sporangia: thin-walled, colorless, ephemeral zoosporangia (ZS) and thick-walled, dark-brown, resistant sporangia (RS). Normal wild-type cultures (strain Portugal IE) under standard conditions produce approximately 90% of their total sporangia as RS. These RS give the cultures a dark-brown color. A mutant was induced with UV irradiation in which the ratio of ZS to RS was shifted so that only 20% of the total sporangia are RS. These cultures are a pale, tan color. Hybrids between the mutants and wild-types produce ca. 65% RS and are also intermediate in the color of the culture. Meiotic segregation in the RS of the hybrid sporophytes gives gametophytes half of which when selfed produce mutant sporophytes and half of which produce wild-type sporophytes. The shift from RS to ZS formation is thus considered to be the result of a one-gene mutation at a locus ‘R.’ The haploid gametophytes of wild-type strains have in addition to male and female gametangia a small number (2-4%) of RS. In mutant gametophytes the percent RS has dropped to 0.1-0.2%. The proposed genotypes at the ‘R’ locus in Allomyces arbuscula are: wild-type sporophytes (RR), hybrid sporophytes (Rr), mutant sporophytes (rr), wild-type gametophytes (R) and mutant gametophytes (r).     

NONHOMOLOGOUS CHROMOSOME PAIRING IN THE FERN CERATOPTERIS

The homosporous fern Ceratopteris thalictroides forms completely homozygous offspring sporophytes in one generation of selfing. Such homozygotes do not breed true and can exhibit structural heterozygosity for chromosomal mutations in some meiocytes. This is due to occasional nonhomologous (presumably homoeologous) chromosome pairing at meiosis in these polyploid ferns.     

Effects of temperature and nutrients on microscopic stages of the bull kelp (Nereocystis luetkeana,Phaeophyceae)

Warming ocean temperatures have been linked to kelp forest declines worldwide, and elevated temperatures can act synergistically with other local stressors to exacerbate kelp loss. The bull kelp Nereocystis luetkeana is the primary canopy-forming kelp species in the Salish Sea, where it is declining in areas with elevated summer water temperatures and low nutrient concentrations. To determine the interactive effects of these two stressors on microscopic stages of N. luetkeana, we cultured gametophytes and microscopic sporophytes from seven different Salish Sea populations across seven different temperatures (10–22°C) and two nitrogen concentrations. The thermal tolerance of microscopic gametophytes and sporophytes was similar across populations, and high temperatures were more stressful than low nitrogen levels. Additional nitrogen did not improve gametophyte or sporophyte survival at high temperatures. Gametophyte densities were highest between 10 and 16°C and declined sharply at 18°C, and temperatures of 20 and 22°C were lethal. The window for successful sporophyte production was narrower, peaking at 10–14°C. Across all populations, the warmest temperature at which sporophytes were produced was 16 or 18°C, but sporophyte densities were 78% lower at 16°C and 95% lower at 18°C compared to cooler temperatures. In the field, bottom temperatures revealed that the thermal limits of gametophyte growth (18°C) and sporophyte production (16–18°C) were reached during the summer at multiple sites. Prolonged exposure of bull kelp gametophytes to temperatures of 16°C and above could limit reproduction, and therefore recruitment, of adult kelp sporophytes.     

Natural soil spore banks — can they be used to retrieve lost ferns?

Some fern species form spore banks — reservoirs in the soil of viable spores which remain dormant while buried but germinate in light if brought to the surface. The recently discovered characteristics of these spore banks are described. Enough is now known to suggest that they might have a role in the conservation of endangered fern species as alternatives toex situ collections of sporophytes, gametophytes and spores, the relative merits of which are also considered. Mature sporophytes of several British species have now been raised from natural spore banks in soil samples; if this proves to be possible also for endangered species, some interesting options become available. The possibilities are discussed of augmenting surviving populations and increasing their genetic diversity, even perhaps of retrieving lost populations, by reintroduction of spore bank-derived plants or by stimulating regeneration from spore banksin situ. Botanic gardens are well placed to provide the further research, the regular monitoring of endangered populations, and the taxonomic and horticultural support required to realise these possiblities.     

ALGAL COMPETITION IN AN EPILITHIC RIVER FLORA1

Field and laboratory procedures demonstrate that the effluent from the Willimantic, Connecticut, sewage treatment plant changes the numerical composition of the dominant epilithic algae in the Shetucket River. This effect is due to the effluent altering the competitive balance between chlorella sp. and Achnanthes deflexa Reim. The chlorella completely dominates highly polluted stretches of the river, and shares dominance with A. deflexa in other areas. The degree of river pollution can be estimated by enumerating the natural standing crops of these 2 algae. A 2-membered assay utilizing these organisms is a sensitive laboratory pollution monitor and a useful predictive tool. Dominant stands of these epilithic species develop in polluted and unpolluted zones in this river due to competition for the limited area on the cobbles.     

CONTROL OF SEX RATIOS IN HAPLOID POPULATIONS OF THE MOSS,CERATODON PURPUREUS

Female biased sex ratios occur in a number of unrelated mosses. Such ratios refer to the relative numbers of male and female gametophytes in moss populations and are therefore more comparable to the numbers of pollen grains and ovules in populations of seed plants than to the numbers of male (microsporangiate) and female (megasporangiate) sporophytes. A survey of 11 populations of the moss, Ceratodon purpureus, showed that sex ratios are heterogeneous, but that female biases occur in more than half the populations. One hundred and sixty single spore isolates representing 40 sporophytes from one population demonstrated that female gametophytes outnumbered males by a ratio of 3:2 at the time of germination. Female gametophytic clones formed significantly more biomass than male clones, and individual female shoots were more robust. Male clones, however, produced more numerous stems. These sexually dimorphic traits may be related to life history differences between male and female gametophytes since females must provide nutritional support to the “parasitic” sporophyte generation, a burden that males do not share.     

ENVIRONMENTAL CONTROL OF GAMETOGENESIS IN LAMINARIA SACCHARINA. IV. IN SITU DEVELOPMENT OF GAMETOPHYTES AND YOUNG SPOROPHYTES1,2

Meiospores of Laminaria saccharina labeled with a fluorescent stain were put into the sea and recovered at frequent intervals. They developed into gametophytes which produced gametes throughout the year. Antheridia were produced earlier than oogonia. Gametogenesis was slowest during the late autumn and early winter. On the basis of nutritional studies and the seasonal distribution of nutrients, it was concluded that the rate of gametogenesis was not due to nutrient deficiency but most likely to light. Production of macroscopic sporophytes appeared to be limited by survival of microscopic sporophytes rather than gametogenesis. Macroscopic sporophytes became evident during the late winter and autumn. The greater light and perhaps higher temperatures of summer were suggested as agents limiting macroscopic sporophyte production during this period.     

Die-Away Kinetic Analysis of the Capacity of Epilithic and Planktonic Bacteria from Clean and Polluted River Water To Biodegrade Sodium Dodecyl Sulfate

The capacities of epilithic and planktonic river bacterial populations to degrade sodium dodecyl sulfate (SDS) in samples taken at two times during 1987 from one clean and four polluted sites in a South Wales river were estimated in die-away tests under simulated environmental conditions. There was a relatively slow disappearance of SDS in die-away tests for both planktonic and epilithic populations taken from the clean source site, as compared with those taken from the downstream polluted sites, for which the rate of biodegradation was accelerated, sometimes after an apparent initial lag period. The kinetic components contributing to the die-away curves were quantified by nonlinear regression analysis in which the experimental data were fitted to a variety of possible kinetic models. All samples except for one from the polluted sites best fitted a model which describes the biodegradation of SDS at concentrations well below its K_m by bacteria whose growth is exponential and unaffected by the addition of a test substrate. Each sample from the clean source site fitted a different model, but there was generally little or no growth on endogenous carbon. A consideration of the numerical values of constants derived from the modeling of epilithic and planktonic populations from polluted sites indicated clearly that the biodegradative capacity of epilithic bacterial populations towards SDS is more stable than that of planktonic bacterial populations.     

Bathymetric origin shapes the physiological responses of Pterygophora californica (Laminariales,Phaeophyceae) to deep marine heatwaves

Kelp communities are experiencing exacerbated heat-related impacts from more intense, frequent, and deeper marine heatwaves (MHWs), imperiling the long-term survival of kelp forests in the climate change scenario. The occurrence of deep thermal anomalies is of critical importance, as elevated temperatures can impact kelp populations across their entire bathymetric range. This study evaluates the impact of MHWs on mature sporophytes of Pterygophora californica (walking kelp) from the bathymetric extremes (8–10 vs. 25–27 m) of a population situated in Baja California (Mexico). The location is near the southernmost point of the species's broad distribution (from Alaska to Mexico). The study investigated the ecophysiological responses (e.g., photobiology, nitrate uptake, oxidative stress) and growth of adult sporophytes through a two-phase experiment: warming simulating a MHW and a post-MHW phase without warming. Generally, the effects of warming differed depending on the bathymetric origin of the sporophytes. The MHW facilitated essential metabolic functions of deep-water sporophytes, including photosynthesis, and promoted their growth. In contrast, shallow-water sporophytes displayed metabolic stress, reduced growth, and oxidative damage. Upon the cessation of warming, certain responses, such as a decline in nitrate uptake and net productivity, became evident in shallow-water sporophytes, implying a delay in heat-stress response. This indicates that variation in temperatures can result in more prominent effects than warming alone. The greater heat tolerance of sporophytes in deeper waters shows convincing evidence that deep portions of P. californica populations have the potential to serve as refuges from the harmful impacts of MHWs on shallow reefs.