Distribution of Pfiesteria piscicida cyst populations in sediments of the Delaware Inland Bays, USA

The toxic dinoflagellate, Pfiesteria piscicida, is a common constituent of the phytoplankton community in the Delaware Inland Bays, USA. In this study, molecular methods were used to investigate the distributions of benthic stages (cysts) of P. piscicida in sediment cores from the Delaware Inland Bays. Cores from 35 sites were partitioned into nephloid and anoxic layers and analyzed for P. piscicida by nested amplification of the 18S rDNA gene using P. piscicida-specific primers. The presence of inhibitory substances in the PCR reaction was evaluated by inclusion of an exogenous control DNA in the extraction buffer, thus eliminating samples that may yield false-negative results. Our results indicate a patchy distribution of P. piscicida in sediments of the Delaware Inland Bays, with distinct differences between each of the three bays. Overall, P. piscicida was found more frequently in sediments from Rehoboth Bay compared to Indian River and Little Assawoman Bays. These differences suggest (i) that populations of P. piscicida may be more widely distributed in Rehoboth Bay, (ii) that populations of P. piscicida may have been introduced to Rehoboth Bay at an earlier time, (iii) that past blooms of P. piscicida in Rehoboth Bay estuaries may have seeded the sediments with higher numbers of cysts, and/or (iv) that Rehoboth Bay sediments may be more resistant to clearing due to storm turbulence.     

Thymol derivatives from hairy roots of Arnica montana

Five known thymol derivatives were isolated from roots of Arnica montana transformed with Agrobacterium rhizogenes LBA 9402. The compounds were characterized by spectral methods. The pattern of thymol derivatives in light-grown hairy roots was slightly different from that in dark-grown ones. This is the first report on the presence of thymol derivatives in hairy roots of the plant.     

The biology ofAzospirillum-sugarcane association II. Ultrastructure

Summary Tissue cultures of sugarcane support abundant growth ofAzospirillum brasilense (SP 7). Visible after 1–2 weeks as a white or pink slime, this growth reaches 2×10⁸ bacteria/mm² on the surface of callus. Growth of the bacterium is strictly extracellular in viable callus, and instances of intracellular growth result from rupture of the cell wall during senescence of callus tissue. A significant proportion of the bacterial population on callus is pleomorphic. Varying the nitrogen source in the nutrient medium caused no obvious effect on callus cell structure. The presence of the bacterium caused structural alterations in callus cells which did not inhibit overall growth of the bacterium. Growth of callus as tight groups of cells lacking intercellular spaces may be important for the establishment of a long-term association withAzospirillum. The interface of bacteria and live callus tissue is at the surface of tight cell groups. Browning of the surface cell layers of these groups in the presence ofAzospirillum is not of the rapid nature known for hypersensitivity reactions. Rather, this production of phenolics appears to be due to the accumulation of extracellular bacterial metabolites. The ultrastructure of this and other callus reactions is described. As evidenced by organogenesis, the associated cultures have remained viable for at least 18–20 months.Florida Agricultural Experiment Station Journal Series No. 1695.     

Ultrastructure of mitosis inAmoeba proteus

Summary The fine structure ofAmoeba proteus nuclei has been studied during interphase and mitosis. The interphase nucleus is discoidal, the nuclear envelope is provided with a honeycomb layer on the inside. There are numerous nucleoli at the periphery and many chromatin filaments and nuclear helices in the central part of nucleus.In prophase the nucleus becomes spherical, the numerous chromosomes are condensed, and the number of nucleoli decreases. The mitotic apparatus forms inside the nucleus in form of an acentric spindle. In metaphase the nuclear envelope loses its pore complexes and transforms into a system of rough endoplasmic reticulum cisternae (ERC) which separates the mitotic apparatus from the surrounding cytoplasm; the nucleoli and the honeycomb layer disappear completely. In anaphase the half-spindles become conical, and the system of ERC around the mitotic spindle persists. Electron dense material (possibly microtubule organizing centers—MTOCs) appears at the spindle pole regions during this stage. The spindle includes kinetochore microtubules attached to the chromosomes, and non-kinetochore ones which pierce the anaphase plate. In telophase the spindle disappears, the chromosomes decondense, and the nuclear envelope becomes reconstructed from the ERC. At this stage, nucleoli can already be revealed with the light microscope by silver staining; they are visible in ultrathin sections as numerous electron dense bodies at the periphery of the nucleus.The mitotic chromosomes consist of 10 nm fibers and have threelayered kinetochores. Single nuclear helices still occur at early stages of mitosis in the spindle region.     

Observations on the social waspRopalidia montana from peninsular India

Four colonies ofRopalidia montana collected in August in the Nilgiri Hills of southern India had adult populations of 32,000-61,000. Queens represented 0·46–1·40% of the populations. All colonies contained males, but in widely varying proportions (1·47–27·00%). The large adult populations and the fact that the nests were largely filled with brood in all stages of development indicate that the colonies were in a stage of active growth in this season. In December, however, colonies have been reported to have smaller adult populations and little brood. Thus brood production appears to be seasonal in southern India. Predation by the hornetVespa tropica was observed in 4 of 31 active colonies.Ropalidia montana adults were defenseless in the face of these depredations, which appear to continue at a low level for days or weeks on a given nest.     

Antimicrobial properties of essential oil of Satureja montana L. on pathogenic and spoilage yeasts

The essential oil of Satureja montana L. had a broad-spectrum of antimicrobial activity against 46 species of yeasts. This high and diffused activity could be used to control potential pathogenic and spoilage yeasts. The assay of MIC toward some pathogenic and spoilage yeasts showed a range values from 0.10 to 0.25 l ml^–1. The MIC and growth rate reduction assay were effective tests for quantitative evaluation of antimicrobial activity.     

Reproductive success and clonal genetic structure of the rareArnica montana (Compositae) in The Netherlands

In a medium-sized population ofArnica montana, a threatened species in The Netherlands, the breeding system, reproductive success and genetic clonal structure were studied. Pollination experiments suggested thatA. montana is largely self-incompatible. Inbreeding depression was observed for seedling weight but not for fruit weight and germination rate. Although genetic variation is rather low in this population, the data suggest an outcrossing mating system. Analysis of the genotype of all mapped rosettes in a plot of 100 m² indicated that dense clusters often consist of identical genotypes, suggesting a clonal structure. Open clusters frequently contained several different genotypes. This may be caused by limited fruit dispersal, since seedlings were found mainly within or in the near surroundings of the clusters.     

Unusual oleanane-type saponins from Arenaria montana

Three oleanane-type saponins, 3-O-β-d-glucopyranosylechinocystic acid 28-O-β-d-xylopyranosyl-(1→4)-[α-l-rhamnopyranosyl-(1→2)]-α-l-rhamnopyranosyl ester (1), 3-O-β-d-glucopyranosylechinocystic acid 28-O-α-l-arabinopyranosyl-(1→3)-β-d-xylopyranosyl-(1→4)-[α-l-rhamnopyranosyl-(1→2)]-α-l-rhamnopyranosyl ester (2), 3-O-β-d-glucopyranosylcaulophyllogenin 28-O-β-d-apiofuranosyl-(1→3)-β-d-xylopyranosyl-(1→4)-[β-d-apiofuranosyl-(1→3)]-α-l-rhamnopyranosyl-(1→2)-α-l-rhamnopyranosyl ester (3) were isolated from the whole plant of Arenaria montana. Their unusual structures for the Caryophyllaceae family were established mainly by 2D NMR techniques and mass spectrometry.     

Ultrastructure and ontogeny of a new type of eyespot in dinoflagellates

Summary Ultrastructure and ontogeny of a new type of eyespot in dinoflagellates is described. A marine tidal poolGymnodinium natalense is found to possess a highly organized eyespot whose structure is unique among dinoflagellates. The eyespot is rectangular in ventral view, C-shaped in apical view, and is located posterior to the sulcus. The eyespot is independent of the chloroplast and consists of several (typically six) layers of hemi-cylindrical walls which are concentrically arranged with narrow spacing between them. Each hemicylindrical wall is enclosed by a single unit membrane and is composed of many regularly arranged rectangular crystalline bricks. These crystalline bricks are produced in small vesicles which are formed in the invaginations of the chloroplast. The vesicles containing newly formed crystalline bricks are then transported to the sulcal area to assemble the eyespot. The crystalline bricks are arranged in a neat row within the vesicle termed “eyespot forming vesicle” (EFV), which is located near the sulcus. The hemi-cylindrical wall is constructed within the EFV. Based on the structure of the eyespot, viz. consisting of concentric multi-layered walls, the eyespot is thought to act as a quarter-wave stack antenna.     

Pollination biology and breeding system of Zeyheria montana (Bignoniaceae)

The reproductive biology of Zeyheria montana was studied through field observations of flower visitors and floral events, controlled manual pollinations, and observations of pollen tube growth and ovule penetration by fluorescence microscopy. Analysis of secretory areas of the nectar chamber was made by flower dissections and histology of serial sections. The flower lasted 6–8 days, but pollen exposure and stigma receptivity occurred only up to the end of the first and second days, respectively. Pollination was effected by several species of hummingbirds, especially Colibri serrirostris. The flowers present a rudimentary, non-functional disc, and secretion of nectar is performed by corolla-borne glandular trichomes. Only hand cross-pollinated and natural-pollinated flowers set fruits. Artificially self-pollinated and non-pollinated flowers dried off after anthesis without presenting any swelling of the ovary. Almost all the ovules in selfed and crossed pistils were penetrated 96 h after pollination. However, a delay in ovule penetration in self-pollinated pistils was verified, which indicates the occurrence of late-acting self-incompatibility.We are grateful to Fundo de Apoio ao Ensino e à Pesquisa (UNICAMP) for financial support, and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for the fellowship granted to the first author (this work is part of a Ph.D. thesis presented to UNICAMP, Campinas, SP, Brazil).     

Studies on cyst formation inPhysarum polycephalum myxamoebae

Encystment of myxamoebae ofPhysarum polycephalum was induced by transferring the amoebae to a high salt medium of 1/60 M Sørensen buffer (pH 6.0) containing 0.125 M NaCl, 1.6 mM MgCl₂ and 0.18 mM CaCl₂. The induction of cysts was blocked by inhibitors of protein synthesis, such as puromycin, cycloheximide and streptomycin. However, inhibitors of RNA synthesis, such as actinomycin D, proflavin and 8-azaguanine did not block the transformation. These results suggest that in the cyst formation,de novo RNA synthesis is not involved, whereas protein synthesis is required. Cyst formation was more strongly inhibited by inhibitors of oxidative phosphorylation than by other respiratory poisons. It seems that oxidative phosphorylation takes part in the energy supply of this differentiation.     

Distribution of alkanes and fatty acids in theCondalia montana (Rhamnaceae) species complex

The alkane and fatty acid composition of the lipid extracts from leaves of 14 populations ofCondalia: C. microphylla f.xanthocarpa, C. microphylla f.erythrocarpa, C. microphylla f.melanocarpa, C. montana, andC. buxifolia, were determined by gas chromatography. Seventeen alkanes and 14 fatty acids were investigated and a predominance of nonacosane, hentriacontane, palmitic and linoleic acids in all species was found.     

Habitat characteristics and the effect of various nutrient solutions on growth and mineral nutrition ofArnica montana L. grown on natural soil

Arnica montana, one of the character species of the replacement plant communityViolion caninae on sandy acid humic podzol, is declining in the Netherlands since 1950. Locally, it is even extinct.This process of decline may be attributed to (i) autonomic succession; (ii) increased rate of acidification of the soil and/or (iii) change in competitive relations amongArnica and more productive plant species, such as the successive dominantDeschampsia flexuosa. This paper examines the last two hypothesized factors, supposedly being regulated by atmospheric input of N-input, by (a) describing the habitat characteristics of a seemingly still healthy individual-rich population ofArnica and (b) growingArnica andDeschampsia in mixed humic podzol soil (mainly A₁ horizon) fertilized with variously-composed nutrient solutions in order to assess the nutrient supplying capacity of the podzol soil and species-specific nutritional demands related to their respective growth capabilities.The results suggest that an increased rate of soil acidification yielding extra supply of ionic Al and Mn is presumably of less importance. The implications of growth rate differences amongArnica andDeschampsia as related to their nutritional demands are likely far more important.Arnica grows more slowly thanDeschampsia in fertilized humus podzol. The latter species has a much more efficient use of nutrients. Furthermore, both species differ in K-nutrition if NH₄ ⁺ is the dominant N-source, a situation normally occurring in acid podzolic soils.Deschampsia possesses the capability to adapt its metabolic nutrition (avoiding nutritional imbalances) to a wide range of nutrient supplies thereby maintaining a rather constant level of growth.Arnica, on the contrary, lacks this capability. These results are discussed in the framework of competitive relations among co-occurring plant species in the plant communityViolion caninae. It is concluded that maintenance ofArnica and likely other character species of that alliance — all characterized by a low growth rate — will only be achieved when the plant community is properly managed by trampling, mowing or light levels of grazing (low stocking rate). Of prime importance is to maintain a low and relatively open vegetation structure at a relatively low level of nutrient supply.     

Ultrastructure ofCyanoptyche gloeocystis f.dispersa (Glaucocystophyceae)

Cyanoptyche gloeocystis f.dispersa (Geitler)Starmach is a palmelloid colonial alga that contains prokaryotic blue-green endocytobionts (cyanelles) instead of chloroplasts. The periphery of the host cell shows a peculiar lacunae system with underlying microtubules. Vegetative cells possess two rudimentary flagella. Zoospores are dorsiventrally shaped with two heterokont and heterodynamic flagella which originate from a subapical depression. This depression can also be seen in vegetative cells. Both flagella possess non-tubular mastigonemes. Main reserve product is starch lying freely in the cytoplasm. Cyanelles, enclosed singly in a host vesicle, are provided with a remnant cell wall. Thylakoids are arranged concentrically. The central part of each cyanelle harbours its DNA and one large polyhedral body, probably a carboxysome.Cyanoptyche gloeocystis f.dispersa shares all taxonomically essential characters with the monadoidCyanophora, the palmelloidGloeochaete, and the coccoidGlaucocystis. All of them are members of the cyanelle-bearing small algal classGlaucocystophyceae. Members of this class serve as model organisms for the evolution of chloroplasts from cyanophycean ancestors.Dedicated to Prof. DrLothar Geitler on the occasion of his 90th birthday.     

Observations on the fine structure of the cercaria of Notocotylus attenuatus and formation of the cyst wall of the metacercaria

Summary The dorsal tegument of the mature cercaria of Notocotylus attenuatus is a syncytial, cytoplasmic layer, containing two types of secretory granule which are identifiable ultrastructurally. The type 1 secretory bodies are electron lucid, whereas most type 2 granules have a banded appearance. The ventral tegument contains granules which are secreted from the type 3 cells; the type 3 granules are membrane bound, electron dense, and consist of both an amorphous and a finely striated zone. The type 4 cells mainly contain cigar-shaped granules consisting of an amorphous core surrounded by concentric striations. The granules exhibit structural variability in shape and content. The type 4 cells undergo a cellular migration to the tegument during encystment. The structure of the posterior-lateral glands and mode of secretion of the granules are described. Possible functions of microtubules are discussed for each cell type. Details of some secretory processes involved in the formation of the hemispherical cyst wall are described. The layers of the cyst wall may be related to the granular contents of the various parenchymal cells of the cercaria. The tegument of the metacercaria originates primarily from the cytoplasm of the type 1, type 2, type 3 and type 4 cells.     

Multinucleate cyst formation in Acetabularia mediterranea after x-irradiation

Cells of Acetabularia mediterranea were irradiated with increasing doses of X-rays (64.5–258·10^-4 kC kg^-1). The cells are radioresistant up to 193.5·10^-4 kC kg^-1 in terms of growth and progression through he life cycle but the morphogenesis of whorls, caps, and cysts is accompanied by morphological alterations. Microscopical examination of cyst bearing caps in irradiated cells has shown the presence of giant cysts neighboring particularly small ones. Photographic recording of cyst development showed that the multinucleate cap cytoplasm partitions into multinucleate portions rather than uninucleate ones as in the control cells. After complete cleavage a cyst wall is deposited onto the multinucleate cytoplasm. In contrast to uninucleate cysts with one lid the wall contains multiple lids. Their number appears to correspond to the number of nuclei in the cytoplasm compartment during cleavage. The results indicate that X-rays preferentially inhibit the synthesis of a factor which plays a role in establishing the normal spatial morphogenetic pattern necessary for cyst formation.     

Comparative analysis of two algicidal bacteria active against the red tide dinoflagellate Karenia brevis

The red tide dinoflagellate Karenia brevis blooms annually along the eastern Gulf of Mexico, USA, and is often linked to significant economic losses through massive fish kills, shellfish harvest closures, and the potential threat to humans of neurotoxic shellfish poisonings as well as exposure to aerosolized toxin. As part of an effort to enhance the strategies employed to manage and mitigate these events and their adverse effects, several approaches are being investigated for controlling blooms. Previous studies have established the presence of algicidal bacteria lethal to K. brevis in these waters, and we aim to characterize bacterial–algal interactions, evaluate their role as natural regulators of K. brevis blooms, and ultimately assess possible management applications. Herein, the algicidal activity of a newly isolated Cytophaga/Flavobacterium/Bacteroidetes (CFB)-bacterium, strain S03, and a previously described CFB-bacterium, strain 41-DBG2, was evaluated against various harmful algal bloom (HAB) and non-HAB species (23 total), including multiple clones of K. brevis, to evaluate algal target specificity. Strains S03 and 41-DBG2, which employ direct and indirect modes of algicidal lysis, respectively, killed 20% and 40% of the bacteria-containing isolates tested. Interestingly, no bacteria-free algal cultures were resistant to algicidal attack, whereas susceptibility varied occasionally among bacteria-containing isolates of a single algal taxon originating from either the same or different geographic location. The dynamics of K. brevis culture death appeared to differ according to whether the algicidal bacterium did or did not require direct contact with algal cells, with the former most rapidly affecting K. brevis morphology and causing cell lysis. Both bacterial strains promoted the formation of a small number of cyst-like structures in the K. brevis cultures, possibly analogous to temporary cysts formed by other dinoflagellates exposed to certain types of stress. Results were also consistent with earlier work demonstrating that bacterial assemblages from certain cultures can confer resistance to attack by algicidal bacteria, again indicating the complexity and importance of microbial interactions, and the need to consider carefully the potential for using such bacteria in management activities.     

Effects of temperature and light on cyst germination and germinated cell survival of the noxious raphidophyte Heterosigma akashiwo

The effects of temperature and light on the germination of Heterosigma akashiwo cysts were examined using bottom sediments collected from Hakata Bay, Japan. In a suspension of mixed sediment and seawater in the temperature range of 5–30 °C, motile cells emerged within 3 weeks, but at ≤12 °C the cell numbers were markedly lower and the emergence of motile cells delayed. When suspension samples incubated at various temperatures were moved to 20 °C and incubated, only a few additional motile cells emerged. The number of motile cells germinated in the dark was significantly lower than under light conditions. When suspension samples incubated in the dark were exposed to light, only a few additional motile cells emerged. These results indicate that the initiation of germination in Heterosigma cysts suspended in seawater is not dependent on temperature and light conditions, although the speed of the germination process is affected by temperature, and cell survival just after germination is strongly affected by temperature and light.     

Ultrastructure of the early stages of carposporophyte development in the red algaChondria tenuissima (Rhodomelaceae,Ceramiales)

The ultrastructure of the early stages of carposporophyte development in the marine red algaChondria tenuissima has been studied. The diploid carposporophyte grows on the gametophyte. Apical gonimoblast cells develop into diploid carpospores. The basal gonimoblast cells cease to divide and undergo considerable cytoplasmic changes before they become incorporated into the expanding fusion cell. Nucleus and plastids degenerate gradually, while mitochondria remain intact. The smooth endoplasmic reticulum becomes prominent, it seems to produce small vesicles with electron dense contents. Simultaneously, numerous mucilage sacs are formed, presumably from dilating ER cisternae. The contents of the mucilage sacs are secreted by exocytosis. The pit connections between gonimoblast cells flare out. They remain as isolated bodies without connection to a wall after fusion. Secondary pit connections occur between vegetative gametophyte cells and sterile carposporophyte cells. There are three different morphological types of pit connections.     

The cytoskeleton of spreadingDictyostelium amoebae

Summary The cytoarchitectural elements ofDictyostelium discoideum amoeba have been visualized by light and electron microscopy in cells prepared with mixtures of glutaraldehyde and Triton-X-100. After negative staining, the peripheral regions of spreading amoebae show a complex meshwork of actin filaments, the majority of which were less than 0.25 microns in length. Multiple branch points, end to side abutments and cross-overs were characteristic features of the actin meshworks. Filopodia extending from the cell periphery consisted of bundles of actin filaments that penetrated into and merged with the actin meshworks in the spreading lamellae. Microtubules emanating from the nucleus associated body penetrated to differing extents into the actin meshworks, sometimes extending close to the cell periphery.Dictyostelium cytoskeletons preparted as described here should prove useful for further studies on the locomotory mechanism.