The ultrastructure of cell walls in Enteromorpha intestinalis (L.) link

Using transmission electron microscopy, the process of cell wall development following cytokinesis is described. Three distinct components appear to be localized in the wall and these are discussed in relation to previous work. In older cells the walls are stratified and the origin of the layers is described. The outer layers of wall are continually being shed and it is suggested that this process prevents epiphytes persisting on this species of alga.     

The motile period of swarmers of Enteromorpha intestinalis (L.) link

Experiments investigating the effects of light and temperature on swarmers of Enteromorpha intestinalis are described. A maximum period of motility of 8 days is recorded and the possible significance of this and other results is discussed.     

Preparation of protoplasts from the green alga Enteromorpha intestinalis (L.) Link

Protoplasts have been obtained from vegetative thallus of the green seaweed Enteromorpha following enzymic digestion with driselase and pectinase. The viability of purified protoplast fractions was assessed by staining and measurements of O₂ uptake and evolution.Abbreviations MES 2-(N-morpholino) ethanesulphonic acid - TES N-tris(hydoxymethyl) methyl-2 aminoethanesulphonic acid     

Ecotypic Variation in the Osmotic Responses of Enteromorpha intestinalis (L.) Link.

Young, A. J., Collins, J. C. and Russell, G. 1987. Ecotypicvariation in the osmotic responses of Enteromorpha intestinalis(L.) Link.—J. exp. Bot. 38: 1309–1324. The physiological basis for salt tolerance has been studiedin the euryhaline marine alga Enteromorpha intestinalis. Adaptationto dilute and concentrated seawaters has been investigated inthree separate populations of this alga: marine, rock pool andestuarine. Internal K⁺, Na⁺ and Cl^– levels have been determined usingtracer efflux analyses. K⁺ has been shown to be the major osmoticsolute within this alga. Cellular levels of Cl^– and, inparticular, Na⁺ are low although levels in the cell wall arehigh. Levels of these ions varied considerably between the separateplants; K⁺ levels within marine plants of E. intestinalis aretwo to four times those found in the other populations. Thetertiary sulphonium compound ß-dimethylsulphonio-propionateis maintained at relatively high levels, although it remainsfairly insensitive to change in the external salinity. Changes in the tissue water content and cell volume are large,particularly within the estuarine plants. The thin cell wallsof these plants allow large changes in volume in the diluteconditions experienced in an estuary, while low turgor preventscell rupture. Thicker cell walls and small cells of the marineand rock pool plants assist in tolerating high and low externalosmotic potential—the estuarine plants respond poorlyto concentrated seawater. Key words: Enteromorpha, osmoregulation, ecotypes     

A fluctuating salinity regime mitigates the negative effects of reduced salinity on the estuarine macroalga, Enteromorpha intestinalis (L.) link

We tested the response of Enteromorpha intestinalis to fluctuating reduced salinity regimes which may occur in coastal estuaries due to both natural and anthropogenic influences. In a fully crossed two factor experiment, we subjected E. intestinalis to 0, 5, 15 and 25 psu water enriched with nutrients for 1-, 5-, 11- and 23-day periods. Each period was followed by 24 h of exposure to 25 psu (ambient) water that was not nutrient enriched. Following 24 h in ambient salinity water, algae were returned to reduced salinity conditions for the appropriate period and the cycle continued over the 24 days for which all treatments were maintained. Exposure to 0 psu for 5 days or longer resulted in loss of pigmentation, decreased wet and dry biomass, increased wet wt:dry wt ratios, decreased removal of nitrogen (N) and phosphorus (P) from the water column and an accumulation of NH(4) in the water column. More frequent exposure to ambient salinity in the 1-day treatment mitigated these effects. Across all salinity levels tested, biomass increased as frequency of exposure to ambient salinity increased. At all durations of exposure to low salinity tested, biomass increased as salinity level increased. We conclude that growth of E. intestinalis is decreased by reduced salinity. E. intestinalis is able to withstand exposure to 0 psu but there is a temporal limit to this tolerance that is somewhere between 1 and 5 days. Populations of E. intestinalis in coastal estuaries may suffer from freshwater inputs if salinity conditions are persistently reduced.     

A simple method for the freeze-preservation of zoospores of the green macroalga Enteromorpha intestinalis

Settled zoospores of the green macroalga Enteromorpha intestinalis were subjected to several different freezing and storing treatments at both cryogenic and non-cryogenic temperatures after which their viability was assessed using a spore germination bioassay. Three different cooling rates were tested: slow cooling at –1°C min⁻¹ and –0.5°C min⁻¹ to end temperatures in the range –20°C to –40°C, and a two-step procedure whereby the spores were frozen to –30°C at a rate of –1°C min⁻¹ prior to immersion in liquid nitrogen at –196°C. Spore viability was also investigated using the cryoprotectants glycerol and dimethyl suphoxide (DMSO), a reduced saline medium and various storage times. In the majority of experiments, the use of a cryoprotectant during the freezing process significantly increased the viability of the spores, with DMSO affording slightly greater protection than glycerol. All treatments produced high viabilities (ranging from 75.3–100.0%) after 5-min storage at the different end temperatures. However, progressively longer storage up to 7 days generally resulted in a marked reduction in viability. This was with the exception of spores frozen in a reduced saline medium; a medium of 75% seawater and either 5 or 10% DMSO greatly increased spore viability, with values of > 40% recorded for spores stored at –20°C for up to 5 weeks. This revised version was published online in July 2006 with corrections to the Cover Date.     

Cation Exchange Properties of the Cell Walls of Enteromorpha intestinalis (L.) Link. (Ulvales, Chlorophyta

The cell wall of Enteromorpha intestinalis (a marine alga) hasbeen found to behave as a weakly cross-linked cation exchangerin NaCl solutions from 0.1–1020 mMolal (0.1–1000mMolar). Anion adsorption could be described by Freundlich isothermsover this concentration range. The large anion, inulin carboxylate,was found to be a tracer of the anion free space of plant tissuesonly in salt solutions above 10 mMolal. The cell wall of Enteromorphahas a cation exchange capacity of about 2500 µ mol g^–1dry wt. (Na+ form). The cell wallvolume is a complex functionof p^H and the NaCl concentration. As a result, the cation exchangecapacity is only predictable on a dry weight basis. The fixednegative charges of the cell wall have a pK_a of2 in situ and1.75 in vitro, and seem to be a mixture of sulphate and carboxylsugar esters. The applicability of the Donnan equation to plant cell wallsis discussed. Interpretation of the cell wall as a single thermodynamicphase is shown to be inappropriate. A large proportion of thecell wall solution is unaffected by the fixed anions.     

Crossing experiments with Enteromorpha intestinalis and E. compressa from different European localities

Sexual compatibility in the marine algae Enteromorpha intestinalis and E. compressa has been studied using laboratory cultures from various geographical areas in Europe. The two species are found to be totally separated by a sterility barrier. Well branched plants always fall in the same of the two genetically isolated groups and should be called E. compressa s.str., while most other plants should be referred to E. intestinalis s.l. The early development of sporophyte germlings may follow two different patterns, both of which have been observed in a single combination of parental plants, though not at the same time.     

Potassium Transport in Enteromorpha intestinalis (L.) Link: II. EFFECTS OF MEDIUM COMPOSITION AND METABOLIC INHIBITORS

In springwater (25.5 mol m^–3 Cl^–, 20.4 mol m^–3Na⁺, 0.14 mol m^–3 K⁺) Enteromorpha intestinalis couldnot survive for more than a few weeks unless provided with 0.5mol m^–3 K⁺ in the medium or alternatively exposed to seawaterfor 1 day per week. Maintenance of a cytoplasmic K⁺ level ofabout 200 mol m^–3 is critical for the maintenance of normalmetabolic activity. Net gains of intracellular K⁺ occurred whenthe plants were transferred from low-salinity to seawater; converselylarge net losses occurred when plants were transferred fromseawater to springwater. These two processes were not simplythe reverse of one another; net gain of K⁺ involved a largeincrease in the tracer flux both into and out of the cell butnet loss of K⁺ virtually halted the tracer flux into the cell.Any injury incurred by rapid salinity changes was short-lived;plants were rapidly able to adjust intracellular [K₁.K⁺). K⁺(orto some extent Rb⁺) was found to be necessary in the effluxmedium for ⁴²K⁺ exchange to occur. The osmotic concentrationof the medium was also important but extracellular Na⁺ and Cl^–concentrationswere not critical. K⁺ influx and efflux in both springwaterand seawater were largely independent of light and were sensitivein varying degrees to a range of common metabolic inhibitorsand uncouplers. The results are best explained by the presenceof an active K⁺ influx, generated by an ATP-dependent K⁺ pumpat the plasmalemma. Key words: Enteromorpha, Potassium transport, Salinity changes, Uncouplers, Inhibitors     

Potassium Transport in Enteromorpha intestinalis (L.) Link: MEASUREMENT OF INTRACELLULAR K+, EXCHANGE FLUXES AND THERMODYNAMIC ANALYSIS

Potassium transport has been studied in the marine euryhalinealga, Enteromorpha intestimlis cultured in seawater and in low-salinitymedium (Artificial Cape Banks Spring Water, ACBSW; 25·5mol m^–3 Cl^–, 20·4 mol m^–3 Na⁺, 0·5mol m^–3 K⁺). K⁺ fluxes were measured using ⁴²K⁺ and ⁸⁶Rb⁺although ⁸⁶Rb⁺ does not act as an efficient K⁺ analogue in thisplant. ⁴²K⁺ experiments on seawater plants typically exhibiteda single protoplasmic exchange phase whereas ⁸⁶Rb⁺ exhibitedtwo exchange phases. Compartmental analysis of ⁸⁶Rb⁺ effluxexperiments on seawater-grown Enteromorpha plants were usedto deduce the intracellular partition of K⁺ between the cytoplasm(279±38 mMolal) and vacuole (405±68 mMolal). Theplasmalemma K⁺ flux in plants in seawater was greater in thelight than in the dark (563±108 nmol m^–2 s^–1versus 389±66·7 nmol m^–2 s^–1). Inlow-salinity plants, separate cytoplasmic and vacuolar exchangephases were apparent. Analysis of ⁴²K⁺ efflux experiments onlow-salinity plants yielded a cytoplasmic K⁺ of 222±38mMolal and a vacuolar K⁺ of 82±11 mMolal. The plasmalemmaand tonoplast flux was 23±4·5 nmol m^–2 s^–1. The Nernst equation showed that, although K⁺ was close to electrochemicalequilibrium, active accumulation of K⁺ across the plasmalemmaoccurred in plants in seawater and ACBSW both in the light anddark. K⁺ was also actively transported inwards across the tonoplastin low-salinity plants. The electrochemical potential for K⁺across the plasmalemma ranged from 2·41±0·60kJ mol^–1 in plants grown in seawater in the light to 5·79±0·87kJ mol^–1 for plants in ACBSW in the light. Although K⁺is close to electrochemical equilibrium, the flux of K⁺ in plantsin both seawater and ACBSW media is high, hence the power consumptionof K⁺ transport is high. The permeability of K⁺ (P_K⁺) was significantlyhigher in the light than in the dark in plants in seawater (about7·0 versus 2·5 nm s^–1) but in plants inlow-salinity (ACBSW) medium the permeability was independentof light (about 12 nm s^–1). The energy requirements ofactive K⁺ transport by ATP-dependent pumps is discussed. Key words: Enteromorpha, Potassium transport, Ionic relations, Saltwater, Low salinity, Thermodynamics     

Host-constrained epidemiology of the fish tapeworm Ligula intestinalis (L.)

Investigations into the biology of the roach, Rutilus rutilus (L.), and Ligula intestinalis (L.) populations at Slapton Ley, Devon were carried out between October 1982 and December 1984, and additional data collected from the lake since 1977 have been re-analysed. The Ligula population exhibited some unusual features: a limited specificity, a persistently low abundance and a scarcity of multiple infections. The population dynamics of the roach were also unusual in that the individual growth rate was one of the fastest in Britain, the survival was very poor and the year classes tended to alternate in strength. Whether the unusual epidemiology of Ligula could be explained by the unusual population dynamics of the roach is investigated. The transmission period of Ligula to the roach was limited to their first few months of life, probably due to a limited period of feeding on copepods by the fry. This narrow transmission window was almost certainly the major factor that has prevented the abundance of Ligula plerocercoids attaining high levels in the fry or of increasing in subsequent years, and has resulted in their abundance being determined entirely by transmission events to the fry. The limited transmission period has also caused the life-cycles of the roach and Ligula populations to become synchronized. At other localities, the abundance of Ligula is typically high, and can have a significant effect on the mortality of the host population, but, with the low abundance at Slapton, Ligula-induced roach mortality was insignificant. The rapid growth of the roach fry was the most likely explanation for the limited period of feeding on copepods, and the large size, short life-span and low abundance of the roach probably also constrained the build-up of the Ligula population in the lake. The random frequency distribution and scarcity of multiple infections was not considered unusual in view of the very low plerocercoid abundance. The very low abundance of Ligula, the separation of the spawning of roach and rudd in time and space, and the scarcity of rudd may account for the absence of infection in this species. It was concluded that the Ligula population in Slapton Ley was being constrained by the atypical population dynamics of the roach, and not vice versa as theory predicts.     

Ion Exchange Fluxes of the Cell Walls of Enteromorpha intestinalis (L. ) Link (Ulvales, Chlorophyta)

The ²²Na⁺ and ³⁶CI^– exchange properties of the cell wallsof Enteromorpha intestinalis (L. ) Link in simple monovalentsalt systems have been shown to be similar to a ‘leaky’cation exchange membrane rather than a homogeneous membrane.The ion exchange properties of the cation and anion cell wallcontents are what would be expected of a cation exchange membranei. e. anion exchange is strongly dependent on the bathing electrolyteconcentration and becomes very slow in dilute salt. This wouldlead to the cell wall becoming a barrier to anions in dilutesalt. However, measurements of the anion flux across cell wallsin living and dead tissues show that anion exchange across cellwalls is facilitated by pores. The exchange kinetics of thebulk of the cell wall anions does not limit the anion flux acrosscell walls of this plant. It is concluded that the cell wallis not a critical limitation to plasmalemma fluxes of the livingplant and that unstirred layers are more important than cellwalls in the measurement of anion flux rates.     

The attachment of Enteromorpha zoospores to a bacterial biofilm assemblage

This study has investigated the relationship between bacterial biofilms and the attachment of zoospores of the green macroalga Enteromorpha. Zoospore attachment to glass slides was enhanced in the presence of a bacterial biofilm assemblage, and the number attaching increased with the number of bacteria present. Zoospores also attached to control surfaces, but at lower numbers; glass surfaces conditioned in autoclaved seawater had the same number of zoospores attached as new glass surfaces. The spatial relationship between bacterial cells and attached zoospores was quantified by image analysis. The hypothesis tested was that zoospores attached preferentially to, or in the very close vicinity of, bacterial cells. Spatial microscopic analysis showed that more bacteria were covered by zoospores than would be expected if zoospore attachment was a random process and zoospores appeared to attach to bacterial clusters. The most likely explanation is that zoospores are attracted to bacterial cells growing on surfaces and the presence of a bacterial biofilm enhances their settlement. The possibility is discussed that Enteromorpha zoospores respond to a chemical signal produced by bacteria, i.e. that there may be prokaryote‐eukaryote cell signalling.     

Transmission experiments with pike fry (Esox Indus L.) rhabdovirus

Experimental infection of fertilized pike eggs with 'red-disease' virus produced 100% mortality in the fry. This mortality was associated with a disease that had previously been described as hydrocephalus internus, indicating that 'red-disease' and hydrocephalus are different manifestations of the same disease. The name pike fry rhabdovirus disease (PFRD) is suggested for the disease complex, and the name pike fry rhabdovirus (PFR) for the causative agent. Exposure of PFR to a Wescodyne * solution containing 25 ppm of iodine resulted in an inactivation of at least 99–99% of viral activity within 30 sec. Experimental egg transmission of PFR could be interrupted by disinfecting the eggs in a Wescodyne solution, suggesting that the virus was located on the egg surface. Conclusive evidence of a naturally occurring vertical transmission in pike culture is still lacking because, using FHM cells as a detection system, PFR could not be found in spawners and their sexual products. The susceptibility of pike fry to PFR rapidly decreases at increasing age.     

Chemical and physical-chemical characteristics of dietary fibres from Ulva lactuca (L.) Thuret and Enteromorpha compressa (L.) Grev.

Dietary fibres from Ulva lactuca (L.) Thuret (sea lettuce) and Enteromorpha compressa (L.) Grev. (A.O. nori) were measured according to a ‘standard’ method and a ‘physiological’ protocol simulating the gastric and intestinal environments. U. lactuca contained 15.8–8.0% soluble and 24.2–32.6% insoluble fibres according to the ‘standard’ and ‘physiological’ methods, respectively. For E. compressa, these values were 14.9–15.9 and 21.6–28.7%, respectively. For both algae, the composition suggests that the soluble fibres were xylorhamnoglycuronans sulphates and insoluble fibres were essentially composed of glucans. No marked chemical compositional variation was observed between soluble fractions extracted under the simulated gastric and intestinal conditions. Fibres in both algae are hydrophilic but the water holding capacities were higher after extraction of soluble fibres (5.5–9.5 g g⁻¹ for the dry algae; 14.0–16.0 g g⁻¹ for the standard insoluble fibres). Water soluble fibres demonstrated low intrinsic viscosities at 37 °C in buffers, particularly those from E. compressa (36.0–36.5 ml g⁻¹), and was affected by pH for those of U. lactuca (147.5 ml g⁻¹ at pH 3.0 and 175.0 ml g⁻¹ at pH 7.3).     

Phenotypic modification of roach (Rutilus rutilus L.) infected with Ligula intestinalis L. (Cestoda: Pseudophyllidea).

In European freshwater, cyprinid fish may be heavily infected by plerocercoids of the pseudophyllidea cestode Ligula intestinalis (L.). During their development, these parasites grow rapidly to a large size in the fish's body cavity, characteristically distending the abdomen. In this study, the influence of this tapeworm on roach (Rutilus rutilus L.) morphology was analyzed. Forty-five infected and 45 uninfected roach were collected from the Lavernose-Lacasse gravel pit in Toulouse, south western France and examined for 40 morphological measurements to study phenotypic modification of the body and 14 bilateral characters for an analysis of asymmetry. Results indicate that the degree of bilateral asymmetry does not change between infected and uninfected roach, despite the strong host-morphological modifications such as deformation of the abdomen, fin displacements at the level of the tail, and sagging of the vertebral column. The intensity of abdominal distension and fish morphology changes depends on the total parasite biomass present. Differences were observed in morphology at different levels of infection, which relate to established effects of L. intestinalis on the physiology and behavior of intermediate hosts. These morphological changes induced by the parasite could increase trophic transmission to the definitive avian hosts.     

Some competition experiments with alang-alang (Imperata cylindrica (L.) Beauv.) in replacement series

Summary The interaction between alang-alang (Imperata cylindrica) and maize or sorghum was studied in competition experiments according to the replacement principle. Dry matter yield of alang-alang in these experiments appeared hardly affected by the presence of maize or sorghum, while this yield of the latter two was strongly reduced by the presence of alang-alang. The relative yield total (RYT) reached unity except in one experiment in which a value of 0.6 was obtained.The results suggest that the allelopathic activity of alang-alang will find expression in an RYT deviating from one only if alang-alang is not able to utilize all available space.Part of the research supported by the Netherlands Foundation for Advancement of Tropical Research (WOTRO) grant W86-34     

Antitumor and immunomodulating activity of polysaccharides from Enteromorpha intestinalis

Two polysaccharides (WEA and WEB) were isolated from Enteromorpha intestinalis by hot water extraction, anion-exchange, and gel-permeation chromatography. The average molecular weights (Mw) of the two fractions were 72.03 kDa (WEA) and 60.12 kDa (WEB). WEA was composed of Rha, Xyl, Man, and Glc in a molar ratio of 1.39:1.00:0.13:3.23. WEB consisted of Rha, Xyl, Gal, and GlcA (glucuronic acid) in a molar ratio of 7.32:1.00:0.51:1.28. Both polysaccharides could inhibit tumor growth in S180 tumor-bearing mice, and increased the relative spleen and thymus weight. They also increased the expression of tumor necrosis factor-alpha (TNF-α) in serum. WEA and WEB induced lymphocyte proliferation, increased the production of TNF-α in macrophages, and stimulated macrophages to produce nitric oxide dose-dependently through the up-regulation of inducible NO synthase activity. However, no direct cytotoxicity against Sarcoma 180 was investigated in vitro. These results indicate that the antitumor effects of these polysaccharides are associated with immunostimulation.     

Development of microsatellite markers in the green algae Enteromorpha intestinalis (Chlorophyta)

The fouling green algae Enteromorpha intestinalis is a cosmopolitan benthic species, which causes green tides in many coastal areas and is used as an indicator species for eutrophication in the Baltic Sea area. The life cycle of E. intestinalis alternates between two morphologically identical reproductive stages, a haploid gametophyte phase and a diploid sporophyte phase. However, it also reproduces through asexual propagation. The reproductive cycles of E. intestinalis in the Baltic Sea and elsewhere are largely unknown. Here we report five polymorphic microsatellite markers developed from enriched genomic libraries. The number of alleles per locus ranged from 7 to 25.