Antisettlement defence of Fucus vesiculosus Chemistry and Ecology

Summary Bacteria are omnipresent in the marine environment. For example, in the Kiel Fjord (Baltic Sea) 0.7 to 2.24 x 106 bacterial cells are present in one ml of seawater. Marine organisms are thus continuously exposed to high densities of bacteria, some of which tend to settle and colonise living surfaces. Some associations between host and bacteria could be neutral or mutualistic, for example algae associated bacteria may protect their host from fouling. But mostly, bacterial epibionts have a number of negative impacts on their hosts, such as providing positive settlement cues to macrofoulers. Although macroalgae constitute a potential substratum for surface colonisation, many of them remain largely free from heavy fouling, presumably due to surface properties or release of antifouling chemicals. In the Baltic Sea this has also been observed in the ecologically important alga Fucus vesiculosus (Phaeophyceae), which remains free from heavy fouling during most of the year. So far there was no indication of a mechanical or structural defence against epibionts in this alga, which suggested that one or several chemical defence mechanisms may exist. The chemical defence of macroalgae has been a theme of literature over two decades. But so far, most of these effects have been demonstrated in studies investigating total tissue extracts instead of testing ecologically relevant surface extracts or surface metabolites at natural concentrations, which actually affect the fouling organisms in nature. Further, only very few among the studies which so far tested surface based compounds at ecologically relevant concentrations targeted the first phase of fouling, i.e. bacterial settlement. It was a main aim of my work to study the effect of surface-associated metabolites on microbial settlement, using the rockweed Fucus vesiculosus as a model organism. Since the control of bacterial fouling on macroalgae is of substantial ecological importance, it is highly relevant to characterize the nature and dynamics of potential bacterial settlement inhibitors - the first line of defensive compounds against microfoulers. In F. vesiculosus, polyphenols have been suggested as fouling deterrents. However, a bioassay-guided structure elucidation of the alga’s metabolites deployed in defence against bacterial settlement has been realized for the first time in my study, and a role of polyphenols in antimicrobial defence of F. vesiculosus was not detected. Using different chromatographic techniques followed by bioassays, I was instead able to isolate and identify an active lipophilic metabolite: fucoxanthin (Chapter I). Subsequently, I proved its surface presence. This was followed by an investigation of the metabolic provenance through segregation of fucoxanthin originating from Fucus and fucoxanthin originating from surface associated diatoms. Further I quantified the compound on algal tips and on whole individuals, in order to know whether it is sufficiently concentrated on the algal surface to warrant an inhibitory effect on bacterial settlers. With the help of this comprehensive study, I was able to report a novel defence strategy of Fucus. In chapter II, I proceeded to investigate the active polar metabolites on Fucus surfaces and reported DMSP and proline to be the hydrophilic metabolites contributing to an inhibition of bacterial settlement. This was the first study reporting the role of DMSP and of the amino acid proline as algal antifoulants. The combined results of chapter I and II provides a picture of multiple chemical defence strategies of Fucus vesiculosus in an ecological context. In chapter III, I show how the algal defence based on these inhibitors may be affected by environmental factors/ shifts such as high temperature and low light. Based on the surface concentrations of the three settlement inhibitors the defence capacity of F. vesiculosus appears to be only moderately affected by potential stressors. Finally, I report on the seasonal and geographical variation of antibacterial defence of the alga (Chapter IV). The anti-settlement defence showed a temporal variation with a peak activity in late summer/ autumn and also showed a strong and consistent difference between sites throughout the year. In summary this thesis highlights the capability of the Baltic foundation species Fucus vesiculosus for chemical defence against microfoulers, as well as the natural defence variability with site and season and the alga’s responses toward simulated environmental conditions. The study thus provides numerous new insights into algae-bacteria interactions and their dynamics and it helps in better understanding of the alga’s reaction towards bacterial epibiosis in response to potential abiotic stressors

Minireview: algal natural compounds and extracts as antifoulants

Marine biofouling is a paramount phenomenon in the marine environment and causes serious problems to maritime industries worldwide. Marine algae are known to produce a wide variety of chemical compounds with antibacterial, antifungal, antialgal, and anti-macrofouling properties, inhibiting the settlement and growth of other marine fouling organisms. Significant investigations and progress have been made in this field in the last two decades and several antifouling extracts and compounds have been isolated from micro- and macroalgae. In this minireview, we have summarized and evaluated antifouling compounds isolated and identified from macroalgae and microalgae between January 2010 and June 2016. Future directions for their commercialization through metabolic engineering and industrial scale up have been discussed. Upon comparing biogeographical regions, investigations from Southeast Asian waters were found to be rather scarce. Thus, we have also discussed the need to conduct more chemical ecology based research in relatively less explored areas with high algal biodiversity like Southeast Asia

Large scale patterns of antimicrofouling defenses in the hard coral Pocillopora verrucosa in an environmental gradient along the Saudi Arabian coast of the Red Sea

Large scale patterns of ecologically relevant traits may help identify drivers of their variability and conditions beneficial or adverse to the expression of these traits. Antimicrofouling defenses in scleractinian corals regulate the establishment of the associated biofilm as well as the risks of infection. The Saudi Arabian Red Sea coast features a pronounced thermal and nutritional gradient including regions and seasons with potentially stressful conditions to corals. Assessing the patterns of antimicrofouling defenses across the Red Sea may hint at the susceptibility of corals to global change. We investigated microfouling pressure as well as the relative strength of 2 alternative antimicrofouling defenses (chemical antisettlement activity, mucus release) along the pronounced environmental gradient along the Saudi Arabian Red Sea coast in 2 successive years. Microfouling pressure was exceptionally low along most of the coast but sharply increased at the southernmost sites. Mucus release correlated with temperature. Chemical defense tended to anti-correlate with mucus release. As a result, the combined action of mucus release and chemical antimicrofouling defense seemed to warrant sufficient defense against microbes along the entire coast. In the future, however, we expect enhanced energetic strain on corals when warming and/or eutrophication lead to higher bacterial fouling pressure and a shift towards putatively more costly defense by mucus releas

Defence Chemistry Modulation by Light and Temperature Shifts and the Resulting Effects on Associated Epibacteria of Fucus vesiculosus

The goals of this study were (1) to investigate whether Fucus vesiculosus regulates the production of its antifouling defence chemicals against epibacteria in response to light limitation and temperature shifts and (2) to investigate if different surface concentrations of defence compounds shape epibacterial communities. F. vesiculosus was incubated in indoor mesocosms at five different temperature conditions (5 to 25°C) and in outdoor mesocosms under six differently reduced sunlight conditions (0 to 100%), respectively. Algal surface concentrations of previously identified antifouling compounds - dimethylsulphopropionate (DMSP), fucoxanthin and proline – were determined and the bacterial community composition was characterized by in-depth sequencing of the 16S-rRNA gene. Altogether, the effect of different treatment levels upon defence compound concentrations was limited. Under all conditions DMSP alone appeared to be sufficiently concentrated to warrant for at least a partial inhibitory action against epibiotic bacteria of F. vesiculosus. In contrast, proline and fucoxanthin rarely reached the necessary concentration ranges for self-contained inhibition. Nonetheless, in both experiments along with the direct influence of temperature and light, all three compounds apparently affected the overall bacterial community composition associated with F. vesiculosus since tendencies for insensitivity towards all three compounds were observed among bacterial taxa that typically dominate those communities. Given that the concentrations of at least one of the compounds (in most cases DMSP) were always high enough to inhibit bacterial settlement, we conclude that the capacity of F. vesiculosus for such defence will hardly be compromised by shading or warming to temperatures up to 25°C

Industrial Digitization, the Use of Real-Time Information, and Operational Agility : Digital and Information Perspectives for Supply Chain Resilience

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Polaribacter septentrionalilitoris sp. nov., isolated from the biofilm of a stone from the North Sea

A new member of the family Flavobacteriaceae was isolated from the biofilm of a stone at Nordstrand, a peninsula at the German North Sea shore. Phylogenetic analysis of the 16S rRNA gene sequence showed that strain ANORD1T was most closely related to the validly described type strains Polaribacter porphyrae LNM-20T (97.0 %) and Polaribacter reichenbachii KMM 6386T (96.9 % 16S rRNA gene sequence similarity) and clustered with Polaribacter gangjinensis K17-16T (96.0 %). Strain ANORD1T was determined to be mesophilic, Gram-negative, non-motile and strictly aerobic. Optimal growth was observed at 20–30 °C, within a salinity range of 2–7 % sea salt and from pH 7–10. Like other type strains of the genus Polaribacter, ANORD1T was tested negative for flexirubin-type pigments, while carotenoid-type pigments were detected. The DNA G+C content of strain ANORD1T was 30.6 mol%. The sole respiratory quinone detected was menaquinone 6 (MK-6). The major fatty acids identified were C15 : 0, iso-C15 : 0, C15 : 1 ω6c and iso-C15 : 0 3-OH. Based on the polyphasic approach, strain ANORD1T represents a novel species in the genus Polaribacter, with the name Polaribacter septentrionalilitoris sp. nov. being proposed. The type strain is ANORD1T (=DSM 110039T=NCIMB 15081T=MTCC 12685T)

Salinity and time can alter epibacterial communities of an invasive seaweed

The establishment of epibacterial communities is fundamental to seaweed health, and fitness, in modulating ecological interactions and may also facilitate adaptation to new environments. Few recent studies have investigated the influence of abiotic factors like light, temperature as drivers of epibacterial community composition on seaweeds. Although salinity can determine bacterial abundance, growth and community composition, influence of salinity as a driver of epibacterial community composition (until species level) has not been investigated for seaweeds and especially under long time scales. We also do not know how abiotic stressors may influence the ‘core’ bacterial species of seaweeds. Following an initial (immediately after field collection, T0) sampling of epibacterial community of an invasive red seaweed Agarophyton vermicullophylum, we conducted a long term mesocosm experiment for 5 months, to examine the influence of three different salinities (low, medium and high) at two different time points (T1, T2) on the epibacterial community richness and composition of Agarophyton. Metagenomic sequencing showed that epibacterial communities changed significantly according to salinity and time points sampled. Epibacterial richness was significantly different between low and high salinities at both time points. Epibacterial richness also varied significantly among T1 and T2 within low, medium and high salinity level. Irrespective of salinity levels and time points sampled 727 taxa consistently appeared in all Agarophyton samples hinting at the presence of core bacterial species on the surface of the alga. Our results indicate that both salinity and time can be major driving forces in structuring epibacterial communities of seaweeds with respect to richness and β diversity. We highlight the necessity of conducting long term experiments allowing us to detect and understand epibacterial succession over time on seaweeds

Response of foundation macrophytes to near‐natural simulated marine heatwaves

Marine heatwaves have been observed worldwide and are expected to increase in both frequency and intensity due to climate change. Such events may cause ecosystem reconfigurations arising from species range contraction or redistribution, with ecological, economic and social implications. Macrophytes such as the brown seaweed Fucus vesiculosus and the seagrass Zostera marina are foundation species in many coastal ecosystems of the temperate northern hemisphere. Hence, their response to extreme events can potentially determine the fate of associated ecosystems. Macrophyte functioning is intimately linked to the maintenance of photosynthesis, growth and reproduction, and resistance against pathogens, epibionts and grazers. We investigated morphological, physiological, pathological and chemical defence responses of western Baltic Sea F. vesiculosus and Z. marina populations to simulated near‐natural marine heatwaves. Along with (a) the control, which constituted no heatwave but natural stochastic temperature variability (0HW), two treatments were applied: (b) two late‐spring heatwaves (June, July) followed by a summer heatwave (August; 3HW) and (c) a summer heatwave only (1HW). The 3HW treatment was applied to test whether preconditioning events can modulate the potential sensitivity to the summer heatwave. Despite the variety of responses measured in both species, only Z. marina growth was impaired by the accumulative heat stress imposed by the 3HW treatment. Photosynthetic rate, however, remained high after the last heatwave indicating potential for recovery. Only epibacterial abundance was significantly affected in F. vesiculosus. Hence both macrophytes, and in particular F. vesiculosus, seem to be fairly tolerant to short‐term marine heatwaves at least at the intensities applied in this experiment (up to 5°C above mean temperature over a period of 9 days). This may partly be due to the fact that F. vesiculosus grows in a highly variable environment, and may have a high phenotypic plasticity

Geographic variation in fitness-related traits of the bladderwrack Fucus vesiculosus along the Baltic Sea-North Sea salinity gradient

In the course of the ongoing global intensification and diversification of human pressures, the study of variation patterns of biological traits along environmental gradients can provide relevant information on the performance of species under shifting conditions. The pronounced salinity gradient, co-occurrence of multiple stressors, and accelerated rates of change make the Baltic Sea and its transition to North Sea a suitable region for this type of study. Focusing on the bladderwrack Fucus vesiculosus, one of the main foundation species on hard-bottoms of the Baltic Sea, we analyzed the phenotypic variation among populations occurring along 2,000 km of coasts subjected to salinities from 4 to >30 and a variety of other stressors. Morphological and biochemical traits, including palatability for grazers, were recorded at 20 stations along the Baltic Sea and four stations in the North Sea. We evaluated in a common modeling framework the relative contribution of multiple environmental drivers to the observed trait patterns. Salinity was the main and, in some cases, the only environmental driver of the geographic trait variation in F. vesiculosus. The decrease in salinity from North Sea to Baltic Sea stations was accompanied by a decline in thallus size, photosynthetic pigments, and energy storage compounds, and affected the interaction of the alga with herbivores and epibiota. For some traits, drivers that vary locally such as wave exposure, light availability or nutrient enrichment were also important. The strong genetic population structure in this macroalgae might play a role in the generation and maintenance of phenotypic patterns across geographic scales. In light of our results, the desalination process projected for the Baltic Sea could have detrimental impacts on F. vesiculosus in areas close to its tolerance limit, affecting ecosystem functions such as habitat formation, primary production, and food supply.Peer reviewe