Genotypic variation in tolerance and resistance to fouling in the brown alga <Emphasis Type="Italic">Fucus vesiculosus</Emphasis>

In this study, we examined genetic variation in resistance and tolerance to fouling organisms in the brown alga Fucus vesiculosus. We first grew 30 algal genotypes in the field, where we allowed fouling organisms to colonise the genotypes at natural levels. We then conducted a manipulative experiment, where we grew 20 genotypes of algae in aquaria with or without fouling organisms. We measured host resistance as the load of fouling organisms and tolerance as the slope of the regression of algal performance on fouling level. Fouling organisms decreased host growth and contents of phlorotannins and thus have the potential to act as selective agents on algal defenses. We found significant among-genotype variation in both resistance and tolerance to fouling. We did not find a trade-off between resistance and tolerance. We found a marginally significant cost of resistance, but no cost of tolerance. Our results thus indicate that both the tolerance and resistance of F. vesiculosus can evolve as a response to fouling and that the costs of resistance may maintain genetic variation in resistance.     

Responses of growth and phlorotannins in <Emphasis Type="Italic">Fucus vesiculosus</Emphasis> to nutrient enrichment and herbivory

Discharge from anthropogenic sources may modify both macroalgal growth patterns and resource allocation to carbon based secondary compounds, thereby affecting their susceptibility for herbivory. We tested the effect of eutrophication in terms of nutrient enhancement on growth and phlorotannin concentration of Fucus vesiculosus by conducting manipulative experiments in the field and mesocosms. In the field experiment we utilised fish farms as nutrient sources and in the mesocosm-experiment we manipulated ambient nutrient levels and occurrence of the herbivorous isopod Idotea baltica. Vicinity of a fish farm affected neither growth nor the phlorotannin concentration of Fucus but increased the amount of epiphytes growing on Fucus. Other organisms such as epiphytic filamentous algae and periphyton, which are more capable of quickly utilizing excess nutrients, may restrain the direct effects of nutrient enhancement on Fucus. In a manipulative mesocosm experiment, neither nutrient enrichment nor occurrence of herbivores affected phlorotannin concentration implying lack of induced defences, at least in terms of increasing phlorotannin concentration. Feeding of thallus decreased the growth rate of algae, but the number of reproductive organs, receptacles, was not affected by herbivory. The negative effect of herbivory on the amount of apical tips tended to be stronger under nutrient enriched conditions. We conclude that eutrophication processes, in terms of nutrient enrichment, does not have strong direct effect on growth or phlorotannin production of F. vesiculosus. However, there may be important indirect consequences. First, herbivory may be targeted more to apical parts of the thallus under eutrophicated conditions. Second, the result that Fucus growing close to nutrient sources were smaller than those in control areas may reflect differences in mortality schedules of algae between eutrophicated and control areas.     

Human dignity, bioethics, and human rights

The authors analyse and assess the Universal Draft Declaration on Bioethics and Human Rights published by UNESCO. They argue that the Draft has two main weaknesses. It unnecessarily confines the scope of bioethics to life sciences and their practical applications. And it fails to spell out the intended role of human dignity in international ethical regulation.     

Ultrasound stimulates proteoglycan synthesis in bovine primary chondrocytes

Mechanical forces can stimulate the production of extracellular matrix molecules. We tested the efficacy of ultrasound to increase proteoglycan synthesis in bovine primary chondrocytes. The ultrasound-induced temperature rise was measured and its contribution to the synthesis was investigated using bare heat stimulus. Chondrocytes from five cellular isolations were exposed in triplicate to ultrasound (1 MHz, duty cycle 20%, pulse repetition frequency 1 kHz) at average intensity of 580 mW/cm2 for 10 minutes daily for 1-5 days. Temperature evolution was recorded during the sonication and corresponding temperature history was created using a controllable water bath. This exposure profile was used in 10-minute-long heat treatments of chondrocytes. Heat shock protein 70 (Hsp70) levels after one-time treatment to ultrasound and heat was analyzed by Western blotting, and proteoglycan synthesis was evaluated by 35S-sulfate incorporation. Ultrasound treatment did not induce Hsp70, while heat treatment caused a slight heat stress response. Proteoglycan synthesis was increased approximately 2-fold after 3-4 daily ultrasound stimulations, and remained at that level until day 5 in responsive cell isolates. However, chondrocytes from one donor cell isolation out of five remained non-responsive. Heat treatment alone did not increase proteoglycan synthesis. In conclusion, our study confirms that pulsed ultrasound stimulation can induce proteoglycan synthesis in chondrocytes.     

The proteomics of feather development in pied flycatchers (Ficedula hypoleuca) with different plumage coloration

The genetic theory of morphological evolution postulates that form evolves largely by changing the expression proteins that are functionally conserved. It follows that understanding the function of proteins during different phases of development as well as the mechanisms by which the functions are modified is a prerequisite for understanding evolutionary change. Male pied flycatchers exhibit marked phenotypic variation in their breeding plumage. This variation has repeatedly been shown to have adaptive significance, but the molecular basis of this variation is not known. Here, we characterize the proteome of developing pied flycatcher feathers from differently pigmented males and also introduce a new method for examining the effect sizes of expression differences in protein interaction networks. Approximately 300 proteins were identified in the developing feathers of males. Gene products associated with cellular transport, cell metabolism and protein synthesis formed a large part of the developing feather proteome. Sixty‐five proteins associated with the development of the epidermis and/or pigmentation were detected in the data. The examination of expression level differences of protein–protein interaction networks revealed an immunological signalling–related network to exhibit significantly higher expression in black compared to brown males. Additionally, indications of differences in energy balance and oxidative stress related characteristics were detected. Together, these results provide new insight into the molecular mechanisms and evolutionary significance of plumage colour variation.     

Automated dead-end ultrafiltration of large volume water samples to enable detection of low-level targets and reduce sample variability

Aims

A Portable Multi‐use Automated Concentration System (PMACS) concentrates micro‐organisms from large volumes of water through automated dead‐end ultrafiltration and backflushing. The ability to detect microbial targets from ground, surface and cooling tower waters collected using standard methods was compared with samples from the PMACS in this study.

Methods and Results

PMACS (100 l) and standard grab samples (100–500 ml) were collected from sites in Florida and South Carolina, USA. Samples were analysed for the presence of faecal indicator bacteria (FIB; ground and surface water) or Legionella pneumophila (Lp; cooling tower water). FIB were enumerated by growth on selective media following membrane filtration or in IDEXX defined substrate media. Lp cells were detected by direct fluorescence immunoassay using FITC‐labelled monoclonal antibodies targeting serogroups 1, 2, 4 and 6. FIB were found in PMACS samples from ground and surface waters when their concentrations were below detection limits in grab samples. The concentrations of Lp in cooling tower samples collected over 5 months were more consistent in PMACS samples than grab samples.

Conclusions

These data demonstrate that PMACS concentration is advantageous for water monitoring. FIB were detected in PMACS samples when their concentrations were below the detection limits of the standard methods used. PMACS processing provided more representative samples of cooling tower waters reducing sample variability during long‐term monitoring.

Significance and Impact of the Study

This study highlights the utility of PMACS processing for enhanced monitoring of water for low‐level microbial targets and for reducing sample variability in long‐term monitoring programmes.     

Fouling mediates grazing: intertwining of resistances to multiple enemies in the brown alga Fucus vesiculosus

Macroalgae have to cope with multiple natural enemies, such as herbivores and epibionts. As these are harmful for the host, the host is expected to show resistance to them. Evolution of resistance is complicated by the interactions among the enemies and the genetic correlations among resistances to different enemies. Here, we explored genetic variation in resistance to epibiosis and herbivory in the brown alga Fucus vesiculosus, both under conditions where the enemies coexisted and where they were isolated. F. vesiculosus showed substantial genetic variation in the resistance to both epibiosis and grazing. Grazing pressure on the alga was generally lower in the presence than in the absence of epibiota. Furthermore, epibiosis modified the susceptibility of different algal genotypes to grazing. Resistances to epibiosis and grazing were independent when measured separately for both enemies but positively correlated when both these enemies coexisted. Thus, when the enemies coexisted, the fate of genotypes with respect to these enemies was intertwined. Genotypic correlation between phlorotannins, brown-algal phenolic secondary metabolites, and the amount of epibiota was negative, indicating that these compounds contribute to resistance to epibiosis. In addition, phlorotannins correlated also with the resistance to grazing, but this correlation disappeared when grazing occurred in the absence of epibiota. This indicates that the patterns of selection for the type of the resistance as well as for the resistance traits vary with the occurrence patterns of the enemies.     

Structural insights into steroid hormone binding: the crystal structure of a recombinant anti-testosterone Fab fragment in free and testosterone-bound forms.

The monoclonal anti-testosterone antibody (3-C(4)F(5)) has a relatively high affinity (3 x 10(8) m(-1)) with an overall good specificity profile. However, the earlier characterized binding properties have shown that both the affinity and specificity of this antibody must be improved if it is intended for use in clinical immunoassays. In this paper, the crystal structures of the recombinant anti-testosterone (3-C(4)F(5)) Fab fragment have been determined in the testosterone-bound and free form at resolutions of 2.60 and 2.72 A, respectively. The high affinity binding of the (3-C(4)F(5)) Fab is mainly determined by shape complementarity between the protein and testosterone. Only one direct hydrogen bond is formed between the hydroxyl group of the testosterone D-ring and the main-chain oxygen of Gly100(J)H. The testosterone is deeply bound in a hydrophobic pocket, and the close shape complementarity is mainly formed by the third complementarity-determining regions (CDR) of the heavy and light chain. Comparison of the bound structure with the free structure indicates conformational changes in the protein upon testosterone binding. The conformational changes of the side chains of two residues Glu95H and Tyr99H in the CDR-H3 are particularly essential for the binding. Interesting similarities in the binding of different steroids were also observed upon comparison of the available structures of anti-steroid antibodies.     

Bloom forming Alexandrium ostenfeldii (Dinophyceae) in shallow waters of the Åland Archipelago,Northern Baltic Sea

In the past years, late summer blooms of the bioluminescent dinoflagellate Alexandrium ostenfeldii have become a recurrent phenomenon in coastal waters of the central and Northern Baltic Sea. This paper reports exceptionally high cell concentrations (10⁵ to 10⁶ cells L^?1) of the species found during bioluminescent blooms in 2003 and 2004 in a shallow embayment of the Åland archipelago at the SW coast of Finland. Clonal cultures were established for morphological, molecular, toxicological and ecophysiological investigations to characterize the Finnish populations and compare them to other global A. ostenfeldii isolates. The Finnish isolates exhibited typical morphological features of A. ostenfeldii such as large size, a prominent ventral pore and an orthogonally bent first apical plate. However, unambiguous differentiation from closely related Alexandrium peruvianum was difficult due to considerable variation of sulcal anterior plate shapes. The Finnish strains were genetically distinct from other isolates of the species, but phylogenetic analyses revealed a close relationship to isolates from southern England and an A. peruvianum morphotype from the Spanish Mediterranean. Together these isolates formed a distinct clade which was separated from a clade containing other Northern European, North American and New Zealand populations. Toxin analyses confirmed the presence of the PSP toxins GTX2, GTX3 and STX in both Finnish isolates with GTX3 being the dominant toxin. Total relative PSP toxin contents were moderate, ranging from approximately 6 to 15 fmol cell^?1 at local salinities of 5 and 10 psu, respectively. Spirolides were not detected. Salinity tolerance experiments showed that the Finnish isolates were well adapted to grow at the low salinities of the Baltic Sea. With a salinity range of approximately 6 to 20–25 psu, Baltic populations are physiologically distinct from their marine relatives. Vigorous production of different cyst types in the cultures suggest that cysts may play a crucial role in the survival and retainment of A. ostenfeldii populations in the Baltic Sea.