Effect of Bacteria Associated with the Green Alga Ulva reticulata on Marine Micro- and Macrofouling

The green alga Ulva reticulata (Forsskal) is often free from biofouling in Hong Kong waters. An early study indicated that bioactive substances from this alga inhibit settlement of the polychaete Hydroides elegans (Haswell). It is also predicted that epibiotic bacteria protect this alga from micro- and macrofouling. In this study, bacterial strains from the surface of U. reticulata were isolated and their inhibitive activities on micro- and macrofouling assayed. The strains were identified by 16S rRNA analysis as belonging to the genera Alteromonas , Pseudoalteromonas and Vibrio . There was no significant effect of these strains or their extracts (aqueous and ethanol) on the growth of five Vibrio strains isolated from natural biofilm. Two bacterial strains ( Alteromonas sp. and Vibrio sp. 3) were non-toxic to the benthic diatom Nitzschia paleacea (Grunow) while the other five strains caused a low level of mortality. No one bacterial strain was toxic to the larvae of H. elegans . Aqueous extract of one of the isolated bacterial species, i.e. Vibrio sp. 2, significantly ( p <0.00001) inhibited the settlement and metamorphosis of H. elegans larvae. The putative antifouling compounds have a molecular weight of >100 kD. On the other hand, biofilm of Pseudoalteromonas sp. 2 and aqueous extract of Vibrio sp. 2 suppressed the settlement of larvae induced by 3-isobutyl-1-methylxanthine (IBMX). Other epibiotic bacteria and their extracts had neither inhibitive nor inductive effects on larval settlement of H. elegans . The results indicate that the antifouling mechanism of U. reticulata may be dependent not only on materials from the macroalga itself but also on the epibiotic bacteria on the algal surface.     

Antifouling potential of Subtilisin A immobilized onto maleic anhydride copolymer thin films

The proteinaceous nature of the adhesives used by most fouling organisms to attach to surfaces suggests that coatings incorporating proteolytic enzymes may provide a technology for the control of biofouling. In the present article, the antifouling (AF) and fouling release potential of model coatings incorporating the surface-immobilized protease, Subtilisin A, have been investigated. The enzyme was covalently attached to maleic anhydride copolymer thin films; the characteristics of the bioactive coatings obtained were adjusted through variation of the type of copolymer and the concentration of the enzyme solution used for immobilization. The bioactive coatings were tested for their effect on the settlement and adhesion strength of two major fouling species: the green alga Ulva linza and the diatom Navicula perminuta. The results show that the immobilized enzyme effectively reduced the settlement and adhesion strength of zoospores of Ulva and the adhesion strength of Navicula cells. The AF efficacy of the bioactive coatings increased with increasing enzyme surface concentration and activity, and was found to be superior to the equivalent amount of enzyme in solution. The results provide a rigorous analysis of one approach to the use of immobilized proteases to reduce the adhesion of marine fouling organisms and are of interest to those investigating enzyme-containing coating technologies for practical biofouling control.     

A model that predicts the attachment behavior of Ulva linza zoospores on surface topography

A predictive model for the attachment of spores of the green alga Ulva on patterned topographical surfaces was developed using a constant refinement approach. This ‘attachment model’ incorporated two historical data sets and a modified version of the previously-described Engineered Roughness Index. Two sets of newly-designed surfaces were used to evaluate the effect of two components of the model on spore settlement. Spores attached in fewer numbers when the area fraction of feature tops increased or when the number of distinct features in the design increased, as predicted by the model. The model correctly predicted the spore attachment density on three previously-untested surfaces relative to a smooth surface. The two historical data sets and two new data sets showed high correlation (R ² = 0.88) with the model. This model may be useful for designing new antifouling topographies.     

Cryptic diversity of Ulva (Ulvales,Chlorophyta) in the Great Bay Estuarine System (Atlantic USA): introduced and indigenous distromatic species

Distromatic foliose blades of the algal genus Ulva are notoriously difficult to identify due to their simple morphologies and few diagnostic characteristics that often exhibit intraspecific variation and interspecific overlap. Hence, species differentiation is difficult and diversity estimates are often inaccurate. Two major goals of this study were to assess the diversity of distromatic Ulva spp. in the Great Bay Estuarine System (GBES) of New Hampshire and Maine, USA, and to compare historical and present day records of these species. Molecular analysis (using ITS sequences) of field-collected specimens revealed four distinct taxa: Ulva lactuca, U. rigida, U. compressa, and U. pertusa. Prior to molecular screening, Ulva lactuca was the only distromatic Ulva species reported for the GBES. Ulva pertusa and the foliose form of U. compressa are newly recorded for the Northwest Atlantic, and the range of U. rigida has been extended. Molecular analysis of historical herbarium voucher specimens indicates that U. rigida, U. pertusa, and the foliose form of U. compressa have been present in the GBES since at least 1966, 1967, and 1972, respectively. The distromatic morphotype of U. compressa is found only in low salinity areas, which suggests that salinity may influence its morphological development. Molecular and morphological evaluations are critical if we are to distinguish between cryptic taxa, accurately assess biodiversity, and effectively monitor the spread of non-indigenous macroalgae.     

Free-floating Ulva in the southwest Netherlands: species or morphotypes? a morphological,molecular and ecological comparison

Free-floating Ulva L. biomass in the eutrophic brackish ‘Veerse Meer’ lagoon (southwest Netherlands) consists of four morphologically identified species: U. curvata (Kützing) De Toni, U. lactuca L., U. rigida C. Agardh and U. scandinavica Bliding. U. curvata could be recognized easily because of the characteristic central cavity in the holdfast of the attached plants, the arrangement of cells in rows and the single pyrenoid in each cell. U. rigida was distinguished by the thick thallus and the large number of pyrenoids. The Veerse Meer isolate, however, was slightly different from the isolate from the Oosterschelde estuary (the Netherlands). U. lactuca and U. scandinavica showed a high degree of overlap in thallus thickness and cell size, but U. scandinavica usually possessed more pyrenoids. However, doubts have frequently been expressed about the use of some morphological characters in Ulva taxonomy. To determine the validity of such characters in the identification of Ulva species, the morphological variation within and between morphological species was recorded and a molecular data set generated. To detect possible ecophysiological differences between species, optimum temperatures and salinities for growth were determined experimentally. The sequences of the nuclear ribosomal DNA internal transcribed spacer 2 (ITS2) and flanking regions of U. lactuca, U. rigida and U. scandinavica from the Veerse Meer were all identical, but differed from that of U. rigida from the Oosterschelde estuary. Ulva species from the Veerse Meer were most closely related to U. armoricana and U. rigida from Brittany (2.9% and 3.5% divergence respectively); the difference between U. rigida from the Veerse Meer and from the Oosterschelde estuary was 7.5%. Rooted trees, based on a comparison of these sequences with sequences of other Ulva and Enteromorpha species obtained from the literature, using Monostroma arcticum as outgroup, suggested that Ulva is paraphyletic with respect to Enteromorpha. The optimum temperature for growth of U. curvata was 25?°C; for all other species it was 10?°C. The optimum salinity for growth was 30?°C for all isolates. It is concluded that U. lactuca, U rigida and U. scandinavica from the Veerse Meer are all members of one highly polymorphic species.     

REGULATION OF BIOSYNTHESIS OF DIMETHYLSULFONIOPROPIONATE AND ITS UPTAKE IN STERILE MUTANT OF ULVA PERTUSA (CHLOROPHYTA)1

It has been shown that marine algae produce the compatible solute dimethylsulfoniopropionate (DMSP) from methionine (Met) via four enzymatic reactions in which the third step, synthesis of 4‐dimethylsulfonio‐2‐hydroxy‐butyrate (DMSHB) from 4‐methylthio‐2‐hydroxybutyrate (MTHB), is the committing step. However, regulation of the biosynthetic pathways and transport properties of DMSP is largely unknown. Here, the effects of sulfur and sodium concentrations on the uptake and synthesis of DMSHB and DMSP were examined in a sterile mutant of Ulva pertusa Kjellm. Sulfur deficiency increased the activity of the sulfur assimilation enzyme O‐acetyl serine sulfhydrylase but decreased the MTHB S‐methyltransferase activity, suggesting the preferential utilization of sulfur atoms for Met metabolites other than DMSP. Uptake of DMSP and DMSHB was enhanced by S deficiency. High salinity enhanced the MTHB S‐methyltransferase activity as well as the uptake of DMSHB. The MTHB S‐methyltransferase activity was inhibited by its product DMSP. These data demonstrate the importance of MTHB S‐methyltransferase activity and uptake of DMSHB for the regulation of DMSP.     

连续性干出对孔石莼生长和生化组成的影响

以孔石莼(Ulva pertusa)为试验材料,研究了连续性干出对其生长和生化组成的影响.结果表明:与对照组(0 h)相比,短时间的连续性干出促进孔石莼生长,表现在特定生长率( SGR)升高、叶绿素a含量上升;随着连续性干出时间延长,孔石莼SGR、叶绿素a、蛋白质含量下降,叶绿素b、可溶性糖含量上升;叶绿素a/b比值变化和可溶性糖含量变化有利于抵抗干出胁迫.     

Eyespot behavior during the fertilization of gametes in Ulva arasakii Chihara (Ulvophyceae, Chlorophyta)

Behavior of the eyespots during the fertilization of Ulva arasakii Chihara was studied using field emission scanning electron microscopy (FE‐SEM). FE‐SEM enabled the visualization of the eyespot of biflagellate male and female gametes. The smaller male gamete has one protruded smaller (1.3 ± 0.15 μm× 1.0 ± 0.29 μm) eyespot and the larger female gamete has a larger (1.6 ± 0.2 μm× 1.1 ± 0.13 μm) one on a posterior position of the cell. The cell membrane over the eyespot region is relatively smooth compared to other parts of the cell body and exhibits hexagonal arranged lipid globules. Because the size of the cell and the morphology of the eyespot are different between male and female gametes, we could follow the fate of the eyespots during the fertilization. The initial cytoplasmic contact and fusion of the gametes takes place at their anterior end, slightly posterior to the flagellar base. The morphology of the fusing gametes followed two clearly distinguishable patterns. About half the gamete pairs lie side‐by‐side with their longitudinal axes nearly parallel, while the rest are oriented anti‐parallel to each other. In all cases, the larger female gamete fused along the same side as the eyespot, while the smaller male gamete fused along the side away from its eyespot. As fusion proceeds, the gamete pair is transformed into the quadriflagellate planozygote, in which the eyespots are positioned side‐by‐side on the region of cell fusion. These observations indicated that the opposite positioning of the eyespot relative to the cell fusion site in male and female gametes is important for the proper arrangement of the eyespots in the planozygote. The significance of this feature in advanced green algae is briefly discussed.     

Conditioning of self-assembled monolayers at two static immersion test sites along the east coast of Florida and its effect on early fouling development

Among the first events after immersion of surfaces in the ocean is surface ‘conditioning’. Here, the accumulation and composition of the conditioning films formed after immersion in the ocean are analyzed. In order to account for different surface chemistries, five self-assembled monolayers that differ in resistance to microfouling and wettability were used. Water samples from two static immersion test sites along the east coast of Florida were collected at two different times of the year and used for experiments. Spectral ellipsometry revealed that conditioning films were formed within the first 24 h and contact angle goniometry showed that these films changed the wettability and rendered hydrophobic surfaces more hydrophilic and vice versa. Infrared reflection adsorption spectroscopy showed that the composition of the conditioning film depended on both the wettability and immersion site. Laboratory and field assays showed that the presence of a conditioning film did not markedly influence settlement of microorganisms.