Marine Processes and Biotechnology

The bacterial communities associated with healthy and diseased colonies of the cold-water gorgonian coral Eunicella verrucosa at three sites off the south-west coast of England were compared using denaturing gradient gel electrophoresis (DGGE) and clone libraries. Significant differences in community structure between healthy and diseased samples were discovered, as were differences in the level of disturbance to these communities at each site; this correlated with depth and sediment load. The majority of cloned sequences from healthy coral tissue affiliated with the Gammaproteobacteria. The stability of the bacterial community and dominance of specific genera found across visibly healthy colonies suggest the presence of a specific microbial community. Affiliations included a high proportion of Endozoicomonas sequences, which were most similar to sequences found in tropical corals. This genus has been found in a number of invertebrates and is suggested to have a role in coral health...

Sediments play a key role in the marine nitrogen cycle and can act either as a source or a sink of biologically available (fixed) nitrogen. This cycling is driven by a number of microbial remineralization reactions, many of which occur across the oxic/anoxic interface near the sediment surface. The presence and activity of large burrowing macrofauna (bioturbators) in the sediment can significantly affect these microbial processes by altering the physicochemical properties of the sediment. For example, the building and irrigation of burrows by bioturbators introduces fresh oxygenated water into deeper sediment layers and allows the exchange of solutes between the sediment and water column. Burrows can effectively extend the oxic/anoxic interface into deeper sediment layers, thus providing a unique environment for nitrogen-cycling microbial communities. Recent studies have shown that the abundance and diversity of micro-organisms can be far greater in burrow wall sediment than in the surrounding surface or subsurface sediment; meanwhile, bioturbated sediment supports higher rates of coupled nitrification-denitrification reactions and increased fluxes of ammonium to the water column. In the present paper we discuss the potential for bioturbation to significantly affect marine nitrogen cycling, as well as the molecular techniques used to study microbial nitrogen cycling communities and directions for future study.

Ulva zoospores preferentially settle on N-acylhomoserine lactone (AHL) producing marine bacterial biofilms. To investigate whether AHL signal molecules also affect the success and rate of zoospore germination in addition to zoospore attraction, the epiphytic bacteria associated with mature Ulva linza were characterized and bacterial isolates representative of this community tested for the ability to produce AHLs. Two of these AHL-producing isolates, Sulfitobacter spp. 376 and Shewanella spp. 79, were transformed with plasmids expressing the Bacillus spp. AHL lactonase gene aiiA to generate AHL-deficient variants. The germination and growth of U. linza zoospores was studied in the presence of these AHL-deficient strains and their AHL-producing counterparts. This revealed that the AHLs produced by Sulfitobacter spp. and Shewanella spp. or the bacterial products they regulate have a negative impact on both zoospore germination and the early growth of the Ulva germling. Further experiments with Escherichia coli biofilms expressing recombinant AHL synthases and synthetic AHLs provide data to demonstrate that zoospores germinated and grown in the absence of AHLs were significantly longer than those germinated in the presence of AHLs. These results reveal an additional role for AHLs per se in the interactive relationships between marine bacteria and Ulva zoospores.

Zoospores of the eukaryotic green seaweed Ulva respond to bacterial N -acylhomoserine lactone (AHL) quorum sensing signal molecules for the selection of surface sites for permanent attachment. In this study we have investigated the production and destruction of AHLs in biofilms of the AHL-producing marine bacterium, Vibrio anguillarum and their stability in seawater. While wild type V. anguillarum NB10 was a strong attractor of zoospores, inactivation of AHL production in this strain by either expressing the recombinant Bacillus lactonase coding gene aiiA , or by mutating the AHL biosynthetic genes, resulted in the abolition of zoospore attraction. In seawater, with a pH of 8.2, the degradation of AHL molecules was temperature-dependent, indicating that the AHLs produced by marine bacterial biofilms have short halflives. The Ulva zoospores sensed a range of different AHL molecules and in particular more zoospores settled on surfaces releasing AHLs with longer ( > six carbons) N -linked acyl chains. However, this finding is likely to be influenced by the differential diffusion rates of AHLs from the experimental surface matrix. Molecules with longer N -acyl chains, such as N-(3oxodecanoyl)-L -homoserine lactone, diffused more slowly than those with shorter N -acyl chains such as N-(3-hydroxy-hexanoyl)-L -homoserine lactone. Image analysis using GFP-tagged V. anguillarum biofilms revealed that spores settle directly on bacterial cells and in particular on microcolonies which we show are sites of concentrated AHL production.

Several studies suggest that coral disease may be attributed to the success of vibrios in out-competing other bacteria in the mucus and tissues of corals. Vibrios utilize a variety of quorum sensing (QS) signal molecules to regulate processes that could be used to colonize corals during adverse environmental conditions. We therefore screened a range of Vibrios isolated from a variety of healthy and diseased corals, for the production of the QS signal molecules, N-acylhomoserine lactones (AHLs) and the AI-2 (autoinducer-2) small furanone signal molecule. All 29 strains examined activated the AI-2 biosensor, but only 17 activated an AHL biosensor. Using reverse phase thin-layer chromatography, we showed that the effect of temperature on AHL production varied considerably among the isolates. For the first time, the QS inhibition by Vibrio harveyi is reported. This only occurred at higher temperatures and does not appear to be due to degradation of AHLs. The large diversity of vibrios and the different effects of temperature on signal production may partly explain the complexity of coral-associated community changes in response to environmental factors.

Arctic surface sediment was analysed in detail using 16S rRNA 454 pyrosequencing. Intact sediment cores were collected and exposed to one of five different pCO2 concentrations [380 (present day), 540, 750, 1120 and 3000 μatm] and RNA extracted after a period of 14 days exposure. Measurements of diversity and multivariate similarity indicated very little difference between pCO2 treatments. Only when the highest and lowest pCO2 treatments were compared were significant differences evident, namely increases in the abundance of operational taxonomic units most closely related to the Halobacteria and differences to the presence/absence structure of the Planctomycetes. The relative abundance of members of the classes Planctomycetacia and Nitrospira increased with increasing pCO2 concentration, indicating that these groups may be able to take advantage of changing pH or pCO2 conditions. The modest response of the active microbial communities associated with these sediments may be due to the low and fluctuating pore-water pH already experienced by sediment microbes, a result of the pH buffering capacity of marine sediments, or due to currently unknown factors. Further research is required to fully understand the impact of elevated CO2 on sediment physicochemical parameters, biogeochemistry and microbial community dynamics.

In marine environments, macrofauna living in or on the sediment surface may alter the structure, diversity and function of benthic microbial communities. In particular, microbial nitrogen (N)-cycling processes may be enhanced by the activity of large bioturbating organisms. Here, we study the effect of the burrowing mud shrimp Upogebia deltaura upon temporal variation in the abundance of genes representing key N-cycling functional guilds. The abundance of bacterial genes representing different N-cycling guilds displayed different temporal patterns in burrow sediments in comparison with surface sediments, suggesting that the burrow provides a unique environment where bacterial gene abundances are influenced directly by macrofaunal activity. In contrast, the abundances of archaeal ammonia oxidizers varied temporally but were not affected by bioturbation, indicating differential responses between bacterial and archaeal ammonia oxidizers to environmental physicochemical controls. This study highlights the importance of bioturbation as a control over the temporal variation in nitrogen-cycling microbial community dynamics within coastal sediments.

Biofilms present complex assemblies of micro-organisms attached to surfaces. They are dynamic structures in which various metabolic activities and interactions between the component cells occur. When phage come in contact with biofilms, further interactions occur dependent on the susceptibility of the biofilm bacteria to phage and to the availability of receptor sites. If the phage also possess polysaccharide-degrading enzymes, or if considerable cell lysis is effected by the phage, the integrity of the biofilm may rapidly be destroyed. Alternatively, coexistence between phage and host bacteria within the biofilm may develop. Although phage have been proposed as a means of destroying or controlling biofilms, the technology for this has not yet been successfully developed. ß 2004 Published by Elsevier B.V. on behalf of the Federation of European Microbiological Societies.

1] Ammonia oxidation is a key microbial process within the marine N-cycle. Sediment and water column samples from two contrasting sites in the English Channel (mud and sand) were incubated (up to 14 weeks) in CO 2 -acidified seawater ranging from pH 8.0 to pH 6.1. Additional observations were made off the island of Ischia (Mediterranean Sea), a natural analogue site, where long-term thermogenic CO 2 ebullition occurs (from pH 8.2 to pH 7.6). Water column ammonia oxidation rates in English Channel samples decreased under low pH with near-complete inhibition at pH 6.5. Water column Ischia samples showed a similar though not statistically significant trend. However, sediment ammonia oxidation rates at all three locations were not affected by reduced pH. These observations may be explained by buffering within sediments or low-pH adaptation of the microbial ammonia oxidizing communities. Our observations have implications for modeling the future impact of ocean acidification on marine ecosystems. Citation: Kitidis, V.,

Stenotrophomonas maltophilia PML168 was isolated from Wembury Beach on the English Coast from a rock pool following growth and selection on agar plates. Here we present the permanent draft genome sequence, which has allowed prediction of function for several genes encoding enzymes relevant to industrial biotechnology, including a novel flavoprotein monooxygenase.

Bacteria are capable of complex assemblage behavior through cell-to-cell communication using diffusible chemical signal molecules that accumulate to a threshold concentration that activates target genes (1). This process, termed "quorum sensing," has not been shown to cross the prokaryote-eukaryote boundary. Here, we show that diffusible signal molecules produced by bacterial biofilms are exploited by the planktonic phase of a marine eukaryotic organism-the green seaweed Enteromorpha.

Bioturbation is a key process in coastal sediments, influencing microbially driven cycling of nutrients as well as the physical characteristics of the sediment. However, little is known about the distribution, diversity and function of the microbial communities that inhabit the burrows of infaunal macroorganisms. In this study, terminal-restriction fragment length polymorphism analysis was used to investigate variation in the structure of bacterial communities in sediment bioturbated by the burrowing shrimp Upogebia deltaura or Callianassa subterranea. Analyses of 229 sediment samples revealed significant differences between bacterial communities inhabiting shrimp burrows and those inhabiting ambient surface and subsurface sediments. Bacterial communities in burrows from both shrimp species were more similar to those in surface-ambient than subsurface-ambient sediment (R ¼ 0.258, Po0.001). The presence of shrimp was also associated with changes in bacterial community structure in surrounding surface sediment, when compared with sediments uninhabited by shrimp. Bacterial community structure varied with burrow depth, and also between individual burrows, suggesting that the shrimp's burrow construction, irrigation and maintenance behaviour affect the distribution of bacteria within shrimp burrows. Subsequent sequence analysis of bacterial 16S rRNA genes from surface sediments revealed differences in the relative abundance of bacterial taxa between shrimp-inhabited and uninhabited sediments; shrimp-inhabited sediment contained a higher proportion of proteobacterial sequences, including in particular a twofold increase in Gammaproteobacteria. Chao1 and ACE diversity estimates showed that taxon richness within surface bacterial communities in shrimp-inhabited sediment was at least threefold higher than that in uninhabited sediment. This study shows that bioturbation can result in significant structural and compositional changes in sediment bacterial communities, increasing bacterial diversity in surface sediments and resulting in distinct bacterial communities even at depth within the burrow. In an area of high macrofaunal abundance, this could lead to alterations in the microbial transformations of important nutrients at the sediment-water interface.