Rapid tissue reduction and recovery in the sponge Aplysinella sp.

We observed a pronounced, yet reversible tissue reduction in the tropical sponge Aplysinella sp. under non-experimental conditions in its natural habitat, after transfer into seawater tanks, as well as after transplantation from deep to shallow water in the field. Tissue reduction resulted in the formation of small “reduction bodies” tightly attached to the sponge skeleton. Although volume loss and gain were substantial, both tissue reduction and regeneration were often remarkably rapid, occurring within few hours. Microscopic analysis of the reduction bodies revealed morphological similarities to previously described sponge primmorphs, with densely packed archaeocytes and spherulous cells enclosed by a thin layer of epithelial-like cells. Denaturing gradient gel electrophoresis (DGGE) revealed pronounced changes in the sponge-associated microbial community upon tissue reduction during laboratory and field experiments and following changes in ambient conditions after transplantation in the field. Generally, the microbial community associated with this sponge proved less stable, less abundant, and less diverse than those of other, previously investigated Verongid sponges. However, one single phylotype was consistently present in DGGE profiles of Aplysinella sp. This phylotype clustered with γ-proteobacterial sequences found previously in other sponge species of different taxonomic affiliations and geographic provenances, as well as in sponge larvae. No apparent changes in the total secondary metabolite content (per dry weight) occurred in Aplysinella sp. upon tissue reduction; however, comparative analysis of intact and reduced tissue suggested changes in the concentrations of two minor compounds. Besides being ecologically interesting, the tissue reduction phenomenon in Aplysinella sp. provides an experimentally manipulable system for studies on sponge/microbe symbioses. Moreover, it may prove useful as a model system to investigate molecular mechanisms of basic Metazoan traits in vivo, complementing the in vitro sponge primmorph system currently used in this context.     

Monitoring microbial diversity and natural product profiles of the sponge<Emphasis Type="Italic"> Aplysina cavernicola</Emphasis> following transplantation

In order to assess the stability of the microbial community of the sponge Aplysina cavernicola under in situ conditions, sponges were transplanted from their original location (>40 m depth) to shallower, more light-exposed sites (7-15 m depth). Transmission electron microscopy revealed that the microbial community remained visually unchanged and free of cyanobacteria over the experimental time period of 3 months. Denaturing gradient gel electrophoresis (DGGE) of PCR-amplified partial 16S rRNA gene sequences allowed a distinction between the variable and permanent fraction of the bacterial community. Comparative sequence analysis of four variable DGGE bands revealed high sequence similarity to representatives of the Alpha- and Gammaproteobacteria and the phylum Bacteroidetes, which have been recovered previously from Mediterranean seawater as clone sequences or by cultivation. Seven (out of 12) permanent DGGE bands showed high sequence similarity to a sponge-specific, monophyletic 16S rRNA gene sequence cluster within the Acidobacteria division, and to a sequence cluster of uncertain affiliation. These sequence clusters represent members of a common microbial community that is shared among distantly related sponges from different, non-overlapping geographic regions. Four additional permanent DGGE bands showed high sequence similarity to a Betaproteobacterium, Burkholderia cepacia, which is not typically known as a marine bacterium. High-performance liquid chromatography analyses of sponge tissues revealed no changes in metabolite pattern, indicating that these compounds are expressed constitutively irrespective of the variations resulting from the transplantation experiment.     

Impact of the polychaete Capitella sp. I on microbial activity in an organic-rich marine sediment contaminated with the polycyclic aromatic hydrocarbon fluoranthene

Polychaetes belonging to the genus Capitella are often present in high numbers in organic-rich sediments polluted with, e.g., oil components, and Capitella spp. may have a great impact on the biogeochemistry of these sediments. We examined the influence of Capitella sp. I on microbial activity in an organic-rich marine sediment contaminated with the polycyclic aromatic hydrocarbon, fluoranthene. Capitella sp. I were added to microcosms (10 000 ind m⁻²) and the impact of a pulse-sedimentation of fluoranthene-contaminated sediment (3 mm layer) was studied for a period of 12 d after sedimentation. The sediment oxygen uptake and total sediment metabolism (TCO₂ production) increased in cores with worms (71 to 131%), whereas the anaerobic activity, measured as sulfate reduction rate 12 d after sedimentation, was lower compared to cores without worms. The effect of fluoranthene on sulfate reduction was most pronounced in the presence of worms, with a 34% reduction versus 16% in cores without worms. The reduced sulfur pools in cores with worms were smaller than in cores without worms, suggesting that the reduced anaerobic activity was caused by increased oxidation of the sediment, which may favor O₂ and other electron-acceptors (e.g. NO₃ ⁻, Fe³⁺, Mn⁴⁺) in organic matter decomposition. The sediment oxygen uptake and TCO₂ production did not show significant changes due to fluoranthene treatment, indicating that these parameters were either less sensitive to fluoranthene stress or recovered more rapidly (i.e. within 48 h) than sulfate reduction rates. Bioturbation by Capitella sp. I altered the depth profile of fluoranthene such that fluoranthene was found in deeper sediment layers (down to 2 cm) where diffusional loss and microbial breakdown probably are reduced relative to surface layers. In cores without worms, fluoranthene was found down to 1 cm, with 75% remaining in the upper 5 mm. Received: 5 December 1996 / Accepted: 11 February 1997     

Oxygen dynamics and transport in the Mediterranean sponge Aplysina aerophoba

The Mediterranean sponge Aplysina aerophoba kept in aquaria or cultivation tanks can stop pumping for several hours or even days. To investigate changes in the chemical microenvironments, we measured oxygen profiles over the surface and into the tissue of pumping and non-pumping A. aerophoba specimens with Clark-type oxygen microelectrodes (tip diameters 18–30 μm). Total oxygen consumption rates of whole sponges were measured in closed chambers. These rates were used to back-calculate the oxygen distribution in a finite-element model. Combining direct measurements with calculations of diffusive flux and modeling revealed that the tissue of non-pumping sponges turns anoxic within 15 min, with the exception of a 1 mm surface layer where oxygen intrudes due to molecular diffusion over the sponge surface. Molecular diffusion is the only transport mechanism for oxygen into non-pumping sponges, which allows total oxygen consumption rates of 6–12 μmol cm⁻³ sponge day⁻¹. Sponges of different sizes had similar diffusional uptake rates, which is explained by their similar surface/volume ratios. In pumping sponges, oxygen consumption rates were between 22 and 37 μmol cm⁻³ sponge day⁻¹, and the entire tissue was oxygenated. Combining different approaches of direct oxygen measurement in living sponges with a dynamic model, we can show that tissue anoxia is a direct function of the pumping behavior. The sponge-microbe system of A. aerophoba thus has the possibility to switch actively between aerobic and anaerobic metabolism by stopping the water flow for more than 15 min. These periods of anoxia will greatly influence physiological variety and activity of the sponge microbes. Detailed knowledge about the varying chemical microenvironments in sponges will help to develop protocols to cultivate sponge-associated microbial lineages and improve our understanding of the sponge-microbe-system.     

Arsenic detoxification potential of <Emphasis Type="Italic">aox</Emphasis> genes in arsenite-oxidizing bacteria isolated from natural and constructed wetlands in the Republic of Korea

Arsenic is subject to microbial interactions, which support a wide range of biogeochemical transformations of elements in natural environments such as wetlands. The arsenic detoxification potential of the bacterial strains was investigated with the arsenite oxidation gene, aox genotype, which were isolated from the natural and constructed wetlands. The isolates were able to grow in the presence of 10 mM of sodium arsenite (As(III) as NaAsO₂) and 1 mM of d+glucose. Phylogenetic analysis based on 16S rRNA gene sequencing indicated that these isolated strains resembled members of the genus that have arsenic-resistant systems (Acinetobacter sp., Aeromonas sp., Agrobacterium sp., Comamonas sp., Enterobacter sp., Pantoea sp., and Pseudomonas sp.) with sequence similarities of 81–98%. One bacterial isolate identified as Pseudomonas stutzeri strain GIST-BDan2 (EF429003) showed the activity of arsenite oxidation and existence of aoxB and aoxR gene, which could play an important role in arsenite oxidation to arsenate. This reaction may be considered as arsenic detoxification process. The results of a batch test showed that P. stutzeri GIST-BDan2 (EF429003) completely oxidized in 1 mM of As(III) to As(V) within 25–30 h. In this study, microbial activity was evaluated to provide a better understanding of arsenic biogeochemical cycle in both natural and constructed wetlands, where ecological niches for microorganisms could be different, with a specific focus on arsenic oxidation/reduction and detoxification.     

Culturable actinobacteria isolated from marine sponge <Emphasis Type="Italic">Iotrochota</Emphasis> sp.

The study describes the diversity of actinobacteria isolated from the marine sponge Iotrochota sp. collected in the South China Sea. Species and natural product diversity of isolates were analyzed, including screening for genes encoding polyketide synthases (PKS) and nonribosomal peptide synthetase (NRPS), and 16S rRNA gene restriction fragment length polymorphism (RFLP). PKS and NRPS sequences were detected in more than half of the isolates and the different “PKS-I–PKS-II–NRPS” combinations in different isolates belonging to the same species indicated a potential natural product diversity and divergent genetic evolution. The phylogenetic analysis based on 16S rRNA gene sequencing showed that the isolates belonged to genera Streptomyces, Cellulosimicrobium, and Nocardiopsis. The majority of the strains tested belonged to the genus Streptomyces and one of them may be a new species. To our knowledge, this is the first report of a bacterium classified as Cellulosimicrobium sp. isolated from a marine sponge. Key Laboratory of Marine Bio-recourses Sustainable Utilization (LMB-CAS), Guangdong Key Laboratory of Marine Materia Medica (LMMM-GD), South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, People’s Republic of China.     

Spatial distribution of bacteria associated with the marine sponge Tethya californiana

Microbial diversity and spatial distribution of the diversity within tissue of the marine sponge Tethya californiana was analyzed based on 16S rRNA gene sequences. One candidate division and nine bacterial phyla were detected, including members of all five subdivisions of Proteobacteria. Moreover, chloroplast-derived Stramenopiles- and Rhodophyta-affiliated 16S rRNA gene sequences were found and Stramenopiles represented the most abundant clones (30%) in the clone library. On the phylum-level, the microbial fingerprint of T. californiana showed a similar pattern as its Mediterranean relative T. aurantium. An interesting difference was that Cyanobacteria that were abundantly present in T. aurantium were not found in T. californiana, but that the latter sponges harbored phototrophic Stramenopiles instead. Surprisingly, the phototrophic microorganisms were evenly distributed over the inner and outer parts of the sponge tissue, which implies that they also reside in regions without direct light exposure. The other phyla were also present in both the outer cortex and the mesohyl of the sponges. These results were confirmed by analysis on the operational taxonomic unit level. This leads to the conclusion that from a qualitative point of view, spatial distribution of microorganisms in T. californiana tissue is quite homogeneous. Thirty-two percent of the operational taxonomic units shared less than 95% similarity with any other known sequence. This indicates that marine sponges are a rich source of previously undetected microbial life.     

Distribution of secondary metabolites in the sponge Oceanapia sp. and its ecological implications

The Micronesian sponge Oceanapia sp. has an unusual growth form that consists of an irregular turnip-shaped base, which is buried in the substrate. One to several fistules, which protrude through the sand, are attached to the base of the sponge. On top of each fistule is a small fragile capitum. We examined whether this conspicuous red-colored sponge was chemically defended and if intraspecimen variation existed in the distribution of secondary metabolites between different parts of the sponge. Furthermore we assessed the deterrent properties of the secondary metabolites to generalist and more specialized fish predators. We also wanted to see if the optimal defense theory holds in the case of a marine invertebrate. According to the theory, organisms evolve and allocate defenses in a way that maximizes individual fitness, assuming that defenses are costly to the fitness of the organisms. We were able to evaluate this hypothesis, since the different sponge parts in Oceanapia sp. were at different risk to damage by predators and had a different value in terms of fitness loss to the sponge (the capitum probably plays a role in asexual propagation). Concentrations of crude organic extract increased from the base to the capitum of the sponge. The major secondary metabolites kuanoniamine C and D also showed a sharp increase from the basal root to the capitum. There was no difference in structural material or ash content between the base and the fistule of the sponge, but fiber and protein content were significantly higher in the fistule. The methanol fraction was highly deterrent in field feeding assays towards generalist reef fish at base concentration. It also deterred feeding by the spongivorous angelfish Pomacanthus imperator in laboratory feeding experiments at the same concentration. The field feeding assays with pure compounds showed that kuanoniamine C and D deterred feeding by natural assemblages of reef fishes at fistule concentrations, confirming their role as defensive agents. The intraspecimen variation of secondary metabolites in Oceanapia sp. supports the optimal defense theory by showing the highest concentrations in those parts of the sponge that are most visible to predators and are likely to be most important for inclusive fitness. Received: 5 May 1999 / Accepted: 16 September 1999     

Linking abundance and diversity of sponge-associated microbial communities to metabolic differences in host sponges

Many sponge species contain large and diverse communities of microorganisms. Some of these microbes are suggested to be in a mutualistic interaction with their host sponges, but there is little evidence to support these hypotheses. Stable nitrogen isotope ratios of sponges in the Key Largo, Florida (USA) area grouped sponges into species with relatively low δ¹⁵N ratios and species with relatively high δ¹⁵N ratios. Using samples collected in June 2002 from Three Sisters Reef and Conch Reef in the Key Largo, Florida area, transmission electron microscopy (TEM) and denaturing gradient gel electrophoresis were performed on tissues of the sponges Ircinia felix and Aplysina cauliformis, which are in the low δ¹⁵N group, and on tissue of the sponge Niphates erecta, which is in the high δ¹⁵N group. Results showed that I. felix and A. cauliformis have large and diverse microbial communities, while N. erecta has a low biomass of one bacterial strain. As the low δ¹⁵N ratios indicated a microbial input of nitrogen, these results suggested that I. felix and A. cauliformis were receiving nitrogen from their associated microbial community, while N. erecta was obtaining nitrogen solely from external sources. Sequence analysis of the microbial communities showed a diversity of metabolic capabilities among the microbes of the low δ¹⁵N group, which are lacking in the high δ¹⁵N group, further supporting metabolic differences between the two groups. This research provides support for hypotheses of mutualisms between sponges and their associated microbial communities.     

Endofauna differences between two temperate marine sponges (Demospongiae; Haplosclerida; Chalinidae) from southwest Australia

The endofaunal assemblages associated with two species of sponge from the family Chalinidae (Haliclona sp. 1 and Haliclona sp. 2) were studied at four locations along the south west coast of Australia. The species have distinct morphologies and inhabit similar microhabitats; there is also considerable scientific interest in Haliclona sp. 1 (green Haliclona) due to the unique bioactive compound it produces. A total of 948 and 287 endofaunal individuals were found associated with 16 specimens of both the green Haliclona and Haliclona sp. 2 (brown Haliclona), respectively. Twenty-four endofaunal taxa were found (from mysid shrimps to teleost fish), with the brown Haliclona having a greater density of endofaunal species and individuals than the green Haliclona. The endofaunal assemblages of both species of sponge were significantly different, but only the endofaunal assemblage within the green Haliclona differed significantly among locations. Differences in the abundance and biomass of associated endofauna of each species of sponge can be related to differences in their morphologies, size and internal structure. In the green Haliclona, differences in endofaunal assemblages among locations are unlikely to be due to environmental influences as taxa discriminating each locations assemblage were common to both species of sponge. Numerous endofaunal individuals were found to be reproductively active, and it is clear that the species of sponge provide important habitats for their associated endofauna. This provision of habitat needs to be taken into account when harvesting green Haliclona biomass for supply of its target bioactive compound for further pharmaceutical development.     

Decreased lipid storage in juvenile Atlantic cod (<Emphasis Type="Italic">Gadus morhua</Emphasis>) during settlement in cold-water eelgrass habitat

We characterized the prey field and the lipid classes/fatty acids in the flesh of age 0 juvenile cod (Gadus morhua) during their late-summer/fall arrival and settlement into eelgrass (Zostera marina) in coastal Newfoundland. Examination of available prey demonstrated a high abundance of small zooplankton (Acartia, Microsetella and Oithona sp.) with no larger Calanus sp. prey. Breakpoint analysis showed significant changes in the accumulation of relative (mg g⁻¹ wet weight) and absolute (μg fish⁻¹) amounts of lipid with standard length at the time of settlement (~60 mm standard length). Settling juvenile cod showed an alternate lipid utilization strategy where they catabolized phospholipids (PL) to a greater extent than triacylgylcerols (TAG). Polyunsaturated fatty acids (PUFA) content in cod flesh decreased as fish grew indicating that nearshore zooplankton quality was not optimal for PL formation. The dramatic reduction in cod PL was likely due to both catabolism of muscle and a lack of dietary PUFA suitable for PL synthesis. However, juvenile cod continued to grow, leading to decreased lipid stores and suggesting that cod settling into eelgrass are under intense selection pressure for growth prior to the onset of winter, possibly as a means of escaping gape-limited predation. These data contrast better-studied freshwater and estuarine systems in which lipid storage is critical for successful overwintering.     

Alterations inras-gene expression and intracellular distribution of protein kinase C in the spongeGeodia cydonium in response to marine pollution

The siliceous spongeGeodia cydonium Jameson was used to study the influence of pollution in marine environments on selected parameters of the intracellular signal transduction pathway. The parameters chosen were: intracellular distribution of protein kinase C (PK-C),ras-gene expression and DNA polymerasealpha (DNA Polalpha) activity. Both PK-C andras-gene product (ras-protein) have previously been established to be key molecules in the intracellular signalling pathway in sponges; increased level ofras-protein mediates events following sponge cell-cell contact. Three unpolluted and three polluted sites in the off-shore seawater around Rovinj (Yugoslavia) were selected for the study in 1989. The state of pollution of these sites has been well-defined in a series of previous studies (1976 to 1989). Transplantation of regenerating sponge cubes ofG. cydonium to the polluted sites resulted in pronounced changes in the parameters chosen, compared to controls exposed to unpolluted environments. Expression ofras gene was increased by three- to five-fold after exposure of regenerating sponge tissue to the impacted sites. At the site with the highest pollutional load,ras mRNA level was about 50% of that at the reference sites. In parallel experiments it was established that, in response to pollution, a translocation of PK-C from the cytosolic to membrane fraction occurred. At the most impacted site, most of the enzyme activity was cytosolic. DNA Polalpha-activity, as a measure of sponge cell proliferation, decreased in a pollutioncorrelated manner. Our results indicate that the intracellular signalling system within sponge cells is activated in response to moderate pollution but is depressed in heavily polluted environments.     

Cellular localisation of secondary metabolites isolated from the Caribbean sponge <Emphasis Type="Italic">Plakortis simplex</Emphasis>

The Caribbean sponge, Plakortis simplex, is known to contain a large array of secondary metabolites, including the antimalarial polyketide plakortin, several unusual glycolipids, and some hopanoids, which closely resemble typical bacterial metabolites. The hypothesis that they could be products of bacterial metabolism was tested by localizing specific metabolites in cells using physical separation of sponge cells, bacterial symbionts and supernatant by differential centrifugation. The obtained fractions were analysed separately for the typical P. simplex metabolites by NMR and mass spectrometry, and most of them were shown to be present in the bacterial cells but not in the sponge cells. In addition, PCR screening showed that the biosynthetic pathway for glycosphingolipids was present in the bacterial cells. Isolation of a Sphingomonas strain PS193 from P. simplex and subsequent glycosphingolipid analysis resulted in the detection of a known glycosphingolipid, GSL-1, that did, however, not match the glycosphingolipid profile of P. simplex. Therefore, it is unlikely that Sphingomonas strain PS193 is an abundant member of the microbial community associated with P. simplex. Other glycosphingolipid producing bacteria in P. simplex remain to be identified. In conclusion, this study provides experimental evidence that the glycolipids and hopanoids and possibly also the polyketide plakortin are produced by microbial symbionts rather than the sponge from which the metabolites were originally isolated.     

Evidence of nitrification and denitrification in high and low microbial abundance sponges

Aerobic and anaerobic microbial key processes were quantified and compared to microbial numbers and morphological structure in Mediterranean sponges. Direct counts on histological sections stained with DAPI showed that sponges with high microbial abundances (HMA sponges) have a denser morphological structure with a reduced aquiferous system compared to low microbial abundance (LMA) sponges. In Dysidea avara, the LMA sponge, rates of nitrification and denitrification were higher than in the HMA sponge Chondrosia reniformis, while anaerobic ammonium oxidation and sulfate reduction were below detection in both species. This study shows that LMA sponges may host physiologically similar microbes with comparable or even higher metabolic rates than HMA sponges, and that anaerobic processes such as denitrification can be found both in HMA and LMA sponges. A higher concentration of microorganisms in the mesohyl of HMA compared to LMA sponges may indicate a stronger retention of and, hence, a possible benefit from associated microbes.     

Identification of the antifungal peptide-containing symbiont of the marine sponge Theonella swinhoei as a novel δ-proteobacterium, “Candidatus Entotheonella palauensis”

 The Palauan sponge Theonella swinhoei (class Demospongiae, order Lithistida, family Theonellidae) harbors filamentous bacterial symbionts that contain theopalauamide, an antifungal, bicyclic glycopeptide. In this study, the filamentous symbionts were shown to be novel bacteria belonging to the δ-subdivision of proteobacteria. The 16S rRNA gene sequence was determined using a combination of denaturing gradient-gel electrophoresis (DGGE) and specific polymerase chain-reaction (PCR) primers, and its source was confirmed by in situ hybridization. In a series of culture experiments, the filamentous bacteria were propagated in a mixed culture on agar plates. Related 16S rRNA gene sequences were isolated from related sponges with slightly different chemistry. The taxonomic status “Candidatus Entotheonella palauensis” is proposed for the theopalauamide-containing filamentous bacteria from T. swinhoei. Received: 12 June 1999 / Accepted: 22 January 2000     

Transformations of <Emphasis Type="Italic">n</Emphasis>-alkanes from petroleum pollutants in alluvial groundwaters

Investigations presented in this paper were aimed at defining the alterations of n-alkane composition in cases of oil-polluted alluvial sediments. Therefore, oil-polluted groundwater samples, taken in five different time intervals during a period of 28 months, were investigated. Samples of alluvial sediments were taken from two boreholes within an oil refinery at Pancevo, Yugoslavia. In both boreholes significant alterations with characteristic degradation of "oil" n-alkanes with no odd- or even-member predominance were observed, as well as subsequent synthesis of new ones with pronounced even-member predominance, and with maxima at C₁₆ and C₁₈. Since no additional contamination of boreholes was observed by analyses of steranes and triterpanes, the observed changes can only be attributed to microbial activity. It is assumed that for the degradation of oil n-alkanes, as well as for the synthesis of "new" n-alkanes, algae such as dinoflagellates are responsible. This assumption was confirmed by identification of n-alcohols with even-member predominance (C₁₄–C₂₀), by identification of cholesterol, as well as of n-fatty acids with even-member predominance (C₁₄–C₁₈) in the extract with n-alkane even-member predominance. Electronic Publication     

Glass sponges arrest pumping in response to sediment: implications for the physiology of the hexactinellid conduction system

The hexactinellid sponge Rhabdocalyptus dawsoni propagates electrical signals to arrest its feeding current in response to mechanical stimuli and sediment. The deepwater habitat of other species of glass sponge, and the difficulty of working with the tissue in vitro have so far prevented confirmation of electrical signaling in other members of the Class. Here we show in laboratory experiments (ex situ) that mechanical and sediment stimuli trigger immediate arrest in R. dawsoni and in a second species of hexactinellid, Aphrocallistes vastus, suggesting that rapid signaling may be a general feature of glass sponge tissue. Further, responses of the two species differed, suggestive of underlying physiological differences in the conduction system. R. dawsoni and A. vastus were sensitive to sediment but arrests were often prolonged in R. dawsoni, whereas in A. vastus pumping resumed immediately following each arrest. Fine sediment (<25 μm) caused immediate arrests in R. dawsoni and A. vastus, but with a higher stimulus threshold in A. vastus. Large amounts of sediment triggered repeated arrests in both species, and prolonged exposure to sediment (over 4 h) caused a gradual reduction in pumping, with recovery taking up to 25 h. During recovery, both species of sponge carried out repeated arrests, which had a precise periodicity indicative of pacemaker activity. Scanning electron microscopy of the tissue of these specimens showed many chambers were clogged. The results suggest that the glass sponge conduction system generates arrest of the feeding current that prevent uptake of small amounts of sediment, and that each species has different threshold sensitivities. However, ongoing exposure to sediment can clog the filtration apparatus.     

The culturable microbial community of the Great Barrier Reef sponge Rhopaloeides odorabile is dominated by an α-Proteobacterium

The microbial community cultured from the marine sponge Rhopaloeides odorabile Thompson et al. is dominated by a single bacterium, designated strain NW001. Sequence analysis of 1212 bp of the16S rRNA gene of strain NW001 indicates that it is a member of the α-subgroup of the class Proteobacteria. The association between this bacterium and its host sponge was observed in healthy R. odorabile collected from six different reefs in the Great Barrier Reef representing a geographic distance of 460 km, and in four collections in different seasons in 1997–1998 at Davies Reef (18°49.6′S; 147°34.49′E). The proportion of colonies of strain NW001 in samples from R. odorabile, expressed as a percentage of the total heterotrophic bacterial colony count, showed no significant spatial (range: 81–98%) or temporal differences (range: 81–99%), although colony counts of strain NW001 varied by up to two orders of magnitude between reef sites and sampling periods. The location of strain NW001 within the sponge mesohyl was visualized by in situ hybridization, using fluorescently labeled probes based on the 16S rRNA gene sequence of this strain. Cells of strain NW001 surround the choanocyte chambers, suggesting that these bacteria may play a role in nutrient uptake by the sponge. The absence of strain NW001 from corresponding seawater samples indicates that it has a specific, intimate relationship with R. odorabile and is not being utilized as a food source. A unique cyanobacterium related to the genera Leptolyngbya and Plectonema was also isolated from R. odorabile and characterized by 16S rRNA gene sequencing. Received: 19 May 2000 / Accepted: 18 November 2000     

Transmission,plasticity and the molecular identification of cyanobacterial symbionts in the Red Sea sponge <Emphasis Type="Italic">Diacarnus erythraenus</Emphasis>

Cyanobacterial symbionts in the sponge Diacarnus erythraenus from the Red Sea were identified in both adult sponges and their larvae by 16S rDNA sequencing. A single cyanobacterial type was found in all samples. This cyanobacterial type is closely related to other sponge cyanobacterial symbionts. The cyanobacterial rDNA, together with the morphological analysis by electron and fluorescence microscopy, provided evidence for vertical transmission of the symbionts in this sponge. In addition, we show phenotypic plasticity of the symbionts inside the sponge, probably as a result of variability in light availability inside the sponge tissue. Finally, the reproduction of Diacarnus erythraenus is also described.Matan Oren, Laura Steindler have contributed equally to the work.     

Feeding on intertidal microbial mats by postlarval tiger shrimp, <Emphasis Type="Italic">Penaeus semisulcatus</Emphasis> De Haan

A series of experiments investigated the potential role of microbial mats in nutrition of the early settlement stages of Penaeus semisulcatus. From 3 days post-metamorphosis, the microbial mat supported high growth and survival rates in postlarvae, equivalent to that supported by a control diet of Artemia nauplii and mussel. Examination of gut contents indicated that benthic postlarvae feed indiscriminately on the microbial mat. However, when postlarvae were fed separated size-fractions of the microbial mat, only the fraction containing a high concentration of infauna (mainly nematodes) was able to support the same growth as intact microbial mat. This appears to be due to the low nitrogen content (0.4–0.9 mmol g⁻¹) of the various size-fractions, compared to that of infauna (4.0 mmol g⁻¹). The stable isotope composition of the dietary size-fractions and postlarval shrimp tissue supports the hypothesis that the shrimp assimilated C and N primarily from the associated infauna. This may be due to selective feeding that is not apparent from stomach contents, due to rapid digestion of fauna soft tissues, or to differential assimilation of infaunal prey relative to other microbial mat components. The results demonstrate that microbial mats may support survival and growth in early-stage penaeid shrimp postlarvae on intertidal mud flats.