RESUMEN
Bacteria isolated from marine sponges, including the Silicibacter-Ruegeria (SR) subgroup of the Roseobacter clade, produce N-acylhomoserine lactone (AHL) quorum sensing signal molecules. This study is the first detailed analysis of AHL quorum sensing in sponge-associated bacteria, specifically Ruegeria sp. KLH11, from the sponge Mycale laxissima. Two pairs of luxR and luxI homologues and one solo luxI homologue were identified and designated ssaRI, ssbRI and sscI (sponge-associated symbiont locus A, B and C, luxR or luxI homologue). SsaI produced predominantly long-chain 3-oxo-AHLs and both SsbI and SscI specified 3-OH-AHLs. Addition of exogenous AHLs to KLH11 increased the expression of ssaI but not ssaR, ssbI or ssbR, and genetic analyses revealed a complex interconnected arrangement between SsaRI and SsbRI systems. Interestingly, flagellar motility was abolished in the ssaI and ssaR mutants, with the flagellar biosynthesis genes under strict SsaRI control, and active motility only at high culture density. Conversely, ssaI and ssaR mutants formed more robust biofilms than wild-type KLH11. AHLs and the ssaI transcript were detected in M. laxissima extracts, suggesting that AHL signalling contributes to the decision between motility and sessility and that it may also facilitate acclimation to different environments that include the sponge host.
Asunto(s)
Biopelículas/crecimiento & desarrollo , Poríferos/microbiología , Percepción de Quorum/fisiología , Roseobacter/metabolismo , 4-Butirolactona/análogos & derivados , 4-Butirolactona/metabolismo , Animales , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Regulación Bacteriana de la Expresión Génica/genética , Regulación Bacteriana de la Expresión Génica/fisiología , Percepción de Quorum/genética , Roseobacter/genética , Roseobacter/crecimiento & desarrollo , Transducción de SeñalRESUMEN
Aerobic ammonia-oxidizing bacteria (AAOB) are known to have an important function in the marine nitrogen cycle. Anaerobic ammonium oxidation (anammox) carried out by some members of Planctomycetales is also an important process in marine ecosystems. Ammonia-monooxygenase gene (amoA) fragments were amplified to investigate the potential for nitrification and the diversity of the AAOB in two marine sponges Ircinia strobilina and Mycale laxissima. All of the AmoA sequences obtained from the two sponges clustered with the AmoA sequences of the Betaproteobacteria Nitrosospira spp. To investigate the anaerobic ammonia-oxidizing bacteria (AnAOB) in sponges, 16S rRNA gene fragments of Planctomycetales and anammox bacteria were also amplified with specific primers, and clone libraries were constructed. The Planctomycetales diversity detected in the two sponges was different. The Planctomycetales community in M. laxissima was affiliated with Pirellula, Planctomyces and anammox bacteria, while all of the I. strobilina Planctomycetales clones were solely affiliated with the candidate phylum 'Poribacteria'. Interestingly, sequences related to anammox genera were recovered only from M. laxissima. This is the first report of anammox bacteria in marine sponges. It is intriguing to find AAOB and AnAOB in M. laxissima, but the nature of their interaction with the sponge host and with each other remains unclear. This work further supports the potential of sponge-associated microorganisms for nitrification and sheds light on anammox as a new aspect of the nitrogen cycle in marine sponges.
Asunto(s)
Amoníaco/metabolismo , Bacterias Aerobias/clasificación , Bacterias Aerobias/metabolismo , Bacterias Anaerobias/clasificación , Bacterias Anaerobias/metabolismo , Biodiversidad , Poríferos/microbiología , Animales , Bacterias Aerobias/aislamiento & purificación , Bacterias Anaerobias/aislamiento & purificación , ATPasas de Translocación de Protón Bacterianas , Análisis por Conglomerados , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Ribosómico/química , ADN Ribosómico/genética , Datos de Secuencia Molecular , Oxidación-Reducción , Oxidorreductasas/genética , Filogenia , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADN , Homología de SecuenciaRESUMEN
Red Sea sponges offer potential as sources of novel drugs and bioactive compounds. Sponges harbor diverse and abundant prokaryotic communities. The diversity of Egyptian sponge-associated bacterial communities has not yet been explored. Our study is the first culture-based and culture-independent investigation of the total bacterial assemblages associated with two Red Sea Demosponges, Hyrtios erectus and Amphimedon sp. Denaturing gradient gel electrophoresis fingerprint-based analysis revealed statistically different banding patterns of the bacterial communities of the studied sponges with H. erectus having the greater diversity. 16S rRNA clone libraries of both sponges revealed diverse and complex bacterial assemblages represented by ten phyla for H. erectus and five phyla for Amphimedon sp. The bacterial community associated with H. erectus was dominated by Deltaproteobacteria. Clones affiliated with Gammaproteobacteria were the major component of the clone library of Amphimedon sp. About a third of the 16S rRNA gene sequences in these communities were derived from bacteria that are novel at least at the species level. Although the overall bacterial communities were significantly different, some bacterial groups, including members of Alphaproteobacteria, Gammaproteobacteria, Acidobacteria, and Actinobacteria, were found in both sponge species. The culture-based component of this study targeted Actinobacteria and resulted in the isolation of 35 sponge-associated microbes. The current study lays the groundwork for future studies of the role of these diverse microbes in the ecology, evolution, and development of marine sponges. In addition, our work provides an excellent resource of several candidate bacteria for production of novel pharmaceutically important compounds.
Asunto(s)
Bacterias/aislamiento & purificación , Poríferos/microbiología , Animales , Bacterias/genética , Secuencia de Bases , Biodiversidad , Clonación Molecular , Electroforesis en Gel Bidimensional , Datos de Secuencia Molecular , Filogenia , ARN Ribosómico 16S/genética , SimbiosisRESUMEN
Marine sponges contain complex assemblages of bacterial symbionts, the roles of which remain largely unknown. We identified diverse bacterial nifH genes within sponges and found that nifH genes are expressed in sponges. This is the first demonstration of the expression of any protein-coding bacterial gene within a sponge. Two sponges Ircinia strobilina and Mycale laxissima were collected from Key Largo, Florida and had delta(15)N values of c. 0-1 per thousand and 3-4 per thousand respectively. The potential for nitrogen fixation by symbionts was assessed by amplification of nifH genes. Diverse nifH genes affiliated with Proteobacteria and Cyanobacteria were detected, and expression of nifH genes affiliated with those from cyanobacteria was detected. The nifH genes from surrounding seawater were similar to those of Trichodesmium and clearly different from the cyanobacterial nifH genes detected in the two sponges. This study advances understanding of the role of bacterial symbionts in sponges and suggests that provision of fixed nitrogen is a means whereby symbionts benefit sponges in nutrient-limited reef environments. Nitrogen fixation by sponge symbionts is possibly an important source of new nitrogen to the reef environment that heretofore has been neglected and warrants further investigation.
Asunto(s)
Bacterias/genética , Bacterias/metabolismo , Expresión Génica , Variación Genética , Fijación del Nitrógeno/genética , Nitrógeno/metabolismo , Poríferos/microbiología , Animales , Bacterias/clasificación , Bacterias/aislamiento & purificación , Fenómenos Fisiológicos Bacterianos , Proteínas Bacterianas/genética , Cianobacterias/clasificación , Cianobacterias/genética , Cianobacterias/aislamiento & purificación , Cianobacterias/metabolismo , ADN Bacteriano/química , ADN Bacteriano/genética , Florida , Datos de Secuencia Molecular , Oxidorreductasas/genética , Filogenia , Reacción en Cadena de la Polimerasa/métodos , Poríferos/fisiología , Proteobacteria/clasificación , Proteobacteria/genética , Proteobacteria/aislamiento & purificación , Proteobacteria/metabolismo , Análisis de Secuencia de ADN , Homología de Secuencia , SimbiosisRESUMEN
Marine sponges in the genus Ircinia are known to be good sources of secondary metabolites with biological activities. A major obstacle in the development of sponge-derived metabolites is the difficulty in ensuring an economic, sustainable supply of the metabolites. A promising strategy is the ex situ culture of sponges in closed or semiclosed aquaculture systems. In this study, the marine sponge Ircinia strobilina (order Dictyoceratida: family Irciniidae) was collected from the wild and maintained for a year in a recirculating aquaculture system. Microbiological and molecular community analyses were performed on freshly collected sponges and sponges maintained in aquaculture for 3 months and 9 months. Chemical analyses were performed on wild collected sponges and individuals maintained in aquaculture for 3 months and 1 year. Denaturing gradient gel electrophoresis was used to assess the complexity of and to monitor changes in the microbial communities associated with I. strobilina. Culture-based and molecular techniques showed an increase in the Bacteroidetes and Alpha- and Gammaproteobacteria components of the bacterial community in aquaculture. Populations affiliated with Beta- and Deltaproteobacteria, Clostridia, and Planctomycetes emerged in sponges maintained in aquaculture. The diversity of bacterial communities increased upon transfer into aquaculture.
Asunto(s)
Acuicultura , Bacterias/aislamiento & purificación , Variación Genética , Poríferos/crecimiento & desarrollo , Poríferos/microbiología , Animales , Bacterias/clasificación , Bacterias/genética , Bacterias/crecimiento & desarrollo , Recuento de Colonia Microbiana , ADN Bacteriano/clasificación , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Ecosistema , Genes de ARNr , Datos de Secuencia Molecular , Filogenia , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADNRESUMEN
Marine sponges are hosts to diverse and dense bacterial communities and thus provide a potential environment for quorum sensing. Quorum sensing, a key factor in cell-cell communication and bacterial colonization of higher animals, might be involved in the symbiotic interactions between bacteria and their sponge hosts. Given that marine Proteobacteria are known to produce N-acyl homoserine lactone (AHL) signal molecules, we tested the production of AHLs by Alpha- and Gammaproteobacteria isolated from marine sponges Mycale laxissima and Ircinia strobilina and the surrounding water column. We used three different AHL biodetection systems in diffusion assays: Chromobacterium violaceum, Agrobacterium tumefaciens and Sinorhizobium meliloti with optimal sensitivity to short-chain (C4-C6), moderate-chain (C8-C12) and long-chain (>or= C14) AHLs respectively. Thirteen of 23 isolates from M. laxissima and five of 25 isolates from I. strobilina were found to produce AHLs. Signals were detected from two of eight proteobacterial strains from the water column. Thin-layer chromatographic assays based on the A. tumefaciens reporter system were utilized to determine the AHL profiles of the positive isolates. The types and amounts of AHLs synthesized varied considerably among the strains. Small ribosomal rRNA gene sequencing revealed that the AHL-producing alphaproteobacterial isolates were mainly from the Silicibacter-Ruegeria subgroup of the Roseobacter clade. Two-dimensional gel electrophoresis (2DGE)-based proteomic analyses were congruent with phylogenetic relationships but provided higher resolution to differentiate these closely related AHL-producing strains.
Asunto(s)
Poríferos/microbiología , Proteobacteria/citología , Percepción de Quorum/fisiología , Acil-Butirolactonas/metabolismo , Animales , Cromatografía en Capa Delgada/métodos , Variación Genética , Biología Marina , Filogenia , Proteobacteria/genética , Proteobacteria/aislamiento & purificación , Proteobacteria/metabolismo , Proteómica , ARN Ribosómico 16S/genética , Transducción de SeñalRESUMEN
The changes in bacterial communities associated with the marine sponge Mycale laxissima on transfer to aquaculture were studied using culture-based and molecular techniques. M. laxissima was maintained alive in flowthrough and closed recirculating aquaculture systems for 2 years and 1 year, respectively. The bacterial communities associated with wild and aquacultured sponges, as well as the surrounding water, were assessed using 16S rRNA gene clone library analysis and denaturing gradient gel electrophoresis (DGGE). Bacterial richness and diversity were measured using DOTUR computer software, and clone libraries were compared using S-LIBSHUFF. DGGE analysis revealed that the diversity of the bacterial community of M. laxissima increased when sponges were maintained in aquaculture and that bacterial communities associated with wild and aquacultured M. laxissima were markedly different than those of the corresponding surrounding water. Clone libraries of bacterial 16S rRNA from sponges confirmed that the bacterial communities changed during aquaculture. These communities were significantly different than those of seawater and aquarium water. The diversity of bacterial communities associated with M. laxissima increased significantly in aquaculture. Our work shows that it is important to monitor changes in bacterial communities when examining the feasibility of growing sponges in aquaculture systems because these communities may change. This could have implications for the health of sponges or for the production of bioactive compounds by sponges in cases where these compounds are produced by symbiotic bacteria rather than by the sponges themselves.
Asunto(s)
Acuicultura , Bacterias/genética , Ecosistema , Variación Genética , Filogenia , Poríferos/microbiología , Animales , Secuencia de Bases , Clonación Molecular , Análisis por Conglomerados , Biología Computacional , Cartilla de ADN/genética , Electroforesis , Florida , Biblioteca de Genes , Datos de Secuencia Molecular , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADNRESUMEN
Actinobacteria exclusively within the sub-class Acidimicrobidae were shown by 16S rDNA community analysis to be major components of the bacterial community associated with two sponge species in the genus Xestospongia. Four groups of Actinobacteria were identified in Xestospongia spp., with three of these four groups being found in both Xestospongia muta from Key Largo, Florida and Xestospongia testudinaria from Manado, Indonesia. This suggests that these groups are true symbionts in these sponges and may play a common role in both the Pacific and Atlantic sponge species. The fourth group was found only in X. testudinaria and was a novel assemblage distantly related to any previously sequenced actinobacterial clones. The only actinobacteria that were obtained in initial culturing attempts were Gordonia, Micrococcus and Brachybacterium spp., none of which were represented in the clone libraries. The closest cultured actinobacteria to all the Acidimicrobidae clones from Xestospongia spp. are 'Microthrix parvicella' and Acidimicrobium spp. Xestospongia spp. can now be targeted as source material from which to culture novel Acidimicrobidae to investigate their potential as producers of bioactive compounds. Isolation of sponge-associated Acidimicrobidae will also make it possible to elucidate their role as sponge symbionts.