Genome Sequence of Ruegeria sp. Strain KLH11, anN-Acylhomoserine Lactone-Producing Bacterium Isolated from the Marine Sponge Mycale laxissima

Abstract

Ruegeria sp. strain KLH11, isolated from the marine sponge Mycale laxissima, produces a complex profile of N-acylhomoserine lactone quorum-sensing (QS) molecules. The genome sequence provides insights into the genetic potential of KLH11 to maintain complex QS systems, and this is the first genome report of a cultivated symbiont from a marine sponge.Sponges can harbor diverse assemblages of microbes, which in some cases can comprise up to 60% of the total biomass of the holobiont (6). This dense microbial community associated with sponges provides conditions in which quorum sensing (QS) may be important (3,8). QS is a process by which bacteria coordinate group activities such as bioluminescence, antibiotic production, and virulence at high cell density via signal molecules, typified by N-acylhomoserine lactone (AHL) in Proteobacteria (2). We isolated a Ruegeria sp. strain, KLH11 [99% 16S rRNA gene sequence similarity with Ruegeria lacuscaerulensis (GenBank accession no. HQ908678)], which can produce a complex profile of long-chain AHLs (3). The Silicibacter-Ruegeria subgroup belongs to the Roseobacter clade, which is widespread and numerous in surface waters of the oceans and includes organisms that are involved in global sulfur cycling (1). Many roseobacters have been fully sequenced and annotated (5). Ruegeria sp. strain KLH11 is the first readily culturable sponge symbiont to have its genome sequenced. This microorganism has been isolated from several samples of the sponge Mycale laxissima and is termed a symbiont using the term as defined by Taylor et al. (6).The genome was sequenced by a combination of Roche/454 and Sanger whole-genome shotgun sequencing. Sequences were assembled with a Celera assembler (4), and genes were predicted and annotated using the automated CloVR-Microbe pipeline (7).The draft genome has a total assembly size of ca. 4.5 Mb, with an average GϩC content of 57.3 mol%, and is predicted to have 4,493 coding sequences (CDS), of which 3,243 (ca. 72.0%) were functionally annotated. This assembly is comprised of 13 scaffolds, of which the largest is ca. 3.1 Mb, having 3,193 CDS. One scaffold of 81 kbp shows similarity to the megaplasmid identified in Ruegeria pomeroyi DSS-3 (accession number CP000032) and could indicate the presence of extrachromosomal elements in the genome. Approximately 82.0% of all the predicted CDS have ATG sequences at the presumptive start position, and 17.8% may use GTG or TTG. One complete rRNA operon, one partial 16S rRNA gene, two 5S rRNA genes, and 52 tRNAs were identified. About 34 genes were found to be related to the synthesis of bacterial flagella and regulation of bacterial motility. No candidates for chemotaxis functions were observed in the genome. Two sets of quorum-sensing circuits were identified, implying the potential for a complex quorum-sensing mechanism. Roughly 44 genes were predicted to function in the biosynthesis and degradation of polysaccharides.Comparison of KLH11 genome sequences to closely related free-living bacteria, such as R. pomeroyi ...

The CckA-ChpT-CtrA Phosphorelay System Is Regulated by Quorum Sensing and Controls Flagellar Motility in the Marine Sponge Symbiont Ruegeria sp. KLH11

“…Annotation of the KLH11 genome revealed that KLH11 has homologues to each of the cckA , chpT and ctrA genes [11]. The putative KLH11 cckA gene (GenBank No.…”

“…S2A). Antiserum raised against whole flagella from C. crescentus , a related alpha-proteobacterium, was able to recognize KLH11 flagellin protein encoded by the fliC gene, of approximately 41.5 kDa in size [11], [19] and was used in the western blot assay. Results showed that none of the three null mutants produced any detectable flagellin protein (Fig.…”

“…Phospho-CtrA, relieved of CpdR inhibition, is an active transcriptional regulator that controls about 26% (144/553) of the genes involved in cell cycle progression and also controls flagellar motility in C. crescentus [10]. Members of the Rhodobacterales such as KLH11 encode cckA , chpT , and ctrA homologues [11], but generally no cpdR homologue [12]. In Silicibacter sp.…”

“…For example, in Burkholderia glumae the tofRI QS system regulates the expression of flhDC , which in turn directly controls motility [20]. Although the FlhDC and FleQ/FlrA homologues are not present in Ruegeria pomeroyi DSS-3 or in KLH11 genome sequences [11], [21], it is possible that the cckA-chpT - ctrA pathway acts in this capacity. In this study we examined whether 1) cckA , chpT and ctrA genes are essential for the viability of KLH11; 2) they can control flagellar motility; 3) they are influenced by the SsaRI system.…”

A solo luxI-type gene directs acylhomoserine lactone synthesis and contributes to motility control in the marine sponge symbiont Ruegeria sp. KLH11

“…Genome sequencing of KLH11 revealed the presence of a luxI homologue, designated sscI, encoded on a large assembled sequence scaffold (.700 kb) that contains neither the ssaRI nor the ssbRI operons (Zan et al, 2011). The sscI and ssbI genes and their translation products are highly similar (70 % and 76 % nucleotide and amino acid identity, respectively) (Fig.…”