Phaeobacter

Abstract: Members of the Roseobacter lineage of marine bacteria are prolific surface colonizers in marine coastal environments, and antimicrobial secondary metabolite production has been hypothesized to provide a competitive advantage to colonizing roseobacters. Here, we report that the roseobacter Phaeobacter sp. strain Y4I produces the blue pigment indigoidine via a nonribosomal peptide synthase (NRPS)-based biosynthetic pathway encoded by a novel series of genetically linked genes: igiBCDFE. A Tn5-based random mutagenesis library of Y4I showed a perfect correlation between indigoidine production by the Phaeobacter strain and inhibition of Vibrio fischeri on agar plates, revealing a previously unrecognized bioactivity of this molecule. In addition, igiD null mutants (igiD encoding the indigoidine NRPS) were more resistant to hydrogen peroxide, less motile, and faster to colonize an artificial surface than the wild-type strain. Collectively, these data provide evidence for pleiotropic effects of indigoidine production in this strain. Gene expression assays support phenotypic observations and demonstrate that igiD gene expression is upregulated during growth on surfaces. Furthermore, competitive cocultures of V. fischeri and Y4I show that the production of indigoidine by Y4I significantly inhibits colonization of V. fischeri on surfaces. This study is the first to characterize a secondary metabolite produced by an NRPS in roseobacters.

Abstract: Phaeobacter gallaeciensis can antagonize fish-pathogenic bacteria in vitro, and the purpose of this study was to evaluate the organism as a probiont for marine fish larvae and their feed cultures. An in vivo mechanism of action of the antagonistic probiotic bacterium is suggested using a non-antagonistic mutant. P. gallaeciensis was readily established in axenic cultures of the two microalgae Tetraselmis suecica and Nannochloropsis oculata, and of the rotifer Brachionus plicatilis. P. gallaeciensis reached densities of 107 cfu/ml and did not adversely affect growth of algae or rotifers. Vibrio anguillarum was significantly reduced by wild-type P. gallaeciensis, when introduced into these cultures. A P. gallaeciensis mutant that did not produce the antibacterial compound tropodithietic acid (TDA) did not reduce V. anguillarum numbers, suggesting that production of the antibacterial compound is important for the antagonistic properties of P. gallaeciensis. The ability of P. gallaeciensis to protect fish larvae from vibriosis was determined in a bath challenge experiment using a multidish system with 1 larva per well. Unchallenged larvae reached 40% accumulated mortality which increased to 100% when infected with V. anguillarum. P. gallaeciensis reduced the mortality of challenged cod larvae (Gadus morhua) to 10%, significantly below the levels of both the challenged and the unchallenged larvae. The TDA mutant reduced mortality of the cod larvae in some of the replicates, although to a much lesser extent than the wild type. It is concluded that P. gallaeciensis is a promising probiont in marine larviculture and that TDA production likely contributes to its probiotic effect.

Abstract: The production of N-acyl homoserine lactones (AHLs) is widely distributed within the marine Roseobacter clade, and it was proposed that AHL-mediated quorum sensing (QS) is one of the most common cell-to-cell communication mechanisms in roseobacters. The traits regulated by AHL-mediated QS are yet not known for members of the Roseobacter clade, but production of the antibiotic tropodithietic acid (TDA) was supposed to be controlled by AHL-mediated QS in Phaeobacter spp. We describe here for the first time the functional role of luxR and luxI homologous genes of an organism of the Roseobacter clade, i.e., pgaR and pgaI in Phaeobacter gallaeciensis. Our results demonstrate that the AHL synthase gene pgaI is responsible for production of N-3-hydroxydecanoylhomoserine lactone (3OHC(10)-HSL). Insertion mutants of pgaI and pgaR are both deficient in TDA biosynthesis and the formation of a yellow-brown pigment when grown in liquid marine broth medium. This indicates that in P. gallaeciensis the production of both secondary metabolites is controlled by AHL-mediated QS. Quantitative real-time PCR showed that the transcription level of tdaA, which encodes an essential transcriptional regulator for TDA biosynthesis, decreased 28- and 51-fold in pgaI and pgaR genetic backgrounds, respectively. These results suggest that both the response regulator PgaR and the 3OHC(10)-HSL produced by PgaI induce expression of tdaA, which in turn positively regulates expression of the tda genes. Moreover, we confirmed that TDA can also act as autoinducer in P. gallaeciensis, as previously described for Silicibacter sp. strain TM1040, but only in the presence of the response regulator PgaR.

Abstract: A total of 523 bacterial strains were isolated during a 4-year period from mollusc hatcheries (flat oyster and clams) in Galicia (NW Spain). All of the strains were tested for their antibacterial activity against three larval pathogens (Vibrio anguillarum USC-72, V. neptunius PP-145.98, and Vibrio sp. PP-203). Of the isolates, 52 inhibited at least one of the target strains, and 11 inhibited all of them. The main source of active strains was oyster larvae, followed by water, tank surfaces, spat, and broodstock. Four similar strains, belonging to the genus Phaeobacter, showed the strongest activity. Strain PP-154, selected as representative of this group, displayed a wide spectrum of inhibitory activity against aquaculture pathogens, especially against members of the genus Vibrio, which is responsible for the most larval deaths. The inhibitory ability of such strain on solid medium was confirmed in seawater experiments, and the optimal conditions for antibacterial activity were established. These strains are promising probiotics for aquaculture facilities. Their potential benefit is based on the capacity to control the proliferation of a variety of aquaculture bacterial pathogens in mollusc larval cultures. [Int Microbiol 2009; 12(2):107-114]