Roseobacter Clade Bacteria Research Articles

Algicidal lactones from the marine Roseobacter clade bacterium Ruegeria pomeroyi

Volatiles released by the marine Roseobacter clade bacterium Rugeria pomeroyi were collected by use of a closed-loop stripping headspace apparatus (CLSA) and analysed by GC–MS. Several lactones were found for which structural proposals were derived from their mass spectra and unambiguously verified by the synthesis of reference compounds. An enantioselective synthesis of two exemplary lactones was performed to establish the enantiomeric compositions of the natural products by enantioselective GC–MS analyses. The lactones were subjected to biotests to investigate their activity against several bacteria, fungi, and algae. A specific algicidal activity was observed that may be important in the interaction between the bacteria and their algal hosts in fading algal blooms.

Comparative genomics of methylated amine utilization by marine Roseobacter clade bacteria and development of functional gene markers (tmm, gmaS)

The marine Roseobacter clade bacteria comprise up to 20% of the microbial community in coastal surface seawater. Marine Roseobacter clade bacteria are known to catalyse some important biogeochemical transformations in marine carbon and sulfur cycles. Using a comparative genomic approach, this study revealed that many marine Roseobacter clade bacteria have the genetic potential to utilize methylated amines (MAs) as alternative nitrogen sources. These MAs represent a significant pool of dissolved organic carbon and nitrogen in the marine environment. The marine Roseobacter clade bacterial genomes also encode full sets of genes providing them with the potential to generate energy from complete oxidation of the methyl groups of MAs. Representative species of the marine Roseobacter clade were tested and their abilities to use MAs are directly linked to the presence in their genomes of genes encoding key enzymes involved in MA metabolism, including trimethylamine monooxygenase (tmm) and gamma-glutamylmethylamide synthetase (gmaS). These two genes were chosen as functional markers for detecting MA-utilizing marine Roseobacter clade bacteria in the environment. PCR primers targeting these two genes were designed and used successfully to retrieve corresponding gene sequences from MA-utilizing isolates of the marine Roseobacter clade, as well as directly from DNA extracted from surface seawater obtained from Station L4 off the coast of Plymouth, UK. Taken together, the results suggest that MAs may serve as important nitrogen and possibly energy sources for marine Roseobacter clade bacteria, which helps to explain their global success in the marine environment.

Benefits of live phytoplankton, Chlorella vulgaris, as a biocontrol agent against fish pathogen Vibrio anguillarum

The ability of the probiotic bacterium Sulfitobacter to inhibit the growth of two virulent strains (psh-9019 and ATCC 43306) of the fish pathogenic bacterium Vibrio anguillarum was tested. Probiotic and pathogenic bacteria were inoculated individually or together in three different types of media. Two of the phytoplankton media tested were filtrates of phytoplankton Chlorella vulgaris cultures, either the live phytoplankton (live-CV) or a condensed phytoplankton (condensed-CV). Phytoplankton culture medium, ESM, was used as a control medium without phytoplankton. In ESM, Sulfitobacter decreased the viable cell counts of both V. anguillarum strains by tenfold. In the live-CV filtrate, V. anguillarum was eradicated by Sulfitobacter within one week. Although colony counts of strain ATCC 43306 declined during the two-week co-incubation with Sulfitobacter, its growth was not fully inhibited; however, the counts were tenfold lower than that in control ESM medium. Neither of the pathogenic V. anguillarum strains were inhibited nor eradicated by Sulfitobacter in the condensed-CV filtrate medium. Our study indicates that commercially available condensed phytoplankton can enhance the growth of V. anguillarum. Thus, the addition of live phytoplankton, including the introduction of Roseobacter clade bacteria to fish larvae tanks, leads to better biocontrol of the fish pathogen V. anguillarum.

Genetic tools for the investigation of Roseobacter clade bacteria

BackgroundThe Roseobacter clade represents one of the most abundant, metabolically versatile and ecologically important bacterial groups found in marine habitats. A detailed molecular investigation of the regulatory and metabolic networks of these organisms is currently limited for many strains by missing suitable genetic tools.ResultsConjugation and electroporation methods for the efficient and stable genetic transformation of selected Roseobacter clade bacteria including Dinoroseobacter shibae, Oceanibulbus indolifex, Phaeobacter gallaeciensis, Phaeobacter inhibens, Roseobacter denitrificans and Roseobacter litoralis were tested. For this purpose an antibiotic resistance screening was performed and suitable genetic markers were selected. Based on these transformation protocols stably maintained plasmids were identified. A plasmid encoded oxygen-independent fluorescent system was established using the flavin mononucleotide-based fluorescent protein FbFP. Finally, a chromosomal gene knockout strategy was successfully employed for the inactivation of the anaerobic metabolism regulatory gene dnr from D. shibae DFL12T.ConclusionA genetic toolbox for members of the Roseobacter clade was established. This provides a solid methodical basis for the detailed elucidation of gene regulatory and metabolic networks underlying the ecological success of this group of marine bacteria.

Carbohydrate Metabolism and Carbon Fixation in Roseobacter denitrificans OCh114

The Roseobacter clade of aerobic marine proteobacteria, which compose 10–25% of the total marine bacterial community, has been reported to fix CO2, although it has not been determined what pathway is involved. In this study, we report the first metabolic studies on carbohydrate utilization, CO2 assimilation, and amino acid biosynthesis in the phototrophic Roseobacter clade bacterium Roseobacter denitrificans OCh114. We develop a new minimal medium containing defined carbon source(s), in which the requirements of yeast extract reported previously for the growth of R. denitrificans can be replaced by vitamin B12 (cyanocobalamin). Tracer experiments were carried out in R. denitrificans grown in a newly developed minimal medium containing isotopically labeled pyruvate, glucose or bicarbonate as a single carbon source or in combination. Through measurements of 13C-isotopomer labeling patterns in protein-derived amino acids, gene expression profiles, and enzymatic activity assays, we report that: (1) R. denitrificans uses the anaplerotic pathways mainly via the malic enzyme to fix 10–15% of protein carbon from CO2; (2) R. denitrificans employs the Entner-Doudoroff (ED) pathway for carbohydrate metabolism and the non-oxidative pentose phosphate pathway for the biosynthesis of histidine, ATP, and coenzymes; (3) the Embden-Meyerhof-Parnas (EMP, glycolysis) pathway is not active and the enzymatic activity of 6-phosphofructokinase (PFK) cannot be detected in R. denitrificans; and (4) isoleucine can be synthesized from both threonine-dependent (20% total flux) and citramalate-dependent (80% total flux) pathways using pyruvate as the sole carbon source.

Genetic Determinants ofSilicibactersp. TM1040 Motility

Silicibacter sp. TM1040 is a member of the marine Roseobacter clade of Alphaproteobacteria that forms symbioses with unicellular eukaryotic phytoplankton, such as dinoflagellates. The symbiosis is complex and involves a series of steps that physiologically change highly motile bacteria into cells that readily form biofilms on the surface of the host. The initial phases of symbiosis require bacterial motility and chemotaxis that drive the swimming bacteria toward their planktonic host. Cells lacking wild-type motility fail to establish biofilms on host cells and do not produce effective symbioses, emphasizing the importance of understanding the molecular mechanisms controlling flagellar biosynthesis and the biphasic "swim-or-stick" switch. In the present study, we used a combination of bioinformatic and genetic approaches to identify the genes critical for swimming of Silicibacter sp. TM1040. More than 40 open reading frames with homology to known flagellar structural and regulatory genes were identified, most of which are organized into approximately eight operons comprising a 35.4-kb locus, with surprising similarity to the fla2 locus of Rhodobacter sphaeroides. The genome has homologs of CckA, CtrA, FlbT, and FlaF, proteins that in Caulobacter crescentus regulate flagellum biosynthesis. In addition, we uncovered three novel genes, flaB, flaC, and flaD, which encode flagellar regulatory proteins whose functions are likely to involve regulation of motor function (FlaD) and modulation of the swim-or-stick switch (FlaC). The data support the conclusion that Silicibacter sp. TM1040 uses components found in other Alphaproteobacteria, as well as novel molecular mechanisms, to regulate the expression of the genes required for motility and biofilm formation. These unique molecular mechanisms may enhance the symbiosis and survival of Roseobacter clade bacteria in the marine environment.