"Pseudoalteromonas januaria" SUT 11 as the source of rare lipodepsipeptides.

The marine bacterium "Pseudoalteromonas januaria" SUT 11 isolated from a seawater sample produced the rare cell-bound cyclic lipodepsipeptides A/A', B/B', and C/C'. The matrix-assisted laser desorption/ionization mass spectra indicated that one bromine atom presented in the peptides B/B' and C/C', whereas the component A/A' contained no bromine atom. The acyldepsipeptides A/A'-C/C' have an identical amino acid sequence, Thr-Val-Asn-Asn-Leu/allo-Ile, but differed in C-terminal amino acid and acyl moieties. Peptides A-C have Leu as a C-terminal amino acid, whereas peptides A'-C' have allo-Ile. Acyl moieties in peptides A/A', B/B', and C/C' have been found to consist of 11-(4'-hydroxyphenyl)-undeca-2,4,6,8,10-pentaenic acid, 9-(3'-bromo-4'-hydroxyphenyl)-nona-2,4,6,8-tetraenic acid, and 11-(3'-bromo-4'-hydroxyphenyl)-undeca-2,4,6,8,10-pentaenic acid, respectively. The structure of a main pair of peptides B/B' with molecular masses 843/845 Da has been determined by means of ultraviolet, infrared, and two-dimensional nuclear magnetic resonance spectroscopy. We have demonstrated that tandem nano-electrospray ionization mass spectrometry is a very efficient way for the fast and sensitive investigation of lipopeptides A/A' and C/C' with molecular masses 791 and 869/871 Da, respectively, which have been isolated in small amounts.

ATP level variations in heterotrophic bacteria during attachment on hydrophilic and hydrophobic surfaces.

A survey of the extracellular ATP levels of 86 heterotrophic bacteria showed that gram-negative bacteria of the genera Sulfitobacter, Staleya, and Marinobacter secreted elevated amounts of extracellular ATP, ranging from 6.0 to 9.8 pM ATP/colony forming unit (cfu), and that gram-positive bacteria of the genera Kocuria and Planococcus secreted up to 4.1 pM ATP/cfu. Variations in the levels of extracellular and intracellular ATP-dependent luminescence were monitored in living cells of Sulfitobacter mediterraneus ATCC 700856T and Planococcus maritimus F 90 during 48 h of attachment on hydrophobic (poly[tert-butyl methacrylate], PtBMA) and hydrophilic (mica) surfaces. The bacteria responded to different polymeric surfaces by producing either intracellular or extracellular ATP. The level of intracellular ATP in S. mediterraneus ATCC 700856T attached to either surface was as high as 50-55 pM ATP/cfu, while in P. maritimus F 90 it was 120 and 250 pM ATP/cfu on PtBMA and mica, respectively. S. mediterraneus ATCC 700856T generated about 20 and 50 pM of extracellular ATP/cfu on PtBMA and mica, respectively, while the amount generated by P. maritimus F 90 was about the same for both surfaces, 6 pM ATP/cfu. The levels of extracellular ATP generated by S. mediterraneus during attachment on PtBMA and mica were two to five times higher than those detected during the initial screening. High-resolution atomic force microscopy imaging revealed a potentially interesting correlation between the porous cell-surface of certain alpha- and gamma-proteobacteria and their ability to secrete high amounts of ATP.

ATP level variations in heterotrophic bacteria during attachment on hydrophilic and hydrophobic surfaces / Variaciones en los niveles de ATP en bacterias heterotrofas durante la adherencia a superficies hidrofílicas e hidrofóbicas

A survey of the extracellular ATP levels of 86 heterotrophic bacteria showed that gram-negative bacteria of the genera Sulfitobacter, Staleya, and Marinobacter secreted elevated amounts of extracellular ATP, ranging from 6.0 to 9.8 pM ATP/colony forming unit (cfu), and that gram-positive bacteria of the genera Kocuria and Planococcus secreted up to 4.1 pM ATP/cfu. Variations in the levels of extracellular and intracellular ATP-dependent luminescence were monitored in living cells of Sulfitobacter mediterraneus ATCC 700856T and Planococcus maritimus F 90 during 48 h of attachment on hydrophobic (poly[tert-butyl methacrylate], PtBMA) and hydrophilic (mica) surfaces. The bacteria responded to different polymeric surfaces by producing either intracellular or extracellular ATP. The level of intracellular ATP in S. mediterraneus ATCC 700856T attached to either surface was as high as 50-55 pM ATP/cfu, while in P. maritimus F 90 it was 120 and 250 pM ATP/cfu on PtBMA and mica, respectively. S. mediterraneus ATCC 700856T generated about 20 and 50 pM of extracellular ATP/cfu on PtBMA and mica, respectively, while the amount generated by P. maritimus F 90 was about the same for both surfaces, 6 pM ATP/cfu. The levels of extracellular ATP generated by S. mediterraneus during attachment on PtBMA and mica were two to five times higher than those detected during the initial screening. High-resolution atomic force microscopy imaging revealed a potentially interesting correlation between the porous cell-surface of certain a- and g-proteobacteria and their ability to secrete high amounts of ATP (AU)

Un estudio de los niveles del ATP extracelular de 86 bacterias heterotrofas mostró que las bacterias gram-negativas de los géneros Sulfitobacter, Staleya y Marinobacter secretaron grandes cantidades de ATP extracelular 6,0 a 9,8 pM ATP/unidad formadora de colonia (cfu) y las bacterias gram-positivas de los géneros Kocuria y Planococcus secretaban cantidades de hasta 4,1 pM ATP/cfu. Las variaciones de los niveles extracelulares e intracelulares de luminiscencia dependiente de ATP fueron controlados en células vivas de Sulfitobacter mediterraneus ATCC 700856T y Planococcus maritimus F 90 durante 48 h de adherencia a superficies hidrofóbicas (poli[tert-butil metacrilato], PtBMA) e hidrofílicas (mica). Las bacterias respondieron a las diferentes superficies produciendo ATP intracelular o extracelular. El nivel de ATP intracelular en S. mediterraneus ATCC 700856T llegó hasta 50-55 pM ATP/cfu en ambas superficies, mientras que para P. maritimus F 90 fue de 120 pM ATP/cfu en PtBMA y de 250 pM ATP/cfu en mica. El ATP extracelular generado por las células de S. mediterraneus ATCC 700856T se situó entre 20 pM ATP/cfu en PtBMA y 50 pM ATP/cfu en mica, mientras que para P. maritimus F 90 era casi igual en ambas superficies, 6 pM ATP/cfu. Los niveles detectados de ATP extracelular generados por S. mediterraneus durante la adherencia a PtBMA y a mica fueron de 2 a 5 veces los detectados en el cribado inicial. Las imágenes obtenidas mediante microscopia de fuerza atómica de alta resolución pusieron de manifiesto una correlación de posible interés entre la superficie celular porosa de ciertas a- y g-proteobacterias y la capacidad de secretar grandes cantidades de ATP (AU)

Brevibacterium celere sp. nov., isolated from degraded thallus of a brown alga.

Two whitish yellow, Gram-positive, non-motile, aerobic bacteria were isolated from enrichment culture during degradation of the thallus of the brown alga Fucus evanescens. The bacteria studied were chemo-organotrophic, mesophilic and grew well on nutrient media containing up to 15 % (w/v) NaCl. The DNA G+C content was 61 mol%. The two isolates exhibited a conspecific DNA-DNA relatedness value of 98 %, indicating that they belong to the same species. A comparative analysis of 16S rRNA gene sequences revealed that strain KMM 3637(T) formed a distinct phyletic lineage in the genus Brevibacterium (family Brevibacteriaceae, class Actinobacteria) and showed the highest sequence similarity (about 97 %) to Brevibacterium casei. DNA-DNA hybridization experiments demonstrated 45 % binding with the DNA of B. casei DSM 20657(T). Physiological and chemotaxonomic characteristics (meso-diaminopimelic acid in the peptidoglycan, major cellular fatty acids 15 : 0ai and 17 : 0ai) of the bacteria studied were consistent with the genomic and phylogenetic data. On the basis of the results of this study, a novel species, Brevibacterium celere sp. nov., is proposed. The type strain is KMM 3637(T) (=DSM 15453(T)=ATCC BAA-809(T)).

Bacillus algicola sp. nov., a novel filamentous organism isolated from brown alga Fucus evanescens.

A slightly yellowish, Gram-positive, filamentous with 'cross-like' branching, aerobic, spore-forming bacterium was isolated from enrichment culture during degradation of the thallus of the brown alga Fucus evanescens. The bacterium studied was chemoorganotrophic, tolerant to 3% NaCl, alkalitolerant, and alginolytic. The predominant cellular fatty acid was ai15:0 which accounted more than 65% of total fatty acids, while i14:0, il5:0 i16:0, and ai17:0 made up 25%. DNA base composition was 37 mol% GC. Phylogenetic analysis of 16S rDNA gene revealed that this isolate was a member of the genus Bacillus, with no close relatives at the species level (16S rRNA gene sequence similarity less 97%). On the basis of the significant differences demonstrated in the phenotypic and chemotaxonomic characteristics, it is suggested that the bacterium be classified as a novel species; the name Bacillus algicola sp. nov. is proposed. The type strain is KMM 3737T (= CIP 107850T).

Low-molecular-weight, biologically active compounds from marine Pseudoalteromonas species.

We have examined the ability of marine Proteobacteria from the Pseudoalteromonas genus and Alteromonas macleodii to produce low-molecular-weight, biologically active compounds with antimicrobial and surface-active properties. A new marine bacterium, Pseudoalteromonas issachenkonii, exhibited a high level of biological activity and produced antifungal and hemolytic compounds. A detailed spectroscopic investigation based on UV, IR, high-resolution mass spectrometry, and 2D 1H and 13C nuclear magnetic resonance revealed that the former was indole-2,3-dione (isatin). The chemical structure of red-brown pigment (C9H7N3OS3) responsible for hemolytic activity remained unclear. Four of the 15 strains studied (P. luteoviolacea, P. rubra, P. undina, and P. issachenkonii) produced cell-bound, two (P. elaykovii and P. carrageenovora) produced extracellular, and one strain (P. citrea) produced cell-bound and extracellular fatty acids and phospholipids with surface activity. Neither peptides nor glycolipids with surface activity were detected.

Formosa algae gen. nov., sp. nov., a novel member of the family Flavobacteriaceae.

Four light-yellow-pigmented, Gram-negative, short-rod-shaped, non-motile isolates were obtained from enrichment culture during degradation of the thallus of the brown alga Fucus evanescens. The isolates studied were chemo-organotrophic, alkalitolerant and mesophilic. Polar lipids were analysed and phosphatidylethanolamine was the only phospholipid identified. The predominant cellular fatty acids were 15 : 0, i15 : 0, ai15 : 0, i15 : 1 and 15 : 1(n-6). The DNA G+C contents of the four strains were 34.0-34.4 mol%. The level of DNA relatedness of the four isolates was conspecific (88-98 %), indicating that they belong to the same species. The 16S rDNA sequence of strain KMM 3553(T) was determined. Phylogenetic analysis revealed that KMM 3553(T) formed a distinct phyletic line in the phylum Bacteroidetes, class Flavobacteria in the family Flavobacteriaceae and that, phylogenetically, this strain could be placed almost equidistant from the genera Gelidibacter and Psychroserpens (16S rRNA gene sequence similarities of 94 %). On the basis of significant differences in phenotypic and chemotaxonomic characteristics, it is suggested that the isolates represent a novel species in a new genus; the name Formosa algae gen. nov., sp. nov. is proposed. The type strain is KMM 3553(T) (=CIP 107684(T)).

Sulfitobacter delicatus sp. nov. and Sulfitobacter dubius sp. nov., respectively from a starfish (Stellaster equestris) and sea grass (Zostera marina).

On the basis of data from phenotypic and genotypic characterization and analysis of 16S rRNA gene sequences, two novel species belonging to the genus Sulfitobacter are described. Strains KMM 3584(T), a pale-yellowish, non-motile strain isolated from a starfish (Stellaster equestris), and KMM 3554(T), which is motile by means of a single subpolar flagellum and was isolated from sea grass (Zostera marina), are marine, Gram-negative, aerobic, rod-shaped organisms. Both strains have the ability to degrade gelatin, but not casein, chitin, agar, DNA, Tween 80 or starch. Strain KMM 3584(T) decomposed alginate and grew at NaCl concentrations of 1-8 % and temperatures of 12-37 degrees C, whereas strain KMM 3554(T) grew in 1-12 % NaCl and at temperatures of 10-30 degrees C. The predominant fatty acid was 18 : 1omega7, amounting to up to 80 % of the total fatty acids. The other characteristic feature was the presence of 18 : 2 isomers. The DNA G+C contents of KMM 3584(T) and KMM 3554(T) were respectively 60.0 and 63.7 mol%. The level of DNA similarity between the two strains was 33 %. DNA from KMM 3584(T) and KMM 3554(T) had hybridization values of 5-24 % and 10-41 %, respectively, with DNA from the type strains of Sulfitobacter pontiacus, Sulfitobacter brevis, Sulfitobacter mediterraneus and Staleya guttiformis. It is proposed that strains KMM 3584(T) (=LMG 20554(T)=ATCC BAA-321(T)) and KMM 3554(T) (=LMG 20555(T)=ATCC BAA-320(T)) represent two novel species, Sulfitobacter delicatus sp. nov. and Sulfitobacter dubius sp. nov., respectively.

Marinobacter excellens sp. nov., isolated from sediments of the Sea of Japan.

Five strains of halophilic, Gram-negative marine bacteria (KMM 3809(T), KMM 3814, KMM 3815, KMM 3817 and KMM 3818) were isolated from sediments collected from Chazhma Bay, Sea of Japan. Phylogenetic 16S rRNA gene sequence-based analysis placed these bacteria in a clade within the genus Marinobacter in the gamma-Proteobacteria. KMM 3809(T) showed highest 16S rRNA gene sequence similarity of 97.3 % to Marinobacter litoralis and 96.9 % to Marinobacter hydrocarbonoclasticus and Marinobacter aquaeolei. DNA-DNA hybridization between the five isolates was at the conspecific level (94-96 %) and that among the closest phylogenetic neighbours ranged from 45.0 to 62.5 %. The new organisms were susceptible to polymyxin. Predominant fatty acids were C(16 : 0), C(16 : 1)omega9c, C(16 : 1)omega7c and C(18 : 1)omega9c. Phylogenetic evidence, along with phenotypic and genotypic characteristics, showed that the bacteria constituted a novel species of the genus Marinobacter. The name Marinobacter excellens sp. nov. is proposed for this species, with the type strain KMM 3809(T) (=CIP 107686(T)).

Ecophysiological variabilities in ectohydrolytic enzyme activities of some Pseudoalteromonas species, P. citrea, P. issachenkonii, and P. nigrifaciens.

The ecophysiological variabilities in the ectohydrolytic enzyme profiles of the three species of Pseudoalteromonas, P. citrea, P. issachenkonii, and P. nigrifaciens, have been investigated. Forty-one bacteria isolated from several invertebrates, macroalgae, sea grass, and the surrounding water exhibited different patterns of hydrolytic enzyme activities measured as the hydrolysis of either native biopolymers or fluorogenic substrates. The activities of the following enzymes were assayed: proteinase, tyrosinase, lipase, amylase, chitinase, agarase, fucoidan hydrolase, laminaranase, alginase, pustulanase, cellulase, beta-glucosidase, alpha- and beta-galactosidases, beta-N-acetylglucosaminidase, beta-glucosaminidase, beta-xylosidase, and alpha-mannosidase. The occurrence and cell-specific activities of all enzymes varied over a broad range (from 0 to 44 micromol EU per hour) and depended not only on taxonomic affiliation of the strain, but also on the source/place of its isolation. This suggests 'specialization' of different species for different types of polymeric substrates as, for example, all strains of P. citrea and P. issachenkonii hydrolyzed alginate and laminaran, while strains of P. nigrifaciens were lacking the ability to hydrolyze most of the algal polysaccharides. The incidence of certain enzymes such as fucoidan hydrolases, alginate lyases, agarases, and alpha-galactosidases might be strain specific and reflect its particular ecological habitat.