[Lipid composition of novel Shewanella species isolated from far Eastern seas].

A comparative study of the lipid composition of 26 strains (including type strains) of marine Gammaproteobacteria belonging to the genera Shewanella, Alteromonas, Pseudoalteromonas, Marinobacterium, Microbulbifer, and Marinobacter was carried out. The bacteria exhibited genus-specific profiles of ubiquinones, phospholipids, and fatty acids, which can serve as reliable chemotaxonomic markers for tentative identification of new isolates. The studied species of the genus Shewanella were distinguished by the presence of two types of isoprenoid quinones, namely, ubiquinones Q-7 and Q-8 and menaquinones MK-7 and MMK-7; five phospholipids typical of this genus, namely, phosphatidylethanolamine (PE), phosphatidylglycerol (PG), diphosphatidylglycerol (DPG), lyso-PE, and acyl-PG; and the fatty acids 15:0, 16:0, 16:1 (n-7), 17:1 (n-8), i-13:0, and i-15:0. The high level of branched fatty acids (38-45%) and the presence of eicosapentaenoic acid (4%) may serve as criteria for the identification of this genus. Unlike Shewanella spp., bacteria of the other genera contained a single type of isoprenoid quinone: Q-8 (Alteromonas, Pseudoalteromonas, Marinobacterium, and Microbulbifer) or Q-9 (Marinobacter). The phospholipid compositions of these bacteria were restricted to three components: two major phospholipids (PE and PG) and a minor phospholipid, bisphosphatidic acid (Alteromonas and Pseudoalteromonas) or DPG (Marinobacterium, Microbulbifer, and Marinobacter). The bacteria exhibited genus-specific profiles of fatty acids.

[Amylases of the fungus Aspergillus flavipes associated with Fucus evanescens]. / Amilazy griba Aspergillus flavipes, assotsiirovannogo s Fucus evanescens.

A promising producer of extracellular amylases, Aspergillus flavipes, was selected from 245 strains of marine fungi. Depending on the conditions of growth, this strain produced diverse amylolytic complexes. When grown on medium containing peptone and yeast extract (pH 7.0), A. flavipes synthesized three forms of amylase, differing in pH optimum (5.5, 6.0, and 7.5). A single form of the enzyme was synthesized either in the absence of peptone from the medium or at the initial pH value of the medium, equal to 8.6. The activity of the isolated amylase forms decreased in the presence of proteolytic enzymes. New, highly stable forms of amylase (with pH optima of 5.5 and 7.5 and maximum activity at 60-80 degrees C) were synthesized in the presence of diisopropyl fluorophosphate, an inhibitor of proteases.

Retrieval of the species Alteromonas tetraodonis Simidu et al. 1990 as Pseudoalteromonas tetraodonis comb. nov. and emendation of description.

A polyphasic taxonomy study was undertaken of three strains of Pseudoalteromonas haloplanktis subsp. tetraodonis (Simidu et al. 1990) Gauthier et al. 1995. DNA was prepared from each of the strains and genomic relatedness was measured by DNA-DNA hybridization. Strains KMM 458T and IAM 14160T shared 99% genetic relatedness, but were only 48-49% related to the type strain of Pseudoalteromonas haloplanktis subsp. haloplanktis, IAM 12915T. The third strain, P. haloplanktis subsp. tetraodonis A-M, showed 83% genetic similarity with P. haloplanktis subsp. haloplanktis IAM 12915T and 32% with KMM 458T. From these results, it is concluded that strains KMM 458T and IAM 14160T comprise a separate species, originally described as Alteromonas tetraodonis, whereas strain A-M belongs to the species Pseudoalteromonas haloplanktis. Based on phenotypic and chemotaxonomic data, genomic fingerprint patterns, DNA-DNA hybridization data and phylogenetic analysis of 16S rRNA, it is proposed that the species Alteromonas tetraodonis be retrieved and recognized as Pseudoalteromonas tetraodonis comb. nov. (type strain IAM 14160T = KMM 458T).

Arenibacter gen. nov., new genus of the family flavobacteriaceae and description of a new species, Arenibacter latericius sp. nov.

Five dark-orange-pigmented, Gram-negative, rod-shaped, non-motile, aerobic bacterial strains were isolated from sandy sediment samples collected in the South China Sea in the Indian Ocean, from a holothurian, Apostichopus japonicus, in the Sea of Japan and from a brown alga, Chorda filum, from the Sea of Okhotsk in the Pacific Ocean. Phenotypic data were collected, demonstrating that the bacteria are chemo-organotrophic and require seawater-based media for growth. Polar lipids were analysed and 27% of the total extract comprised phosphatidylethanolamine as the major component. The predominant cellular fatty acids were branched-chain saturated and unsaturated [i-C15:0, i-C15:1, a-C15:0, C15:0, C16:1(n-7)]. The DNA base composition was 37.5-38.2 mol % G+C. The level of DNA homology of the five isolates was 83-94%, indicating that these isolates belong to the same species. A 16S rDNA sequence of the type strain KMM 426T was determined and phylogenetic analysis, based on neighbour-joining and Fitch-Margoliash methods, revealed that the type strain formed a distinct phyletic line in a clade corresponding to the family Flavobacteriaceae and represented a new genus. From the results of this polyphasic taxonomic analysis, it is proposed that the bacterial strains be classified in a new genus, Arenibacter gen. nov., and species, Arenibacter latericius sp. nov. The type strain is KMM 426T (VKM B 2137DT = LMG 19694T = CIP 106861T).

Evaluation of phospholipid and fatty acid compositions as chemotaxonomic markers of Alteromonas-like proteobacteria.

The cellular phospholipids (PLs) and fatty acids (FAs) were investigated in type and environmental strains of Pseudoalteromonas, Alteromonas macleodii, A. infernus, and in three type strains of Marinomonas, M. communis, M. vaga, M. mediterranea. A total of 40 strains (19 strains in this study and 21 reported in previous papers), including Idiomarina abyssalis, I. zobellii, and Glaciecola punicea, G. pallidula, aerobic Alteromonas-like proteobacteria showed genus-characteristic patterns of phospholipids and fatty acids useful for genera discrimination. The PL patterns of surface cultures of alteromonads, pseudoalteromonads, and marinomonads consisted almost entirely of phosphatidyl ethanolamine and phosphatidyl glycerol presented in different proportions. Neither diphosphatidyl glycerol nor glycophospholipids were found in bacteria studied. In addition, the minor amount of a glycolipid was found in all strains studied. Bacteria of the genera Marinomonas, Idiomarina, and Glaciecola were clearly distinguished by presence of one of the major FAs: 18:1 (n-7), i15:0, and 16:1 (n-7), respectively. The amounts of these FAs reached up to 40-60% of total FAs. Members of Alteromonas and Pseudoalteromonas were characterized by different ratio of the following major FAs:16:1(n-7), 16:0, 17:1 (n-8), and 18:1 (n-7).

[Features of phospholipid composition of marine proteobacteria of Pseudoalteromonas species]. / Osobennosti fosfolipidnogo sostava morskikh proteobakterii roda Pseudoalteromonas.

The study of the phospholipid composition of 14 type strains of marine proteobacteria of the genus Pseudoalteromonas showed that phospholipids are the main polar lipid constituents of membranes in these proteobacteria. The phospholipid patterns of the strains studied were found to be similar and involved five phospholipids typical of gram-negative bacteria, namely, phosphatidylethanolamine, phosphatidylglycerol, bisphosphatidic acid, lysophosphatidylethanolamine, and phosphatidic acid. The major phospholipids were phosphatidylethanolamine and phosphatidylglycerol, which add up to 89-97% of total phospholipids; bisphosphatidic acid was dominant among minor phospholipids. The prevalence of phosphatidylethanolamine (62-77% of total phospholipids) and the absence of diphosphatidylglycerol are the characteristic features of most bacteria of this genus. As in Escherichia coli, the phospholipid composition of the marine proteobacteria depended on the presence of magnesium in the medium.

[Immunoenzyme method for detecting microbial producers of palytoxin]. / Immunofermentnyi metod obnaruzheniia mikrobnogo produtsenta palitoksina.

A competitive ELISA using the intact toxin as a coating antigen for detecting palytoxin was developed. This immunoassay allows palytoxin (PTX) to be determined in the range of 6-250 ng/ml. In sensitivity, this determination is comparable with RIA but is three times inferior to ELISA using monoclonal antibodies. Inhibition experiments using some toxins of marine invertebrates proved the serological specificity of the palytoxin binding to antibodies. Both the indirect and competitive ELISA were used to find PTX-producing bacteria among 420 isolates of sea bacteria. It was found that gram-negative bacteria Aeromonas sp. and Vibrio sp. associated with toxic samples of the soft coral Palythoa sp. produced compounds antigenically related to PTX.

Reclassification of Alteromonas distincta Romanenko et al. 1995 as Pseudoalteromonas distincta comb. nov.

The 16S rRNA gene of Alteromonas distincta KMM 638T was amplified, cloned and sequenced. The nucleotide sequence was aligned with sequences of representative strains of Alteromonas, Moritella, Pseudoalteromonas and Shewanella. Results of phylogenetic analysis, using neighbour-joining and Fitch-Margoliash methods, clearly indicated that this species should be assigned to the genus Pseudoalteromonas. On the basis of polyphasic data obtained from previous work and this study, it is proposed that the species Alteromonas distincta be reclassified as Pseudoalteromonas distincta comb. nov. with type strain KMM 638T (= ATCC 700518T).

[The temperature-induced early and late development of resistance to the bactericidal action of blood serum in Yersinia pseudotuberculosis]. / Indutsirovannoe temperaturoi rannee i pozdnee razvitie ustoichivosti k bakteritsidnomu deistviiu syvorotki krovi u Yersinia pseudotuberculosis.

Y. pseudotuberculosis strains were grown at 6 degrees-8 degrees C and then incubated at 37 degrees C. 3-6 hours later serum resistance appeared in the strains having plasmid virulence and producing outer membrane polypeptide with a molecular weight of 120 kD, known as P1. 10-12 hours later serum resistance appeared in the strain having the virulence plasmid, but not producing P1, as well as in strains in which the plasmid was eliminated.

[Features of a protein component of the endotoxin from Yersinia pseudotuberculosis]. / Kharakteristika belkovogo komponenta éndotoksina iz Yersinia pseudotuberculosis.

The protein moiety of endotoxin from Yersinia pseudotuberculosis was found to consist of two polypeptides with apparent molecular masses 40 and 14.5 kDa (4:1 w/w). The major protein (40 kDa) was isolated from the endotoxin pretreated with sodium deoxy cholate by gel chromatography on the Sephadex G-200 column. Comparative study of this protein and oligomeric form of porin from the outer membrane of Y. pseudotuberculosis using SDS--PAGE, velocity sedimentation, lipid bilayer experiments, chemical and serological analyses revealed their identity. The deoxycholate treatment of the endotoxin does not affect complexes of the major protein and LPS.

[Conformational stability and immunochemical properties of yersinin--a basic protein of the outer membrane of the Pseudotuberculosis microbe]. / Konformatsionnaia stabil'nost' i immunokhimicheskie svoistva iersinina--osnovnogo belka vneshnei membrany psevotuberkuleznogo mikroba.

Using spectroscopic methods (circular dichroism and intrinsic protein fluorescence) and immunoenzyme assay, changes in the spatial and antigenic structure of yersinin, porin from outer membrane of Yersinia pseudotuberculosis, were studied in solutions of ionic and non-ionic detergents at various temperatures and low pH values. Yersinin was shown to retain its secondary structure under various denaturation conditions, the content of regular structural patterns depending on specific action of the denaturation agent. Process of yersinin denaturation similarly to other membrane proteins appears to occur via two structural transitions: dissociation of oligomers and denaturation of monomers. At the first stage changes of quaternary structure accompanied by the loss of so called conformational determinants were observed. Temperature-dependent changes of monomers' tertiary structure affect antigenic activity of yersinin in a smaller degree.

[Immunochemical study of the lipopolysaccharides of Yersinia enterocolitica serovars O5 and O5.27]. / Immunokhimicheskoe izuchenie lipopolisakharidov Yersinia enterocolitica serovarov O5 i O5,27.

Y. enterocolitica lipopolysaccharides (LPS), serovars O5 and O5.27, have similar antigenic specificity. The LPS of serovar O5.27 has been shown to contain no factor O27. Residual alpha-L-rhamnose, glycosylated by D-xylulose, has been found to play an important role in the formation of factor O5, common for both serovars.

[The structure of O-specific polysaccharide isolated from the lipopolysaccharide of Yersinia intermedia strain 180]. / Struktura O-spetsificheskogo polisakharida, vydelennogo iz lipopolisakharida Yersinia intermedia shtamma 680.

An O-specific polysaccharide from lipopolysaccharide of Yersinia intermedia pathogenic strain 680 has been isolated and shown to be a serologically active fructane. Serological specificity of the lipopolysaccharide and the fructane was studied by reactions of precipitation and of inhibition of passive hemolysis. On the basis of methylation studies, 13C NMR spectroscopy, and immunochemical data the following structure was proposed for the repeating unit of the O-specific polysaccharide: (Formula: see text)

[Different forms of a lipopolysaccharide-protein complex from Yersinia pseudotuberculosis]. / O razlichnykh formakh lipopolisakharid-belkovogo kompleksa iz Yersinia pseudotuberculosis.

Two forms of lipopolysaccharide-protein complex with buoyant densities of 1,43 and 1,40 g/cm3 were found in the Yersinia pseudotuberculosis cell wall. These forms have the similar monosaccharide, fatty acid and polypeptide compositions, but differ in the length of O-specific chains. The differences in density are stipulated by the different contents of the main components of the complex. Both forms contain the related antigenic determinants but have some differences in the antigenic structure. The ability of the two forms to produce a hybrid form with the intermediate density of 1,41 g/cm3 has been shown.

Studies on a lipopolysaccharide-protein complex from Yersinia pseudotuberculosis. 1 isolation and characterization.

A comparative study of various procedures of a lipopolysaccharide-protein complex (LPPC) from Yersinia pseudotuberculosis was carried out. The materials obtained were fractionated by molecular-sieve chromatography on Sepharose 2B resulting in highly aggregated complexes with antigen activity. LPPC aggregates dissociated in the presence of sodium dodecylsulphate (SDS) and urea. The chemical composition and serologic properties of fractions obtained are under consideration. The protein component of the complex consists of two major polypeptides (molecular weights--45,000 and 20,000) and some minor ones. The LPS component appeared to give 2--3 narrow bands in gel under conditions of SDS-polyacrylamide gel electrophoresis. It is suggested that such fractionation is caused by LPS association-dissociation in the course of electrophoresis.