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Structural analysis of sphingophospholipids derived from Sphingobacterium spiritivorum, the type species of genus Sphingobacterium.

The unique feature of the genus Sphingobacterium is the presence of sphingophospholipids and ceramides, besides diacylglycerophospholipids. As major cellular lipid components, five kinds of sphingophospholipids were purified from Sphingobacterium spiritivorum ATCC 33861(T), the type species of genus Sphingobacterium. They were identified as ceramide phosphorylethanolamines (CerPE-1 and CerPE-2), ceramide phosphoryl-myo-inositols (CerPI-1 and CerPI-2), and ceramide phosphorylmannose (CerPM-1). The ceramide of CerPE-1, CerPI-1, and CerPM-1 was composed of 15-methylhexadecasphinganine (isoheptadeca sphinganine, iso-C17:0) and 13-methyltetradecanoic acid (isopentadecanoic acid, iso-C15:0), whereas that of CerPE-2 and CerPI-2 was composed of isoheptadeca sphinganine and 2-hydroxy-13-methyltetradecanoic acid (2-hydroxy isopentadecanoic acid, 2-OH iso-C15:0). These sphingophospholipids were also found in cellular lipids of Sphingobacterium multivorum ATCC 33613(T), Sphingobacterium mizutaii ATCC 33299(T), Sphingobacterium faecium IFO 15299(T), Sphingobacterium thalpophilum ATCC 43320(T), and Sphingobacterium antarcticum ATCC 51969(T). To our knowledge, the existence of CerPM-1 is a novel sphingophospholipid through eukaryotic and prokaryotic cells.

Ceramides↗

Carotenoids of an Antarctic psychrotolerant bacterium, Sphingobacterium antarcticus, and a mesophilic bacterium, Sphingobacterium multivorum.

The major carotenoid pigments of an Antarctic psychrotolerant bacterium, Sphingobacterium antarcticus, and a mesophilic bacterium, Sphingobacterium multivorum, were identified as zeaxanthin, beta-cryptoxanthin, and beta-carotene. Analysis was based on ultraviolet-visible spectroscopy, mass spectroscopy, and reversed-phase HPLC. Photoacoustic spectroscopy of intact bacterial cells revealed that the bulk of the pigments in S. antarcticus and S. multivorum was associated with the cell membrane. In vitro studies with synthetic membranes of phosphatidylcholine demonstrated that the major pigment was bound to the membranes and decreased their fluidity. The relative amounts of polar pigments were higher in cells grown at 5 degrees C than in cells grown at 25 degrees C. In the mesophilic strain, the synthesis of polar carotenoids was quantitatively less than that of the psychrotolerant strain.

Antarctic Regions↗

Sphingobacterium daejeonense sp. nov., isolated from a compost sample.

A Gram-negative, strictly aerobic, rod-shaped, non-motile, non-spore-forming bacterial strain, designated TR6-04(T), was isolated from compost and characterized taxonomically by using a polyphasic approach. The organism grew optimally at 30 degrees Celsius and at pH 6.5-7.0. The isolate was positive for catalase and oxidase tests but negative for gelatinase, indole and H(2)S production. Comparative 16S rRNA gene sequence analysis showed that strain TR6-04(T) fell within the radiation of the cluster comprising Sphingobacterium species and clustered with Sphingobacterium mizutaii ATCC 33299(T) (96.7 % sequence similarity); the similarity to sequences of other species within the family Sphingobacteriaceae was less than 92.0 %. The G+C content of the genomic DNA was 38.7 mol%. The predominant respiratory quinone was MK-7. The major fatty acids were iso-C(15 : 0), iso-C(17 : 0) 3-OH and summed feature 4 (iso-C(15 : 0) 2-OH and/or C(16 : 1)omega7c). These chemotaxonomic data supported the affiliation of strain TR6-04(T) to the genus Sphingobacterium. However, on the basis of its phenotypic properties and phylogenetic distinctiveness, strain TR6-04(T) (=KCTC 12579(T)=LMG 23402(T)=CCUG 52468(T)) should be classified as the type strain of a novel species, for which the name Sphingobacterium daejeonense sp. nov. is proposed.

Bacterial Typing Techniques↗

Thioglucosidase activity from Sphingobacterium sp. strain OTG1.

Screening for novel thioglucoside hydrolase activity resulted in the isolation of Sphingobacterium sp. strain OTG1 from enrichment cultures containing octylthioglucoside (OTG). OTG was hydrolysed into octanethiol and glucose by cell free extracts. Besides thioglucoside hydrolysis, several other glucoside hydrolase activities were detected in the Sphingobacterium sp. strain OTG1 cell free extract. By adding beta-glucosidase inhibitors it was possible to discriminate between these different activities. Ascorbic acid and D-gluconic acid lactone inhibited the hydrolysis of p-nitrophenyl beta-glucoside, but did not affect octyl- and octylthioglucoside hydrolase activity. Besides OTG, various other thioglucosides were hydrolysed by the novel thioglucosidase, with almost the same activities regardless of the nature of the aglycone, including the myrosinase model substrate sinigrin (a glucosinolate). Sinigrin could also be used as a growth substrate by Sphingobacterium sp. strain OTG1, although at concentrations exceeding 0.15 mM degradation was not complete.

Bacteria↗

Bacteremia caused by a novel species of Sphingobacterium.

A Gram-negative rod was isolated from the blood cultures of an 84-year-old man with foot cellulitis. The bacterium was first identified as Sphingobacterium spiritivorum on the basis of standard assimilation tests. However, sequencing analysis of its 16S rRNA genes and whole genome hybridization studies with other related bacteria showed that this isolate belongs to a so far undescribed species of Sphingobacterium, close to S. mizutae. This bacterium was susceptible to most of the antibiotics tested, including glycopeptides, but was resistant to aminoglycosides and polymyxins. Treatment with amoxicillin-clavulanate cured the infection.

Aged↗

Polyamine distributions in the Falvobacterium-Cytophaga-Sphingobacterium complex.

Homospermidine was found as the major polyamine in one newly described species of Flavobacterium (F. indologenes), in three species of Sphingobacterium (S. mizutae, S. multivorium, and S. spiritivorum), and in 10 species of Cytophaga (C. aquatilis, C. arvensicola, C. heparina, C. hutchinsonii, C. johnsonae, "C. keratolytica," C. lytica, C. marinoflava, C. uliginosa, and "C. xantha"). These bacteria also all contain putrescine and agmatine as minor components. Flavobacterium indologenes and C. johnsonae contain an unusual diamine, 2-hydroxyputrescine, as a major polyamine. The polyamine distributions of four other species originally included in Flavobacterium (F. acidurans, "F. dormitator," "F. tirrenicum," and Halomonas halmophila), whose taxonomic positions are or were uncertain, were different from the group mentioned above. They either contain spermidine as the major polyamine or lack any polyamine. These results suggest that homospermidine can serve as a chemotaxonomic marker to delineate true members of the Flavobacterium-Cytophaga-Sphingobacterium complex.

Bacteria↗

Polyamine distribution profiles in newly validated genera and species within the Flavobacterium-Flexibacter-Cytophaga-Sphingobacterium complex.

Cellular polyamines of 58 strains belonging to the Flavobacterium-Flexibacter-Cytophaga-Sphingobacterium complex were analysed by HPLC. Homospermidine was found in all species of Flavobacterium, Chryseobacterium, Empedobacter, Myroides, Cellulophaga, Salegentibacter, Psychroserpens and Gelidibacter of the family Flavobacteriaceae. Flavobacterium ferrugineum located outside of this family also contained homospermidine. Cytophaga fermentans and C. xylanolytica belonging to the family Bacteroidaceae contained spermidine. Cytophaga marinoflava and C. latercula belonging to Flavobacteriaceae contained homospermidine. The Cytophaga hutchinsonii/C. aurantiaca group contained homospermidine which was the major polyamine in Flexibacter maritimus/ F. ovolyticus of the family Flavobacteriaceae. The Flexibacter sancti/F filiformis/ Cytophaga arvensicola group, F. elegans, F. ruber, F. canadensis, F. flexilis and F. tractuosus, were located separately in different six clusters, and contained homospermidine. The Flexibacter litoralis/F. polymorphus/F. aggregans group contained spermidine, which was detected in Flexibacter roseolus belonging to a divergent cluster. Sphingobacterium and Pedobacter species of the family Sphingobacteriaceae contained homospermidine. Polyamine profiles serve, as a phenotypic chemotaxonomic marker, for the classification of this complex.

Bacteria↗

[The effect of cefepime on ceftazidime, cefotaxime and imipenem resistant strains of Acinetobacter, Xanthomonas, Pseudomonas, Flavobacterium, Sphingobacterium and on producers of extended spectrum beta-lactamases (ESBL) with resistance transfer].

BACKGROUND: The new cephalosporins with heterocyclic positively charged substituent at position C3 belong to the 4th generation cephalosporins. It is not evident, whereas these antibiotics are effective against cefotaxime-ceftazidime-resistant bacteria. We investigated the efficiency of the cefepime (Maxipime, BMS) against Acinetobacter sp., X. maltophilia, Pseudomonas sp., Flavobacterium sp., Sphingobacterium sp. and against strains with production of ESBL (Extended Spectrum beta-Lactamases) (Enterobacter sp., Citrobacter sp., Klebsiella sp.). METHODS AND RESULTS: We determined the susceptibility of the 54 strains resistant to 50 mg/l and more of ceftazidime. The majority of the strains (35) are susceptible to cefepime and MIC is the range 1.65-3.15 mg/l of cefepime. 17 strains have MIC = 6.25 mg/l. Two strains (one strain P. aeruginosa a one strain K. pneumoniae) were resistant to cefepime and MIC = 12.5 mg/l. The activity of beta-lactamases was determined by quantitative macroiodometric method in one strain of P. cepacia and two strains of Sphingobacterium multivorum. They hydrolyse actively imipenem (they produce metallo-beta-lactamase) and ceftazidime (they are highly resistant to this antibiotic), but cefepime is more slowly hydrolysed than ceftazidime. CONCLUSIONS: At the present time cefepime is effective against ceftazidime-resistant strains in so-called "problematic" species of bacteria. However, it is important to protect its efficiency in the future, this requires its rational use.

Acinetobacter↗

Bioconversion of isopropanol by a solvent tolerant Sphingobacterium mizutae strain.

The bioconversion of high concentration isopropanol (2-propanol, IPA) was investigated by a solvent tolerant strain of bacteria, which was identified as Sphingobacterium mizutae ST2 by partial 16S rDNA gene sequencing. This strain of bacteria exhibited the ability to utilise high concentration isopropanol as the sole carbon source, with mineralization occurring via an acetone intermediate into central metabolism. The biodegradative performance of this strain for IPA was examined over a 2-38 g l(-1) concentration range, using specific growth rate (mu) and conversion rate analysis. Maximum specific growth rates (mu(max)) of 0.0045 h(-1 )were routinely obtainable on IPA. In addition, the highest specific IPA degradation rate was obtained at a concentration of 7.5 g l(-1) with a corresponding value of 0.045 g IPA g cells(-1) h(-1). While the highest acetone yield reached its maximum value of 0.940 g acetone g IPA(-1) at 7.5 g IPA l(-1). This is the first report on bioconversion of isopropanol at such high concentration by this solvent tolerant strain of S. mizutae and may allow its application in novel biocatalytic processes for effective biological conversion in two-phase solvent systems.

2-Propanol↗

Degradation characteristics and metabolic pathway of 17alpha-ethynylestradiol by Sphingobacterium sp. JCR5.

A 17alpha-ethynylestradiol (EE2)-degrading bacterium was isolated from the activated sludge of the wastewater treatment plant (WWTP) of an oral contraceptives producing factory in Beijing, China. On the basis of its morphology, biochemical properties and the 16S rDNA sequence analysis, this strain was identified as Sphingobacterium sp. JCR5. This strain grew on EE2 as sole source of carbon and energy, and metabolized up to 87% of the substrate added (30 mgl-1) within 10 d at 30 degrees C. In addition to EE2 the strain could be cultivated on steroidal estrogens like estrone (E1), 17beta-estradiol (E2), estriol (E3) and mestranol (MeEE2), the intermediates of contraceptive medicine processing and on some aromatic compounds. Mass spectrum analysis of the EE2 degradation showed that in the first step it is oxygenized to E1, 2-hydroxy-2,4-dienevaleric acid and 2-hydroxy-2,4-diene-1,6-dioic acid, which are the main catabolic intermediates. The former was analogous to the pathway of a previously reported testosterone-degrading bacterium Comamonas testosteroni TA441 and the latter is a metabolite with a different cleavage position of 3-hydroxy-4,5-9,10-disecoestrane-1(10),2-diene-5,9,17-trione-4-oic acid from the former.

Ethinyl Estradiol↗

Rapid purification, characterization and substrate specificity of heparinase from a novel species of Sphingobacterium.

A type of heparinase (heparin lysase, no EC number) was isolated from the periplasmic space of a novel species of Sphingobacterium by three-step osmotic shock. It was further purified to apparent homogeneity by a combination of SP-sepharose and Source 30S chromatographies with a final specific activity of 17.6 IU/mg protein and purification factor of 13-fold. MALDI-TOF mass spectrum of the purified heparinase gave a molecular mass of 75,674 Da of the native enzyme. Peptide mass spectrum showed poor homogeneity with the database in the peptide bank. Inhibition of the enzyme activity by N-acetylimidazole indicated that tyrosine residues were necessary for enzyme activity. K(m) and V(max) of the heparinase for de-o-sulfated-N-acetyl heparin were 42 micro M and 166 microM/min/mg protein, respectively. The heparinase showed similar activity on both heparin and heparan sulfate, except for the heparin from bovine lung. The heparinase exhibited only 8.3% of the activity when de-N-sulfated heparin was used as the substrate, but N-acetylation of the de-N-sulfated heparin restored the activity to 78.4%. Thus modification of N-site in heparin structure was favorable for heparinase activity. On the other hand, de-o-sulfation in heparin showed positive effects on the heparinase activity, since the enzyme activity for N-acetyl-de-o-sulfated heparin was increased by 150%. Based on the present findings, the sphingobacterial heparinase differed from flavobacterial and other reported heparinases in molecular mass, composition, charge properties, active site, substrate specificities and other important characteristics, suggesting that it a novel heparin lysase distinct from those from other sources.

Amino Acids↗

Characterization of a strain of Sphingobacterium sp. and its degradation to herbicide mefenacet.

A bacterium (designated strain Y1) degrading acetanilide herbicide mefenacet was isolated from aerobic sludge. Based on the analyses of partial 16S rRNA gene, cellular fatty acid and BIOLOG-GN, and general physiological and biochemical characteristics, strain Y1 was identified as Sphingobacterium multivolum. Strain Y1 was able to degrade mefenacet used as sources of carbon and energy. Degradation of mefenacet was accompanied by producing the metabolites N-methylaniline and an unidentified compound with molecular weight 205, indicating a metabolic pathway of mefenacet initiated by hydrolysis of amido bond.

Acetamides↗

[Production of a novel heparinase from Sphingobacterium sp].

The novel heparinase-producing bacterial strain Sphingobacterium sp. was isolated and screened from soil. The optimum medium composition is (g/L): Soytone 20, NaCl 1, K2HPO4 2.5, MgSO4 0.5, Heparin 2, Sucrose 15, pH 7.5. The optimum temperature for growth and enzyme production was 32 degrees C. When cultured at a rotating shaker at 30 degrees C for 36 hours, 200 r/min, 50 mL medium in 500 mL flask, the production of heparinase reached 4000 U/L.

Bacterial Proteins↗

Induction of KDNase Sm, a deaminoneuraminic acid (KDN) residue-specific sialidase from Sphingobacterium multivorum, using synthetic KDN-glycosides.

Various aryl and alkyl alpha-glycosides of KDN were synthesized and tested as substrates for their susceptibility to a deaminoneuraminic acid (KDN)-specific sialidase from Sphingobacterium multivorum, designated KDNase Sm. The synthetic KDN-glycosides were all hydrolyzed by the action of KDNase Sm. A hydroxyl group at C-5 position of KDN was required for the recognition by the enzyme, and was shown not to be replaced by an amino- or an acylamino group for the enzymatic recognition. These synthetic KDN-glycosides were also examined for their inducing activity of KDNase in S. multivorum and were shown to induce the KDNase activity effectively when the bacterium was cultured minimum salt medium containing both 0.1% glucose and 0.1% various KDN-glycosides. No KDNase activity was induced by the KDN-glycosides without 0.1% glucose. This is the first case of using synthetic KDN-glycosides as inducers of KDNase Sm.

Carbohydrate Conformation↗

Oleic acid transformations by selected strains of Sphingobacterium thalpophilum and Bacillus cereus from composted manure.

In a survey of 186 randomly selected microbial strains isolated from composted manure, 63 transformed oleic acid into three types of products: hydroxy fatty acid, fatty amide, and less polar oleyl lipid. Selection of oleic acid-transforming microorganisms was enhanced in nutrient agar supplemented with 0.1% (vol/vol) oleic acid at pH 7.2. Most of the 63 diverse isolates elicited inconsistent and poorly reproduced transformations. However, strains 142b (NRRL B-14797) transformed oleic acid to 10-hydroxystearic acid consistently, and strain 229b (NRRL B-14812) produced an octadecenamide. Taxonomic studies indicated that NRRL strain B-14797, possessing 1,3-dihydroxy-2-amino-15-methylhexadecane and sphinganine bases, was closely related to Sphingobacterium thalpophilum, and NRRL B-14812 was identified as Bacillus cereus.

Bacillus cereus↗