Discrimination between oral streptococci by pyrolysis gas-liquid chromatography

Washed organisms, including strains of Streptococcus mitior, S. mutans, and S. sanguis, were examined by curie-point pyrolysis gas-liquid chromatography. A linear discriminant function based upon three items from the output data was adequate for segregating the strains according to species. Strains with intermediate properties were also encountered. Sources of variability in cultures were evaluated, chromatographic performance was maintained throughout the investigation, and matching performance from a duplicate pair of chromatographic columns was demonstrated.     

Iron-binding compounds and related outer membrane proteins in Vibrio cholerae non-O1 strains from aquatic environments

A total of 156 strains of Vibrio cholerae non-O1 from aquatic origins were examined for the presence of iron uptake mechanisms and compared with O1 strains and other Vibrio species. All non-O1 strains were able to grow in iron-limiting conditions, with MICs of ethylenediaminedi (O-hydroxyphenylacetic acid) ranging from 20 microM to 2 mM. The production of siderophores was demonstrated by growth in chrome azurol S agar and cross-feeding assays. All strains produced phenolate-type compounds, as assessed by the chemical tests and by bioassays with Salmonella typhimurium enb-7. Some of the strains also promoted the growth of S. typhimurium enb-1 (which can use only enterobactin as a siderophore) as well as some strains of Vibrio anguillarum deficient in the anguibactin-mediated system. The chromatographic analyses and absorption spectra of siderophores extracted from culture supernatants suggest that vibriobactin may be produced by the strains examined. Interestingly, some strains also produced hydroxamate-type compounds, as determined by chemical tests, and were able to promote the growth of an aerobactin-deficient strain of Escherichia coli. These results were confirmed by the absorption spectra and chromatographic analyses of the culture extracts. The synthesis of iron-regulated outer membrane proteins in representative strains was also examined. The molecular sizes of the main induced proteins ranged from 70 to 78 kilodaltons. These results indicate that several iron uptake mechanisms which could be involved in environmental survival and pathogenicity are present in environmental V. cholerae non-O1 strains.     

Identification of Trichoderma strains by image analysis of HPLC chromatograms

Forty-four Trichoderma strains from water-damaged building materials or indoor dust were classified with chromatographic image analysis on full chromatographic matrices obtained by high performance liquid chromatography with UV detection of culture extracts. The classes were compared with morphological identification and rDNA sequence data, and for each class all strains were of the same identity. With all three techniques each strain--except one--was identified as the same species. These strains belonged to Trichoderma atroviride (nine strains), Trichoderma viride (three strains), Trichoderma harzianum (10 strains), Trichoderma citrinoviride (12 strains), and Trichoderma longibrachiatum (nine strains). The odd strain was identified as Trichoderma hamatum by morphology and rDNA sequencing, but not by image analysis as no reference strains of this species were included. It is concluded that the secondary metabolite profile contains sufficient information for classification and species identification.     

Iron-binding compounds and related outer membrane proteins in Vibrio cholerae non-O1 strains from aquatic environments.

A total of 156 strains of Vibrio cholerae non-O1 from aquatic origins were examined for the presence of iron uptake mechanisms and compared with O1 strains and other Vibrio species. All non-O1 strains were able to grow in iron-limiting conditions, with MICs of ethylenediaminedi (O-hydroxyphenylacetic acid) ranging from 20 microM to 2 mM. The production of siderophores was demonstrated by growth in chrome azurol S agar and cross-feeding assays. All strains produced phenolate-type compounds, as assessed by the chemical tests and by bioassays with Salmonella typhimurium enb-7. Some of the strains also promoted the growth of S. typhimurium enb-1 (which can use only enterobactin as a siderophore) as well as some strains of Vibrio anguillarum deficient in the anguibactin-mediated system. The chromatographic analyses and absorption spectra of siderophores extracted from culture supernatants suggest that vibriobactin may be produced by the strains examined. Interestingly, some strains also produced hydroxamate-type compounds, as determined by chemical tests, and were able to promote the growth of an aerobactin-deficient strain of Escherichia coli. These results were confirmed by the absorption spectra and chromatographic analyses of the culture extracts. The synthesis of iron-regulated outer membrane proteins in representative strains was also examined. The molecular sizes of the main induced proteins ranged from 70 to 78 kilodaltons. These results indicate that several iron uptake mechanisms which could be involved in environmental survival and pathogenicity are present in environmental V. cholerae non-O1 strains.     

Analysis of whole-cell fatty acid profiles of verotoxigenic Escherichia coli and Salmonella enteritidis with the microbial identification system.

Differentiation of strains within bacterial species, based on gas chromatographic analysis of whole-cell fatty acid profiles, was assessed with 115 strains of verotoxigenic Escherichia coli and 315 strains of Salmonella enteritidis. Fatty acid-based subgroups within each of the two species were generated. Variability of fatty acid profiles observed in repeat preparations from the same strain approached that observed between subgroups, limiting the usefulness of using fatty acid profiles to subgroup verotoxigenic E. coli and S. enteritidis strains.     

Comparison of antibiotic complexes formed by cultures of Streptomyces griseoruber

Physico-chemical properties of antibiotic complexes formed by various strains of Streptomyces griseoruber: VNIIA 1195, VNIIA 1193-INA 2022/55 and IMiV 1618 were compared. It was shown that the antibiotic complex 1195 differed by the electron absorption spectrum, chromatographic mobility and indicator properties from lateriomycins A and B produced by the type culture of S. griseoruber, Jamaguchi et Saburi, 1955 and strain 70717, as well as from prodigiozin produced by S. griseoruber, VNIIA 1193. By the absorption spectra and chromatographic mobility the components of the antibiotic complex 1195 were identical with cinerubins A and B, pyrromycin and eta- and xi-pyrromycinones included in the composition of antibiotic 1618 produced by S. griseoruber 1618 and differed in their quantitative contents and absence of epsilon-pyrromycinones.     

Streptomyces species with madurose (3-O-methyl-D-galactose) as a whole-cell sugar

Madurose, an actinomycete whole-cell sugar, was found in the strains of the genus Streptomyces: three strains of S. platensis, one strain each of S. platensis subsp. malvinus, and S. albus subsp. albus. The sugar was isolated from the hydrolysate of S. platensis IFO 14008 cells, and was identified as madurose (3-O-methyl-D-galactose) by chromatographic analyses, ¹H-NMR spectrometry, mass spectrometry as its alditol acetate, and demethylation with boron trichloride. Cell walls of the strain contained peptidoglycan and teichoic acids. ll-Diaminopimelic acid, glycine, glutamic acid, and alanine were present in the peptidoglycan fraction in molar ratios of 1.0:1.3:1.2:2.3. Madurose was detected in the teichoic acid fraction, which was composed of phosphorus, glycerol, galactose, and madurose in molar ratios of 9.3:8.5:2.9:1.0. Thus, madurose was found in the glycerol teichoic acid moiety of the cell walls of this strain.Abbreviations PC paper chromatography - TLC thin-layer chromatography - HPLC high performance liquid chromatography - GLC gas-liquid chromatography - TCA trichloroacetic acid     

Early identification of streptothricin group antibiotics]

Methodical approaches to detecting cultures producing streptothricins at the early stages of screening new antibiotics were developed. The approaches are based on chromatographic and electrophoretic mobility of streptothricins and the products of their hydrolysis in the extracts from agar cultures of actinomycetes. Application of the method for screening new antibiotics is illustrated with an example. Nine strains of actinomyces with broad antibacterial spectra isolated from soil samples were studied and 6 of them belonging to 4 species were shown to produce streptothricins in agar cultures. The new streptothricin-producing culture S. roseolilacinus was isolated.     

东乡野生稻内生放线菌分离及菌株S123次级代谢产物分析

【目的】从东乡野生稻(Oryza rufipogon)中分离和鉴定内生放线菌,对其进行抗菌活性筛选,并分析高抗菌活性菌株S123的次级代谢产物。【方法】采用S培养基对东乡野生稻内生放线菌进行分离、纯化,并构建16S rRNA基因序列系统发育进化树进行菌株鉴定。以琼脂扩散法和菌丝生长速率法进行抗菌活性筛选,同时设计简并引物检测菌株I型聚酮合酶(PKS-I)基因。对具广谱抗菌活性的菌株S123进行分批大量发酵,运用多种色谱方法对发酵产物进行分离、纯化,利用MS和NMR分析鉴定化合物的结构。【结果】从东乡野生稻中共分离到11株内生放线菌,分别属于链霉菌属(8株)和假诺卡氏属(3株)。其中有8株具有抗菌活性,8株呈现I型PKS阳性。从高抑菌活性菌株S123中分离到化合物Nigericin和17-O-demethylgeldanamycin,其中Nigericin对金黄色葡萄球菌、枯草芽孢杆菌及水稻纹枯病菌均有抑制活性。【结论】对东乡野生稻内生放线菌进行了分离、鉴定和抗菌活性筛选,并从中分到两种与I型PKS基因相关活性的化合物Nigericin和17-O-demethylgeldanamycin,为研究东乡野生稻内生放线菌的多样性和次级代谢产物的分离提供依据。     

A possible role of poly-3-hydroxybuiyric acid in antibiotic production in<Emphasis Type="Italic">Streptomyces</Emphasis>

The occurrence of poly-3-hydroxybutyric acid (PHB) in 12 different strains of the genus Streptomyces was investigated. Gas chromatographic estimation indicated that all the strains produced PHB and the range of maximum PHB accumulation was between 1.5 and 11.8% dry cell weight. PHB was isolated from Streptomyces coelicolor A3(2) M145 and characterized using Fourier transform-infrared (FT-IR) spectroscopy. The correlation between PHB utilization and antibiotic production in S. coelicolor A3(2) M145, was studied; results indicated a possible role of PHB as a carbon reserve material used for antibiotic production.     

A Thin-Layer Chromatographic Technique for Rapid Estimation of Bacterial Lipases

S ummary . A thin-layer chromatographic technique for the estimation of lipase in Pseudomonas aeruginosa and staphylococci is described. The technique needs a short incubation period (4 h), while the subsequent procedure is quite simple and rapid. The validity of the technique was established by testing 56 strains of Ps. aeruginosa and 35 strains of staphylococci. The results strongly support the usefulness of the technique which can be quite easily applied to routine use.     

Characterization of facultative oligotrophic bacteria from polar seas by analysis of their fatty acids and 16S rDNA sequences

One hundred and seventy three bacterial strains, isolated previously after enrichment under oligotrophic, psychrophylic conditions from Arctic (98 strains) and Antarctic seawater (75 strains), were characterized by gas-liquid chromatographic analysis of their fatty acid compositions. By numerical analysis, 8 clusters, containing 2 to 59 strains, could be delineated, and 8 strains formed separate branches. Five clusters contained strains from both poles, two minor clusters were confined to Arctic isolates, and one cluster consisted of Antarctic isolates only. The 16S rRNA genes from 23 strains, representing the different fatty acid profile clusters and including the unclustered strains, were sequenced. The sequences grouped with the alpha and gamma Proteobacteria, the high percent G+C gram positives, and the Cytophaga-Flavobacterium-Bacteroides branch. The sequences of strains from 4 clusters and of 7 unclustered strains were closely related (sequence similarities above 97%) to reference sequences of Sulfitobacter mediterraneus, Halomonas variabilis, Alteromonas macleodii, Pseudoalteromonas species, Shewanella frigidimarina, and Rhodococcus fascians. Strains from the other four clusters and an unclustered strain showed sequence similarities below 97% with nearest named neighbours, including Rhizobium, Glaciecola, Pseudomonas, Alteromonas macleodii and Cytophaga marinoflava, indicating that the clusters which they represent form as yet unnamed taxa.     

中国酱香型白酒酿造酿酒酵母的独特生理代谢特征

【目的】酿酒酵母是白酒发酵的主要微生物,对白酒的产量及质量都有重要影响。而酱香酒酿造环境具有高温、酸性、高乙醇等胁迫因素,研究其中酿酒酵母的生理代谢特征并有目的地应用于白酒实际生产中的意义。【方法】从酱香酒酿造环境中筛选一株性能优良的酿酒酵母菌株,比较其与酿酒酵母模式菌株S288c和商业酵母的生理代谢特征。【结果】从酱香型酒醅中筛选得到性能优良的Saccharomyces cerevisiae MT1,该菌株可耐受高温42 °C,高浓度乙醇(16%,体积比),低pH (2.0),其最大比生长速率和最大比产乙醇速率分别达到了S288c的125%和114%,其乙醇转化率也要高于其他菌株。一些挥发性物质只有在MT1的发酵液中可检测到,包括苯并噻唑、2,3-二氢苯并呋喃、4-乙烯基愈创木酚以及丁羟甲苯等,MT1的苯乙醇、法呢醇、橙花叔醇、乙偶姻和大马酮量也要高于S288c。另外,MT1可以利用多种碳源发酵产乙醇,如半乳糖、麦芽糖、蜜二糖、松二糖、海藻糖和棉子糖等。【结论】来源于酱香酒酿造的S. cerevisiae MT1具有高耐受力、高效的发酵性能及更广泛的碳源利用图谱,并能生成多种挥发性物质。     

Isolation and characterization of Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus from plants in Bulgaria

One of the traditional ways of preparation of yogurt starter in Bulgaria is placing a branch of a particular plant species into boiled sheep's milk maintained at about 45°C, which is further incubated until a dense coagulum is obtained. To investigate the possible origin of the yogurt starter bacteria, Lactobacillus delbrueckii ssp. bulgaricus ( L. bulgaricus ) and Streptococcus thermophilus ( S. thermophilus ), the traditional way of yogurt-starter preparation was followed. Hundreds of plant samples were collected from four regions in Bulgaria and incubated in sterile skim milk. The two target bacteria at low frequencies from the plant samples collected were successfully isolated. Phenotypic and genotypic characteristics of these bacterial isolates revealed that they were identified as L. bulgaricus and S. thermophilus . Twenty isolates of L. bulgaricus and S. thermophilus , respectively, were selected from the isolated strains and further characterized with regard to their performance in yogurt production. Organoleptic and physical properties of yogurt prepared using the isolated strains from plants were not significantly different from those prepared using commercial yogurt-starter strains.
It was therefore suggested that L. bulgaricus and S. thermophilus strains widely used for commercial yogurt production could have originated from plants in Bulgaria. To our knowledge, this is the first report on the isolation and characterization of L. bulgaricus and S. thermophilus strains from plants.     

宿主遗传背景对重组酿酒酵母木糖共发酵影响的研究进展

木糖是纤维素原料水解液中最主要的五碳糖成分,由于野生的酿酒酵母缺乏有效的木糖利用途径,将外源木糖代谢途径整合至酿酒酵母中使其具有发酵木糖生产乙醇的能力是构建纤维素乙醇发酵菌株的关键。国内外学者的研究表明,同一木糖代谢途径导入不同酿酒酵母菌株中,所得到的重组菌发酵性能存在明显差异,表明宿主的遗传背景对菌株利用木糖能力和发酵性能具有重要的影响。就酿酒酵母宿主对重组菌株的木糖发酵性能的影响进行了综述,分析了产生宿主差异的内在机理,为进一步选育高效木糖共发酵菌种提供借鉴。     

Isolation and characterization of mucous exopolysaccharide (EPS) produced by Vibrio furnissii strain VB0S3

Marine bacterial strains were isolated from coastal regions of Goa and screened for the strains that produce the highest amount of mucous exopolysaccharide (EPS). Our screening resulted in the identification of the strain Vibrio furnissii VB0S3 (hereafter called VB0S3), as it produced the highest EPS in batch cultures during the late logarithmic growth phase. The isolate was identified as VB0S3 based on morphological and biochemical properties. Growth and EPS production were studied in mineral salts medium supplemented with NaCl (1.5%) and glucose (0.2%). The exopolymer was recovered from the culture supernatant by using three volumes of cold ethanol precipitation and dialysis procedure. Chemical analyses of EPS revealed that it is primarily composed of neutral sugars, uronic acids, and proteins. Fourier-transform infrared (FT-IR) spectroscopy revealed the presence of carboxyl, hydroxyl, and amide groups, which correspond to a typical heteropolymeric polysaccharide, and the EPS also possessed good emulsification activity. The gas chromatographic analysis of an alditol-acetate derivatized sample of EPS revealed that it was mainly composed of galactose and glucose. Minor components found were mannose, rhamnose, fucose, ribose, arabinose, and xylose. EPS was readily isolated from culture supernatants, which suggests that the EPS was a slime-like exopolysaccharide. This is the first report of exopolysaccharide characterization that describes the isolation and characterization of an EPS expressed by Vibrio furnissii strain VB0S3. The results of the study contribute significantly and go a long way towards an understanding of the correlation between growth and EPS production, chemical composition, and industrial applications of the exopolysaccharide in environmental biotechnology and bioremediation.     

链格孢霉醇和链格孢霉醇单甲醚产生菌株的筛选

本文对分离自小麦、马铃薯、番茄和茄子上链格孢霉属(Alternaria)2个种(链格孢和茄链格孢)的96个菌株,用枯草杆菌生长抑制试验筛选链格孢霉醇(AOH)和链格孢霉醇单甲醚(AME)的产生菌株,有48株产生毒性作用(占所测菌株的50％)。18株产强、中毒性菌用高效液相色谱分析,有13株产AOH和AME(占所测菌株的72.2％)。链格孢的产毒素菌株率比茄链格孢低。但产毒素含量却是前者明显高于后者。其中产AOH和AME的最高含量,链格孢菌株XA-8分别为280和5140mg/kg,而茄链格孢菌株SA-10分别为95.9和94.3mg/kg。     

High resolution "ultra performance" liquid chromatography coupled to oa-TOF mass spectrometry as a tool for differential metabolic pathway profiling in functional genomic studies

The combination of a new 1.7 mum reversed-phase packing material, and a chromatographic system, operating at ca. 12,000 psi, (so-called ultra performance liquid chromatography, UPLC) has enabled dramatic increases in chromatographic performance to be obtained for complex mixture separation. This increase in performance is manifested in improved peak resolution, together with increased speed and sensitivity. Here, we show that UPLC offers significant advantages over conventional reversed-phase HPLC amounting to a more than doubling of peak capacity, an almost 10-fold increase in speed and a 3- to 5-fold increase in sensitivity compared to that generated with a conventional 3.5 microm stationary phase. The first functional genomic application of UPLC-MS technology is illustrated here with respect to multivariate metabolic profiling of urines from males and females of two groups of phenotypically normal mouse strains (C57BL19J and Alpk:ApfCD) and a "nude mouse" strain. We have also compared this technology to conventional HPLC-MS under similar analytical conditions and show improved phenotypic classification capability of UPLC-MS analysis together with increased ability to probe differential pathway activities between strains as a result of improved analytical sensitivity and resolution.     

Strain specificity and cholinergic modulation of visuospatial attention in three inbred mouse strains

The tremendous increase in the use of mouse inbred strains and mutant mice to study the molecular basis of psychiatric disorders urges for a better understanding of attentional performance in mice. To this aim, we investigated possible strain differences in performance and cholinergic modulation of visuospatial attention in three widely used mouse inbred strains (129S2/SvHsd, C57BL/6JOlaHsd and DBA/2OlaHsd) in the five-choice serial reaction time task. Results indicated that after extended training, performance of 129S2/SvHsd mice was superior to that of C57BL/6JOlaHsd and DBA/2OlaHsd mice in terms of attention, omission errors, inhibitory control and latencies to correct choice. Increasing the attentional load resulted in comparable decrements in attention in all strains and inhibitory control impairments that were most pronounced in DBA/2OlaHsd mice. Further pharmacological evaluation indicated that all strains showed attentional impairments after treatment with the muscarinic and nicotinic antagonists scopolamine and mecamylamine, respectively. 129S2/SvHsd mice were less sensitive, whereas DBA/2OlaHsd mice appeared more sensitive to the detrimental effects of mecamylamine. In addition, subchronic, but not acute, nicotine treatment slightly improved attentional performance in all strains to the same extent. In conclusion, our data indicate strain specificity with particularly good performance of 129S2/SvHsd mice and strong cholinergic involvement in visuospatial attention in mice.     

Studies on the mutagenicity of p-phenylenediamine in Salmonella typhimurium. Presence of PCB's in rat-liver microsomal fraction induced by Aroclor.

The mutagenicity of fresh solutions of p-phenylenediamine (PPD) and Aroclor 1254 was investigated. The histidine-requiring strains of Salmonella typhimurium were used in the absence and presence of uninduced and/or Aroclor-induced rat-liver homogenate. The presence of polychlorinated biphenyls (PCBs) was also examined by chromatographic methods in Aroclor-induced rat-liver homogenate. In the absence of metabolic activation, as well as in the presence of uninduced rat-liver homogenate, PPD was not mutagenic in the strains used. In the presence of Aroclor-induced S9 a twofold increase (or less) was observed in the number of revertant colonies over those of the controls in TA1538 and TA98. There was no increase in the number of revertant colonies over those of the controls when PPD was dissolved in NH4OH solution and the solution mixed with H2O2 before the addition of S9 mix. Aroclor 1254 was not mutagenic in TA1538 or TA98. However, the presence of PCBs in Aroclor-induced rat-liver homogenate (induced S9) was identified by gas-liquid chromatography (GLC), high-performance liquid chromatography (HPLC) and gas--liquid chromatography/mass spectrometry (GC/MS).