Relatedness of Strains of Xanthomonas fragariae by Restriction Fragment Length Polymorphism, DNA-DNA Reassociation, and Fatty Acid Analyses

The levels of relatedness of strains of Xanthomonas fragariae collected over several years from locations in Canada and the United States were compared by determining fatty acid methyl ester profiles, restriction fragment length polymorphisms (RFLP) based on pulsed-field gel electrophoresis (PFGE) analysis, and DNA-DNA reassociation values. Based on qualitative and quantitative differences in fatty acid profiles, the strains were divided into nine groups and four groups by the MIDI “10% rule” and unweighted pair analysis, respectively. Restriction analysis of genomic DNA by PFGE with two endonucleases (XbaI and SpeI) revealed four distinct profiles. When a third endonuclease (VspI) was used, one group was divided into three subgroups. The profile of the American Type Culture Collection type strain differed from the profile of every other strain of X. fragariae. Considerable diversity was observed within X. fragariae, although the majority of the strains represented a clonal population. The four groups based on fatty acid profiles were similar to the four groups based on RFLP, but neither method related groups to the geographic origins of the strains. The DNA-DNA reassociation values were high for representative strains, providing evidence that all of the strains belong to the same species.     

Intraspecies variability of cellular fatty acids among soil and intestinal strains of Desulfovibrio desulfuricans.

A comparison of cellular fatty acid profiles of Desulfovibrio desulfuricans DSM 642 and 14 wild strains of this species, isolated from two completely different environments, soil and the human intestine, was carried out. All the D. desulfuricans strains grown on lactate and sulfate indicated the presence of considerable amounts of i-C15:0, i-C17:1 and C16:0. Although differences in the quantities of individual fatty acids present in each strain were clear in the group of soil strains (similarity, 67.6%), in contrast to almost identical fatty acid patterns (similarity, near 100%) in the intestinal strains, the results were variable within the limits acceptable for species demonstration. The higher similarity of the fatty acid profiles of intestinal strains may be a result of the similarity of biocenoses in the human digestive tract. The coefficients of variability of i-C17:1 and i-C15:0 (the major branched-chain fatty acids), as well as clustering of the investigated strains compared with strains described in the literature after plotting percentages of i-C17:1 fatty acid against i-C15:0 fatty acid, confirmed a certain heterogeneity of cellular fatty acid profiles within the group of soil strains, in contrast to almost ideal homogeneity within the group of intestinal isolates. Intestinal strains contained a higher ratio of saturated to unsaturated fatty acids (2.2 +/- 0.14) than did soil strains (1.6 +/- 0.2; in one case, 2.7). We propose that intestinal D. desulfovibrio bacteria should be assumed to be a highly homogeneous group and should be represented by the strain D. desulfuricans subsp. intestinus in collections of microbial cultures.     

Whole cell fatty acid patterns of Xenorhabdus species

Thirty-three strains of the nematode-associated bacterium Xenorhabdus were characterized by traditional biochemical tests and whole cell fatty acid analysis. In traditional tests 26 strains were found to belong to X. luminescens and 7 to X. nematophilus (sensu latu). No further subdivision could be made. In fatty acid analysis, however, X. luminescens strains could be divided into three subgroups. The amount of distinction in fatty acids is similar to that at subspecies or species level found in other bacteria. Xenorhabdus nematophilus could be clearly differentiated from X. luminescens , key acids are 12: 0, 15: 0 iso, 16: 0, 17: 0 iso, 17: 0 cyclo, 18: 1 cis 11 and 19: 0 cyclo. Separation is almost at genus level. The presence of branched and hydroxy acids in Xenorhabdus and its aberrant morphology make the placement of this genus in the Enterobacteriaceae questionable. This is the first report on fatty acid profiles of Xenorhabdus species.     

Subgrouping of bacterial populations by cellular fatty acid composition

Abstract The cellular fatty acid composition of six bacterial species isolated from the seeds and leaves of sugar beet ( Beta vulgaris ) and from soil were analysed. The quantitative data from the fatty acid methyl ester (FAME) profiles were highly reproducible. Numerical analysis of Xanthomonas maltophilia . FAME profiles sub-grouped strains according to when they were isolated in the growing season. The analytical method used was sensitive enough to differentiate strains of Klebsiella terrigena isolated from either soil or leaves. The results from this study confirm reports that analyses of bacterial FAME composition were rapid to perform, specific and allowed differentiation of strains within the same species.     

Ethanol-mediated variations in cellular fatty acid composition and protein profiles of two genotypically different strains of Escherichia coli O157:H7

Two strains of Escherichia coli O157:H7 were grown in tryptic soy broth (TSB, pH 7.1) supplemented with 0, 2.5, 5.0, 7.5, and 10% ethanol at 30 degrees C for up to 54 h. Growth rates in TSB supplemented with 0, 2.5, and 5.0% ethanol decreased with an increase in ethanol concentration. Growth was not observed in TSB supplemented with 7.5 or 10% ethanol. The pH of TSB containing 5.0% ethanol decreased to 5.8 within 12 h and then increased to 7.0 at 54 h. The ethanol content in TSB supplemented with 2.5 or 5.0% ethanol did not change substantially during the first 36 h of incubation but decreased slightly thereafter, indicating utilization or degradation of ethanol by both strains. Glucose was depleted in TSB supplemented with 0, 2.5, or 5.0% ethanol within 12 h. Cells grown under ethanol stress contained a higher amount of fatty acids. With the exceptions of cis-oleic acid and nonadecanoic acid, larger amounts of fatty acid were present in stationary-phase cells of the two strains grown in TSB supplemented with 5.0% ethanol for 30 h than in cells grown in TSB without ethanol for 22 h. The trans-oleic acid content was 10-fold higher in the cells grown in TSB with 5.0% ethanol than those grown in TSB without ethanol. In contrast, cis-oleic acid was not detected in ethanol-stressed cells but was present at concentrations of 0.32 and 0.36 mg/g of cells of the two strains grown in TSB without ethanol. Protein content was higher in ethanol-stressed cells than in nonstressed cells. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis protein profiles varied qualitatively as affected by the strain and the presence of ethanol in TSB. An ethanol-mediated protein (28 kDa) was observed in the ethanol-stressed cells but not in control cells. It is concluded that the two test strains of E. coli O157:H7 underwent phenotypic modifications in cellular fatty acid composition and protein profiles in response to ethanol stress. The potential for cross protection against subsequent stresses applied in food preservation technologies as a result of these changes is under investigation.     

Cellular fatty acid composition in film-forming strains of two physiological races of Saccharomyces cerevisiae

Eleven strains belonging to two physiological races of Saccharomyces cerevisiae endowed with different abilities of forming films at air-liquid interfaces were analysed in relation to cell fatty acid composition and cell hydrophobicity. Extensive individual differences in fatty acid profiles were observed both in the film and in the non-film phase. The ability of the cells to form a floating film seems to be an implicit strain character associated with an elevated unsaturation level and a mean chain length of fatty acid residues, as well as cellular hydrophobicities higher than those shown by non-film-forming strains belonging to the same species.     

Polar lipid and fatty acid profiles – Re-vitalizing old approaches as a modern tool for the classification of mycoplasmas?

A set of 20 Mollicutes strains representing different lines of descent, including the type species of the genus Mycoplasma, Mycoplasma mycoides, Acholeplasma laidlawii and a strain of Mesoplasma, were subjected to polar lipid and fatty acid analyses in order to evaluate their suitability for classification purposes within members of this group. Complex polar lipid and fatty acid profiles were detected for each examined strain. All strains contained the polar lipids phosphocholine-6'-alpha-glucopyranosyl-(1'-3)-1, 2-diacyl-glycerol (MfGL-I), 1-O-alkyl/alkenyl-2-O-acyl-glycero-3-phosphocholine (MfEL), sphingomyelin (SphM), 1-O-alkyl/alkenyl-glycero-3-phosphocholine (lysoMfEL), the unknown aminophospholipid APL1 and the cholesterol Chol2. A total of 19 strains revealed the presence of phosphatidylethanolamine (PE) and/or phosphatidylglycerol (PG), and the presence of diphosphatidylglycerol (DPG) was detected in 13 strains. The unknown aminolipid AL1 was found in the extracts of 17 strains. Unbranched saturated and unsaturated compounds predominated in the fatty acid profiles. Major fatty acids were usually C16:0, C18:0, C18:1 omega9c and 'Summed feature 5' (C18:2 omega6, 9c/C18:0 anteiso). Our results demonstrated that members of the M. mycoides cluster showed rather homogenous polar lipid and fatty acid profiles. In contrast, each of the other strains was characterized by a unique polar lipid profile and significant quantitative differences in the presence of certain fatty acids. These results indicate that analyses of both polar lipid and fatty acid profiles could be a useful tool for classification of mycoplasmas.     

Molecular phylogenetic typing of pandemic isolates of Salmonella enteritidis

Salmonella enteritidis is now the most common Salmonella serovar in many countries. We have used cloned DNA probes to analyze genome interrelationships between strains chosen to represent the current S. enteritidis pandemic, and included designated type strains of the seven subspecies of Salmonella in order to compare the levels of discrimination of probes. DNA sequence divergence and rearrangements were analyzed in and around the rfa, fim and umuDC loci, and around insertion sites of the Salmonella-specific DNA insertion element, IS200. The S. enteritidis isolates showed a high degree of genome homogeneity. Chromosomal genetic loci exhibited characteristic DNA sequence divergence between subspecies of Salmonella, but no intraserovar divergence or difference with the subspecies I type strain was observed for S. enteritidis. The locus umuDC was not found in S. enteritidis. S. enteritidis contains a conserved and a variable site of insertion of insertion sequence IS200 and the analysis of DNA rearrangements around the second of these sites showed that three distinct evolutionary lines or races exist within pandemic isolates associated with human gasteroenteritis. IS200 profiles of a range of U.K. isolates of the epidemic phage type PT4 showed that all belonged to a single clonal line.     

relA gene control of the synthesis of lipid A fatty acyl moieties.

The incorporation of [14C]acetate into the fatty acid moieties of lipid A was measured during amino acid starvation of rel+ and relA strains of Escherichia coli K-12. The synthesis of the beta-hydroxymyristate and other fatty acid moieties was inhibited two- to fourfold in rel+ strains, whereas no inhibition was observed in relA strains. The fatty acid compositions of the phospholipids synthesized after amino acid starvation or rel+ and relA strains were also determined.     

Distinction of Prevotella intermedia and Prevotella nigrescens from endodontic bacteremia through their fatty acid contents

Prevotella nigrescens has recently been recognized as a new species distinct from Prevotella intermedia. The distinction is based largely on DNA-DNA hybridization, electrophoretic migration of malate and glutamate dehydrogenase, and peptidase and lipase activities of type strains. Gas chromatography of cellular fatty acids can be a useful adjunct for characterization and identification of bacterial species. In the present study, cellular fatty acid profiles were determined for seven strains of P. intermedia and six strains of P. nigrescens. Six of these 13 strains were isolated from the root canal and blood of three patients during endodontic therapy of teeth with Asymptomatic apical periodontitis. The bacteria were cultivated anaerobically in 10 mL prereduced anaerobically sterilized peptone-yeast extract-glucose broth for 24 h. Dried cells of each isolate were methanolysed and their fatty acid contents determined by the Microbial Identification System software package by MIDI. The data were treated by principal component analysis, which distinguished P. nigrescensfromP. intermedia. Cellular fatty acid profiles of these strains of the species in blood matched the profiles of their respective root canal isolates, as demonstrated by Euclidean Distance Square assessment. This suggested that the organisms in the root canal had spread to the bloodstream during endodontic treatment.     

Evaluation of a commercial beta-glucuronidase test for the rapid and economical identification of Escherichia coli

A commercial beta-glucuronidase (beta-GUR) test for the rapid and economical identification of Escherichia coli was evaluated. A total of 762 clinical strains and 228 environmental isolates were studied. More than 95% of the E. coli strains were found to be beta-GUR positive. Thirty-one clinical isolates of Shigella sonnei, 10 of Enterobacter cloacae, eight of Enterobacter aerogenes, nine of Citrobacter freundii and one of Salmonella enteritidis also gave positive results. The enzyme beta-GUR was also detected in two environmental strains of E. cloacae and one C. freundii. A comparative study between the beta-GUR test and the conventional identification system was carried out in 233 consecutive isolates of lactose positive enterobacteria. Agreement was observed in 223 cases and 190 E. coli strains were correctly identified using this test. Discrepancies were found in 10 cases: nine E. coli were beta-GUR negative and one C. freundii was beta-GUR positive. Escherichia coli was the only species positive for both beta-GUR and indole tests. This procedure permits a rapid, easy, precise and inexpensive identification of E. coli. beta-GUR positive Enterobacter strains have not previously been described.     

Inhibition of pathogenic Salmonella enteritidis growth mediated by Escherichia coli microcin J25 producing strains

For the first time, microcin-producing strains showing inhibitory activities against enteropathogen Salmonella enteritidis were isolated from poultry intestinal contents. Among the numerous strains isolated, two strains of Escherichia coli, named J02 and J03, showing the greatest activities against S. enteritidis, were studied. Biochemical tests and purification identified the main antagonist compound produced as microcin J25. In order to evaluate the protective potential of E. coli J02 and J03 against S. enteritidis infection, the ability of these strains to inhibit growth of S. enteritidis was investigated in mixed culture. A strong antagonist activity was obtained with a preculture phase of the active strain in minimal medium before incubation with S. enteritidis. In a bioreactor experiment simulating the chicken gastric and intestinal tract environment, a mixture of the two strains E. coli J02 and J03, provided an enhanced inhibitory effect. Microcinogenic strain activities were not affected by bile, pancreatic enzymes addition, or acidic conditions. These results suggest the relevant role of microcin-producing microorganisms in microbial intestinal ecology. To conclude, this study shows that microcin J25 strains could exert a beneficial protective effect against S. enteritidis growth in situ.     

Analysis of cellular fatty acids by gas chromatography as a tool in the identification of medically important coryneform bacteria

The fatty acid methyl esters of nineteen unidentified pathogenic coryneform bacteria were analysed by gas-liquid chromatography and the resulting profiles were compared with those of type or reference strains of possibly related species, namely Caseobacter polymorphus, Corynebacterium bovis, C. diphtheriae, C. xerosis and Rhodococcus equi. All of the strains had distinct fatty acid profiles but most of them conformed to a general pattern, with high levels of octadecanoic acids and only trace amounts of 10-methyl octadecanoic acid (tuberculostearic acid). These profiles were very similar to those from C. diphtheriae and C. xerosis but could be differentiated from C. bovis, Cas. polymorphus, R. equi and two unidentified pathogenic strains which had significantly higher levels of tuberculostearic acid.     

Analysis of cellular fatty acids by gas chromatography as a tool in the identification of medically important coryneform bacteria

The fatty acid methyl esters of nineteen unidentified pathogenic coryneform bacteria were analysed by gas-liquid chromatography and the resulting profiles were compared with those of type or reference strains of possibly related species, namely Caseobacter polymorphus, Corynebacterium bovis, C. diphtheriae, C. xerosis and Rhodococcus equi. All of the strains had distinct fatty acid profiles but most of them conformed to a general pattern, with high levels of octadecanoic acids and only trace amounts of 10-methyl octadecanoic acid (tuberculostearic acid). These profiles were very similar to those from C. diphtheriae and C. xerosis but could be differentiated from C. bovis, Cas. polymorphus, R. equi and two unidentified pathogenic strains which had significantly higher levels of tuberculostearic acid.     

Identification of atypical Frankia strains by fatty acid analysis

Abstract: Ineffective, non-infective actinomycetous isolates obtained from actinorhizal nodules of Coriaria nepalensis and Datisca cannabina were identified as Frankia using whole cell fatty acid analysis. The isolates exhibited fatty-acid patterns very similar to those of confirmed Frankia strains from other host plants ( Alnus, Casuarina, Colletia, Comptonia, Elaeagnus and Hippophae ). All Frankia strains, including Coriaria and Datisca isolates, showed fatty-acid profiles very distinct from those of other actinomycetes used as controls ( Actinomyces, Geodermatophilus, Nocardia, Mycobacterium and Streptomyces ). For the genus Frankia , a characteristic pattern of five fatty acids (15:0; 15:1; 16:0 iso; 17:0 and 17:1) was found. These fatty acids comprised 75% or more of the total content. All Frankia strains could be placed into three subgroups. Coriaria isolates were found in the largest subgroup which contained most Frankia strains from other hosts while ineffective strains from Alnus, Elaeagnus and Datisca were distributed in all three subgroups of Frankia .     

Characterization of Flavobacterium species by analysis of volatile fatty acid production

Seventy-four Flavobacterium strains were characterized by gas-liquid chromatographic analysis of volatile fatty acids produced in the culture medium. Principal components analysis permitted the graphic representation of the relative positions of the different strains, and aggregation according to the variance enabled a hierarchical classification to be established. The study revealed three subgroups each for F. meningosepticum and F. odoratum. Our F. breve, Flavobacterium sp. group IIb and F. multivorum strains appeared to be homogeneous. These results tallied with those of previous studies on DNA base composition and reassociation, electrophoretic protein profiles and cellular fatty acid composition.     

The redfish species Sebastes viviparus, Sebastes marinus and Sebastes mentella have different composition of their tissue fatty acids

The fatty acid profile in the tissue of heart, gill, skull- and otolith-oil of the three redfish species, Sebastes viviparus, S. marinus and S. mentella was determined by a chemometric method, consisting of methanolysis of samples of the tissues and of the oils and gas chromatography of the resulting fatty acid methyl esters. The analytical data were treated by multivariate statistics comprising principal component analysis (PCA) and soft independent modeling of class analogy (SIMCA). Although the differences in fatty acid composition among the two tissues and two oils were the dominating features of the data, the three species had significantly different fatty acid profiles within each tissue. Variation among the individuals was considerable. The fatty acid profiles appeared to be species specific. The mutual relationship between S. marinus and S. mentella is closer than the relationship between either of them and S. viviparus. A comparison with a corresponding investigation on the same three species from another location indicates that the observed differences do not seem to be affected by biotic or abiotic factors.     

Characterization of Bacteroides gingivalis by direct fluorescent antibody staining and cellular fatty acid profiles

Bacteroides gingivalis is a newly proposed species which includes strains isolated from the mouth. Thirteen strains of B. gingivalis isolated from three geographic locations in the United States and France were examined with direct fluorescent antibody staining and analysis of total cellular fatty acids and compared with 16 strains of B. asaccharolyticus of nonoral origin by the same methods. Bacteroides gingivalis strains reacted with the B. gingivalis conjugate (fluorescein isothiocyanate labeled antibody reagent) only, while the B, asaccharolyticus strains reacted with the B. asaccharolyticus conjugate only. The B. gingivalis strains showed negative fluorescence with fluorescein isothiocyanate conjugates for other black-pigmented Bacteroides species. The specificity of the B. gingivalis conjugate was demonstrated by its failure to stain 88 strains of aerobic and anaerobic bacteria other than B. gingivalis. The fatty acid profiles of B. gingivalis and B. asaccharolyticus were readily distinguishable. The B. gingivalis profile was also distinguishable from those of other pigmenting Bacteroides species on the basis of concentration ratios among the characteristic components. These results support the species separation of B. gingivalis and B. asaccharolyticus. Further, they indicate the usefulness of cellular fatty acid profiles as an adjunct to the use of specific fluorescent antibody conjugates for identification of Bacteroides species.     

集胞藻PCC6803膜脂循环关键基因酶学性质和生理功能

集胞藻PCC6803野生型和其脂酰ACP合酶敲除突变株的自由脂肪酸含量和组成表明膜脂的重构和降解是细胞内自由脂肪酸的来源之一。在这一过程中脂肪酶起到关键性作用。通过基因组数据库检索,发现集胞藻PCC6803基因组中只有一个脂肪酶编码基因sll1969,但是还没有其功能相关的生化证据。为了确定该基因的功能及其在脂肪酸代谢途径中的作用,加深对集胞藻PCC6803脂肪酸代谢途径的了解,文中将sll1969基因在大肠杆菌中过表达和体外纯化,得到重组蛋白Sll1969,并对其酶学性质进行初步分析。在30℃条件下,测得Sll1969以对硝基苯丁酸酯作为底物时的Km和kcat值分别为(1.16±0.01)mmol/L和(332.8±10.0)/min;该脂肪酶的最适反应温度为55℃。通过比较分析sll1969突变株中脂肪酸含量和组成变化,发现sll1969的表达量与细胞自由脂肪酸的产量呈正相关,但Sll1969不是细胞中唯一的脂肪酶。