Control of Colanic Acid Synthesis

The nucleotide pools of certain mucoid, colanic acid-synthesizing strains of Escherichia coli, Salmonella typhimurium, and Aerobacter cloacae were examined, and in all cases the nucleotide sugars uridine-5'-diphosphate glucose (UDPG), uridine-5'-diphosphate galactose (UDPGal), guanosine-5'-diphosphate fucose (GDPF), and uridine-5'-diphosphate glucuronic acid (UDPGA) were detected. It is postulated that these nucleotide sugars are precursors in the synthesis of colanic acid. The levels of these nucleotide sugars and of the enzymes involved in their synthesis were examined in a number of mucoid strains and compared with the levels found in certain strains which were repressed in the synthesis of colanic acid, only becoming mucoid when grown in the presence of p-fluorophenylalanine (PFA). The levels of UDPG and UDPGal and the enzymes involved in their synthesis were substantially the same in both mucoid and repressed types, but the levels of UDPGA and GDPF and of some of the enzymes involved in their synthesis were much higher in mucoid strains. When repressed strains were grown in the presence of PFA, the levels of UDPGA and GDPF approached those found in mucoid strains. The existence of an operon, containing genes coding for certain key enzymes involved in colanic acid synthesis has been suggested.     

Role of Colanic Acid Polysaccharide in Serum Resistance In Vivo and in Adherence

The production of an extracellular layer of polysaccharide, termed the capsule, is a common feature of many bacteria. Capsules play a vital role in permitting evasion of the host immune specific and nonspecific defenses as well as helping in adhesion for colonization of host tissue. Colanic acid capsule is usually produced in low quantities and is a common feature of several enteric bacteria. The role of the colanic acid capsule in aiding adhesion and virulence was investigated. Encapsulated and unencapsulated cells were injected into granuloma pouches that were formed on the backs of rats. During the first 50 h, the viability of the matched encapsulated and unencapsulated cells decreased. These studies showed that colanic acid capsule does not confer resistance to the bactericidal activity of serum or to phagocytosis in vivo, since there was no significant difference in the survival rates of both strains over time. Adherence studies were conducted in monolayers of human carcinoma intestinal cells (T84) with the same matched strains. After incubating radioactively labeled bacteria with the colon cells, the level of adherence was determined by measuring the radioactivity remaining in the tissue culture wells. The results of these experiments indicated that the unencapsulated cells adhered more readily to the intestinal cells, suggesting that colanic acid capsule interferes with adherence in this model system. Received: 7 June 1996 / Accepted: 10 July 1996     

Role of Capsular Colanic Acid in Adhesion of Uropathogenic Escherichia coli

Urinary tract infections are the most common urologic disease in the United States and one of the most common bacterial infections of any organ system. Biofilms persist in the urinary tract and on catheter surfaces because biofilm microorganisms are resistant to host defense mechanisms and antibiotic therapy. The first step in the establishment of biofilm infections is bacterial adhesion; preventing bacterial adhesion represents a promising method of controlling biofilms. Evidence suggests that capsular polysaccharides play a role in adhesion and pathogenicity. This study focuses on the role of physiochemical and specific binding interactions during adhesion of colanic acid exopolysaccharide mutant strains. Bacterial adhesion was evaluated for isogenic uropathogenic Escherichia coli strains that differed in colanic acid expression. The atomic force microscope (AFM) was used to directly measure the reversible physiochemical and specific binding interactions between bacterial strains and various substrates as bacteria initially approach the interface. AFM results indicate that electrostatic interactions were not solely responsible for the repulsive forces between the colanic acid mutant strains and hydrophilic substrates. Moreover, hydrophobic interactions were not found to play a significant role in adhesion of the colanic acid mutant strains. Adhesion was also evaluated by parallel-plate flow cell studies in comparison to AFM force measurements to demonstrate that prolonged incubation times alter bacterial adhesion. Results from this study demonstrate that the capsular polysaccharide colanic acid does not enhance bacterial adhesion but rather blocks the establishment of specific binding as well as time-dependent interactions between uropathogenic E. coli and inert substrates.     

Relation of Lipopolysaccharide and Fatty Acid Ester Release to the Ethylenediaminetetraacetic Acid Alteration of Permeability in Enterobacteriaceae

Escherichia coli subjected to cold osmotic shock released 30 to 40% of their fatty acid esters and 42% of their cellular hexosamine. In contrast, Enterobacter, although they released 40% of fatty acid esters, release only 25% of hexosamine. Proteus released less than 15% of either fatty acid esters or hexosamine. These differences are taken to explain the differences among the Enterobacteriaceae in releasing surface enzymes after osmotic shock. It is felt that the release of additional lipopolysaccharide after osmotic shock is necessary for the release of surface enzymes that are not freed by ethylenediaminetetraacetic acid-tris(hydroxymethyl)aminomethane exposure.     

Occurrence and characteristics of the cytolysin A gene in Shigella strains and other members of the family Enterobacteriaceae

Cytolysin A (ClyA, HlyE, SheA) is a hemolytic pore-forming toxin found in Escherichia coli and Salmonella enterica serovars Typhi and Paratyphi A. In the present study, analysis of several Shigella strains revealed that they harbor only nonfunctional clyA gene copies that have been inactivated either by the integration of insertion sequence (IS) elements (Shigella dysenteriae, Shigella boydii, and Shigella sonnei strains) or by a frameshift mutation (Shigella flexneri). Shigella dysenteriae and S. boydii strains also exhibited IS-associated deletions at the clyA locus. PCR and Southern blot analyses as well as database searches indicated that clyA-related DNA sequences are completely absent in strains belonging to various other genera of the family Enterobacteriaceae. According to these data, ClyA may play a role only for a rather small subset of the enteric bacteria.     

Occurrence of Cyclopiazonic Acid in Peanuts

Samples of segregation 3 farmer stock peanuts from the 1980 southeastern United States growing season were analyzed for the presence of cyclopiazonic acid and aflatoxins. Cyclopiazonic acid appeared in 21 of 27 loose-shell kernel fractions at a range of 32 to 6,525 μg/kg and in 4 of 21 sound mature kernel fractions at a range of 32 to 130 μg/kg. Aflatoxins were detected in 26 of 27 loose-shell kernel fractions and in 20 of 21 sound mature kernel fractions. Cyclopiazonic acid used at 105 and 210 μg/kg to spike peanut samples was recovered at an average rate of 93.3%, with ranges of 89 to 119 and 166 to 221 μg/kg, respectively. The minimum detection limit on oxalic acid-impregnated silica gel plates was 26 ng.     

Occurrence of P.fimbrii in strains from selected genera of Enterobacteriaceae]

This study was aimed at recognition of frequency of occurrence of P fimbriae in strains of Escherichia coli isolated from samples of feces of children with symptoms of diarrhoea and at search of these adhesions in strains representing other than Escherichia genera of Enterobacteriaceae strains. One hundred forty laboratory strains were investigated. They belonged to genus Citrobacter, Enterobacter, Hafnia, Klebsiella, Morganella, Proteus, Providencia, Salmonella, Shigella, and Yersinia. Also were tested 1277 colonies of enteric rods isolated from the MacConkey's medium inoculated with samples of feces from 163 children with symptoms of diarrhoea. Mannose-resistant active hemagglutination test was performed with human group O erythrocytes and guinea pig erythrocytes stabilized with glutaraldehyde. Presence of P fimbriae was detected by the slide latex test with latex covered by P1 glycoprotein. Among 140 laboratory strains of Enterobacteriaceae in 21 strains (3-E. cloaceae, 2-Hafnia, 13-K. pneumoniae, 2-P. rettgeri and in one strains of Providencia) presence of MRHA adhesins was demonstrated. Nine of these strains (2-Hafnia, 5-K. pneumoniae and 2-P. rettgeri) reacted specifically in the latex test. Among 1142 colonies of E. coli isolated from children with symptoms of diarrhoea, 326 colonies belonged to 13 EPEC serotypes. With 118 (36.2%) of EPEC colonies a positive result of MRHA reaction was found with human erythrocytes and 34 (10.4%) with guinea pig erythrocytes. Positive latex test was obtained with 77 (23.6%) colonies. All these colonies possessed MRHA adhesins. Remaining 816 colonies of E. coli strains did not represent microorganisms belonging to serotypes accepted as enteropathogenic. From 112 (13.7%) colonies out of 816 not belonging to EPEC, positive results was obtained in the MRHA test with human erythrocytes and this was the case also with 41 (5.0%) colonies in MRHA reaction with application of guinea pig erythrocytes. The latex test was positive in 65 (7.9%) colonies of E. coli. From remaining 135 colonies other than E. coli, positive result of latex test of presence of P fimbriae was obtained with 54 (40.0%) colonies, including 14 colonies of E. cloacae, 23-K. pneumoniae and 17-K. oxytoca. In all these strains presence of MRHA adhesins was demonstrated. These investigations demonstrated that among EPEC strains significantly more frequently, than not belonging to these serotypes of E. coli, MRHA adhesins, including P fimbriae was observed.(ABSTRACT TRUNCATED AT 400 WORDS)     

Occurrence of Strains Producing Specific Antibacterial Inhibitory Agents in Five Genera of Enterobacteriaceae

Striking differences in the production of specific inhibitory agents affecting other strains of the same (or of related) species were found between genera of the family Enterobacteriaceae. We tested 50–163 strains each of the potentially pathogenic genera: Escherichia, Citrobacter, Enterobacter, Kluyvera, and Leclercia for their ability to produce bacteriophages, high-molecular-weight (HMW) and low-molecular-weight (LMW) bacteriocins and siderophores against the same sets of strains, using the cross-test method. The genus Escherichia differs substantially from all other Enterobacteriaceae, harboring a notable proportion of lysogenic (36.6%) and colicinogenic (13.9%) strains. Only 18.2% of the Citrobacter strains are lysogenic and only rarely are they colicinogenic, although in 7.3%, they produce phage tail-like bacteriocins. On the other hand, Kluyvera strains were only in 1.8% lysogenic, no colicinogenic strains were found, but in 7.3%, they produced siderophores causing zones of growth inhibition in agar cultures of strains of the same genus. In Leclercia, 10.0% of the strains were lysogenic, 2.0% produced HMW bacteriocins, no colicinogenic strains were found and 2.0% produced siderophores. Enterobacter has shown 23.1% of strains producing siderophores, whereas merely 7.7% were lysogenic, 1.9% colicinogenic and 3.8% formed phage tail-like bacteriocins. HMW bacteriocins of Enterobacter strains disposed of an unusually wide spectrum of activity. The siderophore activity spectrum was rather wide in any genus, but the siderophores were usually not produced by strains producing phages or colicins.     

galU基因在乳酸乳球菌中的克隆及其对胞外多糖产生的影响

从EcoliBL21克隆到UDP-葡萄糖焦磷酸化酶（UGPase）基因galU,与pNZ8048载体连接构建重组表达质粒pNZ8048-galU,进而导入乳酸乳球菌L.lactisL18中,得到重组菌L.lactisL18／pNZS048-galU,研究galU插入对该菌产生胞外多糖的影响。结果显示,在含葡萄糖和乳糖（20：20g／L）的MRS培养基中,重组菌L.lactisL18／pNZ8048-galU在30℃,pH6．5的条件下培养26h,EPS产量最高,为1489．54mg／L;而相同条件下,L．lactisL18培养28h产量最高,为848．93mg／L。二者相比,EPS产量增加了1．75倍。     

Occurrence of Glucomuramic Acid in Gram-Positive Bacteria

Analyses of 48 gram-positive bacteria indicate only glucomuramic acid and no galactomuramic acid in cell walls.     

Occurrence of beta-Aminoglutaric Acid in Marine Bacteria

beta-Aminoglutaric acid, a nonprotein amino acid isomer of glutamic acid, was found in the free amino acid pool of a marine bacterium, Alteromonas luteoviolacea. It was also found in a mixed culture of fermenting bacteria enriched from an anoxic marine sediment.     

Distribution of Indole-3-Acetic Acid and the Occurrence of Its Alkali-Labile Conjugates in the Extraxylary Region of Pinus sylvestris Stems

Free and conjugated indole-3-acetic acid (IAA) were measured by quantitative gas chromatography-selected ion monitoringmass spectrometry in the extraxylary region of the stem of large Pinus sylvestris (L.) trees during the annual cycle of cambial activity and dormancy. The extraxylary region at the stem top and bottom was divided into 3 and 4 fractions, respectively, for the free IAA measurements, while the entire extraxylary region was extracted when the IAA-conjugates were analyzed. The effect on the distribution pattern of expressing IAA level as a concentration (per gram fresh weight or dry weight) and as total amount (per square centimeter) was examined. The IAA level was much higher in the cambial region than in the fractions that contained the nonfunctional phloem and the periderm. The largest IAA concentration occurred in the fraction that included the cambium, whereas the total amount of IAA was greatest in the phloemcontaining fraction. The significance of the nonuniform radial distribution of IAA for estimating the IAA concentration in the cambial region is discussed in relation to how the cambial region is sampled. A slight Iongitudinal gradient in IAA concentration, decreasing from the top to the bottom of the stem, was observed in the cambial region when the cambium was in the grand period of activity, but not at the end of the cambial growing period. In all fractions, the total amount of IAA was highest when the cambium was active. However, the IAA concentration in the cambial region did not follow the same pattern, actually being lowest during the tracheid production period at the stem bottom. IAA conjugates were detected on all sampling dates except June 23, but their concentrations were always less than 14% of that of free IAA, and their occurrence did not obviously vary during the year. In general, there was a higher concentration of ester conjugates than of amide conjugates, and the ester conjugates were more abundant at the top of the stem than at the bottom.     

Occurrence of Uronic Acid in Lipopolysaccharides of Vibrionaceae

The occurrence of uronic acid as a sugar constituent of lipopolysaccharides (LPS) in Vibrionaceae was demonstrated for the first time. More than 100 strains were examined. Of five genera constituting Vibrionaceae, i.e., Vibrio, Aeromonas, Plesiomonas, Photobacterium, and Lucibacterium, the latter three contained uronic acid in LPS of all of their constituting members examined, while it was totally lacking in Aeromonas LPS so far tested. Only the members of genus Vibrio were found to be divided into uronic acid-containing and -lacking groups; V. parahaemolyticus, V. alginolyticus, V. fisheri, V. costicola, Beneckea (‘Vibrio’), and V. fluvialis belonged to the former, while all four biotypes of V. cholerae regardless of their serotypes, V. vulnificus and V. anguillarum, belonged to the latter group. The uronic acid content of V. parahaemolyticus O1 to O12 LPS ranged from 1.6 to 4.2%. The uronic acid residue released from V. parahaemolyticus O1, O4, O10, and O12 LPS by heating in 5% acetic acid at 100 C for 2 hr was identified as galacturonic acid; in particular, that from 012 LPS was characterized as d-galacturonic acid.     

Occurrence of Crithidia Factors and Folic Acid in Various Bacteria

Crithidia factors and folic acid were found to be widely distributed in culture fluids and in cells of 27 species of bacteria, when cultured under aerobic conditions into the stationary phase. Most bacteria excreted more Crithidia factors and folic acid than they retained in their cells. One Crithidia factor produced by Serratia indica and one produced by Bacillus cereus differed from biopterin in their chromatographic behavior. The factor excreted by S. indica appeared to be a 2-amino-4-hydroxy-6-substituted pteridine on the basis of KMnO(4) oxidation and ultraviolet absorption spectra. One of the folate compounds excreted by this organism was shown to be identical to 5,10-methylidynetetrahydrofolic acid by bioautography.