The impact of biofilm-forming potential and tafi production on transport of environmental <Emphasis Type="Italic">Salmonella</Emphasis> through unsaturated porous media

Understanding the factors influencing the transport of microbial pathogens, such as Salmonella and Escherichia coli, through porous media is critical to protecting drinking water supplies. The production of biofilms, along with individual biofilm-associated components, such as tafi, is believed to hinder transport of microorganisms through soil. This study investigated the relationship between biofilm formation and tafi production and the transport of environmental Salmonella through porous media. Thirty-two Salmonella isolates were initially assayed for their ability to form biofilms, from which a subset of these was selected to represent a range of high and low biofilm-formation potential and tafi formation capabilities. These were subsequently examined in unsaturated sand columns for transport characteristics. No obvious correlation was observed between Salmonella phenotypes and column retention. The results indicated that while transport of well-characterized laboratory E. coli strains can often be hindered by the presence of tafi and the potential to form biofilms, the presence of tafi did not retard the transport of the Salmonella strains.     

Interactions Between Escherchia coli and the Colloids of Three Variable Charge Soils and Their Effects on Soil Surface Charge Properties

The adhesion of Escherchia coli (E. coli) to the colloids of three variable charge soils and its effect on surface charge properties and potassium adsorption of these soil colloids were investigated. The adhesion isotherms of E. coli by soil colloids can be described using the Langmuir equation. The amount of E. coli adhered by the soil colloids varied with soil type and followed the order: Ultisol from Guangxi > Oxisol from Yunnan > Ultisol from Jiangxi. The iron and aluminum oxide contents and CECs of the soils are the important factors affecting the adhesion of E. coli to soil colloids. The relatively lower iron and aluminum oxide contents and higher CEC of the Ultisol from Jiangxi led to the lower adhesion of E. coli to the soil colloids compared to the Ultisol from Guangxi and the Oxisol from Yunnan. The amount of E. coli adhered to the soil colloids decreased with increasing pH, which was consistent with the results predicted from the DLVO theory. E. coli adhesion made the zeta potential of the soil colloids more negative and reduced the isoelectric point of the soil colloids, suggesting that E. coli decreased the surface positive charge and increased negative charge of the soil colloids. In addition, E. coli adhesion increased K⁺ adsorption by the soil colloids. Therefore, bacterial adhesion improves the fertility of variable charge soils by increasing soil CEC because the CECs of variable charge soils are usually low.     

Iron content of sediment and phosphate adsorption properties

Phosphorus can occur in sediments in different forms and accordingly its availability varies. The distinction between the phosphorus fractions is made with two chemical extraction methods; an ammonium oxalate-oxalic acid extraction and an extraction according to Hieltjes & Lijklema (1980).The iron and aluminum liberated with the ammonium oxalate-oxalic acid extraction method is linearly correlated (r ² = 0.73) with the phosphorus liberated in the first two steps of the Hieltjes and Lijklema extraction by: P = 0.035 (Fe + Al) + 0.001 (P, Fe and Al in mmol g^–1).The iron and aluminum (hydr)oxides are very important fractions in the sediment adsorption capacity for phosphorus. The phosphorus sorption capacity (PSC) is 0.080 mol P (mol (Fe + Al))^–1 and the adsorption constant (k) is 11.9 µmol P l^–1. Here it is assumed that iron and aluminum (hydr)oxide have the same affinity for phosphorus.     

Multiple copies of the insertion-DNA sequences IS1 and IS2 in the chromosome of E. coli K-12

Summary About 8 copies of the DNA sequence IS1 (which consists of 800 nucleotide pairs) and about 5 copies of the DNA sequence IS2 (1400 nucleotide pairs) have been detected by DNA-DNA hybridization in the chromosome of E. coli K-12. No homology is observed between IS1 and IS2. Both IS1 and IS2 are also found in the DNA of the F plasmid.     

Purification and properties of the Escherichia coli nucleoside transporter NupG,a paradigm for a major facilitator transporter sub-family

NupG from Escherichia coli is the archetype of a family of nucleoside transporters found in several eubacterial groups and has distant homologues in eukaryotes, including man. To facilitate investigation of its molecular mechanism, we developed methods for expressing an oligohistidine-tagged form of NupG both at high levels (>20% of the inner membrane protein) in E. coli and in Xenopus laevis oocytes. In E. coli recombinant NupG transported purine (adenosine) and pyrimidine (uridine) nucleosides with apparent K_m values of ～20–30 μM and transport was energized primarily by the membrane potential component of the proton motive force. Competition experiments in E. coli and measurements of uptake in oocytes confirmed that NupG was a broad-specificity transporter of purine and pyrimidine nucleosides. Importantly, using high-level expression in E. coli and magic-angle spinning cross-polarization solid-state nuclear magnetic resonance, we have for the first time been able directly to measure the binding of the permeant ([1′-¹³C]uridine) to the protein and to assess its relative mobility within the binding site, under non-energized conditions. Purification of over-expressed NupG to near homogeneity by metal chelate affinity chromatography, with retention of transport function in reconstitution assays, was also achieved. Fourier transform infrared and circular dichroism spectroscopy provided further evidence that the purified protein retained its 3D conformation and was predominantly α-helical in nature, consistent with a proposed structure containing 12 transmembrane helices. These findings open the way to elucidating the molecular mechanism of transport in this key family of membrane transporters.     

Study on the toxic effect of lead(II) ion on <Emphasis Type="Italic">Escherichia coli</Emphasis>

The toxic effects of lead(II) have been studied in Escherichia coli cells. Using microcalorimetric analysis, it was shown that E. coli growth was inhibited in the presence of Pb²⁺ resulting from damage to the cell membrane and that Pb²⁺ takes part in the metabolism of cells. Treatment with lysozyme confirms damage to the cell’s outer membrane. Similarities between the ionic radii and charge/radius ratio cause Pb(II) to replace Ca(II) at the binding sites of lipopolysacharides, leading to rupture of protecting areas on the cell’s surface. Consequently, the protection and functionality of outer membrane is lost, thus becoming the basis for the biological effect of Pb²⁺ on E. coli.     

Effect of copper hydroxide sprays for citrus canker control on wild-type Escherichia coli

Aims: To show that application of copper hydroxide citrus sprays mixed with field source water (possibly contaminated) will not support Escherichia coli on plant surfaces. Environmental stresses of transient phyllosphere bacteria and presence of copper will eradicate these bacteria before harvest. Methods and Results: Studies were performed in vitro with bacteria grown in broth and then subjected to field spray copper hydroxide concentrations in the broth and on citrus leaves. Escherichia coli exposed to copper hydroxide in vitro were eradicated from the broth within 6–8 h depending on the broth pH. Even with near neutral pH (7·2), cells began to die immediately after exposure to copper. No E. coli survived on leaf surfaces sprayed with copper. Conclusions: Copper field sprays mixed with water that may contain E. coli can help eliminate E. coli from plant surfaces. Significance and Impact of the Study: HACCP mandates are becoming more restrictive because of the increased illness resulting from food pathogens on fresh produce. Use of potable water in fields, a proposed mandate, is not feasible for large grove owners. These data show that copper sprays aimed at reducing citrus canker also affect E. coli and may help to ease water quality mandates.     

Transport of vanadium (V) in saturated porous media: effects of pH,ionic-strength and clay mineral

Vanadium, a hazardous pollutant, has been frequently detected in soil and groundwater, however, its transport behavior in porous media were not clearly understood. In this study, the effects of solution pH, ionic strength (IS) and the effect of clay mineral on the transport of vanadium in saturated porous media were investigated. Laboratory experiments using a series of columns packed with quartz sand were carried out to explore the retention and transport of vanadium with a range of ionic-strength (0.001–0.1 M) and pH (4–8) and two different types of clay minerals montmorillonite and kaolinite. Results of the breakthrough experiments showed that vanadium was highly mobile in the saturated porous media. The increase in pH rendered a higher transport of vanadium in saturated porous media. The study also indicated an easier transfer of vanadium with an increase in IS. Montmorillonite enhanced the mobility of vanadium in the column when compared to kaolinite. A mathematical model based on advection-dispersion equation coupled with equilibrium and kinetic reactions was used to describe the retention and transport of vanadium in the columns very well.     

The influence of biofilm structure and total interaction energy on Escherichia coli retention by Pseudomonas aeruginosa biofilm

The retention of a surrogate pathogenic bacterium, Escherichia coli^T , in Pseudomonas aeruginosa biofilms (with various EPS excreting capacities) was investigated using a laboratory flow cell system. The structural characteristics of the biofilm, as well as the quantity of E. coli^T retained in the biofilm, were assessed using confocal laser scanning microscopy coupled with image analysis. In addition, the total interaction energy between E. coli^T and the P. aeruginosa biofilm was computed with the Derjaguin-Landau-Verwey-Overbeek (DLVO) theory, which provided an additional context to explain the pathogen interaction in aquatic biofilms. The correlations between the quantity of detained E. coli^T cells and the structural characteristics of the biofilm were analysed and the results indicated that the heterogeneity of the biofilm could create a quiescent zone leading to temporary retention of E. coli^T within the biofilm. Overall, this study provided insights toward understanding the retention of pathogenic bacteria in environmental biofilms.     

Study of iron homeostasis following partial hepatectomy in rats with chronic aluminum intoxication

Effects of both chronic aluminum (Al) exposure and partial hepatectomy on iron (Fe) homeostasis were studied. Male Wistar rats were intraperitoneally administered either 27 mg Al/kg body weight (as aluminum hydroxide) or the vehicle saline, three times a week for 3 mo. After this time, half of the rats of each group was sham operated (SH) and the other half was partially hepatectomized (PH). Animals of the four experimental groups (vehicle+SH [SH]; Al+SH; vehicle+PH [PH], and Al+PH) were killed 48h after the surgical procedure. Serum, hepatic, and intestinal Al levels were found to be increased both for Al+SH and Al+PH. The serum Fe concentration and transferrin saturation percentage were significantly diminished in the rats of the Al+PH group, thus showing interaction between Al administration and PH. The ⁵⁰Fe mucosal-to-serosal transport, studied in the intestinal loop in situ, was not affected by Al or PH. The malregulation of intestinal Fe absorption in Al exposure and/or PH when the serum Fe concentration was diminished could be the result of the increased lipid peroxidation (thiobarbituric acid-reactive substances [TBARS]) observed in this tissue. Mucosal TBARS were increased by Al exposure (+26%) and PH (+37%) and interaction between Al and PH was observed (+44%). These results show that when liver surgery is performed after prolonged Al exposure, it leads to impairment of Fe homeostasis. We underline the importance of the exposure to Al, a potentially toxic element, in the study of risk assessment in patients who must be submitted to major liver resection.     

Genomic comparisons among <Emphasis Type="Italic">Escherichia coli</Emphasis> strains B,K-12, and O157:H7 using IS elements as molecular markers

Background  

Insertion Sequence (IS) elements are mobile genetic elements widely distributed among bacteria. Their activities cause mutations, promoting genetic diversity and sometimes adaptation. Previous studies have examined their copy number and distribution in Escherichia coli K-12 and natural isolates. Here, we map most of the IS elements in E. coli B and compare their locations with the published genomes of K-12 and O157:H7.     

Identification of the ferrioxamine B receptor,FoxB, inEscherichia coli K12

The photoreactivep-azidobenzoyl analog of ferrioxamine B was used to show that ferrioxamine-B-mediated iron transport is separate and distinct from coprogen-mediated iron transport inEscherichia coli. Photolysis of this analog inhibited uptake of [⁵⁹Fe]ferrioxamine B but not [⁵⁹Fe]coprogen or [⁵⁹Fe]ferrichrome. Conversely, photolysis of thep-azidobenzoyl analog of coprogen B inhibited uptake of [⁵⁹Fe]coprogen but not [⁵⁹Fe]ferrioxamine B or [⁵⁹Fe]ferrichrome. Photolabeling of outer membranes withp-azidobenzoyl-[⁵⁹Fe]ferrioxamine B resulted in the labeling of two iron-regulated peptides with molecular masses of about 66 and 26 kDa. Expression of these peptides was increased when ferrioxamine B was the sole iron source. Both peptides were present in outer membrane preparations of thefhuF mutant H1717, but the 66 kDa peptide was not inducible. These results are evidence for an outer membrane receptor inE. coli unique for linear ferrioxamines.     

Cloning of the nupC gene of Escherichia coli encoding a nucleoside transport system, and identification of an adjacent Insertion element, IS 186

Escherichia coli is known to contain more than one active transport system for nucleoside uptake. In the present study we report the sequence of a gene encoding a second nucleoside transport system, nupC (in addition to nupG.) An open reading frame (ORF) of 1200bp was identified that codes for a hydrophobic polypeptide of 43 560 Da and an NupC fusion protein was shown to be membrane associated. The native NupC protein is also identified, following over-expression. NupC exhibits short regions of homology to several membrane-associated proteins, including LacY and Cyd. Analysis of the nupC promoter region revealed the presence of at least two putative CRP-binding sites, centred at–40bp and–89bp, which probably flank a CytR-binding site. In addition, an adjacent IS186 element was identified and found to reside within a putative terminator structure, downstream from the nupC ORF. This arrangement is shown to reflect the previously established gene order on the E. coli chromosome.     

Cyclic antimicrobial R-, W-rich peptides: the role of peptide structure and <Emphasis Type="Italic">E. coli</Emphasis> outer and inner membranes in activity and the mode of action

This study compares the effect of cyclic R-, W-rich peptides with variations in amino acid sequences and sizes from 5 to 12 residues upon Gram negative and Gram positive bacteria as well as outer membrane-deficient and LPS mutant Escherichia coli (E. coli) strains to analyze the structural determinants of peptide activity. Cyclo-RRRWFW (c-WFW) was the most active and E. coli-selective sequence and bactericidal at the minimal inhibitory concentration (MIC). Removal of the outer membrane distinctly reduced peptide activity and the complete smooth LPS was required for maximal activity. c-WFW efficiently permeabilised the outer membrane of E. coli and promoted outer membrane substrate transport. Isothermal titration calorimetric studies with lipid A-, rough-LPS (r-LPS)- and smooth-LPS (s-LPS)-doped POPC liposomes demonstrated the decisive role of O-antigen and outer core polysaccharides for peptide binding and partitioning. Peptide activity against the inner E. coli membrane (IM) was very low. Even at a peptide to lipid ratio of 8/1, c-WFW was not able to permeabilise a phosphatidylglycerol/phosphatidylethanolamine (POPG/POPE) bilayer. Low influx of propidium iodide (PI) into bacteria confirmed a low permeabilising ability of c-WFW against PE-rich membranes at the MIC. Whilst the peptide effect upon eukaryotic cells correlated with the amphipathicity and permeabilisation of neutral phosphatidylcholine bilayers, suggesting a membrane disturbing mode of action, membrane permeabilisation does not seem to be the dominating antimicrobial mechanism of c-WFW. Peptide interactions with the LPS sugar moieties certainly modulate the transport across the outer membrane and are the basis of the E. coli selectivity of this type of peptides.     

The sequence of IS4

Summary IS-elements are devoid of easily recognizable transacting functions and exert their visible effects in the position cis only (recent reviews Calos and Miller 1980; Starlinger 1980). It has been a matter of debate, whether these elements encode functions for their own transposition. In the case of the E. coli IS-elements this could not easily be determined by genetic methods, because most of these elements are present in several copies (Saedler and Heiss 1973; Deonier et al. 1979). In the case of the IS-elements flanking transposons, evidence has recently been brought forward that these carry the transposition specificity (Rothstein et al. 1980; Kleckner 1980; Grindley 1981).IS4 is present in one copy only in several E. coli K12 strains and should, therefore, be suitable for genetic and physiological studies (Chadwell et al. 1979). It has been cloned from several sites on the E. coli chromosome in pBR322 (Klaer and Starlinger 1980). Here we report the DNA sequence of IS4 which contains an open reading frame for 442 amino acids, and of the junctions of this element with surrounding DNA at three different sites in E. coli chromosome.     

Distinct roles for U‐type proteins in iron–sulfur cluster biosynthesis revealed by genetic analysis of the Bacillus subtilis sufCDSUB operon

The biosynthesis of iron–sulfur (Fe–S) clusters in Bacillus subtilis is mediated by the SUF‐like system composed of the sufCDSUB gene products. This system is unique in that it is a chimeric machinery comprising homologues of E. coli SUF components (SufS, SufB, SufC and SufD) and an ISC component (IscU). B. subtilis SufS cysteine desulfurase transfers persulfide sulfur to SufU (the IscU homologue); however, it has remained controversial whether SufU serves as a scaffold for Fe–S cluster assembly, like IscU, or acts as a sulfur shuttle protein, like E. coli SufE. Here we report that reengineering of the isoprenoid biosynthetic pathway in B. subtilis can offset the indispensability of the sufCDSUB operon, allowing the resultant Δsuf mutants to grow without detectable Fe–S proteins. Heterologous bidirectional complementation studies using B. subtilis and E. coli mutants showed that B. subtilis SufSU is interchangeable with E. coli SufSE but not with IscSU. In addition, functional similarity in SufB, SufC and SufD was observed between B. subtilis and E. coli. Our findings thus indicate that B. subtilis SufU is the protein that transfers sulfur from SufS to SufB, and that the SufBCD complex is the site of Fe–S cluster assembly.     

Escherichia coli exhibiting gram positivity after cetyl pyridinium chloride treatment

Summary Treatment with a solution of the detergent cetyl pyridinium chloride develops gram positivity in E. coli as is evident from the increased retention of crystal violet by the bacteria. Since cetyl pyridinium chloride has been found to remove larger quantity of lipid from E. coli than from the normal gram positive bacteria S. aureus, it is reasonable to think that lipid content in a bacteria may have an important role in determining the gram character of it. Gram negative bacteria generally contain more lipid than the gram positive ones.     

Aluminium in tea: SEC-ICP-MS speciation studies of infusions and simulated gastrointestinal digests

Abstract

The speciation of aluminium in tea infusions and in vitro gastrointestinal digests of tea infusions has been investigated using size exclusion chromatography coupled to inductively coupled plasma mass spectrometry (SEC-ICP-MS). At pH 2.5, following simulated gastric treatment, Al from tea eluted at a similar retention volume to that obtained for an aqueous Al standard. At pH 5.5, an aqueous Al standard was eluted from an SEC column in Tris buffer (Al recovery ≈ 100%) only in the presence of a complexing agent (NaF), and at a retention volume corresponding to a molecular mass greater than that expected for an ionic species. Aluminium associated with a tea infusion eluted in two fractions: a higher molecular weight fraction corresponding to Al strongly bound to ligands in the tea, and a lower molecular weight fraction probably comprised of labile Al eluting as Al-F complexes. Simulated gastrointestinal digestion produced three Al-bearing fractions, of which the two at higher molecular mass represented ligand-bound Al. The molarity of the Tris buffer strongly influenced the retention volume of the Al fractions, particularly for the ligand-bound Al. After simulated gastrointestinal digestion, the soluble labile fraction (15% of the Al from the tea infusion) was considered to be potentially available for absorption. The actual proportion of the fraction that might be absorbed would depend upon a number of physiological and nutritional factors.     

Host dependenet inactivation by IS2 of induced E.coli penicillin amidase gene cloned on multicopy plasmids

Structural instability of the cloned penicillin acylase gene (pac) from E.coli ATCC11105 was studied under various physiological conditions. Structural changes which adversely affect the expression of penicillin acylase gene were selected for only under conditions in which the gene was active and fully induced. In E.coli strain YMC the predominant mutations were the insertions of the IS2 element at various sites within the 700 bp proximal portion of the pac gene. The results indicated that the induction of the plasmid cloned gene was the main factor which rendered it a target for inactivation by insertions of the host encoded IS2 elements. The mutational events were host specific and they were not influenced by mutual positions and orientations of key marker genes on the plasmid.