Activity of p-aminobenzoic acid compared with other organic acids against selected bacteria

The antibacterial activity of p -aminobenzoic acid against Listeria monocytogenes, Salmonella enteritidis and Escherichia coli was compared with the activity of commonly used acidulants: formic, propionic, acetic, lactic and citric acids. Viable count evaluations and MIC determinations indicated that p -aminobenzoic acid caused greater inhibitory effects than the other organic acids. The activity of p -aminobenzoic acid on the growth of the test organisms at selected pH values indicated that p -aminobenzoic acid was more active at low pH than at high pH. Uptake studies showed that the uptake of p -aminobenzoic acid by E. coli was markedly decreased as the pH values increased. Electron micrographs of E. coli cells grown in the presence of p -aminobenzoic acid indicate that p -aminobenzoic acid caused marked damage to the cell envelope. It is suggested that p -aminobenzoic acid has at least two mechanisms of action: one mechanism in common with other organic acids and the other mechanism by interfering with the synthesis of the peptidoglycan layer by an action on the dihydrofolate reductase enzyme.     

The Activity of Pipemidic Acid and Piromidic Acid against Escherichia coli In Static Culture and in Conditions Simulating the Treatment of Bacterial Cystitis

The activity of the new pyridopyrimidine compounds, pipemidic and piromidic acids, against Escherichia coli was compared with that of nalidixic acid in vitro . In a static turbidimetric system nalidixic and pipemidic acids were found to be more active than piromidic acid when tested against sensitive E. coli strains, but pipemidic acid was the most active of the three compounds against nalidixic acid-resistant clinical isolates. When tested in an in vitro model designed to mimic the conditions in which bacteria and drug interact in the treatment of bacterial cystitis, all three drugs were found to be able to suppress bacterial growth for long periods, but a high, sustained drug level was necessary in order to prevent the emergence of resistant variants.     

Antibacterial activity of fresh and processed red muscadine juice and the role of their polar compounds on Escherichia coli O157:H7

Aims:  The objectives of this research were to show the anti- Escherichia coli O157:H7 effect of fresh (FRMJ) and processed red muscadine (V itis rotundifolia ) juice (PRMJ) and to discern the active compounds responsible for anti- E . coli O157:H7.
Methods and Results:  Polar and phenolic compounds of FRMJ and PRMJ were analysed by high-performance liquid chromatography. Antibacterial activity of FRMJ, PRMJ, their polar and polyphenol fractions, individual synthetic acids and their mixture with or without sugars were investigated on E . coli O157:H7. FRMJ and PRMJ inactivated ( P  ≤ 0·05) 5-log cocktail cells of E. coli O157:H7 within 4 h at 37°C. Polar fractions that contained malic, tartaric and tannic acids showed strong antimicrobial activity ( P  ≤ 0·05) against E . coli O157:H7. Tannic acid among the synthetic acids showed the highest antimicrobial activity against E. coli O157:H7.
Conclusions:  FRMJ, PRMJ and their polar compounds showed strong anti- E . coli O157:H7 activity.
Significance and Impact of the Study:  Earlier findings have failed to show any anti -E . coli O157:H7 effect of grape juice without adding preservatives. Our findings show that red muscadine juice has natural antibacterial substances and suggest that these can be used as active antimicrobial ingredients against E . coli O157:H7 in nonalcoholic beverages.     

Studies on the antibacterial activity of phanquone: chelating properties in relation to mode of action against Escherichia coli and Staphylococcus aureus

Phanquone is active against a wide range of Gram-positive and Gram-negative organisms. Its activity is affected by the nature of the suspending fluid, pH and anaerobic growth conditions. Its ability to chelate metal ions was examined and found to be related to its antibacterial activity, which was reduced by the presence of added metal ions, e.g. Co (II), Cu(II), Fe(II) and Fe(III) in nutrient media for both E. coli and S. aureus. When antibacterial activity was examined in dis-nutrient media for both E. coli and S. aureus. When antibacterial activity was examined in distilled water, then certain added metal ions, whilst antagonizing activity was examined in distilled water, then certain added metal ions, whilst antagonizing the activity of Phanquone against E. coli, exerted a co-operative effect in the case of S. aureus. The addition of EDTA and NTA lowered the activity of Phanquone against S. aureus, but not E. coli, while the addition of thiol-containing compounds lowered its activity against E. coli but not S. aureus. concentration quenching was observed for S. aureus but not for E. coli, while overnight pre-incubation at 4 degrees C resulted in the appearance of a growth zone inside the zone of inhibition in the case of S. aureus but not E. coli. Phanquone may have a different mode of action against the two organisms.     

Cross-reconstitution of the F0F1-ATP synthases of chloroplasts and Escherichia coli with special emphasis on subunit delta

F0F1-ATP synthases catalyse ATP formation from ADP and Pi by using the free energy supplied by the transmembrane electrochemical potential of the proton. The delta subunit of F1 plays an important role at the interface between the channel portion F0 and the catalytic portion F1. In chloroplasts it can plug the protonic conductance of CF0 and in Escherichia coli it is required for binding of EF1 to EF0. We wanted to know whether or not delta of one species was effective between F0 and F1 of the other species and vice versa. To this end the respective coupling membrane (thylakoids, everted vesicles from E. coli) was (partially) depleted of F1 and purified F1, F1(-delta), and delta were added in various combinations to the F1-depleted membranes. The efficiency or reconstitution was measured in thylakoids via the rate of phenazinemethosulfate-mediated cyclic photophosphorylation and in E. coli everted vesicles via the degree of 9-amino-6-chloro-2-methoxyacridine fluorescence quenching. Addition of CF1 to partially CF1-depleted thylakoid vesicles restored photophosphorylation to the highest extent. CF1(-delta)+chloroplast delta, EF1, EF1(-delta)+E. coli delta were also effective but to lesser extent. CF1(-delta)+E. coli delta and EF1(-delta)+chloroplast delta restored photophosphorylation to a small but still significant extent. With F1-depleted everted vesicles prepared by repeated EDTA treatment of E. coli membranes, addition of CF1, CF1 (-delta)+chloroplast delta and CF1(-delta)+E. coli delta gave approximately half the extent of 9-amino-6-chloro-2-methoxyacridine fluorescence quenching as compared to EF1 or EF1(-delta)+E. coli delta by energization of the vesicles with NADH, while Ef1(-delta)+chloroplast delta was ineffective. All 'mixed' combinations were probably reconstitutively active only by plugging the protonic leak through the exposed F0 (structural reconstitution) rather than by catalytic activity. Nevertheless, the cross-reconstitution is stunning in view of the weak sequence similarity between chloroplast delta and E. coli delta. It favors a role of delta as a conformational transducer rather than as a proton conductor between F0 and F1.     

Microbiology of Saturated Salt Solutions and Other Harsh Environments. IV. New Observations of Ribonucleotide-Induced Recovery of KCl-Habituated Penicillium notatum

Salt-tolerant mutant Penicillium notatum sub-cultured in a glucose-peptone broth saturated with KCl shows continued attenuated growth when transferred to salt-free broth. Additional tests have shown E. coli S-RNA to be inferior to yeast RNA preparations, that base-free phosphate sources are inactive, but that nicotinamide adenine dinucleotide and flavine adenine dinucleotide are moderately active. All phosphate derivatives of adenine, cytosine and guanosine and inosine were active including 5'-polyphosphates, 3'(2')-monophosphates 5'-monophosphates, and adenine 3', 5'-cyclic monophosphate. Uracil derivatives were of low activity at best.Among base precursors, orotic acid was moderately active whereas imidazoles were not. The high activity of inosine 5'-phosphate a precursor of other purine nucleotides suggested that one mode of KCl action might involve a block in conversion of 4-amino-5-imidazole carboxamide ribonucleoside to the hypoxanthine nucleotide.     

Differential effects of prostaglandin synthetase stimulators on inhibition of cyclooxygenase.

The different effects of prostaglandin synthetase stimulators on inhibition of the cyclooxygenase by structurally distinct classes of nonsteroidal antiinflammatory agents suggest that the enzyme is altered by interaction with these stimulators. Reversible stimulation of prostaglandin synthetase activity by phenols and some other compounds and the relative influence of these stimulators on inhibitors of the cyclooxygenase were determined quantitatively. Two distinct classes of inhibitors were established. The fenamates were relatively weak inhibitors alone but were much more potent in the presence of phenolic compounds. In contrast, ibuprofen, indomethacin, and flurbiprofen were more potent than the fenamates and were reduced in effectiveness by the stimulators, as expected on the basis of two opposing actions. The relative potency of the cyclooxygenase stimulators (phenol greater than norepinephrine greater than tryptophan greater than benzoquinone greater than anisole) paralleled their synergistic action on the fenamates and their antagonist action on the nonfenamates. This correlation suggests that an enzyme alteration which leads to cyclooxygenase stimulation may also result in increased sensitivity to fenamates and decreased sensitivity to the other inhibitors, possibly by altering their capacity to bind.     

Cell-free complements in vivo expression of the E. coli membrane proteome

Reconstituted cell-free (CF) protein expression systems hold the promise of overcoming the traditional barriers associated with in vivo systems. This is particularly true for membrane proteins, which are often cytotoxic and due to the nature of the membrane, difficult to work with. To evaluate the potential of cell-free expression, we cloned 120 membrane proteins from E. coli and compared their expression profiles in both an E. coli in vivo system and an E. coli-derived cell-free system. Our results indicate CF is a more robust system and we were able to express 63% of the targets in CF, compared to 44% in vivo. To benchmark the quality of CF produced protein, five target membrane proteins were purified and their homogeneity assayed by gel filtration chromatography. Finally, to demonstrate the ease of amino acid labeling with CF, a novel membrane protein was substituted with selenomethionine, purified, and shown to have 100% incorporation of the unnatural amino acid. We conclude that CF is a novel, robust expression system capable of expressing more proteins than an in vivo system and suitable for production of membrane proteins at the milligram level.     

Bacterial ethylene synthesis from 2-oxo-4-thiobutyric acid and from methionine

The ability of selected bacterial cultures to synthesize ethylene during growth in nutrient broth supplemented with methionine or 2-oxo-4-methylthiobutyric acid (KMBA) was examined. Although most cultures transformed KMBA into ethylene, only those of Escherichia coli SPAO and Chromobacterium violaceum were able to convert exogenously added methionine to ethylene. In chemically defined media, E. coli SPAO produced the highest amounts of ethylene from methionine and KMBA. This capability was affected by the nature of the carbon source and the type and amount of nitrogen source used for growth. When glutamate was used as sole source of carbon and nitrogen for growth, the activity of the ethylenogenic enzymes was reduced to 25% of that observed with cultures grown with glucose and NH4Cl. Neither methionine nor KMBA significantly affected the ethylenogenic capacity of E. coli SPAO. Menadione and paraquat, compounds that generate superoxide radicals, stimulated ethylene synthesis by harvested cells, but not by cell-free extracts of E. coli SPAO. In addition, cells of Pseudomonas aeruginosa, which produced no ethylene in culture in the presence of exogenously added KMBA, yet possessed the necessary enzymes in an active form, were able to synthesize ethylene from KMBA when incubated with menadione or paraquat.     

Purification and characterization of the chaperonin 10 and chaperonin 60 proteins from Rhodobacter sphaeroides.

Two heat-shock proteins that show high identity with the Escherichia coli chaperonin 60 (groEL) and chaperonin 10 (groES) chaperonin proteins were purified and characterized from photolithoautotrophically grown Rhodobacter sphaeroides. The proteins were purified by using sucrose density gradient centrifugation and Mono-Q anion-exchange chromatography. In the presence of 1 mM ATP, the chaperonin 10 and chaperonin 60 proteins bound to each other and comigrated as a large complex during sucrose density gradient centrifugation. The native molecular weights of each protein as determined by gel filtration chromatography were 889,200 for chaperonin 60 and 60,000 for chaperonin 10. Chaperonin 60 is comprised of monomers with a molecular weight of 61,000 and chaperonin 10 is comprised of monomers with a molecular weight of 12,700 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Chaperonin 60 was 9.3% of the total soluble cell protein during photolithoautotrophic growth which increased to 28.5% following heat-shock treatment. When cells were grown photoheterotrophically or chemoheterotrophically, chaperonin 60 was reduced to 6.7% and 3.5%, respectively, of the total soluble protein. The N-terminal amino acid sequence of each protein was determined; chaperonin 60 of R. sphaeroides showed 72% identity to E. coli chaperonin 60 protein, and R. sphaeroides chaperonin 10 showed 45% identity with E. coli chaperonin 10. R. sphaeroides chaperonin 60 catalyzed ATP hydrolysis with a specific activity of 134 nmol min-1 mg-1 (kcat = 0.13 s-1) and was inhibited by R. sphaeroides chaperonin 10, but not E. coli chaperonin 10. The E. coli chaperonin 60 ATPase activity was inhibited by chaperonin 10 from both R. sphaeroides and E. coli.     

In vitro efficacy of bioactive extracts of 15 medicinal plants against ESbetaL-producing multidrug-resistant enteric bacteria

Alcoholic crude extracts and some fractions from 15 traditionally used Indian medicinal plants were investigated for their ability to inhibit the growth of extended spectrum beta-lactamases (ESbetaL)-producing multidrug-resistant enteric bacteria. The test bacteria Eschrichia coli and Shigella were resistant to 16-23 antibiotics with intermediate or resistance to beta-lactams (minimum inhibitory concentration (MIC) value range 16-1024 microg/ml). The crude plant extracts demonstrated zone of inhibition in the range of 11-29 mm against one or more test bacteria. On the basis of promising activity, 12 plants were selected to determine their efficacy in terms of MIC, which ranged from 0.64 mg/ml to 10.24 mg/ml. The extracts of Acorus calamus, Hemidesmus indicus, Holarrhena antidysenterica and Plumbago zeylanica demonstrated relatively high activity as compared to other plant extracts and were fractionated into acetone, ethyl acetate and methanol. Acetone fraction in most of the cases exhibited higher potency (low MIC value) as compared to ethyl acetate and methanol fraction. However, in Plumbago zeylanica, ethyl acetate fraction was most active. Synergistic interactions among crude extracts were demonstrated in the 12 different combinations against ESbetaL-producing E. coli (ESbetaL-02). Certain combinations exhibited significant synergy with enlargement of combined inhibition zone size by 5 mm. Interaction of crude extracts with five antibiotics (Tetracycline, ciprofloxacin, nalidixic acid, chloramphenicol and streptomycin) demonstrated synergistic interaction with tetracycline and ciprofloxacin by 10 and 3 plant extracts respectively. Phytochemical analysis and thin layer chromatography (TLC) bioautography of crude extracts showed the presence of alkaloids, phenols and flavonoids as active phytoconstituents. Most active fractions of four plants were subjected to Infrared spectroscopy and the major groups of compounds were detected. The plant extracts were further tested for their in vitro haemolytic activity to sheep erythrocytes and demonstrated no haemolysis at recommended doses. Further activity-guided fractionation of active fractions is needed to isolate and characterize the active principle in order to establish the mode of action against the ESbetaL-producing multidrug-resistant enteric bacteria and the mechanism of synergy.     

Expression of synthetic genes encoding bovine and human basic fibroblast growth factors (bFGFs) in Escherichia coli

Synthetic genes encoding bovine and human basic fibroblast growth factors (bFGFs) were assembled and cloned using established Escherichia coli expression plasmids. Transformed E. coli cells were able to synthesize either a fusion protein, comprising the first seven amino acids of beta-galactosidase, a linker fragment and bovine FGF, or genomic human bFGF. The two growth factors were purified from E. coli lysates by cation exchange and heparin-Sepharose affinity chromatography. The purified recombinant proteins were biologically active as monitored by their mitogenic activity for bovine aortic endothelial cells and their angiogenic capacity in the rabbit cornea.     

Tryptophan-dependent production of indole-3-acetic acid (IAA) affects level of plant growth promotion by Bacillus amyloliquefaciens FZB42

Phytohormone-like acting compounds previously have been suggested to be involved in the phytostimulatory action exerted by the plant-beneficial rhizobacterium Bacillus amyloliquefaciens FZB42. Analyses by high-performance liquid chromatography and gas chromatography-mass spectrometry performed with culture filtrates of FZB42 demonstrated the presence of indole-3-acetic acid (IAA), corroborating it as one of the pivotal plant-growth-promoting substances produced by this bacterium. In the presence of 5 mM tryptophan, a fivefold increase in IAA secretion was registered. In addition, in the trp auxotrophic strains E101 (deltatrpBA) and E102 (deltatrpED), and in two other strains bearing knockout mutations in genes probably involved in IAA metabolism, E103 (deltaysnE, putative IAA transacetylase) and E105 (deltayhcX, putative nitrilase), the concentration of IAA in the culture filtrates was diminished. Three of these mutant strains were less efficient in promoting plant growth, indicating that the Trp-dependent synthesis of auxins and plant growth promotion are functionally related in B. amyloliquefaciens.     

Only one of the two annotated Lactococcus lactis fabG genes encodes a functional beta-ketoacyl-acyl carrier protein reductase

The small genome of the Gram-positive bacterium Lactococcus lactis ssp. lactis IL1403 contains two genes that encode proteins annotated as homologues of Escherichia coli beta-hydroxyacyl-acyl carrier protein (ACP) reductase. E. coli fabG encodes beta-ketoacyl-acyl carrier protein (ACP) reductase, the enzyme responsible for the first reductive step of the fatty acid synthetic cycle. Both of the L. lactis genes are adjacent to (and predicted to be cotranscribed with) other genes that encode proteins having homology to known fatty acid synthetic enzymes. Such relationships have often been used to strengthen annotations based on sequence alignments. Annotation in the case of beta-ketoacyl-ACP reductase is particularly problematic because the protein is a member of a vast protein family, the short-chain alcohol dehydrogenase/reductase (SDR) family. The recent isolation of an E. coli fabG mutant strain encoding a conditionally active beta-ketoacyl-ACP reductase allowed physiological and biochemical testing of the putative L. lactishomologues. We report that expression of only one of the two L. lactis proteins (that annotated as FabG1) allows growth of the E. coli fabG strain under nonpermissive conditions and restores in vitro fatty acid synthetic ability to extracts of the mutant strain. Therefore, like E. coli, L. lactis has a single beta-ketoacyl-ACP reductase active with substrates of all fatty acid chain lengths. The second protein (annotated as FabG2), although inactive in fatty acid synthesis both in vivo and in vitro, was highly active in reduction of the model substrate, beta-ketobutyryl-CoA. As expected from work on the E. coli enzyme, the FabG1 beta-ketobutyryl-CoA reductase activity was inhibited by ACP (which blocks access to the active site) whereas the activity of FabG2 was unaffected by the presence of ACP. These results seem to be an example of a gene duplication event followed by divergence of one copy of the gene to encode a protein having a new function.     

The action of human and rabbit serum phospholipase A2 on Escherichia coli phospholipids

We have compared the properties of phospholipase A (E.C. 3.1.1.4) activity in whole human and rabbit serum toward the phospholipids of Escherichia coli. Using as substrate E. coli labeled during growth with either [1-(14)C]-palmitic acid or [1-(14)C]oleic acid, and then autoclaved to inactivate E. coli phospholipases and to render the labeled phospholipids accessible to exogenous phospholipases, we show that the deacylating activity in both human and rabbit serum is almost exclusively of the A(2) type. Rabbit serum is at least 20-fold more active than human serum. Activity in both sera is maximal at physiological Ca(2+) concentrations (2 mM) and is abolished by ethylenediaminetetraacetic acid. To examine hydrolysis of intact (unautoclaved) E. coli treated with 25% serum, use was made of a phospholipase A-deficient E. coli strain (E. coli S17), thereby eliminating the possible contribution of bacterial phospholipases to degradation. Human and rabbit serum are about equally bactericidal toward E. coli and cause comparable structural damage. However, only rabbit serum produces substantial hydrolysis of the phospholipids of intact E. coli S17. Heated (56 degrees C, 30 min) rabbit serum is non-bactericidal and retains phospholipase A(2) activity toward autoclaved, but not intact E. coli. The ability of heated serum to degrade phospholipids of intact E. coli S17 is restored, however, by adding 25% normal human serum, which is bactericidal. In this combination, doses of heated rabbit serum containing as much phospholipase A(2) activity (toward autoclaved E. coli) as is present in 25% unheated rabbit serum, produce roughly the same extent of hydrolysis of intact E. coli as does normal rabbit serum alone. Low doses with a phospholipase A(2) activity comparable to that of normal human serum elicit little or no hydrolysis. These findings indicate that hydrolysis of the phospholipids of intact E. coli S17 by serum occurs when: 1) the serum is bactericidal, and 2) when sufficient phospholipase A(2) is present. The difference in phospholipid hydrolysis that accompanies killing of E. coli by human or rabbit serum appears to reflect, therefore, the different amounts of phospholipase A(2) activity in the two sera. Phospholipid degradation is not required for the bactericidal action of serum. Bacterial phospholipid breakdown may be important, however, in the overall destruction and digestion of invading bacteria by the host.-Kaplan-Harris, L., J. Weiss, C. Mooney, S. Beckerdite-Quagliata, and P. Elsbach. The action of human and rabbit serum phospholipase A(2) on Escherichia coli phospholipids.     

Interference with valine and isoleucine biosynthesis by cyclic hydroxamic acids

Hogg, Robert W. (University of Illinois, Urbana), Chitra S. Biswas, and Harry P. Broquist. Interference with valine and isoleucine biosynthesis by cyclic hydroxamic acids. J. Bacteriol. 90:1265-1270. 1965.-The introduction of a hydroxyl group in a number of common barbiturates, a substitution which converts these compounds to cyclic hydroxamic acids, gives rise to compounds which inhibit growth of Escherichia coli. The toxicity of these hydroxybarbiturates appears to be associated, at least in part, with interference with valine and isoleucine biosynthesis, as a combination of these amino acids counteracts their toxicity. A subinhibitory level of 1-hydroxy-5-ethyl-5-isopropylbarbituric acid (hydroxyipral) was counteracted either by valine or by its early precursor, alpha-acetolactate, and led to a study of the effect of these cyclic hydroxamic acids on acetolactate synthesis in a cell-free enzyme system of E. coli. In this system, the parent barbiturates and their respective hydroxy derivatives were only moderately active in blocking acetolactate synthesis. Detailed kinetic studies of the most active compound, hydroxyipral showed no obvious relationship to the substrate or cofactors of the system. The inhibitory effects of hydroxyipral, either on growth of E. coli or in the acetolactate-forming system, could not be counteracted by Fe(++), but the toxic effect of aspergillic acid and o-phenanthroline in these instances was reversed by Fe(++), which implies an iron involvement in the acetolactate-forming system of E. coli.     

Glutamate synthase. Properties of the glutamine-dependent activity.

Properties of glutamine-dependent glutamate synthase have been investigated using homogeneous enzyme from Escherichia coli K-12. In contrast to results with enzyme from E. coli strain B (Miller, R. E., and Stadtman, E. R. (1972) J. Biol. Chem. 247, 7407-7419), this enzyme catalyzes NH3-dependent glutamate synthase activity. Selective inactivation of glutamine-dependent activity was obtained by treatment with the glutamine analog. L-2-amino-4-oxo-5-chloropentanoic acid (chloroketone). Inactivation by chloroketone exhibited saturation kinetics; glutamine reduced the rate of inactivation and exhibited competitive kinetics. Iodoacetamide, other alpha-halocarbonyl compounds, and sulfhydryl reagents gave similar selective inactivation of glutamine-dependent activity. Saturation kinetics were not obtained for inactivation by iodoacetamide but protection by glutamine exhibited competitive kinetics. The stoichiometry for alkylation by chloroketone and iodoacetamide was approximately 1 residue per protomer of molecular weight approximately 188,000. The single residue alkylated with iodo [1-14C]acetamide was identified as cysteine by isolation of S-carboxymethylcysteine. This active site cysteine is in the large subunit of molecular weight approximately 153,000. The active site cysteine was sensitive to oxidation by H2O2 generated by autooxidation of reduced flavin and resulted in selective inactivation of glutamine-dependent enzyme activity. Similar to other glutamine amidotransferases, glutamate synthase exhibits glutaminase activity. Glutaminase activity is dependent upon the functional integrity of the active site cysteine but is not wholly dependent upon the flavin and non-heme iron. Collectively, these results demonstrate that glutamate synthase is similar to other glutamine amidotransferases with respect to distinct sites for glutamine and NH3 utilization and in the obligatory function of an active site cysteine residue for glutamine utilization.     

Endonucleases for UV-irradiated and depurinated DNA in barley chloroplasts.

Lysates of barley chloroplasts release more radioactivity into acid soluble form from UV-irradiated and alkylated-depurinated E. coli [3H] DNA than from intact DNA. By means of affinity chromatography on depurinated DNA-cellulose and/or UV irradiated DNA-cellulose and by electrophoresis in polyacrylamide gels, four activities on depurinated DNA were separated. One of these contained activity against heavily UV-irradiated /270 J.m-2/ native DNA. In addition, two other nucleases specific towards UV-DNA were separated. One of them was active on native and heat denatured DNA irradiated with 10 J . m-2 UV, whereas the other was predominantly active on native UV-irradiated DNA.     

Inhibitory action of a non-metabolizable fatty acid on the growth of Escherichia coli: role of metabolism and outer membrane integrity.

The inhibitory action of decanoic acid on both Escherichia coli K-12/154 (normal lipopolysaccharide) and E. coli RC59 (defective lipopolysaccharide) was studied. A correlation was found between the doubling time of E. coli 154 growing in different media and the lethal effect of 0.4% decanoic acid on this bacterium. Decanoic acid (0.4%) exerted a lytic action on glucose-starved and NaN3-inhibited cells of E. coli 154 and RC59. Exponentially growing cultures of both strains were not affected by the addition of 0.4% methyldecanoate, but cells of E. coli RC59 reaching the stationary phase were attacked by that compound. A bactericidal action of 0.4% methyldecanoate on exponential E. coli 154 and RC59 was observed when sodium azide was also present in the media. Concentrations lower than 0.01% methyldecanoate had a lytic effect on spheroplasts from E. coli 154 and RC59. These results indicate that the inhibitory action of a non-metabolizable fatty acid on E. coli depends on the cellular metabolic activity and the outer membrane integrity.     

Effect of various nonionic surfactants on growth of Escherichia coli

Rose, Michael J., Jr. (Veterans Administration Hospital, Washington, D.C.), Stephen A. Aron, and Bernard W. Janicki. Effect of various nonionic surfactants on growth of Escherichia coli. J. Bacteriol. 91:1863-1868. 1966.-Escherichia coli cultivated in media containing 0.5, 1.0, 2.0, or 4.0% concentrations of surface-active polyoxyethylene derivatives of formaldehyde polymers of octyl phenol (Triton WR-1339; Macrocyclon) or of sorbitan mono-fatty acid esters (Tween 20, 40, 60, and 80) exhibited significantly retarded growth only at the highest concentration. To determine the mechanism of bacteriostasis, certain derivatives and compounds related to the surfactants were investigated. Experiments with compounds related to the Triton-type agents demonstrated that incorporation of monomeric substances (Triton X-205, X-305, Igepal CA-730, or Dowfax 9N20) into the medium at a concentration of 4.0% did not inhibit the growth of E. coli. It was concluded that the formaldehyde polymer was essential for growth inhibition by the polyoxyethylene derivatives of octyl phenol. The inhibitory activity of the Tween compounds, in contrast, appeared to result from the unesterified fatty acids which contaminate the commercial preparations. Polyol (60), the sorbitan polyoxyethylene derivative of Tween 60 and the basic structural unit of all the Tween-type compounds, and a Tween 80 preparation which was purified by extraction of the unesterified oleic acid, were not inhibitory. Moreover, the amount of free oleic acid present as a contaminant of Tween 80 was found to be sufficient to cause significant growth inhibition. These results and the observation that E. coli does not appear to hydrolyze the esterified fatty acid of Tween 80 led to the conclusion that growth inhibition obtained with various Tween compounds probaby is a function of their respective fatty acid contaminants.