sodium-hypochlorite and Starvation

To examine the effect that starvation and sodium hypochlorite stress have on virulence of Escherichia coli O157:H7 and the influence of conditioned media on recovery of stressed cells.. Escherichia coli O157:H7 was starved for 5 days then exposed to 1 microg ml(-1) sodium hypochlorite, suspended in defined media Dulbecco's Modified Eagle's Medium supplemented with conditioned media, sampled over a 12-h period; and assayed for growth, production of Shiga toxin (Stx), and attachment to HCT-8 cells. During recovery, stressed and control cells grown in conditioned media exhibited greater attachment efficiencies to HCT-8 cells then cells in DMEM alone. Production of Stx by treated cells mimicked Stx production by control cells suggesting that components of conditioned media assist in recovery. Results showed that levels of autoinducer-2 fluctuate during recovery and growth suggesting involvement of a quorum sensing mechanism during the recovery of stressed E. coli O157:H7.. The recovery of stressed E. coli O157:H7 exposed to starvation conditions and HOCl is positively affected by the presence of autoinducer-2 thereby influencing virulence factor production.. Food-borne pathogens in a stressed state prior to ingestion can rapidly recover in the presence of bacterial by-products; exhibiting virulence characteristics and presenting a microbial food safety hazard.

Topics: Bacteriological Techniques; Culture Media, Conditioned; Disinfectants; Enterobacteriaceae; Escherichia coli Infections; Escherichia coli O157; Food Microbiology; Homoserine; Lactones; Microbial Viability; Quorum Sensing; Shiga Toxin; Sodium Hypochlorite; Starvation; Virulence

To investigate the effect of starvation, surface attachment and growth in a biofilm on the susceptibility of Aureobasidium pullulans to the biocides 2-n-octyl-4-isothiazolin-3-one (OIT) and sodium hypochlorite (NaOCl).. Fluorescence loss from a green fluorescent protein (GFP)-transformed strain was used to monitor real-time loss in viability as previously described in situ in 96-well plates. Exponential phase, yeast-like (YL) cells were settled in the bottom of the wells as a low-density monolayer (LDM) and were susceptible to all biocide concentrations (25-100 mug ml(-1)). The exponential phase YL cells were either starved for 48 h in suspension or starved for 48 h as LDMs in the wells. Starvation in both cases led to a small reduction in susceptibility to the biocides. In contrast, 48-h biofilms grown in malt extract broth showed an apparent lack of susceptibility to 25 and 50 mug ml(-1) OIT and to 25-100 mug ml(-1) NaOCl. However, when the OIT concentration was increased to compensate for the higher cell density in the biofilm, the biofilms were found to be equally susceptible to the LDM.. Starvation of A. pullulans YL cells either in suspension or as attached LDM resulted in a decrease in susceptibility to low concentrations of both OIT and NaOCl while the apparent reduced susceptibility of mature biofilms was due to the increase in biofilm cell density rather than true biofilm resistance per se.. Monitoring fluorescence loss from the GFP-transformed strain of A. pullulans can be used as a fast and reliable method for monitoring cell death in real time as a response to biocide and antimicrobial challenge.

Topics: Biodegradation, Environmental; Biofilms; Colony-Forming Units Assay; Disinfectants; Green Fluorescent Proteins; Microbiological Techniques; Microscopy, Fluorescence; Sodium Hypochlorite; Starvation; Thiazoles; Yeasts

The concentration of cytoplasmic free pyrophosphate was calculated in freeze-clamped livers of rats from the measured concentration of reactants and K(eq.) of the UDP-glucose pyrophosphorylase reaction (UDP-alpha-d-glucose 1-phosphate uridylyltransferase, EC 2.7.7.9). The K(eq.) of the UDP-glucose pyrophosphorylase reaction was redetermined at 38 degrees C, pH7.0, I=0.25mol/l and free [Mg(2+)]=1mm, and was 4.55 in the direction of glucose 1-phosphate formation. The activity of UDP-glucose pyrophosphorylase in rat liver was between 46 and 58mumol of glucose 1-phosphate formed/min per g fresh wt. in the four dietary conditions studied. A fluorimetric assay with enzymic cycling was developed for the measurement of glucose 1-phosphate in HClO(4) extracts of rat liver. The calculated free cytoplasmic PP(i) concentration in nmol/g fresh wt. of liver was 2.3+/-0.3 in starved, 3.8+/-0.4 in fed, 4.9+/-0.6 in meal-fed and 5.2+/-0.4 in sucrose-re-fed animals. These values agree well with recently determined direct measurements of total PP(i) in rat liver and suggest that there is not a large amount of bound or metabolically inert PP(i) in rat liver. The cytoplasmic [ATP]/[AMP][PP(i)] ratio is 10(3) times the cytoplasmic [ATP]/[ADP][P(i)] ratio and varies differently with dietary state. The reaction PP(i)+H(2)O-->2P(i) catalysed by inorganic pyrophosphatase (EC 3.6.1.1) does not attain near-equilibrium in vivo. PP(i) should be considered as one of the group of small inorganic ions which is metabolically active and capable of exerting a controlling function in a number of important metabolic reactions.

Topics: Adenosine Diphosphate; Adenosine Monophosphate; Adenosine Triphosphate; Animals; Cytoplasm; Diet; Diphosphates; Fluorometry; Glucosephosphates; Kinetics; Liver; Male; Nucleotidyltransferases; Phosphates; Pyrophosphatases; Rats; Sodium Hypochlorite; Spectrophotometry; Starvation; Uracil Nucleotides