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At least 739 records · Page 41Linked to original sources

PubMed2000

Use of hydrostatic pressure for inactivation of microbial contaminants in cheese.

The objective of this study was to determine the effect of high pressure (HP) on the inactivation of microbial contaminants in Cheddar cheese (Escherichia coli K-12, Staphylococcus aureus ATCC 6538, and Penicillium roqueforti IMI 297987). Initially, cheese slurries inoculated with E. coli, S. aureus, and P. roqueforti were used as a convenient means to define the effects of a range of pressures and temperatures on the viability of these microorganisms. Cheese slurries were subjected to pressures of 50 to 800 MPa for 20 min at temperatures of 10, 20, and 30 degrees C. At 400 MPa, the viability of P. roqueforti in cheese slurry decreased by >2-log-unit cycles at 10 degrees C and by 6-log-unit cycles at temperatures of 20 and 30 degrees C. S. aureus and E. coli were not detected after HP treatments in cheese slurry of >600 MPa at 20 degrees C and >400 MPa at 30 degrees C, respectively. In addition to cell death, the presence of sublethally injured cells in HP-treated slurries was demonstrated by differential plating using nonselective agar incorporating salt or glucose. Kinetic experiments of HP inactivation demonstrated that increasing the pressure from 300 to 400 MPa resulted in a higher degree of inactivation than increasing the pressurization time from 0 to 60 min, indicating a greater antimicrobial impact of pressure. Selected conditions were subsequently tested on Cheddar cheese by adding the isolates to cheese milk and pressure treating the resultant cheeses at 100 to 500 MPa for 20 min at 20 degrees C. The relative sensitivities of the isolates to HP in Cheddar cheese were similar to those observed in the cheese slurry, i.e., P. roqueforti was more sensitive than E. coli, which was more sensitive than S. aureus. The organisms were more sensitive to pressure in cheese than slurry, especially with E. coli. On comparison of the sensitivities of the microorganisms in a pH 5.3 phosphate buffer, cheese slurry, and Cheddar cheese, greatest sensitivity to HP was shown in the pH 5.3 phosphate buffer by S. aureus and P. roqueforti while greatest sensitivity to HP by E. coli was exhibited in Cheddar cheese. Therefore, the medium in which the microorganisms are treated is an important determinant of the level of inactivation observed.

Cheese↗

PubMed1988

In vitro methods for determination of viability of mycobacteria: comparison of ATP content, morphological index and FDA-EB fluorescent staining in Mycobacterium leprae.

Explore the source record for details and available documents.

Adenosine Triphosphate↗

PubMed2004

The effect of systemic antibiotics on the microbiological diagnosis of experimental foreign body infections caused by Staphylococcus epidermidis.

Qualitative broth culture and quantitative culture on agar were compared with quantitative polymerase chain reaction (PCR) for the diagnosis of foreign body infections (FBI) in a rat model with and without exposure to systemic antibiotics (teicoplanin and rifampin). The 3 methods had a similar and high yield without antibiotics. Antibiotics decreased the number of CFU/foreign body and increased the number of culture-negative foreign bodies and the variability of the results in quantitative culture. The yield of broth culture remained high under antibiotics although prolonged incubation (2-5 days) was required. The yield of the PCR was equivalent or even superior (for teicoplanin) to the yield of broth culture. Quantitative PCR had a higher yield and lower variability than quantitative culture and was not affected by antibiotics. The simultaneous isolation of RNA from all samples indicated viability of the bacteria. Quantitative PCR seems a promising method for the diagnosis of FBI.

Animals↗

PubMed1999

Reverse transcription polymerase chain reaction method for the detection of glycopeptide resistance in enterococci.

In this work we have developed reverse transcription polymerase chain reaction (RT-PCR) methods for detecting specific mRNA from enterococci, particularly vanA and vanB genes, responsible for glycopeptide resistance in this genus. mRNA from the two genes was detected immediately after RNA extraction of a midlog phase culture, determined by growth rate analysis. Because of the short half-life associated with many bacterial RNA species (1.5-2 min), time is an important factor in obtaining RNA of good yield and high purity. Our results showed that: (i) the transcription of mRNA related to vanA ligase in enterococci showing Van A phenotype happens only after induction with both vancomycin and teicoplanin; (ii) the transcription of mRNA related to vanB ligase happens only in the presence of vancomycin and (iii) there was no transcription of mRNA in the two strains positive to vanA gene after PCR experiments. RT-PCR methodology can have numerous applications in microbiology for studying gene expression in isolated bacteria and also in nonculturable cells in environmental samples, for studies of mechanisms and/or as an indicator of viability in bacterial communities.

Anti-Bacterial Agents↗

PubMed1999

Effectiveness of cleaning techniques used in the food industry in terms of the removal of bacterial biofilms.

The effectiveness of cleaning was investigated through food factory trials and laboratory experiments using a naturally occurring biofilm from a food factory environment and generated biofilms. The efficacy of factory cleaning and disinfection programmes was assessed by swabbing and total viable count (TVC) analysis of surfaces before cleaning, after cleaning and after disinfection. Cleaning produced a 0.91 log reduction in the attached population. Investigation of the effectiveness of a variety of cleaning methods in the removal of a naturally occurring food factory biofilm showed that the high pressure spray and the mechanical floor scrubber, which use a high degree of mechanical action, were most effective. Cleaning trials with biofilms of Pseudomonas aeruginosa or Staphylococcus aureus showed that spraying with water at pressures of 34.5, 51.7 and 68.9 bar did not significantly increase the removal, as assessed by direct epifluorescent microscopy (DEM) and swabbing and TVC analysis, beyond the three log reduction observed at 17.2 bar. The effect of spray time at 17.2 bar showed that increasing spray time from 1 to 10 s did not significantly increase removal of Ps. aeruginosa biofilm. Investigation of the optimum distance of the spray lance from the surface at 17.2 bar was found to be between 125 and 250 mm. The use of an alkaline, acidic or neutral detergent prior to spraying with water at 17.2 bar did not significantly increase the removal of Ps. aeruginosa or Staph. aureus. However, the acidic and alkaline products significantly (P = 0.05) affected the viability of Staph. aureus and Ps. aeruginosa, respectively, thereby minimizing the potential for the spread of contamination.

Bacterial Adhesion↗

PubMed2000

Acid resistance of Helicobacter pylori depends on the UreI membrane protein and an inner membrane proton barrier.

ureI encodes an inner membrane protein of Helicobacter pylori. The role of the bacterial inner membrane and UreI in acid protection and regulation of cytoplasmic urease activity in the gastric microorganism was studied. The irreversible inhibition of urease when the organism was exposed to a protonophore (3,3',4', 5-tetrachlorsalicylanide; TCS) at acidic pH showed that the inner membrane protected urease from acid. Isogenic ureI knockout mutants of several H. pylori strains were constructed by replacing the ureI gene of the urease gene cluster with a promoterless kanamycin resistance marker gene (kanR). Mutants carrying the modified ureAB-kanR-EFGH operon all showed wild-type levels of urease activity at neutral pH in vitro. The mutants resisted media of pH > 4.0 but not of pH < 4.0. Whereas wild-type bacteria showed high levels of urease activity below pH 4.0, this ability was not retained in the ureI mutants, resulting in inhibition of metabolism and cell death. Gene complementation experiments with plasmid-derived H. pylori ureI restored wild-type properties. The activation of urease activity found in structurally intact but permeabilized bacteria treated with 0.01% detergent (polyoxy-ethylene-8-laurylether; C12E8), suggested a membrane-limited access of urea to internal urease at neutral pH. Measurement of 14C-urea uptake into Xenopus oocytes injected with ureI cRNA showed acid activation of uptake only in injected oocytes. Acceleration of urea uptake by UreI therefore mediates the increase of intracellular urease activity seen under acidic conditions. This increase of urea permeability is essential for H. pylori survival in environments below pH 4.0. ureI-independent urease activity may be sufficient for maintenance of bacterial viability above pH 4.0.

Acids↗

PubMed2004

In vitro antimicrobial effect of bacteriophages on human dentin infected with Enterococcus faecalis ATCC 29212.

This study assessed the effect of bacteriophages on the viability of Enterococcus faecalis. Human dental roots were inoculated with a suspension of E. faecalis at three different multiplicities of infection - 0.1, 1.0 and 10.0. The phage lysate was able to significantly inhibit bacteria growth when incubated at the multiplicities of infection of 1.0, 10.0 and 0.1. The dental roots were also inoculated with bacteria for 6 days to allow bacterial penetration into the teeth tubules. Addition of the phage lysate to the roots following the 6-day incubation period led to a substantial reduction in bacteria viability. Phage therapy may be an important alternative for the treatment of root canal infections refractory to conventional endodontic therapy.

Bacteriophages↗

PubMed1982

Effect of enzymatic disaggregation on proliferation of human tumor cells in soft agar.

Cell suspensions were prepared by either mechanical or enzymatic disaggregation methods from biopsy specimens from 54 patients with various tumors. The biologic activities of cells derived from the two suspensions were then examined. Biopsy specimens of solid tumors were minced, and one-half of each specimen was further processed by being teased with needles, whereas the other half was exposed to a combination of collagenase, hyaluronidase, and DNase. The enzymatic disaggregation method yielded fewer cells per gram of tissue than the mechanical method. However, the percentage of dye-excluding cells was increased by the enzymatic procedure in 93% of the cases. Cells obtained by enzymatic means also had higher cloning efficiencies than those obtained by mincing. The histologic types of cells present in the initial cell suspensions were the same for cells obtained by the enzymatic or mechanical disaggregation methods. The number of colonies obtained was linearly related to the number of cells plated in both cases. The tritiated thymidine suicide indices (estimates of the percentage of cells in the S-phase of the cell cycle) were the same for the two cell populations obtained by the two methods. The results indicate that cells obtained from solid tumors by enzymatic dissociation methods did not differ significantly from cells obtained by the more conventional mechanical techniques. However, cell viabilities and cloning efficiencies were significantly improved by the enzymatic technique.

Agar↗

PubMed1998

Systematic identification of essential genes by in vitro mariner mutagenesis.

Although the complete DNA sequences of several microbial genomes are now available, nearly 40% of the putative genes lack identifiable functions. Comprehensive screens and selections for identifying functional classes of genes are needed to convert sequence data into meaningful biological information. One particularly significant group of bacterial genes consists of those that are essential for growth or viability. Here, we describe a simple system for performing transposon mutagenesis on naturally transformable organisms along with a technique to rapidly identify essential or conditionally essential DNA segments. We show the general utility of this approach by applying it to two human pathogens, Haemophilus influenzae and Streptococcus pneumoniae, in which we detected known essential genes and assigned essentiality to several ORFs of unknown function.

Base Sequence↗

PubMed1977

The activity of the pneumococcal autolytic system and the fate of the bacterium during ingestion by rabbit polymorphonuclear leukocytes.

The extent to which autolytic microbial enzymes are involved in the fate of microorganisms ingested by phagocytes has not been determined. It is known, however, that activation of degradative enzymes occurs during certain microbicidal events. We examined the possible role of the pneumococcal autolytic enzyme (an N-acetylmuramyl-L-alanine amidase) in the loss of viability and degradation of pneumococci during phagocytosis by rabbit polymorphonuclear leukocytes. Three bacterial systems were compared: (a) wild type pneumococci with an active autolytic system; (b) wild type bacteria grown under conditions that block the endogenous autolytic activity and (c) a mutant strain defective in the major autolytic enzyme of this bacterium. No differences could be detected between the autolysis-positive and negative bacteria in the rate of killing and in the fate of macromolecular cell constituents during ingestion by rabbit peritoneal polymorphonuclear leukocytes.

Amidohydrolases↗

PubMed2004

A fibrous-bed bioreactor for continuous production of monoclonal antibody by hybridoma.

A fibrous-bed bioreactor (FBB) has been developed to culture hybridoma cells for long-term continuous production of monoclonal antibody (MAb). A non-woven polyester fibrous matrix was used to immobilize the cells to reach a high viable cell density of 3 x 10(8) cells cm(-3) packed bed, which gave a high volumetric MAb productivity of 1 g L(-1) day(-1) under continuous feed conditions with the medium containing 10% serum. Reducing the medium serum content to 1% increased MAb production to 6.5 g L(-1) day(-1) in a repeated batch FBB culture. MAb production was higher at higher dissolved oxygen (DO) levels in the range between 10% and 70% of air saturation, although DO did not significantly affect glucose metabolism and lactate production. The medium LDH (lactate dehydrogenase) level increased dramatically when the DO level was decreased from 30% to 10%, suggesting that a critical DO level of approximately 30% is necessary for maintaining the FBB culture for long-term operation. Compared with suspension cultures in T-flasks and spinner flasks, the FBB culture had a lower lactate yield from glucose (0.80 vs. 0.91 g g(-1), produced MAb at a higher concentration (up to 442 mg L(-1) vs. 83.5 mg L(-1), and was stable for continuous long-term operation (more than 1 month). The superior FBB performance was attributed to the highly porous fibrous matrix that enabled the efficient mass transfer, cell immobilization, and continued growth and regeneration that are critical to maintaining a high density of viable and productive cell populations. The cells immobilized in the fibrous matrix had high viability (>85%) even though many of them were in growth arrest (G1/G0 phase) as indicated by their smaller cell size (<10 microm). Scanning electron microscopic studies of the cell-matrix showed that the high density of cells formed large clumps within the interstitial spaces of the fibrous matrix. Their close contact and interaction with each other might have contributed to their ability to survive well under adverse conditions such as low DO and low serum content in the medium. It was also found that the cells present inside the fibrous matrix had a higher viability and lower apoptosis than those present in the liquid suspension, indicating that the fibrous matrix had selectively retained healthy, nonapoptotic cells and dislodged apoptotic and dead cells; this also might have contributed to the stability of the long-term culture. This work demonstrated that the FBB originally developed for microbial fermentation also gave excellent results in achieving high cell density, productivity, and product concentrations, and should have a good potential for industrial animal cell culture applications.

Antibodies, Monoclonal↗

PubMed2005

Dissemination and survival of non-indigenous bacterial genomes in pristine Antarctic environments.

Continental Antarctic is perceived as a largely pristine environment, although certain localized regions (e.g., parts of the Ross Dependency Dry Valleys) are relatively heavy impacted by human activities. The procedures imposed on Antarctic field parties for the handling and disposal of both solid and liquid wastes are designed to minimise eutrofication and contamination (particularly by human enteric bacteria). However, little consideration has been given to the significance, if any, of less obvious forms of microbial contamination resulting from periodic human activities in Antarctica. The predominant commensal microorganism on human skin, Staphylococcus epidermidis, could be detected by PCR, in Dry Valley mineral soils collected from heavily impacted areas, but could not be detected in Dry Valley mineral soils collected from low impact and pristine areas. Cell viability of this non-enteric human commensal is rapidly lost in Dry Valley mineral soil. However, S. epidermidis can persist for long periods in Dry Valley mineral soil as non-viable cells and/or naked DNA.

Antarctic Regions↗

PubMed2004

The interplay between classical and alternative isoprenoid biosynthesis controls gammadelta T cell bioactivity of Listeria monocytogenes.

Isoprenoids are synthesised either through the classical, mevalonate pathway, or the alternative, non-mevalonate, 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway. The latter is found in many microbial pathogens and proceeds via (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP), a potent activator of human Vgamma9/Vdelta2 T cells. Listeria monocytogenes is the only pathogenic bacterium known to contain both pathways concurrently. Strategic gene knockouts demonstrate that either pathway is functional but dispensable for viability. Yet, disrupting the mevalonate pathway results in a complementary upregulation of the MEP pathway. Vgamma9/Vdelta2 T cell bioactivity is increased in DeltalytB mutants where HMB-PP accumulation is expected, and lost in DeltagcpE mutants which fail to produce HMB-PP.

Diphosphates↗

PubMed1989

Comparison of properties of collected cells and cells from the culture vessel during continuous culture of Streptococcus mutans Ingbritt.

In continuous-culture studies chemostat effluents are usually collected into a receiving flask in an ice bath to obtain enough cells for an experiment. It is assumed that the properties of these are not significantly different from those of the culture in the chemostat vessel. This assumption has been tested for the dental pathogen Streptococcus mutans Ingbritt. Collected supernatant fluid and cells were compared with supernatant fluid and cells taken directly from the culture vessel, for four major groups of culture properties: viability and biomass, concentrations of metabolites and nutrients, activities of selected enzymes, and glycolytic rates. The assumption held true except for glycolytic rate during endogenous metabolism. It is suggested that comparison of collected and culture vessel cells is an important control which should be done in all continuous culture studies of microbial physiology and biochemistry, but that the properties of Strep. mutans cells collected on ice up to 16 h do reflect those of cells actively growing in the chemostat.

Bacteriological Techniques↗

PubMed1968

Viability in fertile stocks of Pseudomonas aeruginosa.

Diluent composition, time consumed in experimental manipulations, and the presence of pyocine-like lethal agents affected the viability of sexual fertility factor-positive and fertility factor-negative auxotrophic stocks of Pseudomonas aeruginosa. In the same cultures, recovery of prototrophic revertants increased as the number of viable cells dispensed per plate was reduced. As a result, the number of revertants recovered was indirectly determined by the combined activities of the three conditions affecting viability. Possible modifications by these conditions affecting viability on the expression and interpretation of the fertility factor-positive sex factor-mediated system of genetic recombination are presented.

Bacteriocins↗

PubMed1969

Accumulation of 14C-streptomycin by streptomycin-sensitive and streptomycin-resistant group H streptococci.

Three streptomycin-resistant mutants of group H streptococcus, strain Challis, were examined for ability to accumulate (14)C-streptomycin. Although the mutants exhibited different levels of transformation, only the streptomycin-sensitive parent Challis strain accumulated significant amounts of (14)C-streptomycin. It appears that impermeability to streptomycin does not necessarily result in reduction or loss of transformability. The amount of label accumulated by strain Challis was correlated with a loss of viability. In addition, accumulation of label was influenced by the concentration of (14)C-streptomycin, the time of exposure, and the type of medium employed.

Carbon Isotopes↗

PubMed1994

Adherence of Legionella pneumophila to U-937 cells, guinea-pig alveolar macrophages, and MRC-5 cells by a novel, complement-independent binding mechanism.

In the absence of serum, Legionella pneumophila demonstrated wash-resistant adherence to U-937 cells, primary guinea-pig alveolar macrophages, and MRC-5 cells. Neither complement nor antibody was required for binding. The dynamics of adherence following inoculation of L. pneumophila at increasing 10-fold multiplicities of infection to each of the three host cell types resulted in a first-order kinetic relationship of binding, indicative of one bacterial adhesin molecule recognized by one host cell receptor moiety. Host cell receptor saturation studies showed that depending on the cell type, 2-8% of the bacterial inoculum adhered to cells under these nonopsonic conditions. Preliminary adhesin and receptor characterization studies were performed to define the chemical composition of the binding structures on both the organism and the three different host cell surfaces. The adherence phenomenon was investigated using competitive binding assays in the presence of putative adhesin analogs as well as following treatments modifying the microbial and host cell surface membranes. Attachment was evaluated both by viable bacterial cell colony counts and by indirect immunofluorescent assay. With the exception of aldehyde treatments, the various membrane-modifying regimes and the presence of the adhesin analogs were shown to have no effect on organism or host cell viability. Data suggested that the L. pneumophila adhesin responsible for opsonin-independent binding to these host cells was a protein structure with lectin-like properties. Furthermore, this protein would appear to be intimately associated with carbohydrate or lipid structures located on the bacterial outer membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

Adhesins, Bacterial↗

PubMed2001

Direct microscopic observation of lettuce leaf decontamination with a prototype fruit and vegetable washing solution and 1% NaCl-NaHCO3.

Efficacy of a prototype, food-grade alkaline surfactant washing solution and 1% NaCl-NaHCO3 (pH 10.0) against Escherichia coli O157:H7 cells on lettuce leaves was evaluated. Lettuce was inoculated with 10(9) CFU/ml of E. coli O157:H7 for 24 +/- 1 h at 4 degrees C. Samples were rinsed and treated with the prototype washing solution containing lauryl sodium sulfate or NaCl-NaHCO3 for 3 min at 22 degrees C. Viability of E. coli O157:H7 cells was examined by plate counts at the surface and cut edge, and by confocal scanning microscopic (CSLM) observation of samples stained with Sytox green and Alexa 594 conjugated antibody against E. coli O157:H7 at intact leaf surface, stomata, and damaged tissue (0 to 10, 30 to 40, and 0 to 40 microm from the cut surface). Although both treatments caused significant log reductions in CFU at the surface and cut edge, log reductions were greater for the prototype washing solution (0.7 to 1.1 log CFU/cm2) than for NaCl-NaHCO3 (0.2 to 0.4 log CFU/cm2) (P < 0.05). Percentage of viability determined by CSLM for prototype washing solution was significantly greater at 30 to 40 microm from cut surfaces than at 0 to 10 and 0 to 40 microm from cut surfaces and intact surfaces (P < 0.05). Stomata provided moderate protection. NaCl-NaHCO3 was less effective than the prototype washing solution, and high percentages of E. coli O157:H7 cells remained viable at all sites except at the surface. The percent viabilities determined by CSLM were not significantly different from those determined by plate counts for NaCl-NaHCO3 treatment (P > 0.05). However, CSLM indicated significantly greater percent viability than plate counts for lettuce treated with the prototype washing solution (P < 0.05). Surfactant-containing washing solutions warrant additional testing for decontamination of fresh produce.

Colony Count, Microbial↗