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B J Wilkinson

Publications and source records attributed to B J Wilkinson.

At least 55 records · Page 3Linked to original sources
PubMed1987

Hydrophobicity-hydrophilicity of staphylococci.

The hydrophobicity-hydrophilicity of various strains of Staphylococcus has been studied by a technique involving partitioning of the cells between aqueous and hydrocarbon phases. S. aureus was typically hydrophobic, and to a greater degree in stationary- than in exponential-phase cultures. Mutants that lacked teichoic acid, protein A or coagulase production were hydrophobic, indicating that none of these factors was responsible for hydrophobicity. The presence of a capsule rendered strains hydrophilic. Thus, determination of hydrophobicity may be a useful additional test for capsulation. However, a non-capsulate S. aureus strain was hydrophilic. Trypsin treatment converted strains from hydrophobic to hydrophilic. Isolated bacterial cell wall preparation, either crude or purified, and peptidoglycan were hydrophilic. These results indicate that the determinant of hydrophobicity is a protein or protein-associated molecule localised at the cell surface of the organism, i.e., a component of either the cell wall, cell membrane, or both. Twenty-five strains of twelve coagulase-negative species were examined and most (18) were hydrophobic, again indicating that protein A is not a major determinant of hydrophobicity in these staphylococci. Four of seven hydrophilic strains were capsulate; three strains of S. sciuri were hydrophilic but non-capsulate.

Bacterial Proteins↗
PubMed1987

Effects of temperature, NaCl, and methicillin on penicillin-binding proteins, growth, peptidoglycan synthesis, and autolysis in methicillin-resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus strains produce a fifth penicillin-binding protein (PBP), PBP 2', with low affinity for beta-lactam antibiotics that is believed to represent a beta-lactam-insensitive peptidoglycan transpeptidase. In an effort to evaluate the adequacy of PBP 2' as an explanation of methicillin resistance, PBP 2' production and the responses of growth and peptidoglycan synthesis to methicillin under different environmental conditions have been compared. In the heterogeneous methicillin-resistant strain DU4916-K7, less PBP 2' was produced at 40 degrees C than at 30 degrees C, but inclusion of 5% (wt/vol) NaCl in the medium at 40 degrees C boosted PBP 2' production and allowed growth of the organism in the presence of 10 micrograms of methicillin per ml. When exponential-phase cultures were challenged with methicillin, growth and peptidoglycan synthesis were much more resistant at 30 degrees C than at 40 degrees C. Inclusion of NaCl in medium rendered growth and peptidoglycan synthesis more methicillin resistant at 40 degrees C. Hence, there was a good correlation between PBP 2' production and methicillin-resistant peptidoglycan synthesis under these conditions. However, PBP 2' production was increased by NaCl at 30 degrees C without markedly affecting the susceptibilities of growth and peptidoglycan synthesis to methicillin. Pregrowth of cells with methicillin, which was expected to boost PBP 2' production, seemed to increase the susceptibilities of growth and peptidoglycan synthesis to methicillin. Patterns of growth and peptidoglycan synthesis susceptibilities to methicillin which were similar to those described above were found in chloramphenicol-inhibited cultures, in which presumably no induction of PBP 2' could occur during the methicillin challenge period. Complex effects were noted in the combination of subinhibitory methicillin and NaCl. Growth of cells in the presence of NaCl stimulated their autolytic activity, which was further increased by growth with subinhibitory methicillin in addition to NaCl. It appears that NaCl enhances methicillin resistance by stimulating PBP 2' production and providing osmotic support but opposes it by stimulating autolytic activity which is exacerbated by the very low cross-linking of peptidoglycan in methicillin-resistant strains grown in the presence of methicillin.

Bacterial Proteins↗
PubMed1986

Effects of growth of methicillin-resistant and -susceptible Staphylococcus aureus in the presence of beta-lactams on peptidoglycan structure and susceptibility to lytic enzymes.

Growth of methicillin-resistant Staphylococcus aureus DU4916 in the presence of methicillin yielded crude cell walls that showed an increased rate of autolysis and purified cell walls (PCW) and peptidoglycan (PG) that had increased susceptibilities to autolysin extracted with LiCl and to lysozyme. The PG of cells grown in the presence of methicillin had markedly decreased cross-linking and O acetylation. Growth of the methicillin-susceptible strain H in the presence of subinhibitory concentrations of cefoxitin, a specific inhibitor of penicillin-binding protein (PBP) 4, caused a substantial decrease in PG cross-linking and O acetylation and increased susceptibilities of PCW and PG to LiCl-extracted autolysin and to lysozyme. Strain DU4916 cells grown in the presence of methicillin did not show an increased rate of autolysis or an increased susceptibility to vancomycin- or D-cycloserine-induced lysis, even though their PG was hypo-cross-linked. This implies that the potential for increased autolysis is controlled in intact cells and that this regulation may be involved in the methicillin resistance phenomenon. Growth of the methicillin-susceptible strain DU4916S in the presence of methicillin yielded PCW and PG that showed small increases in susceptibilities to LiCl-extracted autolysin and to lysozyme and a small decrease in PG cross-linking. Comparison of the PBPs of a penicillinase-nonproducing derivative of strain DU4916 (DU4916-K7) with those of strain DU4916S in intact cells and isolated membranes revealed that PBPs 1 to 4 had similar high beta-lactam antibiotic affinities in both strains and identified an additional PBP, PBP2(1), with low beta-lactam affinity in the methicillin-resistant strain DU4916-K7. The low degree of cross-linking of PG in strain DU4916 cells grown with methicillin was probably due mainly to inhibition of the secondary cross-linking function of PBP 4.

Acetylation↗
PubMed1986

Iron-dependent production of a heat-modifiable, 23,000-Mr outer membrane protein in Paracoccus denitrificans.

Production of a 23,000-Mr major outer membrane protein of Paracoccus denitrificans ATCC 13543 was dependent upon the addition of iron to a succinate-salts medium. The 23,000-Mr protein was not produced in an iron-deficient medium, but production of five outer membrane proteins in the 85,000- to 72,000-Mr range and of catechol were induced. The 23,000-Mr protein was not produced in a complex medium even when ferric citrate was added to the medium. Production of the protein was influenced by the carbon source and was decreased by peptone.

Bacterial Outer Membrane Proteins↗
PubMed1986

Slime production by bovine milk Staphylococcus aureus and identification of coagulase-negative staphylococcal isolates.

Staphylococcus aureus isolates from bovine milk were assessed for capsule or slime production. When pure S. aureus cultures in milk were inoculated directly into serum-soft agar constituted with a modified staphylococcus 110 medium, 100% of the isolates grew with diffuse colony morphology. Diffuse colony morphology was rapidly lost on subculture and was more rapidly lost in brain heart infusion-serum-soft agar. No evidence was seen for encapsulation in India ink preparations or by the clumping factor test. It was concluded that freshly isolated S. aureus strains produce slime, not true capsules. During examination of the 84 milk samples that grew staphylococci in addition to S. aureus (27.4%), a significant number of coagulase-negative staphylococcal species were encountered and identified by conventional tests as S. simulans (41.7%), S. xylosus (11.9%), S. epidermidis (3.6%), S. saprophyticus (3.6%), S. hyicus (2.9%), S. cohnii (1.2%), S. haemolyticus (1.2%), and S. warneri (1.2%). Five isolates (6.0%) were not identified. Attempts were also made to identify the isolates by the API Staph-Ident system, which gave an overall accuracy of 45.2%. The susceptibilities of the isolates to a variety of antibiotics were determined, and they appeared to be less resistant than human clinical isolates.

Animals↗
PubMed1984

Sodium-dependent uptake of taurine in encapsulated Staphylococcus aureus strain M.

A novel uptake system for the unusual sulfonated amino acid taurine was discovered in the prokaryote, encapsulated Staphylococcus aureus strain M. This strain has been shown previously to contain taurine in its capsular polysaccharide. Taurine uptake by whole cells incubated in buffer showed a saturable dependency upon Na+ and taurine uptake was itself a saturable process, stimulated by glucose, and markedly affected by temperature. No evidence was found for the inducibility of taurine uptake. In the presence of 10 mM NaCl Lineweaver-Burk plots revealed a Km of 42 microM and Vmax of 4.6 nmol/min per mg dry weight for taurine uptake at 37 degrees C. Increasing concentrations of Na+ decreased the Km of the system and appeared to increase the Vmax. Of various other cations tested only Li+ supported marked taurine uptake. Excess unlabelled taurine did not cause efflux of radioactivity taken up. Taurine was taken up into cold trichloroacetic acid-soluble material and did not chromatograph as taurine, indicating rapid metabolism during or closely following uptake. Taurine uptake appeared to occur via a highly specific system because amino acids representing the major known groups of amino acid transport systems in S. aureus did not inhibit taurine uptake, and uptake was only slightly diminished by the structurally closely related compounds hypotaurine and 3-amino-1-propane sulfonic acid. Sulfhydryl group reagents, electron transport inhibitors, an uncoupler and inhibitors of Na+-linked transport processes inhibited taurine uptake. A variety of other metabolic inhibitors had little effect on taurine uptake.

Amino Acids↗
PubMed1983

Antibody response to peptidoglycan during staphylococcal infections.

Levels of antibodies to peptidoglycan were measured by solid-phase radioimmunoassay in 76 patients with Staphylococcus aureus infections and 44 patients with infections caused by other bacteria. Levels of IgM antibodies to peptidoglycan were elevated in 24% of the patients with endocarditis or complicated bacteremia caused by S. aureus, 21% of those with serious infections caused by other gram-positive cocci, none of those with less serious infections caused by S. aureus or gram-negative bacilli, and 5% of the normal controls. Levels of IgG antibodies to peptidoglycan were elevated in 50% of the patients with endocarditis or complicated bacteremia caused by S. aureus, 3% of those with less severe infections caused by S. aureus, 17% of those with infections caused by other gram-positive cocci, 20% of those infections caused by gram-negative bacilli, and 5% of the normal controls.

Antibodies, Bacterial↗
PubMed1983

Serology of Staphylococcus aureus infections using multiple antigens and serial serum samples.

The detection of antibody to S aureus in human serum can aid in the management of staphylococcal diseases [1]. RIAs and ELISAs can detect low levels of antibody and demonstrate increased antibody production during serious staphylococcal infections [2,3]. We compared four S aureus constituents--cell-wall peptidoglycan and teichoic acid, and extracellular alpha-toxin and nuclease--as antigens in a sensitive ELISA. The value of testing more than a single serum sample was also determined. Elevated IgG antibody to peptidoglycan, present in one or more serum samples of 13 (50%) of 26 patients with complicated bacteremia, was found to be the most sensitive test. All 26 patients had a significant IgG antibody response to peptidoglycan. Three (27%) of 11 patients with uncomplicated bacteremia had elevated levels of antibody to peptidoglycan in their serum, and seven (64%) showed a significant change in titer when serial serum samples were tested. Maximum detection rates for the other antigens in complicated and uncomplicated bacteremia were, respectively, 62% and 37% for teichoic acid, 38% and 37% for nuclease, and 54% and 13% for alpha-toxin. In single serum samples, the detection rate for all four antigens marginally improved the results, with detection rates of 62% and 36% for complicated and uncomplicated bacteremia, respectively. Cross-reactive antibody to peptidoglycan but not to the other three antigens was present in six (75%) of eight patients with long-standing subacute bacterial endocarditis due to either viridans streptococci or Staphylococcus epidermidis (data not shown).(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Bacterial↗
PubMed1983

Survey of taurine uptake and metabolism in Staphylococcus aureus.

Taurine has been reported to be a component of the capsular polysaccharide of the encapsulated M strain of Staphylococcus aureus. This led to a study of the uptake and metabolism of [1,2-14C]taurine in a variety of encapsulated and unencapsulated S. aureus strains. Taurine was taken up by all strains studied. A discrepancy between uptake measured as depletion of radioactivity from growth medium and as cell-associated radioactivity suggested that taurine may be catabolized to CO2 in some strains. In most strains, cell-associated radioactivity was located mainly in cold TCA-soluble (pool metabolites) fractions. About 90% of the cell-associated radioactivity was present in the pool metabolites fraction in the M strain, and about 10% in hot TCA-soluble (nucleic acid-teichoic acid-capsular polysaccharide) fraction. Radioactivity in spent medium and the capsular polysaccharide-containing fraction appeared to be present as taurine in this strain. Radioactivity in the pool metabolites fraction of three of the strains examined did not chromatograph as taurine, indicating that taurine was converted into other cell metabolites. One strain incorporated radioactivity from taurine into cellular macromolecules, thus revealing a heterogeneity of staphylococcal taurine metabolism.

Chromatography, Paper↗
PubMed1983

Methicillin-resistant septal peptidoglycan synthesis in a methicillin-resistant Staphylococcus aureus strain.

In a methicillin-resistant Staphylococcus aureus strain, electron micrographs showed that cell wall septa continued to be formed in the presence of methicillin, although they became distorted and enlarged. The results indicated that peripheral cell wall synthesis was inhibited. It is concluded that a methicillin-resistant mode of septal peptidoglycan synthesis is an important determinant of methicillin resistance.

Bacterial Proteins↗
PubMed1983

Biology and clinical significance of peptidoglycan antibody response in staphylococcal infections.

Peptidoglycan, the basic structure of the staphylococcal cell wall, is a matrix of glycan strands that are cross-linked through short peptide side chains. Many of the biological activities of staphylococcal cells can be ascribed to the peptidoglycan moiety of their cell walls. Staphylococcal peptidoglycan can be shown to be immunogenic in laboratory animals; both humoral and cellular immune responses have been noted. Sensitive techniques, such as radio- or enzyme-immunoassay, have recently shown that virtually all normal human donors have detectable peptidoglycan IgG antibodies in their serum. Peptidoglycan IgG can be transplacentally transferred. The titers of peptidoglycan antibody vary widely among healthy donors. Increased production of peptidoglycan antibodies is found in most patients with complicated S. aureus septicaemia and also in many with uncomplicated bacteremia. Nonbacteremic S. aureus infections usually do not stimulate peptidoglycan antibody production. Compared to other S. aureus products such as teichoic acid, nuclease, and alpha-toxin, peptidoglycan may be the most sensitive antigen for detecting antibody responses during staphylococcal infections. Peptidoglycan antibodies may neutralize some of the toxic effects of the staphylococcal cell wall and promote phagocytosis of the organisms. However, increased peptidoglycan antibody titers with immuno-complex disease have also been associated with longstanding infections due to S. epidermidis, Streptococci and with rheumatoid arthritis. Thus, peptidoglycan antibodies may cross-react among Gram-positive bacterial species and have detrimental effects as well.

Antibodies, Bacterial↗
PubMed1983

Recognition of Staphylococcus aureus by human phagocytes. Signals and disguises of the bacterial surface.

Phagocytic cells provide the host its major defense against invasive Staphylococcus aureus, and the staphylococcal surface, by its influence on phagocyte recognition, is a primary determinant of the function of these cells. The peptidoglycan component of the cell wall plays a key role in both opsonic and chemotactic recognition, mediated by IgG, C3b, and C5a, respectively. While cell wall protein A inhibits opsonic recognition by polymorphonuclear leukocytes, it promotes an opsonin-independent mechanism of phagocytosis by human macrophages which possess cytophilic IgG. By masking cell wall-associated opsonic molecules, capsular polysaccharides inhibit recognition, a phenomenon that is overcome by specific anti-capsular antibodies. It is proposed that impaired phagocyte recognition is a basic element in the pathogenesis of staphylococcal endocarditis, and progress in the prevention and treatment of this infection may depend on understanding the basis for this host defense defect.

Animals↗
PubMed1982

Role of peptidoglycan from Staphylococcus aureus in leukopenia, thrombocytopenia, and complement activation associated with bacteremia.

The role of the major cell wall components of Staphylococcus aureus in the leukopenia, thrombocytopenia, and complement activation associated with S. aureus bacteremia was studied in a guinea pig model. Formalin-killed S. aureus strains HSmR, 52A5, Cowan I, and Cowan EMS and purified peptidoglycan were used. Normal animals given peptidoglycan developed early (5-min) leukopenia, thrombocytopenia, and depletion of C3-C9 hemolytic activity similar to values in animals given killed S. aureus organisms and C4-deficient animals challenged with peptidoglycan. Cobra venom factor-treated animals challenged with peptidoglycan did not develop early leukopenia and thrombocytopenia, but all animal groups persistently had late (greater than 1-hr) leukopenia and thrombocytopenia. This observation suggests that peptidoglycan may play a major role in the early leukopenia and thrombocytopenia associated with S. aureus bacteremia in the guinea pig and that these effects can be mediated by activation of the alternative complement pathway alone. Peptidoglycan also causes a late leukopenia and thrombocytopenia which may occur independently of complement activation.

Animals↗
PubMed1981

Removal of Paracoccus denitrificans outer membrane material by sodium chloride.

The effects of water washing and NaCl treatment on the cell surface of P. denitrificans were studied. Both treatments caused a release of material from cells. Chemical studies showed that NaCl treatment released material containing components characteristic of outer membrane. This treatment also increased the susceptibility of the organism to lysozyme. Scanning electron microscopy was used to monitor the effects of water washing and NaCl treatment on the cell surface. Both treatments were shown to alter the appearance of the cell surface. The disruptive effects of these procedures were found to be dependent upon the age of the culture.

Cell Membrane↗
PubMed1981

Endogenous pyrogen production by human blood monocytes stimulated by staphylococcal cell wall components.

To determine the properties of Staphylococcus aureus contributing to its pyrogenicity, we compared, in human monocytes, endogenous pyrogen production stimulated by heat-killed S. aureus with that stimulated by purified S. aureus cell walls or by particulate peptidoglycan prepared from the same strain. Peptidoglycan, but not the purified cell wall preparation, was found comparable to S. aureus as an endogenous pyrogen stimulus. This finding was associated with a more effective monocyte phagocytosis of S. aureus and peptidoglycan as compared with that of purified cell walls. Lysostaphin digestion of peptidoglycan markedly reduced its pyrogenicity. To test whether the chemical composition of the ingested particles is important, latex particles were tested as possible stimuli for monocyte endogenous pyrogen release. Although 40 to 68% of monocytes ingested latex particles during the first hour, there was no evidence of endogenous pyrogen activity in the supernatant even when supernatants equivalent to 5.2 X 10(6) monocytes were tested. This study demonstrates that the pyrogenic moiety of the S. aureus cell wall resides in the peptidoglycan component. Phagocytosis is not in itself a pyrogenic stimulus, but rather serves as an effective mechanism to bring about contact between the chemical stimulus and the monocyte.

Cell Wall↗
PubMed1981

Factors affecting complement activation by Staphylococcus aureus cell walls, their components, and mutants altered in teichoic acid.

In a previous study, Staphylococcus aureus purified cell walls (PCW), consisting of peptidoglycan (PG) plus covalently linked teichoic acid (TA), were found to be more active in complement consumption than isolated PG. Isolated TA has now been shown to be capable of activating complement. Mild sonication markedly increased the ability of PG to activate complement but had essentially no effect on the activities of PCW and TA. Optimal sonication of PG did not yield activities equal to those of PCW in dose-response and kinetic studies, which may imply that TA plays some role in complement consumption. Sonication did not lead to solubilization of PCW or PG but may have enhanced the activity of PG in complement consumption by better dispersing PG particles, thereby exposing more surface area. Lysostaphin solubilization of PCW and PG markedly decreased their activities in complement consumption. The PCW of an S. aureus TA-deficient mutant, which were mostly PG, caused similar amounts of complement consumption as the parent strain PCW. Of the treatments of PCW commonly used to isolate PG, formamide and periodate extractions in particular led to PG preparations with lower activities in complement consumption than the PCW from which they were prepared, although these activities were stimulated by sonication. When whole organisms were studied by using a TA-deficient mutant, a mutant with an additional cell surface polymer, and the TA-containing parent strains and complement consumption by these strains was compared, no difference was found in either the rate or the degree of complement activation. This led to experiments demonstrating that both material released extracellularly from staphylococci and the cytoplasmic fraction of S. aureus were active in complement consumption. The results of these experiments indicate that both physical and chemical factors must be considered in studies of complement activation by isolated bacterial cell wall components. Under certain conditions, staphylococcal TA may enhance complement activation, but studies with whole organisms clearly show that this cell wall constituent does not play an essential role in this process. In addition, studies of complement consumption with intact organisms have demonstrated that there may be contributions both from cell surface components and from material released by the cells.

Cell Wall↗
PubMed1981

Binding of human immunoglobulin G to protein A in encapsulated Staphylococcus aureus.

Recent studies of the mechanism of resistance to phagocytosis in encapsulated Staphylococcus aureus have suggested that the capsule is readily penetrated by high-molecular-weight proteins such as antibodies and complement components. S. aureus strains contain a cell wall protein, protein A, that reacts with the Fc portion of immunoglobulins. The binding of immunoglobulin G (IgG) to encapsulated and unencapsulated S. aureus strains has been studied to assess the penetrability of the S. aureus capsule by IgG. Encapsulated S. aureus strains M and Smith diffuse bound large amounts of human IgG which were comparable to amounts bound by the unencapsulated strains Cowan I, M variant, and Smith compact. Trypsin treatment of bacteria reduced their ability to bind IgG. Bound IgG was not removed by extensive washing of bacteria with buffer. A non-protein A-containing, coagulase-negative, encapsulated staphylococcal strain did not bind IgG. These observations suggest that IgG is binding to cell wall protein A in encapsulated S. aureus. No differences in the rates of IgG binding by encapsulated and unencapsulated S. aureus strains were observed. It is concluded that the S. aureus capsule is freely permeable to IgG. This is of importance in considerations of the mechanisms of resistance to phagocytosis and antigen masking in encapsulated microorganisms.

Humans↗