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Biomedical subjects

G M Wiseman

Publications and source records attributed to G M Wiseman.

At least 19 recordsLinked to original sources
PubMed1991

Stimulation by gonococci of chloride ion uptake in human leucocytes in relation to other properties of phagocytosis.

Human polymorphonuclear leucocytes (PMNL) underwent several changes in response to challenge with Neisseria gonorrhoeae, namely (1) an increase in oxygen uptake, (2) changes in membrane electrical properties, and (3) increased transport of chloride ions (Cl-) across the membrane. Mean oxygen consumption and Cl- uptake by PMNL were stimulated by both pilate (P+) and non-pilate (P-) gonococci, although the levels were much reduced in the presence of P+ organisms. P+ gonococci also initiated low levels of polarisation or depolarisation in contrast with P- cells, which caused hyperpolarisation followed by depolarisation in the PMNL. Most of the strains showed these patterns. High performance liquid chromatography of extracts of unstimulated PMNL and of PMNL challenged with gonococci confirmed production of hypochlorous acid (HOCl) in the leucocyte. Furthermore, addition of radiolabelled Cl- to the PMNLs showed that some of the Cl- taken up by the cells in response to gonococcal challenge was incorporated into the HOCl, suggesting a direct relationship between stimulation of Cl- uptake and production of active chlorine compounds in the leucocyte.

Biological Transport↗
PubMed1987

Relationship between lipopolysaccharide composition and virulence of Haemophilus ducreyi.

The relationship between lipopolysaccharide (LPS) composition and virulence of Haemophilus ducreyi strains was investigated. Glycoses identified in LPS by gas-liquid chromatography were glucose, galactose, and their amino derivatives glucosamine and galactosamine. Fucose was found in trace amounts but mannose and rhamnose, characteristic of the O-side chain of LPS in many species, were not detected. Qualitatively, the LPS composition of the eight strains examined was similar and differences were mainly quantitative. The total glycose:KDO ratio of the LPS of virulent strains exceeded that of avirulent strains. All strains had similar fatty-acid composition but lacked lauric acid. SDS-polyacrylamide gel electrophoresis of the LPS of virulent and avirulent strains also revealed differences in their electrophoretic mobilities. The LPS profiles of avirulent strains were similar, but differed from those of virulent strains. These profiles lacked high mol. wt bands representing O-side chain repeating units. Thus, differences in the electrophoretic mobilities of the LPS of virulent and avirulent strains may reflect differences in the amount of carbohydrates associated with the core polysaccharide.

Electrophoresis, Polyacrylamide Gel↗
PubMed1986

Changes in host cell membrane activities in response to adhesion of Neisseria gonorrhoeae.

Physiological changes in host cell model membranes (intact human erythrocytes and ghosts) as a consequence of bacterial adhesion were studied with special reference to Neisseria gonorrhoeae. Membrane activities examined were transport of K+, Cl- ions, pyruvate kinase, Na-K-dependent ATPase, and cAMP. We found that K+ and Cl- transport were affected, more so in membranes with attached pilated (P+) organisms than in those with apilated (P-) isogenic strains. In N. gonorrhoeae and in several other species of gram-negative bacteria studied, hemagglutination titres were directly correlated with effects on anion transport, suggesting that perturbations in anion transport are an immediate result of adhesion. Of three P+ gonococcus strains tested, two depressed Na-K-ATPase activity in the membrane, indicating a possible effect on the Na-K pump. Pyruvate kinase activity associated with the membrane appeared to be stimulated by attached gonococci, again by P+ strains to higher levels than P- organisms. Clearly, some enzyme properties of host membranes are intrinsically affected by bacterial adhesion. Human polymorphonuclear neutrophils were also investigated, and with some exceptions, changes observed in leukocyte enzyme activities tended to parallel those in erythrocytes. Since hypochlorous acid production is considered to be an important microbicidal mechanism in neutrophils, interference with Cl- transport could jeopardize their role in host defense.

Adhesiveness↗
PubMed1985

Role of lipopolysaccharide and complement in susceptibility of Haemophilus ducreyi to human serum.

The role of lipopolysaccharide (LPS) in the susceptibility of Haemophilus ducreyi to human serum and the mechanism of complement activation by serum-susceptible (Sers) strains were investigated. Serum treated with 2 mM Mg2+ and 20 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid was nonbactericidal, but inulin-treated serum remained bactericidal. Absorption of serum with heat-killed whole cells of an Sers strain removed its bactericidal activity against the absorbing strain and also against other Sers strains. LPS obtained from Sers strains inhibited the bactericidal activity of serum against all Sers strains, whereas LPS from serum-resistant (Serr) strains and an Serr isogenic strain did not. However, high concentrations of LPS from the Serr strain inhibited the bactericidal activity of serum, an indication that part of the structural site involved in serum susceptibility is retained in the LPS of this strain. The LPS of Sers strains exhibited higher anticomplement activity than the LPS of Serr strains. These findings suggest that the classical pathway of complement activation is involved in the serum killing of H. ducreyi and that LPS composition may contribute to their susceptibility to complement-mediated serum bactericidal activity.

Animals↗
PubMed1984

Virulence factors of Haemophilus ducreyi.

We investigated the susceptibility of virulent and avirulent strains of Haemophilus ducreyi to the bactericidal activity of normal human serum and to phagocytosis and killing by human polymorphonuclear leukocytes (PMNL). Strains were defined as virulent if intradermal inoculation into a rabbit produced a typical necrotic lesion. Nonvirulent strains produced no cutaneous lesions in rabbits. Virulent strains were resistant to the complement-mediated lethal action of normal human and rabbit sera, whereas avirulent strains were susceptible (greater than 95% kill, 60 min). Virulent strains were relatively resistant to phagocytosis and killing by human PMNL, in contrast to the avirulent strains. In past studies polymyxin resistance has been correlated with virulence in H. ducreyi. In our studies, polymyxin resistance could not be correlated with virulence, since polymyxin-sensitive mutants obtained from polymyxin-resistant parent strains remained virulent for rabbits and resistant to bactericidal action of normal serum and phagocytosis and killing by human PMNL. Similarly, polymyxin-resistant mutants obtained from polymyxin-sensitive parent strains remained avirulent for rabbits and susceptible to bactericidal action of normal serum and PMNL. The acquisition of polymyxin resistance was accompanied by the loss of a 47,000-molecular-weight protein. The association of serum resistance and resistance to phagocytosis and killing by human PMNL with virulent strains, as defined by the rabbit intradermal test, suggests that these factors may mediate the pathogenicity of H. ducreyi.

Animals↗
PubMed1984

Inhibition of anion transport in human erythrocytes by pilated Neisseria gonorrhoeae.

Pilated gonococci added to human erythrocytes apparently bind to polypeptide band 3 and inhibit chloride-bicarbonate exchange across the membrane in contrast with nonpilated organisms. Known covalent inhibitors of anion transport (4,4'-diisothiocyano-2,2'-disulphonic acid stilbene and alpha-cyano-4-hydroxycinnamic acid) inhibit chloride efflux from erythrocytes and also reduce hemagglutination titres, strongly suggesting that band 3 is the major host-cell receptor involved in all interaction between gonococci and erythrocytes. In studies of other cell species, however, band 3 prepared from human erythrocytes inhibited adherence of pilated gonococci to human foreskin and HeLa cells, but was without effect in human buccal, Vero, and mouse L cells.

Adhesiveness↗
PubMed1982

The nature of human erythrocyte receptors for Neisseria gonorrhoeae.

Normal and trypsinized human erythrocyte membranes were used as a model in the study of host cell receptors for Neisseria gonorrhoeae. Receptor sites were identified by adherence inhibition assays of fractions of membranes eluted from polyacrylamide gel electrophoresis columns. Results indicated that inhibition of gonococcus T1 and T4 adherence was associated with erythrocyte protein band 3 and 4 and glycophorin A, the major sialoglycoprotein. Further investigation revealed that band 3 preparations isolated by affinity chromatography on concanavalin A - Sepharose 4B columns continued to inhibit T1 adherence to erythrocytes but did not inhibit adherence of T4 organisms. It is suggested that protein band 3 is the receptor on erythrocytes for T1 gonococci and that glycophorin A may be the receptor for T4 cells.

Adhesiveness↗
PubMed1981

Some properties of the human erythrocyte receptors for Neisseria gonorrhoeae.

The nature of the receptors for T1 and T4 Neisseria gonorrhoeae on erythrocytes and other cells was investigated. In general, cells of nonprimate origin contained few receptors for gonococci. Receptors for T4 gonococci were only uncovered when host cells were pretreated with trypsin. Trypsinization, while unnecessary for T1 adherence to erythrocytes, enhanced attachment in inverse proportion to original erythrocyte sensitivity. Receptors for T1 and T4 organisms on trypsinized and trypsin-neuraminidase-treated erythrocytes were blocked by concanavalin A and peanut lectins, respectively, but a distinction could be made between them with wheat germ lectin and galactose oxidase. Of a number of sugars tested as inhibitors, only D-galactose blocked adherence of T4 but was without effect on T1. While the identity of erythrocyte receptors is uncertain, likely candidates are "band 3" protein and glycophorin, by virtue of their galactose content, lectin binding capacity, and partial exposure on the outer surface of the erythrocyte.

Adhesiveness↗
PubMed1977

Composition of the lipopolysaccharide of Neisseria gonorrhoeae.

Analysis of glycose and fatty acid content of lipopolysaccharide extracted from 38 strains of Neisseria gonorrhoeae indicated that glycoses common to colonial types 1 to 5 were glucose, mannose, and galactose, N-acetylneuraminic acid, 2-keto-3-deoxyoctulosonic acid (KDO), glucosamine, and galactosamine were also invariably present. Virulent colonial types 1 and 2 contained no rhamnose, in contrast to avirulent types 3 to 5 and several strains of the nonpathogenic species N. sicca and N. lactamica. Fucose, characteristic of these nonpathogenic species, was not present in the gonococci. Variation in the concentration of individual glycoses in different strains was also noted. Mannose-KDO, galactose-KDO, and glucose-KDO ratios of virulent gonococci exceeded those of avirulent organisms, except that the correlation for glucose was not quite so striking. This relationship was not found in N. sicca and N. lactamica strains. Fatty acid analyses of lipid A from gonococci showed that 10-, 12-, 14-, 16-, and 18-carbon acids, as well as 3-hydroxytetradecanoic acid, were present, but differences in concentration between colonial types, although evident in some cases, appeared less significant than glycose content.

Fatty Acids↗
PubMed1976

Production and purification of the gamma haemolysin of Staphylococcus aureus 'Smith 5R'.

The gamma haemolysin of Staphylococcus aureus 'Smith 5R' was produced on Dolman-Wilson agar overlain with cellophane. Maximal yields of crude lysin with titres of 2000 to 4000 haemolytic units/ml were obtained within 24 h at 37 degrees C in 10% (v/v) CO2 in air, on medium adjusted to pH 7-0. The crude lysin was purified 2700-fold (with 75% recovery) by ultrafiltration, gel filtration and ammonium sulphate fractionation. The specific activity of the lysin was 10(5) haemolytic units/mg protein after the dialysed active precipitate was extracted with NaCl and reprecipitated with ammonium sulphate. Purified gamma lysin was homogeneous by disc electrophoresis and immunoelectrophoresis.

Ammonium Sulfate↗
PubMed1976

Properties of the gamma haemolysin of Staphylococcus aureus 'Smith 5R'.

Purified gamma haemolysin of Staphylococcus aureus was characterized in relation to the alpha, beta and delta haemolysins. The sedimentation coefficient of the gamma lysin was 2-65, somewhat higher than the S20,w values of 1-4 for freshly purified alpha lysin and 1-8 for the beta lysin. The molecular weight of gamma lysin determined by gel filtration was 45000 daltons. The pI of gamma lysin was 6-0, while that of the alpha, beta and delta lysins ranged from 8-5 to 9-6. The amino acid analysis of gamma lysin was characterized by low levels of methionine and histidine. Methionine was, however, the N-terminus, which suggested that all of the amino acid might be involved in the N-terminal group. The gamma lysin was immunologically distinct from the alpha, beta and delta lysins by quantitative precipitin tests; in Ouchterlony agar gel diffusion tests, single lines of precipitation were observed which showed no evidence of cross-reactions amongst the four haemolysins. Gamma, beta and delta lysins had no affect in mice when injected at increasing doses ranging from 0 to 100 mug. The alpha lysin killed mice, the LD50 dose being 0-60 +/- 0-12 mug, or 27 to 34 mug/kg mouse tissue. Gamma lysin was, however, lethal for guinea pigs when 50 mug quantities were injected intracradially. Gamma lysin also lysed human leucocytes and destroyed C-6 (human lymphoblast) cells.

Amino Acids↗
PubMed1975

Trypsin-mediated activation of the alpha-haemolysin of Staphylococcus aureus.

Alpha protoxin of Staphylococcus aureus "Wood 46" was activated by trypsin which had been coupled to carboxymethylcellulose, as indicated by the toxin's ability to hydrolyse tosyl-arginine methylester (TAME). A Lineweaver-Burk plot of the degradation of TAME by toxin and trypsin showed that toxin had a greater affinity for the substrate than had trypsin. N-terminal amino-acid analyses of activated toxin suggested that leucine or isoleucine is the N-terminus, in contrast to protoxin, the N-terminus of which is histidine.

Amino Acid Sequence↗
PubMed1975

A new colonial type of N. gonorrhoeae.

A new variant of Neisseria gonorrhoeae, designated Type 1(1), is described. Colonies of the new type resemble those of Types 1 and 2 in physical characteristics but are granular with a slightly crenated edge and are a deeper gold in colour. The virulence of Type 1(1) in the chick embryo is in keeping with that of Types 1 and 2 but is significantly different from Types 3, 4, and 5. Type 1(1) could be maintained in the laboratory for 6 months, provided that daily selective subcultures were performed. In the absence of this, Type 1(1) reverted to Type 5. It was also possible to preserve the stability of Type 1(1) for long periods by immersion in liquid nitrogen. Pili have been demonstrated on the new type.

Animals↗
PubMed1975

Cultivation of type 1 N. Gonorrhoeae in liquid media.

The effect of CO2 concentration on the growth and colonial stability of Type 1 Neisseria gonorrhoeae has been investigated. Carbon dioxide at a concentration of 16 per cent in air above flasks incubated in a shaker was effective in supporting growth and 100 per cent colonial stability of Types 1, 1', 2, and 4. Lower CO2 tensions increased the generation time of the strains and were less effective in maintaining the stability of virulent variants. Of several liquid media tested, Enriched Single Phase medium, which consists of Difco GC medium base (devoid of agar and starch) to which Lankford supplement and "Isovitalex" have been added, was the most suitable for use with small inocula.

Carbon Dioxide↗