Effects of bacillus of Calmette-Guerin, Candida albicans and human recombinant interferon-gamma on RNA metabolism and on in vitro release of tumor necrosis factor-alpha by human monocytes/macrophages.

Bacillus of Calmette-Guerin (BCG) was found to be effective in the therapy of superficial bladder cancer, although the mechanisms by which this occurs have not yet been clarified. One hypothesis is related to the ability of monocytes/macrophages (MN/M phi) to release tumor necrosis factor-alpha (TNF-alpha), a monokine with cytotoxic and cytostatic effects against certain tumor cell lines. The present study demonstrates that BCG and C. albicans are both very efficient inducers of TNF-alpha, while they inhibit uridine uptake and incorporation into human MN/M phi RNA. However, unlike C. albicans, BCG is cytotoxic for MN/M phi, as determined by release of labelled leucine from target cells.

Influence of yeasts and of their constituents on nucleoside uptake in peritoneal murine macrophages.

A marked reduction of [3H]-uridine uptake was observed when mouse peritoneal macrophages (pM phi) were exposed to heat-killed Candida albicans or Saccharomyces cerevisiae. By contrast, an increased nucleoside uptake was promoted by yeast products such as zymosan, laminarin, or yeast cell-wall extracts, which are mainly composed of beta-glucans and alpha-mannans. In a search for the active fungal component(s), the uptake process was shown to be differently affected by monosaccharides and polysaccharides. These findings support the view that a specific recognition of a pM phi membrane receptor is mediating the effect of the various substances.

Nucleoside uptake in macrophages from various murine strains: a short-time and a two-step stimulation model.

Kinetics of [3H]-uridine uptake by murine peritoneal macrophages (pM phi) is early altered after exposure to a variety of stimuli. Alterations caused by Candida albicans, lipopolysaccharide (LPS) and recombinant interferon-gamma (rIFN-gamma) were similar in SAVO, C57BL/6, C3H/HeN and C3H/HeJ mice, and were not correlated with an activation process as shown by the amount of tumor necrosis factor-alpha (TNF-alpha) being released. Short-time exposure to all stimuli resulted in an increased nucleoside uptake by SAVO pM phi, suggesting that the tumoricidal function of this cell either depends from the type of stimulus or the time when the specific interaction with the cell receptor is taking place. Experiments with priming and triggering signals confirmed the above findings, indicating that the increase or the decrease of nucleoside uptake into the cell depends essentially on the chemical nature of the priming stimulus. The triggering stimulus, on the other hand, is only able to amplify the primary response.

Characterisation of Escherichia coli adhesins in patients with symptomatic urinary tract infections.

The ability of bacteria to adhere to the epithelial cells of hosts has been shown to be mediated by adhesins. Many of these show readily demonstrable haemagglutinating activity. Of 109 Escherichia coli strains isolated from patients with symptomatic urinary tract infection, 11 (10.1%) were identified by their haemagglutinating properties as being P fimbriated, which was confirmed by the latex bead test. Other classes of adhesins, termed X and "other", were found in mannose resistant haemagglutinating E coli strains, which represented 4.6% (5) and 0.9% (1), respectively, of all the strains. Type 1 fimbriae were found in 40.4% (44/109) of E coli strains grown on colonising factor agar (CFA) medium. This incidence was 12.8% higher (53.2%, 58/109) when the strains were grown on CFA supplemented with urea, which suggested that urea may modulate the expression of type 1 fimbriae. Conversely, this phenomenon was not seen in P fimbriated E coli. Assays using trypsinised and non-trypsinised human erythrocytes showed no difference in the percentage of strains that haemagglutinated. Regarding the clinical correlation of fimbriated E coli strains, the X mannose resistant haemagglutinating adhesins were also found to be of clinical relevance. P fimbriated E coli strains were isolated from five out of the eight patients with pyelonephritis.

Nucleoside transport in activated macrophages.

The study of [3H]-uridine uptake by mouse peritoneal macrophages showed that this is an active, temperature- and protein synthesis-dependent phenomenon, which is early altered when are exposed to a variety of stimuli. Murine recombinant interferon-gamma, a stimulus able to activate macrophage and to induce the production of tumor necrosis factor-alpha, within few hours markedly increased [3H]-uridine uptake by mouse macrophage. Other stimuli devoid of activation capacity, such as inert phagocytable latex beads, did not affect this phenomenon, which appeared to be related to macrophage activation. The increase in [3H]-uridine uptake may be an useful phenomenon in studying the early biochemical events associated with macrophage activation.

In vitro activity of antibiotics against Ureaplasma urealyticum and Chlamydia trachomatis strains from patients with nongonococcal urethritis.

The activity of minocycline, doxycycline, tetracycline, erythromycin, mepartricin and lincomycin against 35 freshly isolated Ureaplasma urealyticum strains was tested. Doxycycline was the most active. Twelve strains were resistant to minocycline and four of these were sensitive to erythromycin. Mepartricin showed no activity against the organisms at a concentration of 10 micrograms/ml. The susceptibility of 30 low-laboratory-passage Chlamydia trachomatis strains against tetracycline and erythromycin was tested. A variable degree of sensitivity to tetracycline and erythromycin was found, the median MIC values being 0.13 micrograms/ml and 0.025 micrograms/ml respectively. No resistant Chlamydia trachomatis strain was found.

Enzyme-linked immunosorbent assay for detection of Mycoplasma pneumoniae antibodies.

The enzyme-linked immunosorbent assay (ELISA) described by Engvall and Perlmann has been used for the detection of antibodies against Mycoplasma pneumoniae. These organisms, grown in flat-bottom wells of microtiter plates, were used as antigen. Preliminary results suggest that ELISA is specific and that its sensitivity is somewhere in between those of the metabolic inhibition and radioimmunoprecipitation methods. Unlike other serological methods, such as metabolic inhibition and mycoplasmacidal tests, in which the presence of antibiotics in sera may give false results, ELISA provided results that were not influenced by antibiotics. Furthermore, ELISA offers the possiblity of measuring not only immunoglobulin M, but also complement-independent antibodies, especially those of the immunoglobulin A class. Theoretical aspects concerning the different sensitivities of some serological reactions are discussed.

Colony morphology, ultrastructure and morphogenesis in Mycoplasma hominis, Acholeplasma laidlawii and Ureaplasma urealyticum.

Colonies of Mycoplasma hominis, Acholeplasma laidlawii (three strains) and Ureaplasma urealyticum were examined by light and electron microscopy and their characteristic morphology, ultrastructure and morphogenesis are described. Mycoplasma hominis and A. laidlawii, PG8 and oral strains, developed typical 'fried-egg' colonies which were remarkably heterogeneous in size. The colonies of A. laidlawii strain NCTC 10116 were more homogeneous and grew mainly on the surface of the agar showing a fine granular appearance. Ureaplasma urealyticum produced smaller, granular colonies which grew deeply embedded in the agar and generally without much surface growth. The cellular ultrastructure in these colonies was also examined. The results indicate that several aspects of colony morphogenesis and ultrastructure varied for each of the three species examined.

Antibody-coated bacteria in the urinary sediment of rats with experimental pyelonephritis.

The appearance of antibody-coated bacteria in urinary sediment has been evaluated in rats with experimental pyelonephritis. At day 7 after induction of pyelonephritis, 9 out of 11 rats demonstrated antibody-coated bacteria in the urinary sediment. The other 2 rats never had a positive urinary sediment. Following removal of the pyelonephritic kidneys, antibody-coated bacteria disappeared in 7 of 9 previously positive rats. In the 2 rats which continued to show antibody-coated bacteria, the infecting organisms were found in the remaining kidney. The 2 rats which never developed antibody-coated bacteria in urinary sediment had a higher then normal serum antibody titer, and 1 rat with antibody-coated bacteria showed a normal serum antibody titer. It is concluded that although the search for antibody-coated bacteria in urinary sediment is a very useful technique, its negativity does not exclude upper urinary tract involvement; in the case of renal parenchymal infection, immunity is not the same at the systemic and the local site.

[Induction of prophage gamma by means of bleomycin treatment]. / Induzione del profago lambda mediante trattamento con bleomicina

The kinetics of induction of lambda prophage with bleomycin A2-CP3, mitomycin C and a combination of these two antibiotics have been studied. Bleomycin has been found to cause a rapid induction of prophage in exponentially growing cultures of E. coli K12 (lambda), and the maximal values of including activity are obtained after treatment for 16 minutes with a concentration of 10 mug/ml. The per cent of induction, calculated from the total number of surviving cells, including the phage producing cells, better expresses the dynamics of the phenomenon. The number of inducing events in the population of exposed cells is proportional to the length of bleomycin treatment. These values are greater than those observed after treatment with mitomycin, at the same concentration. Inducing activity is enhanced by combination of the two antibiotics and this effect is not additive, but synergic. After the 16 minutes of treatment with bleomycin the burst sizes are lower than those which are obtained with mitomycin or with the combination of the two antibiotics. The results are interpreted as a consequence and a confirmation of the different mechanisms of action that distinguishes bleomycin from mitomycin.

Glutamate dehydrogenase from Escherichia coli: induction, purification and properties of the enzyme.

When Escherichia coli was grown in a minimum medium with glucose as sole carbon source and a proper level of ammonia, NADP+ specific glutamate dehydrogenase (L-glutamate: NADP+ oxidoreductase (deaminating), ED 1.4.1.4) was induced. The enzyme was solubilized by French press treatment and purified to homogeneity by (NH4)2SO4 fractionation, heat treatment followed by DEAE-cellulose, hydroxylapatite and Bio-Gel chromatography with an overall yield of 30%. The enzyme proved to be heat stable and relatively resistant to protein denaturants. The optimum of enzymic activity for the reductive amination is at pH 8 and at pH 9 for the oxidative deamination. The activity is affected by adenine nucleotides. The molecular weight (about 250 000 for the native form and 46 000 for the inactive subunit) and amino acid composition, suggest strict similarities with the NADP+ enzyme from fungal origin.