Biomedical subjects

G Satta

Publications and source records attributed to G Satta.

At least 109 records · Page 6Linked to original sources

PubMed1983

Laboratory and wild-type Klebsiella pneumoniae strains carrying mannose-inhibitable adhesins and receptors for coliphages T3 and T7 are more pathogenic for mice than are strains without such receptors.

We have shown previously that Klebsiella pneumoniae receptors for coliphages T3 and T7 also mediate mannose-inhibitable adherence to human epithelial cells and protect bacteria from phagocytosis and intracellular killing by human polymorphonuclear cells. In this paper we analyze the possible role of such mannose-inhibitable adhesins and T3-T7 receptors (MIAT) in K. pneumoniae intraperitoneal pathogenicity for mice. We showed that intraperitoneal pathogenicity for mice of four different Klebsiella strains (one laboratory and three wild-type) that carry the MIAT was approximately 60-fold higher than that of four derivative strains that lost such receptors by spontaneous mutation. The MIAT could be repressed by Klebsiella phage AP3 lysogenic conversion. Two laboratory and two wild-type strains converted by phage AP3 were also approximately 60-fold less pathogenic for mice than parental strains and showed a pathogenicity level equal to that of the MIAT-negative mutants. Studies of protection in mice with anti-whole cell antisera showed that passive immunization against MIAT-positive cells was more protective than immunization against MIAT-negative cells. Studies of protection in mice by both active and passive immunization with lipopolysaccharide and purified outer membrane proteins have shown that the proteins are the most protective outer membrane components. Since it has been shown previously that the Klebsiella receptors for T3-T7 have a proteic component and that an outer membrane protein is missing in the strains resistant to T3-T7 (C. Pruzzo et al., in R. C. Berkely (ed.), Microbial Adhesion to Surfaces, 1980); the latter finding further supports the role of MIAT in the pathogenicity of Klebsiella for mice.

Adhesiveness↗

PubMed1983

Streptococcus faecium ATCC 9790 penicillin-binding proteins and penicillin sensitivity are heavily influenced by growth conditions: proposal for an indirect mechanism of growth inhibition by beta-lactams.

The effects of variations in growth conditions on the penicillin response of Streptococcus faecium ATCC 9790 were studied. Changes in the growth temperature and medium composition were found to cause striking changes in the bacterial generation time, cellular penicillin sensitivity (minimum inhibitory concentration), sensitivity of peptidoglycan synthesis to inhibition by penicillin, rate of autolysis, and labeling pattern of penicillin-binding proteins. However, no constant relationship between these parameters and the minimum inhibitory concentration could be observed. Similar electrophoretic patterns for penicillin-binding proteins were observed in cells grown in different media at the optimal growth temperature. Inhibition of cell division by penicillin in cells grown at this temperature (but not at higher or lower temperatures) caused filamentation of the bacteria. In cells grown in a chemically defined medium at the optimal temperature (but not at temperatures above or below), complete inhibition of cell division was associated with only partial inhibition (34% after 150 min) of peptidoglycan synthesis. It is suggested that the status and physiological importance of individual penicillin-binding proteins in S. faecium are heavily influenced by growth conditions. Depending on the growth conditions, different penicillin-binding proteins may perform the cellular function, indispensible for bacterial growth.

Bacterial Proteins↗

PubMed1983

Division blocks in temperature-sensitive mutants of Streptococcus faecium (S. faecalis ATCC 9790).

Two hundred nine temperature-sensitive growth or division (or both) mutants of Streptococcus faecium ATCC 9790 were isolated. These strains were examined for timing of the division block in the cell division cycle. About 42% of the isolates were blocked at terminal stages of cell division. A second large group appeared to be blocked at various stages of septation. Only five of the temperature-sensitive isolates were blocked at a stage before the completion of chromosome replication. Thirty temperature-sensitive isolates lysed after one or more doublings at the nonpermissive temperature.

Cell Division↗

PubMed1983

Suppression of electroacupuncture(EA)-induced beta-endorphin and ACTH release by hydrocortisone in man. Absence of effects on EA-induced anaesthesia.

Auricular electroacupuncture (EA) increased plasma ACTH and beta-endorphin levels significantly in 10 patients receiving EA as an analgesic aid during surgery. Pre-treatment with iv hydrocortisone (200 mg) completely suppressed both ACTH and beta-endorphin release in response to EA without significantly affecting EA anaesthesia in 6 other patients and in a patient with Addison's disease.

Acupuncture Therapy↗

PubMed1983

Biochemical and physical properties of the endo-beta-N-acetylglucosaminidases from Staphylococcus aureus, Staphylococcus simulans, and Staphylococcus saprophyticus.

Biochemical and physical properties of the pure bacteriolytic enzymes excreted by three different Staphylococcus species (S. aureus, S. simulans, and S. saprophyticus) were investigated. Although the three enzymes have previously been shown to share the same specificity of action (endo-beta-N-acetylglucosaminidase activity), their biochemical features clearly indicated that they were three different enzymes, confirming what had previously been suggested by the different lytic-activity patterns displayed by each species and the different procedures needed to achieve purification of each enzyme. Very different values resulted from molecular weight determination: 80,000 for the S. aureus enzyme, 45,000 for the S. simulans enzyme and 31,000 for the S. saprophyticus enzyme. Other important differences were observed in their kinetics of activity on Micrococcus luteus purified cell walls; their stability; their bacteriolytic spectrum against heat-killed cells of various microorganisms; and their response to physical and chemical factors, such as temperature, pH, ionic strength, divalent cations, chelating agents, thiol compounds, and glucose derivatives.

Acetylglucosaminidase↗

PubMed1982

[Immunohistochemical observations on tubulin and alpha actinin during mitosis of cultured fibroblasts].

In this work we studied alpha-actinin and tubulin sites in rabbit fibroblasts in culture. Antibodies anti-alpha-actinin were used for indirect PAP-reaction while antibodies anti-tubulin were used for indirect immunofluorescence method. The observations were carried out by light microscopy, phase-contrast and interference-contrast microscopy, with regard to actinin, and by fluorescence microscopy, with regard to tubulin. During the early mitotic phase, alpha-actinin is localized all over the cell membrane of the fibroblasts, forming a sort strong protective cap, while during diacinesis it forms only rings, localizing below the cell membrane and the philopodia. Thus tubulin forms the bundle fibres during mitotic phases.

Actinin↗

PubMed1982

[Dynamic aspects of alpha-actinin in cultured rabbit fibroblasts studied with immunofluorescence].

In this work we study alpha-actinin sites in rabbit fibroblasts in culture. For this purpose we extracted alpha-actinin from rabbit striated skeletal muscle and produced the relative antibodies in sheep. Electrophoresis was performed, on PAA slab-gel and the immunodiffusion test on agarose slab-gel. Antibodies were used for direct and indirect immunofluorescence techniques by TRIC and FITC. We noticed that in young fibroblasts alpha-actinin is concentrated in perinuclear regions, while in adult fibroblasts it is scattered, more or less evenly in the cytoplasm and above all on the plates which link the cell membrane to the substractum. Both the direct immunofluorescence method with antibodies anti-alpha-actinin, marked by TRITC, and the indirect one with sheep IgG, marked by FITC, gave identical results.

Actinin↗

PubMed1982

Mannose-inhibitable adhesins and T3-T7 receptors of Klebsiella pneumoniae inhibit phagocytosis and intracellular killing by human polymorphonuclear leukocytes.

It has recently been shown that Klebsiella pneumoniae strains adhere to human epithelial cells and that adherence is mediated by mannose-inhibitable adhesins which are also receptors for coliphages T3 and T7. We have now found that Klebsiella strain K59, which adheres to human epithelial cells and carries the receptors for coliphages T3 and T7, adheres to human polymorphonuclear leukocytes (PMN) at 4 degrees C. Strains KRTT1 and KRTT2, which are spontaneous mutants unable to adsorb coliphages T3 and T7 and adhere to human epithelial cells, at this temperature did not adhere to PMN. Adherence of K59 cells to PMN at 4 degrees C was inhibited by D-mannose, by UV-inactivated T7 phages, and by pepsin-digested anti-K59 antibodies absorbed with KRTT1 cells. At 37 degrees C the number of PMN with KRTT bacteria associated was fourfold higher than at 4 degrees C. On the contrary, the number of PMN with K59 bacteria associated at this temperature was fourfold lower than at 4 degrees C. Phagocytosis and intracellular killing experiments performed at 37 degrees C showed that KRTT1 and KRTT2 were phagocytized and killed at a higher rate than K59. After blocking of the mannose-inhibitable adhesins and T3-T7 receptors (MIAT) by D-mannose, UV-inactivated bacteriophage T7, or specific antibodies, K59 cells became more sensitive to phagocytosis and intracellular killing at 37 degrees C. K59 cells lysogenic for prophage AP3 were approximately as sensitive to phagocytosis and intracellular killing by human PMN as strains KRTT1 and KRTT2. Unencapsulated Klebsiella strains isolated from clinical specimens were found to carry MIAT most often. Four such strains were found much more resistant to phagocytosis and intracellular killing than their spontaneous mutants resistant to bacteriophages T3 and T7.

Bacterial Proteins↗

PubMed1982

Purification and characterization of three separate bacteriolytic enzymes excreted by Staphylococcus aureus, Staphylococcus simulans, and Staphylococcus saprophyticus.

As a further development of previous investigations showing that different staphylococcal species display different bacteriolytic activity patterns (lyogroups), the bacteriolytic enzymes excreted by three different Staphylococcus species, Staphylococcus aureus (lyogroup I), S. simulans (lyogroup II), and S. saprophyticus (lyogroup IV); have been purified and characterized. A representative strain from each species was grown in a preselected medium made of fully dialyzable products. Culture supernatants were collected in the appropriate growth phase. Two different affinity adsorbents were used for enzyme purification. One was obtained by coupling lysozyme-digested pure peptidoglycan from Micrococcus luteus to cyanogen bromide-activated Sepharose 4B. The second affinity adsorbent used was chitin. The S. aureus bacteriolytic enzyme bound to the solubilized peptidoglycan but not to chitin, whereas the opposite was true for the S. simulans enzyme. The bacteriolytic enzyme from S. saprophyticus did not bind to either the Sepharose 4B-peptidoglycan resin or to chitin, and its purification was achieved by two ion-exchange chromatography steps combined with gel filtration. All three enzymes were purified to apparent homogeneity. Their subsequent characterization indicated that all acted as endo-beta-N-acetylglucosaminidases. However, the three glucosaminidases differed significantly in their kinetics of activity and bacteriolytic spectrum against heat-killed cells of a variety of microorganisms. Very different values also resulted from molecular weight determinations: 80,000 for the S. aureus enzyme, 45,000 for the S. simulans enzyme, and 31,000 for the S. saprophyticus enzyme. Other important differences were observed in their stability, optimal pH and ionic strength for their activity, and their responses to temperature and divalent cations. These results confirmed the previous proposal that different staphylococcal species excrete different lytic enzymes.

Acetylglucosaminidase↗

PubMed1981

Early initiation of deoxyribonucleic acid replication and shortening of generation time associated with inhibition of lateral wall formation by mecillinam.

The effects of mecillinam on the growth of rods of the pH-conditional morphology mutant MirM7 was studied. It has been found that mecillinam causes, coincident with transition to coccal shape, a balanced rise in the rate of viable count increase and the rate of macromolecular synthesis which lasts either until the cells enter a stationary growth phase or indefinitely, in the case of continuously diluted cultures. When the antibiotic is removed from cells which have already become coccoid, cells continue to grow at a faster rate until they resume the rod shape. No change in the per-cell rate of protein synthesis has been seen in untreated or mecillinam-treated cells before or after the change in growth rate. Studies with synchronously growing cells have shown that the antibiotic causes a shortening in the I period (initiation of deoxyribonucleic acid replication). Evaluation of the residual divisions in nalidixic acid-treated, exponential-phase cells has shown that mecillinam also shortens the D period (cell division). It is proposed that, in strain MirM7, inhibition of lateral wall elongation by the antibiotic allows the initiation of a new septum, though inhibition is still in progress. The initiation of a new septum is, in turn, responsible for both the early inibition of deoxyribonucleic acid replication and accelerated division. In the parental strain, MirA12, as well as in other sensitive gram-negative rods which divide, become cocci, and stop dividing after addition of the antibiotic, inhibition of lateral wall formation activates a feedback mechanism which prevents insertion of new septa (Satta et al., J. Bacteriol. 142:43-51, 1980). Consequently, no early initiation of deoxyribonucleic acid replication is observed, and the last division allowed by the antibiotic occurs in due time. This negative control is missing in MirM7.

Amdinocillin↗

PubMed1980

Identification of the lethal target of benzylpenicillin in Streptococcus faecalis by in vivo penicillin binding studies.

The mode of bacterial killing by penicillins is still unknown in spite of many studies on the subject. The recent finding of multiple penicillin binding proteins (PBPs) in sensitive bacteria and the possibility of analysing the binding of the antibiotic to exponentially growing cells have provided new directions for investigating this problem. Sensitivity to lethal and other effects of penicillin varies very significantly with the conditions of growth of the cells. If PBPs were the penicillin target, changes in conditions of growth causing variations in penicillin sensitivity should be accompanied by changes in these proteins. Furthermore, if one of PBPs could be identified as the killing target, it could possibly be demonstrated to show changes in cells growing in different conditions. We show here that in Streptococcus faecalis ATCC 9790 changes in conditions of growth are accompanied by changes in PBPs. Furthermore, in the presence of the minimal dose of 14C-benzylpenicillin causing complete inhibition of cell growth, 100% of the total radioactivity is bound to a single protein (PBP 3).

Bacterial Proteins↗

PubMed1980

Stimulation of spreading of trypsinized human fibroblasts by lysozymes from Staphylococcus aureus, hen egg white, and human urine.

The effect of lysozyme from three different sources--Staphylococcus aureus, hen egg white, and human urine--on adhesion to substrate and spreading of trypsinized human fibroblasts was studied. Several fibroblast strains were tested under various conditions. It was found that the different cell strains did not show the same capability of spreading and stably attaching to substrates when resuspended in media not containing serum. Some strains did not spread, whereas others spread even in the absence of serum. Cell spreading in these strains did not occur when the cells were pregrown for 5 weeks in media supplemented with 1% fetal bovine serum. Lysozyme from S. aureus allowed stable adhesion to substrate and spreading of all the fibroblast strains unable to elongate in nonsupplemented minimal essential medium. This enzyme accelerated and augmented spreading of the strains capable of elongating in the absence of serum. S. aureus lysozyme also allowed spreading and stable adhesion to substrates of all these strains when they were pregrown for 5 weeks in the presence of 1% fetal bovine serum. Furthermore, hen egg white lysozyme and the lysozyme purified from human urine were both capable of stimulating anchorage to substrate and spreading of trypsinized fibroblasts although their effect was less pronounced than that of the S. aureus lysozyme. Some tentative hypotheses for the mechanism of cell spreading in the presence of lysozyme are made. The possibility that lysozymes, virtually ubiquitous enzymes, may play a specific role in nature in the regulation of cell differentiation and tissue development is finally raised and discussed in light of several previous observations and findings.

Animals↗

PubMed1980

Identification of the major adherence ligand of Klebsiella pneumoniae in the receptor for coliphage T7 and alteration of Klebsiella adherence properties by lysogenic conversion.

We have studied the adherence of both laboratory and wild-type Klebsiella pneumoniae strains, isolated from sputum, urine, and stool samples, to human buccal and intestinal and urinary tract epithelial cells. Of 32 unencapsulated strains, 30 adhered to all epithelial cells tested. Four K. pneumoniae strains lysogenic for AP3, a phage which causes conversion to resistance of coliphages T3, T7, and phi I, were all unable to adhere to epithelial cells. One of these strains was cured from phage infection and became capable of adhering, Spontaneous mutants resistant to coliphage T7, as well as K. pneumoniae K59-sensitive cells preadsorbed with inactivated T7 particles, did not adhere to epithelial cells. All strains capable of adhering were able to adsorb coliphage T7 and T3, whereas all nonadhesive strains were not. AP3-like prophages were induced from 7 of 12 nonadhesive Klebsiella strains. A laboratory strain which was able to adhere was lysogenized with 2 of these phages. In both cases, the strain lost its ability to adsorb coliphages T3, T7, and phi I and to adhere to human epithelial cells. All K. pneumoniae adhesive strains agglutinated yeast cells, whereas the nonadhesive strains did not. Competition studies have shown that D-mannose and concanavalin A prevented adherence to human epithelial cells, yeast agglutination, and adsorption of coliphage T7 to K. pneumoniae cells. It is concluded that in K. pneumoniae adherence to epithelial cells is mediated by the receptor for coliphages T7 (and T3), which in turn recognizes D-mannose in the receptors it binds.

Cell Adhesion↗

PubMed1980

Control of cell septation by lateral wall extension in a pH-conditional morphology mutant of Klebsiella pneumoniae.

The pH-conditional morphology mutant of Klebsiella pneumoniae strain MirM7 grows as cocci at pH 7 and as rods at pH 5.8. The mutant has a high-level mecillinam resistance (50% lethal dose greater than 200 micrograms/ml) in both forms. When broth cultures of the rod-shaped mutant were grown with 0.7 microgram of mecillinam per ml, cells assumed a round shape and continued to divided at a higher rate than the untreated control. A MirM7 rod-shaped revertant (MirA12), when treated with the same antibiotic concentration, changed to coccal shape and stopped dividing. The penicillin-binding proteins (PBPs) of strains MirA12 and MirM7 were analyzed. K. pneumoniae had six major PBPs quite similar to those of Escherichia coli. No differences were seen in the PBPs of MirM7 cocci and rods and MirA12 cells. In particular, PBP2 was found to be present and similar in MirM7 rods and cocci and MirA12 cells. We suggest that that in gram-negative rods, a control mechanism exists which prevents further septation in the absence of lateral cell wall elongation. The unique behavior of MirM7 is due to the fact that the control mechanism is not active in this strain. This model allows us to explain the preservation of shape in bacterial rods under various conditions of growth and the mechanism of bacterial killing by mecillinam.

Amdinocillin↗

PubMed1980

Simplified lyogroup system, a new method for routine identification of staphylococci: description and comparison with three other methods.

A simplified system for routine identification of the six lyogroups of human staphylococci is described. The method is based on the determination of the lytic activity on five different test media and of the phosphatase activity of each isolate. A total of 689 staphylococci isolated from clinical material over a 6-month period were identified according to this simplified system. The same strains were identified in parallel using three other methods designed for routine identification of staphylococci: the Kloos and Schleifer abbreviated scheme (J Clin. Microbiol. 1:82-88, 1975) the scheme proposed by the Subcommittee on the Taxonomy of Staphylococci and Micrococci (Int. J. Syst. Bacteriol. 26:332-334, 1976), and the API STAPH micromethod. Isolated whose identification was in disagreement were identified on a taxonomic level according to the extensive classification scheme of Schleifer and Kloos. In addition, 42 reference staphylococci of known identity were reidentified using the simplified lyogroup system as well as the three other routine methods. The results of this comparative study was detailed and discussed. A primary advantage of the simplified lyogroup system is that it also provides separation of staphylococci from micrococci, which must be performed separately when the other methods are used. Other significant advantages concerned with reliability, speed, and practicality are shown.

Bacteriological Techniques↗

PubMed1979

Degradation of Escherichia coli DNA: evidence for limitation in vivo by protein X, the recA gene product.

DNA is more extensively degraded after it is damaged in recA mutants of E. coli than in wild type cells. All data presented here are consistent with the recA gene product, protein X, being an inhibitor of nalidixic acid induced degradation of the bulk DNA (but not of newly replicated DNA). Production of protein X also is correlated with appearance of various "S.O.S." repair functions. Evidence was obtained by comparing the rates of protein X synthesis and solubilization of uniformly-labeled DNA in intact cells, incubated in the presence of nalidixic acid. A set of mutants at the lexA locus produced protein X at different rates and degraded their DNA at rates which were inversely correlated to their rates of protein X production. A low concentration of rifampicin quite specifically inhibited protein X production by wild type E. coli, and allowed more rapid DNA degradation. After the DNA was damaged by the incubation of cells in the presence of nalidixic acid, cells preloaded with protein X degraded their DNA more slowly. We propose that protein X could protect DNA against degradation by binding to single-stranded regions, thereby inhibiting nuclease action.

Bacterial Proteins↗

PubMed1979

Effects of Staphylococcus aureus lysozyme on human fibroblasts.

The purified lysozyme excreted by Staphylococcus aureus strains promotes elongation and spreading on plastic surfaces and stimulates DNA synthesis of human fibroblasts (WI38). This enzyme also raises twofold the saturation density level of cultures of these cells. It is suggested that the primitive and main effect of lysozyme on fibroblasts is the triggering of morphogenesis.

Cell Division↗

PubMed1979

Peptidoglycan synthesis in cocci and rods of a pH-dependent, morphologically conditional mutant of Klebsiella pneumoniae.

Mir M7 is a spontaneous morphologically conditional mutant of Klebsiella pneumoniae which grows as round cells (cocci) at pH 7 and as normal rods at pH 5.8. We studied the rates of peptidoglycan synthesis of cocci and rods growing at pH values of 7 and 5.8, respectively. It was found that exponentially growing cocci produced a reduced amount of peptidoglycan per cell, compared with rods. Moreover, a shift of cocci to the permissive pH (5.8) caused an increase in the rate of peptidoglycan synthesis, whereas the reverse shift of rods to pH 7 determined a twofold reduction in the rate of [(3)H]diaminopimelic acid incorporation. During synchronous growth at pH 7, the rate of peptidoglycan synthesis after cell division decreased with time and rose before and during the first division. The susceptibilities of rods and cocci to beta-lactam antibiotics were also studied. It was found that cocci were more sensitive both to penicillin G and to cephalexin than were rods, but they showed a high level of resistance to mecillinam. The peculiar behavior of this mutant was interpreted as supporting the existence in bacterial rods of two different sites for peptidoglycan synthesis: one responsible for lateral wall elongation and one responsible for septum formation. In Mir M7, shape damage is described as dependent on the specific inhibition, at the nonpermissive pH, of the site for lateral wall extension.

Amdinocillin↗