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R Ramphal

Publications and source records attributed to R Ramphal.

At least 55 records · Page 3Linked to original sources
PubMed1994

Pseudomonas aeruginosa outer membrane adhesins for human respiratory mucus glycoproteins.

The attachment of Pseudomonas aeruginosa to human respiratory mucus represents an important step in the development of lung infection, especially in cases of cystic fibrosis. For this purpose, microtiter plate adhesion assays have been developed and have suggested that nonpilus adhesins of P. aeruginosa are the most important ones for binding to human respiratory mucins. In order to characterize these mucin-binding adhesins, outer membrane proteins (OMP) from two adhesive strains, 1244-NP and PAK-NP, and their poorly adhesive rpoN mutants, 1244-N3 and PAK-N1, were prepared by a mild extraction with Zwittergent 3-14. Mucin-binding adhesins were detected after polyacrylamide gel electrophoresis and blotting of the OMP on nitrocellulose replicas, using human bronchial mucins labeled with 125I. The binding properties of these OMP with lactotransferrin, another glycoprotein abundant in respiratory mucus, were also studied. Radiolabeled mucins detected four bands at 48, 46, 28, and 25 kDa with strain PAK-NP. With the nonmucoid strain 1244-NP, five bands were observed at 48, 46, 42, 28, and 25 kDa. The bands at 48 and 25 kDa were also visualized by radiolabeled lactotransferrin. These bands were partially or completely displaced by nonradiolabeled respiratory mucin glycopeptides but not by tetramethylurea, suggesting that they recognized carbohydrate sites. In contrast, the poorly adhesive strains showed weakly binding bands. These results demonstrate that outer membranes from two different nonpiliated P. aeruginosa strains express multiple adhesins with an affinity for human respiratory mucins and/or lactotransferrin.

Adhesins, Bacterial↗
PubMed1993

Antibiotic therapy for enterobacter meningitis: a retrospective review of 13 episodes and review of the literature.

Enterobacter meningitis is an uncommon form of meningitis whose treatment poses a therapeutic dilemma because of the development of resistance to the third-generation cephalosporins while the patient receives therapy. In recent years, we have been using trimethoprim-sulfamethoxazole (TMP-SMZ) as treatment for this infection. In this report, we reviewed 13 episodes of enterobacter meningitis that were treated with various antibiotic regimens and 33 episodes from the literature. We found that the development of resistance to beta-lactam agents may be much higher than that seen in bacteremias (approximately 30%), that the case-fatality rate is lower among our patients than among those described previously, and that all patients who received TMP-SMZ were cured, compared with about 70% of those receiving beta-lactam agents. TMP-SMZ appears to be an acceptable alternative to the cephalosporins for the treatment of enterobacter meningitis.

4-Quinolones↗
PubMed1993

Altered carbohydrate composition of salivary mucins from patients with cystic fibrosis and the adhesion of Pseudomonas aeruginosa.

We compared the chemical composition of salivary mucin glycopeptides from cystic fibrosis (CF) and from non-CF subjects and the adhesion of Pseudomonas aeruginosa to these different salivary glycopeptides. Three pools of CF saliva, four pools of non-CF saliva, one individual CF saliva, and one individual non-CF saliva were studied. The soluble fraction of the saliva was treated with pronase, and gel filtration was performed to obtain high and low molecular mass salivary mucin glycopeptides. The yield of total glycopeptides was significantly higher from CF than from non-CF saliva. Furthermore, the chemical composition revealed a significantly higher sialic acid content in CF than in non-CF mucin glycopeptides, and higher sulfate and fucose content in CF than in non-CF high molecular mass glycopeptides. We studied the adhesion of a nonmucoid strain of P. aeruginosa (1244), its nonpiliated isogenic derivative, and a mucoid strain (M35) to salivary mucin glycopeptides from patients with CF and from non-CF subjects. The three strains bound significantly more to the CF salivary glycopeptides than to the corresponding non-CF salivary glycopeptides. The nonpiliated isogenic mutant of P. aeruginosa 1244 also bound to CF salivary glycopeptides, suggesting that the adhesion of P. aeruginosa could involve nonpilus adhesions. Furthermore, neuraminidase treatment of CF glycopeptides decreased the adhesion of P. aeruginosa 1244. Altogether these results suggested that differences in mucins may in part explain the specificity of P. aeruginosa for CF.

Bacterial Adhesion↗
PubMed1992

Vancomycin is not an essential component of the initial empiric treatment regimen for febrile neutropenic patients receiving ceftazidime: a randomized prospective study.

The use of vancomycin as part of the initial antibiotic therapy of febrile neutropenic patients has become a controversial issue. Some studies support its incorporation in the initial regimen, and others suggest that vancomycin can be added later. We examined this issue in a prospective, randomized trial. We randomized 127 febrile neutropenic patients to receive either ceftazidime alone or ceftazidime plus vancomycin as the initial empiric antibiotic treatment. We added vancomycin to the ceftazidime arm of the study when fever persisted after 96 h of monotherapy, when new fever occurred after this time, or when a moderately ceftazidime-resistant gram-positive bacterium was isolated. Each of these regimens had similar initial response rates, similar durations of initial fever, similar frequencies of new fever during therapy, similar microbiological cure rates, similar superinfection rates, and similar survival rates. We observed more renal and cutaneous toxicities in patients receiving vancomycin and ceftazidime as initial therapy. We conclude that ceftazidime is appropriate as initial therapy for febrile neutropenic patients and that the addition of vancomycin is appropriate when fever persists after 4 days of monotherapy or when fever recurs following an initial response.

Adolescent↗
PubMed1992

Genetic analysis of Pseudomonas aeruginosa adherence: distinct genetic loci control attachment to epithelial cells and mucins.

Infection of mucosal tissues by the opportunistic pathogen Pseudomonas aeruginosa is initiated by attachment of the bacterium to host tissues. To gain a better understanding of this interaction, we used two methods to isolate mutants of P. aeruginosa with altered adherence to cultured A549 cells and to mucins. First, from a population of nonpiliated mutants of P. aeruginosa mutagenized with transposon Tn5G, we have isolated variants that are defective in binding to both A549 cells and respiratory mucins. Using a cloned transposon plus flanking DNA from one such mutant as a DNA probe, we have isolated plasmids from a cosmid bank, which, upon reintroduction to the original mutants, restored adhesion to both A549 cells and mucin. The second strategy to identify genes involved in adhesion used mutagenesis of P. aeruginosa N1G, an rpoN mutant which is unable to bind to either A549 cells or mucin, with transposon Tn5 containing an outward-directed promoter. From this bank of mutagenized P. aeruginosa N1G, two classes of adhesion variants were isolated; one class attached to A549 cells and to mucin, and the other class restored binding of the rpoN mutant to mucin but not to A549 cells. These findings suggest that P. aeruginosa can express at least two adhesins distinct from pili, one recognizing receptors shared by epithelial cells and mucins and the other recognizing mucins alone.

Bacterial Adhesion↗
PubMed1991

Enterobacter bacteremia: clinical features and emergence of antibiotic resistance during therapy.

OBJECTIVES: To study the effect of previously administered antibiotics on the antibiotic susceptibility profile of Enterobacter, the factors affecting mortality, and the emergence of antibiotic resistance during therapy for Enterobacter bacteremia. DESIGN: Prospective, observational study of consecutive patients with Enterobacter bacteremia. SETTING: Three university tertiary care centers, one major university-affiliated hospital, and two university-affiliated Veterans Affairs medical centers. PATIENTS: A total of 129 adult patients were studied. MEASUREMENTS: The two main end points were emergence of resistance during antibiotic therapy and death. MAIN RESULTS: Previous administration of third-generation cephalosporins was more likely to be associated with multiresistant Enterobacter isolates in an initial, positive blood culture (22 of 32, 69%) than was administration of antibiotics that did not include a third-generation cephalosporin (14 of 71, 20%; P less than 0.001). Isolation of multiresistant Enterobacter sp. in the initial blood culture was associated with a higher mortality rate (12 of 37, 32%) than was isolation of a more sensitive Enterobacter sp. (14 of 92, 15%; P = 0.03). Emergence of resistance to third-generation cephalosporin therapy (6 of 31, 19%) occurred more often than did emergence of resistance to aminoglycoside (1 of 89, 0.01%; P = 0.001) or other beta-lactam (0 of 50; P = 0.002) therapy. CONCLUSIONS: More judicious use of third-generation cephalosporins may decrease the incidence of nosocomial multiresistant Enterobacter spp., which in turn may result in a lower mortality for Enterobacter bacteremia. When Enterobacter organisms are isolated from blood, it may be prudent to avoid third-generation cephalosporin therapy regardless of in-vitro susceptibility.

Adolescent↗
PubMed1991

Arylneuraminidase activity of Pseudomonas aeruginosa does not degrade natural substrates such as human respiratory mucins.

The culture supernatant from a single Pseudomonas aeruginosa strain has been reported to show neuraminidase activity, leading to the speculation that this bacterium may use this enzyme as a virulence factor to act on host macromolecules. In order to extend this finding, we have examined the activity of concentrated P. aeruginosa culture supernatants and cells on synthetic and natural substrates containing sialic acid, such as human respiratory mucins. Four P. aeruginosa strains showed some activity on the synthetic substrate 4-methylumbelliferyl-alpha-D-N-acetylneuraminic acid but failed to liberate N-acetylneuraminic acid from six different natural substrates. Attempts to induce enzyme production by use of human respiratory mucins in the culture medium were also unsuccessful. The supernatants also showed N-acetyl-beta-D-glucosaminidase-like activity on a synthetic substrate but did not liberate N-acetylhexosamines from natural substrates. We conclude that the neuraminidase-like activity observed in P. aeruginosa can be defined as an arylneuraminidase and that the possession of a neuraminidase active on natural substrates is not a common attribute of P. aeruginosa strains.

Extracellular Space↗
PubMed1991

Pseudomonas aeruginosa recognizes carbohydrate chains containing type 1 (Gal beta 1-3GlcNAc) or type 2 (Gal beta 1-4GlcNAc) disaccharide units.

The adhesion of Pseudomonas aeruginosa to type 1 (Gal beta 1-3GlcNAc) and type 2 (Gal beta 1-4GlcNAc) disaccharide determinants was studied in a microtiter adhesion assay and a thin-layer chromatography bacterial overlay assay. The oligosaccharides were prepared from human breast milk and human urine and were conjugated to hexadecylaniline to form neoglycolipids that were used in were used in the assays. Both the mucoid and the nonmucoid strains that were studied recognized the disaccharide determinants Sialylation of the oligosaccharides did not suppress binding in the thin-layer chromatography assay, but alpha 2-6-linked sialic acid blocked binding in the microtiter assay. The use of bovine serum albumin instead of gelatin as a blocking agent against nonspecific binding completely suppressed binding in the thin-layer chromatography assay. Isogenic nonpiliated mutants of nonmucoid strains constructed by interrupting the pilin gene retained their adhesive capacity for the disaccharide units, indicating that binding to the disaccharides was mediated by a nonpilus adhesin(s). Furthermore, two monoclonal antibodies that recognize the type 2 disaccharide determinant (Gal beta 1-4GlcNAc) partially inhibited adhesion of a pair of piliated and nonpiliated isogenic strains to mucin. This study suggests that P. aeruginosa utilizes a nonpilus adhesin(s) to bind to disaccharide units commonly found in mucins, in addition to pili and alginate, two previously described adhesins.

Amino Sugars↗
PubMed1991

Adhesion of Pseudomonas aeruginosa pilin-deficient mutants to mucin.

Attachment of Pseudomonas aeruginosa to epithelial cells or tracheobronchial mucin is mediated by surface adhesins. Pili, composed of monomeric pilin subunits, make up one such class of adhesins. The formation of pili and flagella in P. aeruginosa is under the control of the alternative sigma factor rpoN. Isogenic mutant strains with insertionally inactivated rpoN genes were constructed with strains PAK, 1244, and CF613 and were tested for their ability to adhere to respiratory mucin. All rpoN mutants showed significant reduction of adherence to mucin relative to that of their wild-type parents. In contrast, the adherence of pilin structural gene mutants was similar to the adherence of wild types. These results provide suggestive evidence that P. aeruginosa also binds to mucin via adhesins that are distinct from pilin and are still under the genetic control of rpoN. Unlike the laboratory strain PAK, the clinical strains 1244 and CF613 are capable of agglutinating erythrocytes. The rpoN mutation had a minimal effect on the interaction of bacteria with erythrocytes, indicating that the transcription of a gene(s) specifying the agglutination phenomenon does not utilize rpoN. These findings collectively indicate the existence of several classes of adhesins on the surface of P. aeruginosa that may play an important role in colonization of the human respiratory tract.

Bacterial Adhesion↗
PubMed1991

The carbohydrate diversity of human respiratory mucins: a protection of the underlying mucosa?

Human respiratory mucins consist of a family of glycoproteins with different peptides in which glycosylation, the major post-translational phenomenon, is responsible for about 70 to 80% of the weight of these molecules. This glycosylation generates a remarkable diversity of O-glycosidically linked carbohydrate chains, which are expressed as several hundreds of different chains in a single person. These chains, which can vary from one to about 20 sugars, may be neutral, sialylated, or sulfated. They bear multiple epitopes. Some antigenic determinants such as ABO, Leb antigens in secretor individuals, Lea, or X or Y antigens have been identified. There is increasing evidence that, among other functions, this diversity of chains allows many interactions with microorganisms and may be an important factor in maintaining the sterility of the respiratory tree. In certain pathologic situations such as cystic fibrosis, which is associated with colonization by Pseudomonas aeruginosa, the hypothesis of an alteration of this interaction is open.

Carbohydrate Sequence↗
PubMed1990

Beta-lactam-resistant Enterobacter bacteremia in febrile neutropenic patients receiving monotherapy.

Bacteremia with resistant Enterobacter species has been reported in febrile, neutropenic cancer patients receiving beta-lactam antibiotics. To assess the relationship between enterobacter bacteremia and ceftazidime monotherapy, medical records were reviewed and isolates were tested from 16 neutropenic and 35 nonneutropenic patients with Enterobacter bacteremia. Fifteen isolates from the neutropenic patients were resistant to extended spectrum cephalosporins; only 12 of 35 isolates from the nonneutropenic patients were resistant to Enterobacter species. The neutropenic patients also had more beta-lactam therapy, both immediately before bacteremia and in the preceding year, than did nonneutropenic patients. Prior beta-lactam antibiotic exposure may predispose neutropenic patients to develop resistant Enterobacter bacteremia.

Adolescent↗
PubMed1989

Evidence for the in vivo degradation of human respiratory mucins during Pseudomonas aeruginosa infection.

The comparison of distribution of glycopeptides of sputa from patients suffering from various chronic hypersecretions has already shown an increased acidity with a decreased proportion of neutral glycopeptides in the respiratory secretions of patients suffering from cystic fibrosis, as compared to those of patients with chronic bronchitis. In order to find out whether this decrease is specific to cystic fibrosis mucins or whether it is due to a degradation of mucus by Pseudomonas aeruginosa, which infects most of the sputa from patients with this disease, mucus glycopeptides from patients with different chronic bronchial disorders, infected by Pseudomonas or not, were prepared and fractionated by ion-exchange chromatography. The neutral fraction, which has never been studied in detail, was gel-filtered, and provided two fractions, one containing true mucin glycopeptides and the other containing a mixture of peptides and glycopeptides with a lower molecular mass. In the Pseudomonas-infected samples, the true mucin glycopeptide fraction was greatly diminished as compared to this same fraction in non-Pseudomonas-infected samples; this was not specific to cystic fibrosis secretions. In contrast, the glycopeptide fraction with a lower molecular mass was greatly increased in all the Pseudomonas-infected samples. Polyacrylamide gel electrophoresis of this second fraction showed unique glycopeptide bands between 40-50 kDa in the Pseudomonas-infected samples, regardless of the origin of the samples. These bands were revealed by an antibody directed against whole cystic fibrosis mucin. Infected chronic bronchitis sputa and cystic fibrosis samples without P. aeruginosa did not show these bands. These studies therefore suggest that there are P. aeruginosa-associated changes in mucins which may result from degradation of mucins.

Amino Acids↗
PubMed1989

Differences in adhesion of Pseudomonas aeruginosa to mucin glycopeptides from sputa of patients with cystic fibrosis and chronic bronchitis.

Pseudomonas aeruginosa is the most prominent colonizer of the respiratory tract of patients with cystic fibrosis, but it is not known why this occurs. P. aeruginosa adheres to mucins from normal individuals, but mucins from cystic fibrosis patients have not been studied. To compare adhesion to mucins from cystic fibrosis with other mucins, we prepared highly glycosylated mucin glycopeptides from cystic fibrosis and chronic bronchitis patients by ion-exchange and gel-filtration chromatography and measured the adhesion of P. aeruginosa 1244 to these glycopeptides. We found (i) that the most mucinlike glycopeptides from P. aeruginosa-infected cystic fibrosis sputa showed less bacterial adhesion than did the corresponding bronchitis samples, (ii) that the most adhesive activity in cystic fibrosis samples came from a fraction that contains O and N glycopeptides and may be in part a degradation product of P. aeruginosa infection, and (iii) that highly glycosylated glycopeptides of the most acidic species (sialylated and sulfated) showed no adhesion at all. A single cystic fibrosis sample not infected by P. aeruginosa showed better binding in the adhesion-positive fractions than did the infected sputa. These studies suggest that cystic fibrosis mucins may be altered after infection is established, resulting in less binding to some fragments. However, since the clinical picture shows heavy mucus colonization, other receptors, such as cellular glycolipids which have been shed into mucus, may be contributing to this colonization.

Bacterial Adhesion↗
PubMed1988

The binding of anti-pseudomonal antibiotics to macromolecules from cystic fibrosis sputum.

Antibiotics are known to bind to whole cystic fibrosis sputum. However, the composition of sputum varies from one patient to another, making the interpretation of binding studies difficult. This problem has been examined by standardising the macromolecule concentration of sputum from four cystic fibrosis patients and adding tobramycin or ceftazidime directly to the sputum components. Binding to mucin-rich and DNA-rich fractions of sputum was also studied before and after DNase treatment of these fractions. These studies indicated that (i) the degree of tobramycin binding is dependent on the sputum macromolecule concentration, (ii) a significant proportion of tobramycin is bound even at concentrations of 100 mg/l of drug, (iii) tobramycin binds to both the mucin rich fraction and the DNA rich fraction of sputum and (iv) ceftazidime binding to sputum is negligible. Our data indicate that there is a need to standardise sputum in antibiotic binding studies and they provide another rationale for favouring the use of ceftazidime over aminoglycosides in infectious exacerbations of cystic fibrosis caused by Pseudomonas aeruginosa.

Ceftazidime↗
PubMed1988

Respiratory-mucin inhibition of the opsonophagocytic killing of Pseudomonas aeruginosa.

Pseudomonas aeruginosa is a frequent respiratory tract colonizer in diseases in which mucociliary clearance is defective. The most striking of these is cystic fibrosis. The reasons for this organism's ability to colonize the respiratory tract and to persist there are not fully understood. Earlier studies showed that P. aeruginosa adheres preferentially to tracheobronchial mucin when compared with enterobacteria. We reasoned that if adherence to respiratory mucin protected P. aeruginosa from opsonophagocytic killing, then the ability of this organism to chronically colonize the respiratory tract could be partially explained. Using an opsonophagocytic killing assay with human polymorphonuclear leukocytes, we found that respiratory mucin protected six strains of P. aeruginosa from opsonophagocytic killing but did not protect poorly adhering strains of Escherichia coli, Staphylococcus aureus, or group B streptococci. Incubating P. aeruginosa with the mucin prior to addition to the opsonic assay inhibited phagocytic killing, whereas incubation of polymorphonuclear leukocytes with mucin did not, suggesting that inhibition was not due to an effect of mucin on leukocytes per se but was a consequence of bacterial adherence to mucin. Further studies indicated no decrease in the binding of either antibody or complement component C3 to the bacterial surface in the presence of mucin. This suggests that phagocytic inhibition may be due to a defect in uptake or destruction of mucin-coated bacteria by the leukocytes. Thus, the adherence of P. aeruginosa to respiratory mucin potentially contributes to its persistence in the respiratory tract by interfering with host immune responses.

Adult↗
PubMed1988

Passive serum antibody causes temporary recovery from influenza virus infection of the nose, trachea and lung of nude mice.

BALB/c normal and nude mice were infected with a non-lethal mouse-passaged A/PC/1/73 (H3N2) influenza virus in order to assess the role of T cells on the course of disease of the nose, trachea and lung. The tracheal epithelium of both mouse strains was desquamated by 3 days after infection. Although normal regeneration began, nude mice never completed that regeneration whereas normal mice had fully regenerated tracheas by Day 14. This failure to complete the recovery was also evident from the continued virus shedding by the nude mouse. In order to assess the role of serum antibody on recovery from infection, ferret, goat or mouse antibody to H3N2 influenza virus was passively administered to nude mice after infection. It resulted in a transient decrease in virus shedding from the nose, trachea and lung, and complete but temporary regeneration of the tracheal epithelium. However, later in the course of the infection, when serum antibody levels were no longer detectable, the tracheal epithelium of these animals redesquamated and large amounts of virus were again shed from nose, trachea and lungs. We conclude that: (i) desquamation of the ciliated epithelium of the trachea is not T-cell dependent; and (ii) serum antibody can contribute to temporary recovery from infection, but by itself is insufficient for permanent recovery of the nose, trachea or lung.

Animals↗