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C M Verduin

Publications and source records attributed to C M Verduin.

11 recordsLinked to original sources
PubMed2001

Cellular membrane associated mucins in artificial urine as mediators of crystal adhesion: an in vitro enterocystoplasty model.

PURPOSE: Cells are a major component of mucus in enterocystoplasties. We evaluate the role of secreted mucins on cells and cellular membranes as crystal adhesion and agglomeration mediators in artificial urine infected with Proteus mirabilis. MATERIALS AND METHODS: Five human intestinal cell lines, HT29, HT29-18N2, HT29-FU, HT29-MTX, Caco-2 and 1 ureter cell line, SV-HUC-1, were incubated for 3 hours in artificial urine with P. mirabilis (ATCC49565) in monolayer and scraped conditions. We isolated Triton X-100 soluble membrane proteins from cells to evaluate the effect of MUC2 and MUC 5AC as membrane associated proteins on crystal formation and crystal adherence. Scanning electron microscopy, light microscopy, Coulter Counter measurements and x-ray microanalysis were used to evaluate crystal formation. RESULTS: Brushite crystals were adhered to cellular surface sites rich in sulfur as crystal agglomerates. Smaller and more numerous crystals were observed in the presence of scraped cells. Crystal growth and agglomeration was inhibited by the presence of MUC 5AC, whereas MUC2 had the opposite effect. Both are present on cellular membranes and are rich in sulfur. Cellular invasion by bacteria occurred in all cell lines. CONCLUSIONS: Membrane associated cellular secretions such as MUC2 and 5AC are important crystal adhesion molecules on cells and have a clear effect on urease induced crystallization in vitro. MUC2 and MUC 5AC induce crystal adhesion and mucin type dependent effects on crystal agglomeration. The effects of MUC2 and MUC 5AC may explain the high incidence of bladder calculi in enterocystoplasties and emphasize the role of cellular surfaces in urine.

Cell Adhesion↗
PubMed2000

Complement-resistant Moraxella catarrhalis forms a genetically distinct lineage within the species.

Moraxella catarrhalis is a bacterial species that has been implicated in 15-20% of all cases of otitis media in the USA and the complement-resistant variant of M. catarrhalis has been considered particularly pathogenic. A collection of geographically diverse, complement-sensitive (n=28) and -resistant strains (n=47) of M. catarrhalis was assembled in order to analyse the bacterial population structure. All strains were identified as M. catarrhalis by conventional microbiological and biochemical methods. Amplification of the small subunit (ssu) ribosomal RNA gene followed by restriction fragment length polymorphism (RFLP) analysis did not reveal consistent differences between serum-susceptible and -resistant M. catarrhalis isolates. Interestingly, upon automated ribotyping using the Qualicon RiboPrinter(R) microbial characterisation system, the complement-sensitive and -resistant strains segregated into two groups. This suggested the existence of two clearly distinguishable lineages within the species M. catarrhalis. This observation was corroborated by pulsed field gel electrophoresis (PFGE) of DNA macro-restriction fragments, a non-ribosomal PCR RFLP procedure and random amplification of polymorphic DNA (RAPD) analysis. All procedures grouped the two variants similarly. Redefinition of the taxonomic status of complement-resistant M. catarrhalis or even the definition of a new species may be opportune.

Bacterial Typing Techniques↗
PubMed2000

Distinguishing species of the Burkholderia cepacia complex and Burkholderia gladioli by automated ribotyping.

Several species belonging to the genus Burkholderia are clinically relevant, opportunistic pathogens that inhabit major environmental reservoirs. Consequently, the availability of means for adequate identification and epidemiological characterization of individual environmental or clinical isolates is mandatory. In the present communication we describe the use of the Riboprinter microbial characterization system (Qualicon, Warwick, United Kingdom) for automated ribotyping of 104 strains of Burkholderia species from diverse sources, including several publicly accessible collections. The main outcome of this analysis was that all strains were typeable and that strains of Burkholderia gladioli and of each species of the B. cepacia complex, including B. multivorans, B. stabilis, and B. vietnamiensis, were effectively discriminated. Furthermore, different ribotypes were discerned within each species. Ribotyping results were in general agreement with strain classification based on restriction fragment analysis of 16S ribosomal amplicons, but the resolution of ribotyping was much higher. This enabled automated molecular typing below the species level. Cluster analysis of the patterns obtained by ribotyping (riboprints) showed that within B. gladioli, B. multivorans, and B. cepacia genomovar VI, the different riboprints identified always clustered together. Riboprints of B. cepacia genomovars I and III, B. stabilis, and B. vietnamiensis did not show distinct clustering but rather exhibited the formation of loose assemblages within which several smaller, genomovar-specific clusters were delineated. Therefore, ribotyping proved useful for genomovar identification. Analysis of serial isolates from individual patients demonstrated that infection with a single ribotype had occurred, despite minor genetic differences that were detected by pulsed-field gel electrophoresis of DNA macrorestriction fragments. The automated approach allows very rapid and reliable identification and epidemiological characterization of strains and generates an easily manageable database suited for expansion with information on additional bacterial isolates.

Automation↗
PubMed1999

Identification of Burkholderia spp. in the clinical microbiology laboratory: comparison of conventional and molecular methods.

Cystic fibrosis (CF) predisposes patients to bacterial colonization and infection of the lower airways. Several species belonging to the genus Burkholderia are potential CF-related pathogens, but microbiological identification may be complicated. This situation is not in the least due to the poorly defined taxonomic status of these bacteria, and further validation of the available diagnostic assays is required. A total of 114 geographically diverse bacterial isolates, previously identified in reference laboratories as Burkholderia cepacia (n = 51), B. gladioli (n = 14), Ralstonia pickettii (n = 6), B. multivorans (n = 2), Stenotrophomonas maltophilia (n = 3), and Pseudomonas aeruginosa (n = 11), were collected from environmental, clinical, and reference sources. In addition, 27 clinical isolates putatively identified as Burkholderia spp. were recovered from the sputum of Dutch CF patients. All isolates were used to evaluate the accuracy of two selective growth media, four systems for biochemical identification (API 20NE, Vitek GNI, Vitek NFC, and MicroScan), and three different PCR-based assays. The PCR assays amplify different parts of the ribosomal DNA operon, either alone or in combination with cleavage by various restriction enzymes (PCR-restriction fragment length polymorphism [RFLP] analysis). The best system for the biochemical identification of B. cepacia appeared to be the API 20NE test. None of the biochemical assays successfully grouped the B. gladioli strains. The PCR-RFLP method appeared to be the optimal method for accurate nucleic acid-mediated identification of the different Burkholderia spp. With this method, B. gladioli was also reliably classified in a separate group. For the laboratory diagnosis of B. cepacia, we recommend parallel cultures on blood agar medium and selective agar plates. Further identification of colonies with a Burkholderia phenotype should be performed with the API 20NE test. For final confirmation of species identities, PCR amplification of the small-subunit rRNA gene followed by RFLP analysis with various enzymes is recommended.

Bacterial Typing Techniques↗
PubMed1996

Moraxella catarrhalis in acute laryngitis: infection or colonization?

The complement phenotypes of Moraxella catarrhalis isolates obtained from adult patients with acute laryngitis were investigated using a microliter serum bactericidal assay and compared with those of other donor groups. Laryngitis isolates had a higher proportion (57%) of complement-resistant strains than did carrier strains from healthy 8- to 13-year-old schoolchildren (16%). The difference between these groups was statistically significant (chi2 [3 x 2 table] = 21.55; P < .001). The relatively frequent occurrence of the complement-resistant (virulence-associated) phenotype in adults with acute laryngitis supports the theory of an active role of M. catarrhalis in the pathogenesis of acute laryngitis.

Adolescent↗
PubMed1995

Complement resistance is a virulence factor of Branhamella (Moraxella) catarrhalis.

The purpose of this study was to investigate complement resistance in Branhamella (Moraxella) catarrhalis isolated from healthy schoolchildren or sputum-producing adult patients. Two techniques were used: a serum bactericidal assay as the gold standard and an easier 'culture and spot' test. Children (age 4-13; n = 303) and patients (n = 1047) showed high colonization/infection rates with B. catarrhalis (31% and 19%, respectively). Complement resistance or intermediate sensitivity occurred frequently in patient isolates (62% and 27%, respectively) and less often in children (33% and 8.5%, respectively; P << 0.0001). In young children (age 4-5 years), the proportion of complement-resistant strains was around 50%. Complement resistance in B. catarrhalis is associated with illness and may hence be considered a virulence factor.

Adolescent↗
PubMed1995

Assessment of complement-mediated killing of Moraxella (Branhamella) catarrhalis isolates by a simple method.

Recently, we showed that complement resistance is an important virulence factor of Moraxella (Branhamella) catarrhalis. Our study used a serum bactericidal assay to determine complement resistance in M. catarrhalis. Although the serum bactericidal assay is considered the "gold standard" for determining complement resistance, it is laborious and time-consuming and therefore not well suited for large-scale studies. Using a large number (n = 324) of M. catarrhalis isolates obtained from the sputa of patients with lower respiratory tract infections (n = 200) and young carriers (n = 124), we assessed the value of a simple "culture-and-spot" test as an alternative to the serum bactericidal assay. For both groups of isolates, the degree of concordance between the two tests used was very significant (P < 0.0001). The agreement between the two assays was estimated to be "excellent beyond chance" (as determined by Cohen's kappa test). The culture-and-spot assay is a valuable alternative to the serum bactericidal assay, not only for screening purposes as shown here but also for studying the mechanism of complement resistance in M. catarrhalis at the molecular level.

Bacteriological Techniques↗
PubMed1994

Experimental evidence for Moraxella-induced penicillin neutralization in pneumococcal pneumonia.

Resistance of microorganisms to antimicrobial agents is an increasing problem in the treatment of infectious diseases. In mixed infections, an interesting development can arise when one organism protects another from being killed by an antibiotic. Unfortunately, in the case of respiratory tract infections, experimental evidence of this development is poor. In this study, mice intranasally infected with a lethal number of pneumococci and treated with a curative dose of penicillin or amoxicillin died from pneumococcal pneumonia when they were coinoculated with beta-lactamase-producing Moraxella catarrhalis. beta-lactamase-negative M. catarrhalis did not show a similar indirect pathogenic effect. Treatment with a combination of amoxicillin and the beta-lactamase inhibitor clavulanic acid was not affected by beta-lactamase-producing M. catarrhalis. These findings help explain antibiotic failure in respiratory tract infections, even though the causative microorganism is sensitive to the antibiotic in vitro.

Amoxicillin↗
PubMed1994

Differences in complement activation between complement-resistant and complement-sensitive Moraxella (Branhamella) catarrhalis strains occur at the level of membrane attack complex formation.

The mechanism of resistance to human complement-mediated killing in Moraxella catarrhalis was studied by comparing different complement-sensitive and complement-resistant M. catarrhalis strains in a functional bystander hemolysis assay and an enzyme-linked immunosorbent assay (ELISA) for soluble terminal complement complexes. Complement-resistant stains appeared to activate complement to the same extent as, or even slightly better than, complement-sensitive strains. This indicates that complement-resistant strains do not inhibit classical or alternative pathway activation but interfere with complement at the level of membrane attack complex formation. A clear difference in dose-response curves for resistant and sensitive strains was observed both in the bystander hemolysis assay and in the ELISA. Complement-resistant strains showed optimum curves, whereas complement-sensitive strains gave almost linear curves. We conclude that resistant strains bind and/or inactivate one of the terminal complement components or intermediates involved in membrane attack complex formation. Trypsin, known to abolish complement resistance, changed the optimum dose-response curve of a resistant strain to a linear one, which strongly suggests that complement resistance is mediated by an M. catarrhalis-associated protein. This protein acts directly or through the binding of a terminal complement inhibitor present in serum.

Bacterial Proteins↗