20 research outputs found
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
Fusidic acid cream in the treatment of impetigo in general practice: double blind randomised placebo controlled trial
OBJECTIVE: To test the hypothesis that fusidic acid would not increase the
treatment effect of disinfecting with povidone-iodine alone in children
with impetigo. DESIGN: Randomised placebo controlled trial. SETTING:
General practices in Greater Rotterdam. PARTICIPANTS: 184 children aged
0-12 years with impetigo. MAIN OUTCOME MEASURES: Clinical cure and
bacterial cure after one week. RESULTS: After one week of treatment 55% of
the patients in the fusidic acid group were clinically cured compared with
13% in the placebo group (odds ratio 12.6, 95% confidence interval 5.0 to
31.5, number needed to treat 2.3). After two weeks and four weeks the
differences in cure rates between the two groups had become smaller. More
children in the placebo group were non-compliant (12 v 5) and received
extra antibiotic treatment (11 v 3), and more children in the placebo
group reported adverse effects (19 v 7). Staphylococcus aureus was found
in 96% of the positive cultures; no strains were resistant to fusidic
acid. CONCLUSIONS: Fusidic acid is much more effective than placebo (when
both are given in combination with povidone-iodine shampoo) in the
treatment of impetigo. Because of the low rate of cure and high rate of
adverse events in the placebo group, the value of povidone-iodine in
impetigo can be questioned
Moraxella catarrhalis: from Emerging to Established Pathogen
Moraxella catarrhalis (formerly known as Branhamella catarrhalis) has
emerged as a significant bacterial pathogen of humans over the past two
decades. During this period, microbiological and molecular diagnostic
techniques have been developed and improved for M. catarrhalis, allowing
the adequate determination and taxonomic positioning of this pathogen.
Over the same period, studies have revealed its involvement in respiratory
(e.g., sinusitis, otitis media, bronchitis, and pneumonia) and ocular
infections in children and in laryngitis, bronchitis, and pneumonia in
adults. The development of (molecular) epidemiological tools has enabled
the national and international distribution of M. catarrhalis strains to
be established, and has allowed the monitoring of nosocomial infections
and the dynamics of carriage. Indeed, such monitoring has revealed an
increasing number of B-lactamase-positive M. catarrhalis isolates (now
well above 90%), underscoring the pathogenic potential of this organism.
Although a number of putative M. catarrhalis virulence factors have been
identified and described in detail, their relationship to actual bacterial
adhesion, invasion, complement resistance, etc. (and ultimately their role
in infection and immunity), has been established in a only few cases. In
the past 10 years, various animal models for the study of M. catarrhalis
pathogenicity have been described, although not all of these models are
equally suitable for the study of human infection. Techniques involving
the molecular manipulation of M. catarrhalis genes and antigens are also
advancing our knowledge of the host response to and pathogenesis of this
bacterial species in humans, as well as providing insights into possible
vaccine candidates. This review aims to outline our current knowledge of
M. catarrhalis, an organism that has evolved from an emerging to a
well-established human pathogen
Changes in genetic types and population dynamics of Moraxella catarrhalis in hospitalized children are not associated with an exacerbation of existing disease
Pulsed-field gel electrophoresis typing was performed on a retrospective
set of 129 Moraxella catarrhalis isolates obtained over a 20 month period
from 70 children admitted to, or presenting at, the Erasmus University
Medical Center, Rotterdam, The Netherlands. The mean age of the children
(at the end of the study) was 2.5 years, with a range of 6 months to 15
years. Fifty-one different M. catarrhalis types were isolated from the
hospitalized children, with 31 % (22/70) being infected with tw
Pathological and Therapeutic Significance of Cellular Invasion by Proteus mirabilis in an Enterocystoplasty Infection Stone Model
Proteus mirabilis infection often leads to stone formation. We evaluated how bacterium-mucin adhesion, invasion, and intracellular crystal formation are related to antibiotic sensitivity and may cause frequent stone formation in enterocystoplasties. Five intestinal (Caco-2, HT29, HT29-18N2, HT29-FU, and HT29-MTX) and one ureter cell line (SV-HUC-1) were incubated in artificial urine with five Proteus mirabilis strains. Fluorescence-activated cell sorting (FACS), laser scanning microscopy, and electron microscopy evaluated cellular adhesion and/or invasion, pathologic changes to mitochondria, and P. mirabilis-mucin colocalization (MUC2 and MUC5AC). An MTT (thiazolyl blue tetrazolium bromide) assay and FACS analysis of caspase-3 evaluated the cellular response. Infected cells were incubated with antibiotics at dosages representing the expected urinary concentrations in a 10-year-old, 30-kg child to evaluate bacterial invasion and survival. All cell lines showed colocalization of P. mirabilis with human colonic mucin (i.e., MUC2) and human gastric mucin (i.e., MUC5AC). The correlation between membrane mucin expression and invasion was significant and opposite for SV-HUC-1 and HT29-MTX. Microscopically, invasion by P. mirabilis with intracellular crystal formation and mitochondrial damage was found. Double membranes surrounded bacteria in intestinal cells. Relative resistance to cotrimoxazole and augmentin was found in the presence of epithelial cells. Ciprofloxacin and gentamicin remained effective. Membrane mucin expression was correlated with relative antibiotic resistance. Cell invasion by P. mirabilis and mucin- and cell type-related distribution and response differences indicate bacterial tropism that affects crystal formation and mucosal presence. Bacterial invasion seems to have cell type-dependent mechanisms and prolong bacterial survival in antibiotic therapy, giving a new target for therapeutic optimalization of antibiotic treatment
Coccidioidomycosis Infection of a Total Knee Arthroplasty in a Nonendemic Region
Fungal prosthetic joint infections are rare and difficult to treat. There is an ongoing discussion about the type and duration of antifungal treatment and the necessity of prosthesis removal. We report the first European case of an infected total knee arthroplasty with Coccidioides immitis. Treatment consisted of lifelong treatment with oral fluconazole at a dose of 400 mg/d, without total knee arthroplasty removal. After 6 months, the initial complaints of pain and swelling were completely resolved. This case report clearly states that a travel history and culturing for fungi are helpful in patients with persisting complaints after joint arthroplasty. (C) 2013 Elsevier Inc. All rights reserve
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