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M Altwegg

Publications and source records attributed to M Altwegg.

At least 19 recordsLinked to original sources
PubMed2006

Internal transcribed spacer sequencing versus biochemical profiling for identification of medically important yeasts.

In this study, we established an in-house database of yeast internal transcribed spacer (ITS) sequences. This database includes medically important as well as colonizing yeasts that frequently occur in the diagnostic laboratory. In a prospective study, we compared molecular identification with phenotypic identification by using the ID32C system (bioMérieux) for yeast strains that could not be identified by a combination of CHROMagar Candida and morphology on rice agar. In total, 113 yeast strains were included in the study. By sequence analysis, 98% of all strains were identified correctly to the species level. With the ID32C, 87% of all strains were identified correctly to the species or genus level, 7% of the isolates could not be identified, and 6% of the isolates were misidentified, most of them as Candida rugosa or Candida utilis. For a diagnostic algorithm, we suggest a three-step procedure which integrates morphological criteria, biochemical investigation, and sequence analysis of the ITS region.

Candida albicans↗
PubMed2006

16S rRNA gene sequencing versus the API 20 NE system and the VITEK 2 ID-GNB card for identification of nonfermenting Gram-negative bacteria in the clinical laboratory.

Over a period of 26 months, we have evaluated in a prospective fashion the use of 16S rRNA gene sequencing as a means of identifying clinically relevant isolates of nonfermenting gram-negative bacilli (non-Pseudomonas aeruginosa) in the microbiology laboratory. The study was designed to compare phenotypic with molecular identification. Results of molecular analyses were compared with two commercially available identification systems (API 20 NE, VITEK 2 fluorescent card; bioMérieux, Marcy l'Etoile, France). By 16S rRNA gene sequence analyses, 92% of the isolates were assigned to species level and 8% to genus level. Using API 20 NE, 54% of the isolates were assigned to species and 7% to genus level, and 39% of the isolates could not be discriminated at any taxonomic level. The respective numbers for VITEK 2 were 53%, 1%, and 46%, respectively. Fifteen percent and 43% of the isolates corresponded to species not included in the API 20 NE and VITEK 2 databases, respectively. We conclude that 16S rRNA gene sequencing is an effective means for the identification of clinically relevant nonfermenting gram-negative bacilli. Based on our experience, we propose an algorithm for proper identification of nonfermenting gram-negative bacilli in the diagnostic laboratory.

Bacterial Typing Techniques↗
PubMed2005

[The role of PCR for the diagnosis of Lyme arthritis].

OBJECTIVE, PATIENTS AND METHODS: We studied retrospectively four patients with Lyme arthritis of the knee, the role of PCR for the detection of B. burgdorferi DNA and its influence on further therapeutic decisions. RESULTS: All four patients with Lyme arthritis suffered from knee pain and effusions. None of them remembered having had a tick bite or an erythema migrans. The diagnosis was confirmed by positive serology and in three cases by detection of B. burgdorferi DNA by PCR analysis of the joint fluid. In one patient, PCR was also positive in the synovial tissue. Because of persistent symptoms after adequate antibiotic therapy, PCR was repeated in the joint fluid of two patients. In one patient a positive PCR suggested an ongoing infection. Thus, the antibiotic treatment was changed. A further PCR was negative. Symptoms resolved slowly in all patients over a time of two to seven months after the end of the antibiotic treatment. CONCLUSION: PCR to detect B. burgdorferi DNA in synovial fluid or tissue respectively is a helpful tool for the diagnosis of Lyme arthritis. Moreover, in patients with refractory Lyme arthritis PCR may be helpful in monitoring the course of the disease.

Adult↗
PubMed2005

Rapid detection of diarrheagenic E. coli by real-time PCR.

Enterovirulent Escherichia coli are among the most important causes of acute diarrhea in developing as well as in developed countries. We have adapted classical PCR to detect these organisms in stool specimens to real-time PCR using the LightCycler (LC) SYBR Green format followed by melting curve analysis. With only two different cycling protocols we could detect enteropathogenic E. coli (EPEC) and verocytotoxin-producing E. coli (VTEC) (duplex assay for both Verotoxin 1 (VT1) and Verotoxin 2 (VT2)) in one run and enteroaggregative E. coli (EAEC), enteroinvasive E. coli (EIEC) and enterotoxigenic E. coli (ETEC) (duplex assay detecting both heat-stable enterotoxin (ST) and heat-labile enterotoxin (LT)) in another run. Using serial dilutions of control strains, the LC proved to be clearly more sensitive than conventional PCR for five out of seven investigated targets: VTEC (VT1 and VT2), ETEC (ST and LT) and EIEC. For EPEC and EAEC, LC and conventional PCR had identical sensitivities. With stool samples, we found an optimal agreement between LC-PCR and the conventional PCR when samples were tested in a 1:10 dilution. Only one specimen was discrepant, being repetitively positive for VT by LightCycler but not by conventional PCR. Given the significantly higher sensitivity of the LC-PCR for the VT target (up to a 10(-4) dilution factor by melting curve analysis and up to a 10(-6) dilution factor following gel electrophoresis), this is probably a false negative result by conventional PCR. We conclude that LightCycler PCR is more rapid, easier than and at least as sensitive as our conventional PCR for the detection of enterovirulent E. coli in stool specimens after culture on MacConkey.

Bacterial Toxins↗
PubMed2005

Deadly carousel or difficult interpretation of new diagnostic tools for Whipple's disease: case report and review of the literature.

Whipple's disease is a rare systemic disorder classically presenting with weight loss, arthralgias, and diarrhea, which was first described in 1907. The causative bacterium Tropheryma whipplei, is a fastidious organism not growing on conventional media. Before the introduction of polymerase chain reaction (PCR)-based methods, the diagnostic gold standard was histological detection of diastase-resistant periodic acid Schiff (PAS)-positive macrophages or electron microscopy. As in the present case, contradictory results between the former and new diagnostic methods may obscure the correct diagnosis. We critically summarize the performance of the different diagnostic methods and discuss their impact on the clinical management of patients with suspected Whipple's disease.

Aged↗
PubMed2004

Comparison of conventional and molecular methods for identification of aerobic catalase-negative gram-positive cocci in the clinical laboratory.

Over a period of 18 months we have evaluated the use of 16S ribosomal DNA (rDNA) sequence analysis as a means of identifying aerobic catalase-negative gram-positive cocci in the clinical laboratory. A total of 171 clinically relevant strains were studied. The results of molecular analyses were compared with those obtained with a commercially available phenotypic identification system (API 20 Strep system; bioMérieux sa, Marcy l'Etoile, France). Phenotypic characterization identified 67 (39%) isolates to the species level and 32 (19%) to the genus level. Seventy-two (42%) isolates could not be discriminated at any taxonomic level. In comparison, 16S rDNA sequencing identified 138 (81%) isolates to the species level and 33 (19%) to the genus level. For 42 of 67 isolates assigned to a species with the API 20 Strep system, molecular analyses yielded discrepant results. Upon further analysis it was concluded that among the 42 isolates with discrepant results, 16S rDNA sequencing was correct for 32 isolates, the phenotypic identification was correct for 2 isolates, and the results for 8 isolates remained unresolved. We conclude that 16S rDNA sequencing is an effective means for the identification of aerobic catalase-negative gram-positive cocci. With the exception of Streptococcus pneumoniae and beta-hemolytic streptococci, we propose the use of 16S rDNA sequence analysis if adequate species identification is of concern.

Aerobiosis↗
PubMed2004

Three month treatment of reactive arthritis with azithromycin: a EULAR double blind, placebo controlled study.

OBJECTIVE: To determine the efficacy of weekly treatment with oral azithromycin for 13 weeks on the severity and resolution of reactive arthritis (ReA). METHODS: 186 patients from 12 countries were enrolled in a randomised, double blind, placebo controlled trial. Inclusion criteria were inflammatory arthritis of < or =6 swollen joints, and disease duration of < or =2 months. All patients received a single azithromycin dose (1 g) as conventional treatment for possible Chlamydia infection, and were then randomly allocated to receive weekly azithromycin or placebo. Clinical assessments were made at 4 week intervals for 24 weeks. RESULTS: 152 patients were analysable (34 failed entry criteria), with a mean (SD) age of 33.8 (9.4) and duration of symptoms 30.7 (17.5) days. Mean C reactive protein (CRP) was 48 mg/l, and approximately 50% of those typed were HLA-B27+, suggesting that the inclusion criteria successfully recruited patients with acute ReA. Treatment and placebo groups were well matched for baseline characteristics. There were no statistical differences for changes in any end point (swollen and tender joint count, joint pain, back pain, heel pain, physician and patient global assessments, and CRP) between the active treatment and placebo groups, analysed on an intention to treat basis or according to protocol completion. The time to resolution of arthritis and other symptoms or signs by life table analyses was also not significantly different. Adverse events were generally mild, but were more commonly reported in the azithromycin group. CONCLUSIONS: This large trial has demonstrated that prolonged treatment with azithromycin is ineffective in ReA.

Adolescent↗
PubMed2003

Spondylodiscitis as the first manifestation of Whipple's disease -a removal worker with chronic low back pain.

Whipple's disease is a rare systemic infectious disease caused by the actinobacterium Tropheryma whipplei. Spondylodiscitis is an extremely rare manifestation of the infection and has previously been described in only three case reports. We present a 55-year-old man with persistent lumbago and signs of systemic illness, but without any gastrointestinal symptoms or arthralgia. The signal response in the lumbar spine in magnetic resonance tomography, both native and after intravenous gadolinium administration, was compatible with spondylodiscitis at the L4/L5 level. Culture of a specimen obtained by radiographically guided disc puncture and repeated blood cultures remained sterile. Tropheryma whipplei was detected by PCR amplification in material obtained from the disc specimen, from a biopsy of the terminal ileum and from the stool. The histology of duodenum, terminal ileum, colon and disc material was normal and, in particular, showed no PAS-positive inclusions in macrophages. Long-term antibiotic treatment with sulphamethoxazole and trimethoprim was successful, with marked improvement of the low back pain and normalisation of the systemic inflammatory signs. The possibility of Whipple's disease must be suspected in the case of a 'culture-negative' spondylodiscitis even if there are no gastrointestinal symptoms and no arthralgia present.

Anti-Bacterial Agents↗
PubMed2003

Prevalence of Tropheryma whipplei DNA in patients with various gastrointestinal diseases and in healthy controls.

BACKGROUND: Little is known about the epidemiology of Tropheryma whipplei and its prevalence in people without clinical signs of Whipple's disease. PATIENTS AND METHODS: We screened 239 patients with various gastrointestinal diseases for T. whipplei DNA and compared them with 215 healthy controls in order to check whether T. whipplei might be a risk factor for common gastrointestinal problems or diseases. We detected the 16S rDNA of T. whipplei in salivary and stool samples using a specific seminested PCR. RESULTS: The prevalence of T. whipplei DNA in patients and in controls was 4.2% (95% CI 2.0-7.6% ) and 7.0% (95% CI 4.0-11.3%), respectively. None of the different gastrointestinal diseases was associated with a higher rate of PCR-positive tests, except for the group of patients with reflux syndrome. Five out of 43 patients with reflux were found to be positive, with all five being positive in the salivary sample. This is in contrast to our findings in carriers without reflux with mainly positive stool samples (p < 0.01). CONCLUSION: We conclude that the asymptomatic carrier state of T. whipplei indeed exists and that it is much more frequent than the rare Whipple's disease. The higher prevalence of T. whipplei DNA in the saliva of patients with reflux syndrome suggests that the stomach might be the habitat of the organism.

Actinomycetales↗
PubMed2003

An interlaboratory comparison for the detection of Mycoplasma pneumoniae in respiratory samples by the polymerase chain reaction.

A panel of 78 respiratory samples collected from 43 patients was analyzed in three different Centers for the presence of Mycoplasma pneumoniae DNA by polymerase chain reaction (PCR). One Center collected the samples and extracted the DNA by two different methods. DNA extracted according to the first method was amplified using primers targetting the 16 S rRNA gene. DNA extracted according to the second method was amplified using the same primers in a semi-nested format and was sent to the two other Centers. The latter Centers both used the same primers targetting the P1 gene but with a different detection format. Thirty-nine samples (50%) from 19 patients were positive by at least two PCR assays. None of the laboratories were free of false positive or false negative PCR results. Calculated specificities of the individual PCR assays ranged from 97.4% to 87.2% and sensitivities ranged from 97.4% to 89.2%. Complement fixation was done on sera of 33 patients. The calculated specificity and sensitivity of serology was 100% and 58.8%, respectively. Several aspects concerning false positive and false negative results with PCR are discussed.

DNA, Bacterial↗
PubMed2003

Ribosomal DNA sequencing for identification of aerobic gram-positive rods in the clinical laboratory (an 18-month evaluation).

We have evaluated over a period of 18 months the use of 16S ribosomal DNA (rDNA) sequence analysis as a means of identifying aerobic gram-positive rods in the clinical laboratory. Two collections of strains were studied: (i) 37 clinical strains of gram-positive rods well identified by phenotypic tests, and (ii) 136 clinical isolates difficult to identify by standard microbiological investigations, i.e., identification at the species level was impossible. Results of molecular analyses were compared with those of conventional phenotypic identification procedures. Good overall agreement between phenotypic and molecular identification procedures was found for the collection of 37 clinical strains well identified by conventional means. For the 136 clinical strains which were difficult to identify by standard microbiological investigations, phenotypic characterization identified 71 of 136 (52.2%) isolates at the genus level; 65 of 136 (47.8%) isolates could not be discriminated at any taxonomic level. In comparison, 16S rDNA sequencing identified 89 of 136 (65.4%) isolates at the species level, 43 of 136 (31.6%) isolates at the genus level, and 4 of 136 (2.9%) isolates at the family level. We conclude that (i) rDNA sequencing is an effective means for the identification of aerobic gram-positive rods which are difficult to identify by conventional techniques, and (ii) molecular identification procedures are not required for isolates well identified by phenotypic investigations.

Bacteria, Aerobic↗
PubMed2002

Aerococcus urinae endocarditis: case report and review of the literature.

Aerococcus urinae is a rare cause of urinary tract infections, mainly in elderly men with underlying urinary tract pathologies. In addition, it has been described as a pathogen in balanitis, soft tissue infections, septicemia and endocarditis. To date ten cases of A. urinae endocarditis have been reported in the literature with a high rate of mortality (7/10) and morbidity, as two out of three survivors suffered from neurovascular complications. Here we present the case of an additional patient who was successfully treated with surgical valve replacement and antibiotic therapy consisting of ceftriaxone and netilmicin for 6 weeks. Furthermore, we review all reported cases of A. urinae endocarditis with emphasis on predisposing factors and therapeutic options.

Aged↗
PubMed2001

Antibodies against recombinant heat shock protein 65 of Tropheryma whipplei in patients with and without Whipple's disease.

Tropheryma whipplei is the causative agent of Whipple's disease (WD), a chronic, life-threatening infection. Laboratory diagnosis is mainly based on PCR and histopathological analysis in duodenal biopsies and other specimens requiring invasive procedures. We have examined the presence of antibodies to recombinant heat shock protein (Hsp65) of T. whipplei in patients with Whipple's disease as well as in control subjects by Western blot and enzyme-linked immunosorbent assay (ELISA). A recombinant plasmid carrying the entire T. whipplei hsp65 gene was constructed, and the expression yielded a 65-kDa histidine-tagged protein. Among four patients with Whipple's disease, two showed an IgG- and one an IgA-response, respectively, when analyzed by Western blotting, whereas from 10 patients without Whipple's disease, only two patients showed a positive IgG-response. The differences between the sera from patients and controls were thus not significant. Successful purification of the protein was achieved by Ni-NTA affinity chromatography. Quantitative analysis of serum antibodies by ELISA demonstrated that antibody levels in the sera of 14 patients were not significantly higher than in those of 89 control subjects. The established ELISA test is not useful to clinical diagnostics.

Actinobacteria↗
PubMed2001

Community-acquired pneumonia in children due to Mycoplasma pneumoniae: diagnostic performance of a seminested 16S rDNA-PCR.

A 16S rDNA-PCR assay for Mycoplasma pneumoniae applied to nasopharyngeal secretion (NPS) or pharyngeal swab (PS) from children with community-acquired pneumonia (CAP) was prospectively compared to serological tests including complement fixation (CF) test, a mu-capture enzyme immuno assay (EIA) for the detection of specific IgM, and an EIA for the detection of specific IgG. During a 24-months-period diagnosis of active M. pneumoniae infection was established in 32 (12.6%) of 253 patients for whom paired sera were available. In the acute phase, the sensitivities of PCR from NPS and PS, CF test, IgM EIA, and IgG EIA were 90.0%, 79.3%, 46.9%, 78.1%, and 59.4%, respectively. The corresponding specificities were 98.1%, 98.6%, 97.6%, 87.1%, and 72.4%, respectively. Thus, the 16S rDNA-PCR assay provides a highly sensitive and accurate tool for the rapid diagnosis of M. pneumoniae infection in children with CAP.

Adolescent↗
PubMed2001

Broad range polymerase chain reaction for diagnosis of rat-bite fever caused by Streptobacillus moniliformis.

An 11-year-old boy presented with fever, vomiting, rash, limping and blisters on his feet after a finger bite by his domestic rat. Although cultures from blood, cerebrospinal fluid and urine remained negative, broad range polymerase chain reaction amplification of a part of the 16S rRNA gene followed by sequencing allowed the detection and identification of Streptobacillus moniliformis in blister fluid, thus confirming the suspected clinical diagnosis of rat-bite fever.

Animals↗
PubMed2001

Description of Tropheryma whipplei gen. nov., sp. nov., the Whipple's disease bacillus.

A detailed characterization was performed of the Whipple's disease bacillus, strain Twist-MarseilleT, isolated from the cardiac valve of a patient with Whipple's disease bacillus endocarditis. This strain was isolated and maintained on human embryonic lung fibroblast monolayers, but could not be cultivated in the absence of living eukaryotic cells. Two morphological forms were observed, with differing staining properties; an intracellular form with intact and degenerating bacteria within vacuoles of infected cells and an extracellular form with masses of bacteria embedded in an extracellular matrix. Determination of the DNA G+C content confirmed that it belongs to the high-G+C gram-positive bacteria. Strain Twist-MarseilleT (= CNCM I-2202T) is proposed as the type strain of a new species within a new genus, Tropheryma whipplei gen. nov., sp. nov., that was provisionally created solely on the basis of 16S rRNA gene sequence data.

Actinobacteria↗
PubMed2001

Whipple's disease and "Tropheryma whippelii".

Whipple's disease is a rare bacterial infection that may involve any organ system in the body. It occurs primarily in Caucasian males older than 40 years. The gastrointestinal tract is the most frequently involved organ, with manifestations such as abdominal pain, malabsorption syndrome with diarrhea, and weight loss. Other signs include low-grade fever, lymphadenopathy, skin hyperpigmentation, endocarditis, pleuritis, seronegative arthritis, uveitis, spondylodiscitis, and neurological manifestations, and these signs may occur in the absence of gastrointestinal manifestations. Due to the wide variability of manifestations, clinical diagnosis is very difficult and is often made only years or even decades after the initial symptoms have appeared. Trimethoprim-sulfamethoxazole for at least 1 year is usually considered adequate to eradicate the infection. The microbiological diagnosis of this insidious disease is rendered difficult by the virtual lack of culture and serodiagnostic methods. It is usually based on the demonstration of periodic acid-Schiff-positive particles in infected tissues and/or the presence of bacteria with an unusual trilaminar cell wall ultrastructure by electron microscopy. Recently, the Whipple bacteria have been characterized at the molecular level by amplification of their 16S rRNA gene(s). Phylogenetic analysis of these sequences revealed a new bacterial species related to the actinomycete branch which was named "Tropheryma whippelli." Based on its unique 16S ribosomal DNA (rDNA) sequence, species-specific primers were selected for the detection of the organism in clinical specimens by PCR. This technique is currently used as one of the standard methods for establishing the diagnosis of Whipple's disease. Specific and broad-spectrum PCR amplifications mainly but not exclusively from extraintestinal specimens have significantly improved diagnosis, being more sensitive than histopathologic analysis. However, "T. whippelii" DNA has also been found in persons without clinical and histological evidence of Whipple's disease. It is unclear whether these patients are true asymptomatic carriers or whether differences in virulence exist among strains of "T. whippelii" that might account for the variable clinical manifestations. So far, six different "T. whippelii" subtypes have been found by analysis of their 16S-23S rDNA spacer region. Further studies of the pathogen "T. whippelii" as well as the host immune response are needed to fully understand this fascinating disease. The recent cultivation of the organisms is a promising major step in this direction.

Actinobacteria↗