Seasonality of Clostridium difficile infections in Southern Germany.

Between 2000 and 2009, the total number of patients with Clostridium difficile infections increased considerably in Southeastern Germany. A clear seasonality was observed with a higher number of affected patients occurring in the winter months (January-March). Moxifloxacin and erythromycin-resistant C. difficile PCR ribotypes 001 (72%) and 027 (4·6%) were the most commonly isolated strains.

Recognition of Clostridium difficile PCR-ribotypes 001, 027 and 126/078 using an extended MALDI-TOF MS system.

During the last decade, Clostridium difficile infection (CDI) increased markedly inside as well as outside of hospitals. In association with the occurrence of new hypervirulent C. difficile strains, CDI became more important. Until now typing of C. difficile strains has been enabled by PCR-ribotyping. However, this method is restricted to specialized laboratories combined with high maintenance cost. Therefore, we tested MALDI-TOF mass spectrometry for typing of C. difficile to provide a fast method for surveillance of CDI. Using a standard set of 25 different C. difficile PCR ribotypes a database was made by different mass spectra recorded in the SARAMIS software (AnagnosTec, Zossen, Germany). The database was validated with 355 C. difficile strains belonging to 29 different PCR ribotypes collected prospectively from all submitted feces samples in 2009. The most frequent PCR ribotypes were type 001 (70%), 027 (4.8%) and 078/126 (4.7%). All three types were recognized by MALDI-TOF MS. We conclude that an extended MALDI-TOF system was capable to recognize specific markers for ribotypes 001, 027 and 078/126 allowing an effective identification of these strains.

Association of ciprofloxacin prescriptions to outpatients to Clostridium difficile infections.

To study if antibiotic treatment of outpatients had triggered Clostridium difficile infections (CDI), prescription numbers were compared with CDI-affected patient numbers. A strong correlation was observed for ciprofloxacin (R=0.917), suggesting that increased use of ciprofloxacin by outpatients contributed to increased numbers of CDI. These findings deserve further investigation as they may have an impact on future decisions regarding antibiotic prescribing.

Increased number of Clostridium difficile infections and prevalence of Clostridium difficile PCR ribotype 001 in southern Germany.

In recent years, Clostridium difficile infection (CDI) has emerged as an increasing problem, both in in- and outpatients. In a rural region of southern Germany, the annual number of C. difficile toxin (Tcd)-positive patients has increased from 95 to 796 in the period from 2000 to 2007. Simultaneously, the proportion of positive tests among all Tcd examinations has risen from 7.0% to 12.8%, indicating that the higher number of affected patients was not solely due to an increase in the number of assays. Elevated numbers of CDI have recently been associated with outbreaks of the ribotype 027 strain, particularly in North America. This strain has also been isolated in Europe, including in Germany. Ribotyping and PCR testing for binary toxin genes of C. difficile strains isolated from in- and outpatients demonstrate a predominance (59%) of C. difficile ribotype 001, which exhibits antibiotic resistance to erythromycin, ciprofloxacin, and moxifloxacin, but lacks binary toxin genes. In summary, in our region of Germany, the number of patients affected by CDI has increased, probably due to spread of C. difficile ribotype 001.

Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS).

Ceramide (CER) is an important signaling molecule involved in a variety of cellular processes, including differentiation, cell growth, and apoptosis. Currently, different techniques are applied for CER quantitation, some of which are relatively insensitive and/or time consuming. Tandem mass spectrometry with its high selectivity and sensitivity is a very useful technique for detection of low abundant metabolites without prior purification or derivatization. In contrast to existing mass spectrometry methods, the developed electrospray tandem mass spectrometry (ESI-MS/MS) technique is capable of quantifying different CER species from crude cellular lipid extracts. The ESI-MS/MS is performed with a continuous flow injection and the use of an autosampler, resulting in a high throughput capability. The collision-induced fragmentation of CER produced, in addition to others, a characteristic fragment of m/z 264, making a precursor ion scan of 264 well suited for CER quantitation. Quantitation is achieved by use of a constant concentration of a non-naturally occurring internal standard C8-CER, together with a calibration curve established by spiking different concentrations of naturally occurring CER. The calibration curves showed linearity over a wide concentration range and sample volumes equivalent to 10 microg of cell protein corresponding to about 20, 000 fibroblasts were sufficient for CER analysis. Moreover this assay showed a detection limit at the subpicomole level. In summary, this methodology enables accurate and rapid analysis of CER from small samples without prior separation steps, thus providing a useful tool for signal transduction research.

Molecular cloning of the human ATP-binding cassette transporter 1 (hABC1): evidence for sterol-dependent regulation in macrophages.

We have cloned the full-length cDNA for the human ATP binding cassette transporter 1 (hABC1). The 6603-bp open reading frame encodes a polypeptide of 2201 amino acids resulting in a deduced molecular weight of 220 kDa. The hABC1 cDNA is highly homologous (62%) to the human rim ABC transporter (ABCR). hABC1 is expressed in a variety of human tissues with highest expression levels found in placenta, liver, lung, adrenal glands, and fetal tissues. We demonstrate that the hABC1 expression is induced during differentiation of human monocytes into macrophages in vitro. In macrophages, both the hABC1 mRNA and protein expression are upregulated in the presence of acetylated low-density lipoprotein (AcLDL). The AcLDL-induced increase in hABC1 expression is reversed by cholesterol depletion mediated by the addition of high-density lipoprotein (HDL3). Our data, demonstrating sterol-dependent regulation of hABC1 in human monocytes/macrophages, suggest a novel role for this transporter molecule in membrane lipid transport.

The influence of comonomer composition on dimethacrylate resin properties for dental composites.

During the polymerization of multifunctional monomers for dental restorations, typical final double-bond conversions range from 55 to 75%. The low conversion results in a large amount of extractable monomer, reduced adhesion to the filler, and the potential for increased swelling. In this work, the ability to increase the maximum conversion by optimizing the copolymer composition is explored. A series of multi-ethylene glycol dimethacrylate monomers of various lengths was used as a model system to determine how the copolymer composition affects the final conversion, the mechanical properties, and the predicted shrinkage. It was found that the ultimate conversion can be significantly increased, shrinkage decreased, and mechanical properties maintained. It was found that up to 30 wt% of poly(ethylene glycol) 600 dimethacrylate could be added to diethylene glycol dimethacrylate without reducing the strength and increasing the conversion. Results for other comonomer combinations were similar.

Continuous quantification of urethral competence with a new tube-foil sleeve catheter.

Increasing urethral occlusion pressures were created with an artificial sphincter in bladder-urethra specimens of adult female pigs. Antegrade urethral perfusion pressures, at 19.5 ml./min. through a flexible F8 urethral catheter with an F12 outflow sleeve of tube-foil, closely approximated the associated intravesical pressures at which leakage from the noncatheterized urethra starts. With this new technique, leakage pressure and thus urethral competence can be measured accurately and continuously, almost independently from bladder pressure and pelvic floor movement.