Use of quantitative microbiological analyses to trace origin of contamination of parenteral nutrition solutions.

In the summer of 2010, parenteral nutrition (PN) admixtures were administered to neonates in the Pediatric Department of the University Medical Center Mainz that provoked severe clinical sequelae. Contamination of a dummy infusion with Enterobacter cloacae and Escherichia hermannii was detected on the day of the incident, and the same isolates were subsequently grown from all PN admixtures as well as from the parent amino acid solution from which the admixtures had been prepared. Quantitative microbiological analyses paired with the determination of endotoxin concentrations enabled the conclusion to be reached that the amino acid solution had represented the primary source of contamination, which must have occurred in the distant past and may have derived from passage of the bacteria through a crack in the glass container. The findings have large implications, and the approaches employed should become of value when similar incidents occur again in the future.

Real-time PCR detection of Pseudomonas aeruginosa in clinical and municipal wastewater and genotyping of the ciprofloxacin-resistant isolates.

Real-time quantification of Pseudomonas aeruginosa was performed in various wastewater systems including clinical, municipal wastewaters and inflow from a wastewater treatment plant. The highest concentrations of P. aeruginosa-specific targets were detected in clinical wastewaters. Limitations of the detection system resulting from inhibition or cross-reaction were identified. Ciprofloxacin-resistant P. aeruginosa strains were isolated after specific enrichment from clinical and municipal wastewaters. In some cases they were also cultivated from effluent of a wastewater treatment plant, and from its downstream river water. A total of 119 isolates were phenotypically characterized as ciprofloxacin-resistant via antibiogram testing. Subsequently, the fluoroquinolone-resistance-mediating mutations in the genes gyrA codon positions 83 and 87, gyrB codon position 466 and parC codon positions 87 and 91 were determined by mini-sequencing. Ciprofloxacin resistance was mainly associated with mutations in gyrA codon position 83 and parC mutation in codon positions 87 or 91 of the bacterial gyrase and topoisomerase II genes. All ciprofloxacin-resistant P. aeruginosa strains were compared with genotypes from clinical data of fluoroquinolone-resistant P. aeruginosa infections. The results were in agreement with data from clinical analyses, with the exception that no gyrA 87 and no gyrB mutations were found in ciprofloxacin-resistant P. aeruginosa wastewater isolates.

The "Mainzer EMF-Wachhund": results from a watchdog project on self-reported health complaints attributed to exposure to electromagnetic fields.

The "Mainzer EMF-Wachhund," a watchdog project, offered a system of self-notification of health complaints attributed to exposures to electromagnetic fields (EMFs) to a population of a part of Germany with about 4 million inhabitants. By using a self-administered questionnaire, which was provided online and for download from the Internet, 192 persons reported such health complaints in the period from October 2003 to March 2005. Of these, 56% classified themselves as electromagnetic hypersensitive (EH). Predictors of this self classification were being affected by all kinds of EMF rather than single EMF sources and being female. On average, EH subjects reported a high degree of suffering, 77% of whom had already sought advice from physicians. An Internet-based standardized questionnaire is an economic way of offering affected persons a direct link to scientific institutions to establish contact. However, the study base obtained by such an approach is not representative to estimate a population-based prevalence. As a large number of subjects did not classify themselves as EH and reported very specific links between exposure and symptoms, they may provide a very distinct and interesting group for future research.

Microbiological quality of carbonated drinking water produced with in-home carbonation systems.

The microbiological quality of carbonated water produced with tap water in commercial in-home carbonation systems was determined, the results being discussed in the context of the microbiological quality of the tap water used, the properties of the drink makers, and the procedures of preparation and washing of various parts of the appliance. The last-mentioned data were received from each participant of the study by questionnaire. Escherichia coli, coliforms, fecal streptococci and spore-forming sulphite-reducing anaerobes were used as indicators for the hygienic quality of the water. Tap-water samples were collected according to the usual procedure when filling the carbonating bottle, i.e., without previous flushing and disinfection of the faucet. In 12% of tap-water samples, coliforms could be detected. On the other hand, in 20 of 52 carbonated waters (39%), coliforms as indicators of water pollution were found. By means of fecal streptococci and Pseudomonas aeruginosa, it was possible to establish additional contamination not involving E. coli or coliforms alone. Analysis revealed that, in addition to contaminated tap water, a bacterial biofilm on the inner surface of the re-usable bottles had a predominant influence on the microbiological quality of the carbonated water.

In vitro evaluation of the biocompatibility of contaminated implant surfaces treated with an Er : YAG laser and an air powder system.

Titanium platelets with a sand-blasted and acid-etched surface were coated with bovine serum albumin and incubated with a suspension of Porphyromonas gingivalis (ATCC 33277). Four groups with a total of 48 specimens were formed. Laser irradiation of the specimens (n = 12) was performed on a computer-controlled XY translation stage at pulse energy 60 mJ and frequency 10 pps. Twelve specimens were treated with an air powder system. After the respective treatment, human gingival fibroblasts were incubated on the specimens. The proliferation rate was determined by means of fluorescence activity of a redox indicator (Alamar Blue Assay) which is reduced by metabolic activity related to cellular growth. Proliferation was determined up to 72 h. Contaminated and non-treated as well as sterile specimens served as positive and negative controls. Proliferation activity was significantly (Mann-Whitney U-test, P < 0.05) reduced on contaminated and non-treated platelets when compared to sterile specimens. Both on laser as well as air powder-treated specimens, cell growth was not significantly different from that on sterile specimens. Air powder treatment led to microscopically visible alterations of the implant surface whereas laser-treated surfaces remained unchanged. Both air powder and Er : YAG laser irradiation have a good potential to remove cytotoxic bacterial components from implant surfaces. At the irradiation parameters investigated, the Er : YAG laser ensures a reliable decontamination of implants in vitro without altering surface morphology.

Infections associated with medical devices: pathogenesis, management and prophylaxis.

The insertion or implantation of foreign bodies has become an indispensable part in almost all fields of medicine. However, medical devices are associated with a definitive risk of bacterial and fungal infections. Foreign body-related infections (FBRIs), particularly catheter-related infections, significantly contribute to the increasing problem of nosocomial infections. While a variety of micro-organisms may be involved as pathogens, staphylococci account for the majority of FBRIs. Their ability to adhere to materials and to promote formation of a biofilm is the most important feature of their pathogenicity. This biofilm on the surface of colonised foreign bodies is regarded as the biological correlative for the clinical experience with FBRI, that is, that the host defence mechanisms often seem to be unable to handle the infection and, in particular, to eliminate the micro-organisms from the infected device. Since antibacterial chemotherapy is also frequently not able to cure these infections despite the use of antibacterials with proven in vitro activity, removal of implanted devices is often inevitable and has been standard clinical practice. However, in specific circumstances, such as infections of implanted medical devices with coagulase-negative staphylococci, a trial of salvage of the device may be justified. All FBRIs should be treated with antibacterials to which the pathogens have been shown to be susceptible. In addition to systemic antibacterial therapy, an intraluminal application of antibacterial agents, referred to as the 'antibiotic-lock' technique, should be considered to circumvent the need for removal, especially in patients with implanted long-term catheters. To reduce the incidence of intravascular catheter-related bloodstream infections, specific guidelines comprising both technological and nontechnological strategies for prevention have been established. Quality assurance, continuing education, choice of the catheter insertion site, hand hygiene and aseptic techniques are aspects of particular interest. Furthermore, all steps in the pathogenesis of biofilm formation may represent targets against which prevention strategies may be directed. Alteration of the foreign body material surface may lead to a change in specific and nonspecific interactions with micro-organisms and, thus, to a reduced microbial adherence. Medical devices made out of a material that would be antiadhesive or at least colonisation resistant would be the most suitable candidates to avoid colonisation and subsequent infection. Another concept for the prevention of FBRIs involves the impregnation of devices with various substances such as antibacterials, antiseptics and/or metals. Finally, further studies are needed to translate the knowledge on the mechanisms of biofilm formation into applicable therapeutic and preventive strategies.

Evaluation of different primers for DNA fingerprinting of Legionella pneumophila serogroup 1 strains by polymerase chain reaction.

A DNA fingerprinting method for the characterization of Legionella pneumophila serogroup 1 strains was established. This method was based on the DNA extraction using Chelex 100 and subsequent PCR analysis using primers under conditions of low stringency. Sixteen single primers were tested for the typing of the 10 epidemiologically unrelated reference strains of L. pneumophila serogroup 1 as well as patient isolates and environmental strains isolated from the water system of a hospital where patients with legionellosis were treated. In addition, a combination of two primers (Lpm-1 and Lpm-2) originally established for the specific detection of Legionella strains was tested. The PCR results were compared with two further subtyping methods, i.e. monoclonal antibody analysis and pulsed-field gel electrophoresis. The type strains Philadelphia 1, Knoxville 1, Allentown 1, Benidorm 0303E, Bellingham 1, and France 5811 could be distinguished clearly in experiments using all of the primers. Depending on the primer used, Heysham 1 and Oxford 4032E showed different DNA profiles. The strains Olda and Camperdown 1 were nearly indistinguishable. In contrast, the analysis by PFGE and MAb subtyping revealed distinct types for all 10 reference strains. The discrimination of the patient isolates from two suspected cases of nosocomial legionellosis and environmental isolates was not possible with the 16 single primers used in the study. However, the PCR assay with the combination of Lpm-1 and Lpm-2 as well as the PFGE and MAb analysis were able to differentiate distinct types. The use of the sequence-specific primers under low-stringency annealing conditions allowed both simultaneous gene detection as well as epidemiological typing of Legionella strains.

Development of a long-lasting ventricular catheter impregnated with a combination of antibiotics.

A ventricular silicone catheter impregnated with a combination of rifampin and a quinolone was developed in order to prevent ventricular shunt related infections. As model substance for the quinolones we used sparfloxacin, because of its specific physicochemical properties resulting in a quantitative detection also in the presence of a second antibiotic. In our study we focused especially on an optimization of the antibiotic release out of the impregnated catheters in order to develop long lasting devices with a broad antimicrobial spectrum. A release-optimized catheter was tested with an in vitro colonization test and additionally with a method developed to examine the spread of bacteria on a catheter surface. In vitro experiments showed that the impregnated catheters reduce the colonization with Staphylococcus epidermidis for at least 1 year and prevent the spread of bacteria along the catheter surface.