The impact of a register on the management of neonatal cooling in Switzerland.

BACKGROUND: Therapeutic hypothermia following hypoxic ischaemic encephalopathy in term infants was introduced into Switzerland in 2005. Initial documentation of perinatal and resuscitation details was poor and neuromonitoring insufficient. In 2011, a National Asphyxia and Cooling Register was introduced. AIMS: To compare management of cooled infants before and after introduction of the register concerning documentation, neuromonitoring, cooling methods and evaluation of temperature variability between cooling methods. STUDY DESIGN: Data of cooled infants before the register was in place (first time period: 2005-2010) and afterwards (second time period: 2011-2012) was collected with a case report form. RESULTS: 150 infants were cooled during the first time period and 97 during the second time period. Most infants were cooled passively or passively with gel packs during both time periods (82% in 2005-2010 vs 70% in 2011-2012), however more infants were cooled actively during the second time period (18% versus 30%). Overall there was a significant reduction in temperature variability (p < 0.001) comparing the two time periods. A significantly higher proportion of temperature measurements within target temperature range (72% versus 77%, p < 0.001), fewer temperature measurements above (24% versus 7%, p < 0.001) and more temperatures below target range (4% versus 16%, p < 0.001) were recorded during the second time period. Neuromonitoring improved after introduction of the cooling register. CONCLUSION: Management of infants with HIE improved since introducing the register. Temperature variability was reduced, more temperature measurements in the target range and fewer temperature measurements above target range were observed. Neuromonitoring has improved, however imaging should be performed more often.

RNA/DNA co-analysis from blood stains--results of a second collaborative EDNAP exercise.

A second collaborative exercise on RNA/DNA co-analysis for body fluid identification and STR profiling was organized by the European DNA Profiling Group (EDNAP). Six human blood stains, two blood dilution series (5-0.001 µl blood) and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by the participating laboratories using a RNA/DNA co-extraction or solely RNA extraction method. Two novel mRNA multiplexes were used for the identification of blood: a highly sensitive duplex (HBA, HBB) and a moderately sensitive pentaplex (ALAS2, CD3G, ANK1, SPTB and PBGD). The laboratories used different chemistries and instrumentation. All of the 18 participating laboratories were able to successfully isolate and detect mRNA in dried blood stains. Thirteen laboratories simultaneously extracted RNA and DNA from individual stains and were able to utilize mRNA profiling to confirm the presence of blood and to obtain autosomal STR profiles from the blood stain donors. The positive identification of blood and good quality DNA profiles were also obtained from old and compromised casework samples. The method proved to be reproducible and sensitive using different analysis strategies. The results of this collaborative exercise involving a RNA/DNA co-extraction strategy support the potential use of an mRNA based system for the identification of blood in forensic casework that is compatible with current DNA analysis methodology.

Selection of highly specific and sensitive mRNA biomarkers for the identification of blood.

In the present work, we have evaluated eight reportedly blood-specific mRNA markers (HBB, HBA, ALAS2, CD3G, ANK1, PBGD, SPTB, AQP9) in an attempt to determine the most suitable ones for use in forensic applications based on their sensitivities, specificities and performance with casework samples. While varying levels of expression were observed, all markers were relatively sensitive requiring as little as 1ng of RNA input into the reverse transcription (RT) reaction. In singleplex reactions, seven of the eight analyzed blood markers (all except AQP9) demonstrated a high degree of specificity for blood. In multiplex reactions, non-reproducible cross-reactivity was observed for several of the mRNA markers, which was reduced and, in most cases, eliminated when less input total RNA was used. Additionally, some cross-reactivity was observed with tissue and animal samples. Despite differences in the observed sensitivity and specificity of the blood markers examined in this study, a number of the candidates appear to be suitable for inclusion in appropriately validated multiplex mRNA-based body fluid identification systems.

mRNA profiling for the identification of blood--results of a collaborative EDNAP exercise.

A collaborative exercise on mRNA profiling for the identification of blood was organized by the European DNA Profiling Group (EDNAP). Seven blood samples and one blood dilution series were analyzed by the participating laboratories for the reportedly blood-specific markers HBB, SPTB and PBGD, using different kits, chemistries and instrumentation. The results demonstrate that HBB is expressed abundantly in blood, SPTB moderately and PBGD significantly less. All but one of the 16 participating laboratories were able to successfully isolate and detect RNA from the dried bloodstains even though a majority of the laboratories had no prior experience with RNA. Despite some expected variation in sensitivity between laboratories, the method proved to be reproducible and sensitive using different analysis strategies. The results of this collaborative exercise support the potential use of mRNA profiling as an alternative to conventional serological tests.

mRNA profiling for body fluid identification by reverse transcription endpoint PCR and realtime PCR.

mRNA profiling is a promising new method for the identification of body fluids from biological stains. Major advantages of mRNA profiling are the possibility of detecting several body fluids in one multiplex reaction and of simultaneously isolating DNA without loss of material. A reverse transcription endpoint polymerase chain reaction (PCR) method and a realtime PCR assay were established for the identification of blood, saliva, semen, vaginal secretions and menstrual blood, and were compared to conventional enzymatic and immunologic tests. The results for specificity, sensitivity and suitability to biological stains were satisfying and mRNA stability was demonstrated for up to 2-year-old stains. Two novel multiplex assays were created with the endpoint PCR primers: multiplex 1 amplifies two markers for each of the above mentioned body fluids and is suited for screening; multiplex 2 was designed for the detection of blood, vaginal secretions and menstrual blood. The results demonstrate that both endpoint PCR and realtime PCR are suitable for the identification of body fluids in forensic stains and represent an effective alternative to conventional enzymatic and immunologic tests.

[Detection of papillomavirus DNA in the prostate: a virus with underestimated clinical relevance?]. / Die Detektion von Papillomavirus-DNA in der Prostata : Ein Virus mit unterschätzter klinischer Relevanz?

BACKGROUND: Human papillomaviruses (HPV) are the most frequent pathogens of sexually transmitted diseases. They have been associated with an increased incidence of several anogenital tumors. Whether oncogenic HPV are involved in the pathogenesis of prostate cancer has been a subject of great controversy. This study's purpose was to investigate the association between HPV infection and prostate cancer (PCA). MATERIAL AND METHODS: The study included 213 consecutive patients with an average age of 65.7 (+/-8.4) years. Within the framework of transrectal, ultrasonic-guided multibiopsy of the prostate, one additional core was examined by means of polymerase chain reaction (PCR) in relation to bacterial, fungal, and viral (including HPV) DNA, with subsequent DNA sequencing. The collected data were correlated with the histological results and with diverse clinical variables. The influence of several predictors for the existence of PCA was verified with a logistic regression model. RESULTS: No general bacterial DNA (16S rDNA) was detected. Of the 213 patients, 145 (68.1%) showed HPV DNA. In 64% (n=137), high-risk HPV DNA were depicted; these were 18% of the total in each case of HPV genotypes 16 and 18. From our examinations, no significant positive correlation existed between the HPV evidence and the histologically verified PCA that was found in 23.5% of the patients (n=50; odds ratio 1.45; 95% confidence interval 0.71-2.91). The BK virus was not found in any of the cores confirmed through PCR. CONCLUSION: Although no positive correlation between HPV infection and PCA existed in our study, data from the literature suggest an influence of the papillomavirus on PCA oncogenesis. Future studies should highlight to what extent HPV DNA is inserted in the genome of prostate cells and is able to cause subsequent malignant transformation of particular genes.

Y-chromosome STR haplotypes in a population sample from Switzerland (Zurich area).

Allele frequencies and haplotypes for 12 Y-chromosome STR loci (DYS19, DYS385a/b, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438 and DYS439), included in the PowerPlex Y multiplex kit, were determined for a Swiss population sample of 150 male individuals. The gene diversities for the different loci were in the same range as shown for other European Population samples. The haplotype diversity was 0.9922. Pairwise haplotype analysis showed no significant differences in comparison with other European Population samples.

[Forgoing life-sustaining measures in patients at the end of life in the German-speaking part of Switzerland: results of a death certificate study]. / Begrenzung lebenserhaltender Massnahmen bei Patienten am Lebensende in der deutschsprachigen Schweiz -- Resultate einer Todesfallstudie.

BACKGROUND AND OBJECTIVE: There have not been any comprehensive data from German-speaking countries on the medical practice of withholding or withdrawing treatment at the end of life. This study from the German-speaking part of Switzerland provides the first in-depth analysis in this field. This study is based on data from this region and is a contribution to a large empirical research project on medical end-of-life decisions in 6 European countries (EURELD). METHODS: Continuous random samples (n = 4991) were taken from all deaths in the German-speaking part of Switzerland that had been reported to the Swiss Federal Office of Statistics between June and October 2001. Doctors who had been attending the deceased person were asked to complete mailed questionnaires, their replies being kept strictly anonymous. RESULTS: The response rate was 67 %. Medications were withheld or withdrawn in 48 % of all treatments forgone: among these, antibiotics accounted for 17 %. Other potentially life-sustaining medical measures forgone included artificial hydration (12 %), surgery (7 %), artificial feeding (6 %), chemotherapy (6 %), diagnostic tests (4 %), hospital admissions (3 %), renal dialysis (2 %), blood product infusions (2 %), intubation (2 %), ventilation (2 %), resuscitation (2 %), and radiotherapy (1 %). 43 % of all treatments were forgone in patients who died in hospital, 42 % in nursing homes, and 15 % at home. In almost three-quarters (73 %) of the treatments forgone, a primary-care doctor had ordered the treatment to be withheld or withdrawn. On average, forgoing treatment led to a life-shortening effect of more than one month in 8 % of all cases. The proportion was higher for renal dialysis (25 %), blood products infusion (18 %), and diagnostic tests (16 %). CONCLUSIONS: Forgoing life-sustaining medical treatment comprises a wide range of decisions taken in many different clinical settings. In most cases the likely lifespan is only slightly shortened.

DNA mixtures: biostatistics for mixed stains with haplotypic genetic markers.

The conventional theory for interpreting forensic DNA evidence developed for the autosomal genetic markers is not applicable in the case of haplotypic markers, specifically for Y-STR based data. The reason is, that in contrast to the case of autosomal markers, single alleles found in the mixed stain cannot be assigned to unknown stain contributors independently of each other, while the assignable entities are sets of linked alleles which should be treated as non-separable units. It is shown that the conventional theory cannot be extended to this situation. A novel theory which accounts for the features of haplotypic markers has been developed within the general framework of the hypotheses testing approach. This theory opens the way for the use of haplotypic markers in the analysis of mixed stains with the arbitrary numbers of unknown contributors and linked loci. A numerical example demonstrates the application of the theory.

[Diagnosis of systemic Candida infections. Evaluation of serology, molecular biology and D-arabinitol detection]. / Diagnose systemischer Candida-Infektionen: Bewertung der Serologie, der Molekularbiologie und des D-Arabinitol-Nachweises.

In order to diagnose systemic Candida infection in ICU patients, several methods were compared. In the first round, antigens and antibodies of 104 patients were investigated. Seventeen patients were infected; in nine of these patients a positive antigen was detected; elevated antibodies were detected in 11 patients. Only 42 patients were colonized. In this group, one positive antigen and six elevated antibody titres were detected. Combining these results, the sensitivity of the antigen only (58.5%) or the antibodies only (52.9%) increased to 100%. In the second round (n = 83 patients) the D-arabinitol/L-arabinitol quotient was also determined in the urine. 18.1% of the patients had elevated antibodies and 26.5% elevated D-arabinitol/L-arabinitol quotients. In the third round, PCR was also applied (n = 27 patients). Five patients had elevated D-arabinitol/L-arabinitol quotients and one of these had a positive PCR result. In conclusion, serological methods (antigen and antibody detection) should only be applied in strictly selected patients. In uncertain cases, the addition of PCR or determination of D-arabinitol/L-arabinitol might be helpful.

[Unusual gram positive rods, causing pneumonia]. / Ungewöhnliche grampositive Stäbchenbakterien als Auslöser von Pneumonien.

In the last decade, a growing number of patients with pneumonia, caused by unusual gram positive rods have been observed. Mostly, the patients had been infected as a consequence of impaired immunity. In some cases, bioterrorist activities may also induce pneumonia by gram positive rods (B. anthracis). In order to bring these organisms to the attention of the medical community, we present three clinical cases and describe six species of gram positive rods, known to provoke this kind of pneumonias. Case 1 was a 84 years old patient with impaired lung function. He was suspicious of tuberculosis (Tbc). Nocardia spec. was isolated. Case 2 was an alcoholic of 46 years with pneumonia. Reactivation of Tbc was suspected. Actinomadura madurae has been isolated. Case 3 was a patient of 58 years with myelodysplastic syndrome (MDS) and pneumonia. N. asteroides was isolated. All patients shared impaired immunity (age, alcoholism, MDS) with impaired lung functions; Tbc had been suspected (Case 1 + 2). Infection by A. madurae was contained by Clindamycin. Therapy of Nocardia with Moxifloxacin (Case 1) or Bactrim (Case 3) was only partly effective. In the appendix, six species of gram positive rods which are known to cause pneumonia, are summarized (Nocardia, Actinomyceta, Actinomadura, Rhodococcus, Corynebacterium and Bacillus).

Assisted suicide bordering on active euthanasia.

A 44-year-old woman was almost completely paralysed after a severe brainstem haemorrhage. Even after several years of efforts at rehabilitation, she remained completely dependent on the help of others. However, a special device enabled her to administer (after careful preparation) liquids through the PEG catheter despite her poorly coordinated movements. Four years after the stroke, the woman joined a right-to-die society with the wish to bring her life to an end. A doctor working with this organisation prescribed her a lethal dose of pentobarbital. In the presence of her husband and her companion from the organisation, the woman administered herself the lethal substance by means of the device. On the basis of the fact that she herself had switched the device on this death was classed as (assisted) suicide.

[How stable is the attitude of aged people towards life-extending measures? Results of a 3-year follow-up in nursing home residents]. / Wie stabil ist die Einstellung Betagter zu lebensverlängernden Massnahmen? Resultate einer 3-Jahres-Katamnese von Pflegeheimbewohnern.

OBJECTIVE: To explore the stability of attitude towards life-extending measures in aged people. METHOD: Face-to-face interviews of nursing home residents at an interval of 3 years (prospective longitudinal survey). RESULTS: During the first interview period in 1997 (n=50), 19 nursing home residents (38%) had advocated treatment with antibiotics in a hypothetical scenario of acute life-threatening pneumonia. 15 individuals (30%) had refused such treatment, while statements from a further 16 (32%) had been ambivalent. Three years later, 19 individuals could be re-interviewed (28 had died in the meantime, 3 suffered from advanced dementia). 16 of these expressed the same or a similar attitude towards the above scenario as they did three years earlier (correspondence 84.2%, p<0.01). CONCLUSION: The results indicate a high stability of attitude towards life-extending measures in aged people. This finding may have implications for the meaning of advance directives in geriatric long-term care.

Diagnosis of systemic Candida infections in patients of the intensive care unit. Significance of serum antigens and antibodies.

The combined detection of Candida antigen and antibody for the determination of systemic Candida infections (SCI) was investigated. One hundred and four patients from the intensive care unit (ICU) were analysed. Seventeen of the patients were suspected of having SCI, based on clinical and laboratory criteria. In these patients, Candida antigens and antibodies were analysed extensively. Ten patients had a positive Candida antigen (titre >1:16) determined by the latex agglutination assay Cand-Tec(R) and their median antibody titre was 1:160 in the indirect haemagglutination test (HAT). Seven antigen-negative patients had a median titre of 1:1280 (HAT). Forty-one of 42 colonized control patients had negative antigen titres and a median antibody titre of 1:160. The sensitivities and specificities were 58.8% and 97.6% for antigenemia, and 52.9% and 85.7% for antibody detection. These values reached 100.0% and 83.3%, respectively, when the results of both tests were combined. This indicates a high degree of concordance between serological results (Candida antigen and/or antibodies) and clinical presentation. We conclude, that the combined investigation of antigen and antibody titres might be a helpful tool in the characterization of SCI in ICU patients, if antigen titres are >or=1:16 or antibody titres (HAT) are >or=1:640.

Open regulation and practice in assisted dying.

The Netherlands, Oregon and Switzerland are the only areas in the world where assistance in dying has legally been practised in recent years. This article provides a detailed comparison of the history of the origins, legislation, monitoring systems and the extent of assistance in dying in these three places. It shows that the actual practice in Switzerland which, unlike Oregon, also allows assistance in suicide by means of infusions or gastric tubes, can today be technically quite similar to the permitted practice of active euthanasia on request in the Netherlands. Considering the preconditions restricting these practices, Swiss regulations are the most open, in that the law requires neither a medical second opinion (as in both the Netherlands and Oregon) nor the existence of a terminal illness (as in Oregon) as prerequisite to assistance in dying. In 2001, the proportion of assisted deaths (as reported to the authorities) in all deaths was almost ten times higher in the Netherlands (1.5% of all deaths) then in Oregon (<0.1% of all deaths) or Switzerland (0.2% of all deaths). The analysis of the different normative concepts underlying legislation reveals that in the Netherlands the basis for non-prosecution lies in the conflict of the physician's duties to respect life versus relief of suffering, while in the USA and in Switzerland the right-to-die concept plays a major role. These two concepts allow appreciation of distinctions between the roles of the physician in end-of-life practices and between assisted suicide and voluntary active euthanasia.

Contamination of bronchoscopes with Mycobacterium tuberculosis and successful sterilization by low-temperature hydrogen peroxide plasma sterilization.

BACKGROUND: The transmission of mycobacteria by bronchoscopes has been reported several times in the last years. To explore methods to prevent transmission of tuberculosis in this way, we sterilized contaminated bronchoscopes with low-temperature hydrogen peroxide gas plasma sterilization. METHODS: Bronchoscopes were contaminated with Mycobacterium tuberculosis and decontaminated with a washer/disinfector ("normal washing"). Some were additionally disinfected with glutaraldehyde ("intensive washing"). Afterward the bronchoscopes were sterilized by low-temperature hydrogen peroxide plasma sterilization. RESULTS: After normal washing, 8/17 samples had positive results by culture, and 7/17 had positive results by nucleic acid amplification technique. After intensive washing, all samples had negative results by culture, and 10/25 had positive results by nucleic acid amplification technique; after sterilization with low-temperature hydrogen peroxide plasma sterilization, all samples had negative results by culture and nucleic acid amplification technique. CONCLUSION: Washing of bronchoscopes, as performed normally, is not sufficient for decontamination of bronchoscopes. Additional disinfection is recommended. If the nucleic acid amplification technique is used for diagnostic procedures, sterilization by low-temperature hydrogen peroxide plasma sterilization is recommended to avoid false-positive results.

A variable fluence step clustering and segmentation algorithm for step and shoot IMRT.

A step and shoot sequencer was developed that can be integrated into an IMRT optimization algorithm. The method uses non-uniform fluence steps and is adopted to the constraints of an MLC. It consists of a clustering, a smoothing and a segmentation routine. The performance of the algorithm is demonstrated for eight mathematical profiles of differing complexity and two optimized profiles of a clinical prostate case. The results in terms of stability, flexibility, speed and conformity fulfil the criteria for the integration into the optimization concept. The performance of the clustering routine is compared with another previously published one (Bortfeld et al 1994 Int. J. Radiat. Oncol. Biol. Ph.vs. 28 723-30) and yields slightly better results in terms of mean and maximum deviation between the optimized and the clustered protile. We discuss the specific attributes of the algorithm concerning its integration into the optimization concept.

Second primary glioblastoma.

Although characterized by a highly variable phenotype and multiple genetic alterations, glioblastomas are considered monoclonal in origin. We here report on a 64-yr-old patient who developed a second glioblastoma in the left frontal lobe 10 yr after surgical resection of a glioblastoma of right frontal lobe. The first tumor contained 2 p53 mutations, in codon 213 (CGA-->TGA, Arg-->stop) and codon 306 (CGA-->TGA, Arg-->stop), further, 1 missense PTEN mutation (codon 257, TTC-->TTA, Phe-->Leu) and a silent PTEN mutation (codon 154, TTC-->TTT, Phe-->Phe). The second glioblastoma also contained multiple, but different mutations: p53 mutations in codons 158 (CGC-->CAC, Arg-->His) and 273 (CGT-->TGT, Arg-->Cys), and a PTEN mutation in codon 233 (CGA-->TGA, Arg-->Stop). Both neoplasms had a homozygous p16 deletion. The discordant pattern of mutations indicates that the second glioblastoma was not a recurrence but an independent second glioblastoma. The presence in these neoplasms of multiple mutations in tumor suppressor genes suggests the involvement of a novel disease mechanism but there was no indication of a DNA mismatch repair deficiency or of an inherited tumor syndrome.