Recombinant Reporter Phage rTUN1::nLuc Enables Rapid Detection and Real-Time Antibiotic Susceptibility Testing of Klebsiella pneumoniae K64 Strains.

The emergence of multi-drug-resistant Klebsiella pneumoniae (Kp) strains constitutes an enormous threat to global health as multi-drug resistance-associated treatment failure causes high mortality rates in nosocomial infections. Rapid pathogen detection and antibiotic resistance screening are therefore crucial for successful therapy and thus patient survival. Reporter phage-based diagnostics offer a way to speed up pathogen identification and resistance testing as integration of reporter genes into highly specific phages allows real-time detection of phage replication and thus living host cells. Kp-specific phages use the host's capsule, a major virulence factor of Kp, as a receptor for adsorption. To date, 80 different Kp capsule types (K-serotypes) have been described with predominant capsule types varying between different countries and continents. Therefore, reporter phages need to be customized according to the locally prevailing variants. Recently, we described the autographivirus vB_KpP_TUN1 (TUN1), which specifically infects Kp K64 strains, the most predominant capsule type at the military hospital in Tunis (MHT) that is also associated with high mortality rates. In this work, we developed the highly specific recombinant reporter phage rTUN1::nLuc, which produces nanoluciferase (nLuc) upon host infection and thus enables rapid detection of Kp K64 cells in clinical matrices such as blood and urine. At the same time, rTUN1::nLuc allows for rapid antibiotic susceptibility testing and therefore identification of suitable antibiotic treatment in less than 3 h.

Occupational physicians dealing with mental health: between employee and company interests: a qualitative study.

BACKGROUND: Occupational health physicians are increasingly confronted with mental health issues at their workplace. Facing them, most of them feel insecure and not sufficiently trained. Employee's mental well-being depends at the same time on individual and significantly on organizational variables. This complicates the physician's position, since they have to serve many interests. The focus of the present study is to investigate what difficulties occupational health physicians face and how organizational culture and management influence their work. METHODS: Interviews were conducted with N = 25 physicians as part of a training for basic mental health care. Interviews were interpreted using qualitative content analysis. RESULTS: Working with mentally ill employees was difficult for the physicians interviewed. Many felt insecure managing and preventing mental health issues. A need for further education was observed. Environmental factors (organizational culture, management) have a strong impact on the work of an occupational health physician and highlight its systemic dimension. Even though many of our participants report a meanwhile more open attitude towards mental disorders at their workplace, on the level of direct contact to the management prevail descriptions of little acceptance and a high priority of economic outcomes. CONCLUSIONS: More education on topics of mental health is needed for occupational health physicians. Future trainings should consider the intertwined nature of their work and enable them in dealing consciously with other actors in the company. For enhancing employee's mental well-being occupational health physicians could be granted a strengthened position in companies or be supported through more exchange with colleagues in other companies.

Psychische Erkrankungen im Arbeitskontext ­ eine Verortung im Versorgungssystem.

Work and mental health are closely interrelated. Sick leave rates due to mental illness have constantly been rising for years. Thus, not only companies but also the society as a whole have become more and more aware of the relevance of this topic.At first, various work stress models are outlined in this article. This is followed by an overview of how to classify work-related mental disorders in the ICD and DSM systems. Burnout is explained with regard to terminology, origin and adequate use in clinical practice. Furthermore, the ILO (International Labour Organization) classification of work-related mental disorders is presented. Possible intervention models for the prevention and therapy of work-related mental health problems are discussed. In addition, the most essential components of work-related psychotherapy are pointed out. Finally, corresponding guidelines are put into an international context.

Characterization of distribution of pesticide residues in crop units.

The characteristic features of distribution of pesticide residues in crop units and single sample increments were studied based on more than 19,000 residue concentrations measured in root vegetables, leafy vegetables, small-, medium- and large-size fruits representing 20 different crops and 46 pesticides. Log-normal, gamma and Weibull distributions were found to provide the best fit for the relative frequency distributions of individual residue data sets. The overall best fit was provided by lognormal distribution. The relative standard deviation of residues (CV) in various crops ranged from 15-170%. The 100-120 residue values being in one data set was too small to identify potential effects of various factors such as the chemical and physical properties of pesticides and the nature of crops. Therefore, the average of CV values, obtained from individual data sets, were calculated and considered to be the best estimate for the likely variability of unit crop residues for treated field (CV = 0.8) and market samples (CV = 1.1), respectively. The larger variation of residues in market samples was attributed to the potential mixing of lots and varying proportion of non-detects. The expectable average variability of residues in composited samples can be calculated from the typical values taking into account the sample size.

Assessment of skeletal muscle microcirculation in type 2 diabetes mellitus using dynamic contrast-enhanced ultrasound: a pilot study.

PURPOSE: To investigate muscular micro-perfusion by employing dynamic contrast-enhanced ultrasound (CEUS) and performing transient arterial occlusion in patients with type 2 diabetes mellitus (DM-2). METHODS: Twenty DM-2 patients (mean age, 58 ± 8.6 years; duration of diabetes, 15.4 ± 12.1 years) and 20 healthy volunteers (mean age, 54 ± 5.4 years) participated. CEUS was applied to the calf, while 4.8 mL of SonoVue(®) was injected intravenously. At the thigh level, arterial occlusion (60 s) was performed. CEUS parameters (tmax, max, AUCpost and m) were evaluated and Pearson-product-moment correlation coefficients were computed. RESULTS: A moderate negative correlation of HbA1c and max was established (-0.53). Max in patients with DM-2 >10 years was 79.89 ± 37.4. Max in patients with DM-2 duration <10 years was 137.62 ± 71.72 (p = 0.04). AUCpost in patients with DM-2 duration >10 years was 3924.01 ± 1630.52. AUCpost in patients with DM-2 duration <10 years was 6453.59 ± 3206.23 (p = 0.04). CONCLUSION: Patients with long history of DM-2 present with impaired muscular perfusion. CEUS and transient arterial occlusion may provide appropriate methods for semi-quantitative evaluation of muscular micro-perfusion in patients with DM-2.

Dynamic contrast-enhanced ultrasound for assessment of skeletal muscle microcirculation in peripheral arterial disease.

OBJECTIVE: : This feasibility study was performed to assess whether dynamic contrast-enhanced ultrasound (CEUS) and transient arterial occlusion are able to detect alterations in the microvascular perfusion and arterial perfusion reserve in patients suffering from peripheral arterial disease (PAD) in comparison with healthy volunteers. MATERIALS AND METHODS: : Twenty patients with PAD, Rutherford classification grade I, category III (mean age, 64 years; mean height, 173 cm; mean weight, 81.8 kg), and 20 volunteers (mean age, 50 years; mean height, 174 cm; mean weight, 77.8 kg) participated in the study. Low-mechanical index CEUS (7 MHz; MI, 0.28) was performed to the dominant lower leg after start of a continuous automatic intravenous injection of 4.8 mL suspension with microbubbles containing sulfur hexafluoride (SonoVue) within 5 minutes. Perfusion of the calf muscle was monitored by CEUS before, during, and after release of arterial occlusion at the thigh level lasting for 60 seconds. Several parameters, especially the time to maximum enhancement after release of occlusion (tmax), the maximum enhancement after release of occlusion (maxenh), the total vascular response after release of occlusion (AUCpost), and the resulting slope (m2) to maximum enhancement were calculated. RESULTS: : After release of the occlusion, a significantly delayed increase of the CEUS signal to maxenh was observed in the patients with PAD (32 ± 17 seconds) compared with volunteers (17 ± 8 seconds, P = 0.0009). maxenh was 66.5 ± 36.6 (∼mL) in PAD versus 135.6 ± 75.1 (∼mL) in volunteers (P = 0.0016). AUCpost was 3016.5 ± 1825.8 (∼mL·s) in PAD versus 5906.4 ± 3173.1 (∼mL·s) in volunteers (P = 0.0013), and m2 was significantly lower in PAD (3.8 ± 5.2 vs. 14.8 ± 9.7 [∼mL/s], P = 0.0001). CONCLUSIONS: : Microvascular perfusion deficits and reduced arterial perfusion reserve in patients with PAD are clearly detectable with dynamic CEUS after transient arterial occlusion.

Analyzing fractal dynamics employing R.

Recent empirical studies from cognitive, social and biological psychology revealed the fractal properties of many psychological phenomena. Employing methodologies from time- and frequency-domain analyses enabled detecting persistent long-range dependencies in various psychological and behavioral time series. These very slowly decaying autocorrelations are known as 1/f noise and typical for self-similar long memory processes. This paper evaluated different estimators of long memory parameters commonly available in the open source statistical software R concerning their ability to distinguish between fractional Brownian motions and fractional Gaussian noises, stationary and nonstationary fractal processes, short and long memory series. The following procedures implemented in the R packages fractal and fracdiff were considered: PSD (hurstSpec), DFA, the Whittle method (FDWhittle), semiparametric estimators of Reisen (fdSperio) and Geweke & Porter-Hudak (fdGPH) as well as the approximate ML algorithm of Haslett and Raftery (fracdiff). The key finding of the study was that the performance of the methods strongly depends on the complexity of the underlying process and parameterizations. Since in empirical settings the true structure is never known, an elaborated strategy for the estimation of the long memory parameter d combining different techniques was developed and demonstrated on an empirical example.

DNA-binding properties of Drosophila ecdysone receptor isoforms and their modification by the heterodimerization partner ultraspiracle.

Transcriptional activity of ecdysone receptor (EcR) isoforms varies considerably and is modified further by the heterodimerization partner and hormone treatment. To investigate whether differences in DNA binding of receptor complexes are responsible for these variations in transcriptional activity, interaction of Drosophila EcR isoforms, and variants of Ultraspiracle (Usp), the orthologue of RXR, with the ecdysone response elements (EcRE) hsp 27, PAL-1, and DR-1, were determined by electrophoretic mobility shift assays. Receptor proteins were expressed in vertebrate cells (CHO-K1) in order to rule out an influence of endogenous receptor proteins. In the absence of a heterodimerization partner, weak DNA binding of EcR was detected even without hormone with EcR-A and -B1, but not EcR-B2. In the presence of hormone, all three isoforms show increased binding to the hsp 27 EcRE. The heterodimerization partner Usp increased DNA binding considerably. The hormone effect of heterodimers is more pronounced with both EcR-B isoforms compared to EcR-A. Two specific bands were obtained for EcR-A and B1 but only one band is visible with EcR-B2. Deletion of the C-domain of Usp still allows basal DNA binding of the heterodimer, but in contrast to full-length Usp, addition of hormone decreases the intensity of the retarded receptor band of all EcR isoforms and the EcREs hsp27 and DR-1 considerably, whereas interaction with the EcRE PAL-1 is only slightly affected. Synergistic effects on transcriptional activity are associated with the formation of different receptor DNA-complexes observed with 1xhsp27 and 3xhsp27. Comparison of DNA-binding properties of EcR isoforms and EcR/Usp heterodimers revealed that binding of receptor complexes to hsp 27 EcRE is dependent on the AB domain of EcR and the AB-, C-, and D-domains of the heterodimerization partner. Interaction with the hsp 27 EcRE correlates neither with ligand binding nor with transcriptional activity of the various receptor complexes. We, therefore, conclude that the different receptor functions are regulated separately, for example, by interaction with co-modulators or post-transcriptional modifications.

Estimating long-range dependence in time series: an evaluation of estimators implemented in R.

Recent studies have shown that many physiological and behavioral processes can be characterized by long-range correlations. The Hurst exponent H of fractal analysis and the fractional-differencing parameter d of the ARFIMA methodology are useful for capturing serial correlations. In this study, we report on different estimators of H and d implemented in R, a popular and freely available software package. By means of Monte Carlo simulations, we analyzed the performance of (1) the Geweke-Porter-Hudak estimator, (2) the approximate maximum likelihood algorithm, (3) the smoothed periodogram approach, (4) the Whittle estimator, (5) rescaled range analysis, (6) a modified periodogram, (7) Higuchi's method, and (8) detrended fluctuation analysis. The findings-confined to ARFIMA (0, d, 0) models and fractional Gaussian noise-identify the best estimators for persistent and antipersistent series. Two examples combining these results with the step-by-step procedure proposed by Delignières et al. (2006) demonstrate how this evaluation can be used as a guideline in a typical research situation.

Comparison of automated procedures for ARMA model identification.

This article evaluates the performance of three automated proceduresfor ARMA modelidentification commonly available in current versions of SAS for Windows: MINIC, SCAN, and ESACF. Monte Carlo experiments with different model structures, parameter values, and sample sizes were used to compare the methods. On average, the procedures either correctly identified the simulated structures or selected parsimonious nearly equivalent mathematical representations in at least 60% of the trials conducted. For autoregressive models, MINIC achieved the best results. SCAN was superior to the other two procedures for mixed structures. For moving-average processes, ESACF obtained the most correct selections. For all three methods, model identification was less accurate for low dependency than for medium or high dependency processes. The effect of sample size was more pronounced for MINIC than for SCAN and ESACE SCAN and ESACF tended to select higher-order mixed structures in larger samples. These findings are confined to stationary nonseasonal time series.

Model Identification of Integrated ARMA Processes.

This article evaluates the Smallest Canonical Correlation Method (SCAN) and the Extended Sample Autocorrelation Function (ESACF), automated methods for the Autoregressive Integrated Moving-Average (ARIMA) model selection commonly available in current versions of SAS for Windows, as identification tools for integrated processes. SCAN and ESACF can be applied to either nontransformed or differenced series, so the advantages and drawbacks of both procedures were compared. The best results were 79% of correct identifications for SCAN and 80% for ESACF. For some models and parameterizations, the accuracy of SCAN and ESACF was disappointing. The key finding of the study is that both human experts and automated methods provide inconsistent model identifications. Hence an elaborated strategy for model selection combining different techniques was developed and demonstrated on 2 empirical examples.

Active succinate dehydrogenase (SDH) and lack of SDHD mutations in sporadic paragangliomas.

BACKGROUND: Paragangliomas are benign, slow-growing tumours of the head and neck region. The candidate gene for familial and some sporadic paragangliomas, SDHD (succinate dehydrogenase, subunit D), has been mapped to the PGL1 locus in 11q23.3. MATERIALS AND METHODS: Normal and tumour DNA of 17 patients with sporadic paragangliomas were analysed by sequencing (SDHD, SDHB and SDHC genes), fluorescence in situ hybridisation (FISH). In addition, loss of heterozygosity (LOH) and succinate dehydrogenase (SDH) enzyme activity assays were performed. RESULTS AND CONCLUSION: Only two patients from our collective showed SDH gene mutations, one in SDHD and one in SDHB, respectively. Moreover, SDH activity detected in 5/8 patients confirmed the fact that SDH inactivation is not a major event in sporadic paragangliomas. LOH and FISH analysis demonstrated a frequent loss of regions within chromosome 11, indicating that additional genes in 11q may play a role in tumour genesis of sporadic paragangliomas.

GJB2 mutations in patients with non-syndromic hearing loss from Northeastern Hungary.

Mutations in the GJB2 gene encoding the gap-junction protein connexin 26 have been identified in many patients with childhood hearing impairment (HI). One single mutation, c.35delG, accounts for the majority of mutations in Caucasian patients with HI. In the present study we screened 500 healthy control individuals and a group of patients with HI from Northeastern Hungary for GJB2 mutations. The patients' group consisted of 102 familial from 28 families and 92 non-familial cases. The most common mutation in the Hungarian population is the c.35delG, followed by the c.71G>A (p.W24X) mutation. 34.3% of the patients in the familial group were homozygous, and 17.6% heterozygous for 35delG. In the non-familial group the respective values were 37% and 18% (allele frequency: 46.2%). In the general population an allele frequency of 2.4% was determined. Several patients were identified with additional, already described or new GJB2 mutations, mostly in heterozygous state. The mutation c.380G>A (p.R127H) was formerly found only in heterozygous state and its disease relation was controversial. We demonstrated the presence of this mutation in a family with three homozygous patients and 4 heterozygous unaffected family members, a clear indication of recessively inherited HI. Furthermore, we provided evidence for the pathogenic role of two new mutations, c.51C>A (p.S17Y) and c.177G>T (p.G59V), detected in the present study. In the latter case the pattern of inheritance might be dominant. Our results confirm the importance of GJB2 mutations in the Hungarian population displaying mutation frequencies that are comparable with those in the Mediterranean area.

Chromosome 11 monosomy in conjunction with a mutated SDHD initiation codon in nonfamilial paraganglioma cases.

Paragangliomas of the head and neck region are a group of rare, usually benign, slow-growing tumors developing from paraganglionic chemoreceptors in most patients. Mutations in a subunit of the mitochondrial enzyme II complex (succinate dehydrogenase [SDHD]) were shown to be responsible for the formation of paragangliomas. In addition, loss of heterozygosity (LOH) on chromosome 11, mainly in 11q23 (PGL1), was observed recently. We analyzed DNA derived from tumor sections of three unrelated paraganglioma patients (one case with multiple paragangliomas, two cases with single tumors; all of them sporadic cases) for mutations in the SDHD gene by direct sequencing. Microsatellite-based LOH was performed, and events of chromosomal loss were validated by fluorescence in situ hybridization (FISH) on paraffin-embedded tumor and normal tissue by using centromeric satellite DNA. Sequence analysis revealed mutations in SDHD exon 1 in all patients, affecting the initiation codon (M1V). Another alteration was detected in exon 2 but was lacking in tumor DNA and therefore classified as polymorphism (H50R). LOH and FISH analyses demonstrated partial/total monosomy for chromosome 11 in the tumor samples tested. A common genetic mechanism appears to be the pathophysiologic basis for sporadic tumor development because the proposed two-hit model comprising both LOH and point mutation is manifest in our patients. Loss of chromosome 11 regions, including the deletion of PGL1 and PGL2 loci, may result in a more severe phenotype, as exemplified by the development of multiple tumors in one of the patients.

Frequencies of GJB2 mutations in German control individuals and patients showing sporadic non-syndromic hearing impairment.

Mutations in the GJB2 gene encoding the gap-junction protein connexin 26 have been identified in many patients with childhood hearing impairment (HI). One single mutation, 35delG (30delG), accounts for up to 70% of all analyzed European patients with autosomal recessive inherited HI and 10% of patients with HI of unknown origin, respectively. We screened 188 control individuals and 342 German patients with non-syndromic sporadic HI for the 35delG, compound heterozygosity and other GJB2 mutations by PCR, restriction enzyme based screening, SSCP and sequencing. In all patients, non-progressive hearing impairment varied from moderate to profound involving all frequencies. This study revealed one novel silent mutation (438C/T), three novel gene variants resulting in amino acid substitutions (K112E, T123S, K223R) and two novel HI-related mutations (I82M, 313del14).