Genomic and phenotypic diversity of Clostridium difficile during long-term sequential recurrences of infection.

Infection with the emerging pathogen Clostridioides (Clostridium) difficile might lead to colonization of the gastrointestinal tract of humans and mammals eventually resulting in antibiotic-associated diarrhea, which can be mild to possibly life-threatening. Recurrences after antibiotic treatment have been described in 15-30% of the cases and are either caused by the original (relapse) or by new strains (reinfection). In this study, we describe a patient with ongoing recurrent C. difficile infections over 13 months. During this time, ten C. difficile strains of six different ribotypes could be isolated that were further characterized by phenotypic and genomic analyses including motility and sporulation assays, growth fitness and antibiotic susceptibility as well as whole-genome sequencing. PCR ribotyping of the isolates confirmed that the recurrences were a mixture of relapses and reinfections. One recurrence was due to a mixed infection with three different strains of two different ribotypes. Furthermore, genomes were sequenced and multi-locus sequence typing (MLST) was carried out, which identified the strains as members of sequence types (STs) 10, 11, 14 and 76. Comparison of the genomes of isolates of the same ST originating from recurrent CDI (relapses) indicated little within-patient microevolution and some concurrent within-patient diversity of closely related strains. Isolates of ribotype 126 that are binary toxin positive differed from other ribotypes in various phenotypic aspects including motility, sporulation behavior and cell morphology. Ribotype 126 is genetically related to ribotype 078 that has been associated with increased virulence. Isolates of the ribotype 126 exhibited elongated cells and a chaining phenotype, which was confirmed by membrane staining and scanning electron microscopy. Furthermore, this strain exhibits a sinking behavior in liquid medium in stationary growth phase. Taken together, our observation has proven multiple CDI recurrences that were based on a mixture of relapses and reinfections.

Steep rise in norovirus cases and emergence of a new recombinant strain GII.P16-GII.2, Germany, 2016.

Since early November 2016, the number of laboratory-confirmed norovirus infections reported in Germany has been increasing steeply. Here, we report the detection and genetic characterisation of an emerging norovirus recombinant, GII.P16-GII.2. This strain was frequently identified as the cause of sporadic cases as well as outbreaks in nine federal states of Germany. Our findings suggest that the emergence of GII.P16-GII.2 contributed to rising case numbers of norovirus gastroenteritis in Germany.

Natural zeolites in diet or litter of broilers.

This study aims to analyse the influence of adding natural zeolites (clinoptilolite) to the diet or litter of broilers and their effects on growth performance, carcass yield and litter quality. Three consecutive flocks of broilers were raised on the same sawdust litter, from d 1 to d 42 of age, and distributed in three treatments (control with no added zeolites, addition of 5 g/kg zeolite to diet and addition of 100 g/kg zeolites to litter). The addition of zeolites to the diet or litter did not affect growth performance or carcass yield. The addition of zeolites to the diet did not influence moisture content of the litter, ammonia volatilisation was reduced only in the first flock and pH of litter was reduced in the second and third flock. However, the addition of zeolites to the litter reduced moisture content, litter pH and ammonia volatilisation in all flocks analysed. The addition of 5 g/kg zeolite to the diet in three consecutive flocks was not effective in maintaining litter quality, whereas the addition of 100 g/kg natural zeolites to sawdust litter reduced litter moisture and ammonia volatilisation in three consecutive flocks raised on the same litter.

Clostridium difficile infections among adults and children in Mwanza/Tanzania: is it an underappreciated pathogen among immunocompromised patients in sub-Saharan Africa?

Little is known regarding the epidemiology Clostridium difficile in developing countries. Fresh stool samples from patients with diarrhoea were cultured anaerobically. C. difficile was detected in nine (6.4%) of 141 (95% confidence interval 4.2-13.1), of which seven (77.8%) were from children. HIV infection, prolonged hospitalization and antibiotic use were independent factors associated with the occurrence of C. difficile in the gastrointestinal tract. Two of the toxigenic isolates were typed as ribotype 045, and the other two had unknown ribotype. All C. difficile isolates were susceptible to metronidazole, moxifloxacin and clarithromycin, while three isolates were resistant to clarithromycin. C. difficile may be an important pathogen causing diarrhoea in sub-Saharan Africa among immunocompromised patients.

[Endophthalmitis, liver abscess, and cerebral and pulmonary emboli in a 48-year-old Vietnamese man]. / Endophthalmitis, Leberabszess sowie zerebrale und pulmonale Embolien bei einem 48-jährigen Vietnamesen.

We present the case of a 48-year-old Vietnamese man with endophthalmitis, liver abscess, and pulmonary and cerebral septic emboli. Klebsiella pneumoniae was isolated as the causative organism; there were no laboratory findings suggestive of invasive fungal infection. Klebsiella pneumoniae invasive syndrome is a rare disease in Germany. This case report exemplifies the necessity of a dedicated diagnostic approach that takes into consideration factors such as ethnic origin and accompanying diseases of the patient.

GMP-adapted overexpression of CXCR4 in human mesenchymal stem cells for cardiac repair.

BACKGROUND: Human mesenchymal stem cells (MSC) have been utilized for cardiac regeneration after myocardial damage. Their clinical effects are marginal and only a minority of administered cells could make their way into the myocardium. The chemokine receptor CXCR4 has been identified as crucial for migration and homing of stem cells. In this study we overexpressed CXCR4 on human MSC to improve cell trafficking and tissue repair. METHODS: Human MSC were isolated from the spongiosa of tibia and femur as well as from pelvic bone marrow. MSC were characterized by differentiation assays and FACS analysis. CXCR4 was overexpressed by mRNA-nucleofection. Intracellular signaling was analyzed to demonstrate functionality of CXCR4. The modified Boyden chamber, wounding assays and time lapse microscopy were utilized to investigate MSC migration. RESULTS: MSC did not express relevant amounts of CXCR4 spontaneously. CXCR4 could be overexpressed in 93% of MSC with a cell viability of 62%. Functionality of the overexpressed CXCR4 was demonstrated by a significant cytosolic Ca(2+) increase and activation of different MAP kinases followed by SDF-1α stimulation. In contrast no improvement of cell migration could be observed. There was a strong basal MSC chemokinesis independent from CXCR4 expression. CONCLUSIONS: CXCR4 could be effectively overexpressed in human MSC by mRNA-nucleofection. Despite functionality of CXCR4 MSC were characterized by a strong basal chemokinesis that could not be further enhanced by CXCR4 overexpression. As isolation, culture and nucleofection of pelvic bone marrow-derived MSC basically fulfill the GMP-requirements our approach seems suited for an in vivo application in patients.

Atrial fibrillation-induced cardiac shock: first manifestation of a congenitally corrected transposition of the great arteries in a 45-year-old man.

Background. The congenitally corrected transposition of the great arteries (L-TGA) is a very rare congenital heart defect, which often remains undetected for several decades of life. Case Presentation. We report on a 45-year-old man without prior history of heart disease, presenting with cardiac shock related to a first episode of tachycardic atrial fibrillation. The diagnostic work-up identified a L-TGA as the underlying cause for acute heart failure. Discussion. L-TGA is a very rare congenital heart defect, which is characterized by an atrioventricular as well as a ventriculoarterial discordance. By this means, the physiological sequence of pulmonary and systemic circulation is still maintained. On the basis of an ongoing strain of the right ventricle, which has to carry the burden of the systemic blood pressure, after more than four decades without symptoms, acute heart failure was triggered by a tachycardic atrial fibrillation.

Ring polymer simulations with global radius of curvature.

We simulate three-dimensional flexible off-lattice ring polymers of length L up to L=4000 for various values of the global radius of curvature Rgrc=0.25 , 0.48, and 1.0 and Rgrc=2.0 . We utilize two different ensembles: one with a delta -function constraint on the radius, and the other with a theta -function. For both cases the global radius of curvature provides a valid regularization of polymers with thickness D=2Rgrc . The Flory-type critical exponent nu SAW of self-avoiding rings at D=2 is found to be nu SAW=0.5869(5) from the radii of gyration chain length scaling, while other D values produce consistent results. For our current implementation, the numerical effort of chain thickness calculations is bounded by a number O(LlnL) per single update. We also study low-temperature configurations of spatially dense Lennard-Jones homopolymers on a ring and identify some conformational building blocks.

Active control of type-I edge-localized modes with n=1 perturbation fields in the JET tokamak.

Type-I edge-localized modes (ELMs) have been mitigated at the JET tokamak using a static external n=1 perturbation field generated by four error field correction coils located far from the plasma. During the application of the n=1 field the ELM frequency increased by a factor of 4 and the amplitude of the D(alpha) signal decreased. The energy loss per ELM normalized to the total stored energy, DeltaW/W, dropped to values below 2%. Transport analyses shows no or only a moderate (up to 20%) degradation of energy confinement time during the ELM mitigation phase.

Improved confinement due to open ergodic field lines imposed by the dynamic ergodic divertor in TEXTOR.

The ergodization of the magnetic field lines imposed by the dynamic ergodic diverter (DED) in TEXTOR can lead both to confinement improvement and to confinement deterioration. The cases of substantial improvement are in resonant ways related to particular conditions in which magnetic flux tubes starting at the X points of induced islands are connected with the wall. This opening process is connected with a characteristic modification of the heat deposition pattern at the divertor target plate and leads to a substantial increase and steepening of the core plasma density and pressure. The improvement is tentatively attributed to a modification of the electric potential in the plasma carried by the open field lines. The confinement improvement bases on a spontaneous density built up due to the application of the DED and is primarily a particle confinement improvement.

Forced magnetic reconnection and field penetration of an externally applied rotating helical magnetic field in the TEXTOR tokamak.

The magnetic field penetration process into a magnetized plasma is of basic interest both for plasma physics and astrophysics. In this context special measurements on the field penetration and field amplification are performed by a Hall probe on the dynamic ergodic divertor (DED) on the TEXTOR tokamak and the data are interpreted by a two-fluid plasma model. It is observed that the growth of the forced magnetic reconnection by the rotating DED field is accompanied by a change of the plasma fluid rotation. The differential rotation frequency between the DED field and the plasma plays an important role in the process of the excitation of tearing modes. The momentum input from the rotating DED field to the plasma is interpreted by both a ponderomotive force at the rational surface and a radial electric field modified by an edge ergodization.

Properties of geodesic acoustic modes and the relation to density fluctuations.

The geodesic acoustic mode (GAM) is a high frequency branch of zonal flows, which is observed in toroidal plasmas. Because of toroidal curvature effects, density fluctuations are excited, which are investigated with the O-mode correlation reflectometer at TEXTOR. This Letter reports on the poloidal distribution of GAM induced density fluctuation and compares them with theoretical predictions. The influence of the GAM flows on the ambient turbulence is studied, too.

The ability of selected oral microorganisms to emit red fluorescence.

Some novel caries detection and excavation devices rely on the ability of bacteria to produce red fluorescing compounds. The aim of this study was to examine the ability of selected oral microorganisms to emit red fluorescence. Streptococcus mutans, S. oralis, S. salivarius, S. sobrinus, Lactobacillus fermentans, L. casei, L. rhamnosus, Actinomyces naeslundi, A. israelii, Prevotella intermedia, and Fusobacterium nucleatum were inoculated onto Columbia agar with haemin and vitamin K and incubated anaerobically for up to 7 days in the dark. The resulting bacterial colonies were excited using filtered xenon light (405 +/- 20 nm) and digitally photographed through a 530-nm high-pass filter. The red and green portions of the colony fluorescence were analyzed using a computer program and the red/green ratio was calculated. All colonies emitted both red and green fluorescence. The green outweighed the red portion for the following species (in descending order) S. oralis, S. salivarius, S. mutans, F. nucleatum and S. sobrinus. The red portion was higher for the following species (in descending order) P. intermedia, A. naeslundi, A. israelii, L. fermentans, L. rhamnosus and L. casei. With all the bacteria examined, one color portion generally outweighed the other, giving the visual impression of either red or green fluorescence. We conclude that red fluorescence is well suited to detection of the bacteria which cause dentin caries but it is not suitable as an indicator of the presence and activity of the streptococci involved in initial caries.

Labeling of adult stem cells for in vivo-application in the human heart.

Tissue regeneration with human hematopoietic or mesenchymal stem cells has become a fashionable research topic. In cardiology, intracoronary injection of adult stem cells has already been used for the treatment of human myocardial infarction and ischemic cardiomyopathy. The experimental background of such therapies, however, i.e. the potential of adult stem cells to regenerate myocardium through "transdifferentiation" of hematopoietic or mesenchymal stem cells into cardiomyocytes described in animal models, has recently been challenged by other experimental data. Nonetheless, clinical trials are continuing. This may be due to the fact that, in open-labeled pilot trials, a benefit of intracoronary injection of adult stem cells for the treatment of myocardial infarction has been described. As pilot trials may overemphasize the beneficial effects of intracoronary injection of bone marrow stem cells, controlled double-blinded randomised multicenter studies are warranted. Furthermore, a careful characterization of the cells involved in the proposed cardiac repair as well as in vivo-monitoring of such cells following intracoronary injection in humans might help to answer many essential questions linked to this important research topic. The latter requires biocompatible labeling. This review focuses on the technologies available for stem cell labeling and summarizes the arguments and contra-arguments to use these labeling technologies for application in humans.

Influence of the dynamic ergodic divertor on the density limit in TEXTOR.

A significant influence of the dynamic ergodic divertor (DED) on the density limit in TEXTOR has been found. In Ohmic discharges, where without DED detachment normally arises at the density limit, a MARFE (multifaceted asymmetric radiation from the edge) develops when the DED is operated in a static regime. The threshold of the MARFE onset in the neutral beam heated plasmas is increased by applying 1 kHz ac DED at the high-field side. The theoretical predictions based on the parallel energy balance taking poloidal asymmetries into account agree well with the experimental observation.

Toroidal plasma rotation induced by the dynamic ergodic divertor in the TEXTOR tokamak.

The first results of the Dynamic Ergodic Divertor in TEXTOR, when operating in the m/n=3/1 mode configuration, are presented. The deeply penetrating external magnetic field perturbation of this configuration increases the toroidal plasma rotation. Staying below the excitation threshold for the m/n=2/1 tearing mode, this toroidal rotation is always in the direction of the plasma current, even if the toroidal projection of the rotating magnetic field perturbation is in the opposite direction. The observed toroidal rotation direction is consistent with a radial electric field, generated by an enhanced electron transport in the ergodic layers near the resonances of the perturbation. This is an effect different from theoretical predictions, which assume a direct coupling between rotating perturbation and plasma to be the dominant effect of momentum transfer.

Dimerisation mutants of Lac repressor. I. A monomeric mutant, L251A, that binds Lac operator DNA as a dimer.

Dimer formation between monomers of the Escherichia coli Lac repressor is substantially specificed by the interactions between three alpha-helices in each monomer which form a hydrophobic interface. As a first step in analysing the specificity of this interaction, we examined the mutant L251A. LacR bearing this mutation in a background lacking the C-terminal heptad repeats is completely incapable of forming dimers in solution, with a dimer-monomer equilibrium dissociation constant, or Kd, higher than 10(-5)M. This correlates with a 200-fold decrease in its ability to repress the lac operon in vivo compared to dimeric LacR. Surprisingly, the mutant is still capable of forming dimers upon binding to short operator DNA in vitro. Analysis of the kinetic parameters of binding of the mutant to operator DNA reveals a 2000 to 3000-fold increase in the equilibrium dissociation constant (Kd) of the mutant-DNA complex in comparison to dimeric LacR-operator complexes, with the change almost entirely due to a greater than 1000-fold decrease in association rate. The dissociation rate varies only by a factor of about two, in comparison to dimeric LacR. This change reflects a kinetic pathway in which dimer formation, in solution or on DNA, is the rate-limiting step. These findings have implications for the specificity and stability of the protein-protein interface in question.

Corynebacterium thomssenii sp. nov., a Corynebacterium with N-acetyl-beta-glucosaminidase activity from human clinical specimens.

A strain of a previously undescribed non-lipophilic coryneform bacterium was isolated from pleural fluids of a patient with chronic renal failure, stroke and pneumonia. Slow fermentative acid production from glucose, maltose and sucrose, and strong N-acetyl-beta-glucosaminidase activity were the most characteristic features of the bacterium. Chemotaxonomic characterization unambiguously indicated that the organism belonged to the genus Corynebacterium. The results of comparative 16S rRNA gene sequence analysis revealed that the isolate represented a new species within the genus, for which the name Corynebacterium thomssenii sp. nov. is proposed. The type strain is DSM 44276.