Diversität von psychrophilen sulfatreduzierenden und thermophilen sporenbildenden Mikroorganismen im arktischen Meeressediment vor Spitzbergen

Mehr als 90 % des Meeresbodens weist eine Temperatur von unter 4 ° C auf und stellt somit einen wichtigen Lebensraum für marine Mikroorganismen dar. Durch DsrAB-Stammbaumanalyse konnte ein Einblick in die Diversität von sulfatreduzierenden Mikroorganismen (SRM) in diesem Lebensraum gewonnen werden. Der Vergleich zweier mariner Standorte vor der Westküste von Svalbard ergab eine unterschiedliche phylogenetische Zusammensetzung. So bildeten die Mehrheit der gefundenen OTUs (67 % der Sequenzen) des Sediment des nördlicher gelegenen Smeerenburgfjorden (Station J) eine tiefzweigende, monophyletische Gruppe, welche keine Verwandtschaft zu kultivierbaren SRM aufwies, während der Großteil der erhaltenen OTUs (52 % der Sequenzen), isoliert aus dem Sediment von van Mijenfjorden (Station AH) den Deltaproteobacteria zugeordnet werden konnte. Ein weiteres Ziel dieser Arbeit war die Durchführung einer 16S-rRNS-Stammbaumanalyse sowie die Quantifizierung von sporenbildenden, thermophilen Mikroorganismen in arktischem Sediment, isoliert aus zwei Inkubationsreihen bei 50 °C mit, bzw. ohne initialer Zugabe von organischen Substraten. Die Quantifizierung von thermophilen SRM zeigte für beide Inkubationen nach einer anfänglichen Erhöhung der Gen-Abundanz, eine gleichbleibende Kopienzahl der DNS-Matrize für die restliche Inkubation. Die initiale Substratzugabe schien dabei keinen Einfluss auf das Wachstum der SRM, aber auf ihre metabolische Aktivität zu haben. Anhand der phylogenetische Analyse konnte die Mehrheit der Sequenzen (54 %), welche mit allgemeinen 16S-rRNS-Primern amplifiziert wurden, der Klasse der Clostridia zugeordnet werden. 20 % aller Sequenzen gruppierten dabei innerhalb der Gattung Desulfotomaculum, welche Vertreter beinhaltet, die in der Lage sind durch Endosporen extreme Bedingungen zu überdauern. Des Weiteren wurden Sequenzen identifiziert, welche eine hohe Sequenzähnlichkeit zu Vertretern der Gattungen Tepidibacter, Caloranaerobacter und Garciella aufwiesen. Diese Linien beinhalten thermophile Organismen, welche Kohlenhydrate und Proteine zu kurzkettigen freien Fettsäuren abbauen und somit SRM Elektronendonoren zu Verfügung stellen. Einige Vertreter dieser Linien sind mit Ölreservoirs oder heißen marinen Quellen assoziiert, welche möglicherweise den Ursprung der thermophilen Organismen in diesem Habitat darstellen.Since 90 % of the marine sediment has a temperature below 4 °C it is a very important habitat for microorganisms. It was possible to gain insight into the diversity of sulfate reducing bacteria (SRB) in this habitat by analysing DsrAB-sequences. The comparison of two different fjord sediments on the west coast of Svalbard in the Arctic showed differences in the phylogeny of SRB. Most (67 %) of the sequences obtained from the northern Smeerenburgfjorden sediment (station J), showed no affinity to cultivated SRB and generated a monophyletic, deeply branching group within the tree. However, the majority the sequences (52 %) obtained from sediment of the southern van Mijenfjorden (Station AH) were related to the Deltaproteobacteria. Another aim of this study was the phylogenetic analysis and quantification of thermophilic, spore-forming microorganisms obtained from two incubations of arctic sediment at 50 °C. Volatile fatty acids (VFA) were added to one incubation at the beginning of the experiment. The quantification of the SRB showed an increase of 16S-rRNA gene abundance at the beginning of incubation. After this the gene abundance remained constant. Initial adding VFA seemed to have no effect on the growth rate but on the metabolic activity of the bacteria. The phylogenetic analysis showed that most of the sequences (54 %) - amplified with general 16S-rRNS-primers - belong to the class Clostridia. 20 % of all sequences grouped within the genus Desulfotomaculum, which are able to form endospores and are assumed to cause the increase of sulphate reduction at 50 °C. Moreover, sequences where identified which show a high similarity to bacteria of the genera Tepidibacter, Caloranaerobacter and Garciella. These lineages contain thermophilic microorganisms able to catabolize carbohydrates and proteins to VFA that serve as electron-donors for SRB. Some microorganisms of these genera are associated with oil reservoirs or hydrothermal vents, which may serve as sources for the thermophilic population in the arctic sediment

Increased plasma vaspin concentration in patients with sepsis: an exploratory examination

Introduction: Vaspin (visceral adipose tissue-derived serpin) was first described as an insulin-sensitizing adipose tissue hormone. Recently its anti-inflammatory function has been demonstrated. Since no appropriate data is available yet, we sought to investigate the plasma concentrations of vaspin in sepsis. Materials and methods: 57 patients in intensive care, fulfilling the ACCP/SCCM criteria for sepsis, were prospectively included in our exploratory study. The control group consisted of 48 critically ill patients, receiving intensive care after trauma or major surgery. Patients were matched by age, sex, weight and existence of diabetes before statistical analysis. Blood samples were collected on the day of diagnosis. Vaspin plasma concentrations were measured using a commercially available enzyme-linked immunosorbent assay. Results: Vaspin concentrations were significantly higher in septic patients compared to the control group (0.3 (0.1-0.4) ng/mL vs. 0.1 (0.0-0.3) ng/mL, respectively; P < 0.001). Vaspin concentration showed weak positive correlation with concentration of C-reactive protein (CRP) (r = 0.31, P = 0.002) as well as with SAPS II (r = 0.34, P = 0.002) and maximum of SOFA (r = 0.39, P < 0.001) scoring systems, as tested for the overall study population. Conclusion: In the sepsis group, vaspin plasma concentration was about three-fold as high as in the median surgical control group. We demonstrated a weak positive correlation between vaspin and CRP concentration, as well as with two scoring systems commonly used in intensive care settings. Although there seems to be some connection between vaspin and inflammation, its role in human sepsis needs to be evaluated further

Increased plasma zonulin in patients with sepsis

Introduction: Zonulin is a eukaryotic protein structurally similar to Vibrio cholerae’s zonula occludens toxin. It plays an important role in the opening of small intestine tight junctions. The loss of gut wall integrity during sepsis might be pivotal and has been described in various experimental as well as human studies. Increased levels of zonulin could be demonstrated in diseases associated with increased intestinal inflammation, such as celiac disease and type 1 diabetes. We therefore investigated the role of plasma levels of zonulin in patients with sepsis as a non-invasive marker of gut wall integrity. Materials and methods: Plasma level of zonulin was measured in 25 patients with sepsis, severe sepsis or septic shock according to ACCP/SCCM criteria at the first day of diagnosed sepsis. 18 non-septic post-surgical ICU-patients and 20 healthy volunteers served as control. Plasma levels were determined by using commercially available ELISA kit. Data are given as median and interquartile range (IQR). Results: Significantly higher plasma concentration of zonulin were found in the sepsis group: 6.61 ng/mL (IQR 3.51-9.46), as compared to the to the post-surgical control group: 3.40 ng/mL (IQR 2.14-5.70) (P = 0.025), as well as to the healthy group: 3.55 ng/mL (IQR 3.14-4.14) (P = 0.008). Conclusion: We were able demonstrate elevated levels of plasma zonulin, a potential marker of intestinal permeability in septic patients. Increased zonulin may serve as an additional mechanism for the observed increased intestinal permeability during sepsis and SIRS

Liver transplantation reverses hypergammaglobulinemia in patients with chronic hepatic failure

Introduction: Sparse data are available about the effect of therapy methods on antibody levels in patients with liver failure. The aim of this study was to determine serum immunoglobulin concentrations in patients with chronic hepatic failure (CHF), acute- (ALF), or acute-on-chronic liver failure (ACLF) and to evaluate the impact of MARS treatment or liver transplantation (LT) on antibody levels. Materials and methods: We followed ten patients with ALF, twelve with ACLF and 18 with CHF. Eight patients with ALF and seven with ACLF underwent MARS therapy, whereas the rest received LT. 13 healthy volunteers served as controls. Serum antibody concentrations were measured using ELISA-technique. Results: Median serum levels of IgA, IgG and IgM were significantly increased in patients with CHF compared to ALF or controls (P < 0.02, P < 0.01, and P < 0.01). IgM and IgG concentrations were also significantly elevated in patients with CHF compared to ACLF (IgM, 3.7 vs. 1 g/L, P < 0.001; IgG, 8.7 vs. 3.1 g/L, P = 0.004). Immediately after LT a significant decrease of IgA (6.9 vs. 3.1 g/L, P = 0.004), IgG (8.7 vs. 5.1 g/L, P = 0.02) and IgM (3.7 vs. 1.8 g/L, P = 0.001) was detected in patients with CHF and antibody levels further decreased the days after LT reaching levels comparable to healthy individuals. MARS treatment had no apparent effect on the immunoglobulin profile in patients with ALF or ACLF. Conclusion: We provide evidence that LT reverses hypergammaglobulinemia in patients suffering from CHF within one day, which could be explained to a reconstituted hepatic antibody clearance, whereas MARS treatment has no immediate effect on immunoglobulin levels

A922 Sequential measurement of 1 hour creatinine clearance (1-CRCL) in critically ill patients at risk of acute kidney injury (AKI)

Meeting abstrac

The potential of chemokines and trefoil factor peptides as biomarkers for the early detection of chronic kidney disease and for monitoring the course of renal diseases

Patienten mit chronischem Nierenversagen (CNV) weisen eine hohe Mortalitätsrate und ein erhöhtes Risiko für weitere Erkrankungen auf. CNV schreitet langsam voran und erste Symptome treten zumeist kurz vor einem manifesten Nierenversagen auf, weswegen viele Patienten erst im fortgeschrittenen Stadium als solche erkannt werden. Da jedoch ein möglichst frühzeitiger Therapiebeginn die besten klinischen Ergebnisse erzielt, ist die Identifizierung von Patienten mit eingeschränkter Nierenfunktion von großer Wichtigkeit, um das Voranschreiten der Erkrankung rechtzeitig einzudämmen. Einerseits ist CNV mit einer anhaltenden Entzündungsreaktion assoziiert, andererseits jedoch auch mit einer erhöhten Aktivität zellulärer Reparaturmechanismen. Während der Entzündungsreaktion kommt es zu einer Aktivierung von Leukozyten sowie zu einer leukozytären Infiltration in das betroffene Gewebe.^ ^Dabei werden die Leukodiapedese und die weitere Migration der Leukozyten von sogenannten Chemokinen bestimmt. Daher war ein Ziel dieser Dissertation, die Chemokine CXCL11, CCL20, CCL22, und CCL17 im Harn und Serum von Patienten mit chronischem Nierenversagen nachzuweisen, sowie deren fraktionelle Exkretion zu berechnen, um das Potential dieser Proteine als Biomarker für CNV zu ermitteln. Mit zunehmendem Verlust der Nierenfunktion werden verschiedene Regulationsmechanismen aktiviert, um die renale Entzündungsreaktion einzudämmen und dabei unnötigen Zelltod und Gewebedestruktion zu verhindern. Einer davon ist die Hochregulierung von Trefoil factor family (TFF) Peptiden, denen eine besondere Bedeutung in der Regenerierung von Schleimhäuten zukommt. In den menschlichen Harnwegen konnte bereits die Sekretion von TFF Peptiden nachgewiesen werden, sowie deren vermehrte Expremierung bei verschiedenen Nierenerkrankungen.^ Ein weiteres Ziel dieser Arbeit war, das Potential von TFF Peptiden als Biomarker zur Früherkennung von Patienten mit renalen Einschränkungen und zur Abschätzung des Voranschreitens des Nierenversagens zu ermitteln. Die Konzentration von TFF Peptiden und den zuvor genannten Chemokinen wurde im Serum und im Harn von Patienten mit chronischem Nierenversagen der Stadien 1 bis 5 mittels ELISA gemessen. Gemessene Konzentrationen wurden anschließend mit verschiedenen klinischen Parametern korreliert sowie die fraktionelle Exkretion ermittelt, um die Proteinexkretion unabhängig von der glomerulären Funktion abschätzen zu können. Bei Patienten mit CNV konnten wir eine signifikant erhöhte fraktionelle Chemokinexkretion im Vergleich zur gesunden Kontrollgruppe nachweisen. Die Analyse mittels ROC-Kurve ergab, dass durch die Ermittlung der fraktionellen Chemokinexkretion verschiedene Stadien des CNV abgeschätzt werden können.^ Des Weiteren hatten Patienten mit fortgeschrittenen CNV Stadien signifikant erhöhte TFF3-Konzentrationen in Serum und Harn, was auf eine Hochregulation von Reparaturmechanismen in höheren CNV Stadien schließen lässt. Vergleichbar zu TFF3 konnten wir höhere Serumkonzentrationen von TFF1 und TFF2 in Patienten mit fortgeschrittener Nierenerkrankung nachweisen. Deren Konzentration im Harn war jedoch am höchsten bei Patienten mit niedrigeren CNV Stadien was auf das Potential dieser Proteine hindeutet, frühzeitig gefährdete Personen zu identifizieren. Mit Voranschreiten der Erkrankung sanken die Harnkonzentrationen von TFF1 und TFF2 wieder auf Werte vergleichbar mit gesunden Kontrollen.^ Eine Analyse mittels ROC-Kurve zeigte, dass mittels der Messung von TFF-Peptidkonzentrationen verschiedene CNV Stadien ermittelt und Patienten in einem frühen Stadium diagnostiziert werden könnten. Zusammenfassend konnten wir eine sich verändernde Expressionsrate von Chemokinen und TFF Peptiden während des Voranschreitens des CNV nachweisen, wobei TFF1 und TFF2 im Vergleich zu TFF3 unterschiedlich reguliert waren.Patients suffering from chronic kidney disease (CKD) have high mortality rates and increased risks of serious complications, particularly of developing cardiovascular disease. Even a minimal deterioration in renal function can entail severe complications like anaemia, malnutrition, bone disease, or neuropathy. Moreover, CKD progression is silent. Consequently, many patients are identified shortly before the onset of symptomatic kidney failure. At that state, only a few therapeutic options are left to avert detrimental outcomes or progression to end-stage renal disease (ESRD), necessitating kidney replacement therapy. Therefore, the early identification of persons at risk would help to initiate an appropriate treatment on time and could help to slow the progression of renal failure to ESRD. CKD is associated with ongoing inflammation, on the one hand, and initiation of repair mechanisms, on the other.^ ^Continuous inflammation is triggered among others by leukocyte activation and the migration of immunocompetent cells towards the site of damage. In doing so, chemokines are locally upregulated to facilitate leukocyte trafficking. An increase in chemokine expression has already been proven for different renal diseases. Therefore, one aim of this thesis was to measure chemokine levels in urine and serum of CKD patients, to calculate fractional chemokine expression, and to examine the potential of chemokine levels as biomarkers for CKD. Analyzed chemokines include CXCL11, CCL20, CCL22, and CCL17 which are essentially involved in the development and progression of CKD. During chronic inflammation, counterregulatory mechanisms are initiated to contain increased cell death and overcome continuous tissue destruction. One of these is the upregulation of Trefoil factor family (TFF) peptides, which are involved in the regeneration of mucus-containing epithelia.^ In the human urinary tract TFF peptides are secreted in a site-specific manner and have been shown to be upregulated in renal diseases. Therefore, the second aim of this thesis was to investigate the potential of TFF peptides as biomarker for CKD. The serum and urinary concentrations of TFF peptides and the above mentioned chemokines have been measured in patients with CKD stages 1 - 5 by using the ELISA technique. The obtained concentration profiles were subsequently correlated with several clinical parameters. Furthermore, fractional chemokine and TFF peptide expression rates were calculated to evaluate protein excretion irrespective of glomerular function. We found significantly elevated fractional chemokine expression rates in CKD patients as compared to healthy probands. Furthermore, fractional chemokine expression was able to predict various CKD stages, as depicted by ROC curve analysis.^ The TFF3 levels were significantly increased in later CKD stages, signifying the upregulation of counterregulatory processes during CKD progression. Comparable to TFF3, serum levels of TFF1 and TFF2 levels constantly increased with significantly elevated concentrations in higher CKD stages as compared to controls. In contrast, urinary levels of TFF1 and TFF2 were elevated with the onset of CKD and diminished with disease progression to levels comparable to healthy individuals, indicating the potential of these two proteins to identify patients with early CKD stages. The ROC curve analysis revealed that measurement of TFF peptide levels could help to predict different CKD stages and to identify individuals at risk. In summary, we found a changed expression rate of chemokines and TFF peptides during CKD progression, with TFF1 and TFF2 being differently regulated to TFF3.^ However, further clinical studies are obligatory to unravel the role of chemokines and TFF peptides in CKD and to ultimately evaluate their potential as biomarkers for CKD.submitted by Diana Lebherz-EichingerZusammenfassung in deutscher SpracheMedizinische Universität Wien, Diss., 201

Alterations of Endothelial Glycocalyx During Orthotopic Liver Transplantation in Patients With End-Stage Liver Disease.

BACKGROUND Endothelial glycocalyx participates in the maintenance of vascular integrity, and its perturbations cause capillary leakage, loss of vascular responsiveness, and enhanced adhesion of leukocytes and platelets. We hypothesized that marked shedding of the glycocalyx core protein, syndecan-1, occurs in end-stage liver disease (ESLD) and that it increases during orthotopic liver transplantation (OLT). We further evaluated the effects of general anesthesia on glycocalyx shedding and its association with acute kidney injury (AKI) after OLT. PATIENTS AND METHODS Thirty consecutive liver transplant recipients were enrolled in this prospective study. Ten healthy volunteers served as a control. Acute kidney injury was defined by Acute Kidney Injury Network criteria. RESULTS Plasma syndecan-1 was significantly higher in ESLD patients than in healthy volunteers (74.3 ± 59.9 vs 10.7 ± 9.4 ng/mL), and it further increased significantly after reperfusion (74.3 ± 59.9 vs 312.6 ± 114.8 ng/mL). The type of general anesthesia had no significant effect on syndecan-1. Syndecan-1 was significantly higher during the entire study in patients with posttransplant AKI stage 2 or 3 compared to patients with AKI stage 0 or 1. The area under the curve of the receiver operating characteristics curve of syndecane-1 to predict AKI stage 2 or 3 within 48 hours after reperfusion was 0.76 (95% confidence interval, 0.57-0.89, P = 0.005). CONCLUSIONS Patients with ESLD suffer from glycocalyx alterations, and ischemia-reperfusion injury during OLT further exacerbates its damage. Despite a higher incidence of AKI in patients with elevated syndecan-1, it is not helpful to predict de novo AKI. Volatile anesthetics did not attenuate glycocalyx shedding in human OLT