A better understanding of the function of neutrophil extracellular traps (NETs) may facilitate the development of interventions for sepsis. The study aims to investigate the formation and degradation of NETs in three murine sepsis models and to analyze the production of reactive oxygen species (ROS) during NET formation. Murine sepsis was induced by midgut volvulus (720° for 15 min), cecal ligation and puncture (CLP), or the application of lipopolysaccharide (LPS) (10 mg/kg body weight i.p.). NET formation and degradation was modulated using mice that were genetically deficient for peptidyl arginine deiminase-4 (PAD4-KO) or DNase1 and 1L3 (DNase1/1L3-DKO). After 48 h, mice were killed. Plasma levels of circulating free DNA (cfDNA) and neutrophil elastase (NE) were quantified to assess NET formation and degradation. Plasma deoxyribonuclease1 (DNase1) protein levels, as well as tissue malondialdehyde (MDA) activity and glutathione peroxidase (GPx) activity, were quantified. DNase1 and DNase1L3 in liver, intestine, spleen, and lung tissues were assessed. The applied sepsis models resulted in a simultaneous increase in NET formation and oxidative stress. NET formation and survival differed in the three models. In contrast to LPS and Volvulus, CLP-induced sepsis showed a decreased and increased 48 h survival in PAD4-KO and DNase1/1L3-DKO mice, when compared to WT mice, respectively. PAD4-KO mice showed decreased formation of NETs and ROS, while DNase1/1L3-DKO mice with impaired NET degradation accumulated ROS and chronicled the septic state. The findings indicate a dual role for NET formation and degradation in sepsis and ischemia-reperfusion (I/R) injury: NETs seem to exhibit a protective capacity in certain sepsis paradigms (CLP model), whereas, collectively, they seem to contribute adversely to scenarios where sepsis is combined with ischemia-reperfusion (volvulus).

Introduction: An early and accurate diagnosis of early onset neonatal sepsis (EONS) and late onset neonatal sepsis (LONS) is essential to improve the outcome of this devastating conditions. Especially, preterm infants are at risk. Reliable biomarkers are rare, clinical decision-making depends on clinical appearance and multiple laboratory findings. Markers of NET formation and NET turnover might improve diagnostic precision. Aim of this study was to evaluate the diagnostic value of NETs in sepsis diagnosis in neonatal preterm infants. Methods: Plasma samples of neonatal preterm infants with suspected sepsis were collected. Blood samples were assayed for markers of NET formation and NET turnover: cfDNA, DNase1, nucleosome, NE, and H3Cit. All clinical findings, values of laboratory markers, and epidemiological characteristics were collected retrospectively. Two subpopulations were created to divide EONS from LONS. EMA sepsis criteria for neonatal sepsis were used to generate a sepsis group (EMA positive) and a control group (EMA negative). Results: A total of 31 preterm neonates with suspected sepsis were included. Out of these, nine patients met the criteria for sepsis according to EMA. Regarding early onset neonatal sepsis (3 EONS vs. 10 controls), cfDNA, DNase I, nucleosome, and CRP were elevated significantly. H3Cit and NE did not show any significant elevations. In the late onset sepsis collective (6 LONS vs. 12 controls), cfDNA, DNase I, and CRP differed significantly compared to control group.

Background: Neutrophil extracellular traps (NETs)—as double-edged swords of innate immunity—are involved in numerous processes such as infection, inflammation and tissue repair. Research on neutrophil granulocytes is limited because of their short lifetime of only a few hours. Several attempts have been made to prolong the half-life of neutrophils using cytokines and bacterial products and have shown promising results. These long-term surviving neutrophils are reported to maintain phagocytic activity and cytokine release; however, little is known regarding their capability to release NETs.Methods: We analysed the prolongation of neutrophil survival in vitro under various culture conditions using granulocyte colony-stimulating factor (G-CSF), lipopolysaccharide (LPS) or tumour necrosis factor alpha (TNF-α) by flow cytometry and a viability assay. Additionally, we assessed NET formation following stimulation with phorbol 12-myristate 13-acetate (PMA) by immunofluorescence staining, myeloperoxidase (MPO)-DNA sandwich-ELISA and fluorometric assays for cell-free DNA (cfDNA), neutrophil elastase (NE) and myeloperoxidase (MPO).Results: Untreated neutrophils could form NETs after stimulation with PMA for up to 24 h. Incubation with LPS extended their ability to form NETs for up to 48 h. At 48 h, NET release of neutrophils cultured with LPS was significantly higher compared to that of untreated cells; however, no significantly different enzymatic activity of NE and MPO was observed. Similarly, incubation with G-CSF resulted in significantly higher NET release at 48 h compared to untreated cells. Furthermore, NETs showed significantly higher enzymatic activity of NE and MPO after incubation with G-CSF. Lastly, incubation with TNF-α had no influence on NET release compared to untreated cells although survival counts were altered by TNF-α.Conclusions: G-CSF, LPS or TNF-α each at low concentrations lead to prolonged survival of cultured neutrophils, resulting in considerable differences in NET formation and composition. These results provide new information for the use of neutrophils in long-term experiments for NET formation and provide novel insights for neutrophil behaviour under inflammatory conditions.

Background: Increasing resistance to antibiotics poses medical challenges worldwide. Prospective data on carriage prevalence of multidrug resistant organisms (MDRO) in children at hospital admission are limited and associated risk factors are poorly defined. Aim: To determine prevalence of MDRO carriage in children at admission to our paediatric hospital in Hamburg and to identify MDRO carriage risk factors. Methods: We prospectively obtained and cultured nasal/throat and inguinal/anal swabs from children (≤ 18 years) at admission between September 2018 and May 2019 to determine prevalence of meticillin-resistant Staphylococcus aureus (MRSA), multidrug-resistant Gram-negative bacteria (MRGN) and vancomycin-resistant enterococcus (VRE) and associated species. We collected medical histories using a questionnaire and evaluated 31 risk factors using logistic regression models. Results: MDRO carriage prevalence of 3,964 children was 4.31% (95% confidence interval (CI): 3.69–5.00). MRSA carriage prevalence was 0.68% (95% CI: 0.44–0.99), MRGN prevalence was 3.64% (95% CI: 3.07–4.28) and VRE prevalence 0.08% (95% CI: 0.02–0.22). MDRO carriage was associated with MRGN history (odds ratio (OR): 6.53; 95% CI: 2.58–16.13), chronic condition requiring permanent care (OR: 2.67; 95% CI: 1.07–6.13), antibiotic therapy (OR: 1.92, 95% CI: 1.24–2.94), living in a care facility (OR: 3.34; 95% CI: 0.72–12.44) and refugee status in previous 12 months (OR: 1.91; 95% CI: 0.27–8.02). Compared to established practice, screening using risk-factors had better diagnostic sensitivity (86.13%; 95% CI: 80.89–91.40) and specificity (73.54%; 95% CI: 72.12–74.97). Conclusion: MRGN carriage was higher than MRSA and VRE. Extended risk-factor-based admission screening system seems warranted.

Infant mortality due to congenital malformations, deformities and chromosomal abnormalities is approximately 700 per 100 000 live births in Germany. In addition, congenital malformations are frequently associated with high morbidity or even lifelong...

Background: Ilioinguinal-iliohypogastric block (ILIHB) is a well-established procedure for postoperative analgesia after open inguinal surgery in children. This procedure is effective and safe, especially when ultrasound is used. Data availability for comparing ultrasound-guided blocks versus wound infiltration is still weak. The study was designed to determine the efficacy of ultrasound-guided ILIHB (US-ILIHB) on postoperative pain control in pediatric patients following a inguinal daycase surgery, compared with perifocal wound infiltration (PWI) by the surgeon. Methods: This randomized, double-blinded trail was conducted in pediatric patients aged from 6 months to 4 years. The total number of children included in the study was 103. Patients were allocated at random in two groups by sealed envelopes. The ILIHB group recieved 0,2% ropivacain for US-ILIHB after anesthesia induction. The PWI group recieved 0,2% ropivacain for PWI performed by a surgeon before wound closure. Parameters recorded included the postoperative pain score, pain frequency, time to first analgesics and consumption of analgesics. Results: US-ILIHB significantly reduced the occurrence of pain within the first 24 h after surgery (7.7%, p = 0.01). Moreover, the pain-free interval until administration of the first dose of opioids was 21 min longer, on average (p = 0.003), following US-ILIHB compared to perifocal wound infiltration. 72% of children who received US-ILIHB did not require additional opioids, as compared to 56% of those who received PWI. Conclusion: Thus our study demonstrates that US-ILIHB ensures better postoperative analgesia in children and should be prioritized over postoperative PWI. Trail registration: UIHBOPWIIC, DRKS00020987. Registered 20 March 2020 – Retrospectivley registered.