Abundance and heterotrophic activity of Bacteria and Archaea and bacterial community structure in the water column of the eastern Atlantic

Im Osten des Atlantiks wurden entlang eines Transekts die Abundanzen und Leucin-Aufnahmeraten von Bakterien und Archaea in der Wassersäule ermittelt. Außerdem wurde die Zusammensetzung von aktiven und gesamten Bakteriengemeinschaften durch molekulare Fingerprinting-Techniken analysiert. Die Häufigkeiten von Bakterien, Crenarchaeota und Euryarchaeota wurden mittels dem Catalyzed Reporter Deposition-Fluorescence In Situ Hybridization (CARD-FISH) Verfahren ermittelt. Obwohl der Anteil der Bakterien an der gesamten Prokaryoten-Abundanz im Bathypelagial (1000 – 4000 m) geringer war als an der Oberfläche und dem Mesopelagial (100 – 1000 m), dominierten sie dennoch in der gesamten Wassersäule. Crenarchaota trugen 30 ± 12% zu der gesamten Prokaryoten-Abundanz bei, wobei sie eine generell höhere Verbreitung im Bathypelagial hatten als in der Oberflächenschicht und dem Mesopelagial. Euryarchaeota überstiegen in keiner Probe 5 % der gesamten Prokaryoten-Abundanz. Zusätzlich zu den Leucin-Aufnahmeraten der Gesamtprokaryotengemeinschaft wurden auch die Leucin-Aufnahmeraten für Archaea und Bakterien ermittelt. Um die verschiedenen Aktivitätsraten von Bakterien und Archaea in Bezug zu der gesamten Prokaryoten-Produktivität zu bestimmen, wurde der bakterielle Hemmstoff Erythromycin und der Archaea Hemmstoff Diphtheria Toxin eingesetzt. Die Spezifität des Hemmstoffs Erythromycin wurde durch die Kombination von Microautoradiographie mit CARD-FISH getestet. Dabei wurde festgestellt, dass Erythromycin selektiv die bakterielle Produktivität unterdrückt. In der euphotischen Zone und dem Mesopelagial war der Beitrag der Bakterien an der gesamten Leucin-Aufnahmerate 65 ± 15% der gesamten Prokaryoten Leucin-Aufnahmerate und nahm mit zunehmender Tiefe ab. Die durchschnittliche Zellen-spezifische Aktivitätsrate der Crenarchaeota (3.7 ± 2.0 x 10-5 fmol cell-1 d-1) war 5-mal geringer als die der Bakterien (18.3 ± 17.6 x 10-5 fmol cell-1 d-1). Im Bathypelagial, jedoch, war die Zell-spezifischen Leucin-Aufnahmerate der Archaea vergleichbar mit der der Bakterien (1.8 x 10-5 fmol cell-1 d-1 for Crenarchaeota, 2.3 x 10-5 fmol cell-1 d-1 for Bacteria). Diese Ergebnisse deuten darauf hin, dass im Bathypelagial des Atlantiks Crenarchaeota und Bakterien bezogen auf Zellniveau vergleichbare heterotrophe Aktivitäten haben, in den Oberflächengewässern und dem oberen Mesopelagial jedoch die heterotrophe Aktivität der Archaea um eine Größenordnung geringer als die der Bakterien ist. Um zwischen der Zusammensetzung der Gemeinschaft von metabolisch aktiven Bakterien und der Gesamtbakteriengemeinschaft zu unterscheiden, wurde der Einbau von Bromodeoxyuridine (BrdU) in die aktive Gemeinschaft mit einer Kombination von immunocapturing der DNA in die BrdU eingebaut wurde und Automated Ribosomal Intergenic Spacer Analysis (ARISA), als molekulare Fingerprinting-Technik, verwendet. Insgesamt wurden dabei 655 verschiedene operational taxonomic units (OTUs) detektiert. Die Ähnlichkeitsanalyse der bakteriellen Gemeinschaften ergab zwischen den aktiven (BrdU-markierte Bakterien) und gesamten Bakterien einen signifikanten (ANOSIM, r = 0.361, p = 0.001) Unterschied in der Zusammensetzung der Gemeinschaften in der gesamten Wassersäule. Die Ähnlichkeiten zwischen Oberflächengewässer und Mesopelagial (100 – 1000 m) und Bathypelagial (1000 – 3000 m) war für die BrdU-markierte Bakteriengemeinschaft größer (r = 0.181, p = 0.007) als für die Gesamtbakteriengemeinschaft (r = 0.332, p = 0.003). Der Shannon-Wiener index (H’), als Index für die Diversität einer Gemeinschaft, war für die Gemeinschaft der Gesamtbakteriengemeinschaft größer als für die aktive (paired t-test; p = 0.002). Diese Ergebnisse deuten darauf hin, dass die Diversität von Gemeinschaften im Wesentlichen durch die sich im Ruhestand befindenden Bakterien gebildet wird, die in einer sich verändernden Umwelt eine Art „Samenbank“ bilden könnte.The abundance and leucine incorporation rate of Archaea and Bacteria were determined throughout the water column along a transect in the eastern Atlantic. Additionally, the composition of the active and total bacterial community was analyzed using molecular fingerprinting techniques. The abundance of Bacteria, Crenarchaeota and Euryarchaeota was determined by Catalyzed Reporter Deposition - Fluorescence In Situ Hybridization (CARD-FISH). Bacteria dominated throughout the water column, although their contribution to total prokaryotic abundance in the bathypelagic layer (1000 - 4000 m) was lower than in the surface and mesopelagic layers (0 - 1000 m). Crenarchaeota contributed about 30 ± 12% to the total prokaryotic abundance with a generally higher contribution in the bathypelagic layer than in the surface and mesopelagic layers. Euryarcheota contributed less than 5% to the total prokaryotic abundance throughout the water column. Leucine incorporation rates were determined for the total prokaryotic community as well as for Bacteria and Archaea separately using selective inhibitors. The bacterial inhibitor erythromycin and the archaeal inhibitor diphtheria toxin were used to determine the contribution of Archaea and Bacteria, respectively, to total heterotrophic activity. Using microautoradiography in combination with CARD-FISH, the specificity of erythromycin was tested. Erythromycin was found to selectively inhibit bacterial activity. The contribution of Bacteria to total leucine incorporation amounted to 65 ± 15% of total prokaryotic leucine incorporation decreasing in their contribution with depth. The mean cell-specific leucine incorporation rate of Crenarchaeota (3.7 ± 2.0 x 10-5 fmol cell-1 d-1) was 5 times lower than that of Bacteria (18.3 ± 17.6 x 10-5 fmol cell-1 d-1) in the surface and mesopelagic layer. In the bathypelagic layer, however, cell-specific leucine incorporation rates of Crenarchaeota were similar to those of Bacteria (1.8 x 10-5 fmol cell-1 d-1 for Crenarchaeota, 2.3 x 10-5 fmol cell-1 d-1 for Bacteria). These results indicate that Crenarchaeota and Bacteria exhibit similar heterotrophic activity on a per-cell level in the bathypelagic waters of the Atlantic, while in the surface and upper mesopelagic waters, cell-specific heterotrophic archaeal activity is about one order of magnitude lower than that of Bacteria. To differentiate the community composition of metabolically active Bacteria from the total bacterial community, bromodeoxyuridine (BrdU) incorporation into the active bacterial community was used in combination with immunocapturing and automated ribosomal intergenic spacer analysis (ARISA) as a molecular fingerprinting approach. In total, 655 operational taxonomic units (OTUs) were found. Results of the similarity analysis (ANOSIM, r = 0.361, p = 0.001) of the obtained clusters showed a significant difference in the community composition throughout the water column between active (Bacteria incorporating BrdU) and the total bacterial community. The similarity between surface and mesopelagic communities (100 – 1000 m) and bathypelagic communities (1000 – 3000 m) was higher for the active (r = 0.181, p = 0.007) than for the total (r = 0.332, p = 0.003) bacterial community. The Shannon-Wiener diversity index (H’) was significant higher for the total Bacteria (paired t-test; p = 0.002) than for the active ones. These findings suggest that the diversity of communities is mainly driven by the dormant fraction of the bacterial community, potentially forming a ‘seed bank’ in changing environmental conditions

A Mitochondrial Polymorphism Alters Immune Cell Metabolism and Protects Mice from Skin Inflammation

Several genetic variants in the mitochondrial genome (mtDNA), including ancient polymorphisms, are associated with chronic inflammatory conditions, but investigating the functional consequences of such mtDNA polymorphisms in humans is challenging due to the influence of many other polymorphisms in both mtDNA and the nuclear genome (nDNA). Here, using the conplastic mouse strain B6-mtFVB, we show that in mice, a maternally inherited natural mutation (m.7778G > T) in the mitochondrially encoded gene ATP synthase 8 (mt-Atp8) of complex V impacts on the cellular metabolic profile and effector functions of CD4+ T cells and induces mild changes in oxidative phosphorylation (OXPHOS) complex activities. These changes culminated in significantly lower disease susceptibility in two models of inflammatory skin disease. Our findings provide experimental evidence that a natural variation in mtDNA influences chronic inflammatory conditions through alterations in cellular metabolism and the systemic metabolic profile without causing major dysfunction in the OXPHOS system

Nucleocapsid-specific T cell responses associate with control of SARS-CoV-2 in the upper airways before seroconversion

Despite intensive research since the emergence of SARS-CoV-2, it has remained unclear precisely which components of the early immune response protect against the development of severe COVID-19. Here, we perform a comprehensive immunogenetic and virologic analysis of nasopharyngeal and peripheral blood samples obtained during the acute phase of infection with SARS-CoV-2. We find that soluble and transcriptional markers of systemic inflammation peak during the first week after symptom onset and correlate directly with upper airways viral loads (UA-VLs), whereas the contemporaneous frequencies of circulating viral nucleocapsid (NC)-specific CD4+ and CD8+ T cells correlate inversely with various inflammatory markers and UA-VLs. In addition, we show that high frequencies of activated CD4+ and CD8+ T cells are present in acutely infected nasopharyngeal tissue, many of which express genes encoding various effector molecules, such as cytotoxic proteins and IFN-γ. The presence of IFNG mRNA-expressing CD4+ and CD8+ T cells in the infected epithelium is further linked with common patterns of gene expression among virus-susceptible target cells and better local control of SARS-CoV-2. Collectively, these results identify an immune correlate of protection against SARS-CoV-2, which could inform the development of more effective vaccines to combat the acute and chronic illnesses attributable to COVID-19

ZMYND10 Is Mutated in Primary Ciliary Dyskinesia and Interacts with LRRC6

Defects of motile cilia cause primary ciliary dyskinesia (PCD), characterized by recurrent respiratory infections and male infertility. Using whole-exome resequencing and high-throughput mutation analysis, we identified recessive biallelic mutations in ZMYND10 in 14 families and mutations in the recently identified LRRC6 in 13 families. We show that ZMYND10 and LRRC6 interact and that certain ZMYND10 and LRRC6 mutations abrogate the interaction between the LRRC6 CS domain and the ZMYND10 C-terminal domain. Additionally, ZMYND10 and LRRC6 colocalize with the centriole markers SAS6 and PCM1. Mutations in ZMYND10 result in the absence of the axonemal protein components DNAH5 and DNALI1 from respiratory cilia. Animal models support the association between ZMYND10 and human PCD, given that zmynd10 knockdown in zebrafish caused ciliary paralysis leading to cystic kidneys and otolith defects and that knockdown in Xenopus interfered with ciliogenesis. Our findings suggest that a cytoplasmic protein complex containing ZMYND10 and LRRC6 is necessary for motile ciliary function

Abstracts from the 8th International Conference on cGMP Generators, Effectors and Therapeutic Implications

This work was supported by a restricted research grant of Bayer AG

Time to Treatment Discontinuation in German Patients with Schizophrenia: Long-Acting Injectables versus Oral Antipsychotics

Background and objective!#!Long-acting injectable antipsychotics (LAIs) are associated with better treatment adherence and persistence than oral antipsychotics (OAPs) in patients with schizophrenia. However, real-world evidence assessing the impact of treatment with LAIs in Germany is limited. To fill this gap, we compared antipsychotic medication adherence and risk of treatment discontinuation (TD) among schizophrenia patients newly initiated on LAI or who switched their OAP regimen (overall cohort; OC).!##!Methods!#!Claims data of German schizophrenia patients who initiated LAIs or switched their OAP during 2012-2016 (index date) were retrospectively analyzed. Treatment switch was defined as add-on medication to existing prescription or terminating the existing prescription and initiating another OAP. Adherence and time to treatment discontinuation (TTD) were estimated. Determinants of treatment discontinuation were analyzed using two Cox regression models. Model 1 controlled for age, sex, and Charlson Comorbidity Index (CCI); model 2 also included insurance status, and medication, visit, and psychiatric inpatient stay costs. Sensitivity analysis on patients who terminated existing prescriptions and initiated new OAPs (complete switch cohort; CSC) was performed.!##!Results!#!In OC (n = 2650), LAI users had better adherence (35.4% vs. 11.6%), persistence (no 60-day gap; 40.7% vs. 19.8%), and longer TTD (median [95% confidence interval (CI)] 216 [193-249] vs. 50 [46-56] days) than OAP users. OAP usage (hazard ratio [HR] 1.89, 95% CI 1.73-2.06; p < 0.001) and greater CCI (HR 1.04, 95% CI 1.00-1.07; p = 0.023) were associated with greater risk of TD in model 1. Model 2 showed similar results. LAI users in CSC also had better adherence, persistence, and longer TTD. In CSC too, OAP usage and greater CCI were associated with greater risk of TD in model 1, but only CCI was significant in model 2. Higher pre-index psychiatric inpatient costs were associated with lower risk of TD (HR 0.99, 95% CI 0.98-1.00; p = 0.014).!##!Limitations!#!Inherent limitations of claims data and lack of control on OAP administration may have influenced the results.!##!Conclusion!#!This real-world study associates LAIs with better medication adherence and lower antipsychotic discontinuation risk than OAPs

Hospitalization Rates and Therapy Costs of German Schizophrenia Patients Who are Initiated on Long-Acting Injectable Medication: A Mirror-Image Study

Background!#!Long-acting injectable (LAI) antipsychotics can reduce relapse, hospitalization, and costs in patients with schizophrenia. However, real-world evidence assessing the impact of treatment with LAIs in Germany is limited.!##!Objective!#!To provide updated evidence on the impact of LAI initiation on hospitalization rates and therapy costs.!##!Methods!#!Using a mirror-image design, claims data of 850 German patients with schizophrenia who initiated treatment with LAIs during 2013-2015 was retrospectively analyzed. For the included patients, costs and resource utilization were compared for the 12 months before the index date (first initiation of LAI) and the 12 months after the index date. Annual treatment costs, hospitalization rates, ambulatory visits, sick leaves and medical aids were assessed. Two models were used to evaluate hospitalization and its costs. In model 1, hospitalization during the index date (first LAI prescription in 2013-2015) was allocated to the 'pre-' time interval, while in model 2 it was neither attributed to the pre- nor to the post-index date. Regression analysis was performed to identify patients who benefited the most in terms of cost reduction from LAI initiation.!##!Results!#!Medication costs were significantly higher post-switching to LAI compared with pre-switching period (€3832 vs €799; p < 0.001). In model 1, number of hospitalizations, days hospitalized, and associated costs were significantly lower post-switching compared with pre-switching (2.3 vs 2.6; 59.2 vs 73.4; and €5355 vs €11,908, respectively; all p < 0.001). Similar results were obtained for costs in model 2 (€5355 vs €10,276; p < 0.001). Mean total costs reduced significantly from pre-switching to post-switching period in model 1 (€13,776 vs €10,418; p < 0.001). Patients with characteristics such as higher number of non-psychiatric and psychiatric inpatient stays during the pre-index period (all p < 0.05) benefited the most from cost reduction after LAI initiation.!##!Conclusion!#!In this cohort of German patients with schizophrenia, treatment initiation with LAI resulted in reduced hospitalization rates and total costs

Systematic Review of Invasive Meningococcal Disease: Sequelae and Quality of Life Impact on Patients and Their Caregivers

Introduction: Invasive meningococcal disease (IMD, septicaemia and/or meningitis) has a severe acute and long-term burden: 5-10% of patients die within 48h, and long-term sequelae have been reported in 10-20% of survivors. Health-related quality of life (HRQoL) is increasingly but inconsistently assessed. Methods: A systematic literature review on Neisseria meningitidis IMD sequelae and HRQoL in survivors of all ages and their caregivers, including family, was conducted for high-income countries from 2001 to 2016 (in Medline and Embase, following Cochrane and PRISMA guidelines). Results: A total of 31 studies, mostly of childhood IMD cases, were included. A broad range of physical, neurological and psychological IMD sequelae were identified. The literature has evolved, with more types of sequelae reported in more recent studies; however, meningococcal disease-specific and sequelae-specific HRQoL data are lacking, and existing studies used a wide variety of instruments. Physical sequelae included: amputations (up to 8% of children, 3% adolescents/adults) and skin scars (up to 55% of children, 18% adolescents, 2% adults). Neurologic sequelae included: hearing loss (up to 19% of infants, 13% children, 12% adolescents, 8% adults). Psychological sequelae included: anxiety, learning difficulties, emotional and behavioural difficulties. IMD negatively affects HRQoL in patients and also in their family and close caregiver network, both in the short- and long-term. Even IMD survivors without sequelae experienced an adverse impact on HRQoL after many years, affecting self-esteem, physical, mental and psychosocial health, and HRQoL was worse in those with cognitive and behavioural sequelae. Conclusion: A high proportion of IMD survivors are affected by a broad range of sequelae and reduced HRQoL that persists years after infection. Childhood IMD survivors had more sequelae and more severe sequelae compared with adult survivors. HRQoL was affected in patients and also in their families, caregivers and surrounding network over the long term. More research is needed to resolve data gaps and to standardise HRQoL assessment

Memorandum Registry for Health Services Research: Update 2019

Health registries could be used to analyze questions concerning routine practice in healthcare. Therefore, registries are a core method in health services research. The German Network for Health Services Research (Deutsches Netzwerk Versorgungsforschung, DNVF) promotes the quality of registries by scientific exchange, organization of advanced training, and recommendations in the form of a memorandum "Registry for Health Services Research". The current recommendations are an update of the memorandum's first version of 2010. The update describes the capabilities and aims of registries in health services research. Furthermore, it illustrates the state-of-the-art in designing and implementing health registries. The memorandum provides developers the methodological basis to ensure high quality health registries. It further provides users of health registries with insights that enable assessing the quality of data and results of health registries. Finally, funding agencies and health policy actors can use the quality criteria to establish a framework for the financing and legislative requirements for health registries. The memorandum provides first a definition of health registries and presents an overview of their utility in health services research and health care improvement. Second, several areas of methodological importance for the development and operation of health registries are presented. This includes the conceptual and preliminary design, implementation, technical organization of a health registry, statistical analysis, reporting of results, and data protection. From these areas, criteria are deduced to allow the assessment of the quality of a health registry. Finally, a checklist is presented

Phospholipase D Activity Is Regulated by Product Segregation and the Structure Formation of Phosphatidic Acid within Model Membranes

Phospholipase D from Streptomyces chromofuscus (scPLD) hydrolyzes phosphatidylcholines (PC) to produce choline and phosphatidic acid (PA), a lipid messenger molecule within biological membranes. To scrutinize the influence of membrane structure on scPLD activity, three different substrate-containing monolayers are used as model systems: pure dipalmitoylphosphatidylcholine (DPPC) as well as equimolar mixtures of DPPC/n-hexadecanol (C16OH) and DPPC/dipalmitoylglycerol (DPG). The activity of scPLD toward these monolayers is tested by infrared reflection-absorption spectroscopy and exhibits different dependencies on surface pressure. For pure DPPC, the catalytic turnover drastically drops above 20 mN/m. On addition of C16OH, this strong decrease starts at 5 mN/m. For the DPPC/DPG system, the reaction yield linearly decreases between 5 and 25 mN/m. The difference in scPLD activity is correlated to the phase state of the monolayers as examined by x-ray diffraction, Brewster angle microscopy, and atomic force microscopy. Because the additives C16OH and DPG mediate the miscibility of PC and PA, only a basal activity of scPLD is observed toward the mixed systems at higher surface pressures. At pure DPPC monolayers, scPLD is activated after the segregation of initially formed PA. Furthermore, scPLD is inhibited when the lipids in the PA-rich domains adopt an upright orientation. This phenomenon offers a self-regulating mechanism for the concentration of the second messenger PA within biological membranes