In situ development of a methanotrophic microbiome in deep-sea sediments.

Emission of the greenhouse gas methane from the seabed is globally controlled by marine aerobic and anaerobic methanotrophs gaining energy via methane oxidation. However, the processes involved in the assembly and dynamics of methanotrophic populations in complex natural microbial communities remain unclear. Here we investigated the development of a methanotrophic microbiome following subsurface mud eruptions at Håkon Mosby mud volcano (1250 m water depth). Freshly erupted muds hosted deep-subsurface communities that were dominated by Bathyarchaeota, Atribacteria and Chloroflexi. Methanotrophy was initially limited to a thin surface layer of Methylococcales populations consuming methane aerobically. With increasing distance to the eruptive center, anaerobic methanotrophic archaea, sulfate-reducing Desulfobacterales and thiotrophic Beggiatoaceae developed, and their respective metabolic capabilities dominated the biogeochemical functions of the community. Microbial richness, evenness, and cell numbers of the entire microbial community increased up to tenfold within a few years downstream of the mud flow from the eruptive center. The increasing diversity was accompanied by an up to fourfold increase in sequence abundance of relevant metabolic genes of the anaerobic methanotrophic and thiotrophic guilds. The communities fundamentally changed in their structure and functions as reflected in the metagenome turnover with distance from the eruptive center, and this was reflected in the biogeochemical zonation across the mud volcano caldera. The observed functional succession provides a framework for the response time and recovery of complex methanotrophic communities after disturbances of the deep-sea bed.

High efficacy of sofosbuvir/velpatasvir and impact of baseline resistance-associated substitutions in hepatitis C genotype 3 infection.

BACKGROUND: Twelve weeks of the pangenotypic direct-acting antiviral (DAA) combination sofosbuvir/velpatasvir (SOF/VEL) was highly efficient in patients with hepatitis C virus (HCV) genotype 3 (GT3) infection in the ASTRAL-3 approval study. However, presence of resistance-associated substitutions (RASs) in the HCV nonstructural protein 5A (NS5A) was associated with lower treatment response. AIM: To assess the efficacy and safety of SOF/VEL ± ribavirin (RBV) and the impact of NS5A RASs and RBV use on treatment outcome in HCV GT3 infection in a real-world setting. METHODS: In this multicentre cohort study, GT3 patients from ten treatment centres across Germany were included. Sustained virological response was assessed 12 weeks after end-of-treatment (SVR12) in modified intention-to-treat (mITT) and per-protocol analysis (PP). NS5A RASs were tested by population-based sequencing. RESULTS: A total of 293 GT3 patients were included. The median age was 48 years, 70% were male, 25.3% were cirrhotic, 9.2% were HCV/HIV co-infected and 21.8% were treatment-experienced, including 4.1% with DAA experience. Baseline NS5A RASs (Y93H, A30K, L31M) were detected in 11.2%. RBV was added in 5% of noncirrhotic and 58.9% of cirrhotic patients, respectively. SVR12 rates for SOF/VEL±RBV were 95.9% (mITT) and 99.5% (PP), respectively. Only 1 virological relapse occurred in a cirrhotic patient previously treated with SOF/RBV. No treatment-related major adverse events occurred. CONCLUSION: Twelve weeks of SOL/VEL±RBV was safe and highly efficient in HCV GT3 across a diverse patient population. Baseline NS5A RASs were rarely observed and presence did not seem to impact SVR, regardless of the use of RBV.

Deficiency of MAPK-activated protein kinase 2 (MK2) prevents adverse remodelling and promotes endothelial healing after arterial injury.

Maladaptive remodelling of the arterial wall after mechanical injury (e. g. angioplasty) is characterised by inflammation, neointima formation and media hypertrophy, resulting in narrowing of the affected artery. Moreover, mechanical injury of the arterial wall causes loss of the vessel protecting endothelial cell monolayer. Mitogen-activated protein kinase (MAPK)-activated protein kinase 2 (MK2), a major downstream target of p38 MAPK, regulates inflammation, cell migration and proliferation, essential processes for vascular remodelling and re-endothelialisation. Therefore, we investigated the role of MK2 in remodelling and re-endothelialisation after arterial injury in genetically modified mice in vivo. Hypercholesterolaemic low-density-lipoprotein-receptor-deficient mice (ldlr-/-) were subjected to wire injury of the common carotid artery. MK2-deficiency (ldlr-/-/mk2-/-) nearly completely prevented neointima formation, media hypertrophy, and lumen loss after injury. This was accompanied by reduced proliferation and migration of MK2-deficient smooth muscle cells. In addition, MK2-deficiency severely reduced monocyte adhesion to the arterial wall (day 3 after injury, intravital microscopy), which may be attributed to reduced expression of the chemokine ligands CCL2 and CCL5. In line, MK2-deficiency significantly reduced the content of monocytes, neutrophiles and lymphocytes of the arterial wall (day 7 after injury, flow cytometry). In conclusion, in a model of endothelial injury (electric injury), MK2-deficiency strongly increased proliferation of endothelial cells and improved re-endothelialisation of the arterial wall after injury. Deficiency of MK2 prevents adverse remodelling and promotes endothelial healing of the arterial wall after injury, suggesting that MK2-inhibition is a very attractive intervention to prevent restenosis after percutaneous therapeutic angioplasty.

Anaerobic methanotrophic community of a 5346-m-deep vesicomyid clam colony in the Japan Trench.

Vesicomyidae clams harbor sulfide-oxidizing endosymbionts and are typical members of cold seep communities where active venting of fluids and gases takes place. We investigated the central biogeochemical processes that supported a vesicomyid clam colony as part of a locally restricted seep community in the Japan Trench at 5346 m water depth, one of the deepest seep settings studied to date. An integrated approach of biogeochemical and molecular ecological techniques was used combining in situ and ex situ measurements. In sediment of the clam colony, low sulfate reduction rates (maximum 128 nmol mL(-1) day(-1)) were coupled to the anaerobic oxidation of methane. They were observed over a depth range of 15 cm, caused by active transport of sulfate due to bioturbation of the vesicomyid clams. A distinct separation between the seep and the surrounding seafloor was shown by steep horizontal geochemical gradients and pronounced microbial community shifts. The sediment below the clam colony was dominated by anaerobic methanotrophic archaea (ANME-2c) and sulfate-reducing Desulfobulbaceae (SEEP-SRB-3, SEEP-SRB-4). Aerobic methanotrophic bacteria were not detected in the sediment, and the oxidation of sulfide seemed to be carried out chemolithoautotrophically by Sulfurovum species. Thus, major redox processes were mediated by distinct subgroups of seep-related microorganisms that might have been selected by this specific abyssal seep environment. Fluid flow and microbial activity were low but sufficient to support the clam community over decades and to build up high biomasses. Hence, the clams and their microbial communities adapted successfully to a low-energy regime and may represent widespread chemosynthetic communities in the Japan Trench. In this regard, they contributed to the restricted deep-sea trench biodiversity as well as to the organic carbon availability, also for non-seep organisms, in such oligotrophic benthic environment of the dark deep ocean.

Niche differentiation among mat-forming, sulfide-oxidizing bacteria at cold seeps of the Nile Deep Sea Fan (Eastern Mediterranean Sea).

Sulfidic muds of cold seeps on the Nile Deep Sea Fan (NDSF) are populated by different types of mat-forming sulfide-oxidizing bacteria. The predominant sulfide oxidizers of three different mats were identified by microscopic and phylogenetic analyses as (i) Arcobacter species producing cotton-ball-like sulfur precipitates, (ii) large filamentous sulfur bacteria including Beggiatoa species, and (iii) single, spherical Thiomargarita species. High resolution in situ microprofiles revealed different geochemical settings selecting for the different mat types. Arcobacter mats occurred where oxygen and sulfide overlapped above the seafloor in the bottom water interface. Filamentous sulfide oxidizers were associated with steep gradients of oxygen and sulfide in the sediment. A dense population of Thiomargarita was favored by temporarily changing supplies of oxygen and sulfide in the bottom water. These results indicate that the decisive factors in selecting for different mat-forming bacteria within one deep-sea province are spatial or temporal variations in energy supply. Furthermore, the occurrence of Arcobacter spp.-related 16S rRNA genes in the sediments below all three types of mats, as well as on top of brine lakes of the NDSF, indicates that this group of sulfide oxidizers can switch between different life modes depending on the geobiochemical habitat setting.