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Agricultural pesticides, nutrients, and habitat degradation are major causes of insect declines in lowland streams. To effectively conserve and restore stream habitats, standardized stream monitoring data and societal support for freshwater protection are needed. Here, we sampled 137 small stream monitoring sites across Germany, 83 % of which were located in agricultural catchments, with >900 citizen scientists in 96 monitoring groups. Sampling was carried out according to Water Framework Directive standards as part of the citizen science freshwater monitoring program FLOW in spring and summer 2021, 2022 and 2023. The biological indicator SPEAR pesticides was used to assess pesticide exposure and effects based on macroinvertebrate community composition. Overall, 58 % of the agricultural monitoring sites failed to achieve a good ecological status in terms of macroinvertebrate community composition and indicated high pesticide exposure (SPEAR pesticides status class: 29 % "moderate", 19 % "poor", 11 % "bad"). The indicated pesticide pressure in streams was related to the proportion of arable land in the catchment areas (R 2  = 0.23, p < 0.001). Also with regards to hydromorphology, monitoring results revealed that 65 % of the agricultural monitoring sites failed to reach a good status. The database produced by citizen science groups was characterized by a high degree of accuracy, as results obtained by citizen scientists and professionals were highly correlated for SPEAR pesticides index (R 2  = 0.79, p < 0.001) and hydromorphology index values (R 2  = 0.72, p < 0.001). Such citizen-driven monitoring of the status of watercourses could play a crucial role in monitoring and implementing the objectives of the European Water Framework Directive, thus contributing to restoring and protecting freshwater ecosystems., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Increasing antibiotic resistances of numerous pathogens mean that myxobacteria, well known producers of new antibiotics, are becoming more and more interesting. More than 100 secondary metabolites, most of them with bioactivity, were described from the order Myxococcales. Especially new myxobacterial genera and species turned out to be reliable sources for novel antibiotics and can be isolated from uncommon neglected habitats like, for example, acidic soils. Almost nothing is known about the diversity of myxobacteria in moors, except some information from cultivation studies of the 1970s. Therefore, we evaluated the myxobacterial community composition of acidic high moor and fen both with cultivation-independent 16S rRNA clone bank analysis and with cultivation. Phylogenetic analyses of clone sequences revealed a great potential of undescribed myxobacteria in high moor and fen, whereby all sequences represent unknown taxa and were detected exclusively by cultivation-independent analyses. However, many clones were assigned to sequences from other cultivation-independent studies of eubacterial diversity in acidic habitats. Cultivation revealed different strains exclusively from the genus Corallococcus. Our study shows that the neglected habitat moor is a promising source and of high interest with regard to the cultivation of prospective new bioactive secondary metabolite producing myxobacteria., (© 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

Myxobacteria harbor an enormous potential for new bioactive secondary metabolites and therefore the isolation of in particular new groups is of great interest. The diversity of myxobacteria present in two ecological habitats, namely sand from Kiritimati Island and German compost, was evaluated by both cultivation-based and cultivation-independent methods. Phylogenetic analyses of the strains in comparison with 16S rRNA gene sequences from cultured and uncultured material in GenBank revealed a great potential of undescribed myxobacteria in both sampling sites. Several OTUs (operational taxonomic units) represent unknown taxa and were detected by clone bank analyses, but not by cultivation. Clone bank analyses indicated that the myxobacterial community is predominantly indigenous. The 16S rDNA libraries from the two samples were generated from total community DNA with myxobacterial specific forward and universal reverse primer sets. The clones were partially sequenced. Cultivation was successful for exclusively bacteriolytic, but not for cellulolytic myxobacteria and revealed 42 strains from the genera Corallococcus, Myxococcus, and Polyangium. The genera of Myxococcaceae family were represented by both approaches. But, even in this well studied family, as well as in the suborders Sorangiineae and Nannocystineae, a considerable number of clones were assigned to, if any, uncultivated organisms. Our study shows an overrepresentation of the genera Myxococcus spp. and Corallococcus spp. with standard cultivation methods. However, high deficits are demonstrated in the cultivation success of the myxobacterial diversity detected by exclusively cultivation-independent approaches. Especially, clades which are exclusively represented by clones are of high interest with regard to the cultivation of new bioactive secondary metabolite producers., (© 2015 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

Granzyme K (GzmK) belongs to a family of trypsin-like serine proteases localized in electron dense cytoplasmic granules of activated natural killer and cytotoxic T-cells. Like the related granzymes A and B, GzmK can trigger DNA fragmentation and is involved in apoptosis. We expressed the Ser(195) --> Ala variant of human pro-GzmK in Escherichia coli, crystallized it, and determined its 2.2-A x-ray crystal structure. Pro-GzmK possesses a surprisingly rigid structure, which is most similar to activated serine proteases, in particular complement factor D, and not their proforms. The N-terminal peptide Met(14)-Ile(17) projects freely into solution and can be readily approached by cathepsin C, the natural convertase of pro-granzymes. The pre-shaped S1 pocket is occupied by the ion paired residues Lys(188B)-Asp(194) and is hence not available for proper substrate binding. The Ser(214)-Cys(220) segment, which normally provides a template for substrate binding, bulges out of the active site and is distorted. With analogy to complement factor D, we suggest that this strand will maintain its non-productive conformation in mature GzmK, mainly due to the unusual residues Gly(215), Glu(219), and Val(94). We hypothesize that GzmK is proteolytically active only toward specific, as yet unidentified substrates, which upon approach transiently induce a functional active-site conformation.

Tryptase-like activities of T and NK cells contribute to the induction of target cell apoptosis, but only granzyme A (GzmA) has been shown to exhibit Z-Lys-SBzl esterase activity in murine T cells. GzmA-deficient mice exhibit residual Z-Lys-SBzl hydrolyzing activity and almost normal levels of lymphocyte-mediated cytotoxicity. Here we report the cloning and biochemical characterization of recombinant mouse granzyme K (GzmK). The purified murine protein shows Z-Lys-SBzl hydrolyzing activity and is inhibited by bikunin, the light chain of inter-alpha-trypsin inhibitor, like the human homolog. We conclude that GzmK expressed by GzmA-deficient T cells accounts for the remaining Z-Lys-SBzl activity. Functional similarities between GzmA and GzmK may explain the subtle immunological deficits observed in GzmA-deficient mice.

We explored expression and possible function of interferon-gamma (IFN-gamma) in cultured fetal (E15) rat dorsal root ganglion neurons combining whole cell patch-clamp electrophysiology with single cell reverse transcriptase polymerase chain reaction and confocal laser immunocytochemistry. Morphologically, we located IFN-gamma protein in the cytoplasm of the neurons in culture as well as in situ during peri- and postnatal development. Transcripts for classic IFN-gamma and for its receptor were determined in probes of cytoplasm sampled from individual cultured neurons, which had been identified by patch clamp electrophysiology. In addition, the cultured neurons expressed both chains of the IFN-gamma receptor. Locally produced IFN-gamma acts back on its cellular source. Phosphorylation and nuclear translocation of the IFN-inducible transcriptional factor STAT1 as well as IFN-gamma-dependent expression of major histocompatibility complex class I molecules on the neuronal membrane were noted in untreated cultures. However, both processes were substantially blocked in the presence of antibodies neutralizing IFN-gamma. Our findings indicate a role of IFN-gamma in autocrine regulation of sensory neurons.