Infectious diseases are influenced by interactions between host and pathogen, and the number of infected hosts is rarely homogenous across the landscape. Areas with elevated pathogen prevalence can maintain a high force of infection and may indicate areas with disease impacts on host populations. However, isolating the ecological processes that result in increases in infection prevalence and intensity remains a challenge. Here we elucidate the contribution of pathogen clade and host species in disease hotspots caused by Ophidiomyces ophidiicola, the pathogen responsible for snake fungal disease, in 21 species of snakes infected with multiple pathogen strains across 10 countries in Europe. We found isolated areas of disease hotspots in a landscape where infections were otherwise low. O. ophidiicola clade had important effects on transmission, and areas with multiple pathogen clades had higher host infection prevalence. Snake species further influenced infection, with most positive detections coming from species within the Natrix genus. Our results suggest that both host and pathogen identity are essential components contributing to increased pathogen prevalence.

• MeSH
• Dermatomycoses * epidemiology   microbiology   MeSH
• Snakes microbiology   MeSH
• Disease Hotspot MeSH
• Prevalence MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Europe epidemiology   MeSH

HotSpot Wizard is a web server for automatic identification of 'hot spots' for engineering of substrate specificity, activity or enantioselectivity of enzymes and for annotation of protein structures. The web server implements the protein engineering protocol, which targets evolutionarily variable amino acid positions located in the active site or lining the access tunnels. The 'hot spots' for mutagenesis are selected through the integration of structural, functional and evolutionary information obtained from: (i) the databases RCSB PDB, UniProt, PDBSWS, Catalytic Site Atlas and nr NCBI and (ii) the tools CASTp, CAVER, BLAST, CD-HIT, MUSCLE and Rate4Site. The protein structure and e-mail address are the only obligatory inputs for the calculation. In the output, HotSpot Wizard lists annotated residues ordered by estimated mutability. The results of the analysis are mapped on the enzyme structure and visualized in the web browser using Jmol. The HotSpot Wizard server should be useful for protein engineers interested in exploring the structure of their favourite protein and for the design of mutations in site-directed mutagenesis and focused directed evolution experiments. HotSpot Wizard is available at http://loschmidt.chemi.muni.cz/hotspotwizard/.

• MeSH
• beta-Lactamases chemistry   MeSH
• Glycoside Hydrolases chemistry   MeSH
• Phosphoric Triester Hydrolases chemistry   MeSH
• Hydrolases chemistry   MeSH
• Internet MeSH
• Protein Engineering * MeSH
• Reproducibility of Results MeSH
• Software * MeSH
• User-Computer Interface MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• beta-Lactamases MeSH
• Glycoside Hydrolases MeSH
• haloalkane dehalogenase MeSH Browser
• Phosphoric Triester Hydrolases MeSH
• Hydrolases MeSH
• licheninase MeSH Browser

Subglacial environments provide conditions suitable for the microbial production of methane, an important greenhouse gas, which can be released from beneath the ice as a result of glacial melting. High gaseous methane emissions have recently been discovered at Russell Glacier, an outlet of the southwestern margin of the Greenland Ice Sheet, acting not only as a potential climate amplifier but also as a substrate for methane consuming microorganisms. Here, we describe the composition of the microbial assemblage exported in meltwater from the methane release hotspot at Russell Glacier and its changes over the melt season and as it travels downstream. We found that a substantial part (relative abundance 27.2% across the whole dataset) of the exported assemblage was made up of methylotrophs and that the relative abundance of methylotrophs increased as the melt season progressed, likely due to the seasonal development of the glacial drainage system. The methylotrophs were dominated by representatives of type I methanotrophs from the Gammaproteobacteria; however, their relative abundance decreased with increasing distance from the ice margin at the expense of type II methanotrophs and/or methylotrophs from the Alphaproteobacteria and Betaproteobacteria. Our results show that subglacial methane release hotspot sites can be colonized by microorganisms that can potentially reduce methane emissions.

• Keywords
• Greenland Ice Sheet, Methanotrophs, Methylotrophs, Subglacial environment,
• MeSH
• Ice Cover * microbiology   MeSH
• Methane * analysis   MeSH
• Climate MeSH
• Seasons MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Greenland MeSH
• Names of Substances
• Methane * MeSH

BACKGROUND: Treating memory and cognitive deficits requires knowledge about anatomical sites and neural activities to be targeted with particular therapies. Emerging technologies for local brain stimulation offer attractive therapeutic options but need to be applied to target specific neural activities, at distinct times, and in specific brain regions that are critical for memory formation. METHODS: The areas that are critical for successful encoding of verbal memory as well as the underlying neural activities were determined directly in the human brain with intracranial electrophysiological recordings in epilepsy patients. We recorded a broad range of spectral activities across the cortex of 135 patients as they memorised word lists for subsequent free recall. FINDINGS: The greatest differences in the spectral power between encoding subsequently recalled and forgotten words were found in low theta frequency (3-5 Hz) activities of the left anterior prefrontal cortex. This subsequent memory effect was proportionally greater in the lower frequency bands and in the more anterior cortical regions. We found the peak of this memory signal in a distinct part of the prefrontal cortex at the junction between the Broca's area and the frontal pole. The memory effect in this confined area was significantly higher (Tukey-Kramer test, p<0.05) than in other anatomically distinct areas. INTERPRETATION: Our results suggest a focal hotspot of human verbal memory encoding located in the higher-order processing region of the prefrontal cortex, which presents a prospective target for modulating cognitive functions in the human patients. The memory effect provides an electrophysiological biomarker of low frequency neural activities, at distinct times of memory encoding, and in one hotspot location in the human brain. FUNDING: Open-access datasets were originally collected as part of a BRAIN Initiative project called Restoring Active Memory (RAM) funded by the Defence Advanced Research Project Agency (DARPA). CT, ML, MTK and this research were supported from the First Team grant of the Foundation for Polish Science co-financed by the European Union under the European Regional Development Fund.

• Keywords
• Anterior prefrontal cortex, Frontal pole, Human verbal memory, Intracranial recordings, Memory encoding,
• MeSH
• Humans MeSH
• Magnetic Resonance Imaging MeSH
• Brain Mapping MeSH
• Brain physiology   MeSH
• Memory * physiology   MeSH
• Prefrontal Cortex * physiology   MeSH
• Mental Recall physiology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

HotSpot Wizard 2.0 is a web server for automated identification of hot spots and design of smart libraries for engineering proteins' stability, catalytic activity, substrate specificity and enantioselectivity. The server integrates sequence, structural and evolutionary information obtained from 3 databases and 20 computational tools. Users are guided through the processes of selecting hot spots using four different protein engineering strategies and optimizing the resulting library's size by narrowing down a set of substitutions at individual randomized positions. The only required input is a query protein structure. The results of the calculations are mapped onto the protein's structure and visualized with a JSmol applet. HotSpot Wizard lists annotated residues suitable for mutagenesis and can automatically design appropriate codons for each implemented strategy. Overall, HotSpot Wizard provides comprehensive annotations of protein structures and assists protein engineers with the rational design of site-specific mutations and focused libraries. It is freely available at http://loschmidt.chemi.muni.cz/hotspotwizard.

• MeSH
• Automation MeSH
• Biocatalysis MeSH
• Databases, Protein MeSH
• Internet * MeSH
• Evolution, Molecular MeSH
• Models, Molecular MeSH
• Mutation * MeSH
• Mutagenesis, Site-Directed methods   MeSH
• Peptide Library * MeSH
• Proteins chemistry   genetics   MeSH
• Software * MeSH
• Protein Stability MeSH
• Amino Acid Substitution MeSH
• Substrate Specificity MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Peptide Library * MeSH
• Proteins MeSH

Polymorphous low-grade adenocarcinoma (PLGA) is the second most frequent type of malignant tumor of the minor salivary glands. We identified PRKD1 hotspot mutations encoding p.Glu710Asp in 72.9% of PLGAs but not in other salivary gland tumors. Functional studies demonstrated that this kinase-activating alteration likely constitutes a driver of PLGA.

• MeSH
• Adenocarcinoma genetics   pathology   MeSH
• NIH 3T3 Cells MeSH
• Immunohistochemistry MeSH
• Immunoprecipitation MeSH
• Microscopy, Confocal MeSH
• Humans MeSH
• Mutation, Missense genetics   MeSH
• Models, Molecular * MeSH
• Molecular Sequence Data MeSH
• Mutagenesis MeSH
• Mice MeSH
• Salivary Gland Neoplasms genetics   pathology   MeSH
• Protein Kinase C chemistry   genetics   MeSH
• Amino Acid Sequence MeSH
• Sequence Analysis, DNA MeSH
• Sequence Alignment MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• protein kinase D MeSH Browser
• Protein Kinase C MeSH

β-N-Acetylhexosaminidase from Talaromyces flavus (TfHex; EC 3.2.1.52) is an exo-glycosidase with dual activity for cleaving N-acetylglucosamine (GlcNAc) and N-acetylgalactosamine (GalNAc) units from carbohydrates. By targeting a mutation hotspot of the active site residue Glu332, we prepared a library of ten mutant variants with their substrate specificity significantly shifted towards GlcNAcase activity. Suitable mutations were identified by in silico methods. We optimized a microtiter plate screening method in the yeast Pichia pastoris expression system, which is required for the correct folding of tetrameric fungal β-N-acetylhexosaminidases. While the wild-type TfHex is promiscuous with its GalNAcase/GlcNAcase activity ratio of 1.2, the best single mutant variant Glu332His featured an 8-fold increase in selectivity toward GlcNAc compared with the wild-type. Several prepared variants, in particular Glu332Thr TfHex, had significantly stronger transglycosylation capabilities than the wild-type, affording longer chitooligomers - they behaved like transglycosidases. This study demonstrates the potential of mutagenesis to alter the substrate specificity of glycosidases.

• Keywords
• Pichia pastoris, Talaromyces flavus, site-directed mutagenesis, site-saturation mutagenesis, substrate specificity, β-N-acetylhexosaminidase,
• MeSH
• Acetylgalactosamine metabolism   MeSH
• Acetylglucosamine * metabolism   MeSH
• Acetylglucosaminidase MeSH
• beta-N-Acetylhexosaminidases * metabolism   MeSH
• Kinetics MeSH
• Mutation MeSH
• Substrate Specificity MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Acetylgalactosamine MeSH
• Acetylglucosamine * MeSH
• Acetylglucosaminidase MeSH
• beta-N-Acetylhexosaminidases * MeSH

HotSpot Wizard is a web server used for the automated identification of hotspots in semi-rational protein design to give improved protein stability, catalytic activity, substrate specificity and enantioselectivity. Since there are three orders of magnitude fewer protein structures than sequences in bioinformatic databases, the major limitation to the usability of previous versions was the requirement for the protein structure to be a compulsory input for the calculation. HotSpot Wizard 3.0 now accepts the protein sequence as input data. The protein structure for the query sequence is obtained either from eight repositories of homology models or is modeled using Modeller and I-Tasser. The quality of the models is then evaluated using three quality assessment tools-WHAT_CHECK, PROCHECK and MolProbity. During follow-up analyses, the system automatically warns the users whenever they attempt to redesign poorly predicted parts of their homology models. The second main limitation of HotSpot Wizard's predictions is that it identifies suitable positions for mutagenesis, but does not provide any reliable advice on particular substitutions. A new module for the estimation of thermodynamic stabilities using the Rosetta and FoldX suites has been introduced which prevents destabilizing mutations among pre-selected variants entering experimental testing. HotSpot Wizard is freely available at http://loschmidt.chemi.muni.cz/hotspotwizard.

• MeSH
• Databases, Protein MeSH
• Internet * MeSH
• Catalytic Domain MeSH
• Models, Molecular MeSH
• Mutation MeSH
• Proteins chemistry   genetics   MeSH
• Amino Acid Sequence MeSH
• Sequence Alignment MeSH
• Software * MeSH
• Protein Stability MeSH
• Thermodynamics MeSH
• Computational Biology * MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Proteins MeSH

'QTL-hotspot' is a genomic region on linkage group 04 (CaLG04) in chickpea (Cicer arietinum) that harbours major-effect quantitative trait loci (QTLs) for multiple drought-adaptive traits, and it therefore represents a promising target for improving drought adaptation. To investigate the mechanisms underpinning the positive effects of 'QTL-hotspot' on seed yield under drought, we introgressed this region from the ICC 4958 genotype into five elite chickpea cultivars. The resulting introgression lines (ILs) and their parents were evaluated in multi-location field trials and semi-controlled conditions. The results showed that the 'QTL-hotspot' region improved seed yield under rainfed conditions by increasing seed weight, reducing the time to flowering, regulating traits related to canopy growth and early vigour, and enhancing transpiration efficiency. Whole-genome sequencing data analysis of the ILs and parents revealed four genes underlying the 'QTL-hotspot' region associated with drought adaptation. We validated diagnostic KASP markers closely linked to these genes using the ILs and their parents for future deployment in chickpea breeding programs. The CaTIFY4b-H2 haplotype of a potential candidate gene CaTIFY4b was identified as the superior haplotype for 100-seed weight. The candidate genes and superior haplotypes identified in this study have the potential to serve as direct targets for genetic manipulation and selection for chickpea improvement.

• Keywords
• Cicer arietinum, Chickpea, drought stress, haplotypes, introgression lines, legume, marker-assisted backcrossing, transpiration efficiency, whole-genome sequencing,
• MeSH
• Cicer * genetics   MeSH
• Genomics MeSH
• Publication type
• Journal Article MeSH

• MeSH
• Incidence MeSH
• Humans MeSH
• Kidney Neoplasms epidemiology   etiology   MeSH
• Colonic Neoplasms epidemiology   etiology   MeSH
• Environmental Exposure adverse effects   MeSH
• Check Tag
• Humans MeSH
• Publication type
• News MeSH
• Geographicals
• Czech Republic epidemiology   MeSH