Refine
Has Fulltext
- yes (285)
Is part of the Bibliography
- yes (285)
Year of publication
Document Type
- Journal article (285) (remove)
Language
- English (285) (remove)
Keywords
- Infektionsbiologie (57)
- Escherichia coli (19)
- Candida albicans (12)
- escherichia coli (11)
- gene expression (10)
- RNA-seq (9)
- expression (8)
- infection (8)
- Biologie (7)
- Staphylococcus aureus (7)
- virulence (7)
- Immunologie (6)
- epithelial cells (6)
- identification (6)
- activation (5)
- antimicrobial resistance (5)
- gene regulation (5)
- messenger RNA (5)
- plasmodium falciparum (5)
- sRNA (5)
- transcriptome (5)
- antibiotics (4)
- antibodies (4)
- antisense RNA (4)
- bacteria (4)
- biofilms (4)
- inhibition (4)
- pathogens (4)
- resistance (4)
- small RNA (4)
- synthesis (4)
- vancomycin (4)
- Expression (3)
- Hfq (3)
- Legionella pneumophila (3)
- Neisseria meningitidis (3)
- RNA (3)
- RNA sequencing (3)
- RNA-binding proteins (3)
- Salmonella (3)
- adaptation (3)
- antibacterial activity (3)
- autophagy (3)
- bacillus subtilis (3)
- biology (3)
- cancer (3)
- genes (3)
- genome sequence (3)
- leishmania major (3)
- macrophages (3)
- mechanisms (3)
- medicine (3)
- mouse models (3)
- protein kinase (3)
- sequence (3)
- signaling pathway (3)
- staphylococcus aureus (3)
- stem cells (3)
- substituent (3)
- transcription (3)
- transcription factors (3)
- transcriptomics (3)
- translation (3)
- Agent (2)
- Antisense RNA (2)
- Aspergillus fumigatus (2)
- Breast-tumors (2)
- COVID-19 (2)
- Campylobacter jejuni (2)
- Cancer (2)
- Candida auris (2)
- DNA transcription (2)
- E. coli serotype 06 (2)
- F8 fimbriae (2)
- Grad-seq (2)
- Growth (2)
- HFQ (2)
- HeLa cells (2)
- Helicobacter pylori (2)
- In-vivo (2)
- Leishmania (2)
- Lung cancer (2)
- MATQ-seq (2)
- Medizin (2)
- Nude-mice (2)
- RNA synthesis (2)
- RNS (2)
- Rhodobacter sphaeroides (2)
- SARS-CoV-2 (2)
- SEQ (2)
- Salmonella Typhimurium (2)
- Salmonella enterica (2)
- Salmonella typhimurium (2)
- Salmonellosis (2)
- Streptomyces (2)
- Therapy (2)
- adhesion (2)
- allelic replacement (2)
- antibiotic resistance (2)
- antifungals (2)
- antigen (2)
- antisense RNAs (2)
- apoptosis (2)
- archaea (2)
- bacterial genetics (2)
- bioluminescence imaging (2)
- biosynthesis (2)
- chaperone (2)
- comparative genomics (2)
- cytokines (2)
- cytotoxicity (2)
- dRNA-Seq (2)
- enterica serovar typhimurium (2)
- epidemiology (2)
- evolution (2)
- fluorescence microscopy (2)
- gastric cancer (2)
- gastrointestinal tract (2)
- gene (2)
- gene cloning (2)
- gene expression profiling (2)
- genome (2)
- genome annotation (2)
- glycopeptide antibiotics (2)
- hemolysin (2)
- host cells (2)
- in vitro (2)
- in vivo imaging (2)
- in-vitro (2)
- in-vivo (2)
- inflammation (2)
- inhibitor (2)
- leishmaniasis (2)
- liraglutide (2)
- magnetic resonance imaging (2)
- malaria (2)
- mechanism (2)
- messenger-RNA (2)
- metabolism (2)
- metastasis (2)
- methionine (2)
- mutation (2)
- neutrophils (2)
- noncoding RNA (2)
- obesity (2)
- oncolytic virotherapy (2)
- oncolytic viruses (2)
- parasitic diseases (2)
- pathogenicity (2)
- plants (2)
- polycationic peptides (2)
- protein kinases (2)
- proteins (2)
- recombination (2)
- red fluorescent protein (2)
- regulator (2)
- regulator genes (2)
- regulatory RNA (2)
- response regulator (2)
- responses (2)
- ribonucleases (2)
- riboswitch (2)
- secretion (2)
- secretion systems (2)
- signal transduction (2)
- single-cell RNA-seq (2)
- small RNAs (2)
- small nucleolar RNAs (2)
- strains (2)
- structure-activity (2)
- subunit (2)
- therapy (2)
- transmission (2)
- transmission electron microscopy (2)
- tumors (2)
- type VII secretion system (2)
- vaccinia virus (2)
- virulence factors (2)
- (+)-limonene (1)
- (Nucleotide sequence (1)
- 0 antigen (1)
- 2-component system (1)
- 3D tissue model (1)
- 3′ UTR (1)
- 5-bromodeoxyuridine (1)
- 6S RNA (1)
- AMP-activated kinases (1)
- ATG (1)
- ATG8 (1)
- AcrAB-TolC efflux pump (1)
- Adherence (1)
- Adhesion (1)
- Adhäsion (1)
- Alkyltransferase Ribozyme SAMURI (1)
- Apicomplexan (1)
- Archaea (1)
- Avirulent Salmonella (1)
- BNIP3 (1)
- BT_1884 (1)
- Bacillus subtilis (1)
- Bacterial conjugation (1)
- Bacterial pathogens (1)
- Bacteriaophage AR9 (1)
- Bakterien (1)
- Biology (1)
- Blood-brain barrier (1)
- Bordeiella pertussis (1)
- Bovis (1)
- Bradyrhizobium (1)
- C-6-H (1)
- C-Terminus (1)
- COX-2 (1)
- CRISPR-Cas system (1)
- CRISPRs (1)
- CTSE (1)
- CUL4-DDB1 ubiquitin ligase (1)
- Carcinoma (1)
- Cas9 (1)
- Cell binding (1)
- Chemical modification (1)
- Chemotherapy (1)
- Combination therapy (1)
- Crohns-disease (1)
- CsrA (1)
- Cutaneous leishmaniasis (1)
- Cyclophosphamide (1)
- Cystic-fibriosis (1)
- DASH (1)
- DNA (1)
- DNA glycosation (1)
- DNA lang range mapping (1)
- DNA methylation (1)
- DNA methylferase homolog (1)
- Deletion (1)
- Dextran sulphate (1)
- Diarrhea (1)
- Disease gene prioritization (1)
- Division (1)
- Dual 3'seq (1)
- E. coli hemolysin (1)
- E. coli virulence (1)
- ESAT‐6‐like secretion system (1)
- ESS (1)
- Elektronisches Publizieren (1)
- Enterica serovar typhimurium (1)
- Enterobacteriaceae (1)
- Enterococcus faecalis (1)
- Enterococcus faecium (1)
- Environmental isolate (1)
- Epstein-Barr virus (1)
- Erwachsener (1)
- Erwinia amylovora (1)
- EsaA (1)
- Escherichia coli-Hfq (1)
- Escherichia-coli K-12 (1)
- Ethiopia (1)
- Evolution (1)
- Excision (1)
- F-19 MRI (1)
- F1 (1)
- Family (1)
- Fimbria (1)
- Fimbriae (1)
- FinO family (1)
- Foc protein (1)
- Ftsz (1)
- GFP (1)
- GI-101A tumor xenografts (1)
- GLP-1 (1)
- GLV-1 h153 (1)
- GLV-1H68 (1)
- GO1 (1)
- Gene (1)
- Gene cloning (1)
- Gene expression (1)
- Gene mapping (1)
- Gene regulation (1)
- Gene-expression (1)
- Gene-transfer (1)
- Genetik (1)
- Genome analysis (1)
- Genome re-annotation (1)
- Genomic profile (1)
- Germany (1)
- GibS (1)
- Gifsy-1 (1)
- Gram-positive bacteria (1)
- Haemolysin (1)
- Haloferax volcanii (1)
- Harnwegsinfekt (1)
- Helicobacterpylori (1)
- Hemolysin excretion (1)
- Hirnhautentzündung (1)
- Homology (1)
- Hospital water system (1)
- Host adaptation (1)
- Host cells (1)
- Hämolysin (1)
- IL-12 production (1)
- IcaR (1)
- Ileal Crohns-disease (1)
- Images (1)
- In-vitro (1)
- Inactivation (1)
- Infection (1)
- Inflammation (1)
- Inhibitor (1)
- Internal transcription start site (1)
- Invasion genes (1)
- Ire1 (1)
- Iron-oxide (1)
- Iron-uptake (1)
- IsrK (1)
- K279a (1)
- KhpB protein (1)
- Klebsiella pneumoniae (1)
- Krebs <Medizin> (1)
- Langerhans cell (1)
- Leaderless transcript (1)
- Legionella ssp. (1)
- Legionellae (1)
- Leishmania major (1)
- Lesions (1)
- Level (1)
- Listeria monocytogenes (1)
- Lungenkrebs (1)
- Lysis (medicine) (1)
- MACE (1)
- MI-2/NURD complex (1)
- MRI reporter (1)
- MRSA (1)
- MRSA - methicillin-resistant Staphylococcus aureus (1)
- MTOR (1)
- MVT (1)
- Macrophage (1)
- Maintenance (1)
- Malaria (1)
- Malignant effusion (1)
- Medicine (1)
- Methanosarcina mazei (1)
- Mice (1)
- MicroRNAs (1)
- Mig1 (1)
- Minor subunits (1)
- Molekulare Infektionsbiologie (1)
- Mouse (1)
- Mutant (1)
- Mycobacterium (1)
- N-glycans (1)
- NAFLD (1)
- NAS (1)
- NASH (1)
- ND10 complex (1)
- NF-KAPPA-B (1)
- NK cells (1)
- NOTCH (1)
- NRG1 (1)
- NSG (1)
- Neugeborenes (1)
- Nodule (1)
- Non-coding RNAs (1)
- Nonpathogenic Escherichia-coli (1)
- OLFM4 (1)
- Oncolytic virotherapy (1)
- One Health (1)
- Organoids (1)
- Orthogonal field attenuation gel electrophoresis (1)
- OxyR (1)
- P-fimbriae (1)
- P. falciparum (1)
- PCR (1)
- PEGylation (1)
- PETRI-seq (1)
- PFS25 (1)
- PIA/ica (1)
- PML (1)
- PML nuclear-bodies (1)
- PPGPP (1)
- PSI-blast (1)
- PYY3-36 (1)
- Paeonia (1)
- Pan1 (1)
- Particles (1)
- Pathogenicity (1)
- Pathogens (1)
- PfAMA1 (1)
- PfCCp protein (1)
- Pfs230 (1)
- Pilus (1)
- PknB (1)
- Plasmid (1)
- ProQ (1)
- Promoter (1)
- Promoter prediction (1)
- Protein (1)
- Protein function prediction (1)
- Proteins (1)
- Proteogenomics (1)
- Proteomanalyse (1)
- Pseudomonas (1)
- Pseudomonas aeruginosa (1)
- Pseudomonas-aeruginosa (1)
- RFP (1)
- RNA CHAPERONE HFQ (1)
- RNA chaperone Hfq (1)
- RNA denaturation (1)
- RNA expression (1)
- RNA sequence analyses (1)
- RNA structure (1)
- RNA-Seq (1)
- RNA-polymerase-I (1)
- RNA-polymerase-II (1)
- RNA-sequencing (1)
- RNAseq (1)
- RNase E (1)
- RNase III (1)
- ROS (1)
- RTS,S/AS01 malaria vaccine (1)
- RYGB (1)
- Rapid evolution (1)
- Rats (1)
- Recombinant vaccinia (1)
- Recombination directionality factor (1)
- Regression (1)
- Ribosome profiling (1)
- Roux-en-Y gastric bypass surgery (1)
- S fimbrial adhesin (Sfa) (1)
- S-Fimbriae (1)
- S-fimbria (1)
- S-fimbriae (1)
- S. aureus (1)
- SER/THR kinase (1)
- SM-like protein (1)
- SNF1 (1)
- SUBSP carotovora (1)
- SWI2/SNF2-like protein (1)
- Salmonella enterica Typhimurium strain SL1344 (1)
- Salmonella-enteritidis (1)
- Salmonella-typhimurium (1)
- Schistosoma japonicum; Egg antigen; Carbohydrate epitope; Circumoval precipitin test; Immunoassay (1)
- SdsR (1)
- Sequence identity (1)
- Sequencing (1)
- Serogroup (1)
- Serratia marcescens (1)
- Shiga toxin-producing Escherichia coli (1)
- Shigella (1)
- Shigellosis (1)
- Site-specific RNA labelling (1)
- Snf1 (1)
- Soft-tissue infection (1)
- Southern hybridization (1)
- Speichel (1)
- Staphylococcal infection (1)
- Staphylococcus aureus USA300 (1)
- Staphylococcus-aureus (1)
- Stenotrophomonas maltophilia (1)
- Stp (1)
- Strain nissle-1917 (1)
- Strains (1)
- Sugar-transport (1)
- Suicide vector (1)
- SuperSAGE (1)
- T cell (1)
- T cells (1)
- T helper cell (1)
- T lymphocytes (1)
- T-cell (1)
- TEM (1)
- TRNA(ASP) (1)
- Tissue (1)
- Toll-like receptors (1)
- Toxicity ; plasmids (1)
- Toxoplasma gondii (1)
- TraDIS (1)
- Tracking (1)
- Transcription (1)
- Transcription profiling (1)
- Transcription start site (1)
- Transcriptome (1)
- Transcriptome analysis (1)
- Transfer-RNA modification (1)
- Translation efficiency (1)
- Transposon insertion sequencing (1)
- Typhimurium (1)
- UKZ (1)
- UME6 (1)
- Urinary tract (1)
- Uropatbcgenie E. coli (1)
- VEGF (1)
- Vaccinia virus (1)
- Valeriana wallichii (1)
- Variability (1)
- Variants (1)
- Vascular endothelial Growth Factor (1)
- Virologie (1)
- Virulence (1)
- Virusinfektion (1)
- WD40 (1)
- Yersinia (1)
- Yersinia-pseudotuberculosis (1)
- YoelII-Nigeriensis (1)
- Ypk1 (1)
- abscesses (1)
- acetivorans C2A (1)
- acetones (1)
- acid (1)
- acrican trypanosomes (1)
- actinomycetes (1)
- activation mechanism (1)
- acute myeloid leukemia (1)
- adaptation phase (1)
- adults (1)
- agroinfiltration (1)
- alignment (1)
- alignment clustering (1)
- alpha defensins (1)
- alternative pig farming (1)
- alveolar fibrosis (1)
- alveolar regeneration (1)
- animal model (1)
- animal models (1)
- animal-model (1)
- annotation (1)
- anti-biofilm (1)
- anti-schistosomal activity (1)
- anti-sigma factor (1)
- antibacterial (1)
- antibacterial drug resistance (1)
- antibacterials (1)
- antibiotic (1)
- antibiotic production (1)
- antifungal drug (1)
- antigenic variation (1)
- antimicrobial activity (1)
- antimicrobial peptides (1)
- antitermination (1)
- arabidopsis (1)
- aspergillosis (1)
- aspergillus fumigatus (1)
- atypical tetracyclines (1)
- autophagy-related (1)
- azobenzenes (1)
- bacterial biofilms (1)
- bacterial genomes (1)
- bacterial genomics (1)
- bacterial immune evasion (1)
- bacterial infection (1)
- bacterial infection model (1)
- bacterial lipoproteins (1)
- bacterial migration (1)
- bacterial pathogen (1)
- bacterial resistance (1)
- bacterial secretion (1)
- bacterial signal molecule (1)
- bacterial transcription (1)
- bacterial virulence (1)
- bacteriophage (1)
- balancing selection (1)
- beta-glucuronidase (1)
- beta-lactamases (1)
- binding (1)
- binding protein HFQ (1)
- biocide polyhexamethylene biguanide (1)
- biofilm (1)
- biofilm regulation (1)
- biolog (1)
- biological scaffold (1)
- bioluminescence (1)
- bioorthogonal SAM analogue ProSeDMA (1)
- biophotonic imaging (1)
- biosynthetic gene-cluster (1)
- blood (1)
- blood-brain barrier (1)
- blood-cerebrospinal fluid barrier (1)
- blood-stream forms (1)
- brain endothelial cells (1)
- breast-tumors (1)
- broad-spectrum antibiotics (1)
- burkholderia cenocepacia (1)
- caffeic acid bornyl ester (1)
- campestris PV vesicatoria (1)
- campylobacter (1)
- cancer treatment (1)
- cancers and neoplasms (1)
- candida albicans (1)
- carbon metabolism (1)
- carcinoma (1)
- carcinomas (1)
- carvacrol (1)
- casposon (1)
- catalytic (1)
- cell cultures (1)
- cell cycle (1)
- cell death (1)
- cell differentiation (1)
- cell fusion (1)
- cell signalling (1)
- cell vaccines (1)
- cell wall (1)
- cell wall synthesis (1)
- cell-cycle arrest (1)
- cell-line (1)
- cepedia complex (1)
- chaperone HFQ (1)
- chaperone Hfq (1)
- characterization (1)
- chelocardins (1)
- children (1)
- chlamydomonas flagella (1)
- chromosomal genes (1)
- citral (1)
- citrus (1)
- clinical (1)
- clinical isolates (1)
- clonal analysis (1)
- coagulase-negative staphylococci (1)
- cold-shock protein (1)
- coli nissel 1917 (1)
- colony morphotypes (1)
- colorectal (1)
- colorectal cancer (1)
- community settings (1)
- comparative sequence analysis (1)
- complete genome sequence (1)
- complex (1)
- components (1)
- conditional mutants (1)
- conditional promoter replacement (1)
- conservation (1)
- conserving surgery (1)
- consesus (1)
- contact lens (1)
- cost-effectiveness (1)
- crystal structure (1)
- crystal-structures (1)
- cytolethal distending toxin (1)
- dRNA-seq (1)
- dark-matter (1)
- database (1)
- decay (1)
- deep sequencing (1)
- deepSuperSAGE (1)
- deficient mice (1)
- deletion mutagenesis (1)
- dendritic cells (1)
- dependent gene-expression (1)
- dependent protein-kinase (1)
- derivatives (1)
- determines pathgenicity (1)
- diarrhea (1)
- differential (1)
- directional RNA-Seq (1)
- disease (1)
- dogs (1)
- domain (1)
- domain genes A3(2) (1)
- down regulation (1)
- drospophila (1)
- drug design (1)
- drug resistance evolution (1)
- drug-delivery systems (1)
- dual function (1)
- dynamic programming (1)
- eigenvector centrality (1)
- electron tomography (1)
- electrophilic stress (1)
- elements (1)
- emotional behavior (1)
- emulsions (1)
- endocytosis (1)
- endoribonuclease (1)
- energy metabolism (1)
- enteric pathogens (1)
- enterica serovar thphimurium (1)
- enterobacteria (1)
- enterococci (1)
- environment (1)
- environmental regulation (1)
- enzyme APOBEC3G (1)
- enzyme-linked immunoassays (1)
- ephitelial cells (1)
- epithelial-mesenchymal transition (1)
- epithelium (1)
- escherichia coli K-12 (1)
- essential genes (1)
- essential oils (1)
- ethanol (1)
- eugenyl cinnamate (1)
- eukaryotes (1)
- experimental visceral leishmaniasis (1)
- expression mutants (1)
- extracellular domain (1)
- extracellular enzymes (1)
- extraintestinal E. coli (1)
- extraintestinal isolates (1)
- extrapulmonary (1)
- falciparum (1)
- family (1)
- fatty acids (1)
- fibronectin-binding protein (1)
- filamentous Salmonella Typhimurium (1)
- finder using symmetrical best hits (1)
- flagellar basal body (1)
- flagellum (1)
- flotillin (1)
- flow cytometry (1)
- fluconazole resistance (1)
- fluorescence imaging (1)
- fluorescent probe (1)
- fluorescent protein (1)
- flux balance analysis (1)
- food industry (1)
- functional genomics (1)
- fungal genetics (1)
- fungal pathogens (1)
- fungi (1)
- gallotannins (1)
- gamete (1)
- gametes (1)
- gametocyte (1)
- gastric bypass (1)
- gastrointestinal infections (1)
- gene expression heterogeneity (1)
- gene family targeting (1)
- gene probes (1)
- gene therapy (1)
- gene-cloning (1)
- gene-cluster (1)
- gene-expression (1)
- general stress response (1)
- genetic organization (1)
- genetic recombination (1)
- genetic transcription (1)
- genome wide (1)
- genome-wide analysis (1)
- genomes (1)
- genomic deletions (1)
- genomic islands (1)
- genomic libraries (1)
- genomic library construction (1)
- genomics (1)
- genus Aspergillus (1)
- geometry (1)
- germination (1)
- global gene expression (1)
- glv-1h68 (1)
- glycolysis (1)
- glycosome (1)
- glycosyl phosphatidyl-inostitols. (1)
- gram-negative bacteria (1)
- green fluorescent protein (1)
- growth (1)
- growth rate control (1)
- haemoproteus-columbae (1)
- halstedii JM8 (1)
- harmonia axyridis (1)
- harmonine (1)
- hela cells (1)
- hemolytic-uremic syndrome (1)
- heterogeneity (1)
- heterologous production (1)
- heteropathogenicity (1)
- hexose transporter (1)
- hidden markov-models (1)
- histidine kinases (1)
- histology (1)
- homologs (1)
- homology search (1)
- horizontal gene transfer (1)
- hos tcells (1)
- host (1)
- host (organism) (1)
- host pathogen interaction (1)
- host-cell invasion (1)
- human herpesvirus 6 (1)
- human macrophages (1)
- human pathogenic fungi (1)
- human pathogens (1)
- human sodium iodide symporter (hNIS) (1)
- humanized mice (1)
- hydrogen regulation (1)
- hydrogen-peroxide (1)
- hyperexpression techniques (1)
- hypersensitive response (1)
- hyperthermophile (1)
- hyphae (1)
- hypothalamic gene expression (1)
- immune response (1)
- immunity (1)
- immunization (1)
- immunodeficiency-virus type-1 (1)
- in vivo (1)
- in-vitro propagation (1)
- inactivation (1)
- infectious disease (1)
- inflammatory-bowel-disease (1)
- inhibitors (1)
- insect immunity (1)
- insights (1)
- integration (1)
- interaction surfaces (1)
- interactome (1)
- interferon-gamma (1)
- intermediate host (1)
- internalization (1)
- intervention strategies (1)
- interview (1)
- intestinal enteroids (1)
- intracellular pathogen (1)
- intracellular pathogens (1)
- invasion (1)
- iron limitation (1)
- island (1)
- isolation (1)
- kingdom (1)
- latency (1)
- lead structure (1)
- leaves (1)
- legionella pneumophila (1)
- leishmania (1)
- libraries (1)
- library screening (1)
- life cycle (1)
- lines (1)
- linker influence (1)
- lipids (1)
- liquid chromatography-mass spectrometry (1)
- listeria monocytogenes (1)
- livestock-associated staphylococci (1)
- locus (1)
- luciferase (1)
- lux (1)
- lymph nodes (1)
- lymphadenitis (1)
- lytic replication (1)
- mC (1)
- mCherry (1)
- macrophage infection (1)
- malaria parasite (1)
- malaria vaccine (1)
- maps (1)
- marine sponges (1)
- mass spectrometry (1)
- mastectomy (1)
- maternal separation (1)
- mating (1)
- maturation (1)
- mechanism of resistance (1)
- membrane (1)
- membrane proteins (1)
- membrane vesicles (1)
- membrane-protein topology (1)
- meningococcus (1)
- merozoite (1)
- messanger RNA (1)
- messenger-RNA decay (1)
- meta-analysis based orthology (1)
- metabolic adaptation (1)
- metastases (1)
- methanoarchaea (1)
- methanol methyltransferase isozymes (1)
- methanosarcina mazei GO1 (1)
- methicillin (1)
- miRNAs (1)
- microRNAs (1)
- microSPLiT (1)
- microbiology (1)
- microbiome (1)
- microenvironment (1)
- microneme (1)
- microtubule motor (1)
- mitochondria (1)
- model (1)
- modulation of virus replication (1)
- molecular epidemiology (1)
- molecular evolution (1)
- monocyte (1)
- morphogenesis (1)
- mouse (1)
- mucosal inflammation (1)
- multidrug-resistant bacteria (1)
- multivesicular tubule (1)
- mycobacterium tuberculosis (1)
- mycobacterium-tuberculosis (1)
- natively unstructured protein (1)
- natural antisense transcripts (1)
- natural transformation (1)
- ncRNA (1)
- necrosis-factor-alpha (1)
- neisseria gonorrhoeae (1)
- network biology (1)
- non-aureus staphylococci (1)
- noncoding RNAs (1)
- nondocing RNA (1)
- northern blotting (1)
- nosocomial pathogen (1)
- nucleotide sequence (1)
- nude-mice (1)
- nutrients (1)
- olfactomedin 4 (1)
- oligopeptides (1)
- oncolytic viral therapy (1)
- oncolytic virus (1)
- one-health approach (1)
- organic farming (1)
- organization (1)
- organohalide respiration (1)
- organoids (1)
- origin (1)
- orthology (1)
- orthology network (1)
- outbreak (1)
- outer-membrane proteins (1)
- oviedomycin (1)
- oxidative stress (1)
- pangolin (1)
- par-seqFISH (1)
- parallel evolution (1)
- parasexual recombination (1)
- parasite (1)
- paromomycin (1)
- pathogen (1)
- pathogenesis (1)
- pathogenetic organism (1)
- pathogenic bacteria (1)
- pathogenicity island 2 (1)
- peptide conjugates (1)
- peptide tyrosine tyrosine (PYY) (1)
- peptide tyrosine tyrosine 3-36 (PYY\(_{3-36}\)) (1)
- peptido-glycan associated protein (1)
- permease (1)
- persistence (1)
- persister cells (1)
- phage (1)
- phagocytes (1)
- pharmacokinetics (1)
- phased metamorphosis (1)
- phenotypic heterogeneity (1)
- phenotypic microarray (1)
- pheromones (1)
- phosphorylation (1)
- photooxidative stress (1)
- photosynthesis genes (1)
- phototrophic growth (1)
- phylogenetic analysis (1)
- phylogenetic trees (1)
- phylogeny (1)
- phytochemicals (1)
- pig farming methods (1)
- plant-made vaccines (1)
- plant-microbe interaction (1)
- plasmid-chromosome crosstalk (1)
- plasmodium (1)
- plastid targeting (1)
- point mutation (1)
- polyadenylation sites (1)
- polycistronic transcription (1)
- polymerase chain reaction (PCR) (1)
- porphyromonas gingivalis (1)
- positive selection (1)
- post-transcriptional regulation (1)
- post-transcriptionalregulation (1)
- posttranscriptional control (1)
- ppl (1)
- pre-messenger RNA (1)
- promastigotes (1)
- promoters (1)
- prophage (1)
- proteasome (1)
- protein (1)
- protein HFQ (1)
- protein expressions (1)
- protein families database (1)
- protein kinase signaling cascade (1)
- protein synthesis (1)
- protein-RNA recognition (1)
- protozoan parasite (1)
- pseudomas aeruginosa (1)
- pulmonary (1)
- purification (1)
- pyrococcus furiosus (1)
- quanititative proteomics (1)
- quinone (1)
- rRNA depletion (1)
- radiation-therapy (1)
- radioiodine therapy (1)
- ralstonia solanacearum (1)
- rat (1)
- receptor (1)
- receptors (1)
- reciprocal best hit (1)
- recombinant proteins (1)
- regionalization and organoids (1)
- regulation (1)
- regulatory small RNAs (1)
- remote sequence conservation (1)
- replication (1)
- reporter (1)
- reporter genes (1)
- repression (1)
- reputation (1)
- resistance mechanism (1)
- resistance-breaking properties (1)
- resistant Staphylococcus-aureus (1)
- reveals (1)
- rhodobacter sphaeroides (1)
- ribosomal RNA (1)
- sRNA atlas (1)
- sRNA biogenesis (1)
- salmonella enterica (1)
- sationary phase (1)
- scaffold protein (1)
- schistosoma (1)
- schistosomula (1)
- scholarly publishing (1)
- scientific publishing (1)
- secondary structure (1)
- segmentation (1)
- sequence motif analysis (1)
- sequences (1)
- sequencing protocol (1)
- serotonin (1)
- serovar Typhimurium (1)
- serum resistance (1)
- sexual stage (1)
- sfaA gene) (1)
- shiga toxin (1)
- shock sigma factor (1)
- siRNAs (1)
- signal-transduction systems (1)
- simultaneous (1)
- singlet oxygen stress (1)
- sinorhizobium fredii NGR234 (1)
- skin fatty acid (1)
- small interfering RNAs (1)
- small non-coding RNAs (1)
- small noncoding RNAs (1)
- small regulatory RNAs (1)
- sodium-iodide symporter (1)
- solubility (1)
- soluble-RNA (1)
- soluble-RNAs (1)
- sortase A (1)
- specificity (1)
- spores (1)
- stability (1)
- stage-i (1)
- staib agar (1)
- staphilococci (1)
- staphylocccal infection/epidemiology (1)
- staphylococcus (1)
- stationary phase (1)
- stomach (1)
- strain (1)
- strain Newman (1)
- streptococcus pneumoniae (1)
- stress resistance (1)
- stress response (1)
- stringency (1)
- structural elucidation (1)
- structural modification (1)
- structure activity (1)
- structure-activity relationship (1)
- structure–activity (1)
- subcellular localization (1)
- subpopulation (1)
- subtilis genome (1)
- sulfates (1)
- sulfur (1)
- suppressor mutation (1)
- surgery (1)
- synthetase (1)
- tag based (1)
- tandem mass-spectra (1)
- target (1)
- target recognition (1)
- tetrachloroethene (1)
- thymyl cinnamate (1)
- thyroid-cancer (1)
- tobacco (1)
- toll-like receptor 2 (1)
- topical treatment (1)
- toxin (1)
- trans-activation (1)
- transcription factor (1)
- transcription initiation site (1)
- transcription start site (1)
- transcription start sites (1)
- transcriptional control (1)
- transcriptional landscape (1)
- transcriptional regulation (1)
- transcriptional regulator (1)
- transcriptional start site (1)
- transcriptional uni (1)
- transcriptome analysis (1)
- transcriptomeanalysis (1)
- transfer RNA (1)
- translation initiation (1)
- translational initiation (1)
- translocation (1)
- transport systems (1)
- transposable elements (1)
- transposition (1)
- tuberculosis (1)
- two-component systems (1)
- two‐component system (1)
- type III protein secretion system complex (1)
- type III secretion system pathways (1)
- tyrosine kinase (1)
- ubiquitin (1)
- ulcreative colitis (1)
- untranslated regions (1)
- urinary tract infection (1)
- urinary tract infection (UTI) (1)
- urinary-tract-infection (1)
- uropathogenic Escherichia coli (1)
- uropathogenicity (1)
- uropathogens (1)
- vaccines (1)
- vacuoles (1)
- variant surface glycoprotein (1)
- version control (1)
- vibrio parahaemolyticus (1)
- viral protein-R (1)
- viral replication (1)
- virotherapy (1)
- virulence genes (1)
- virulence modulation (1)
- virus-infection (1)
- visceral leishmaniasis (1)
- wikis (1)
- xanthomonas (1)
- xanthomonas campestris (1)
- xanthomonas maltophilia (1)
- yvcK/glmR operon (1)
- zinc cluster transcription factor (1)
- zoonotic (1)
Institute
- Institut für Molekulare Infektionsbiologie (285) (remove)
Sonstige beteiligte Institutionen
- Genelux Corporation, San Diego Science Center, 3030 Bunker Hill Street, Suite 310, San Diego, California 92109, USA (1)
- Institut für Molekulare Infektionsbiologie (MIB) der Universität Würzburg (1)
- MRB Forschungszentrum für Magnet-Resonanz-Bayern e.V., Am Hubland, D-97074 Würzburg (1)
- Research Center for Infectious Diseases (ZINF), University of Wuerzburg, Wuerzburg, Germany, (1)
- Research Center of Infectious Diseases (ZINF) of the University of Wurzburg, Germany (1)
Background
Differential RNA-sequencing (dRNA-seq) is indispensable for determination of primary transcriptomes. However, using dRNA-seq data to map transcriptional start sites (TSSs) and promoters genome-wide is a bioinformatics challenge. We performed dRNA-seq of Bradyrhizobium japonicum USDA 110, the nitrogen-fixing symbiont of soybean, and developed algorithms to map TSSs and promoters.
Results
A specialized machine learning procedure for TSS recognition allowed us to map 15,923 TSSs: 14,360 in free-living bacteria, 4329 in symbiosis with soybean and 2766 in both conditions. Further, we provide proteomic evidence for 4090 proteins, among them 107 proteins corresponding to new genes and 178 proteins with N-termini different from the existing annotation (72 and 109 of them with TSS support, respectively). Guided by proteomics evidence, previously identified TSSs and TSSs experimentally validated here, we assign a score threshold to flag 14 % of the mapped TSSs as a class of lower confidence. However, this class of lower confidence contains valid TSSs of low-abundant transcripts. Moreover, we developed a de novo algorithm to identify promoter motifs upstream of mapped TSSs, which is publicly available, and found motifs mainly used in symbiosis (similar to RpoN-dependent promoters) or under both conditions (similar to RpoD-dependent promoters). Mapped TSSs and putative promoters, proteomic evidence and updated gene annotation were combined into an annotation file.
Conclusions
The genome-wide TSS and promoter maps along with the extended genome annotation of B. japonicum represent a valuable resource for future systems biology studies and for detailed analyses of individual non-coding transcripts and ORFs. Our data will also provide new insights into bacterial gene regulation during the agriculturally important symbiosis between rhizobia and legumes.
Microcin C (McC) is a peptide-nucleotide antibiotic produced by Escherichia coli cells harboring a plasmid-borne operon mccABCDE. The heptapeptide MccA is converted into McC by adenylation catalyzed by the MccB enzyme. Since MccA is a substrate for MccB, a mechanism that regulates the MccA/MccB ratio likely exists. Here, we show that transcription from a promoter located upstream of mccA directs the synthesis of two transcripts: a short highly abundant transcript containing the mccA ORF and a longer minor transcript containing mccA and downstream ORFs. The short transcript is generated when RNA polymerase terminates transcription at an intrinsic terminator located in the intergenic region between the mccA and mccB genes. The function of this terminator is strongly attenuated by upstream mcc sequences. Attenuation is relieved and transcription termination is induced when ribosome binds to the mccA ORF. Ribosome binding also makes the mccA RNA exceptionally stable. Together, these two effects-ribosome induced transcription termination and stabilization of the message-account for very high abundance of the mccA transcript that is essential for McC production. The general scheme appears to be evolutionary conserved as ribosome-induced transcription termination also occurs in a homologous operon from Helicobacter pylori.
A total of 36 Escherichia coli urinary tract isolates (UTI) of serotype 06, with different combinations of capsule ( K) and flagellin ( H) antigens, were analysed according to the outer membrane pattern (OMP), serum resistance properties, mannose-resistant hemagglutination using various types of erythrocytes, and also for the genetic presence and the expression of Pfimbriae. S fimbriae/F1 C fimbriae, Type 1 fimbriae, aerobactin and hemolysin. Twenty selected strains were further analysed by pulsed field gel electrophoresis (PFGE), elaborating genomic profilas by Xba I cleavage and subsequent Southern hybridization to virulence-associated DNA probes. lt could be shown that 06 UTI isolates represent a highly heterogeneaus group of strains according to the occurrence and combination of these traits. Relatedness an the genetic and the phenotypic Ievei was found for some of the strains exhibiting the same 0: K: H: F serotype. DNA Iang-range mapping further indicated some interesting features, according to the copy number and the genomic linkage of virulence genes.
Escherichia coli isolates of serotype 06: K5 are the most common causative agents of cystitis and pyelonephritis in adults. To answer the question, as to whether strains of this particular serotype represent one special clonal group, out of a collection of 34 serotype 06: K5 isolates [Zingler et al. ( 1990) Zentralbl. Bakteriol Mikrobiol Hyg [A] 274:372-381] 15 strains were selected andanalyzed in detail. The flagellar (H) antigen and the outer membrane protein (OMP) pattern were determined. Furtherserum resistance properties and the genetic presence and expression of other virulence factors, including hemolysin, aerobactin, P fimbriae, S/F1C fimbriae and type 1 fimbriae was evaluated. In~laddition the Xbalmacrorestriction pattern of ten representative isolates was elaborated and the fimbrial (F) antigentype ofthe P fimbriae was determined, to obtain the complete 0: K: H: F pattern. These analyses could clearly show that the 06: K5 isolates do not represent one clonal group. The Xbal-macrorestriction profiles were heterogeneaus and marked differences in the hybridization patterns, using virulenceassociated gene probes in Southern hybridization of long-range-separated genomic DNA, were observed among the strains. However, some of strains showed similarities in the genomic profiles, arguing for clonal groupings among the 06: K5 isolates. lnterstingly the strains grouped tagether exhibited the same fimbrial F typethat many indicate a coincidence of this phenotypic trait with clonality.
Bacteria lose or gain genetic material and through selection, new variants become fixed in the population. Here we provide the first, genome-wide example of a single bacterial strain’s evolution in different deliberately colonized patients and the surprising insight that hosts appear to personalize their microflora. By first obtaining the complete genome sequence of the prototype asymptomatic bacteriuria strain E. coli 83972 and then resequencing its descendants after therapeutic bladder colonization of different patients, we identified 34 mutations, which affected metabolic and virulence-related genes. Further transcriptome and proteome analysis proved that these genome changes altered bacterial gene expression resulting in unique adaptation patterns in each patient. Our results provide evidence that, in addition to stochastic events, adaptive bacterial evolution is driven by individual host environments. Ongoing loss of gene function supports the hypothesis that evolution towards commensalism rather than virulence is favored during asymptomatic bladder colonization.
To understand the gene regulation of an organism of interest, a comprehensive genome annotation is essential. While some features, such as coding sequences, can be computationally predicted with high accuracy based purely on the genomic sequence, others, such as promoter elements or noncoding RNAs, are harder to detect. RNA sequencing (RNA-seq) has proven to be an efficient method to identify these genomic features and to improve genome annotations. However, processing and integrating RNA-seq data in order to generate high-resolution annotations is challenging, time consuming, and requires numerous steps. We have constructed a powerful and modular tool called ANNOgesic that provides the required analyses and simplifies RNA-seq-based bacterial and archaeal genome annotation. It can integrate data from conventional RNA-seq and differential RNA-seq and predicts and annotates numerous features, including small noncoding RNAs, with high precision. The software is available under an open source license (ISCL) at https://pypi.org/project/ANNOgesic/.
Agrobacterium species are capable of interkingdom gene transfer between bacteria and plants. The genome of Agrobacterium tumefaciens consists of a circular and a linear chromosome, the At-plasmid and the Ti-plasmid, which harbors bacterial virulence genes required for tumor formation in plants. Little is known about promoter sequences and the small RNA (sRNA) repertoire of this and other α-proteobacteria. We used a differential RNA sequencing (dRNA-seq) approach to map transcriptional start sites of 388 annotated genes and operons. In addition, a total number of 228 sRNAs was revealed from all four Agrobacterium replicons. Twenty-two of these were confirmed by independent RNA gel blot analysis and several sRNAs were differentially expressed in response to growth media, growth phase, temperature or pH. One sRNA from the Ti-plasmid was massively induced under virulence conditions. The presence of 76 cis-antisense sRNAs, two of them on the reverse strand of virulence genes, suggests considerable antisense transcription in Agrobacterium. The information gained from this study provides a valuable reservoir for an in-depth understanding of sRNA-mediated regulation of the complex physiology and infection process of Agrobacterium.
Motivation:
Next generation sequencing technologies have provided us with a wealth of information on genetic variation, but predi cting the functional significance of this variation is a difficult task. While many comparative genomics studies have focused on gene flux and large scale changes, relatively little attention has been paid to quantifying the effects of single nucleotide polymorphisms and indels on protein function, particularly in bacterial genomics.
Results:
We present a hidden Markov model based approach we call delta-bitscore (DBS) for identifying orthologous proteins that have diverged at the amino acid sequence level in a way that is likely to impact biological function. We benchmark this approach with several widely used datasets and apply it to a proof-of-concept study of orthologous proteomes in an investigation of host adaptation in Salmonella enterica. We highlight the value of the method in identifying functional divergence of genes, and suggest that this tool may be a better approach than the commonly used dN/dS metric for identifying functionally significant genetic changes occurring in recently diverged organisms.
FinO domain proteins such as ProQ of the model pathogen Salmonella enterica have emerged as a new class of major RNA-binding proteins in bacteria. ProQ has been shown to target hundreds of transcripts, including mRNAs from many virulence regions, but its role, if any, in bacterial pathogenesis has not been studied. Here, using a Dual RNA-seq approach to profile ProQ-dependent gene expression changes as Salmonella infects human cells, we reveal dysregulation of bacterial motility, chemotaxis, and virulence genes which is accompanied by altered MAPK (mitogen-activated protein kinase) signaling in the host. Comparison with the other major RNA chaperone in Salmonella, Hfq, reinforces the notion that these two global RNA-binding proteins work in parallel to ensure full virulence. Of newly discovered infection-associated ProQ-bound small noncoding RNAs (sRNAs), we show that the 3′UTR-derived sRNA STnc540 is capable of repressing an infection-induced magnesium transporter mRNA in a ProQ-dependent manner. Together, this comprehensive study uncovers the relevance of ProQ for Salmonella pathogenesis and highlights the importance of RNA-binding proteins in regulating bacterial virulence programs.
IMPORTANCE
The protein ProQ has recently been discovered as the centerpiece of a previously overlooked “third domain” of small RNA-mediated control of gene expression in bacteria. As in vitro work continues to reveal molecular mechanisms, it is also important to understand how ProQ affects the life cycle of bacterial pathogens as these pathogens infect eukaryotic cells. Here, we have determined how ProQ shapes Salmonella virulence and how the activities of this RNA-binding protein compare with those of Hfq, another central protein in RNA-based gene regulation in this and other bacteria. To this end, we apply global transcriptomics of pathogen and host cells during infection. In doing so, we reveal ProQ-dependent transcript changes in key virulence and host immune pathways. Moreover, we differentiate the roles of ProQ from those of Hfq during infection, for both coding and noncoding transcripts, and provide an important resource for those interested in ProQ-dependent small RNAs in enteric bacteria.
The transcriptome is a powerful proxy for the physiological state of a cell, healthy or diseased. As a result, transcriptome analysis has become a key tool in understanding the molecular changes that accompany bacterial infections of eukaryotic cells. Until recently, such transcriptomic studies have been technically limited to analyzing mRNA expression changes in either the bacterial pathogen or the infected eukaryotic host cell. However, the increasing sensitivity of high-throughput RNA sequencing now enables “dual RNA-seq” studies, simultaneously capturing all classes of coding and noncoding transcripts in both the pathogen and the host. In the five years since the concept of dual RNA-seq was introduced, the technique has been applied to a range of infection models. This has not only led to a better understanding of the physiological changes in pathogen and host during the course of an infection but has also revealed hidden molecular phenotypes of virulence-associated small noncoding RNAs that were not visible in standard infection assays. Here, we use the knowledge gained from these recent studies to suggest experimental and computational guidelines for the design of future dual RNA-seq studies. We conclude this review by discussing prospective applications of the technique.
In Staphylococcus aureus, de novo methionine biosynthesis is regulated by a unique hierarchical pathway involving stringent-response controlled CodY repression in combination with a T-box riboswitch and RNA decay. The T-box riboswitch residing in the 5′ untranslated region (met leader RNA) of the S. aureus metICFE-mdh operon controls downstream gene transcription upon interaction with uncharged methionyl-tRNA. met leader and metICFE-mdh (m)RNAs undergo RNase-mediated degradation in a process whose molecular details are poorly understood. Here we determined the secondary structure of the met leader RNA and found the element to harbor, beyond other conserved T-box riboswitch structural features, a terminator helix which is target for RNase III endoribonucleolytic cleavage. As the terminator is a thermodynamically highly stable structure, it also forms posttranscriptionally in met leader/ metICFE-mdh read-through transcripts. Cleavage by RNase III releases the met leader from metICFE-mdh mRNA and initiates RNase J-mediated degradation of the mRNA from the 5′-end. Of note, metICFE-mdh mRNA stability varies over the length of the transcript with a longer lifespan towards the 3′-end. The obtained data suggest that coordinated RNA decay represents another checkpoint in a complex regulatory network that adjusts costly methionine biosynthesis to current metabolic requirements.
Converging evidence suggests a role of serotonin (5-hydroxytryptamine, 5-HT) and tryptophan hydroxylase 2 (TPH2), the rate-limiting enzyme of 5-HT synthesis in the brain, in modulating long-term, neurobiological effects of early-life adversity. Here, we aimed at further elucidating the molecular mechanisms underlying this interaction, and its consequences for socio-emotional behaviors, with a focus on anxiety and social interaction. In this study, adult, male Tph2 null mutant (Tph2\(^{-/-}\)) and heterozygous (Tph2\(^{+/-}\)) mice, and their wildtype littermates (Tph2\(^{+/+}\)) were exposed to neonatal, maternal separation (MS) and screened for behavioral changes, followed by genome-wide RNA expression and DNA methylation profiling. In Tph2\(^{-/-}\) mice, brain 5-HT deficiency profoundly affected socio-emotional behaviors, i.e., decreased avoidance of the aversive open arms in the elevated plus-maze (EPM) as well as decreased prosocial and increased rule breaking behavior in the resident-intruder test when compared to their wildtype littermates. Tph2\(^{+/-}\) mice showed an ambiguous profile with context-dependent, behavioral responses. In the EPM they showed similar avoidance of the open arm but decreased prosocial and increased rule breaking behavior in the resident-intruder test when compared to their wildtype littermates. Notably, MS effects on behavior were subtle and depended on the Tph2 genotype, in particular increasing the observed avoidance of EPM open arms in wildtype and Tph2\(^{+/-}\) mice when compared to their Tph2\(^{-/-}\) littermates. On the genomic level, the interaction of Tph2 genotype with MS differentially affected the expression of numerous genes, of which a subset showed an overlap with DNA methylation profiles at corresponding loci. Remarkably, changes in methylation nearby and expression of the gene encoding cholecystokinin, which were inversely correlated to each other, were associated with variations in anxiety-related phenotypes. In conclusion, next to various behavioral alterations, we identified gene expression and DNA methylation profiles to be associated with TPH2 inactivation and its interaction with MS, suggesting a gene-by-environment interaction-dependent, modulatory function of brain 5-HT availability.
Background: In principle, the elimination of malignancies by oncolytic virotherapy could proceed by different mechanisms - e.g. tumor cell specific oncolysis, destruction of the tumor vasculature or an anti-tumoral immunological response. In this study, we analyzed the contribution of these factors to elucidate the responsible mechanism for regression of human breast tumor xenografts upon colonization with an attenuated vaccinia virus (VACV). Methods: Breast tumor xenografts were analyzed 6 weeks post VACV infection (p.i.; regression phase) by immunohistochemistry and mouse-specific expression arrays. Viral-mediated oncolysis was determined by tumor growth analysis combined with microscopic studies of intratumoral virus distribution. The tumor vasculature was morphologically characterized by diameter and density measurements and vessel functionality was analyzed by lectin perfusion and extravasation studies. Immunological aspects of viral-mediated tumor regression were studied in either immune-deficient mouse strains (T-, B-, NK-cell-deficient) or upon cyclophosphamide-induced immunosuppression (MHCII+-cell depletion) in nude mice. Results: Late stage VACV-infected breast tumors showed extensive necrosis, which was highly specific to cancer cells. The tumor vasculature in infected tumor areas remained functional and the endothelial cells were not infected. However, viral colonization triggers hyperpermeability and dilatation of the tumor vessels, which resembled the activated endothelium in wounded tissue. Moreover, we demonstrated an increased expression of genes involved in leukocyte-endothelial cell interaction in VACV-infected tumors, which orchestrate perivascular inflammatory cell infiltration. The immunohistochemical analysis of infected tumors displayed intense infiltration of MHCII-positive cells and colocalization of tumor vessels with MHCII+/CD31+ vascular leukocytes. However, GI-101A tumor growth analysis upon VACV-infection in either immunosuppressed nude mice (MHCII+-cell depleted) or in immune-deficient mouse strains (T-, B-, NK-cell-deficient) revealed that neither MHCII-positive immune cells nor T-, B-, or NK cells contributed significantly to VACV-mediated tumor regression. In contrast, tumors of immunosuppressed mice showed enhanced viral spreading and tumor necrosis. Conclusions: Taken together, these results indicate that VACV-mediated oncolysis is the primary mechanism of tumor shrinkage in the late regression phase. Neither the destruction of the tumor vasculature nor the massive VACV-mediated intratumoral inflammation was a prerequisite for tumor regression. We propose that approaches to enhance viral replication and spread within the tumor microenvironment should improve therapeutical outcome.
Background
Oncolytic virotherapy of tumors is an up-coming, promising therapeutic modality of cancer therapy. Unfortunately, non-invasive techniques to evaluate the inflammatory host response to treatment are rare. Here, we evaluate \(^{19}\)F magnetic resonance imaging (MRI) which enables the non-invasive visualization of inflammatory processes in pathological conditions by the use of perfluorocarbon nanoemulsions (PFC) for monitoring of oncolytic virotherapy.
Methodology/Principal Findings
The Vaccinia virus strain GLV-1h68 was used as an oncolytic agent for the treatment of different tumor models. Systemic application of PFC emulsions followed by \(^1H\)/\(^{19}\)F MRI of mock-infected and GLV-1h68-infected tumor-bearing mice revealed a significant accumulation of the \(^{19}\)F signal in the tumor rim of virus-treated mice. Histological examination of tumors confirmed a similar spatial distribution of the \(^{19}\)F signal hot spots and \(CD68^+\)-macrophages. Thereby, the \(CD68^+\)-macrophages encapsulate the GFP-positive viral infection foci. In multiple tumor models, we specifically visualized early inflammatory cell recruitment in Vaccinia virus colonized tumors. Furthermore, we documented that the \(^{19}\)F signal correlated with the extent of viral spreading within tumors.
Conclusions/Significance
These results suggest \(^{19}\)F MRI as a non-invasive methodology to document the tumor-associated host immune response as well as the extent of intratumoral viral replication. Thus, \(^{19}\)F MRI represents a new platform to non-invasively investigate the role of the host immune response for therapeutic outcome of oncolytic virotherapy and individual patient response.
Background: Recent data suggest that cancer stem cells (CSCs) play an important role in cancer, as these cells possess enhanced tumor-forming capabilities and are responsible for relapses after apparently curative therapies have been undertaken. Hence, novel cancer therapies will be needed to test for both tumor regression and CSC targeting. The use of oncolytic vaccinia virus (VACV) represents an attractive anti-tumor approach and is currently under evaluation in clinical trials. The purpose of this study was to demonstrate whether VACV does kill CSCs that are resistant to irradiation and chemotherapy.
Methods: Cancer stem-like cells were identified and separated from the human breast cancer cell line GI-101A by virtue of increased aldehyde dehydrogenase 1 (ALDH1) activity as assessed by the ALDEFLUOR assay and cancer stem cell-like features such as chemo-resistance, irradiation-resistance and tumor-initiating were confirmed in cell culture and in animal models. VACV treatments were applied to both ALDEFLUOR-positive cells in cell culture and in xenograft tumors derived from these cells. Moreover, we identified and isolated CD44\(^+\)CD24\(^+\)ESA\(^+\) cells from GI-101A upon an epithelial-mesenchymal transition (EMT). These cells were similarly characterized both in cell culture and in animal models.
Results: We demonstrated for the first time that the oncolytic VACV GLV-1h68 strain replicated more efficiently in cells with higher ALDH1 activity that possessed stem cell-like features than in cells with lower ALDH1 activity. GLV-1h68 selectively colonized and eventually eradicated xenograft tumors originating from cells with higher ALDH1 activity. Furthermore, GLV-1h68 also showed preferential replication in CD44\(^+\)CD24\(^+\)ESA\(^+\) cells derived from GI-101A upon an EMT induction as well as in xenograft tumors originating from these cells that were more tumorigenic than CD44\(^+\)CD24\(^-\)ESA\(^+\) cells.
Conclusions: Taken together, our findings indicate that GLV-1h68 efficiently replicates and kills cancer stem-like cells. Thus, GLV-1h68 may become a promising agent for eradicating both primary and metastatic tumors, especially tumors harboring cancer stem-like cells that are resistant to chemo and/or radiotherapy and may be responsible for recurrence of tumors.
Epstein-Barr virus (EBV) is best known for infection of B cells, in which it usually establishes an asymptomatic lifelong infection, but is also associated with the development of multiple B cell lymphomas. EBV also infects epithelial cells and is associated with all cases of undifferentiated nasopharyngeal carcinoma (NPC). EBV is etiologically linked with at least 8% of gastric cancer (EBVaGC) that comprises a genetically and epigenetically distinct subset of GC. Although we have a very good understanding of B cell entry and lymphomagenesis, the sequence of events leading to EBVaGC remains poorly understood. Recently, ephrin receptor A2 (EPHA2) was proposed as the epithelial cell receptor on human cancer cell lines. Although we confirm some of these results, we demonstrate that EBV does not infect healthy adult stem cell-derived gastric organoids. In matched pairs of normal and cancer-derived organoids from the same patient, EBV only reproducibly infected the cancer organoids. While there was no clear pattern of differential expression between normal and cancer organoids for EPHA2 at the RNA and protein level, the subcellular location of the protein differed markedly. Confocal microscopy showed EPHA2 localization at the cell-cell junctions in primary cells, but not in cancer cell lines. Furthermore, histologic analysis of patient tissue revealed the absence of EBV in healthy epithelium and presence of EBV in epithelial cells from inflamed tissue. These data suggest that the EPHA2 receptor is not accessible to EBV on healthy gastric epithelial cells with intact cell-cell contacts, but either this or another, yet to be identified receptor may become accessible following cellular changes induced by inflammation or transformation, rendering changes in the cellular architecture an essential prerequisite to EBV infection.
Background: Searching the orthologs of a given protein or DNA sequence is one of the most important and most commonly used Bioinformatics methods in Biology. Programs like BLAST or the orthology search engine Inparanoid can be used to find orthologs when the similarity between two sequences is sufficiently high. They however fail when the level of conservation is low. The detection of remotely conserved proteins oftentimes involves sophisticated manual intervention that is difficult to automate.
Results: Here, we introduce morFeus, a search program to find remotely conserved orthologs. Based on relaxed sequence similarity searches, morFeus selects sequences based on the similarity of their alignments to the query, tests for orthology by iterative reciprocal BLAST searches and calculates a network score for the resulting network of orthologs that is a measure of orthology independent of the E-value. Detecting remotely conserved orthologs of a protein using morFeus thus requires no manual intervention. We demonstrate the performance of morFeus by comparing it to state-of-the-art orthology resources and methods. We provide an example of remotely conserved orthologs, which were experimentally shown to be functionally equivalent in the respective organisms and therefore meet the criteria of the orthology-function conjecture.
Conclusions: Based on our results, we conclude that morFeus is a powerful and specific search method for detecting remotely conserved orthologs.
Background
During development in human erythrocytes, Plasmodium falciparum parasites display a remarkable number of adhesive proteins on their plasma membrane. In the invasive merozoites, these include members of the PfMSP1 and PfAMA1/RON complexes, which facilitate contact between merozoites and red blood cells. In gametocytes, sexual precursor cells mediating parasite transmission to the mosquito vector, plasma membrane-associated proteins primarily belong to the PfCCp and 6-cys families with roles in fertilization. This study describes a newly identified WD40-repeat protein unique to Plasmodium species that associates with adhesion protein complexes of both merozoites and gametocytes.
Methods
The WD40-repeat protein-like protein PfWLP1 was identified via co-immunoprecipitation assays followed by mass spectrometry and characterized using biochemical and immunohistochemistry methods. Reverse genetics were employed for functional analysis.
Results
PfWLP1 is expressed both in schizonts and gametocytes. In mature schizonts, the protein localizes underneath the merozoite micronemes and interacts with PfAMA1, while in gametocytes PfWLP1 primarily accumulates underneath the plasma membrane and associates with PfCCp1 and Pfs230. Reverse genetics failed to disrupt the pfwlp1 gene, while haemagglutinin-tagging was feasible, suggesting a crucial function for PfWLP1 during blood stage replication.
Conclusions
This is the first report on a plasmodial WD40-repeat protein associating with cell adhesion proteins. Since WD40 domains are known to mediate protein–protein contact by serving as a rigid scaffold for protein interactions, the presented data suggest that PfWLP1 supports the stability of adhesion protein complexes of the plasmodial blood stages.
Our body is colonized by a vast array of bacteria the sum of which forms our microbiota. The gut alone harbors >1,000 bacterial species. An understanding of their individual or synergistic contributions to human health and disease demands means to interfere with their functions on the species level. Most of the currently available antibiotics are broad‐spectrum, thus too unspecific for a selective depletion of a single species of interest from the microbiota. Programmable RNA antibiotics in the form of short antisense oligonucleotides (ASOs) promise to achieve precision manipulation of bacterial communities. These ASOs are coupled to small peptides that carry them inside the bacteria to silence mRNAs of essential genes, for example, to target antibiotic‐resistant pathogens as an alternative to standard antibiotics. There is already proof‐of‐principle with diverse bacteria, but many open questions remain with respect to true species specificity, potential off‐targeting, choice of peptides for delivery, bacterial resistance mechanisms and the host response. While there is unlikely a one‐fits‐all solution for all microbiome species, I will discuss how recent progress in bacterial RNA biology may help to accelerate the development of programmable RNA antibiotics for microbiome editing and other applications.
We investigated the roJe of Escherichia coU expressing mannose-resistant hemagglutination and adhesins with regard to the induction of leukotrienes from a suspension of human lymphocytes, monocytes, and basophils (LMBs) compared with human polymorphonuclear granulocytes (PMNs). Genetically cloned E. coli strains expressing various types of mannose-resistant hemagglutination (MRH+) were phagocytosed to a higher degree by monocytes than the nonadherent E. coli strain. The various strains dUfered in their capacity to induce a chemiluminescence response, which showed the same pattern for LMBs and PMNs. Stimulation of LMBs with bacteria alone, unlike granulocytes, did not activate the cells for the release of leukotrienes. However, preincubation of LMBs with bacteria decreased subsequent leukotriene formation when the cells were stimulated with calcium ionophore. The inhibitory eft'ect was dependent on the concentration of bacteria used for preincubation as weil as on the preincubation temperature. The various bacterial strains dift'ered in inhibitory potency for mediator release. Preincubation of LMBs with zymosan, opsonized zymosan, the bacterfal peptide FMLP, and peptidoglycan bad no inhibitory eft'ect or even increased subsequent IeukotrieDe formation. Opsonized bacteria were far less inhibitory than nonopsonized bacteria. In contrast to human LMBs, preincubation of human PMNs with mannose-resistant bacteria led to increased leukotriene 84 generation and reduced w-oxidation of leukotriene 84 • Our data soggest that phagocytes (neutrophils, monocytes) respond in a different way for leukotriene formation after Interaction with mannose-resistant E. coli.
The eukaryotic unicellular pathogen Plasmodium falciparum tightly regulates gene expression, both during development and in adaptation to dynamic host environments. This regulation is evident in the mutually exclusive expression of members of clonally variant virulence multigene families. While epigenetic regulators have been selectively identified at active or repressed virulence genes, their specific recruitment remains a mystery. In recent years, noncoding RNAs (ncRNAs) have emerged as lynchpins of eukaryotic gene regulation; by binding to epigenetic regulators, they provide target specificity to otherwise non-specific enzyme complexes. Not surprisingly, there is great interest in understanding the role of ncRNA in P. falciparum, in particular, their contribution to the mutually exclusive expression of virulence genes. The current repertoire of P. falciparum ncRNAs includes, but is not limited to, subtelomeric ncRNAs, virulence gene-associated ncRNAs and natural antisense RNA transcripts. Continued improvement in high-throughput sequencing methods is sure to expand this repertoire. Here, we summarize recent advances in P. falciparum ncRNA biology, with an emphasis on ncRNA-mediated epigenetic modes of gene regulation.
F 1 C fimbriae allow uropathogenic Escherichia coli to adhere to specific epithelial surfaces. This adhesive property is probably due to the presence of minor fimbrial components in F1C fimbriae. The foe gene cluster encoding F1C fimbriae has been cloned, as described previously. Here we present the nucleotide sequence (2081 bp) coding for the F 1 C minor fimbria I subunits. The structural genes code for polypeptides of 175 (FocF), 166 (FocG), and 300 (FocH) amino acids. The deduced amino acids of the F 1 C minor subunits were compared with the reported sequences of the minor subunits of other types of fimbriae. The data show that the Foc minor subunits are highly homologous to the corresponding Sfa proteins, whereas homology to the minor subunits of type 1 and P fimbriae is much lower.
As multidrug-resistant bacteria represent a concerning burden, experts insist on the need for a dramatic rethinking on antibiotic use and development in order to avoid a post-antibiotic era. New and rapidly developable strategies for antimicrobial substances, in particular substances highly potent against multidrug-resistant bacteria, are urgently required. Some of the treatment options currently available for multidrug-resistant bacteria are considerably limited by side effects and unfavorable pharmacokinetics. The glycopeptide vancomycin is considered an antibiotic of last resort. Its use is challenged by bacterial strains exhibiting various types of resistance. Therefore, in this study, highly active polycationic peptide-vancomycin conjugates with varying linker characteristics or the addition of PEG moieties were synthesized to optimize pharmacokinetics while retaining or even increasing antimicrobial activity in comparison to vancomycin. The antimicrobial activity of the novel conjugates was determined by microdilution assays on susceptible and vancomycin-resistant bacterial strains. VAN1 and VAN2, the most promising linker-modified derivatives, were further characterized in vivo with molecular imaging and biodistribution studies in rodents, showing that the linker moiety influences both antimicrobial activity and pharmacokinetics. Encouragingly, VAN2 was able to undercut the resistance breakpoint in microdilution assays on vanB and vanC vancomycin-resistant enterococci. Out of all PEGylated derivatives, VAN:PEG1 and VAN:PEG3 were able to overcome vanC resistance. Biodistribution studies of the novel derivatives revealed significant changes in pharmacokinetics when compared with vancomycin. In conclusion, linker modification of vancomycin-polycationic peptide conjugates represents a promising strategy for the modulation of pharmacokinetic behavior while providing potent antimicrobial activity.
Multidrug‐resistant bacteria represent one of the biggest challenges facing modern medicine. The increasing prevalence of glycopeptide resistance compromises the efficacy of vancomycin, for a long time considered as the last resort for the treatment of resistant bacteria. To reestablish its activity, polycationic peptides were conjugated to vancomycin. By site‐specific conjugation, derivatives that bear the peptide moiety at four different sites of the antibiotic were synthesized. The most potent compounds exhibited an approximately 1000‐fold increased antimicrobial activity and were able to overcome the most important types of vancomycin resistance. Additional blocking experiments using d‐Ala‐d‐Ala revealed a mode of action beyond inhibition of cell‐wall formation. The antimicrobial potential of the lead candidate FU002 for bacterial infection treatments could be demonstrated in an in vivo study. Molecular imaging and biodistribution studies revealed that conjugation engenders superior pharmacokinetics.
Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) loci are found in bacterial and archaeal genomes where they provide the molecular machinery for acquisition of immunity against foreign DNA. In addition to the cas genes fundamentally required for CRISPR activity, a second class of genes is associated with the CRISPR loci, of which many have no reported function in CRISPR-mediated immunity. Here, we characterize MM_0565 associated to the type I-B CRISPR-locus of Methanosarcina mazei Gö1. We show that purified MM_0565 composed of a CRISPR-Cas Associated Rossmann Fold (CARF) and a winged helix-turn-helix domain forms a dimer in solution; in vivo, the dimeric MM_0565 is strongly stabilized under high salt stress. While direct effects on CRISPR-Cas transcription were not detected by genetic approaches, specific binding of MM_0565 to the leader region of both CRISPR-Cas systems was observed by microscale thermophoresis and electromobility shift assays. Moreover, overexpression of MM_0565 strongly induced transcription of the cas1-solo gene located in the recently reported casposon, the gene product of which shows high similarity to classical Cas1 proteins. Based on our findings, and taking the absence of the expressed CRISPR locus-encoded Cas1 protein into account, we hypothesize that MM_0565 might modulate the activity of the CRISPR systems on different levels.
Escherichia coli 0139: K82: H1 strains originating from outbreaks and single cases of oedema disease in pigs were characterized by their genomic restriction fragment length polymorphism (RFLP), their virulence pattern, and by the occurrence as well as the genomic distribution of the determinants for hemolysin (hly) and verotoxins (shiga-like toxins; sltI, sltII). Whereas the RFLPs revealed considerable variation among the E. coli 0139: K82: H1 isolates depending the origin and epidemic source of the strains, the virulence gene slt II was found to be present in nearly all strains in a particular chromosomal region. Similar to RFLPs, the plasmid profiles are useful for epidemiological analysis.
Ten monoclonal antibodies (McAbs) raised to Schistosoma japonicum eggs could be assigned using several serological and immunochemical techniques to 3 groups. The McAbs, termed A, B and C-McAbs, apparently recognize carbohydrate epitopes that can be located on the same antigen molecule. The antibodies, generally of IgM isotype, are idiotypically related. They are distinct from another IgM McAb (Group D-McAb) the carbohydrate target epitope of which can also be associated with the epitopes of A. B and C-McAbs. The McAbs produce large vacuolated bleb reactions in the circumoval precipitin test (COPT) and target epitopes have different representations in various life cycle stages such as immature and mature eggs, male and female worms (including S. mansoni). Antigens affinity purified on columns containing A, B, C and D-McAbs stimulate proliferation of T cells from egg-sensitized mice and elicit DTH reactions in such mice. This raises the possibility that the target antigens of these carbohydrate-reactive monoclonal antibodies are immunopathologic and involved in egg-induced granuloma formation.
Many pathogenic bacteria utilize specialized secretion systems to deliver proteins called effectors into eukaryotic cells for manipulation of host pathways. The vast majority of known effector targets are host proteins, whereas a potential targeting of host nucleic acids remains little explored. There is only one family of effectors known to target DNA directly, and effectors binding host RNA are unknown. Here, we take a two-pronged approach to search for RNA-binding effectors, combining biocomputational prediction of RNA-binding domains (RBDs) in a newly assembled comprehensive dataset of bacterial secreted proteins, and experimental screening for RNA binding in mammalian cells. Only a small subset of effectors were predicted to carry an RBD, indicating that if RNA targeting was common, it would likely involve new types of RBDs. Our experimental evaluation of effectors with predicted RBDs further argues for a general paucity of RNA binding activities amongst bacterial effectors. We obtained evidence that PipB2 and Lpg2844, effector proteins of Salmonella and Legionella species, respectively, may harbor novel biochemical activities. Our study presenting the first systematic evaluation of the RNA-targeting potential of bacterial effectors offers a basis for discussion of whether or not host RNA is a prominent target of secreted bacterial proteins.
Background: Gametogenesis and fertilization play crucial roles in malaria transmission. While male gametes are thought to be amongst the simplest eukaryotic cells and are proven targets of transmission blocking immunity, little is known about their molecular organization. For example, the pathway of energy metabolism that power motility, a feature that facilitates gamete encounter and fertilization, is unknown.
Methods: Plasmodium berghei microgametes were purified and analysed by whole-cell proteomic analysis for the first time. Data are available via ProteomeXchange with identifier PXD001163.
Results: 615 proteins were recovered, they included all male gamete proteins described thus far. Amongst them were the 11 enzymes of the glycolytic pathway. The hexose transporter was localized to the gamete plasma membrane and it was shown that microgamete motility can be suppressed effectively by inhibitors of this transporter and of the glycolytic pathway.
Conclusions: This study describes the first whole-cell proteomic analysis of the malaria male gamete. It identifies glycolysis as the likely exclusive source of energy for flagellar beat, and provides new insights in original features of Plasmodium flagellar organization.
Background
Malignant pleural effusion (MPE) is associated with advanced stages of lung cancer and is mainly dependent on invasion of the pleura and expression of vascular endothelial growth factor (VEGF) by cancer cells. As MPE indicates an incurable disease with limited palliative treatment options and poor outcome, there is an urgent need for new and efficient treatment options.
Methods
In this study, we used subcutaneously generated PC14PE6 lung adenocarcinoma xenografts in athymic mice that developed subcutaneous malignant effusions (ME) which mimic pleural effusions of the orthotopic model. Using this approach monitoring of therapeutic intervention was facilitated by direct observation of subcutaneous ME formation without the need of sacrificing mice or special imaging equipment as in case of MPE. Further, we tested oncolytic virotherapy using Vaccinia virus as a novel treatment modality against ME in this subcutaneous PC14PE6 xenograft model of advanced lung adenocarcinoma.
Results
We demonstrated significant therapeutic efficacy of Vaccinia virus treatment of both advanced lung adenocarcinoma and tumor-associated ME. We attribute the efficacy to the virus-mediated reduction of tumor cell-derived VEGF levels in tumors, decreased invasion of tumor cells into the peritumoral tissue, and to viral infection of the blood vessel-invading tumor cells. Moreover, we showed that the use of oncolytic Vaccinia virus encoding for a single-chain antibody (scAb) against VEGF (GLAF-1) significantly enhanced mono-therapy of oncolytic treatment.
Conclusions
Here, we demonstrate for the first time that oncolytic virotherapy using tumor-specific Vaccinia virus represents a novel and promising treatment modality for therapy of ME associated with advanced lung cancer.
Bacterial small RNAs (sRNAs) are widespread post-transcriptional regulators that control bacterial stress responses and virulence. Nevertheless, little is known about how they arise and evolve. Homologs can be difficult to identify beyond the strain level using sequence-based approaches, and similar functionalities can arise by convergent evolution. Here, we found that the virulence-associated CJnc190 sRNA of the foodborne pathogen Campylobacter jejuni resembles the RepG sRNA from the gastric pathogen Helicobacter pylori. However, while both sRNAs bind G-rich sites in their target mRNAs using a C/U-rich loop, they largely differ in their biogenesis. RepG is transcribed from a stand-alone gene and does not require processing, whereas CJnc190 is transcribed from two promoters as precursors that are processed by RNase III and also has a cis-encoded antagonist, CJnc180. By comparing CJnc190 homologs in diverse Campylobacter species, we show that RNase III-dependent processing of CJnc190 appears to be a conserved feature even outside of C. jejuni. We also demonstrate the CJnc180 antisense partner is expressed in C. coli, yet here might be derived from the 3’UTR (untranslated region) of an upstream flagella-related gene. Our analysis of G-tract targeting sRNAs in Epsilonproteobacteria demonstrates that similar sRNAs can have markedly different biogenesis pathways.
Analysis of host microRNA function uncovers a role for miR-29b-2-5p in Shigella capture by filopodia
(2017)
MicroRNAs play an important role in the interplay between bacterial pathogens and host cells, participating as host defense mechanisms, as well as exploited by bacteria to subvert host cellular functions. Here, we show that microRNAs modulate infection by Shigella flexneri, a major causative agent of bacillary dysentery in humans. Specifically, we characterize the dual regulatory role of miR-29b-2-5p during infection, showing that this microRNA strongly favors Shigella infection by promoting both bacterial binding to host cells and intracellular replication. Using a combination of transcriptome analysis and targeted high-content RNAi screening, we identify UNC5C as a direct target of miR-29b-2-5p and show its pivotal role in the modulation of Shigella binding to host cells. MiR-29b-2-5p, through repression of UNC5C, strongly enhances filopodia formation thus increasing Shigella capture and promoting bacterial invasion. The increase of filopodia formation mediated by miR-29b-2-5p is dependent on RhoF and Cdc42 Rho-GTPases. Interestingly, the levels of miR-29b-2-5p, but not of other mature microRNAs from the same precursor, are decreased upon Shigella replication at late times post-infection, through degradation of the mature microRNA by the exonuclease PNPT1. While the relatively high basal levels of miR-29b-2-5p at the start of infection ensure efficient Shigella capture by host cell filopodia, dampening of miR-29b-2-5p levels later during infection may constitute a bacterial strategy to favor a balanced intracellular replication to avoid premature cell death and favor dissemination to neighboring cells, or alternatively, part of the host response to counteract Shigella infection. Overall, these findings reveal a previously unappreciated role of microRNAs, and in particular miR-29b-2-5p, in the interaction of Shigella with host cells.
Staphylococcus aureus is a major human pathogen, which can invade and survive in non-professional and professional phagocytes. Uptake by host cells is thought to contribute to pathogenicity and persistence of the bacterium. Upon internalization by epithelial cells, cytotoxic S. aureus strains can escape from the phagosome, replicate in the cytosol and induce host cell death. Here, we identified a staphylococcal cysteine protease to induce cell death after translocation of intracellular S. aureus into the host cell cytoplasm. We demonstrated that loss of staphopain A function leads to delayed onset of host cell death and prolonged intracellular replication of S. aureus in epithelial cells. Overexpression of staphopain A in a non-cytotoxic strain facilitated intracellular killing of the host cell even in the absence of detectable intracellular replication. Moreover, staphopain A contributed to efficient colonization of the lung in a mouse pneumonia model. In phagocytic cells, where intracellular S. aureus is exclusively localized in the phagosome, staphopain A did not contribute to cytotoxicity. Our study suggests that staphopain A is utilized by S. aureus to exit the epithelial host cell and thus contributes to tissue destruction and dissemination of infection.
Author summary Staphylococcus aureus is an antibiotic-resistant pathogen that emerges in hospital and community settings and can cause a variety of diseases ranging from skin abscesses to lung inflammation and blood poisoning. The bacterium can asymptomatically colonize the upper respiratory tract and skin of humans and take advantage of opportune conditions, like immunodeficiency or breached barriers, to cause infection. Although S. aureus was not regarded as intracellular bacterium, it can be internalized by human cells and subsequently exit the host cells by induction of cell death, which is considered to cause tissue destruction and spread of infection. The bacterial virulence factors and underlying molecular mechanisms involved in the intracellular lifestyle of S. aureus remain largely unknown. We identified a bacterial cysteine protease to contribute to host cell death of epithelial cells mediated by intracellular S. aureus. Staphopain A induced killing of the host cell after translocation of the pathogen into the cell cytosol, while bacterial proliferation was not required. Further, the protease enhanced survival of the pathogen during lung infection. These findings reveal a novel, intracellular role for the bacterial protease staphopain A.
Non-aureus staphylococci (NAS) are ubiquitous bacteria in livestock-associated environments where they may act as reservoirs of antimicrobial resistance (AMR) genes for pathogens such as Staphylococcus aureus. Here, we tested whether housing conditions in pig farms could influence the overall AMR-NAS burden. Two hundred and forty porcine commensal and environmental NAS isolates from three different farm types (conventional, alternative, and organic) were tested for phenotypic antimicrobial susceptibility and subjected to whole genome sequencing. Genomic data were analysed regarding species identity and AMR gene carriage. Seventeen different NAS species were identified across all farm types. In contrast to conventional farms, no AMR genes were detectable towards methicillin, aminoglycosides, and phenicols in organic farms. Additionally, AMR genes to macrolides and tetracycline were rare among NAS in organic farms, while such genes were common in conventional husbandries. No differences in AMR detection existed between farm types regarding fosfomycin, lincosamides, fusidic acid, and heavy metal resistance gene presence. The combined data show that husbandry conditions influence the occurrence of resistant and multidrug-resistant bacteria in livestock, suggesting that changing husbandry practices may be an appropriate means of limiting the spread of AMR bacteria on farms.
H-Y-specific and H-2Db-restricted, Lyt-1 "2+ T-cell clones (CTLL) with graded specific cytotoxic activities on male C57BL/6 (B6) target cells (1E3, +++; 2C5, ++; 2A5, +, 3E6, ±) were tested for their capacity to inhibit the generation of H-Y-specific cytotoxic T lymphocytes (CTL) in vitro. Addition of irradiated lymphocytes of CTLL 1E3 and CTLL 3E6 but not those of CTLL 2A5 or CTLL 2C5 abolished the generation of CTL from in vivo primed H-Y-specific precursor cells (CTLP) when added to fresh mixed-lymphocyte cultures (MLC). Exogenous sources of T-cell growth factors (TCGF) did not overcome suppression. Rather the presence of TCGF resulted in a further enhancement of suppressive activities in CTLL 1E3 and 3E6 and the induction of similar activities in cells from CTLL 2A5 and 2C5, which by themselves were not inhibitory. Moreover when added to similar MLC on Day 1 instead of Day 0, only irradiated cells of CTLL 3E6 but not those of the other three CTLL were suppressive. Induction of suppressive activities in H-Y-specific CTLL was independent of the appropriate male stimulator cells since it was also observed in MLC induced by irrelevant antigens (H-2, trinitrophenol). Furthermore at low cell numbers, irradiated lymphocytes from any of the CTLL consistently enhanced CTL activities generated from H-Y-specific CTLP. This augmenting activity, which was not TCGF, could be transferred by soluble mediators present in antigen-sensitized CTLL cultures. Thus, these data indicate (i) that cytotoxic effector cells can function as suppressor cells in the generation of CTL, (ii) that the cytotoxic activity of cloned CTL does not correlate with their capacity to suppress CTL responses, (iii) that the inhibition of CTL responses by CTLL is not due to simple consumption of T-cell growth factors produced in MLC, and (iv) that different CTL clones may interfere with the generation of CTL at different stages of their maturation. Moreover, the experiments suggest an antigen-independent enhancement of suppression by the interaction of CTL with lymphokines. Together with the augmenting activity evoked by cloned CTL the data provide strong evidence for the expression of multiple immunological functions by one particular subset of T cells and suggest that cytotoxic effector cells can differentially regulate the maturation and/or clonal expression of their precursor cells.
In this study we report that cloned Thy-l +, L3T4-, Lyt-l-, Lyt-2+, H-Y-specific and H-2Db-restricted cytotoxic T ce11 lines (CTLL) when indueed by lectin or antigen secrete a soluble mediator(s) (SF) that inhibits proliferation and generation of cytotoxic lymphocytes (CTL) in mixed lymphocyte cultures (MLC). The biological activity was separable by gel filtration and appeared as a broad peak in the moleeular mass range between 10000 and 50000 kDa. It was found that the suppressive activity released by CTLL neither strictly correlates with their cytotoxic potential nor with their ability to produce immune interferon or Iymphotoxin. SF was shown to elicitits activity in an antigen-nonspeeific manner in that it suppressed the maturation of T lymphocytes responding to both, the appropriate H-Y antigen as weH as to unrelated H_2d alloantigens or to the hapten 2,4,6-trinitrophenyl (TNP). The effect of SF on CTL responses was most pronounced in early phases of primary or secondary MLC. When analyzed for its inhibitory activity on precursor ceHs in populations selected for either Lyt-2- or L3T4- lymphocytes, it was found that SF interfered with the maturation of both subsets. The inhibition of CTL responses elicited by SF could not be reversed by the addition of exogenous interleukin 2. The findtng that SF also inhi. bited the proliferation of some but not a11 antigen-dependent cloned T ceHs with helper or eytc'toxic potential provides evidence that the faetor also may regulate effector lymphl)cytes. In addition, the results support the assumption that SF exerts its effect direetly on the responder rather than the stimulator population, and demonstrate that the development of CTL from their preeursor eeHs is contro11ed at least in part by the eytotoxic effeetor cells themselves via a soluble factor(s) that interferes with distinct stages of T ce11 maturation. These findings again emphasize the expression of multiple functions by CTL and indieate their possible role du ring the course of an immune response by their capability to eliminate target cells and to secrete a soluble product(s) that mediates feedback contro!.
Background
Advances in high-throughput sequencing have led to the discovery of widespread transcription of natural antisense transcripts (NATs) in a large number of organisms, where these transcripts have been shown to play important roles in the regulation of gene expression. Likewise, the existence of NATs has been observed in Plasmodium but our understanding towards their genome-wide distribution remains incomplete due to the limited depth and uncertainties in the level of strand specificity of previous datasets.
Results
To gain insights into the genome-wide distribution of NATs in P. falciparum, we performed RNA-ligation based strand-specific RNA sequencing at unprecedented depth. Our data indicate that 78.3% of the genome is transcribed during blood-stage development. Moreover, our analysis reveals significant levels of antisense transcription from at least 24% of protein-coding genes and that while expression levels of NATs change during the intraerythrocytic developmental cycle (IDC), they do not correlate with the corresponding mRNA levels. Interestingly, antisense transcription is not evenly distributed across coding regions (CDSs) but strongly clustered towards the 3′-end of CDSs. Furthermore, for a significant subset of NATs, transcript levels correlate with mRNA levels of neighboring genes.
Finally, we were able to identify the polyadenylation sites (PASs) for a subset of NATs, demonstrating that at least some NATs are polyadenylated. We also mapped the PASs of 3443 coding genes, yielding an average 3′ untranslated region length of 523 bp.
Conclusions
Our strand-specific analysis of the P. falciparum transcriptome expands and strengthens the existing body of evidence that antisense transcription is a substantial phenomenon in P. falciparum. For a subset of neighboring genes we find that sense and antisense transcript levels are intricately linked while other NATs appear to be regulated independently of mRNA transcription. Our deep strand-specific dataset will provide a valuable resource for the precise determination of expression levels as it separates sense from antisense transcript levels, which we find to often significantly differ. In addition, the extensive novel data on 3′ UTR length will allow others to perform searches for regulatory motifs in the UTRs and help understand post-translational regulation in P. falciparum.
Campylobacter jejuni is currently the leading cause of bacterial gastroenteritis in humans. Comparison of multiple Campylobacter strains revealed a high genetic and phenotypic diversity. However, little is known about differences in transcriptome organization, gene expression, and small RNA (sRNA) repertoires. Here we present the first comparative primary transcriptome analysis based on the differential RNA–seq (dRNA–seq) of four C. jejuni isolates. Our approach includes a novel, generic method for the automated annotation of transcriptional start sites (TSS), which allowed us to provide genome-wide promoter maps in the analyzed strains. These global TSS maps are refined through the integration of a SuperGenome approach that allows for a comparative TSS annotation by mapping RNA–seq data of multiple strains into a common coordinate system derived from a whole-genome alignment. Considering the steadily increasing amount of RNA–seq studies, our automated TSS annotation will not only facilitate transcriptome annotation for a wider range of pro- and eukaryotes but can also be adapted for the analysis among different growth or stress conditions. Our comparative dRNA–seq analysis revealed conservation of most TSS, but also single-nucleotide-polymorphisms (SNP) in promoter regions, which lead to strain-specific transcriptional output. Furthermore, we identified strain-specific sRNA repertoires that could contribute to differential gene regulation among strains. In addition, we identified a novel minimal CRISPR-system in Campylobacter of the type-II CRISPR subtype, which relies on the host factor RNase III and a trans-encoded sRNA for maturation of crRNAs. This minimal system of Campylobacter, which seems active in only some strains, employs a unique maturation pathway, since the crRNAs are transcribed from individual promoters in the upstream repeats and thereby minimize the requirements for the maturation machinery. Overall, our study provides new insights into strain-specific transcriptome organization and sRNAs, and reveals genes that could modulate phenotypic variation among strains despite high conservation at the DNA level.
RNA-binding proteins (RBPs) have been established as core components of several post-transcriptional gene regulation mechanisms. Experimental techniques such as cross-linking and co-immunoprecipitation have enabled the identification of RBPs, RNA-binding domains (RBDs) and their regulatory roles in the eukaryotic species such as human and yeast in large-scale. In contrast, our knowledge of the number and potential diversity of RBPs in bacteria is poorer due to the technical challenges associated with the existing global screening approaches. We introduce APRICOT, a computational pipeline for the sequence-based identification and characterization of proteins using RBDs known from experimental studies. The pipeline identifies functional motifs in protein sequences using position-specific scoring matrices and Hidden Markov Models of the functional domains and statistically scores them based on a series of sequence-based features. Subsequently, APRICOT identifies putative RBPs and characterizes them by several biological properties. Here we demonstrate the application and adaptability of the pipeline on large-scale protein sets, including the bacterial proteome of Escherichia coli. APRICOT showed better performance on various datasets compared to other existing tools for the sequence-based prediction of RBPs by achieving an average sensitivity and specificity of 0.90 and 0.91 respectively. The command-line tool and its documentation are available at https://pypi.python.org/pypi/bio-apricot.
The pathogen Staphylococcus aureus causes a broad range of severe diseases and is feared for its ability to rapidly develop resistance to antibiotic substances. The increasing number of highly resistant S. aureus infections has accelerated the search for alternative treatment options to close the widening gap in anti-S. aureus therapy. This study analyses the humoral immune response to vaccination of Balb/c mice with sublethal doses of live S. aureus. The elicited antibody pattern in the sera of intravenously and intramuscularly vaccinated mice was determined using of a recently developed protein array. We observed a specific antibody response against a broad set of S. aureus antigens which was stronger following i.v. than i.m. vaccination. Intravenous but not intramuscular vaccination protected mice against an intramuscular challenge infection with a high bacterial dose. Vaccine protection was correlated with the strength of the anti-S. aureus antibody response. This study identified novel vaccine candidates by using protein microarrays as an effective tool and showed that successful vaccination against S. aureus relies on the optimal route of administration.
Defeat of the antibiotic resistance of pathogenic bacteria is one great challenge today and for the future. In the last century many classes of effective antibacterials have been developed, so that upcoming resistances could be met with novel drugs of various compound classes. Meanwhile, there is a certain lack of research of the pharmaceutical companies, and thus there are missing developments of novel antibiotics. Gram-positive bacteria are the most important cause of clinical infections. The number of novel antibacterials in clinical trials is strongly restricted. There is an urgent need to find novel antibacterials. We used synthetic chemistry to build completely novel hybrid molecules of substituted indoles and benzothiophene. In a simple one-pot reaction, two novel types of thienocarbazoles were yielded. Both indole substituted compound classes have been evaluated as completely novel antibacterials against the Staphylococcus and Enterococcus species. The evaluated partly promising activities depend on the indole substituent type. First lead compounds have been evaluated within in vivo studies. They confirmed the in vitro results for the new classes of small-molecule antibacterials.
Increasing antibacterial drug resistance threatens global health, unfortunately, however, efforts to find novel antibacterial agents have been scaled back by the pharmaceutical industry due to concerns about a poor return on investment. Nevertheless, there is an urgent need to find novel antibacterial compounds to combat antibacterial drug resistance. The synthesis of novel drugs from natural sources is mostly cost-intensive due to those drugs’ complicated structures. Therefore, it is necessary to find novel antibacterials by simple synthesis to become more attractive for industrial production. We succeeded in the discovery of four antibacterial compound (sub)classes accessible in a simple one-pot reaction based on fluorinated benzothiophene-indole hybrids. They have been evaluated against various S. aureus and MRSA strains. Structure- and substituent-dependent activities have been found within the (sub)classes and promising lead compounds have been identified. In addition, bacterial pyruvate kinase was found to be the molecular target of the active compounds. In conclusion, simple one-pot synthesis of benzothiophene-indoles represents a promising strategy for the search of novel antimicrobial compounds.
Abstract
In the murine model of Leishmania major infection, resistance or susceptibility to the parasite has been associated with the development of a Th1 or Th2 type of immune response. Recently, however, the immunosuppressive effects of IL-10 have been ascribed a crucial role in the development of the different clinical correlates of Leishmania infection in humans. Since T cells and professional APC are important cellular sources of IL-10, we compared leishmaniasis disease progression in T cell-specific, macrophage/neutrophil-specific and complete IL-10-deficient C57BL/6 as well as T cell-specific and complete IL-10-deficient BALB/c mice. As early as two weeks after infection of these mice with L. major, T cell-specific and complete IL-10-deficient animals showed significantly increased lesion development accompanied by a markedly elevated secretion of IFN-γ or IFN-γ and IL-4 in the lymph nodes draining the lesions of the C57BL/6 or BALB/c mutants, respectively. In contrast, macrophage/neutrophil-specific IL-10-deficient C57BL/6 mice did not show any altered phenotype. During the further course of disease, the T cell-specific as well as the complete IL-10-deficient BALB/c mice were able to control the infection. Furthermore, a dendritic cell-based vaccination against leishmaniasis efficiently suppresses the early secretion of IL-10, thus contributing to the control of parasite spread. Taken together, IL-10 secretion by T cells has an influence on immune activation early after infection and is sufficient to render BALB/c mice susceptible to an uncontrolled Leishmania major infection.
Author Summary
The clinical symptoms caused by infections with Leishmania parasites range from self-healing cutaneous to uncontrolled visceral disease and depend not only on the parasite species but also on the type of the host's immune response. It is estimated that 350 million people worldwide are at risk, with a global incidence of 1–1.5 million cases of cutaneous and 500,000 cases of visceral leishmaniasis. Murine leishmaniasis is the best-characterized model to elucidate the mechanisms underlying resistance or susceptibility to Leishmania major parasites in vivo. Using T cell-specific and macrophage-specific mutant mice, we demonstrate that abrogating the secretion of the immunosuppressive cytokine IL-10 by T cells is sufficient to render otherwise susceptible mice resistant to an infection with the pathogen. The healing phenotype is accompanied by an elevated specific inflammatory immune response very early after infection. We further show that dendritic cell-based vaccination against leishmaniasis suppresses the early secretion of IL-10 following challenge infection. Thus, our study unravels a molecular mechanism critical for host immune defense, aiding in the development of an effective vaccine against leishmaniasis.
Many microRNAs (miRNAs) are co-regulated during the same physiological process but the underlying cellular logic is often little understood. The conserved, immunomodulatory miRNAs miR-146 and miR-155, for instance, are co-induced in many cell types in response to microbial lipopolysaccharide (LPS) to feedback-repress LPS signalling through Toll-like receptor TLR4. Here, we report that these seemingly co-induced regulatory RNAs dramatically differ in their induction behaviour under various stimuli strengths and act non-redundantly through functional specialization; although miR-146 expression saturates at sub-inflammatory doses of LPS that do not trigger the messengers of inflammation markers, miR-155 remains tightly associated with the pro-inflammatory transcriptional programmes. Consequently, we found that both miRNAs control distinct mRNA target profiles; although miR-146 targets the messengers of LPS signal transduction components and thus downregulates cellular LPS sensitivity, miR-155 targets the mRNAs of genes pervasively involved in pro-inflammatory transcriptional programmes. Thus, miR-155 acts as a broad limiter of pro-inflammatory gene expression once the miR-146 dependent barrier to LPS triggered inflammation has been breached. Importantly, we also report alternative miR-155 activation by the sensing of bacterial peptidoglycan through cytoplasmic NOD-like receptor, NOD2. We predict that dosedependent responses to environmental stimuli may involve functional specialization of seemingly coinduced miRNAs in other cellular circuitries as well.
Many microRNAs (miRNAs) are co-regulated during the same physiological process but the underlying cellular logic is often little understood. The conserved, immunomodulatory miRNAs miR-146 and miR-155, for instance, are co-induced in many cell types in response to microbial lipopolysaccharide (LPS) to feedback-repress LPS signalling through Toll-like receptor TLR4. Here, we report that these seemingly co-induced regulatory RNAs dramatically differ in their induction behaviour under various stimuli strengths and act non-redundantly through functional specialization; although miR-146 expression saturates at sub-inflammatory doses of LPS that do not trigger the messengers of inflammation markers, miR-155 remains tightly associated with the pro-inflammatory transcriptional programmes. Consequently, we found that both miRNAs control distinct mRNA target profiles; although miR-146 targets the messengers of LPS signal transduction components and thus downregulates cellular LPS sensitivity, miR-155 targets the mRNAs of genes pervasively involved in pro-inflammatory transcriptional programmes. Thus, miR-155 acts as a broad limiter of pro-inflammatory gene expression once the miR-146 dependent barrier to LPS triggered inflammation has been breached. Importantly, we also report alternative miR-155 activation by the sensing of bacterial peptidoglycan through cytoplasmic NOD-like receptor, NOD2. We predict that dosedependent responses to environmental stimuli may involve functional specialization of seemingly coinduced miRNAs in other cellular circuitries as well.
A major obstacle in infection biology is the limited ability to recapitulate human disease trajectories in traditional cell culture and animal models, which impedes the translation of basic research into clinics. Here, we introduce a three-dimensional (3D) intestinal tissue model to study human enteric infections at a level of detail that is not achieved by conventional two-dimensional monocultures. Our model comprises epithelial and endothelial layers, a primary intestinal collagen scaffold, and immune cells. Upon Salmonella infection, the model mimics human gastroenteritis, in that it restricts the pathogen to the epithelial compartment, an advantage over existing mouse models. Application of dual transcriptome sequencing to the Salmonella-infected model revealed the communication of epithelial, endothelial, monocytic, and natural killer cells among each other and with the pathogen. Our results suggest that Salmonella uses its type III secretion systems to manipulate STAT3-dependent inflammatory responses locally in the epithelium without accompanying alterations in the endothelial compartment. Our approach promises to reveal further human-specific infection strategies employed by Salmonella and other pathogens.
IMPORTANCE Infection research routinely employs in vitro cell cultures or in vivo mouse models as surrogates of human hosts. Differences between murine and human immunity and the low level of complexity of traditional cell cultures, however, highlight the demand for alternative models that combine the in vivo-like properties of the human system with straightforward experimental perturbation. Here, we introduce a 3D tissue model comprising multiple cell types of the human intestinal barrier, a primary site of pathogen attack. During infection with the foodborne pathogen Salmonella enterica serovar Typhimurium, our model recapitulates human disease aspects, including pathogen restriction to the epithelial compartment, thereby deviating from the systemic infection in mice. Combination of our model with state-of-the-art genetics revealed Salmonella-mediated local manipulations of human immune responses, likely contributing to the establishment of the pathogen's infection niche. We propose the adoption of similar 3D tissue models to infection biology, to advance our understanding of molecular infection strategies employed by bacterial pathogens in their human host.
Investigations were carried out on the adhesion of cloned S-fimbriated E. coli, labelled with fluoresceinisothiocyanate (FITC) to human buccal epithelial cells. Fluorescence microscopic analysis revealed binding of bacteria to 75-95% of epithelial cells. Inhibition experiments with fetuin, a 1-acid glycoprotein and N-acetyl neuraminic acid confirmed the specificity of bacterial binding to sialoglycoproteins. Further studies using saliva as an inhibitor resulted in a 4-5 times stronger binding inhibition by newborn saliva in comparison to adult saliva coinciding with a 4-5 times higher content of total N-acetyl neuraminic acid in samples of newborn saliva. In Western blot analysis sialoglycoprotein bands with a molecular weight >200 kD reacting with wheat germ agglutinin (WGA), were only identified in samples of newborn saliva. These bands are classified as mucins on account of molecular weight and staining. These data suggest that saliva mucins could represent a major defense mechanism against bacterial infections at a stage of ontogeny where the secretory IgAsystem is not yet developed.
S-fimbriae mediated adhesin of Escherichia coli to human buccal epithelial cells is age independent
(1992)
S-fimbriated Escherichia coli, which cause sepsis and meningitis in the newbom, bind to sialic acid-containing glycoprotein structures on the surface of human buccal epithelial cells. The dependence of · this binding on host age was examined. S-fimbriated · E. coli adhered in comparable numbers to cells in newborns, infants, children and adults (23.0 ± 8.6; 23.1 ± 11.5; 24.7 ± 7.9; 28.9 ± 8.8). Thus, the increased susceptibility of neonates to infections caused by S-fimbriated E. coli cannot be explained by enhanced · adhesion to epithelial cells