• Homophilic protein interactions facilitate bacterial aggregation and IgG-dependent complex formation by the Streptococcus canis M protein SCM.

      Nerlich, Andreas; Lapschies, Antje-Maria; Kohler, Thomas P; Cornax, Ingrid; Eichhorn, Inga; Goldmann, Oliver; Krienke, Petra; Bergmann, Simone; Nizet, Victor; Hammerschmidt, Sven; Rohde, Manfred; Fulde, Marcus; HZI, Helmholtz Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig Germany. (Taylor & Francis, 2019-01-01)
      Streptococcus canis is a zoonotic agent that causes serious invasive diseases in domestic animals and humans, but knowledge about its pathogenic potential and underlying virulence mechanisms is limited. Here, we report on the ability of certain S. canis isolates to form large bacterial aggregates when grown in liquid broth. Bacterial aggregation was attributed to the presence and the self-binding activity of SCM, the M protein of S. canis, as evaluated by bacterial sedimentation assays, immunofluorescence- and electron microscopic approaches. Using a variety of truncated recombinant SCM fragments, we demonstrated that homophilic SCM interactions occur via the N-terminal, but not the C-terminal part, of the mature M protein. Interestingly, when incubated in human plasma, SCM forms soluble protein complexes comprising its known ligands, immunoglobulin G (IgG) and plasminogen (Plg). Co-incubation studies with purified host proteins revealed that SCM-mediated complex formation is based on the interaction of SCM with itself and with IgG, but not with Plg or fibrinogen (Fbg), well-established constituents of M protein-mediated protein complexes in human-associated streptococci. Notably, these soluble, SCM-mediated plasma complexes harbored complement factor C1q, which can induce complement breakdown in the periphery and therefore represent another immune evasion mechanism of SCM.
    • DncV Synthesizes Cyclic GMP-AMP and Regulates Biofilm Formation and Motility in ECOR31.

      Li, Fengyang; Cimdins, Annika; Rohde, Manfred; Jänsch, Lothar; Kaever, Volkhard; Nimtz, Manfred; Römling, Ute; HZI, Helmholtz Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig Germany. (ASM, 2019-03-05)
      Cyclic dinucleotides (cDNs) act as intracellular second messengers, modulating bacterial physiology to regulate the fundamental life style transition between motility and sessility commonly known as biofilm formation. Cyclic GMP-AMP (cGAMP), synthesized by the dinucleotide cyclase DncV, is a newly discovered cDN second messenger involved in virulence and chemotaxis in Vibrio cholerae O1 biovar El Tor. Here we report a novel role for horizontally transferred DncV in cGAMP production and regulation of biofilm formation and motility in the animal commensal strain Escherichia coli ECOR31. ECOR31 expresses a semiconstitutive temperature-independent rdar (red, dry, and rough) morphotype on Congo red agar plates characterized by the extracellular matrix components cellulose and curli fimbriae which requires activation by the major biofilm regulator CsgD and cyclic di-GMP signaling. In contrast, C-terminal His-tagged DncV negatively regulates the rdar biofilm morphotype and cell aggregation via downregulation of csgD mRNA steady-state level. Furthermore, DncV sequentially promotes and inhibits adhesion to the abiotic surface after 24 h and 48 h of growth, respectively. DncV also suppresses swimming and swarming motility posttranscriptional of the class 1 flagellum regulon gene flhD Purified DncV produced different cDNs, cyclic di-GMP, cyclic di-AMP, an unknown product(s), and the dominant species 3'3'-cGAMP. In vivo, only the 3'3'-cGAMP concentration was elevated upon short-term overexpression of dncV, making this work a first report on cGAMP production in E. coli Regulation of rdar biofilm formation and motility upon overexpression of untagged DncV in combination with three adjacent cotransferred gene products suggests a novel temperature-dependent cGAMP signaling module in E. coli ECOR31.IMPORTANCE The ability of bacteria to sense and respond to environmental signals is critical for survival. Bacteria use cyclic dinucleotides as second messengers to regulate a number of physiological processes, such as the fundamental life style transition between motility and sessility (biofilm formation). cGAMP, which is synthesized by a dinucleotide cyclase called DncV, is a newly discovered second messenger involved in virulence and chemotaxis in the Vibrio cholerae biovar El Tor causing the current 7th cholera pandemic. However, to what extent cGAMP exists and participates in physiological processes in other bacteria is still unknown. In this study, we found an elevated cGAMP level to possibly regulate biofilm formation and motility in the animal commensal E. coli strain ECOR31. Thus, we detected a novel role for cGAMP signaling in regulation of physiological processes other than those previously reported in proteobacterial species.
    • Microbiome yarns: the Global Phenotype-Genotype Survey: Episode II: laryngeal microbiota and vocal phenotypes (or diction and addiction).

      Timmis, Kenneth; Jebok, Franziska; Rohde, Manfred; Molinari, Gabriella; HZI, Helmholtz Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig Germany. (Wiley-Blackwell, 2019-03-01)
    • Comparative genomic analysis of eight novel haloalkaliphilic bacteriophages from Lake Elmenteita, Kenya.

      Akhwale, Juliah Khayeli; Rohde, Manfred; Rohde, Christine; Bunk, Boyke; Spröer, Cathrin; Klenk, Hans-Peter; Boga, Hamadi Iddi; Wittmann, Johannes; HZI, Helmholtz Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig Germany. (PLOS, 2019-01-01)
      We report complete genome sequences of eight bacteriophages isolated from Haloalkaline Lake Elmenteita found on the floor of Kenyan Rift Valley. The bacteriophages were sequenced, annotated and a comparative genomic analysis using various Bioinformatics tools carried out to determine relatedness of the bacteriophages to each other, and to those in public databases. Basic genome properties like genome size, percentage coding density, number of open reading frames, percentage GC content and gene organizations revealed the bacteriophages had no relationship to each other. Comparison to other nucleotide sequences in GenBank database showed no significant similarities hence novel. At the amino acid level, phages of our study revealed mosaicism to genes with conserved domains to already described phages. Phylogenetic analyses of large terminase gene responsible for DNA packaging and DNA polymerase gene for replication further showed diversity among the bacteriophages. Our results give insight into diversity of bacteriophages in Lake Elmenteita and provide information on their evolution. By providing primary sequence information, this study not only provides novel sequences for biotechnological exploitation, but also sets stage for future studies aimed at better understanding of virus diversity and genomes from haloalkaline lakes in the Rift Valley.
    • Intracellular Streptococcal Uptake and Persistence: A Potential Cause of Erysipelas Recurrence.

      Jendoubi, Fatma; Rohde, Manfred; Prinz, Jörg Christoph; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Frontiers, 2019-01-01)
      Erysipelas is a severe streptococcal infection of the skin primarily spreading through the lymphatic vessels. Penicillin is the treatment of choice. The most common complication consists in relapses which occur in up to 40% or more of patients despite appropriate antibiotic treatment. They cause lymphatic damage resulting in irreversible lymphedema and ultimately elephantiasis nostras and lead to major health restrictions and high socio-medical costs. Prevention of relapses is an unmet need, because even long-term prophylactic penicillin application does eventually not reduce the risk of recurrence. In this article we assess risk factors and causes of erysipelas recurrence. A systematic literature search for clinical studies addressing potential causes and measures for prevention of erysipelas recurrence was combined with a review of experimental and clinical data assessing the ability and clinical relevance of streptococci for intracellular uptake and persistence. The literature review found that venous insufficiency, lymphedema, and intertrigo from fungal infections are considered to be major risk factors for recurrence of erysipelas but cannot adequately explain the high recurrence rate. As hitherto unrecognized likely cause of erysipelas relapses we identify the ability of streptococci for intracellular uptake into and persistence within epithelial and endothelial cells and macrophages. This creates intracellular streptococcal reservoirs out of reach of penicillins which do not reach sufficient bactericidal intracellular concentrations. Incomplete streptococcal elimination due to intracellular streptococcal persistence has been observed in various deep tissue infections and is considered as cause of relapsing streptococcal pharyngitis despite proper antibiotic treatment. It may also serves as endogenous infectious source of erysipelas relapses. We conclude that the current antibiotic treatment strategies and elimination of conventional risk factors employed in erysipelas management are insufficient to prevent erysipelas recurrence. The reactivation of streptococcal infection from intracellular reservoirs represents a plausible explanation for the frequent occurrence erysipelas relapses. Prevention of erysipelas relapses therefore demands for novel antibiotic strategies capable of eradicating intracellular streptococcal persistence.
    • Molecular typing of Streptococcus suis strains isolated from diseased and healthy pigs between 1996-2016.

      Prüfer, T Louise; Rohde, Judith; Verspohl, Jutta; Rohde, Manfred; de Greeff, Astrid; Willenborg, Jörg; Valentin-Weigand, Peter; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (PLOS, 2019-01-01)
      Streptococcus suis is an economically important pathogen of pigs as well as a zoonotic cause of human disease. Serotyping is used for further characterization of isolates; some serotypes seem to be more virulent and more widely spread than others. This study characterizes a collection of German field isolates of Streptococcus suis from pigs dating from 1996 to 2016 with respect to capsular genes (cps) specific for individual serotypes and pathotype by multiplex PCR and relates results to the clinical background of these isolates. The most prominent finding was the reduction in prevalence of serotype-2/serotype-1/2 among invasive isolates during this sampling period, which might be attributed to widely implemented autogenous vaccination programs in swine against serotype 2 in Germany. In diseased pigs (systemically ill; respiratory disease) isolates of serotype-1/serotype-14, serotype-2/serotype-1/2, serotype 3 to 5 and 7 to 9 were most frequent while in carrier isolates a greater variety of cps types was found. Serotype-1/serotype-14 seemed to be preferentially located in joints, serotype 4 and serotype 3 in the central nervous system, respectively. The virulence associated extracellular protein factor was almost exclusively associated with invasive serotype-1/serotype-14 and serotype-2/serotype-1/2 isolates. In contrast, lung isolates of serotype-2/serotype-1/2 mainly harbored the gene for muramidase-released protein. Serotype 4 and serotype 9 isolates from clinically diseased pigs most frequently carried the muramidase-released protein gene and the suilysin gene. When examined by transmission electron microscopy all but one of the isolates which were non-typable by molecular and serological methods showed various amounts of capsular material indicating potentially new serotypes among these isolates. Given the variety of cps types/serotypes detected in pigs, not only veterinarians but also medical doctors should consider other serotypes than just serotype 2 when investigating potential human cases of Streptococcus suis infection.
    • Day and Night: Metabolic Profiles and Evolutionary Relationships of Six Axenic Non-Marine Cyanobacteria.

      Will, Sabine Eva; Henke, Petra; Boedeker, Christian; Huang, Sixing; Brinkmann, Henner; Rohde, Manfred; Jarek, Michael; Friedl, Thomas; Seufert, Steph; Schumacher, Martin; Overmann, Jörg; Neumann-Schaal, Meina; Petersen, Jörn; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Oxford Academic, 2019-01-01)
      Cyanobacteria are dominant primary producers of various ecosystems and they colonize marine as well as freshwater and terrestrial habitats. On the basis of their oxygenic photosynthesis they are known to synthesize a high number of secondary metabolites, which makes them promising for biotechnological applications. State-of-the-art sequencing and analytical techniques and the availability of several axenic strains offer new opportunities for the understanding of the hidden metabolic potential of cyanobacteria beyond those of single model organisms. Here, we report comprehensive genomic and metabolic analyses of five non-marine cyanobacteria, that is, Nostoc sp. DSM 107007, Anabaena variabilis DSM 107003, Calothrix desertica DSM 106972, Chroococcidiopsis cubana DSM 107010, Chlorogloeopsis sp. PCC 6912, and the reference strain Synechocystis sp. PCC 6803. Five strains that are prevalently belonging to the order Nostocales represent the phylogenetic depth of clade B1, a morphologically highly diverse sister lineage of clade B2 that includes strain PCC 6803. Genome sequencing, light and scanning electron microscopy revealed the characteristics and axenicity of the analyzed strains. Phylogenetic comparisons showed the limits of the 16S rRNA gene for the classification of cyanobacteria, but documented the applicability of a multilocus sequence alignment analysis based on 43 conserved protein markers. The analysis of metabolites of the core carbon metabolism showed parts of highly conserved metabolic pathways as well as lineage specific pathways such as the glyoxylate shunt, which was acquired by cyanobacteria at least twice via horizontal gene transfer. Major metabolic changes were observed when we compared alterations between day and night samples. Furthermore, our results showed metabolic potential of cyanobacteria beyond Synechocystis sp. PCC 6803 as model organism and may encourage the cyanobacterial community to broaden their research to related organisms with higher metabolic activity in the desired pathways.
    • Mycoplasma hyopneumoniae resides intracellularly within porcine epithelial cells.

      Raymond, B B A; Turnbull, L; Jenkins, C; Madhkoor, R; Schleicher, I; Uphoff, C C; Whitchurch, C B; Rohde, M; Djordjevic, S P; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (2018-12-06)
      Enzootic pneumonia incurs major economic losses to pork production globally. The primary pathogen and causative agent, Mycoplasma hyopneumoniae, colonises ciliated epithelium and disrupts mucociliary function predisposing the upper respiratory tract to secondary pathogens. Alleviation of disease is reliant on antibiotics, vaccination, and sound animal husbandry, but none are effective at eliminating M. hyopneumoniae from large production systems. Sustainable pork production systems strive to lower reliance on antibiotics but lack of a detailed understanding of the pathobiology of M. hyopneumoniae has curtailed efforts to develop effective mitigation strategies. M. hyopneumoniae is considered an extracellular pathogen. Here we show that M. hyopneumoniae associates with integrin β1 on the surface of epithelial cells via interactions with surface-bound fibronectin and initiates signalling events that stimulate pathogen uptake into clathrin-coated vesicles (CCVs) and caveosomes. These early events allow M. hyopneumoniae to exploit an intracellular lifestyle by commandeering the endosomal pathway. Specifically, we show: (i) using a modified gentamicin protection assay that approximately 8% of M. hyopneumoniae cells reside intracellularly; (ii) integrin β1 expression specifically co-localises with the deposition of fibronectin precisely where M. hyopneumoniae cells assemble extracellularly; (iii) anti-integrin β1 antibodies block entry of M. hyopneumoniae into porcine cells; and (iv) M. hyopneumoniae survives phagolysosomal fusion, and resides within recycling endosomes that are trafficked to the cell membrane. Our data creates a paradigm shift by challenging the long-held view that M. hyopneumoniae is a strict extracellular pathogen and calls for in vivo studies to determine if M. hyopneumoniae can traffic to extrapulmonary sites in commercially-reared pigs.
    • Virulence factor-dependent basolateral invasion of choroid plexus epithelial cells by pathogenic Escherichia coli in vitro.

      Rose, Rebekah; Häuser, Svenja; Stump-Guthier, Carolin; Weiss, Christel; Rohde, Manfred; Kim, Kwang Sik; Ishikawa, Hiroshi; Schroten, Horst; Schwerk, Christian; Adam, Rüdiger; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Oxford Academic Press, 2018-11-21)
      Escherichia coli is the most common Gram-negative causative agent of neonatal meningitis and E. coli meningitis is associated with high morbidity and mortality. Previous research has been carried out with regard to the blood-brain barrier and thereby unveiled an assortment of virulence factors involved in E. coli meningitis. Little, however, is known about the role of the blood-cerebrospinal fluid (CSF) barrier (BCSFB), in spite of several studies suggesting that the choroid plexus (CP) is a possible entry point for E. coli into the CSF spaces. Here, we used a human CP papilloma (HIBCPP) cell line that was previously established as valid model for the study of the BCSFB. We show that E. coli invades HIBCPP cells in a polar fashion preferentially from the physiologically relevant basolateral side. Moreover, we demonstrate that deletion of outer membrane protein A, ibeA or neuDB genes results in decreased cell infection, while absence of fimH enhances invasion, although causing reduced adhesion to the apical side of HIBCPP cells. Our findings suggest that the BCSFB might constitute an entry point for E. coli into the central nervous system, and HIBCPP cells are a valuable tool for investigating E. coli entry of the BCSFB.
    • Iron Regulation in Clostridioides difficile.

      Berges, Mareike; Michel, Annika-Marisa; Lassek, Christian; Nuss, Aaron M; Beckstette, Michael; Dersch, Petra; Riedel, Katharina; Sievers, Susanne; Becher, Dörte; Otto, Andreas; Maaß, Sandra; Rohde, Manfred; Eckweiler, Denitsa; Borrero-de Acuña, Jose M; Jahn, Martina; Neumann-Schaal, Meina; Jahn, Dieter; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (2018-01-01)
      The response to iron limitation of several bacteria is regulated by the ferric uptake regulator (Fur). The Fur-regulated transcriptional, translational and metabolic networks of the Gram-positive, pathogen Clostridioides difficile were investigated by a combined RNA sequencing, proteomic, metabolomic and electron microscopy approach. At high iron conditions (15 μM) the C. difficile fur mutant displayed a growth deficiency compared to wild type C. difficile cells. Several iron and siderophore transporter genes were induced by Fur during low iron (0.2 μM) conditions. The major adaptation to low iron conditions was observed for the central energy metabolism. Most ferredoxin-dependent amino acid fermentations were significantly down regulated (had, etf, acd, grd, trx, bdc, hbd). The substrates of these pathways phenylalanine, leucine, glycine and some intermediates (phenylpyruvate, 2-oxo-isocaproate, 3-hydroxy-butyryl-CoA, crotonyl-CoA) accumulated, while end products like isocaproate and butyrate were found reduced. Flavodoxin (fldX) formation and riboflavin biosynthesis (rib) were enhanced, most likely to replace the missing ferredoxins. Proline reductase (prd), the corresponding ion pumping RNF complex (rnf) and the reaction product 5-aminovalerate were significantly enhanced. An ATP forming ATPase (atpCDGAHFEB) of the F0F1-type was induced while the formation of a ATP-consuming, proton-pumping V-type ATPase (atpDBAFCEKI) was decreased. The [Fe-S] enzyme-dependent pyruvate formate lyase (pfl), formate dehydrogenase (fdh) and hydrogenase (hyd) branch of glucose utilization and glycogen biosynthesis (glg) were significantly reduced, leading to an accumulation of glucose and pyruvate. The formation of [Fe-S] enzyme carbon monoxide dehydrogenase (coo) was inhibited. The fur mutant showed an increased sensitivity to vancomycin and polymyxin B. An intensive remodeling of the cell wall was observed, Polyamine biosynthesis (spe) was induced leading to an accumulation of spermine, spermidine, and putrescine. The fur mutant lost most of its flagella and motility. Finally, the CRISPR/Cas and a prophage encoding operon were downregulated. Fur binding sites were found upstream of around 20 of the regulated genes. Overall, adaptation to low iron conditions in C. difficile focused on an increase of iron import, a significant replacement of iron requiring metabolic pathways and the restructuring of the cell surface for protection during the complex adaptation phase and was only partly directly regulated by Fur.
    • Type IV Secretion System and Its Adhesin Subunit, CagL, Mediate Potent Inflammatory Responses in Primary Human Endothelial Cells.

      Tafreshi, Mona; Guan, Jyeswei; Gorrell, Rebecca J; Chew, Nicole; Xin, Yue; Deswaerte, Virginie; Rohde, Manfred; Daly, Roger J; Peek, Richard M; Jenkins, Brendan J; Davies, Elizabeth M; Kwok, Terry; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Frontiers, 2018-01-01)
      The Gram-negative bacterium, Helicobacter pylori, causes chronic gastritis, peptic ulcers, and gastric cancer in humans. Although the gastric epithelium is the primary site of H. pylori colonization, H. pylori can gain access to deeper tissues. Concurring with this notion, H. pylori has been found in the vicinity of endothelial cells in gastric submucosa. Endothelial cells play crucial roles in innate immune response, wound healing and tumorigenesis. This study examines the molecular mechanisms by which H. pylori interacts with and triggers inflammatory responses in endothelial cells. We observed that H. pylori infection of primary human endothelial cells stimulated secretion of the key inflammatory cytokines, interleukin-6 (IL-6) and interleukin-8 (IL-8). In particular, IL-8, a potent chemokine and angiogenic factor, was secreted by H. pylori-infected endothelial cells to levels ~10- to 20-fold higher than that typically observed in H. pylori-infected gastric epithelial cells. These inflammatory responses were triggered by the H. pylori type IV secretion system (T4SS) and the T4SS-associated adhesin CagL, but not the translocation substrate CagA. Moreover, in contrast to integrin α5β1 playing an essential role in IL-8 induction by H. pylori upon infection of gastric epithelial cells, both integrin α5β1 and integrin αvβ3 were dispensable for IL-8 induction in H. pylori-infected endothelial cells. However, epidermal growth factor receptor (EGFR) is crucial for mediating the potent H. pylori-induced IL-8 response in endothelial cells. This study reveals a novel mechanism by which the H. pylori T4SS and its adhesin subunit, CagL, may contribute to H. pylori pathogenesis by stimulating the endothelial innate immune responses, while highlighting EGFR as a potential therapeutic target for controlling H. pylori-induced inflammation. Introduction
    • Microbiome yarns: The Global Phenotype-Genotype Survey: Episode I: all my worldly goods, including my microbiome, I thee endow.

      Timmis, Kenneth; Jebok, Franziska; Rohde, Manfred; Molinari, Gabriella; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Wiley and Sons, 2019-01-01)
    • Macrophage entrapped silica coated superparamagnetic iron oxide particles for controlled drug release in a 3D cancer model.

      Ullah, Sami; Seidel, Katja; Türkkan, Sibel; Warwas, Dawid Peter; Dubich, Tatyana; Rohde, Manfred; Hauser, Hansjörg; Behrens, Peter; Kirschning, Andreas; Köster, Mario; Wirth, Dagmar; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (2018-12-23)
      Targeted delivery of drugs is a major challenge in treatment of diverse diseases. Systemically administered drugs demand high doses and are accompanied by poor selectivity and side effects on non-target cells. Here, we introduce a new principle for targeted drug delivery. It is based on macrophages as transporters for nanoparticle-coupled drugs as well as controlled release of drugs by hyperthermia mediated disruption of the cargo cells and simultaneous deliberation of nanoparticle-linked drugs. Hyperthermia is induced by an alternating electromagnetic field (AMF) that induces heat from silica-coated superparamagnetic iron oxide nanoparticles (SPIONs). We show proof-of-principle of controlled release by the simultaneous disruption of the cargo cells and the controlled, AMF induced release of a toxin, which was covalently linked to silica-coated SPIONs via a thermo-sensitive linker. Cells that had not been loaded with SPIONs remain unaffected. Moreover, in a 3D co-culture model we demonstrate specific killing of associated tumour cells when employing a ratio as low as 1:40 (SPION-loaded macrophage: tumour cells). Overall, our results demonstrate that AMF induced drug release from macrophage-entrapped nanoparticles is tightly controlled and may be an attractive novel strategy for targeted drug release.
    • On the relation between filament density, force generation, and protrusion rate in mesenchymal cell motility.

      Dolati, Setareh; Kage, Frieda; Mueller, Jan; Müsken, Mathias; Kirchner, Marieluise; Dittmar, Gunnar; Sixt, Michael; Rottner, Klemens; Falcke, Martin; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Amrican Society for Cell biology, 2018-11-01)
      Lamellipodia are flat membrane protrusions formed during mesenchymal motion. Polymerization at the leading edge assembles the actin filament network and generates protrusion force. How this force is supported by the network and how the assembly rate is shared between protrusion and network retrograde flow determines the protrusion rate. We use mathematical modeling to understand experiments changing the F-actin density in lamellipodia of B16-F1 melanoma cells by modulation of Arp2/3 complex activity or knockout of the formins FMNL2 and FMNL3. Cells respond to a reduction of density with a decrease of protrusion velocity, an increase in the ratio of force to filament number, but constant network assembly rate. The relation between protrusion force and tension gradient in the F-actin network and the density dependency of friction, elasticity, and viscosity of the network explain the experimental observations. The formins act as filament nucleators and elongators with differential rates. Modulation of their activity suggests an effect on network assembly rate. Contrary to these expectations, the effect of changes in elongator composition is much weaker than the consequences of the density change. We conclude that the force acting on the leading edge membrane is the force required to drive F-actin network retrograde flow.
    • The olfactory epithelium as a port of entry in neonatal neurolisteriosis.

      Pägelow, Dennis; Chhatbar, Chintan; Beineke, Andreas; Liu, Xiaokun; Nerlich, Andreas; van Vorst, Kira; Rohde, Manfred; Kalinke, Ulrich; Förster, Reinhold; Halle, Stephan; Valentin-Weigand, Peter; Hornef, Mathias W; Fulde, Marcus; TWINCORE, Zentrum für experimentelle und klinische Infektionsforschung GmbH,Feodor-Lynen Str. 7, 30625 Hannover, Germany. (2018-10-15)
      Bacterial infections of the central nervous system (CNS) remain a major cause of mortality in the neonatal population. Commonly used parenteral infection models, however, do not reflect the early course of the disease leaving this critical step of the pathogenesis largely unexplored. Here, we analyzed nasal exposure of 1-day-old newborn mice to Listeria monocytogenes (Lm). We found that nasal, but not intragastric administration, led to early CNS infection in neonate mice. In particular, upon bacterial invasion of the olfactory epithelium, Lm subsequently spread along the sensory neurons entering the brain tissue at the cribriform plate and causing a significant influx of monocytes and neutrophils. CNS infection required listeriolysin for penetration of the olfactory epithelium and ActA, a mediator of intracellular mobility, for translocation into the brain tissue. Taken together, we propose an alternative port of entry and route of infection for neonatal neurolisteriosis and present a novel infection model to mimic the clinical features of late-onset disease in human neonates.
    • Complete genome sequence of C130_2, a novel myovirus infecting pathogenic Escherichia coli and Shigella strains.

      Sváb, Domonkos; Falgenhauer, Linda; Rohde, Manfred; Chakraborty, Trinad; Tóth, István; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2018-09-20)
      The genome sequence of a novel virulent bacteriophage, named " C130_2", that is morphologically a member of the family Myoviridae is reported. The 41,775-base-pair double-stranded DNA genome of C130_2 contains 59 ORFs but exhibits overall low sequence similarity to bacteriophage genomes for which sequences are publicly available. Phylogenetic analysis indicated that C130_2 represents a new phage type. C130_2 could be propagated well on enterohemorrhagic Escherichia coli (EHEC) O157:H7 and other pathogenic E. coli strains, as well as on strains of various Shigella species.
    • Microbiome yarns: microbiome basis of memory,,.

      Timmis, Kenneth; Jebok, Franziska; Molinari, Gabriella; Rohde, Manfred; Timmis, James Kenneth; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017-01-01)
    • Lipoteichoic acid deficiency permits normal growth but impairs virulence of Streptococcus pneumoniae.

      Heß, Nathalie; Waldow, Franziska; Kohler, Thomas P; Rohde, Manfred; Kreikemeyer, Bernd; Gómez-Mejia, Alejandro; Hain, Torsten; Schwudke, Dominik; Vollmer, Waldemar; Hammerschmidt, Sven; Gisch, Nicolas; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017-12-12)
      Teichoic acid (TA), a crucial cell wall constituent of the pathobiont Streptococcus pneumoniae, is bound to peptidoglycan (wall teichoic acid, WTA) or to membrane glycolipids (lipoteichoic acid, LTA). Both TA polymers share a common precursor synthesis pathway, but differ in the final transfer of the TA chain to either peptidoglycan or a glycolipid. Here, we show that LTA exhibits a different linkage conformation compared to WTA, and identify TacL (previously known as RafX) as a putative lipoteichoic acid ligase required for LTA assembly. Pneumococcal mutants deficient in TacL lack LTA and show attenuated virulence in mouse models of acute pneumonia and systemic infections, although they grow normally in culture. Hence, LTA is important for S. pneumoniae to establish systemic infections, and TacL represents a potential target for antimicrobial drug development.
    • Microbiome Yarns: microbiome of the built environment, paranormal microbiology, and the power of single cell genomics1,2,3,4.

      Timmis, Kenneth; Jebok, Franziska; Rohde, Manfred; Molinari, Gabriella; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2018-05-16)
    • Regulation of Flagellum Biosynthesis in Response to Cell Envelope Stress in Serovar Typhimurium.

      Spöring, Imke; Felgner, Sebastian; Preuße, Matthias; Eckweiler, Denitsa; Rohde, Manfred; Häussler, Susanne; Weiss, Siegfried; Erhardt, Marc (2018-05-01)
      Flagellum-driven motility of serovar Typhimurium facilitates host colonization. However, the large extracellular flagellum is also a prime target for the immune system. As consequence, expression of flagella is bistable within a population of , resulting in flagellated and nonflagellated subpopulations. This allows the bacteria to maximize fitness in hostile environments. The degenerate EAL domain protein RflP (formerly YdiV) is responsible for the bistable expression of flagella by directing the flagellar master regulatory complex FlhDC with respect to proteolytic degradation. Information concerning the environmental cues controlling expression of and thus about the bistable flagellar biosynthesis remains ambiguous. Here, we demonstrated that RflP responds to cell envelope stress and alterations of outer membrane integrity. Lipopolysaccharide (LPS) truncation mutants of Typhimurium exhibited increasing motility defects due to downregulation of flagellar gene expression. Transposon mutagenesis and genetic profiling revealed that σ (RpoE) and Rcs phosphorelay-dependent cell envelope stress response systems sense modifications of the lipopolysaccaride, low pH, and activity of the complement system. This subsequently results in activation of RflP expression and degradation of FlhDC via ClpXP. We speculate that the presence of diverse hostile environments inside the host might result in cell envelope damage and would thus trigger the repression of resource-costly and immunogenic flagellum biosynthesis via activation of the cell envelope stress response. Pathogenic bacteria such as Typhimurium sense and adapt to a multitude of changing and stressful environments during host infection. At the initial stage of gastrointestinal colonization, uses flagellum-mediated motility to reach preferred sites of infection. However, the flagellum also constitutes a prime target for the host's immune response. Accordingly, the pathogen needs to determine the spatiotemporal stage of infection and control flagellar biosynthesis in a robust manner. We found that uses signals from cell envelope stress-sensing systems to turn off production of flagella. We speculate that downregulation of flagellum synthesis after cell envelope damage in hostile environments aids survival of during late stages of infection and provides a means to escape recognition by the immune system.