VAAM-Jahrestagung 2011 Karlsruhe, 3.–6. April 2011

Conclusion: TNF-α is instrumental in inhibiting mycobacterial growth invitro and IL-1 is involved in immunoregulation and inflammation. Ourresults therefore support the notion that different LAM structures areimplicated in the pathogenesis of mycobacterial infections in humans.[1] Hamasur, B. et al (1999): Synthesis and immunologic characterisation of Mycobacteriumtuberculosis lipoarabinomannanspecific oligosaccharide protein conjugates. Vaccine 17, 2853-2861.MPP044A scavenger receptor on nasal epithelial surfaces-Animportant player in Staphylococcus aureus nasalcolonizationS. Baur*, M. Rautenberg, S. Wanner, L. Kull, C. WeidenmaierInstitute of Microbiology and Infection Medicine, Eberhard-Karls-University, Tübingen, GermanyIt has been demonstrated that cell wall glycopolymers (CWGs) are animportant part of the repertoire of adhesins that Gram-positive bacteria useto adhere to and infect host cells. Staphylococcus aureus cell wall teichoicacid (WTA) mediates adhesion to nasal epithelial cells and is a key moleculein a cotton rat model of nasal colonization. However, the appropriatereceptor on nasal epithelial cells remains elusive. Novel advances in the fieldof glycobiology suggest members of the scavenger receptor family as WTAinteraction partners on nasal epithelial surfaces. We tested this hypothesis byinhibitors against scavenger receptors in adhesion assays and detected amarked inhibition of S. aureus to nasal epithelial cells. Recently, epithelialexpression of a receptor belonging to the scavenger receptor family had beendescribed and we demonstrated expression of this receptor in nasal epithelialcells of human origin. Function blocking antibodies to this receptor inhibitedS. aureus adhesion to human epithelial cells under static and mild sheerstress conditions. We therefore propose that the strong influence of WTAmediated adhesion on nasal colonization is also an effect governed by ascavenger receptor. To further elucidate these in vitro findings in the cottonrat model of nasal colonization we established primary cultures of cotton ratnasal epithelial cells. Thereby, we were able to detect the expression of thementioned scavenger receptor. Furthermore, we could prove WTA dependedspecific binding to primary cotton rat epithelial cells by utilizing WTAlabelledlatex beads. Planned studies with a function blocking antibodyagainst the mentioned scavenger receptor should reveal its influence oncolonization in this in vivo model. Thus, we present here the first nasalepithelial WTA receptor.MPP045Response of Candida albicans to nitrogen starvationS. Ramachandra* 1 , S. Brunke 2 , B. Hube 2 , M. Brock 1,21 Department of Microbial Biochemistry and Physiology, Hans-Knöll-Institute (HKI), Jena, Germany2 Department of Microbial Pathogenicty Mechanisms, Hans-Knöll-Institute(HKI), Jena, GermanyCandida albicans is a commensal inhabitant of the intestinal tract of warmbloodedanimals, but also able to cause life-threatening oportunisticinfections in the debilitated or compromised host. During pathogenesis thefungus disseminates via the bloodstream and infections can manifest withinvarious tissues. Regardless the commensal state, in which C. albicans has tocope for nutrients with competing microorganisms, dissemination via thebloodstream with subsequent attack by immune effector cells or growthwithin tissues, acquisition of nutrients is an essential prerequisite to survivewithin the host. It has been assumed that phagocytosis by immune effectorcells generates a carbon and nitrogen limited environment and C. albicansescapes from these cells by switching from the yeast into a hyphal growthform causing mechanical pressure on host cells and eventually leading to thepathogen release. In this respect we monitored the transcriptional responseof C. albicans when shifted from a nutrient rich to a nitrogen-starvedenvironment. As expected, the translational machinery, mainly resembled byribosomal proteins was strongly downregulated within the first 30 - 60 minafter shifting the cells to starvation. However, within four to six hours atranscriptional steady state for these genes was reached. In contrast,transcription of some genes involved in amino acid metabolism was stronglyinduced, among them several genes involved in arginine metabolism. Thisinduction was in agreement with the observation of cell clumping andformation of hyphae, which has been shown to be dependent on theinduction of genes involved in arginine metabolism. In future experimentswe will add different nitrogen sources to nitrogen-starved cells to elucidatespecific adaptation mechanisms in response to the respective nitrogensource. These experiments will allow to study nutrient uptake andinterruption of this process could cause an attenuation of virulence.MPP046Pseudomonas aeruginosa population structure revisitedunder environmental focus: the impact of water qualityand phage pressureJ. Sikorski* 1 , C. Rohde 21 Molecular Systems, German Collection of Microorganisms and CellCultures (DSMZ), Braunschweig, Germany2 DSMZ, Microbiology, Braunschweig, GermanyPseudomonas aeruginosa attracts research attention as a commonopportunistic nosocomial pathogen causing severe health problems inhumans. Nevertheless, the primary habitat is the natural environment. Here,we relate the genetic diversity of 391 environmental isolates from NorthernGermany rivers to ecological factors such as river system, season ofsampling, and different levels of water quality. From representatives of 99environmental clones, also in comparison to 91 clinical isolates, wedetermine motility phenotypes, virulence factors, biofilm formation,serotype, and the resistance to seven environmental P. aeruginosa phages.The integration of genetic, ecologic, and phenotypic data shows (i) thepresence of several extended clonal complexes (ecc) which non-uniformlydistribute across different water qualities, and (ii) a disproportionate phagepressure on the ecc which apparently depends on the diverged serotypecomposition of ecc. For at least one ecc (eccB) we assume theecophysiological differences on environmental water adaptation and phageresistance to be so distinct to reinforce an environmentally driven cladogenicsplit from the remainder of P. aeruginosa. In sum, we conclude that themajority of the microevolutionary population dynamics of P. aeruginosa isshaped by the natural environment and not by the clinical habitatMPP047Expression of Plx1, an ADP-ribosylating toxin ofPaenibacillus larvaeA. Fünfhaus*, L. Poppinga, E. GenerschState Institute for Bee Research, Department of Molecular Microbiologyand Bee Diseases, Hohen Neuendorf, GermanyThe Gram-positive bacterium Paenibacillus larvae is the causative agent ofAmerican Foulbrood, a notifiable epizootic of honey bee larvae. The diseaseprocess in individual larvae can be divided into a non-invasive and aninvasive phase. The non-invasive phase at the beginning of infection ischaracterized by massive proliferation of P. larvae in the larval midgutlumen. During the invasive phase, P. larvae enters the haemocoel bybreaching the intestinal epithelium of honey bee larvae accompanied orinitiated by rounding-up of epithelial cells [4]. Toxins and proteases aremost likely involved in this process. Good toxin candidates are ADPribosylatingAB-toxins, which are expressed by P. larvae (Fünfhaus et al.,2009) and which are known to destroy the cytoskeleton [1] and cell-cellcontacts of the host [3]. AB-toxins are comprised of a catalytic A-subunitand a B-subunit which induces translocation and targeting of the host cell.We recently identified Plx1 (P. larvae toxin 1) as an ADP-ribosylating ABtoxinspecific for P. larvae genotype ERIC I [2]. For functionalcharacterization, Plx1 was cloned in an appropriate expression vector,harbouring promotor-, RBS- and start codon- sequences as well as an N-terminal His-tag. The recombinant plasmid was in vitro-translated using anE. coli-based in vitro expression system. SDS-gel-analyses revealed asuccessful translation of the N-terminal His-tagged protein. Purification ofPlx1 was performed by affinity-binding of the His-tag to magnetic nickelparticles. After binding, washing and elution of the tagged protein withimidazol it could be visualized on SDS-gels with a mass of about 111 kDa.Purified recombinant Plx1 is now available for functional assays includingin vitro cell culture- as well as larval-assays to verify toxin function and toidentify the cellular target molecule.[1] Aktories, K. and A. Wegner (1992): Mechanisms of the cytopathic action of actin-ADPribosylatingtoxins. Mol. Microbiol. 6, 2905-2908.[2] Fünfhaus, A. et al (2009): Use of suppression subtractive hybridization to identify geneticdifferences between differentially virulent genotypes of Paenibacillus larvae, the etiological agent ofAmerican foulbrood of honeybees. Environ. Microbiol. Reports 1, 240-250.[3] Sousa, S. et al (2005): Microbial strategies to target, cross or disrupt epithelia. Curr. Opin. CellBiol. 17, 489-98.spektrum | Tagungsband 2011