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

sequencing of the 16S rRNA genes for hitherto underrepresented termitetaxa. Clear differences in community structure between the bacterial gutmicrobiota of lower and higher termites reflect the importance of the fiberdigestingflagellates as a bacterial habitat (e.g., the loss of the abundantendosymbiotic Endomicrobia). The strong increase of potentially cellulolyticFibrobacteres in wood-feeding taxa and the decrease of Spirochaetes and theconcomitant appearance of apparently alkali-adapted Firmicutes in soilfeedingtaxa correlate with the nutritional specialization of their termite host.Our results document the evolution of specific gut microbial communities ineach lineage of higher termites and will help to better understand thefunction of the gut microbiota in the digestive process.SIP009Analysis of Anthraquinones biosynthesis in PhotorhabdusQ. Zhou*, H.B. BodeInstitute for Molecular Bio Science, Goethe-University, Frankfurt am Main,GermanyThe entomopathogenic bacterium Photorhabdus luminescens lives insymbiosis with nematodes of the genus Heterohabditis. Among other naturalproducts, P. luminescens produces anthraquinones (AQ). Earlier studies inour group have shown that a type II polyketide synthase (PKS) isresponsible for the biosynthesis of AQ, which is only the second example oftype II PKS production in Gram-negative bacteria 1 . Unlike expected fromthe heptaketide backbone of AQ, Brachmann et al. have shown that AQ arederived from an octaketide precursor, as deletion of gene encoding acyclase/aromatase AntH resulted in the formation of octaketides, which werealready known as shunt-products in in the octaketide natural productactinorhodin from Streptomyces coelicolor A3(2) 2 .We continued to investigate the AQ biosynthesis by deletion of cyclaseencoding gene antA, which resulted in production of additional typicalpolyketide shunt-products. Furthermore, AQ biosynthesis was successfullyreconstituted in Escherichia coli. Thus, partial heterologous expression ofthe ant cluster allowed the detailed investigation of the AQ biosynthesis.Recently, the type II PKS components were expressed in E. coli and purifiedfor the activity assays. Therefore, the mechanisms of the biosynthesis andspecificities of the proteins can now be investigated in vitro 3 . The results ofthe analysis will be presented on our poster.[1] Brachmann, A.O. et al (2007): Chembiochem, 8, 1721-1728.[2] Mcdaniel, R. et al (1993): Science, 262, 1546-1550.[3] Zhang, W. and T. Tang (2009): Methods Enzymol. 459, 367-393.SIP010Losing the partner - now what? Effects of host loss ontranscription in arbuscular mycorrhizal fungiF. Zielinski*, T. Netzker, T. FesterDepartment of Environmental Microbiology, Helmholtz Center forEnvironmental Research (USZ), Leipzig, GermanyArbuscular mycorrhizal fungi are an ancient fungal phylum(Glomeromycota) that coevolved with plants for the last 400 million years,assisting the colonization of land masses by higher plants. Today, thesefungi associate with 70-90% of all plant roots and form intimate and, inmost cases, mutualistic symbioses. Collectively referred to as arbuscularmycorrhiza (AM), this association is regarded as the most widespreadterrestrial symbiosis. The interaction is mainly characterized by fungalarbuscules, i.e. tree-shaped subcellular structures within plant root cells thatare the main site of nutrient exchange between the fungal and plantsymbiotic partners. The fungal partner provides water, phosphate and othernutrients which are taken up via its extensive hyphal network from the soil.In return AM fungi obtain carbohydrates from their plant partner. Up to 20%of the photosynthesis products of terrestrial plants (roughly 5 billion tonnesof carbon per year) are estimated to be consumed by AM fungi. Therefore,the AM symbiosis contributes considerably to global phosphate and carboncycling and influences primary productivity in terrestrial ecosystems.While the symbiosis is well characterized in regard to nutrient and signalexchange, little is known regarding the senescence (ageing and death) ofAM fungi. The life time of extraradical hyphae (i.e. hyphae outside the root)has been shown to be relatively short, on average 5-6 days, and intraradicalhyphae (hyphae inside the root) as well as arbuscules have been described tobe turned over equally rapidly. The aim of our experiments is to characterizethis turnover on a molecular level. Using carrot root cultures colonized byGlomus irregulare (DAOM197198) we induced fungal senescence byseparating AM fungi from their plant partner. Host loss eventually resultedin the termination of fungal cytoplasmic streaming after several days.Currently, we compare mRNA extracts of vital and senescent fungal culturesby suppressive subtractive hybridization to detect differentially expressedgenes. Key mRNA transcripts defined in this project will be used asmolecular markers in field studies directed at assessing the presence,abundance, and activity of AM fungi in various ecosystems and undersimulated conditions of climate and land use change.SIP011Detection of Differences between Specific BacterialGroups of the Intestinal Flora of Adipose Personscompared to Normal Weighted PersonsC. Wallendorf*, C. Lange*, J. van Harsselaar, L.E. Weimer*, S. Ratering,S. SchnellInstitut for Applied Microbiology, Justus-Liebig-University, Gießen,GermanyObesity epidemic is global phenomena, affecting both the developed anddeveloping world. Development of obesity is due to many factors such asless exercise, excessive nutrition, genetics or illnesses. Recent work hasshown that the intestinal flora may also have an influence on the energybalance of the host due to interactions of the intestinal flora and the host. Forunderstanding these observation in more detail stool samples were collectedof 10 normal weighted (mean body mass index (kg/m²) of 22.84) and 11obese (mean body mass index (kg/m²) of 40.54) healthy persons in ahomogeneously group regarding to age and gender. After extraction of theDNA from the stool samples real-time quantitative PCRs were performedwith different primers sets detecting mayor groups of intestinal flora e. g.Bacteroidetes and Firmicutes. Further detailed characterizations of thecommunity were done using the same primer sets for cloning and PCR-SCCP with subsequent sequencing of the 16S rRNA gene of the DNA-bandsor the vector inserts. First results showed significantly higher Firmicutes16S rRNA gene targets in the obese group compared to the control groupwhereas no differences in the target numbers were found between the groupsby primer sets for Bacteroidetes and total Bacteria. DNA-Band pattern ofPCR-SCCP with Firmicutes specific primer showed a prominent band in thesamples of the obese.SIP012Biosynthesis of xenocyloins, secondary metabolites fromXenorhabdus bovieniiA. Proschak*, Q. Zhou, H.B. BodeInstitute for Molecular Bio Science, Goethe-University, Frankfurt am Main,GermanyXenorhabdus are Gram-negative bacteria belonging to the family ofEnterobacteriaceae. They live in a symbiotic association with soil dwellingnematodes of the genus Steinernema. Once the nematodes infect an insectlarvae, the bacteria start to proliferate in the hemocoel of the insect andproduce bioactive secondary metabolites to inhibit the insect immune systemand to protect their food source from other microorganisms [1]. Formerwork demonstrated that indole derived compounds show strong antifungaland antibacterial activity [2].As we are interested in the biosynthesis of bioactive compounds from thesebacteria [3] we searched for the biosynthesis gene cluster of these indolederivatives that we named xenocyloins in the producer strain X. bovienii SS-2004. We assumed, an acetolactate synthase like enzyme must be the keystepto the xenocyloin biosynthesis and subsequently could identify onebiosynthesis gene cluster, which was only present in X. bovienii but not innon-producers like X. nematophila or related Photorhabdus species.Heterologous expression of the predicted xenocyloin biosynthesis genecluster in E. coli DH10B confirmed that this cluster is indeed responsible forxenocyloin biosynthesis. Structure elucidation of known and new derivativeswas obtained by detailed NMR and HPLC-MS experiments. Their absolutestereochemistry was determined by CD spectroscopy. Expression of thisgene cluster into a transaminase-deficient E. coli strain DL39 allowed thedetailed elucidation of the biosynthesis via specific feeding experiments [4]and allowed the differentiation between the incorporation of amino acids andtheir respective keto acids. Recently we could demonstrate that not theputative esterase but a gene encoding a putative beta-oxoacyl(ACP)synthaseIII is involved in the esterification of hydroxylated xenoxyloins. Moreover,results from deletion of acetolactatesynthases in X. bovienii genome will bepresented on our poster.[1] Goodrich-Blair, H. and DJ Clarke (2007): Mol Microbiol, 64(2), 260-8.spektrum | Tagungsband 2011