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Lantibiotics (lanthionine-containing antibiotics) from Gram-positive bacteria typically exhibit activity against Gram-positive bacteria. The activity and structure of pinensin A (1) and B (2), lantibiotics isolated from a native Gram-negative producer Chitinophaga pinensis are described. Surprisingly, the pinensins were found to be highly active against many filamentous fungi and yeasts but show only weak antibacterial activity. To the best of our knowledge, lantibiotic fungicides have not been described before. An in-depth bioinformatic analysis of the biosynthetic gene cluster established the ribosomal origin of these compounds and identified candidate genes encoding all of the enzymes required for post-translational modification. Additional encoded functions enabled us to build up a hypothesis for the biosynthesis, export, sensing, and import of this intriguing lantibiotic.

The antibiotic elansolid C1 (8) was isolated from Chitinophaga sancti strain FxGBF13 after fermentation in the presence of anthranilic acid. Remarkably, 8 was also obtained by addition of anthranilic acid to a crude fermentation extract containing the macrolide elansolid A2 (1*). This Michael-type conjugate addition allowed us to generate 21 new derivatives of elansolid C1 (9-29) by using various nucleophiles. Biological activities of all derivatives were evaluated against Staphylococcus aureus, Micrococcus luteus, and the mouse cell line L929.

Sulfangolids are the first sulfate ester containing secondary metabolites from myxobacteria. The metabolites 1-4 and the structurally related kulkenon (5) were isolated from different strains of the species Sorangium cellulosum. In the course of isolation all metabolites proved to be rather sensitive due to their conjugated double bond systems and the strong acidic nature of the sulfate ester in sulfangolids. The relative configuration of sulfangolid C (3) was assigned by extensive 1D and 2D NMR analysis and molecular modelling. In addition, the biosynthesis of 3 was studied by feeding experiments.

Antibiotic TA is a macrocyclic secondary metabolite produced by myxobacteria that has broad-spectrum bactericidal activity. The structure of TA is unique, and its molecular target is unknown. Here, we sought to elucidate TA's mode of action (MOA) through two parallel genetic approaches. First, chromosomal Escherichia coli TA-resistant mutants were isolated. One mutant that showed specific resistance toward TA was mapped and resulted from an IS 4 insertion in the lpp gene, which encodes an abundant outer membrane (Braun's) lipoprotein. In a second approach, the comprehensive E. coli ASKA plasmid library was screened for overexpressing clones that conferred TA r . This effort resulted in the isolation of the lspA gene, which encodes the type II signal peptidase that cleaves signal sequences from prolipoproteins. In whole cells, TA was shown to inhibit Lpp prolipoprotein processing, similar to the known LspA inhibitor globomycin. Based on genetic evidence and prior globomycin studies, a block in Lpp expression or prevention of Lpp covalent cell wall attachment confers TA r by alleviating a toxic buildup of mislocalized pro-Lpp. Taken together, these data argue that LspA is the molecular target of TA. Strikingly, the giant ta biosynthetic gene cluster encodes two lspA paralogs that we hypothesize play a role in producer strain resistance.

Roimatacene (1) was isolated from the myxobacterium Cystobacter ferrugineus strain Cb G35 in a bioactivity-guided process, by following the antimicrobial activity against Escherichia coli. Since 1 was extremely sensitive to oxygen, a protective isolation and handling protocol was developed, by utilizing the free radical scavenger 4-ethoxyphenol. The structure of 1 was determined by HRMS, 1D and 2D NMR spectroscopy and chemical derivatization to acetonides and Mosher esters to finally establish the absolute configuration. Methionine and acetate were identified as building blocks in the biosynthesis of 1 by feeding experiments with differently (13)C-labeled precursors. The antimicrobial activity of 1 was determined in a broad screening revealing 1 to inhibit several Gram-negative bacteria.

The gram-negative myxobacterium Sorangium cellulosum So ce56 bears the largest bacterial genome published so far, coding for nearly 10,000 genes. Careful analysis of this genome data revealed that part of the genes coding for the very well conserved biosynthesis of lipopolysaccharides (LPS) are missing in this microbe. Biochemical analysis gave no evidence for the presence of LPS in the membranes of So ce56. By analyzing the lipid composition of its outer membrane sphingolipids were identified as the major lipid class, together with ornithine-containing lipids (OL) and ether lipids. A detailed analysis of these lipids resulted in the identification of more than 50 structural variants within these three classes, which possessed several interesting properties regarding to LPS replacement, mediators in myxobacterial differentiation, as well as potential bioactive properties. The sphingolipids with the basic structure C9-methyl-C(20)-sphingosine possessed as an unusual trait C9-methylation, which is common to fungi but highly uncommon to bacteria. Such sphingolipids have not been found in bacteria before, and they may have a function in myxobacterial development. The OL, also identified in myxobacteria for the first time, contained acyloxyacyl groups, which are also characteristic for LPS and might replace those in certain functions. Finally, the ether lipids may serve as biomarkers in myxobacterial development.

An expanded neighbour-joining tree of myxobacteria is presented based on the analysis of 16S rRNA gene sequences of 101 strains (including types) representing 3 suborders, 6 families, 20 genera, 46 species, and 12 other novel taxa. The distinctions amongst members of the three suborders (Sorangiineae, Cytobacterineae and Nannocystineae) are reaffirmed. The positions of anaerobic myxobacteria, novel groups (Pyxidicoccus and several Cystobacter species) in Cystobacterineae, the marine genera (Plesiocystis, Haliangium, Enhygromyxa), and two additional novel taxa ('Paraliomyxa miuraensis', brackish-water isolate) were together revealed for the first time. Changes in the nomenclature of several isolates (Polyangium vitellinum Pl vt1(T), Polyangium thaxteri Pl t3, Polyangium cellulosum, NOSO-1, NOCB-2, NOCB-4) are also highlighted. Suborders Sorangiineae and Nannocystineae hold great promise for novel strain discovery. In Sorangiineae, the new family Phaselicystidaceae, with a monotypic genus, was added. Nine additional novel taxa were discovered in this suborder for which new genera or even families may be erected in the near future. These taxa appear to represent the so-called viable but not culturable (VBNC) group of myxobacteria. Based on at least 4% phylogenetic distance, new clades were formed comprising of novel Nannocystineae and Sorangiineae isolates. Overall, the myxobacteria, on the basis of bracket distance, could be divided into 16 clusters, as supported by tree topology and a morphology-based approach.

2-Methyltetrahydrothiophen-3-one (3) is a volatile compound that plays an important role especially in food and flavour chemistry because it contributes to the aroma of several foodstuffs including wine. Although 3 can be formed by chemical reactions during food preparation, it is also produced by microorganisms. Recent studies with yeasts showed that methionine (1) is a potential precursor of 3, but the mechanism of the transformation is unknown. The biosynthetic pathway leading to 3 in the bacterium Chitinophaga Fx7914 was probed. Extensive feeding experiments with differently labelled precursors by using liquid cultures of Chitinophaga Fx7914 were performed. The volatiles released by the bacterium were collected by using a closed loop stripping apparatus (CLSA) and analysed by GC-MS. The observed incorporation pattern of the precursors into 3 led to the elucidation of the biosynthetic pathway. One part of the compound 2 originates from homocysteine (15), which is transformed into 3-mercaptopropanal (17). The second biosynthetic building block is pyruvate (14). An acyloin-forming reaction furnishes the key intermediate 21, which cyclises intramolecularly to a diol. Dehydration followed by tautomerisation lead to the cyclic ketone 3, which is produced by the bacterium in racemic form.

Summary Interruption of the (p)ppGpp synthetase gene (rel) of Sorangium cellulosum So ce56 resulted in loss of ppGpp accumulation after norvaline treatment during exponential growth phase. The rel mutant failed to produce wild-type levels of the polyketides chivosazol and etnangien in production media. In wild-type cells expression of the chivosazol biosynthetic operon can be significantly increased by norvaline or α-methylglucoside. This induction does not occur in the rel mutant. The rel mutant also lost the capability to form multicellular fruiting bodies under nutrient starvation.

Sorangium cellulosum strains produce approximately 50% of the biologically active secondary metabolites known from myxobacteria. These metabolites include several compounds of biotechnological importance such as the epothilones and chivosazols, which, respectively, stabilize the tubulin and actin skeletons of eukaryotic cells. S. cellulosum is characterized by its slow growth rate, and natural products are typically produced in low yield. In this study, biomagnetic bead separation of promoter-binding proteins and subsequent inactivation experiments were employed to identify the chivosazol regulator, ChiR, as a positive regulator of chivosazol biosynthesis in the genome-sequenced strain So ce56. Overexpression of chiR under the control of T7A1 promoter in a merodiploid mutant resulted in fivefold overproduction of chivosazol in a kinetic shake flask experiment, and 2.5-fold overproduction by fermentation. Using quantitative reverse transcription PCR and gel shift experiments employing heterologously expressed ChiR, we have shown that transcription of the chivosazol biosynthetic genes (chiA-chiF) is directly controlled by this protein. In addition, we have demonstrated that ChiR serves as a pleiotropic regulator in S. cellulosum, because mutant strains lack the ability to develop into regular fruiting bodies.

SummaryAn operon consisting of three open reading frames, annotated in silico as methylmalonyl-CoA (mm-CoA) epimerase, mm-CoA mutase (MCM), and meaB, was identified in the sequencing project of the myxobacterium Sorangium cellulosum So ce56. This putative MCM pathway operon was subcloned from a bacterial artificial chromosome by Red/ET recombineering onto a minimal replicon derived from p15A. This plasmid was modified for integration and heterologous expression in Pseudomonas putida to enable the production of complex secondary metabolites requiring mm-CoA as precursor. Methylmalonate was identified in the recombinant P. putida strain by an analysis method based on gas chromatography/mass spectrometry. The engineered strain is able to synthesize polyketides requiring mm-CoA as an extender unit, which was demonstrated by the production of myxothiazol after integration of the biosynthetic gene cluster into the chromosome, followed by induction of expression.

Feeding experiments with isotope-labeled precursors rule out hydroxypyruvate and TCA cycle intermediates as the metabolic source of methoxymalonyl-ACP, the substrate for incorporation of "glycolate" units into ansamitocin P-3, soraphen A, and other antibiotics. They point to 1,3-bisphosphoglycerate as the source of the methoxymalonyl moiety and show that its C-1 gives rise to the thioester carbonyl group (and hence C-1 of the "glycolate" unit), and its C-3 becomes the free carboxyl group of methoxymalonyl-ACP, which is lost in the subsequent Claisen condensation on the type I modular polyketide synthases (PKS). d-[1,2-(13)C(2)]Glycerate is also incorporated specifically into the "glycolate" units of soraphen A, but not of ansamitocin P-3, suggesting differences in the ability of the producing organisms to activate glycerate. A biosynthetic pathway from 1,3-bisphosphoglycerate to methoxymalonyl-ACP is proposed. Two new syntheses of R- and S-[1,2-(13)C(2)]glycerol were developed as part of this work.

Two groups of 3-substituted indole 1, 6–9 as well as 4-substituted quinoline derivatives 2–5 and harman (10) were isolated from various species and genera of myxobacteria. They were identified by their spectroscopic data. Some of the metabolites show moderate antifungal activity. Their biosynthetic origin in myxobacteria was demonstrated for 1, 2, and 3 by incorporation of L-[1'-14C]tryptophan.

Type III polyketide synthases (PKS) were regarded as typical for plant secondary metabolism before they were found in microorganisms recently. Due to microbial genome sequencing efforts, more and more type III PKS are found, most of which of unknown function. In this manuscript, we report a comprehensive analysis of the phylogeny of bacterial type III PKS and report the expression of a type III PKS from the myxobacterium Sorangium cellulosum in pseudomonads. There is no precedent of a secondary metabolite that might be biosynthetically correlated to a type III PKS from any myxobacterium. Additionally, an inactivation mutant of the S. cellulosum gene shows no physiological difference compared to the wild-type strain which is why these type III PKS are assumed to be "silent" under the laboratory conditions administered. One type III PKS (SoceCHS1) was expressed in different Pseudomonas sp. after the heterologous expression in Escherichia coli failed. Cultures of recombinant Pseudomonas sp. harbouring SoceCHS1 turned red upon incubation and the diffusible pigment formed was identified as 2,5,7-trihydroxy-1,4-naphthoquinone, the autooxidation product of 1,3,6,8-tetrahydroxynaphthalene. The successful heterologous production of a secondary metabolite using a gene not expressed under administered laboratory conditions provides evidence for the usefulness of our approach to activate such secondary metabolite genes for the production of novel metabolites.

In the attempt to establish a reliable real-time PCR protocol for transcriptional analysis of secondary metabolism in Sorangium cellulosum strain So ce56, a RNA extraction method and a reverse transcription protocol was developed. In order to validate chivosazol or etnangien gene cluster transcripts as good candidates to develop the real-time PCR protocol, stability measurements of the transcripts were performed proving both transcripts to be very stable. The chivosazol biosynthetic gene cluster was taken as the test case to evaluate the special problems arising from the large size of the transcripts and the high G/C-content of the encoding DNA. A set of primer pairs targeting the presumed 90 kbp chivosazol transcript at different positions was employed. The production rate of chivosazol was compared to the transcription of the operon in time course experiments revealing that during the logarithmic growth phase transcription is maximally induced and levels out during the stationary phase. Some deviations in transcript numbers could be measured depending on the primer pair used, but cross-evaluation strengthened the notion that the measured numbers reflect the whole transcript quantities and the in vivo level. Finally, a putative promoter located between chiA and chiB was examined by using the developed real-time PCR protocol.

Media, completely different with respect to their complexity, were investigated for growth and secondary metabolism with Sorangium cellulosum strain So ce56. While technical substrates are best for metabolite production, these media are not suitable to study gene expressions. Free amino acids as present in soluble media based on peptones were found to inhibit polyketide biosynthesis. Therefore synthetic growth and production media were developed. Different nitrogen- and carbon sources as well as trace elements were studied with respect to growth and regulation of chivosazol biosynthesis. A simple defined medium with asparagine as nitrogen source, glucose as carbon source and zinc ions as essential trace element is proposed for future studies into secondary metabolism using transcriptomics and proteomics.

Antidotal doses of hydroxocobalamin are associated with transient increases in blood pressure in some animals and humans. These studies in anesthetized rabbits were undertaken to explore the possible mechanisms underlying the hemodynamic effects of hydroxocobalamin by investigating 1) possible hemodynamic effects of cyanocobalamin, which is formed on a molar-to-molar basis when hydroxocobalamin binds cyanide, and 2) the interference of hydroxocobalamin with the endothelial nitric oxide system.Study 1 investigated the hemodynamic effects of cyanocobalamin. This study included two treatment arms: 1) cyanocobalamin (75 mg/kg, IV) followed by saline (n = 7) and 2) saline followed by cyanocobalamin (n = 7). Study 2 assessed the hemodynamic effects of hydroxocobalamin (75 mg/kg, IV) in the presence and absence of the nitric oxide synthase inhibitor L-Nomega-nitro-L-arginine methyl ester (L-NAME; 30 mg/kg, IV). Nitric oxide synthase inhibition itself increases blood pressure. Thus, as part of Study 2, the hemodynamic effects of hydroxocobalamin were also investigated in the presence of an equipressor dose of angiotensin II (ANGII; 0.05 microg/kg/min, IV) in order to determine whether elevated blood pressure per se could interfere with hydroxocobalamin's hemodynamic effects. This study included six treatment arms (designated as first treatment + second treatment): saline + saline (n = 5), L-NAME + saline (n = 7), saline + hydroxocobalamin (n = 7), L-NAME + hydroxocobalamin (n = 7), ANGII + hydroxocobalamin (n = 7), and ANGII + saline (n = 7).In Study 1, the effects of cyanocobalamin on hemodynamic parameters were indistinguishable from those of saline. In Study 2, hydroxocobalamin infusion was associated with moderate hemodynamic effects, including an increase in systemic vascular resistance, an increase in blood pressure, and a decrease in cardiac output. Administration of L-NAME abolished the effects of hydroxocobalamin on all hemodynamic parameters. ANGII at a dose producing a pressor response comparable to that of L-NAME did not influence the hydroxocobalamin-associated hemodynamic changes.These studies in anesthetized rabbits demonstrate that the moderate pressor effect of hydroxocobalamin is not related to the formation of cyanocobalamin but is very likely related to the scavenging of nitric oxide by hydroxocobalamin.

Two novel highly cytotoxic metabolites, spirangien A (1) and B (2), were isolated from the myxobacterium Sorangium cellulosum (strain So ce90). The structures were elucidated by detailed NMR spectroscopic analysis. The previously unknown molecular framework common to spirangien A and B includes a highly functionalized spiroketal core structure, a side chain bearing a pentaene chromophore and a terminal carboxyl group, and a total of thirteen stereocenters. The absolute configuration at C-3 was determined by degradation and subsequent fragment analysis by GC. The relative stereochemistry of the spiroketal structure was proposed on the basis of vicinal proton couplings and ROESY data for hydroperoxide derivative 4, obtained by ozonolysis of spirangien A, and for 1,3-diene 5, obtained by cross-metathesis with ethylene. X-ray crystal structure analysis of 5 confirmed its structure and unambiguously provided the complete relative stereochemistry of all twelve stereocenters. The 1,3-diene derivative 5 still shows strong cytotoxic activity. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)

SummaryNovel moderately thermophilic Myxobacteria fromboth known suborders ( Cystobacterineae and Sor-angineae ) were isolated from soil samples of semiaridand warm climates. The addition of the anthelminticand amoebizidal agent levamisole was a new and cru-cial improvement for the fast isolation of overall 37strains, which grew very fast at temperatures of 42–44 ∞ C. When the 16S rDNAs were compared with Gen-Bank data of common Myxobacteria, identities were98–99%, thus not reflecting the physiological differ-ences. Similar to the Myxobacteria described so far,the new isolates are multiresistant against a varietyof antibiotics and are producers of typical myxobac-terial secondary metabolites. Analysis of our previousstrain collection isolated from soil samples takenworldwide revealed a more or less uniform distribu-tion of strains which synthesize specific metabolites.Therefore these moderately thermophilic Myxobacte-ria, which grow 2–3 times faster, have the potential toreplace the slow-growing isolates and provide ameans for fast and cost-saving production of myxo-bacterial metabolites in the future.Introduction

Three novel metabolites, tuscolid (1) and tuscoron A (2) and B (3), were isolated from the myxobacterium Sorangium cellulosum (strains So ce1401 and So ce1383). The structures were elucidated by detailed NMR spectroscopic analysis, and their biosynthesis was studied by feeding 13C-labelled precursors. The results revealed that the macrolide 1 and the acyclic derivative 2 are closely related polyketides containing, as a characteristic structural feature, a furan-3(2H)-one ring system. The related minor component tuscoron B (3) was identified as an unstable structural analogue of tuscoron A. The relative stereochemistry of the tetrahydropyran ring and of C-13 to C-17 of tuscolid (1), and of the dihydropyran ring of tuscoron A (2), was determined on the basis of 1H-1H-coupling constants and NOE correlations. A tentative mechanism for the conversion of 1 into 2 is discussed. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)

Myxobacteria increasingly gain attention as a source of bioactive natural products. The genus Sorangium produces almost half of the secondary metabolites isolated from these microorganisms. Nevertheless, genetic systems for Sorangium strains are poorly developed, which makes the identification of the genes directing natural product biosynthesis difficult. Using biparental and triparental mating, we have developed methodologies for DNA transfer from Escherichia coli via conjugation for the genome sequencing model strain So ce56 and the secondary metabolite multiproducing strain So ce12. The conjugation protocol developed for strain So ce56 is not applicable to other Sorangium strains. Crucial points for the conjugation are the ratio of E. coli and Sorangium cellulosum cells, the choice of liquid or solid medium, the time used for the conjugation process and antibiotic selection in liquid medium prior to the plating of cells. A mariner-based transposon containing a hygromycin resistance gene was generated and used as the selectable marker for S. cellulosum. The transposon randomly integrates into the chromosome of both strains. As a proof of principle, S. cellulosum So ce12 transposon mutants were screened using an overlay assay to target the chivosazole biosynthetic gene cluster.

In addition to previously isolated ratjadone A we describe three new members of this family, ratjadones B, C, and D, from another strain of the myxobacterium Sorangium cellulosum. We have investigated the properties of these ratjadones with respect to their activity on mammalian cell lines. We found IC(50) values in the picomolar range and a significant increase in the size of nuclei. A further examination showed that they inhibit the export of the leucine-rich nuclear export signal (LR-NES) containing proteins in different cell lines. Ratjadones are able to inhibit the formation of the nuclear export complex composed of the CRM1, RanGTP, and the cargo protein, as shown by two different in vitro assays. Finally, the binding of ratjadone C to CRM1 was demonstrated. These ratjadone activities are in the same concentration range as described for the polyketide leptomycin B (LMB) from Streptomyces sp. Like LMB, it seems that the ratjadones covalently bind to CRM1, inhibit cargo protein binding via LR-NES, and thereby block nuclear export. Thus, the ratjadones represent a new class of natural compounds which inhibit proliferation in eukaryotes by blocking nuclear export.

A bioassay-guided fractionation of the crude methanol extract of the myxobacterium Hyalangium minutum, strain NOCB-2(T) (DSM 14724(T)), led to the isolation of hyaladione (1), a novel S-methyl cyclohexadiene-dione. The structure of 1 was established by HRESIMS, NMR, and IR spectroscopy as well as X-ray crystallography. Compound 1 was active against growing mammalian cell lines, with IC(50) values ranging from 1.23 to 3.93 μM, in addition to a broad spectrum of antibacterial and antifungal activities, including inhibition of pathogenic methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa with an MIC of 0.83 and 8.5 μg mL(-1), respectively.

In this study, Sorangium cellulosum So ce56 was phenotypically and genotypically analysed in order to evaluate whether this strain can be used in a comprehensive genome project as a representative of the secondary metabolite-producing myxobacteria. In contrast to many other strains of S. cellulosum, strain So ce56 was found to have various advantageous features, including fast and homogeneous growth in submerged cultures and the ability to complete its morphological differentiation cycle on agar, even when the inoculant originates from a liquid culture. Two groups of secondary metabolites isolated from the culture broth were identified, the polyketides etnangien and chivosazole. The presence of polyketide synthase-encoding genes in the genome of strain So ce56 was demonstrated via PCR. The phenotypic classification was confirmed by comparison of 16S rDNA sequences which showed that S. cellulosum So ce56 clusters within a separate lineage together with S. cellulosum ATCC 25531 and the epothilone producer S. cellulosum So ce90. The genome of S. cellulosum So ce56 belongs to the largest bacterial genomes described so far. It is estimated to be 12.2 Mb in size, by pulsed-field gel electrophoresis. In order to demonstrate that S. cellulosum So ce56 is a convenient strain for molecular biological studies, a genetic manipulation system was developed. Using triparental mating, polyketide synthase-encoding genes were inactivated, leading to chivosazole-negative mutants.

Myxobacteria are Gram-negative soil-dwelling bacteria belonging to the phylum Proteobacteria . They are a rich source of promising compounds for clinical application, such as epothilones for cancer therapy and several new antibiotics. In the course of a bioactivity screening program of secondary metabolites produced by Sorangium cellulosum strains, the macrolide chlorotonil A was found to exhibit promising antimalarial activity. Subsequently, we evaluated chlorotonil A against Plasmodium falciparum laboratory strains and clinical isolates from Gabon. Chlorotonil A was highly active, with a 50% inhibitory concentration between 4 and 32 nM; additionally, no correlations between the activities of chlorotonil A and artesunate (rho, 0.208) or chloroquine (rho, −0.046) were observed. Per os treatment of Plasmodium berghei -infected mice with four doses of as little as 36 mg of chlorotonil A per kg of body weight led to the suppression of parasitemia with no obvious signs of toxicity. Chlorotonil A acts against all stages of intraerythrocytic parasite development, including ring-stage parasites and stage IV to V gametocytes, and it requires only a very short exposure to the parasite to exert its antimalarial action. Conclusively, chlorotonil A has an exceptional and unprecedented profile of action and represents an urgently required novel antimalarial chemical scaffold. Therefore, we propose it as a lead structure for further development as an antimalarial chemotherapeutic.

Seven new polyketides, for which the trivial names hyafurones A1-B (1-3), hyapyrrolines A (4) and B (5), and hyapyrones A (6) and B (7) are proposed, were isolated from the fermentation broth of the myxobacteria Hyalangium minutum, strains NOCB-2(T) and Hym-3. Their structures were elucidated from NMR and HRESIMS data, and their geometric configuration was assigned based on NOE and vicinal (1)H coupling data. Both hyafurone B (3) and hyapyrone B (7) inhibited growth of the Gram-positive bacterium Nocardia flava, while 7 showed antifungal activity against Mucor hiemalis.

In 1937 Imshenetski and Solntseva isolated a new species of cellulose-degrading myxobacteria, which they called Sorangium cellulosum. 16S rRNA gene sequencing of nine isolates of Sorangium and their comparison with Polyangium cellulosum as the reference strain proved a close phylogenetic relationship (evolutionary distance, less than 3% on the nucleotide level). The use of single antibiotics or better combinations of several antibiotics that act on different targets may be helpful, because Sorangium species usually turn out to be multiresistant. Cells of the suborder “Cystobacterineae” on the one hand and “Sorangiineae” and “Nannocystineae” on the other hand can easily be distinguished because they differ in cell morphology, as can be detected using phase-contrast microscopy. Strains of Byssophaga cruenta, characterized by their intense blood-red color, and the very common S. cellulosum strains are the only myxobacteria which degrade crystalline cellulose and can use it as the sole carbon source. At the Gesellschaft fur Biotechnologische Forschung (GBF) an isolation and screening program was initiated in the late 1970s to evaluate the potential of the different genera of myxobacteria as producers of secondary metabolites. A ddc gene product, which was previously found only in eukaryotes, has also been identified in Sorangium strains. As discussed in this chapter, the fascinating microorganisms of the genus Sorangium attract more and more attention, because they undergo a complex life cycle, possess the largest bacterial genomes known to date, and show a high potential as producers of biotechnologically important natural products.

An antifungal activity against Mucor hiemalis was detected in the culture broth of Sorangium cellulosum (Myxococcales) strain So ce90. The activity was excreted into the supernatant during the log and early stationary phase. When the adsorber resin XAD-16 was added to the culture, the active metabolites were quantitatively bound to the resin. The epothilons showed a high cytotoxicity for animal cells and mimic the biological effects of taxol (BOLLAG et al., Cancer Res. 55: 2325 approximately 2333, 1995).

The antifungal metabolite ratjadone (1) was isolated from the myxobacterium Sorangium cellulosum, strain So ce360. Production was achieved by fermentation in the presence of the adsorber resin XAD-16, extraction and separation by RP chromatography. The structure of 1 was determined by spectroscopic methods. It is characterized by a 4-hydroxytetrahydropyran and a 5,6-dihydropyran-2-one ring which are connected by an unsaturated C11 carbon chain. The polyketide origin of 1 was proven by feeding experiments with 13C-labeled precursors and NMR spectroscopic examinations.

A new antibiotic, ripostatin, was isolated from the culture supernatant of the myxobacterium, Sorangium cellulosum strain So ce377. It is a macrocyclic lactone carbonic acid containing an unsubstituted phenyl ring in a side chain. The antibiotic acts especially on Staphylococcus aureus, but seems not to penetrate most bacteria. The MIC values are in the range of 1 μg/ml. Ripostatin is an inhibitor of eubacterial RNA polymerase. It interferes with the initiation of RNA synthesis.

A new antibiotic, disorazol, was isolated from the culture broth of the myxobacterium, Sorangium cellulosum strain So ce 12. It is a macrocyclic compound containing two oxazole rings. The antibiotic acted against many fungi and mammalian cell cultures. The latter responded to extremely low doses (MIC 3-30 pg/ml). None of the tested bacteria and yeasts were inhibited.

New antibiotics were isolated from the culture broth of the myxobacterium, Sorangium cellulosum, strain So ce 678. The antibiotics were active against Gram-positive bacteria and mammalian cells. They were named tartrolon A and B. Tartrolon B contains a boron atom. The boron binding region of tartrolon is identical with that of boromycin and aplasmomycin.

An antifungal activity, ratjadon, was detected in the culture broth of Sorangium cellulosum (Myxococcales) strain So ce360. The metabolite was quantitatively bound to the adsorber resin XAD-16, which was added to the medium at the beginning of the fermentation. The antibiotic spectrum was narrow, but some important phytopathogenic fungi, especially species of Oomycetes, were inhibited at very low concentrations.

The gliding bacterium Sandaracinus amylolyticus, strain NOSO-4T, was recently characterized as the first representative of a new myxobacterial genus. A screening of the culture broth for antibiotically active metabolites followed by isolation and characterization revealed two unique 3-formylindol derivatives, indiacen A (1) and its chloro derivative indiacen B (2). Both are active against Gram-positive and Gram-negative bacteria as well as the fungus Mucor hiemalis. The biosynthetic origin of the isoprene-like side chain in 1 and 2 was studied by in vivo feeding experiments with 13C-labeled precursors.

Theories and methods developed in molecular pharmacology for drug receptor interactions were used to explain artificially induced myxospore formation. The correlation between inducer concentrations and the yield of myxospores, i.e. the experimentally obtained dose-response curves, were much steeper than expected for an interaction based on mass action law. We postulate that the interaction of an inducer with one of the different receptors of the bacterial cell causes the production of a common stimulus which programmes the cells to turn into myxospores, if a threshold value is reached. Simultaneous addition of inducers of the same or of different inducer groups showed synergistic interactions. Inducer concentrations that by themselves were not inducing gave maximum myxospore formation when combined. Addition of pyrrole, a specific inhibitor, caused a concentration-dependent parallel shift of the glycerol dose-response curve. Compounds, inducers or inhibitors, with identical receptor specificities competed for the common receptor according to their affinity. Interactions of inducers with different receptor specificities could be predicted from calculations with a mathematical model of functional synergism.

An antifungal activity was detected in the culture broth of Sorangium cellulosum (Myxococcales), strain So ce26. The activity was excreted into the supernatant during the log and early stationary phase. The active substance was quantitatively bound to XAD absorber resin added to the medium at the beginning of the fermentation. The new secondary metabolite was called soraphen and is of special interest to plant disease control for its inhibitory activity against numerous phytopathogenic fungi.