In filamentous fungi, Sfp-type 4'-phosphopantetheinyl transferases (PPTases) activate enzymes involved in primary (a-aminoadipate reductase [AAR]) and secondary (polyketide synthases and nonribosomal peptide synthetases) metabolism. We cloned the PPTase gene PPT1 of the maize anthracnose fungus Colletotrichum graminicola and generated PPTasedeficient mutants (Dppt1). Dppt1 strains were auxotrophic for Lys, unable to synthesize siderophores, hypersensitive to reactive oxygen species, and unable to synthesize polyketides (PKs). A differential analysis of secondary metabolites produced by wild-type and Dppt1 strains led to the identification of six novel PKs. Infection-related morphogenesis was affected in Dppt1 strains. Rarely formed appressoria of Dppt1 strains were nonmelanized and ruptured on intact plant. The hyphae of Dppt1 strains colonized wounded maize (Zea mays) leaves but failed to generate necrotic anthracnose disease symptoms and were defective in asexual sporulation. To analyze the pleiotropic pathogenicity phenotype, we generated AAR-deficient mutants (Daar1) and employed a melanin-deficient mutant (M1.502). Results indicated that PPT1 activates enzymes required at defined stages of infection. Melanization is required for cell wall rigidity and appressorium function, and Lys supplied by the AAR1 pathway is essential for necrotrophic development. As PPTase-deficient mutants of Magnaporthe oryzea were also nonpathogenic, we conclude that PPTases represent a novel fungal pathogenicity factor.

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SUMMARY Iron is an essential element for the growth of nearly all organisms. In order to overcome the problem of its low bioavailability, microorganisms (including fungi) secrete siderophores, high-affinity iron chelators. As the acquisition of iron is also a key step in infection processes, siderophores have been considered as potential virulence factors in several host-pathogen interactions. Most fungi produce siderophores of the hydroxamate-type, which are synthesized by non-ribosomal peptide synthetases (NRPSs). Magnaporthe grisea, the causal agent of rice blast disease, produces ferricrocin as intracellular storage siderophore and excretes coprogens. In the M. grisea genome we identified SSM1, an NRPS gene, and a gene encoding an l-ornithine N5-monooxygenase (OMO1) that is clustered with SSM1 and responsible for catalysing the first step in siderophore biosynthesis, the N(5) hydroxylation of ornithine. Disruption of SSM1 confirmed that the gene encodes ferricrocin synthetase. Pathogenicity of these mutants towards rice was reduced, suggesting a role of this siderophore in pathogenicity of M. grisea.

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In this work, a systematic approach to plant-wide control design is proposed. The method combines ingredients from process networks, thermodynamics and systems theory to derive robust decentralized controllers that will ensure complete plant stability. As a first step, the considered process system is decomposed into abstract mass and energy inventory networks. In this framework, conceptual inventory control loops are then designed for the mass and energy layers to guarantee that the states of the plant, both in terms of extensive and intensive properties, will converge to a compact convex region defined by constant inventories. This result by itself does not ensure the convergence of intensive variables to a desired operation point as complex dynamic phenomena such as multiplicities may appear in the invariant set. In order to avoid these phenomena, thermodynamics naturally provides the designer, in these convex regions, with a legitimate storage or Lyapunov function candidate, the entropy, that can be employed to ensure global stability. Based on this, the control structure design procedure is completed with the realization of the conceptual inventory and intensive variable control loops over the available degrees of freedom in the system. To that purpose, both PI and feedback linearization control are employed. The different aspects of the proposed methodology will be illustrated on a non-isothermal chemical reaction network.

Abstract. Microsomal preparations of six species of the plant family Fabaceae were screened for high-affinity binding of branched (1 ~ 3), (1 --, 6)-[3-glucans. Oligoglucosides of this type are specific elicitors of phytoalexin accumulation in soybean (Glycine max L.), a member of this family. The species studied were alfalfa (Medicago sativa L.), broadbean (Vicia faba L.), chickpea (Cicer arietinum L.), french bean (Phaseolus vulgaris L.), pea (Pisum sativum L.), and white lupin (Lupinus albus L.). A 125I-labeled 4-(2-aminophenyl)ethylamine conjugate of a (1 ~ 3), (1 ~ 6)-[3-glucan fraction with an average degree of polymerization (DP) of 18, obtained from mycelial walls of Phytophthora sojae, was used as radioligand for initial screening. The structural complexity of this fraction allowed the identification of binding sites with affinities for isomeric structures other than the (1 ~ 3), (1--* 6)-hepta-[3-glucoside for which soybean binding sites display highest affinity. Radioligand competition experiments against unlabeled fungal [3-glucan resulted in the identification of high-affinity binding in alfalfa, bean, lupin, and pea. Half-maximal competition concentrations (ICso) for fungal [3-glucan in these species were between 5 and 30 nM. Pseudoheterologous radioligand competition by unlabeled hepta-[3-glucoside showed that for alfalfa, lupin and pea the ICs0 values for this structure (4 to 16 nM) * Present address: Pontificia Universidad Catolica del Peru, Lima 100, Peru ** Present address: Botariisches Institut der Universitfit, D-80638 Miinchen, Germany Abbreviations: DP = degree of polymerization; ECso = concentration of elicitor necessary to obtain a half-maximal biological response; HG = synthetic (1 ~ 3), (1 ~ 6)-hepta-~-glucoside phytoalexin elicitor; HG-APEA = 1-[4-(2-aminophenyl)ethylamino-1-hexaglucosyl]deoxyglucitol; ICso = ligand concentration necessary to obtain half-maximal displacement of radioligand in competition binding assays; Ka = dissociation constant; OS = branched (1 ~ 3), (1-,6)-~-glucan obtained by hydrolysis of mycelial walls of Phytophthora sojae; OS-APEA = 1-[4-(2-aminophenyl)ethylamino-1-oligoglucosyl]deoxyglucitol conjugate of OS Correspondence to: J. Ebel; FAX: 49(89)1782274; E-mail: j.ebel@botanik.biologie.uni-muenchen.de were similar to those of soybean (7.7 nM). Bean microsomes, however, displayed an IC5o significantly higher than soybean (68 nM) suggesting that the structural motif recognized by its binding sites is not identical to that of soybean or the other three species. Radioligand saturation assays with alfalfa, lupin and pea microsomes using an 125i_labele d aminophenylethylamine hepta-[~-glucoside conjugate gave dissociation constants (Ka) of 5.3, 3.7, and 1.8 nM, respectively. The affinity of these sites for hepta-13-glucoside was in the same range as that of soybean (Ka 1-3 nM), whereas the affinity of the binding sites of bean for the same ligand was significantly lower (Ka = 33 nM). Good correlation was found between the presence of high-affinity binding and the a...

Freeze-drying is considered to be an attractive dehydration method of preserving the quality of high value foods products. Unfortunately, it is an expensive operation, which calls for efficient tools capable of minimizing time and/or energy while preserving product quality. In this work, timescale analysis has been applied to a detailed firstprinciple based model. From such analysis a simplified model, capable of describing freeze-drying at the time scales relevant to quality, has been proposed. The model has been solved by the Finite Element method, showing good agreement with the results in literature. Likewise, the efforts associated with the computation of optimal operation policies have been reduced. In this regard, different operational scenarios that take into account shelf temperature (T L) and chamber pressure (P c) have been considered on a simulation basis. In all cases the resulting optimal control profiles obtained led to significant reductions of cycle time while ensuring product quality.

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