While abscisic acid (ABA) is known as a hormone produced by plants through the carotenoid pathway, a small number of phytopathogenic fungi are also able to produce this sesquiterpene but they use a distinct pathway that starts with the cyclization of farnesyl diphosphate (FPP) into 2Z,4E-α-ionylideneethane which is then subjected to several oxidation steps. To identify the sesquiterpene cyclase (STC) responsible for the biosynthesis of ABA in fungi, we conducted a genomic approach in Botrytis cinerea. The genome of the ABA-overproducing strain ATCC58025 was fully sequenced and five STC-coding genes were identified. Among them, Bcstc5 exhibits an expression profile concomitant with ABA production. Gene inactivation, complementation and chemical analysis demonstrated that BcStc5/BcAba5 is the key enzyme responsible for the key step of ABA biosynthesis in fungi. Unlike what is observed for most of the fungal secondary metabolism genes, the key enzyme-coding gene Bcstc5/Bcaba5 is not clustered with the other biosynthetic genes, i.e., Bcaba1 to Bcaba4 that are responsible for the oxidative transformation of 2Z,4E-α-ionylideneethane. Finally, our study revealed that the presence of the Bcaba genes among Botrytis species is rare and that the majority of them do not possess the ability to produce ABA.

The sequencing of the genomes of the B05.10 and T4 strains of the fungus Botrytis cinerea revealed an abundance of novel biosynthetic gene clusters, the majority of which were unexpected on the basis of the previous analyses of the fermentation of these and closely related species. By systematic alteration of easy accessible cultivation parameters, using chemical induction with copper sulfate, we have found a cryptic sesquiterpenoid family with new structures related to eremophil-9-ene, which had the basic structure of the sesquiterpene (+)-5-epiaristolochene ((+)-4-epieremophil-9-ene). An expression study of the sesquiterpene cyclase genes present in the Botrytis cinerea genome, under culture conditions, is reported. In general, a 3 day delay and a higher BcSTC genes expression were observed when copper (5 ppm) was fed to the fermentation broth. In addition, to the observed effect on the BcBOT2 (BcSTC1) gene, involved in the biosynthesis of the botrydial toxin, a higher expression level for BcSTC3 and BcSTC4 was observed with respect to the control in the strain B05.10. Interestingly, under copper conditions, the BcSTC4 gene was the most expressed gene in the Botrytis cinerea UCA992 strain. In vitro evaluation of the biological role of these metabolites indicates that they contributed to the conidial development in B. cinerea and appear to be involved in self-regulation of the production of asexual spores. Furthermore, they promoted the formation of complex appressoria or infection cushions.

Proteomics has become one of the most relevant high-throughput technologies. Several approaches have been used for studying, for example, tumor development, biomarker discovery, or microbiology. In this “post-genomic” era, the relevance of these studies has been highlighted as the phenotypes determined by the proteins and not by the genotypes encoding them that is responsible for the final phenotypes. One of the most interesting outcomes of these technologies is the design of new drugs, due to the discovery of new disease factors that may be candidates for new therapeutic targets. To our knowledge, no commercial fungicides have been developed from targeted molecular research, this review will shed some light on future prospects. We will summarize previous research efforts and discuss future innovations, focused on the fight against one of the main agents causing a devastating crops disease, fungal phytopathogens.

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Cultivation of the phytopathogenic fungus Botrytis cinerea using sublethal amounts of copper sulfate yielded a cryptic sesquiterpenoids family, which displayed the basic chemical structure of (+)-4-epi-eremophil-9-ene. The biosynthesis pathway was established, and the route involved the likely transformation of the diphosphate of farnesyl (FDP), to give a cis-fused eudesmane cation, through (S)-hedycaryol, finally yielding the (+)-4-epi-eremophil-9-enol derivatives. An expression study of genes that code for the sesquiterpene cyclases (STC), including the recently reported gene Bcstc7 present in the B. cinerea genome, was performed in order to establish the STC involved in this biosynthesis. The results showed a higher expression level for the Bcstc7 gene with respect to the other stc1–5 genes in both wild-type strains, B05.10 and Botrytis cinerea UCA992. Deletion of the Bcstc7 gene eliminated (+)-4-epi-eremophilenol biosynthesis, which could be re-established by complementing the null mutant with the Bcstc7 gene. Chemical analysis suggested that STC7 is the principal enzyme responsible for the key step of cyclization of FDP to eremophil-9-en-11-ols. Furthermore, a thorough study of the two wild-types and the complemented mutant revealed four new eremophilenol derivatives whose structures are reported here.

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Botrytis cinerea is one of the most relevant plant pathogenic fungi. The first step during its infection process is the germination of the conidia. Here, we report on the first proteome analysis during the germination of B. cinerea conidia, where 204 spots showed significant differences in their accumulation between ungerminated and germinated conidia by two-dimensional polyacrylamide gel electrophoresis and qPCR. The identified proteins were grouped by gene ontology revealing that the infective tools are mainly preformed inside the ungerminated conidia allowing a quick fungal development at the early stages of conidial germination. From 118 identified spots, several virulence factors have been identified while proteins, such as mannitol-1-phosphate dehydrogenase, 6,7-dimethyl-8-ribityllumazine synthase or uracil phosphoribosyltransferase, have been disclosed as a new potential virulence factors in botrytis whose role in pathogenicity needs to be studied to gain new insights about the role of these proteins as therapeutic targets and virulence factors.

& Introduction The limited ability of Quercus species to regenerate naturally in Mediterranean forests has led to the development of various artificial regeneration methods; however, there is no general consensus as to what specific method is the best one for this purpose. & Material and methods In this work, we assessed morphology, growth and survival of two Quercus species (Quercus ilex ssp. ballota and Quercus suber) using two different methods of artificial regeneration (viz. direct seeding and planting) and two seedling ages (1-year-old seedlings and 3-year-old seedlings) in southern Spain. & Results and discussion The 1-year-old seedlings of both species were found to exhibit the highest survival percentages and direct-seeded plants intermediate survival values. For direct-seeded plants, seed mass was found to have a significantly positive effect on the establishment success in both species. No clear-cut trend in survival was detected in the 3-year-old seedlings. The survival of the 3-year-old Q. suber seedlings and the direct-seeded plants was similar, but not in Q. ilex, where the survival of the 3-year-old seedlings was the lowest. The latter result may have been a consequence of cultivation in smaller containers leading to root deformation and limiting plant access to water. Differences in survival could not be ascribed to morphological and growth variables or stomatal conductance. & Conclusion Based on the results, all three artificial regeneration methods can be similarly effective provided appropriate nursery cultivation conditions are used and seeds are protected against predators, the best choice in each case being dictated by the particular restoration goals.