RESUMO
Patulin is a mycotoxin that contaminates pome fruits and derived products worldwide. Basidiomycete yeasts belonging to the subphylum Pucciniomycotina have been identified to have the ability to degrade this molecule efficiently and have been explored through different approaches to understand this degradation process. In this study, Sporobolomyces sp. strain IAM 13481 was found to be able to degrade patulin to form two different breakdown products, desoxypatulinic acid and (Z)-ascladiol. To gain insight into the genetic basis of tolerance and degradation of patulin, more than 3,000 transfer DNA (T-DNA) insertional mutants were generated in strain IAM 13481 and screened for the inability to degrade patulin using a bioassay based on the sensitivity of Escherichia coli to patulin. Thirteen mutants showing reduced growth in the presence of patulin were isolated and further characterized. Genes disrupted in patulin-sensitive mutants included homologs of Saccharomyces cerevisiae YCK2, PAC2, DAL5, and VPS8. The patulin-sensitive mutants also exhibited hypersensitivity to reactive oxygen species as well as genotoxic and cell wall-destabilizing agents, suggesting that the inactivated genes are essential for tolerating and overcoming the initial toxicity of patulin. These results support a model whereby patulin degradation occurs through a multistep process that includes an initial tolerance to patulin that utilizes processes common to other external stresses, followed by two separate pathways for degradation.
Assuntos
Basidiomycota/genética , Proteínas Fúngicas/genética , Patulina/metabolismo , Saccharomyces cerevisiae/genética , Acetatos/metabolismo , Sequência de Bases , Basidiomycota/efeitos dos fármacos , Basidiomycota/crescimento & desenvolvimento , Basidiomycota/fisiologia , Caseína Quinase I/genética , Caseína Quinase I/metabolismo , Cromatografia Líquida de Alta Pressão , Escherichia coli/efeitos dos fármacos , Escherichia coli/crescimento & desenvolvimento , Proteínas Fúngicas/metabolismo , Furanos/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Dados de Sequência Molecular , Mutagênese Insercional , Estresse Oxidativo , Patulina/isolamento & purificação , Patulina/farmacologia , Pironas/metabolismo , Espécies Reativas de Oxigênio/farmacologia , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/crescimento & desenvolvimento , Saccharomyces cerevisiae/fisiologia , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Alinhamento de Sequência , Estresse FisiológicoRESUMO
During September 2004, downy mildew of parsley caused by a species of Plasmopara was observed in an experimental field of parsley (Petroselinum crispum subsp. crispum L. cv. Gigante d'Italia/Hilmar) in Borgeby in southern Sweden. The summer of 2004 was exceptionally wet and humid. Disease became widespread throughout the field in just a few days. Local growers reported that symptoms consistent with downy mildew had appeared in their parsley fields every year since 2001. Plasmopara, under P. nivea, has been reported on parsley in Europe since the middle of the 19th century (4). In recent years, this disease has caused severe damage to parsley grown in several European countries, e.g., France, Germany, Switzerland, and Belgium (1,3). The first symptoms appeared as faint chlorotic spots on the upper surfaces of the leaves. On the corresponding lower surfaces, mycelium and sporangiophores grew profusely and developed a white mat that in part turned dark gray. Eventually, the leaves and stalks became necrotic and died. The sporangiophores were monopodially branched, 248.4 ± 13.36 µm long (n = 17), each branch ending in 2 to 5 ultimate branchlets tapered toward the tip. The trunk diameter measured 7.0 ± 0.77 µm (n = 9) above the basal part and 6.1 ± 0.81 µm just below the first branch. The sporangia were broadly ellipsoidal to ellipsoidal, hyaline, 22.5 ± 0.73 µm long and 16.6 ± 0.48 µm wide (n µ 40). They were mostly nonpapillate when young, although exit pores 4.8 ± 0.32 µm (n = 10) were visible. Mature sporangia exhibited a dehiscence apparatus and a plug in the exit pore. On the basis of the characteristics above, the pathogen was identified as P. petroselini (= P. nivea pro parte [2]). Independent verification of the identity was done by O. Constantinescu at the Botanical Museum, Uppsala, Sweden. A voucher specimen was deposited at the Herbarium UPS, in Uppsala under the number UPS F-118873. To our knowledge, this is the first report of P. petroselini on parsley in Sweden. References: (1) E. Bèliard and J. Thibault. Phytoma 554:2, 2002. (2) O. Constantinescu. Taxon 54:813, 2005. (3) C. Crepel and S. Inghelbrecht. Plant Dis. 87:1266, 2003. (4) A. de Bary, Ann. Sci. Nat. Bot., Sér. 4, 20:5, 1863.