Antifungal Activity of Alpha-Mangostin against Colletotrichum gloeosporioides In Vitro and In Vivo.

We investigated alpha-mangostin (α-mangostin, α-MG), a xanthone natural product extracted from the pericarp of mangosteen (Garcinia mangostana), for its antifungal activities and possible mechanism against Colletotrichum gloeosporioides, which causes mango anthracnose. The results demonstrated that α-MG had a relatively high in vitro inhibitory activity against C. gloeosporioides among 20 plant pathogenic fungi. The median effective concentration (EC50) values of α-MG against mycelial growth were nearly 10 times higher than those of spore germination inhibition for both strains of C. gloeosporioides, the carbendazim-sensitive (CBD-s) and carbendazim-resistant (CBD-r). The results suggested that α-MG exhibited a better inhibitory effect on spore germination than on the mycelial growth of C. gloeosporioides. Further investigation indicated that the protective effect could be superior to the therapeutic effect for mango leaves for scab development. The morphological observations of mycelium showed that α-MG caused the accumulation of dense bodies. Ultrastructural observation further revealed that α-MG caused a decrease in the quantity and shape of the swelling of mitochondria in the mycelium cells of C. gloeosporioides. In addition, bioassays disclosed that the inhibitory activity of α-MG on spore germination was reduced by adding exogenous adenosine triphosphate (ATP). These results suggested that the mode of action of α-MG could be involved in the destruction of mitochondrial energy metabolism. The current study supports α-MG as a natural antifungal agent in crop protection.

Protein extract from Cereus jamacaru (DC.) inhibits Colletotrichum gloeosporioides growth by stimulating ROS generation and promoting severe cell membrane damage.

Mandacaru (Cereus jamacaru DC.), is a cactaceous symbol of caatinga vegetation at Brazilian Northeast region, however, there are no much studies about biochemical properties of this species. Here, the pioneering study brings very relevant data to highlight the importance of research with endemic plants of the caatinga. Afterward, the presence of enzymes such as peroxidase, protease, chitinase, ß-1,3-glucanase, and serine (trypsin) and cysteine (papain) protease inhibitors were evaluated. The peroxidase activity was higher in roots than other tissues. The ß-1,3-glucanase and proteolytic activity were prominent in stem and roots. The chitinase activity and protease inhibitor for both classes analyzed were detected in the stem and fruit peel. Antifungal activity against Colletotrichum gloeosporioides showed the root extract has a promising inhibitory activity on this economical important phytopathogenic fungus. After the contact of the hyphae with root extract increase in membrane permeability, based on Propidium Iodide (PI) uptake, and production of reactive oxygen species (ROS) were detected, compared to negative control. In addition, Scanning Electron Microscopy (SEM) analysis showed morphological damage on hyphae structure indicating that the treatment debilitates either cell membrane or cell wall leading to the cell death C. gloeosporioides.

The Glycosylphosphatidylinositol Anchor Biosynthesis Genes GPI12, GAA1, and GPI8 Are Essential for Cell-Wall Integrity and Pathogenicity of the Maize Anthracnose Fungus Colletotrichum graminicola.

Glycosylphosphatidylinositol (GPI) anchoring of proteins is one of the most common posttranslational modifications of proteins in eukaryotic cells and is important for associating proteins with the cell surface. In fungi, GPI-anchored proteins play essential roles in cross-linking of ß-glucan cell-wall polymers and cell-wall rigidity. GPI-anchor synthesis is successively performed at the cytoplasmic and the luminal face of the ER membrane and involves approximately 25 proteins. While mutagenesis of auxiliary genes of this pathway suggested roles of GPI-anchored proteins in hyphal growth and virulence, essential genes of this pathway have not been characterized. Taking advantage of RNA interference (RNAi) we analyzed the function of the three essential genes GPI12, GAA1 and GPI8, encoding a cytoplasmic N-acetylglucosaminylphosphatidylinositol deacetylase, a metallo-peptide-synthetase and a cystein protease, the latter two representing catalytic components of the GPI transamidase complex. RNAi strains showed drastic cell-wall defects, resulting in exploding infection cells on the plant surface and severe distortion of in planta-differentiated infection hyphae, including formation of intrahyphal hyphae. Reduction of transcript abundance of the genes analyzed resulted in nonpathogenicity. We show here for the first time that the GPI synthesis genes GPI12, GAA1, and GPI8 are indispensable for vegetative development and pathogenicity of the causal agent of maize anthracnose, Colletotrichum graminicola.

Cell death of Nicotiana benthamiana is induced by secreted protein NIS1 of Colletotrichum orbiculare and is suppressed by a homologue of CgDN3.

Colletotrichum orbiculare, the causal agent of cucumber anthracnose, infects Nicotiana benthamiana. Functional screening of C. orbiculare cDNAs in a virus vector-based plant expression system identified a novel secreted protein gene, NIS1, whose product induces cell death in N. benthamiana. Putative homologues of NIS1 are present in selected members of fungi belonging to class Sordariomycetes, Dothideomycetes, or Orbiliomycetes. Green fluorescent protein-based expression studies suggested that NIS1 is preferentially expressed in biotrophic invasive hyphae. NIS1 lacking signal peptide did not induce NIS1-triggered cell death (NCD), suggesting apoplastic recognition of NIS1. NCD was prevented by virus-induced gene silencing of SGT1 and HSP90, indicating the dependency of NCD on SGT1 and HSP90. Deletion of NIS1 had little effect on the virulence of C. orbiculare against N. benthamiana, suggesting possible suppression of NCD by C. orbiculare at the postinvasive stage. The CgDN3 gene of C. gloeosporioides was previously identified as a secreted protein gene involved in suppression of hypersensitive-like response in Stylosanthes guianensis. Notably, we found that NCD was suppressed by the expression of a CgDN3 homologue of C. orbiculare. Our findings indicate that C. orbiculare expresses NIS1 at the postinvasive stage and suggest that NCD could be repressed via other effectors, including the CgDN3 homologue.

Systematic analysis of the falcate-spored graminicolous Colletotrichum and a description of six new species from warm-season grasses.

Species limits in the fungal genus Colletotrichum are traditionally distinguished by appressorial and/or conidial morphology or through host plant association, but both criteria are criticized for their inability to resolve distinct taxa. In previous research eight novel falcate-spored Colletotrichum species were identified from graminicolous hosts using multilocus molecular phylogenetic analysis. In the present work formal descriptions and illustrations are provided for six of the new taxa: C. hanaui sp. nov., C. nicholsonii sp. nov., C. paspali sp. nov., C. jacksonii sp. nov., C. miscanthi sp. nov. and C. axonopodi sp. nov.; and an emended description with epitypification is provided for C. eleusines. Comparison of hyphopodial appressoria and host association against phylogenetic species boundaries and evolutionary relationships in the graminicolous Colletotrichum group demonstrate that, while these characters can be useful in combination for the purpose of species diagnosis, erroneous identification is possible and species boundaries might be underestimated if these characters are used independently, as exemplified by the polyphyletic taxa C. falcatum. Appressoria have been subject to convergent evolution and were not predictive of phylogenetic relationships. Despite these limitations, the results of this work establish that in combination appressorial and host range characters could be used to generate informative dichotomous identification keys for Colletotrichum species groups when an underlying framework of evolutionary relationships, taxonomic criteria and nomenclature have been satisfactorily derived from molecular systematic treatments.

Colutellin A, an immunosuppressive peptide from Colletotrichum dematium.

Colletotrichum dematium is an endophytic fungus recovered from a Pteromischum sp. growing in a tropical forest in Costa Rica. This fungus makes a novel peptide antimycotic, colutellin A, with a MIC of 3.6 microg ml(-1) (48 h) against Botrytis cinerea and Sclerotinia sclerotiorum. Collutellin A has a mass of 1127.7 Da and contains residues of Ile, Val, Ser, N-methyl-Val and beta-aminoisobutryic acid in nominal molar ratios of 3 : 2 : 1 : 1 : 1, respectively. Independent lines of evidence suggest that the peptide is cyclic and sequences of Val-Ile-Ser-Ile and Ile-Pro-Val have been deduced by MS/MS as well as Edman degradation methods. Colutellin A inhibited CD4(+) T-cell activation of interleukin 2 (IL-2) production with an IC(50) of 167.3+/-0.38 nM, whereas cyclosporin A in the same test yielded a value of 61.8 nM. Inhibition of IL-2 production by collutellin A at such a low concentration indicates the potential immunosuppressive activity of this compound. In repeated experiments, cyclosporin A at or above 8 microg ml(-1) exhibited high levels of cytotoxicity on human peripheral blood mononuclear cells, whereas collutellin A or DMSO (carrier) alone, after 24 and 48 h of culture, exhibited no toxicity. Because of these properties collutellin A has potential as a novel immunosuppressive drug.