The entomopathogenic fungus Beauveria bassiana produces microsclerotia-like pellets mediated by oxidative stress and peroxisome biogenesis.

Several filamentous fungi are known to produce macroscopic pigmented hyphal aggregates named sclerotia. In recent years, some entomopathogenic fungi were reported to produce small sclerotia termed 'microsclerotia', becoming new potential propagules for biocontrol strategies. In this study, we described the production of microsclerotia-like pellets by the entomopathogenic fungus Beauveria bassiana. The carbon: nitrogen ratio equal to or higher than 12.5:1 amended with Fe2+ induced the germination of conidia, producing hyphal aggregate that formed sclerotial structures in submerged liquid cultures. These aggregates were able to tolerate desiccation as they germinated and subsequently produced viable conidia. Conidia derived from microsclerotial aggregates formulated with diatomaceous earth effectively kill Tribolium castaneum larvae. Optical and transmission microscopical imaging, qPCR and spectrophotometric analysis revealed that an oxidative stress scenario is involved in conidial differentiation into microsclerotia-like pellets, inducing fungal antioxidant response with high peroxidase activity - mainly detected in peroxisomes and mitochondria - and progress with active peroxisome proliferation. The results provide clues about B. bassiana microsclerotial differentiation and indicate that these pigmented aggregates are promising propagules for production, formulation and potentially application in the control of soil-inhabiting arthropod pests.

Cryopreparation and Electron Tomography of Yeast Cells.

Three-dimensional imaging of cells using electron tomography enables analysis of cell structure at unprecedented resolution. The preparation of cells for tomography using rapid freezing followed by freeze-substitution is an essential first step to ensure the optimal preservation of the cell structure for 3D studies. This protocol outlines a method for obtaining well-preserved cells using high-pressure freezing followed by freeze-substitution. We have found that this method is particularly well suited for electron tomography studies and has the added bonus of preserving antigenicity for immuno-electron microscopy. The steps involved in imaging cells and performing tomographic analysis of cellular structures are also outlined.

Building Cell Structures in Three Dimensions: Electron Tomography Methods for Budding Yeast.

Saccharomyces cerevisiae has been an important model system for numerous cellular, genetic, and molecular studies. However, this small eukaryote presents a challenge for imaging at the electron microscope level. Preparation of yeast using high-pressure freezing followed by freeze-substitution (HPF/FS) results in excellent preservation of cell structure in these difficult-to-fix samples. In particular, cells prepared by HPF/FS can be used for 3D electron tomography (ET) studies where optimum cell preservation is critical. Here, we discuss the advantages of using HPF/FS for ET and show examples of the utility of this method for building yeast cell structures in three dimensions.

Ciclo de vida de corynelia tropica, patógeno foliar en podocarpus saligna / Life cycle of corynelia tropica, leaf pathogen on podocarpus saligna

El presente estudio tuvo como objetivo determinar el ciclo de vida de Corynelia tropica, patógeno de hojas y ramillas en mañío de hoja larga (Podocarpus saligna). Mensualmente se recolectaron ramillas de P. saligna. En la medida que se fueron detectando estadíos interesantes dentro del ciclo de vida de este patógeno, la frecuencia de las colectas se intensificó en forma quincenal, e incluso semanal. Se pudo constatar que C. tropica presentó un ciclo de vida anual muy definido, con gran regularidad en la aparición y desarrollo de sus estructuras fructíferas. Invariablemente el ciclo de vida se inició en la primera quincena de noviembre con la inoculación de los nuevos brotes de P. saligna. A inicios de enero se manifestaron los primeros síntomas de la infección y a comienzos de marzo ya eran notorias las estructuras del anamorfo. A fines de mayo las estructuras del teleomorfo se hacían protuberantes y se iniciaba la formación de los ascos. A fines de julio los ascocarpos presentaban forma y tamaño ya adulto, y las ascosporas al interior de los ascos se encontraban en proceso de maduración. A fines de octubre las ascosporas ya estaban maduras y su liberación a comienzos de noviembre, sincronizaba con la emergencia de los brotes de P. saligna, con lo que se daba inicio una vez más a un nuevo ciclo de vida de C. tropica. Se pudo constatar que además del follaje y ramas, este patógeno también atacaba a los frutos de P.saligna.

This study aimed to determine the life cycle of Corynelia tropica, pathogen leaves and twins in longleaf mañío (Podocarpus saligna). Monthly P. saligna twigs were collected. As they were detecting interesting stages in the life cycle of this pathogen, the frequency of collections were intensified fortnightly or even weekly. It was found that C. tropica presented a very defined, with great regularity in the occurrence and development of their fruiting structures annual life cycle. Invariably the life cycle began in the first half of November with the inoculation of new outbreaks of P. saligna. In early January the first symptoms of infections manifested and early March were already notorious the anamorph structure. In late May teleomorph structures became prominent and the formation of the asci began. In late July the ascocarps presented adult size and shape, and the ascospores within the asci were maturing. In late October ascospores were ripe and in early November, release synchronized with outbreaks of P. saligna, which was given start again a new life cycle of C. tropica. It was found that besides the foliage and branches, this pathogen also attacked the fruits of P. saligna.

Tritirachium candoliense sp. nov., a novel basidiomycetous fungus isolated from the anoxic zone of the Arabian Sea.

A fungal culture (FCAS11) was isolated from coastal sediments of the Arabian Sea during the anoxic season. Multigene phylogenetic analyses confidentially place the organism as a novel species within the recently defined class Tritirachiomycetes, subphylum Pucciniomycotina, phylum Basidiomycota. We named the new species Tritirachium candoliense and provide the first description of a member of this class from a marine environment. DNA sequences and morphological characters distinguish T. candoliense from previously described Tritirachium species. Its growth characteristics, morphology, and ultrastructural features showed that under anoxic conditions the species grows slowly and produces mainly hyphae with only few blastoconidia. Electron microscopy revealed differences when the culture was exposed to anoxic stress. Notable ultrastructural changes occur for example in mitochondrial cristae, irregularly shaped fat globules and the presence of intracellular membrane invaginations. We assume that the growth characteristics and substrate utilization patterns are an adaptation to its source location, the seasonally anoxic environment of the Arabian Sea.

Non-developing ascospores in apothecia of asexually reproducing lichen-forming fungi

The presence of apothecia in mixed species (vegetatively reproducing lichens, occasionally producing ascomata) has been interpreted as a mechanism to increase genetic variability in mostly clonal populations. However, spore viability from these apothecia has not been studied. We asked whether ascospores of the mixed species Physconia grisea are viable and thereby contribute to increasing the genetic diversity within populations of this species. An ontogenetic study of spores in cultures of P. grisea and a related sexual species (P. distorta), showed that although mature apothecia from both species produced and discharged meiospores capable of germination, spores from P. grisea were only rarely (0.43 %) able to continue development whereas those from P. distorta germinated and developed successfully. The strongly reduced viability of P. grisea spores suggested that they do not have a strong reproductive function, at least in the two local populations analyzed. Additionally, we show that the segregation of Physconia grisea ssp. lilacina does not have molecular support (AU)

No disponible

Standard and Swedish variant types of the hybrid alder Phytophthora attacking alder in Hungary.

A new Phytophthora disease of common alder (Alnus glutinosa) similar to that previously reported in several countries in Europe has been observed in Hungary. Based on these earlier studies, the alder Phytophthora was considered likely to be a hybrid between P cambivora and a P fragariae-like species: across Europe a range of new alder Phytophthora is spreading that comprise a range of heteroploid hybrids including a 'standard' hybrid type and several other hybrid types termed 'variants'. Phenotypic and molecular features of the pathogen in Hungary were characterised and compared with isolates from elsewhere. The morphologies of five isolates from one region (Hévíz) resembled the common, 'standard' type, whereas the three isolates from another region (Hanság) exhibited traits similar to those of one of the 'variant' types, ie the Swedish 'variant'. Molecular markers of these two groups of Hungarian isolates also represented a good fit to those of the standard type and the Swedish variant, respectively. Isozyme patterns and profiles of restriction fragments of the entire internal transcribed spacer (ITS) region or mitochondrial DNAs and of RAPD-PCR products did not differ within a group, but distinct polymorphisms were exhibited between the two groups of isolates. Southern analysis of random amplified polymorphic DNA (RAPD) revealed the homologous nature of co-migrating bands of P cambivora and the isolates of alder Phytophthora. Furthermore, restriction fragment profiles of the ITS region of ribosomal DNAs and the mtDNAs were consistent with reported biparental origin of alder Phytophthora. The hybrid status of these continuously evolving pathogens raises many issues and challenges concerning efficient control measures.