In planta proteomics and proteogenomics of the biotrophic barley fungal pathogen blumeria graminis f. sp hordei

To further our understanding of powdery mildew biology during infection, we undertook a systematic shotgun proteomics analysis of the obligate biotroph Blumeria graminis f. sp. hordei at different stages of development in the host. Moreover we used a proteogenomics approach to feed information into the annotation of the newly sequenced genome. We analyzed and compared the proteomes from three stages of development representing different functions during the plant-dependent vegetative life cycle of this fungus. We identified 441 proteins in ungerminated spores, 775 proteins in epiphytic sporulating hyphae, and 47 proteins from haustoria inside barley leaf epidermal cells and used the data to aid annotation of the B. graminis f. sp. hordei genome. We also compared the differences in the protein complement of these key stages. Although confirming some of the previously reported findings and models derived from the analysis of transcriptome dynamics, our results also suggest that the intracellular haustoria are subject to stress possibly as a result of the plant defense strategy, including the production of reactive oxygen species. In addition, a number of small haustorial proteins with a predicted N-terminal signal peptide for secretion were identified in infected tissues: these represent candidate effector proteins that may play a role in controlling host metabolism and immunity. Molecular & Cellular Proteomics 8: 2368-2381, 2009.

The effect of carrier substrate, dose rate and time of application on biocontrol efficacy of fungal antagonists against Armillaria root rot of strawberry plants.

In a glasshouse experiment using potted strawberry plants (cv. Cambridge Favourite) as hosts, the effect of selected fungal antagonists grown on 25 or 50 g of mushroom compost containing autoclaved mycelia of Agaricus bisporus, or wheat bran was evaluated against Armillaria mellea. Another glasshouse experiment tested the effect of application time of the antagonists in relation to inoculations with the pathogen. A significant interaction was found between the antagonists, substrates and dose rates. All the plants treated with Chaetomium olivaceum isolate Co on 50 g wheat bran survived until the end of the experiment which lasted 482 days, while none of them survived when this antagonist was added to the roots of the plants on 25 g wheat bran or 25 or 50 g mushroom compost. Dactylium dendroides isolate SP had a similar effect, although with a lower host survival rate of 33.3%. Trichoderma hamatum isolate Tham 1 and T. harzianum isolate Th23 protected 33.3% of the plants when added on 50 g and none when added on 25 g of either substrate, while 66.7% of the plants treated with T. harzianum isolate Th2 on 25 g, or T viride isolate TO on 50 g wheat bran, survived. Application of the antagonists on mushroom compost initially resulted in development of more leaves and healthier plants, but this effect was not sustained. Eventually, plants treated with the antagonists on wheat bran had significantly more leaves and higher health scores. The plants treated with isolate Th2 and inoculated with Armillaria at the same time had a survival rate of 66.7% for the duration of the experiment (475 days), while none of them survived that long when the antagonist and pathogen were applied with an interval of 85 days in either sequence. C. olivaceum isolate Co showed a protective effect only, as 66.7% of the plants survived when they were treated with the antagonist 85 days before inoculation with the pathogen, while none of them survived when the antagonist and pathogen were applied together or the infection preceded protection.

Genotype and temperature influence pea aphid resistance to a fungal entomopathogen

The influence of temperature on life history traits of four Acyrthosiphon pisum clones was investigated, together with their resistance to one genotype of the fungal entomopathogen Erynia neoaphidis . There was no difference among aphid clones in development rate, but they did differ in fecundity. Both development rate and fecundity were influenced by temperature, but all clones showed similar responses to the changes in temperature (i.e. the interaction term was nonsignificant). However, there were significant differences among clones in susceptibility to the pathogen, and this was influenced by temperature. Furthermore, the clones differed in how temperature influenced susceptibility, with susceptibility rankings changing with temperature. Two clones showed changes in susceptibility which mirrored changes in the in vitro vegetative growth rate of E. neoaphidis at different temperatures, whereas two other clones differed considerably from this expected response. Such interactions between genotype and temperature may help maintain heritable variation in aphid susceptibility to fungal pathogen attack and have implications for our understanding of disease dynamics in natural populations. This study also highlights the difficulties of drawing conclusions about the efficacy of a biological control agent when only a restricted range of pest genotypes or environmental conditions are considered.

In planta proteomics and proteogenomics of the biotrophic barley fungal pathogen blumeria graminis f. sp hordei

To further our understanding of powdery mildew biology during infection, we undertook a systematic shotgun proteomics analysis of the obligate biotroph Blumeria graminis f. sp. hordei at different stages of development in the host. Moreover we used a proteogenomics approach to feed information into the annotation of the newly sequenced genome. We analyzed and compared the proteomes from three stages of development representing different functions during the plant-dependent vegetative life cycle of this fungus. We identified 441 proteins in ungerminated spores, 775 proteins in epiphytic sporulating hyphae, and 47 proteins from haustoria inside barley leaf epidermal cells and used the data to aid annotation of the B. graminis f. sp. hordei genome. We also compared the differences in the protein complement of these key stages. Although confirming some of the previously reported findings and models derived from the analysis of transcriptome dynamics, our results also suggest that the intracellular haustoria are subject to stress possibly as a result of the plant defense strategy, including the production of reactive oxygen species. In addition, a number of small haustorial proteins with a predicted N-terminal signal peptide for secretion were identified in infected tissues: these represent candidate effector proteins that may play a role in controlling host metabolism and immunity. Molecular & Cellular Proteomics 8: 2368-2381, 2009.

Biorefinery development through utilization of biodiesel industry by-products as sole fermentation feedstock for 1,3-propanediol production

Rapeseed meal (RSM) hydrolysate was evaluated as substitute for commercial nutrient supplements in 1,3-propanediol (PDO) fermentation using the strain Clostridium butyricum VPI 1718. RSM was enzymatically converted into a generic fermentation feedstock, enriched in amino acids, peptides and various micro-nutrients, using crude enzyme consortia produced via solid state fermentation by a fungal strain of Aspergillus oryzae. Initial free amino nitrogen concentration influenced PDO production in batch cultures. RSM hydrolysates were compared with commercial nutrient supplements regarding PDO production in fed-batch cultures carried out in a bench-scale bioreactor. The utilization of RSM hydrolysates in repeated batch cultivation resulted in a PDO concentration of 65.5 g/L with an overall productivity of 1.15 g/L/h that was almost 2 times higher than the productivity achieved when yeast extract was used as nutrient supplement.