Alpha-proteobacteria cultivated from marine sponges display branching rod morphology

PCR amplification of two CHS gene fragments of the obligate biotroph Plasmopara viticola, the causal agent of downy mildew of grapevine, is described. While one fragment shows homology to fungal class IV chitin synthases, the other fragment groups with other oomycete chitin synthases to form a novel class of chitin synthases most closely related to class I-III. RT-PCR experiments indicate that PvCHS1 is constitutively expressed, whereas PvCHS2 is specifically transcribed in sporangiophores and sporangia. Analyses of wheat germ agglutinin labeling patterns by confocal laser scanning microscopy show that chitin is present on the surface of hyphal cell walls during in planta growth, and of sporangiophores and sporangia.     

A new species of <Emphasis Type="Italic">Brasiliomyces</Emphasis> (Erysiphaceae) on <Emphasis Type="Italic">Dalbergia cultrata</Emphasis> var. <Emphasis Type="Italic">cultrata</Emphasis> from Thailand

A powdery mildew fungus on leaves of Dalbergia cultrata var. cultrata (Fabaceae) collected at the Queen Sirikit Botanical Garden in northern Thailand is proven to be a new species of the genus Brasiliomyces and is described as B. chiangmaiensis sp. nov. with light and SEM micrographs. Differences in known Brasiliomyces species are discussed, and a key to species of this genus is provided.     

Mildew-resistant mutants induced in North American two- and six-rowed malting barley cultivars

Mildew-resistant mutants were induced with sodium azide in three North American malting barley cultivars, two in the six-rowed Ursula (URS1 and URS2), one in the six-rowed Gertrud (GER1), and one in the two-rowed Prudentia (PRU1). Two of the mutants, URS1 and PRU1, showed complete resistance and were shown to have two new alleles at the mlo locus; these were designated, respectively, mlo31 and mlo32. Mutant URS2, showing partial resistance, was inherited as a dominant gene, but was not an allele at the Mla locus. The mean yield of each mutant was higher than that of its parental line, but yield levels varied across environments, although this was independent of the severity of the mildew attack. Other reasons, for example, the severity of the necrotic lesions in the mutants, may account for yield variations. The malting quality of the GER1 mutant proved similar to that of Gertrud, but both URS1 and URS2 showed lower malt extract than Ursula. This lower extract might be due to the smaller grain size of the mutants that could, in turn, result from necrotic lesions in the leaves, as implied by the effects on grain yield.Communicated by G. Wenzel     

Loss of actin cytoskeletal function and EDS1 activity,in combination,severely compromises non-host resistance in Arabidopsis against wheat powdery mildew

Plant immunity against the majority of the microbial pathogens is conveyed by a phenomenon known as non-host resistance (NHR). This defence mechanism affords durable protection to plant species against given species of phytopathogens. We investigated the genetic basis of NHR in Arabidopsis against the wheat powdery mildew fungus Blumeria graminis f. sp. tritici (Bgt). Both primary and appressorial germ tubes were produced from individual Bgt conidia on the surface of the Arabidopsis leaves. Attempted infection occasionally resulted in successful penetration, which led to the development of an abnormal unilateral haustorium. Inoculation of a series of Arabidopsis defence-related mutants with Bgt resulted in the attenuation of reactive oxygen intermediate (ROI) production and salicylic acid (SA)-dependent defence gene expression in eds1, pad4 and nahG plants, which are known to be defective in some aspects of host resistance. Furthermore, Bgt often developed bilateral haustoria in the mutant Arabidopsis lines that closely resembled those formed in wheat. A similar decrease in NHR was observed following treatment of the wild-type Arabidopsis plants with cytochalasin E, an inhibitor of actin microfilament polymerisation. In eds1 mutants, inhibition of actin polymerisation severely compromised NHR in Arabidopsis against Bgt. This permitted completion of the Bgt infection cycle on these plants. Therefore, actin cytoskeletal function and EDS1 activity, in combination, are major contributors to NHR in Arabidopsis against wheat powdery mildew.     

Functional analysis of barley RAC/ROP G-protein family members in susceptibility to the powdery mildew fungus

Small monomeric G-proteins of the plant ras (rat sarcome oncogene product) related C3 botulinum toxin substrate (RAC)/Rho of plants (ROP) family are molecular switches in signal transduction of many cellular processes. RAC/ROPs regulate hormone effects, subcellular gradients of Ca2+, the organisation of the actin cytoskeleton and the production of reactive oxygen intermediates. Therefore, we followed a genetic bottom-up strategy to study the role of these proteins during the interaction of barley (Hordeum vulgare L.) with the fungal biotrophic pathogen Blumeria graminis f.sp. hordei (Bgh). We identified six barley RAC/ROP proteins and studied their gene expression. Five out of six Rac/Rop genes were expressed constitutively in the leaf epidermis, which is the site of interaction with Bgh. None of the genes showed enhancement of mRNA abundance after inoculation with Bgh. After microprojectile mediated transformation of single barley epidermal cells with constitutively activated mutant RAC/ROP proteins, we found an RAC/ROP-specific enhancement of pathogen accessibility, tagging HvRACB, HvRAC3 and HvROP6 as host proteins potentially involved in the establishment of susceptibility to Bgh. Confocal laser scanning microscopy (CLSM) of green fluorescent protein (GFP):HvRAC/ROP-transformed cells revealed varying strengths of plasma membrane association of barley RAC/ROPs. The C-terminal CAAX motif for presumable prenylation or the C-terminal hypervariable region (HVR), respectively, were required for membrane association of the RAC/ROPs. Proper intracellular localisation was essential for HvRACB and HvRAC3 function. Together, our data support the view that different paths of host signal transduction via RAC/ROP G-proteins are involved in processes supporting parasitic entry into epidermal host cells.     

Accumulation of pathogenesis-related proteins in barley leaf intercellular spaces during leaf senescence

Accumulation of the pathogenesis-related (PR) proteins localised in intercellular spaces of barley primary leaves, chlorophyll content, structure of chloroplasts, and photosynthesis were examined during natural and in vitro induced leaf senescence (cultivation of whole plants in the dark or detached leaves under nutrient deficiency). Some of PR proteins accumulated during natural senescence, but their accumulation pattern was different from those of pathogen-induced as well as during in vitro-induced senescence, which indicate different molecular bases of these processes. Photosynthetic rate and chlorophyll content indicate that natural senescence of barley primary leaves began from 15th day after sowing. In 35-d-old first leaves, the chloroplasts showed typical characteristics of senescence as significant decrease of size, greater grana, and prominent plastoglobuli. The chloroplasts contained more grana under in vitro induced senescence and they had reduced length in the dark. Correspondingly, accumulation of PR proteins was detectable on about the 15th day but the content of some PR proteins increased in later stages of senescence. This revised version was published online in July 2006 with corrections to the Cover Date.     

Identification of a QTL decreasing yield in barley linked to Mlo powdery mildew resistance

A molecular marker map, including Mlo mildew resistance, of the spring barley cross Derkado (Mlo-resistant) × B83-12/21/5 (Mlo-susceptible) was scanned for yield QTLs to determine whether the association of Mlo resistance with reduced yield was due to linkage or pleiotropy. Over the mapped portion of the genome of the cross, the QTL with the greatest effect upon yield was located within a 22 cM region between mlo and the simple sequence repeat HVM67 on chromosome 4(4H). The association of Mlo resistance with lower yield was therefore due to a repulsion linkage. Analysis of yield component characters revealed QTL alleles for reduced grain number and earlier heading date in the same region, also associated with Mlo resistance. Genotyping of a range of cultivars and sources of Mlo resistance with the HVM67 simple sequence repeat showed that the Derkado HVM67 allele was rare as it was found only in one other cultivar and four land-races or sources of disease resistance. Grannenlose Zweizeilige, the source, and Salome, the carrier of Mlo resistance in Derkado, have the same HVM67 genotype, although Salome was a mixture of two genotypes. The entire mlo-HVM67 chromosomal segment from Grannenlose Zweizeilige is therefore thought to have been transmitted to Derkado, possibly through joint selection for Mlo resistance and early heading. L92, synonym EP79, was another source of Mlo resistance with the same HVM67 allele as Derkado but recombination must have occurred during the breeding of Atem as it possesses a different HVM67 allele which is present in all the other Mlo sources and cultivars surveyed. Abbreviations: GN, grains per main stem ear; HD, heading date; MSTGW, thousand grain weight derived from GN and MSY; MSY, yield of grain on the main stem; PY, yield of grain from the whole plot; sCIM, simplified compound interval mapping; SIM, simple interval mapping; SPY, single plant yield; S-SAP, sequence-specific amplification polymorphism; TGW, thousand grain weight derived from bulk of plot seed; TN, number of fertile stems per plant.     

Quantitative proteomic analysis of cabernet sauvignon grape cells exposed to thermal stresses reveals alterations in sugar and phenylpropanoid metabolism

Grapes (Vitis vinifera) are a valuable fruit crop and wine production is a major industry. Global warming and expanded range of cultivation will expose grapes to more temperature stresses in future. Our study investigated protein level responses to abiotic stresses, with particular reference to proteomic changes induced by the impact of four different temperature stress regimes, including both hot and cold temperatures, on cultured grape cells. Cabernet Sauvignon cell suspension cultures grown at 26°C were subjected to 14 h of exposure to 34 and 42°C for heat stress, and 18 and 10°C for cold stress. Cells from the five temperatures were harvested in biological triplicates and label‐free quantitative shotgun proteomic analysis was performed. A total of 2042 non‐redundant proteins were identified from the five temperature points. Fifty‐five proteins were only detected in extreme heat stress conditions (42°C) and 53 proteins were only detected at extreme cold stress conditions (10°C). Gene Ontology (GO) annotations of differentially expressed proteins provided insights into the metabolic pathways that are involved in temperature stress in grape cells. Sugar metabolism displayed switching between alternative and classical pathways during temperature stresses. Additionally, nine proteins involved in the phenylpropanoid pathway were greatly increased in abundance at extreme cold stress, and were thus found to be cold‐responsive proteins. All MS data have been deposited in the ProteomeXchange with identifier PXD000977 ( http://proteomecentral.proteomexchange.org/dataset/PXD000977 ).     

Epichloë endophyte affects the ability of powdery mildew (Blumeria graminis) to colonise drunken horse grass (Achnatherum inebrians)

The effect of the systemic seed-borne endophyte Epichloë gansuensis on the colonization by Blumeria graminis, the cause of powdery mildew disease, and the growth of the host grass Achnatherum inebrians, was studied under four soil water conditions. Infection incidence, disease lesion parameters, disease index, biomass production and growth parameters of the grass with and without the fungal endophyte were measured and counted after a period of disease. There was a significantly (P < 0.05) higher disease incidence and disease index for endophyte-free (E−) compared to endophyte-infected (E+) plants under different drought stresses. The presence of the endophyte significantly positively affected all of the host grass growth factors. The results of the present study demonstrate that the presence of the Epichloë endophyte reduced the ability of B. graminis to colonise A. inebrians and also conferred enhanced host plant growth at all soil water conditions tested.