A secreted WY-domain-containing protein present in European isolates of the oomycete Plasmopara viticola induces cell death in grapevine and tobacco species.

Plasmopara viticola is a biotrophic oomycete pathogen causing grapevine downy mildew. We characterized the repertoire of P. viticola effector proteins which may be translocated into plants to support the disease. We found several secreted proteins that contain canonical dEER motifs and conserved WY-domains but lack the characteristic RXLR motif reported previously from oomycete effectors. We cloned four candidates and showed that one of them, Pv33, induces plant cell death in grapevine and Nicotiana species. This activity is dependent on the nuclear localization of the protein. Sequence similar effectors were present in seven European, but in none of the tested American isolates. Together our work contributes a new type of conserved P. viticola effector candidates.

Advances in Diagnostics of Downy Mildews: Lessons Learned from Other Oomycetes and Future Challenges.

Downy mildews are plant pathogens that damage crop quality and yield worldwide. Among the most severe and notorious crop epidemics of downy mildew occurred on grapes in the mid-1880s, which almost destroyed the wine industry in France. Since then, there have been multiple outbreaks on sorghum and millet in Africa, tobacco in Europe, and recent widespread epidemics on lettuce, basil, cucurbits, and spinach throughout North America. In the mid-1970s, loss of corn to downy mildew in the Philippines was estimated at US$23 million. Today, crops that are susceptible to downy mildews are worth at least $7.5 billion of the United States' economy. Although downy mildews cause devastating economic losses in the United States and globally, this pathogen group remains understudied because they are difficult to culture and accurately identify. Early detection of downy mildews in the environment is critical to establish pathogen presence and identity, determine fungicide resistance, and understand how pathogen populations disperse. Knowing when and where pathogens emerge is also important for identifying critical control points to restrict movement and to contain populations. Reducing the spread of pathogens also decreases the likelihood of sexual recombination events and discourages the emergence of novel virulent strains. A major challenge in detecting downy mildews is that they are obligate pathogens and thus cannot be cultured in artificial media to identify and maintain specimens. However, advances in molecular detection techniques hold promise for rapid and in some cases, relatively inexpensive diagnosis. In this article, we discuss recent advances in diagnostic tools that can be used to detect downy mildews. First, we briefly describe downy mildew taxonomy and genetic loci used for detection. Next, we review issues encountered when identifying loci and compare various traditional and novel platforms for diagnostics. We discuss diagnosis of downy mildew traits and issues to consider when detecting this group of organisms in different environments. We conclude with challenges and future directions for successful downy mildew detection.

Genome Sequence and Architecture of the Tobacco Downy Mildew Pathogen Peronospora tabacina.

Peronospora tabacina is an obligate biotrophic oomycete that causes blue mold or downy mildew on tobacco (Nicotiana tabacum). It is an economically important disease occurring frequently in tobacco-growing regions worldwide. We sequenced and characterized the genomes of two P. tabacina isolates and mined them for pathogenicity-related proteins and effector-encoding genes. De novo assembly of the genomes using Illumina reads resulted in 4,016 (63.1 Mb, N50 = 79 kb) and 3,245 (55.3 Mb, N50 = 61 kb) scaffolds for isolates 968-J2 and 968-S26, respectively, with an estimated genome size of 68 Mb. The mitochondrial genome has a similar size (approximately 43 kb) and structure to those of other oomycetes, plus several minor unique features. Repetitive elements, primarily retrotransposons, make up approximately 24% of the nuclear genome. Approximately 18,000 protein-coding gene models were predicted. Mining the secretome revealed approximately 120 candidate RxLR, six CRN (candidate effectors that elicit crinkling and necrosis), and 61 WY domain-containing proteins. Candidate RxLR effectors were shown to be predominantly undergoing diversifying selection, with approximately 57% located in variable gene-sparse regions of the genome. Aligning the P. tabacina genome to Hyaloperonospora arabidopsidis and Phytophthora spp. revealed a high level of synteny. Blocks of synteny show gene inversions and instances of expansion in intergenic regions. Extensive rearrangements of the gene-rich genomic regions do not appear to have occurred during the evolution of these highly variable pathogens. These assemblies provide the basis for studies of virulence in this and other downy mildew pathogens.

Universally Primed-PCR indicates geographical variation of Peronospora farinosa ex. Chenopodium quinoa.

In the Andean region of South America downy mildew, caused by Peronospora farinosa, is the most important disease of quinoa (Chenopodium quinoa). Peronospora farinosa, a highly polyphyletic species, occurs on quinoa and wild relatives on all continents. However, very little is known about the geographic diversity of the pathogen. As the interest in quinoa as a novel crop is increasing worldwide, geographical differences in the population structure of the downy mildew pathogen must be taken into consideration in order to design appropriate control strategies under a variety of circumstances. As a step towards understanding the geographic diversity of P. farinosa from quinoa, 40 downy mildew isolates from the Andean highlands and Denmark were characterized using universally primed PCR (UP-PCR). Eight UP-PCR primers were tested. A combined analysis of markers separated the Danish and Andean isolates in two distinct clusters. This study raises new questions about the origin and spread of P. farinosa on quinoa, its geographic diversity and host specificity.