Abstract
Downy mildews affect important crops and cause severe losses in production worldwide. Accurate identification and monitoring of these plant pathogens, especially at early stages of the disease, is fundamental in achieving effective disease control. The rapid development of molecular methods for diagnosis has provided more specific, fast, reliable, sensitive, and portable alternatives for plant pathogen detection and quantification than traditional approaches. In this review, we provide information on the use of molecular markers, serological techniques, and nucleic acid amplification technologies for downy mildew diagnosis, highlighting the benefits and disadvantages of the technologies and target selection. We emphasize the importance of incorporating information on pathogen variability in virulence and fungicide resistance for disease management and how the development and application of diagnostic assays based on standard and promising technologies, including high-throughput sequencing and genomics, are revolutionizing the development of species-specific assays suitable for in-field diagnosis. Our review provides an overview of molecular detection technologies and a practical guide for selecting the best approaches for diagnosis.
Highlights
The mitochondrial genomes of over 87 species of oomycetes, including several important genera like Phytophthora, Pythium, and a broad range of downy mildew pathogens, are available [82]. These genomic resources are useful for designing specific markers that focus on unique gene order, unique putative ORFs or indels and avoid the dependency on polymerase chain reaction (PCR) stringency while providing flexibility to develop assays for genus, species, or infraspecific taxonomic categories [32,75,82]
One of the best options is to combine Mitochondrial DNA (mtDNA) loci with quantitative PCR (qPCR) to improve the detection of low pathogen levels in environmental samples and early stages of infection in the host [86]. This strategy allowed the development of species-specific markers based on the cytochrome oxidase 2 gene for Ps. cubensis [76] and an mtDNA region for lettuce downy mildew
A similar approach based on metagenomic filtering using the Contig Annotation Tool (CAT) removed contaminating sequences from the Pe. effusa genome [37], generating an assembly comparable to those of pathogens that can be isolated in axenic culture (Figure 5)
Summary
Downy mildew (DM) pathogens include several species of obligate oomycetes that can cause devastating damage to commercial [1], landscape [2], and natural ecosystem plants [3,4,5]. The diagnostics of downy mildew diseases has mainly relied upon direct observation of symptoms and signs using the naked eye or hand lenses and microscopes [27] This is possible after observing their sexual (e.g., antheridia and oogonia) and asexual structures (e.g., sporangiophores, sporangia, and zoospores) (Figure 1) involved in survival and dispersion, and because many downy mildew pathogens produce distinctive foliar signs and symptoms when colonizing a host plant [2,28,29]. Ps. humuli, the downy mildew pathogen of hop, causes systemic infections including the rhizomes used as planting material to start new hop yards [8]. Ps. humuli, the downy mildew pathogen of hop, causes systemic infections tions including the rhizomes used as planting material to start new hop yards [8].
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