Peronospora kuewa, sp. nov., a new downy mildew species infecting the endangered Hawaiian plant Plantago princeps var. princeps.

Plantago princeps var. princeps is an endangered native Hawaiian plant, and part of the recovery plan includes repopulation using plants grown in a nursery. However, disease pressure from downy mildew is hindering repopulation efforts. The organism associated with the downy mildew was determined to be a Peronospora species with brown, ellipsoid conidia measuring 21 by 16 µm on average, which was morphologically different from validly described species of Peronospora that infect Plantago species, but it was morphologically similar to the invalidly published species Peronospora lanceolatae (Art. 40.1). Comparison of mitochondrial cytochrome oxidase subunit I (cox1), mitochondrial cytochrome oxidase subunit II (cox2), nuclear internal transcribed spacer (ITS), and nuclear 28S rRNA D1-D2 (28S) loci revealed the unknown Peronospora to be molecularly divergent from Peronospora alta and Peronsopora plantaginis, but very similar to Peronospora from Plantago lanceolata, the type host of P. lanceolatae. Phylogenetic trees inferred with maximum likelihood and Bayesian inference from a concatenated alignmaent and individual gene trees confirmed the divergence of the unknown Peronospora from P. alta and P. plantaginis and its similarity to P. lanceolatae. However, attempts to inoculate Plantago lanceolata with the strain from Plantago princeps var. princeps were unsuccessful, which, in conjunction with divergence in ITS, suggests that the unknown Peronospora is specific to Plantago princeps var. princeps. Herein, the Peronospora strain on Plantago princeps var. princeps is described as the new species Peronospora kuewa based on morphology, molecular phylogenetics, and host specificity. In addition, Peronospora gaponenkoae is described here to honor Nina Ivanova Gaponenko on the basis of her description of P. lanceolatae.

Development of a robust, field-deployable loop-mediated isothermal amplification (LAMP) assay for specific detection of potato pathogen Dickeya dianthicola targeting a unique genomic region.

Destructive maceration, a wide host range, and longevity in non-plant substrates has established Dickeya dianthicola (blackleg of potato) as a significant threat to potato industries worldwide. To protect these businesses, a specific and sensitive point-of-care D. dianthicola detection tool is necessary. We have developed a loop-mediated isothermal amplification (LAMP) assay for specific, sensitive, and rapid detection of D. dianthicola, which can be streamlined for point-of-care use. The developed LAMP assay targets a unique gene, alcohol dehydrogenase, of D. dianthicola. Assay specificity was assessed using strains present in inclusivity (16 D. dianthicola strains) and exclusivity panels (56 closely related, potato pathogenic, and other bacterial strains). Amplification with strains of inclusivity panel occurred, and cross-reactivity with non-target DNA was not observed. The limit of detection (LOD) was 10 CFU/ml when dilutions were made before isolating the genomic DNA; however, LOD was determined as 1 pg using 10-fold serially diluted D. dianthicola genomic DNA. Similar LOD of 1 pg was observed when serially diluted target genomic DNA was mixed with host genomic DNA. LOD (1 pg) was also calculated with 10-fold serially diluted synthetic DNA fragments containing primer target sites. Naturally and artificially inoculated plant samples were used for field adaptability tests with the field-deployable Optigene Plant Material Lysis Kit and a heat block (65°C); the results were obtained within 20 minutes. Despite the lack of method precision, no false positives or false negatives were observed. Therefore, with prepared reactions and a steady heat source, this assay can be used for rapid point-of-care detection, which is imperative for quarantine, eradication, disease management, and border protection.