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First Report of Pectobacterium parmentieri Causing Bacterial Soft Rot and Blackleg on Potato in Maine

Ge, T. L., Jiang, H. H., Hao, J. J., Johnson, S. B.
Plant disease 2018 v.102 no.2 pp. 437
Dickeya, Pectobacterium, Solanum tuberosum, agar, algorithms, bacteria, blackleg of potato, cultivars, databases, ethanol, financial economics, genes, genomics, gentian violet, greenhouse experimentation, greenhouses, oligodeoxyribonucleotides, pathogenicity, pathogens, plant tissues, planting, polymerase chain reaction, potatoes, potting mix, ribosomal DNA, ribosomal RNA, sequence analysis, sodium hypochlorite, soil, stems, surveys, tubers, Maine
Bacterial soft rot and blackleg of potato (Solanum tuberosum) caused by Pectobacterium spp. and Dickeya spp. can be a problem that causes severe economic losses (Jiang et al. 2016). A survey was conducted in 10 Maine potato fields from May to August 2015, following the outbreak of blackleg in the Northeastern United States (Jiang et al. 2016). Potato plants with blackleg symptoms and darkened and necrotic basal stems were collected from Maine potato fields in 2015. The lower 2 cm of blackleg symptomatic stems were excised, surface sanitized with 75% ethanol for 40 s, transferred to a 0.65% sodium hypochlorite solution for 1 min, and rinsed with sterile distilled water. The stem segment was then placed in a 50-ml tube containing 10 ml of sterile distilled water for 5 min to allow the bacteria to release from the plant tissue. The bacterial suspension was plated on crystal violet polypectate agar (Hélias et al. 2012), and single colonies that formed cavities after 3 days were selected, purified, and stored. Strain ME175 was used in the following studies. Genomic DNA was extracted from culture of strain ME175 using the FastDNA kit (MP Biomedicals, Santa Ana, CA). Polymerase chain reaction (PCR) was performed using the extracted DNA as a template and primer pair fD/rP targeting 16S rRNA gene, and primer pair recA F/R targeting recA gene, resulting in amplicons of 1,417 and 714 bp, respectively (Waleron et al. 2002). These PCR products were sequenced and analyzed using BLAST algorithm against the NCBI database. Results showed that the 16S rDNA region (GenBank accession no. KY930911) shared 100% identity with P. wasabiae strain RNS08.42.1; recA region (GenBank accession no. KY930912) had 100% identity with P. wasabiae strain WPP163. Based on genomic and phenotypical data, the above two reference strains of P. wasabiae isolated from potato plant have been transferred to P. parmentieri (Khayi et al. 2016). Thus, strain ME175 was confirmed to be P. parmentieri. Strain ME175 was also characterized for pathogenicity in a greenhouse study. Potato (cultivar Shepody) tubers were planted in potting mix in a plastic pot. Six weeks after planting, stems of three plants were inoculated by injecting 100 µL of bacterial suspension (10⁷ CFU/ml) of ME175 on the stem 2 cm above the soil line. Three stems were injected with sterile distilled water as a control. Plants were maintained at 25°C (day) and 15°C (night) in a greenhouse. Sixteen days after inoculation, stems inoculated with the pathogen had black-colored and rotten stems, whereas the control plants remained symptomless. Bacteria were reisolated from the symptomatic stems and confirmed to be P. parmentieri using PCR and DNA sequencing as described above. The greenhouse trial was repeated with consistent results. In addition, tuber inoculation with ME175 showed severe soft rot symptoms. Thus, strain ME175 was confirmed as P. parmentieri. To our knowledge, this is the first report of soft rot and blackleg of potato caused by P. parmentieri in Maine.