First report of bacterial leaf spot caused by Pseudomonas cichorii on Thai Basil in Hawaii

Authors: LUIZ, BLAINE - University Of Hawaii; HELLER, WADE - University Of Hawaii; BRILL, EVA - University Of Hawaii; BUSHE, BRIAN - University Of Hawaii; Keith, Lisa

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/17/2018
Publication Date: 10/24/2018
Citation: Luiz, B., Heller, W., Brill, E., Bushe, B., Keith, L.M. 2018. First report of bacterial leaf spot caused by Pseudomonas cichorii on Thai Basil in Hawaii. Plant Disease. 102(12):2637. https://doi.org/10.1094/PDIS-06-18-0995-PDN.
DOI: https://doi.org/10.1094/PDIS-06-18-0995-PDN

Interpretive Summary: Thai basil has unique anise and licorice flavors and is commonly used in Asian cuisine. In October 2017, water soaked, irregular, and grey to black leaf spots were observed in a garden in Hilo, Hawaii. The bacteria causing the disease was identified and its pathogenicity was proven. This is the first report of bacterial leaf spot caused by Pseudomonas cichorii on Thai basil in Hawaii.

Technical Abstract: Thai basil (Ocimum basilicum var. thyrsiflora) has unique anise and licorice flavors and is commonly used in Asian cuisine. In October 2017, leaf spots were observed in a garden in Hilo, Hawaii. The leaf spots were water soaked, irregular, and grey to black. Lesions ranged from 2 to 6 mm spots to marginal necrotic areas of 12 mm or larger. Three leaf samples were submitted to the USDA Agricultural Research Service (ARS) for diagnosis. Isolations were made on Luria-Bertani agar (LB) and nearly pure cultures of bacteria were consistently isolated from diseased tissue. Bacterial isolates were transferred to King’s Medium B agar (KMB) where non-mucoid, cream and fluorescent colonies under 365 nm UV light were obtained. A strain from a single colony (P18-1) was transferred and characterized as gram-negative, oxidase-positive, and produced a hypersensitive reaction in tobacco. The strain was identified as Pseudomonas cichorii based on the LOPAT scheme (Schaad et al. 2001). A partial 16S rRNA gene product (1,372 bp) using primers Y1 and Bacterial 16S reverse was sequenced and compared in GenBank (accession no. MH396006) and was 100% identical to multiple accessions of P. cichorii in the NCBI database, including an isolate from Bridal Bouquet Plumeria (Plumeria pudica) from Hawaii (accession no. MF536096). Bacterial identity was further confirmed by using the P. cichorii-specific primers Hrp1a/Hrp2a to amplify and sequence an 862 bp fragment (accession no. MH396007) which contained 12 single nucleotide polymorphisms relative to the P. pudica Plumeria isolate and shared 99% sequence identity to P. cichorii strain 83-1 (Cottyn et al. 2011). To prove pathogenicity, four healthy Thai basil plants were spray inoculated to run-off with a 107 CFU/ml bacterial suspension, placed in a clear plastic bag, and held at 24°C. Four control plants were sprayed with sterile distilled water. Two days post-inoculation, numerous 2 to 8 mm water soaked greyish-black lesions were observed on the adaxial leaf surface. Irregular leaf spots identical to the original diseased samples continued to develop and by day 3 several had coalesced into larger lesions. Bacterial colonies were consistently re-isolated from the inoculated plants and morphologically and molecularly identified as P. cichorii, while no bacterial colonies were isolated from the control plants, thus fulfilling Koch’s postulates. The test was repeated once. Bacterial leaf spot of sweet basil has been reported in Florida, Louisiana, and Indiana (Webb et al. 2016). To our knowledge, this is the first report of bacterial leaf spot caused by P. cichorii on Thai basil in Hawaii. According to Zhang and Roberts (2009), disease management involves decreasing humidity and standing moisture on plant surfaces through increased spacing of plants and elimination of overhead watering, as well as removal of diseased leaves and plants.