First Report of Leaf Necrosis Caused by Pseudomonas viridiflava on Melon Seedlings in Italy.

Abstract

In April 2001, necrotic lesions surrounded by thin, water-soaked halos were observed on cotyledons of 12-day-old melon seedlings (Cucumis melo var. reticulatus, cv. Baggio, Calipso, and Proteo) grown in plant beds in an unheated greenhouse located in the Province of Perugia (central Italy). The incidence of the disease was approximately 10%, and the economic impact was limited as seedlings recovered from the disease. Cream-colored, mucoid, bacterial colonies were consistently isolated on nutrient agar from the diseased leaf tissues. Two representative strains selected for identification were gram negative, fluorescent on King's medium B, and had oxidative but not fermentative metabolism. They were levan negative, oxidase negative, potato rot positive, arginine dihydrolase negative, and tobacco hypersensitive response positive in LOPAT tests. These isolates showed pectolytic activity at pH 8 but not at pH 4 and utilized L-arabinose, D(-)-tartrate and L-lactate. They did not utilize sucrose, L(+)-tartrate, or trigonelline, and did not produce acid from sucrose after 21 days of incubation. These results were similar to those obtained with the type strain LMG 2352T of Pseudomonas viridiflava (Burkholder) Dowson. Although suitable for strain characterization, we found that repetitive sequence-based polymerase chain reaction (rep-PCR) conducted with primer BOXA1R was not appropriate for identifying P. viridiflava. In fact, each P. viridiflava strain tested, (LMG 2352T, LMG 2353, LMG 5397, and NCPPB 1382) generated unique fingerprints, which differed from the two melon strains. Pathogenicity tests were carried out with 3-week-old melon (cv. Baggio), cucumber (cv. Lungo verde degli ortolani), and zucchini (cv. Consul) plants (three plants for each species and isolate). To prepare the inoculum, the two bacterial strains were grown on nutrient agar for 24 h at 27°C, suspended in sterile deionized water, and adjusted to 1 × 106 CFU ml-1. Young leaves of melon, cucumber, and zucchini plants infiltrated with bacterial suspensions by a glass atomizer at high pressure developed small, chlorotic spots with necrotic centers surrounded by water-soaked halos 5 to 7 days after inoculation. The most severe symptoms were observed on melon plants. The two strains also induced severe symptoms (chlorosis and water soaking) on Arabidopsis thaliana (ecotype Columbia-0) 3 to 4 days after inoculation. No symptoms were observed in control plants. The bacterium was readily recovered from inoculated plants, and their rep-PCR fingerprints were identical to the strains used for inoculation. On the basis of biochemical, physiological, nutritional, and pathogenicity tests, it was concluded that the bacteria isolated from symptomatic melon seedlings were P. viridiflava. To our knowledge, this is the first report of P. viridiflava attacks on melon plants in Italy. The disease was previously recorded in Turkey (1) and Greece (2).