First Report of a Leaf Spot Disease of Golden Dewdrop (Duranta erecta) Caused by Pseudomonas cichorii and a Xanthomonas Species in Florida.

Duranta erecta (Verbenaceae) is used extensively in southern states as an ornamental shrub and has replaced boxwood as the most common short hedge accenting flower beds. Over the past 2 years, during warm wet periods, dark necrotic leaf spots have been observed on golden dewdrop plants in Florida. Isolations from these spots on Difco nutrient agar (NA) consistently yielded two types of bacterial colonies that were not always simultaneously present: 1) round butyrous, bright yellow and 2) flat cream-colored. Both were 2 mm in size after 48 h, gram-negative, and produced a hypersensitivity reaction (HR) on tobacco cv Hicks. Yellow colony bacteria were oxidase negative and non-fluorescent on King's medium B (KMB) (1). Cream-colored colony bacteria were oxidase positive and fluorescent on KMB. Three isolates of both types were selected for further study. Partial 16S rDNA sequencing and fatty acid analysis (FAME) MIDI Microbial Identification System (Microbial ID, Inc., Newark, DE) were used for identification of strains. The 16S rDNA primers used were; forward primer AMB14 5'-TCCAGCAATGCCGCGTGTGT-3' and reverse primer AMB13 5'-CATCCACCGCTTGTGCGGGT-3'. The PCR program consisted of an initial denaturing cycle of 95°C for 2 min followed by 30 cycles of denaturing at 95°C for 30 s, annealing at 60°C for 40 s and extension at 72°C for 1 min and one final extension at 72°C for 10 min. Using FAME analysis, the three strains of the cream-colored colony type were identified as Pseudomonas cichorii with high similarity values (0.907, 0.961, 0.819) and this corresponded well with the 16S rDNA sequences where 99% sequence identity was observed with P. cichorii strain JBC1 16S ribosomal RNA gene, partial sequence GenBank Accession No. JF951725. Two of the three yellow colony strains were identified by MIDI FAME profiles as Xanthomonas axonopodis pv. manihotis with similarity coefficients of 0.767 and 0.826. The third strain had a low similarity match to X. a. pv. carotae (0.541). The 16S rDNA sequencing of these strains showed 98% sequence identity to X. citri subsp. citri strain SA1 16S ribosomal RNA gene only, partial sequence identity JQ890091.1, thus indicating a possible undescribed X. axonopodis pathovar. To satisfy Koch's postulates, three golden dewdrop 'Golden Mound' plants were sprayed with a suspension of 108 CFU/ml of a 2-day NA culture of each strain, bagged for 24 h to raise humidity, and placed in a greenhouse. A strain of P. cichorii (P409) isolated from chrysanthemum was used as a positive control when comparing cream-colored strains. A saline buffered control was used as a negative control. Within 3 weeks, leaf spots developed on plants sprayed with each of the six strains, including positive control strain of P. cichorii. Reisolations yielded the same type of colony as the originally inoculated strain. Inoculation experiments were repeated three times with a minimum of three plants per isolate with similar results. To our knowledge, this is the first report in the United States of bacterial leaf spot caused by P. cichorii and X. axonopodis on golden dewdrop. An earlier morphological and physiological description of a Xanthomonas sp. was done on Duranta in India in 1962 (2). Due to the difficulty in controlling bacterial diseases and the popularity of Duranta spp. in the landscape, these diseases may present a problem in ornamental trade. References: (1) E. O. King et al. J. Lab. Clin. Med. 44:301, 1954. (2) M. C. Srinivasan et al. Proc. Indian Acad. Sci. 56:88, 1962.

HopX1 in Erwinia amylovora functions as an avirulence protein in apple and is regulated by HrpL.

Fire blight is a devastating disease of rosaceous plants caused by the Gram-negative bacterium Erwinia amylovora. This pathogen delivers virulence proteins into host cells utilizing the type III secretion system (T3SS). Expression of the T3SS and of translocated and secreted substrates is activated by the alternative sigma factor HrpL, which recognizes hrp box promoters upstream of regulated genes. A collection of hidden Markov model (HMM) profiles was used to identify putative hrp boxes in the genome sequence of Ea273, a highly virulent strain of E. amylovora. Among potential virulence factors preceded by putative hrp boxes, two genes previously known as Eop3 and Eop2 were characterized. The presence of functionally active hrp boxes upstream of these two genes was confirmed by ß-glucuronidase (GUS) assays. Deletion mutants of the latter candidate genes, renamed hopX1(Ea) and hopAK1(Ea), respectively, did not differ in virulence from the wild-type strain when assayed in pear fruit and apple shoots. The hopX1(Ea) deletion mutant of Ea273, complemented with a plasmid overexpressing hopX1(E)(a), suppressed the development of the hypersensitivity response (HR) when inoculated into Nicotiana benthamiana; however, it contributed to HR in Nicotiana tabacum and significantly reduced the progress of disease in apple shoots, suggesting that HopX1(Ea) may act as an avirulence protein in apple shoots.

Complete genome sequence of the plant pathogen Erwinia amylovora strain ATCC 49946.

Erwinia amylovora causes the economically important disease fire blight that affects rosaceous plants, especially pear and apple. Here we report the complete genome sequence and annotation of strain ATCC 49946. The analysis of the sequence and its comparison with sequenced genomes of closely related enterobacteria revealed signs of pathoadaptation to rosaceous hosts.