Factors Affecting Pseudomonas savastanoi pv. savastanoi Plant Inoculations and Their Use for Evaluation of Olive Cultivar Susceptibility.

ABSTRACT Pseudomonas savastanoi pv. savastanoi causes olive knot disease, which is present in most countries where olive trees are grown. Although the use of cultivars with low susceptibility may be one of the most appropriate methods of disease control, little information is available from inoculation assays, and cultivar susceptibility assessments have been limited to few cultivars. We have evaluated the effects of pathogen virulence, plant age, the dose/response relationship, and the induction of secondary tumors in olive inoculation assays. Most P. savastanoi pv. savastanoi strains evaluated were highly virulent to olive plants, but interactions between cultivars and strains were found. The severity of the disease in a given cultivar was strongly dependent of the pathogen dose applied at the wound sites. Secondary tumors developed in noninoculated wounds following inoculation at another position on the stem, suggesting the migration of the pathogen within olive plants. Proportion and weight of primary knots and the presence of secondary knots were evaluated in 29 olive cultivars inoculated with two pathogen strains at two inoculum doses, allowing us to rate most of the cultivars as having either high, medium, or low susceptibility to olive knot disease. None of the cultivars were immune to the disease.

Tumorigenic Agrobacterium sp. Isolated from Weeping Fig in Spain.

Agrobacterium-like colonies were recovered onto Roy-Sasser's medium from a young tumor (4 cm in diameter) on the stem of weeping fig (Ficus benjamina L.), 10 cm from the crown. The galled plant was collected in 1999 from a garden center in Valencia, Spain. After colony purification and tomato and weeping fig plant inoculations, one nonpathogenic and five Agrobacterium isolates that were tumorigenic in both plant species were characterized. On the basis of biovar classification tests, the nonpathogenic isolate was identified as belonging to biovar 1 of Agrobacterium (now called A. tumefaciens), whereas the tumorigenic isolates could not be assigned to any of the known Agrobacterium biovars. The isolates were positive for oxidase, growth in 2% NaCl, production of alkali from l-tartaric acid, and production of acid from mannitol-CaCO3 and negative for 3-ketolactose production, growth and pigmentation in ferric ammonium citrate, growth at 35°C, citrate utilization, acid production from sucrose and melezitose, and alkali production from malonic acid. Nopaline was the unique opine found in galls induced in weeping fig plants inoculated with the pathogenic isolates. Moreover, all isolates utilize the opine nopaline, but not octopine, manopine, agropine, chrysopine, cucumopine, or mikimopine. They were susceptible to agrocin 84 produced by strain K84. Heat-treated bacterial suspensions of these isolates yielded the expected amplification product using polymerase chain reaction (PCR) with the FGPtmr530/FGPtmr701' primers pair from the tmr gene (3). Aerial gall disease was first reported on F. benjamina in Florida (1), and the isolated agrobacteria belongs to a new species named A. larrymoorei (2). Later, tumorigenic agrobacteria from weeping fig galls were isolated in Italy and the Netherlands (4). Our data suggest that the tumorigenic strains isolated in Spain differ greatly from those first described in the United States (1) on the basis of alkali production from l-tartaric acid, chrysopine detection on tumors, susceptibility to agrocin 84, and tmr amplification, but they might be similar to some of the Italian isolates (4). To our knowledge, this is the first report of isolation of tumorigenic Agrobacterium sp. from F. benjamina L. in Spain. References: (1) H. Bouzar et al. Appl. Environ. Microbiol. 61:65, 1995. (2) H. Bouzar and J. B. Jones. Int. J. Syst. Evol. Microbiol. 51:1023. 2001. (3) X. Nesme et al. Pages 47-50 in: Endocytobiology IV. P. Nardon et al. eds. INRA, France, 1989. (4) A. Zoina et al. Plant Pathol. 50:620, 2001.

Systemic movement of Agrobacterium tumefaciens in several plant species.

AIMS: The systemic movement of Agrobacterium spp. inside plants of different species was studied to determine the most valuable diagnostic methodology for their detection. METHODS AND RESULTS: Pathogenic agrobacteria were detected by isolation and PCR in tissue away from primary tumours in tomato plants grown in the presence of Agrobacterium spp. Moreover, this bacterium was also able to induce secondary tumours beyond the inoculation site. In addition, the capacity of agrobacteria to translocate and induce secondary tumours was analysed in rose, grapevine, chrysanthemum, cherry and peach x almond hybrid GF677. No differences among strains of Agrobacterium spp. were detected in secondary tumour development, although some of them induced a significantly higher number of primary tumours in some species. Movement of inoculated pathogenic cells of four strains was also demonstrated in symptomless portions of the plant stems by isolation and PCR. Finally, pathogenic agrobacteria were detected in root, crown and stem portions of naturally infected walnuts. In all assays, PCR was the most efficient technique for detecting the movement of Agrobacterium spp. within the plants. CONCLUSIONS: Migration of agrobacteria inside plants is a complex phenomenon and more extensive than previously reported. Therefore, efficient and sensitive detection methods such as PCR must be used to select clean plants to avoid latent infections of Agrobacterium spp. SIGNIFICANCE AND IMPACT OF THE STUDY: The results show that migration of Agrobacterium spp. could be relatively frequent in several cultivated fruit trees, and systemic infections should be taken into account when designing strategies for controlling crown gall disease.

Evidence of Biological Control of Agrobacterium tumefaciens Strains Sensitive and Resistant to Agrocin 84 by Different Agrobacterium radiobacter Strains on Stone Fruit Trees.

The effectiveness of Agrobacterium radiobacter K84, 0341, and a K84 non-agrocin-producing mutant (K84 Agr) in biological control of crown gall on rootstocks of stone fruit trees was determined in three experiments. In experiment 1, K84 and 0341 controlled crown gall on plum plants in soil inoculated with two strains of Agrobacterium tumefaciens resistant to agrocin 84. In experiment 2, K84 controlled crown gall on peach plants in soils inoculated with strains of A. tumefaciens sensitive or resistant to agrocin 84 or with a mixture of both. However, the effectiveness of K84 was higher against the sensitive strain than against the resistant strain. There was a residual effect of K84 from one year to another in soil inoculated with the sensitive strains. In experiment 3, K84 and K84 Agr controlled crown gall on plum and peach plants in soils inoculated with strains of A. tumefaciens sensitive or resistant to agrocin 84. The control afforded by K84 was higher than that provided by K84 Agr against the sensitive strain but was similar against the resistant strain.