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Carbohydrate utilization of Erwinia amylovora in vitro
Published May 10, 2004

Nectar is a multi-component aqueous solution that promotes bacterial multiplication. The concentration of nectar in plant flowers is not stable since it is under the influence of environmental conditions, especially free moisture and relative humidity. Experiments were conducted with "artificial nectar" and directed along two lines: (1) determi...nation of the optimal concentrations of carbohydrates for the growth of E. amylovora development (2) consumption of different carbohydrates besides basic sugars.

Solutions of "artificial nectar" were prepared in different compositions by changing the dominance of basic sugars (fructose — glucose —sucrose) in proportions of 2:1:1, 1:2:1, 1:1:2 and between concentrations of 10-0.6% (diluted with Basal minimum broth) in order to determine optimal conditions for the development of E. amylovora.

At a basic sugar concentration of 10% bacterial multiplication started and continued until I log degree (from 106 to 107 cfu/ml). At concentrations of 5% and 2,5 % cells developed with nearly the same kinetics (from 106 to 8x107 cfu/ml and from 106 to 9x107 cfu/ml, respectively). Multiplication was more pronounced and nearly the same at concentrations of 1.2 % and 0.6 % (from106 to 2x108 cfu/ml). At a basic sugar concentration 30% total sugars bacterial multiplication did not occur, while at 20 % it was negligible, not measurable photometrically.

At minimal concentrations of F, G, S (between 1-0.1 %) bacterial cells were still able to multiply, producing organic acids from sugars.

Our study showed that E. amylovora requires only a small amount of sugars (0.1%) for multiplication (acid production) while high concentrations inhibit multiplication. There was a negative correlation between sugar content and cell density. The optimal range of sugar concentration was at about 1%.

Effect of "less frequent carbohydrates" to E. amylovora multiplication was also determined using the API 50 CH strip. We could provide information on utilization of 39 carbohydrates by the bacterium at different categories as follows: Not utilized-, Slowly and weakly utilized-, Slowly and completely utilized-, Quickly and completely utilized carbohydrates. We suppose that carbohydrates that belong to the latter two groups could play an important role as nectar components in promoting E. amylovora multiplication in the blossoms of pome fruit trees.

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Study of Erwinia amylovora colonization and migration on blossoms of susceptible and tolerant apple cultivars
Published May 10, 2004

The stigmata of detached flowers of susceptible and tolerant apple cultivars were inoculated with about 104 gfp labeled Erwinia amylovora . There were no apparent differences in the colonization, multiplication and survival of the bacteria on the stigmatic surface of the culivars. Bacteria were washed down to the hy...panthium surface 24 hours after inoculation. The visual symptoms of the infection were the discoloration and shrinkage of the floral parts. The gradual browning associated with the infection appeared first on the surface of the hypanthium followed by the discoloration of the style. The color of the filaments turned into brown only 120 hours after the inoculation. Bacterial cells were not detected in the tissues of the styles and filaments. The traits of the hypanthium surface are of prominent importance in the progression of the infection. The wrinkled surface, the convex shape of the outer epidermal cell walls with thin cuticle and the sunken stomata helped to preserve a water film for a longer period providing medium for the motility of the bacteria in the susceptible cultivar. Bacteria were restricted to small water droplets on the flat and waxy surface of the hypanthium of the tolerant cultivar and only a few were able to enter the tissues.

Large bacterium aggregations were detected in the intercellular spaces of the parenchyma of the susceptible cultivar 48 hours after the inoculation. In the next period the Erwinia amylovora cells gradually invaded the intercellulars of the hypanthium wall, the wall of the ovary and the pedicel. Low level of bacterium aggregation was found in the intercellulars of the tolerant cultivars. It is suggested that the progression of the infection was inhibited also by physiological factors.

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The effect of modified bacterial virulence to host-pathogen relationship (Phaseolus vulgaris L. Pseudomonas savastanoi pv. phaseolicola)
Published May 24, 1999

The Pseudomonas savastanoi pv. phaseolicola is one of the most expressive biogen stressors of the bean (Phaseolus vulgaris L.) in Hungary. The chemical and agrotechnological defence is inefficient, so breeding is the only workable way. The conventional cultivars are susceptible to PS while most of the new industrial varieties ...have genetic resistance to the pathogen. The genetic background of resistance is, however, a complex system in the bean. Leaf resistance is a monogenic system, but this gene is not expressed in juvenile stage of the host. The pathogen species can be divided into different races. After inoculation with virulent strains, typical symptoms appeared on the leaves. To understand the details of host-pathogen relationships, there were carried out experiments using bacterial strains with altered virulence. Six transposon mutants of the PS were tested. Our main objective was to test these modified bacterial strains on bean cultivars of known genetic background. First we analysed the symptoms, and then the correlation between the symptoms and the multiplication of mutant bacteria. Three cultivars (Cherokee, Inka and Főnix) were tested.

The infection by the virulent PS isolate produced typical symptoms on the three cultivars tested. Mutant bacteria (except strain 756) did not cause any significant symptoms on the hosts. The mutant 756 induced visible symptoms on the cultivars Cherokee and Inka. On Cherokee there were small watersoaked lesions, and HR (hypersensitivity reaction) was detected on Inka, but this was restricted to some cells only (mikro HR). The rate of multiplication of the wild type strain was much higher than the multiplication of the mutants. Bacteria were detected in the cotyledons and primordial leaf, but there is not any substantial number of bacteria in leaves, except for strains 757, 1212 and 1213. The rate of multiplication of strain 756 was intermediate. These, and other experiments can help to understand the genetic background of resistance and the host-pathogen relationship in the Pseudomonas-bean pathosystem.


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