Response of Various Cucurbits to Infection by Plasmid-Harboring Strains of Agrobacterium

Abstract

Tumor formation in cucurbit cultivars resulting from infection by various strains of Agrobacterium tumefaciens and Agrobacterium rhizogenes is environmentally affected. In all instances, tumors could be induced on excised cotyledons while inoculating attached cotyledons or stems resulted in no tumor formation. In addition, buttercup squash (Cucurbita maxima Duch. buttercup) was most susceptible to tumor formation, while butterbush squash (Cucurbita maxima Duch. butterbush) failed to form tumors when inoculated with any of the strains of Agrobacterium. Other tested cucurbit cultivars showed intermediate susceptibility to tumor induction by the various Agrobacterium strains.Dicotyledonous plants are susceptible to infection from two species of Gram-negative soil bacteria, Agrobacterium tumefaciens and Agrobacterium rhizogenes (2,12). The result of infection by these pathogens is a plant neoplasm, caused by the insertion of a segment of DNA, termed T-DNA, from the bacterium into the host genome (2). This T-DNA is contained on the Ti2 plasmid in A. tumefaciens and Ri plasmid in A. rhizogenes (2). These plasmids have been found to share DNA sequence homology (9), and are a requirement for bacterial virulence (16). In addition, genetically engineered forms of these plasmids have served as vehicles for gene transfer and expression in plants (7).The host range for most species ofAgrobacterium is very wide (4). Recently, however, it was reported that certain strains exhibit very high host specificity based primarily on the Ti plasmid, with a role also assigned to the chromosomal background ofthe strain (10). Further, Owens and Cress (13) Moore. In most cases, an overnight culture grown in L-media (10 g/L tryptone, 10 g/L NaCl, 5 g/L yeast extract, pH 7.5) was used as the inoculum. Occasionally, log phase cultures were used. In general, no qualitative differences were observed between these two inocula.Plants and Inoculations. Four varieties of squash (Cucurbita maxima Duch. buttercup, green hubbard, vegetable spaghetti, and butterbush) and one variety of pumpkin (Cucurbita maxima Duch. big max) were grown in a growth chamber (16 h photoperiod, 22C) in vermiculite irrigated with Hoagland solution (6). As positive controls, carrot (Daucus carota) root slices, tomato (Lycopersicum esculentum) stems, and Kalanchoe stems were used.Two protocols were attempted for tumor induction. Virulence was induced on whole plants and detached cotyledons as described by Owens and Cress (13). Stem and/or leaf wounds were made with a sterile toothpick. Approximately 10 ,ul of a bacterial suspension was spread over the wound site. Plant ages ranged form 7 to 21 d after planting. Detached cotyledons from approximately 8 d old plants were excised and surface sterilized in 10% commercial bleach for 15 min. After rinsing with sterile H20, cuts along both sides ofthe main vein were made, and inoculated with approximately 2.5 ,ul of the bacterial suspension. After incubating the cotyledons 3 d on water agar, they were transferred to Murash...

Factors Enhancing Virulence Response of Agrobacteria in Cucumber (<i>Cucumis sativus</i> L.)

“…This variation in virulence may be due to host range limitation. Analogous works on crown gall induction by various strains of A. tumefaciens were reported by other workers (Machado et al, 1997;Unger et al, 1985;Smarrelli et al, 1986). Different rates of crown gall induction obtained from the same explant of cucumber after inoculation with different A. tumefaciens strains suggests that the rates of tumorigenicity are dependent on the strains of Agrobacterium.…”

“…This variation in virulence may be due to host range limitation. Several workers reported hairy root induction by various strains of A. rhizogenes (Katavic et al, 1991;Smarrelli et al, 1986).…”

Hormone-Resistant Mutants of Arabidopsis Have an Attenuated Response to Agrobacterium Strains

“…Naturally occurring variation in susceptibility to Agrobacterium inoculation has been described in cucurbits (20), pea (16), soybean (14), grapevine (24), and interspecific hybrids ofNicotiana (1 1). In grapevine, resistance to crown gall disease seems to be inherited as a single gene, but in the other species differences in susceptibility are not inherited in a simple fashion.…”

Use of a Root Tumorigenesis Assay to Detect Genotypic Variation in Susceptibility of Thirty-four Cultivars of Pisum sativum to Crown Gall

“…Genetic variation in susceptibility to crown gall has been exploited to analyze resistance genes in grapevine, an economically important host of A. tumefaciens (31 (24), alfalfa (21), kalanchoe (3), castor bean (7), sunflower (4), soybean (15,27), and squash (29,33). The results indicate that genetic variation in response to A. tumefaciens is readily detectable in natural populations.…”