SUMMARY: Sixty-nine Agrobacterium strains, including representatives of six current species, were tested: for colony formation on sucrose + salts medium, 3-ketoglycoside production, response on litmus milk medium, infectivity on carrot-root discs, infectivity and rate of tumour appearance on primary pinto bean leaves. These tests separated the majority of the strains into species. Forty-four strains were tumourigenic on pinto bean leaves, ten of which did not form colonies on a sucrose + salts medium and nine of these showed a rate of tumour appearance typical of auxotrophs. Glutamate was required for colony formation by seven of the ten auxotrophs, two required only pantothenate and one strain required either glutamate or pantothenate. The two strains among the ten auxotrophs required nicotinic acid + pantothenate + glutamate. Strains that required either pantothenate or pantothenate + nicotinic acid for colony formation also required these compounds in liquid medium. None of the strains which required glutamate for growth on plates required it in liquid medium though glutamate shortened lag and division times. The growth requirements of the auxotrophs did not obviously correlate with their specific infectivity. Six of the ten auxotrophs produced no 3-ketoglycosides, but this response did not correlate with infectivity or nutritional requirements. Three auxotrophs initiated more tumours when appropriate growth factors were added to inoculated leaves. The agrobacteria that induced pinto bean leaf tumours fell into four groups: I, strains which infected all wound sites with equal efficiency regardless of wound size; II, strains which infected all but the smallest wound sites with equal efficiency; III, phototrophic strains which infected only larger wound sites; IV, auxotrophic strains which, in the typical examples, infected only the larger wound sites. Nineteen of the tumourigenic strains (43%) were restricted in their ability to infect small wound sites and were assigned to groups III and IV. Because loss of the ability to initiate tumours at small wound sites is due to nutritional limitations imposed on the infecting bacteria by the wound site medium, these group III and IV strains were nutritionally limited in their ability to infect this host. The large proportion of strains which showed this defect indicates that variation in the ability of tumefacient strains to adapt to the wound medium supplied by a host is quite common and hence represents one major pathway by which pathogenicity within the genus may have evolved.


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