Hypersensitive response and acyl-homoserine lactone production of the fire blight antagonists Erwinia tasmaniensis and Erwinia billingiae

Abstract

Fire blight caused by the Gram-negative bacterium Erwinia amylovora can be controlled by antagonistic microorganisms. We characterized epiphytic bacteria isolated from healthy apple and pear trees in Australia, named Erwinia tasmaniensis, and the epiphytic bacterium Erwinia billingiae from England for physiological properties, interaction with plants and interference with growth of E. amylovora. They reduced symptom formation by the fire blight pathogen on immature pears and the colonization of apple flowers. In contrast to E. billingiae, E. tasmaniensis strains induced a hypersensitive response in tobacco leaves and synthesized levan in the presence of sucrose. With consensus primers deduced from lsc as well as hrpL, hrcC and hrcR of the hrp region of E. amylovora and of related bacteria, these genes were successfully amplified from E. tasmaniensis DNA and alignment of the encoded proteins to other Erwinia species supported a role for environmental fitness of the epiphytic bacterium. Unlike E. tasmaniensis, the epiphytic bacterium E. billingiae produced an acyl-homoserine lactone for bacterial cell-to-cell communication. Their competition with the growth of E. amylovora may be involved in controlling fire blight.

Figure 1

Antagonistic effects of E. billingiae, E. tasmaniensis on pear slices and apple flowers inoculated with E. amylovora.
A. Symptoms after treatment of pear slices with Eb660, Eb661, FLA03, Et1/99 and Et2/99. One slice from a set of four similar slices with the same treatment is shown.
B. Fire blight symptoms on apple flowers sprayed with Eb660, Eb661, Et1/99 and Et2/99, and then inoculated with 500 cells of E. amylovora Ea1/79Sm. Control was treatment with water before inoculation with E. amylovora (three flowers top right). Evaluation for growth of Ea1/79Sm is given in Table 1. The experiments were repeated at least twice producing similar results.

Figure 2

Sequence comparison of parts from lsc genes of the E. tasmaniensis strains Et1/99, Et2/99 and Et4/99 with lsc of E. amylovora strain Ea1/79.

Figure 3

Induction of HR on tobacco leaves. Ea1/79, Et1/99, Et2/99 and Et4/99 were cultured in inducing medium and produced local necrotic HR lesions (right) in a leaf of cultivar ‘SR1’. No hypersensitive response was found for E. billingiae Eb661 or with the E. tasmaniensis strains Et1/99, Et2/99 and Et4/99 without conditioning in IM (left).

Figure 4

Alignment of partial amino acid sequences HrcC, HrcR and HrpL from E. tasmaniensis, E. amylovora and E. pyrifoliae strains. HrcC (187 aa): E. tasmaniensis Et2/99, E. amylovora Ea1/79 and E. pyrifoliae Ep4/96; HrcR (146 aa): same but Et4/99; HrpL (182 aa): same but Et1/99.

Figure 5

AHL assays with C. violaceum CV026 as a sensor (vertical streak). a, upper streak; b, lower streak; 1a, E. persicina; 1b, E. rhapontici; 2a, E. billingiae Eb1261; 2b, E. billingiae Eb661; 3a, P. stewartii DC283; 3b, D. chrysanthemi 3937; 4a, E. billingiae Eb660; 4b, E. mallotivora; 5a, E. amylovora Ea1/79; 5b, E. pyrifoliae Ep1/96.