The Phytoalexin-Inducible Multidrug Efflux Pump AcrAB Contributes to Virulence in the Fire Blight Pathogen, Erwinia amylovora

Abstract

The enterobacterium Erwinia amylovora causes fire blight on members of the family Rosaceae, with economic importance on apple and pear. During pathogenesis, the bacterium is exposed to a variety of plant-borne antimicrobial compounds. In plants of Rosaceae, many constitutively synthesized isoflavonoids affecting microorganisms were identified. Bacterial multidrug efflux transporters which mediate resistance toward structurally unrelated compounds might confer tolerance to these phytoalexins. To prove this hypothesis, we cloned the acrAB locus from E. amylovora encoding a resistance nodulation division-type transport system. In Escherichia coli, AcrAB of E. amylovora conferred resistance to hydrophobic and amphiphilic toxins. An acrB-deficient E. amylovora mutant was impaired in virulence on apple rootstock MM 106. Furthermore, it was susceptible toward extracts of leaves of MM 106 as well as to the apple phytoalexins phloretin, naringenin, quercetin, and (+)-catechin. The expression of acrAB was determined using the promoterless reporter gene egfp. The acrAB operon was up-regulated in vitro by the addition of phloretin and naringenin. The promoter activity of acrR, encoding a regulatory protein involved in acrAB expression, was increased by naringenin. In planta, an induction of acrAB was proved by confocal laser scanning microscopy. Our results strongly suggest that the AcrAB transport system plays an important role as a protein complex required for virulence of E. amylovora in resistance toward apple phytoalexins and that it is required for successful colonization of a host plant.

Effector-Repressor Interactions, Binding of a Single Effector Molecule to the Operator-bound TtgR Homodimer Mediates Derepression

“…Therefore, the TtgABC efflux pump may be a critical element in the competitive colonization of plant roots by P. putida, similarly to other MDR transporters found in soil-and plant-associated bacteria (28 -30). In addition, the fact that a mutant E. amylovora deficient in the MDR efflux pump AcrAB (60% identical to TtgAB) was impaired in its virulence against plants (27), emphasizes our hypothesis.…”

“…However, the particularly broad substrate specificity of this efflux system and the multidrug binding capacity of its cognate repressor TtgR raised the question concerning their physiological role(s). Based on the fact that the plant rhizosphere is one of the natural habitats of P. putida, and considering that the AcrB transporter from the plant pathogen Erwinia amylovora, involved in the resistance of this strain to various plant defense metabolites (27), is 60% identical to the TtgB transporter from P. putida DOT-T1E, we investigated the potential role of TtgABC in the resistance to plant-derived antimicrobials. To verify this hypothesis, the MICs of five plant secondary metabolites known to have antibacterial activities (45)(46)(47) were determined for both, P. putida DOT-T1E and its derivative mutant strain DOT-T1E18, deficient in the TtgABC efflux pump (Table 2).…”

“…Some MDR efflux pumps present in soil-living or plant-associated bacteria have been shown to confer resistance to plant defense molecules and seem to play an important role in the efficient colonization of plant-influenced microenvironments by these microorganisms and/or in their virulence, in the case of plant pathogens (27)(28)(29)(30).…”

Floral traits affecting fire blight infection and management

“…The antibiotic activity of their respective aglucones, phloretin and hydroquinone, against E. amylovora has long been demonstrated (Römmelt et al 2003b;Pontais et al 2008). Burse et al (2004) identified a multidrug efflux pump in E. amylovora, which confers resistance to apple phenolics like phloretin, naringenin, quercetin and catechin, contributing to bacterial virulence, which seems to confirm the role of phenolic compounds during the infection process (Pontais et al 2008). Phloretin was detected in all the hypanthium samples of various apple and pear cultivars, its levels in apple samples being approximately three times higher than in pear samples (Horváth et al 2004a).…”

Detection of potential AcrAB-TolC multidrug efflux pump inhibitor in calyces extract of Hibiscus sabdariffa

“…In E. coli, AcrAB-TolC mediate resistance toward metabolic byproducts such as bile salt, environmental antimicrobials, toxins, dyes, and detergents [5]. AcrAB-TolC of E. amylovora plays an important role in resistance toward phytoalexins, as virulence and fitness factors that are required for successful colonization of a host plant [6,7].…”