Mapping the Efficacy and Mode of Action of Ethylzingerone [4-(3-Ethoxy-4-Hydroxyphenyl) Butan-2-One] as an Active Agent against Burkholderia Bacteria

Abstract

Burkholderia cepacia complex (Bcc) bacteria are intrinsically antimicrobial resistant opportunistic pathogens and key risk species in the contamination of non-food industrial products. New agents and formulations to prevent growth of Burkholderia in home care (cleaning agents) and personal care (cosmetics and toiletries) products are required. We characterised how ethylzingerone [4-(3-ethoxy-4-hydroxyphenyl) butan-2-one)] (HEPB) acts as a preservative with activity against Burkholderia species encountered in industry. Burkholderia (n = 58) and non-Burkholderia (n = 7) bacteria were screened for susceptibility to HEPB, and its mode of action/resistance determined for a model B. vietnamiensis strain using transposon mutagenesis, transcriptomics and genome resequencing analysis. The susceptibility of Burkholderia spp. to HEPB (MIC = 0.45 ±0.11 % w/v; MBC = 0.90 ±0.3 % w/v) was characterised, with limited inter-/intra-species differences. HEPB (1% w/v) was rapidly bactericidal producing a 6-Log reduction in viability within 4 hours. Spontaneous resistance to HEPB did not develop, but transient phenotypes with altered growth characteristics and susceptibility to antibiotics were identified after prolonged exposure to sub-lethal HEPB concentrations. Transposon mutagenesis and RNA-sequencing analysis identified multiple genetic pathways associated with HEPB exposure, including stress response mechanisms, altered permeability, regulation of intracellular pH, damage/repair of intracellular components and alteration/repair of lipopolysaccharides. Key pathways included the stringent response, homeostasis of intracellular pH by the kdp operon, protection against electrophiles by KefC, and repair of oxidised proteins by methionine sulfoxide reductase enzymes. In summary, we show that HEPB has potent, targeted efficacy against Burkholderia bacteria without promoting wider stable antimicrobial resistance. The mode of action of HEPB against Burkholderia is multifactorial but killing by intracellular oxidation is a key mechanism of this promising agent. Importance. Burkholderia bacteria are opportunistic pathogens that can overcome preservatives used in the manufacture of non-sterile industrial products, and occasionally cause contamination. Consequently, new preservatives to prevent the growth of key risk Burkholderia cepacia complex bacteria in non-food industrial products are urgently required. Here we show that ethylzingerone is active against these problematic bacteria, killing them via a multifactorial mode of action which involves intracellular oxidation.

Ethylated analogue of Zingerone: A new and eco‐respectful preservative in cosmetics

“…In addition, EZ has shown very significant activity against problematic bacteria such as Burkholderia spp., killing them via a multifactorial mode of action which involves intracellular oxidation [37].…”

“…Moreover, recent analyses have revealed that C. albicans phenotypic and transcriptomic responses to EZ treatment were distinguishable from those of two widely used preservatives, triclosan and methylparaben. Chemogenomic analyses have demonstrated that EZ antifungal activity strongly interfered with the biosynthesis of aromatic amino acids [36,37].…”