Quorum sensing asaI mutants affect spoilage phenotypes, motility, and biofilm formation in a marine fish isolate of Aeromonas salmonicida

Abstract

Microbial spoilage is associated with the regulation of quorum sensing (QS). A. salmonicida AE03 with QS mediated acylated homoserine lactones (AHLs) activity was isolated from spoiled large yellow croaker (Pseudosciaena crocea). In this study the activity and role of AHLs in spoilage phenotypes, motility and biofilm formation of AE03 were investigated. The strain AE03 could induce Chromobacterium violaceum CV026 to produce the violacein pigment both at 28 °C and 4 °C in a density-dependent manner. Five types of AHLs were detected in AE03 culture by LC-MS/MS analysis, and N-butanoyl-l-homoserine lactone (C4-HSL) was a major signal molecule, reaching the highest concentration when incubated for 30 h at 28 °C. An asaI-mutant, constructed by a suicide plasmid, failed to produce short chain AHLs signal. Compared with wild type (WT) strain, the production of trimethylamine (TMA), biogenic amino and protease significantly increased in asaI-mutant during the exponential and stationary phase, while the growth rate did not differ. Swimming motility in asaI-mutant was comparatively stronger than that of WT strain, whereas, asaI-mutant resulted in the decrease of maturing biofilm. Furthermore, supplementation of exogenous C4-HSL restored the production of spoilage metablites, protease and biofilm formation in mutant. In accordance with the effect of asaI deletion on the spoilage phenotypes and motility, asaI-mutant was showed to significantly up-regulate the transcript levels of torA, cadA and fliR, as well as asaR, indicating that C4-HSL could be involved in the modulation of the spoilage related enzymes and flagella. Indeed, the asaI-mutant promoted the spoilage progress of fish fillets stored at 4 °C, while exogenous C4-HSL repressed the sensory change and TVB-N accumulation. The present study highlighted that AsaI/C4-HSL was an important regulator in spoilage, motility and biofilm formation of A. salmonicida, and spoilage potential was under the negative control of AsaI/AsaR-type system.