No abstract

and are among the predominant spoilage bacteria in refrigerated shrimp (). are incapable of producing acyl-homoserine lactone (AHL) quorum sensing signals, but can respond to environmental AHLs. In this paper, was found to produce three AHLs, i.e. -butanoyl-l-homoserine lactone (C4-HSL),-(3-oxohexanoyl)-l-homoserine lactone (O-C6-HSL) and -(3-oxooctanoyl)-l-homoserine lactone (O-C8-HSL), according to thin-layer chromatography using the bioreporter A136. The agar diffusion and β-galactosidase assays revealed that could eavesdrop on these three AHLs from. Eavesdropping on AHLs especially C4-HSL was found to boost the growth of particularly under nutrient limiting conditions (up to 40-fold increase) in the co-culture experiments. The azocasein assay revealed that produced fourfold more extracellular proteases in response to AHLs. As demonstrated by the biofilm crystal violet staining assay and confocal laser scanning microscopy, eavesdropping also decreased the biofilm-forming capacity of . By inoculation of and onto surface-sterilized shrimp, eavesdropping was found to endow a growth advantage to in vivo, resulting in a 0.5 day shortened shelf life of shrimp according to total volatile basic nitrogen levels and sensory analysis. Overall, the AHL-dependent eavesdropping increased the spoilage potential of , providing a fresh perspective on the spoilage process of refrigerated, and this may inspire the development of novel preservation techniques in the future to further reduce post-harvest loss of shrimp.

Numerous bacterial species utilize quorum sensing molecules acyl-homoserine-lactones (AHLs) to communicate, however, crosstalk often complicates the dynamics and behaviors of mixed populations. In this study, we developed a mutant of wild type SA03 (WT SA03), and aimed to investigate the role of LuxR (AHLs receptor) involved in the spoilage of refrigerated shrimp () by inoculating WT SA03 and mutant of SA03 ( SA03), respectively. The results indicated the maximum growth rate of total viable bacteria in shrimp inoculated with SA03 was 73.34% lower than that of WT SA03. The lag time of total bacteria in shrimp treated with SA03 were 87.6 h, significantly longer than that of WT SA03. Meanwhile, the total volatile basic nitrogen concentrations of shrimp treated with WT SA03 were significantly higher than that of SA03 after 2 days of storage, which were in agreement with the decrease of the content of AHLs of the shrimp. The results indicated might utilize AHLs produced by other bacteria and accelerate the shrimp spoilage process through LuxR receptor system.

The spoilage potential of Shewanella putrefaciens and S. baltica isolated from spoiled refrigerated Litopenaeus vannamei was evaluated by in vitro assays for trimethylamine oxide reduction, extracellular hydrolytic enzymes and biofilm formation, and in vivo inoculation into surface-sterilised shrimp followed by microbial, biochemical and sensory analyses during storage for 5 days at 4°C. S. baltica displayed higher spoilage potential than S. putrefaciens both in vitro and in vivo. Shrimp co-inoculated with them had one-day shorter shelf life than those mono-inoculated, based on the results of bacterial density, volatile base nitrogen, trimethylamine, volatile organic compounds and sensory analysis, which strongly suggests cooperation of Shewanella species in shrimp spoilage. Exogenous cyclo-(L-Pro-L-Leu) boosted bacterial growth, extracellular protease and collagenase activities, and biofilm formation of S. putrefaciens and S. baltica at least before they entered the stationary phase, indicating that cyclo-(L-Pro-L-Leu)-dependent quorum sensing, a recently suggested communication mechanism between them, contributes to the cooperation. ). We previously discovered three types of autoinducers, i.e. AHLs, AI-2 and DKPs, in refrigerated Litopenaeus vannamei; nevertheless, QS circuits based on AHLs and AI-2 for the SSO (i.e. S. baltica and S. putrefaciens) have been found to be incomplete (Zhu et al., 2015). The SSO can sense AHLs, but are incapable of producing these molecules. The exact reverse is the case for AI-2. Cyclo-(L-Pro-L-Leu), the most abundant DKP detected in refrigerated shrimp, can be secreted by S. baltica, and responded by both S. baltica and S. putrefaciens in quite distinct ways. Cyclo-(L-Pro-L-Leu)-dependent QS thus seems to be the interspecies communication mechanism between S. baltica and S. putrefaciens in refrigerated Litopenaeus vannamei. The current research evaluated cooperation of the SSO in shrimp spoilage by inoculating S. putrefaciens and S. baltica into surface-sterilised shrimp, and to examine contribution of QS-based communication to the cooperation, the effects of exogenous cyclo-(L-Pro-L-Leu) on the spoilage potential of S. putrefaciens and S. baltica were also determined. Materials and methodsCooperation and QS in shrimp spoilage S. Zhu et al. Figure 1 (a) TVB-N and (b) TMA levels in shrimp with or without inoculation.S. putrefaciens + S. balticarepresents co-inoculationat the CFU ratio of 1. The results are presented as the mean AE standard deviation (n = 3). Different letters indicate significant differences in mean values (P < 0.05).