The involvement of bacterial quorum sensing in the spoilage of refrigerated Litopenaeus vannamei

Quorum-sensing signals in refrigerated shrimp (Litopenaeus vannamei) undergoing spoilage were examined using bioreporter assays, thin-layer chromatography and gas chromatography-mass spectrometry, and the results revealed the presence of three types of autoinducers including acetylated homoserine lactones (AHLs) (i.e., N-hexanoyl-homoserine lactone, N-oxohexanoyl-homoserine lactone and N-octanoyl-homoserine lactone), autoinducer-2, and cyclic dipeptides (i.e., cyclo-(l-Pro-l-Leu), cyclo-(l-Leu-l-Leu) and cyclo-(l-Pro-l-Phe)). Autoinducer-2, rather than any AHL, was detected in extracts from pure cultures of the specific spoilage organisms (SSO), i.e., Shewanella putrefaciens (SS01) and Shewanella baltica (SA02). As for the cyclic peptides, only SA02 was determined to produce cyclo-(l-Pro-l-Leu). According to the transcription levels of LuxR (the master quorum-sensing regulator) in the SSO in response to exogenous autoinducers, the SSO could sense AHLs and cyclo-(l-Leu-l-Leu), rather than autoinducer-2, cyclo-(l-Leu-l-Leu) and cyclo-(l-Pro-l-Phe). In accordance with the results of LuxR expression, the production of biofilm matrixes and extracellular proteases in the SSO was regulated by exogenous AHLs and cyclo-(l-Pro-l-Leu), rather than 4,5-dihydroxy-2,3-pentanedione (the autoinducer-2 precursor), cyclo-(l-Leu-l-Leu) and cyclo-(l-Pro-l-Phe). Exogenous N-hexanoyl-homoserine lactone and cyclo-(l-Pro-l-Leu) increased the growth rates and population percentages of the SSO in shrimp samples under refrigerated storage, and interestingly, exogenous 4,5-dihydroxy-2,3-pentanedione also increased the population percentages of the SSO in vivo by inhibiting the growth of the competing bacteria. However, according to the levels of TVB-N and the volatile organic components in the shrimp samples, exogenous 4,5-dihydroxy-2,3-pentanedione did not accelerate the shrimp spoilage process as N-hexanoyl-homoserine lactone and cyclo-(l-Pro-l-Leu) did. In summary, our results suggest that quorum sensing involves the spoilage of refrigerated Litopenaeus vannamei.