Interaction and cross-contamination potential of prepared beef steak isolates Pseudomonas weihenstephanensis and Macrococcus caseolyticus in biofilms of dual-species

This study evaluated the interactions between single or dual-species biofilms formed by dominant spoilage bacteria P. weihenstephanensis and M. caseolyticus isolated from refrigerated, spoilage prepared beef steaks at 4 °C and elucidated the interactive behavior of biofilm development in dual species. In addition, the relationship between biofilm formation capacity and cross-contamination was analyzed by simulating surface to food contact transfer. The results showed that the two species exhibited synergism as biofilms developed, which was the main mode of interaction observed. Under aerobic conditions, Pseudomonas weihenstephanensis and Macrococcus caseolyticus co-cultured for 96 h showed obvious biofilm formation ability, resulting in greater cross-contamination. Scanning electron microscopy and Confocal laser scanning microscopy showed the formation of flattened dense biofilms in the co-culture. The significant increase in Fe content and decrease in siderophore content of the dual-species biofilm as determined by ICP-MS was attributed to respiratory inhibition resulting in a decrease in the transcription of genes regulating the two-component regulatory system of Macrococcus tyrolyticus SrrAB and an increase in the expression of cytoplasmic hydrolase leading to the rupture of the release of hemoglobin to provide a source of iron for P. weihenstephanensis. The increase of heme content in the supernatant of dual-species and the results of RT-qPCR showed that the gene expression of the heme transport system of P. weihenstephanensis was significantly up-regulated and the siderophore gene expression was decreased, which further revealed that P. weihenstephanensis preferentially uses the heme uptake system to take up the iron source provided by M. caseolyticus for P. weihenstephanensis. Overall, our results provide insight into the complex dynamics of biofilms formed by P. weihenstephanensis and M. caseolyticus, emphasizing that the iron reaction pathway may be a key factor influencing the growth of P. weihenstephanensis biofilms, and that these results will provide a theoretical basis for the control of spoilage of refrigerated foods.