Single- and mixed-species biofilm formation by Escherichia coli O157:H7 and Salmonella, and their sensitivity to levulinic acid plus sodium dodecyl sulfate

•3% LVA +2% SDS was effective against both single- and dual-species biofilms.•E. coli O157:H7 and Salmonella were antagonistic to each other in mixed biofilms.•Microbial behavior in biofilms can be studied using FP-labeled bacteria.

The development of single- and mixed-species biofilms formed by Escherichia coli O157:H7 and Salmonella was observed, and the antimicrobial effectiveness of levulinic acid (LVA) plus sodium dodecyl sulfate (SDS) on the cells in single- and dual-species biofilms was determined. Biofilm-forming ability of single- and mixed-species cultures was observed by crystal violet staining and their resistance to levulinic acid plus SDS was determined by enumeration. Fluorescent protein-labeled E. coli O157:H7 and Salmonella were constructed and the bacterial composition of the biofilms after treatment with levulinic acid plus SDS was visualized by confocal laser scanning microscopy (CLSM). E. coli O157:H7 and Salmonella were antagonistic to each other, being more sensitive to levulinic acid plus SDS in mixed-species biofilms. Images captured by CLSM revealed that E. coli O157:H7 and Salmonella were distributed evenly in the single- and dual-species biofilms, and confirmed that the combination of levulinic acid and SDS was effective in inactivating bacterial cells in biofilms. Results revealed that levulinic acid with SDS may be used as a potential biofilm control intervention.