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Effects of alternative sweeteners on lactic acid bacteria growth and the antibacterial mechanisms of their cell-free supernatants: Transcriptomic insights into Escherichia coli O157:H7 inhibition

ABSTRACT: The use of probiotics in the food industry is increasing; however, the application of strong antibiotics or preservatives to maintain strain functionality is limited, which may compromise product safety. This study aimed to evaluate the effects of alternative sweeteners on the growth and antimicrobial activity of lactic acid bacteria used in this study (Lactobacillus acidophilus, Lacticaseibacillus casei, and Lactiplantibacillus plantarum) and to elucidate their antimicrobial mechanisms against pathogens, thereby exploring the potential application of sweeteners in probiotic-based foods. L. casei exhibited stable growth with aspartame (ASP), acesulfame potassium (ACE-K), erythritol (ERY), sucrose (SUC), and saccharin (SAC). Among them, the cell-free supernatant (CFS) of L. casei cultured with 1% (w/v) SAC showed significant antimicrobial activity against Escherichia coli O157:H7 and Salmonella Typhimurium. SAC alone inhibited pathogen growth, and the effect was further enhanced in co-culture with L. casei. Transcriptome analysis revealed that SAC and L. casei exposure upregulated genes related to sulfur metabolism, cationic antimicrobial peptide resistance, and two-component systems, while downregulating genes associated with flagellar assembly, chemotaxis, and biofilm formation. These findings indicate that SAC and L. casei trigger stress responses in pathogens while suppressing motility and virulence. Phenotypic assays of motility and biofilm formation supported the transcriptome data, confirming that SAC-induced gene expression changes translated into observable phenotypes. Importantly, SAC did not affect the growth or antimicrobial activity of L. casei but selectively inhibited E. coli O157:H7 under co-culture conditions. Overall, SAC shows promise as an antimicrobial agent for controlling foodborne pathogens in the food industry.

ORGANISM(S): Escherichia coli O157:H7

PROVIDER: GSE295793 | GEO | 2026/03/31

REPOSITORIES: GEO

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Project description:Pathogenic biofilms have been associated with persistent infections due to high resistance to antimicrobial agents while commensal biofilms often fortify host immune system. Hence, controlling biofilm formation of both pathogenic bacteria and commensal bacteria is important in bacteria-related diseases. We investigated the effect of plant flavonoids on biofilm formation of both enterohemorrhagic Escherichia coli O157:H7 and three commensal E. coli K-12 strains. Phloretin abundant in apples markedly reduced E. coli O157:H7 biofilm formation without affecting the growth of planktonic cells while phloretin did not harm commensal E. coli K-12 biofilms. Also, phloretin reduced E. coli O157:H7 attachment to human colon epithelial cells. Global transcriptome analyses revealed that phloretin repressed toxin genes (hlyE and stx2), autoinducer-2 importer genes (lsrACDBF), a curli gene (csgA), and a dozens of prophage genes in E. coli O157:H7 cells. Electron microscopy confirmed that phroretin reduced the curli production in E. coli O157:H7. In addition, phloretin suppressed TNF-Î±-induced inflammatory response in vitro using human colonic epithelial cells. Moreover, in the trinitrobenzene sulfonic acid (TNBS)-induced rat colitis model, phloretin significantly ameliorated colon inflammation and body weight loss. Taken together, our results suggest that phloretin may act as an inhibitor of E. coli O157:H7 biofilm formation as well as anti-inflammatory agent on inflammatory bowel diseases while leaving beneficial commensal E. coli biofilm intact. For the microarray experiments, E. coli O157:H7 EDL933 was inoculated in 25 ml of LB in 250 ml flasks with overnight cultures that were diluted at 1:100. Cells were shaken at 100 rpm and 37Â°C for 7 hrs. Cells were immediately chilled with dry ice and 95% ethanol (to prevent RNA degradation) for 30 sec before centrifugation in 50 ml centrifuge tubes at 13,000 g for 2 min; cell pellets were frozen immediately with dry ice and stored -80Â°C. RNA was isolated using Qiagen RNeasy mini Kit (Valencia, CA, USA). To eliminate DNA contamination, Qiagen RNase-free DNase I was used to digest DNA. RNA quality was assessed by Agilent 2100 bioanalyser using the RNA 6000 Nano Chip (Agilent Technologies, Amstelveen, The Netherlands), and quantity was determined by ND-1000 Spectrophotometer (NanoDrop Technologies, Inc., DE, USA).

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Effects of alternative sweeteners on lactic acid bacteria growth and the antibacterial mechanisms of their cell-free supernatants: Transcriptomic insights into Escherichia coli O157:H7 inhibition

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