Dual-Species Biofilms Formed by Escherichia coli and Salmonella Enhance Chlorine Tolerance.

In this research, mono- and dual-species biofilms of Escherichia coli (O45:H2 and O121:H19) and Salmonella enterica serovar Typhimurium formed on stainless-steel coupons were treated with 100 mg/L NaClO for 1 min. Confocal laser scanning microscopy (CLSM) was applied to investigate the spatial structural dynamics of mono- and dual-species biofilms, and nuclear magnetic resonance (NMR) spectroscopy was employed to further investigate their metabolic responses toward chlorine. CLSM results indicated that mixed-species biofilms (total biovolume, 148,000 to 167,000 µm3) stimulated the growth of biomass 2 to 6 times that of single-species biofilms. Upon chlorine treatment, E. coli O45 and S. Typhimurium achieved less reduction (P < 0.05) when coexisting in mixed biofilms (0.70 and 1.17 log CFU/coupon reductions, respectively) compared with their corresponding single-species biofilms (1.97 and 2.01 log CFU/coupon reductions, respectively), while for E. coli O121, more reduction (P < 0.05) was achieved in a mixed biofilm (1.37 log CFU/coupon reductions) compared with its single-species biofilm (0.59 log CFU/coupon reductions). Moreover, NMR results suggested that the increase of putrescine (antioxidation regulator) and the decrease of glucose (enhanced glycolysis for energy replenishment) might contribute to the improved chlorine tolerance in mixed biofilms. Overall, dual-species biofilms promoted biofilm growth and their chlorine tolerance. This study improved our knowledge of the metabolic difference of single- and mixed-species biofilms of E. coli and Salmonella to chlorine sanitization and raised an urgency to investigate the effectiveness of common disinfectants against multispecies consortia. IMPORTANCE Outbreaks of Escherichia coli and Salmonella in food might be associated with the cross-contamination of biofilms on food-contact surfaces. The knowledge of the sanitization of mono-species biofilm on the food-contact surface is well established, while mixed-species biofilm occurs more naturally, which could profoundly affect the efficacy of sanitizer. Therefore, this research aims to evaluate the efficacy of using chlorine against single- and dual-species biofilms of E. coli and Salmonella along with the underlying bacterial metabolic responses. The responses of a mixed biofilm of E. coli and Salmonella to chlorine sanitization were clarified, providing insights to develop a targeted and green sanitization strategy against specific pathogens by perturbing their most susceptible metabolism pathway without sanitizer residue.

Effect of chlorine sanitizer on metabolic responses of Escherichia coli biofilms "big six" during cross-contamination from abiotic surface to sponge cake.

The effect of chlorine on Escherichia coli biofilm O157:H7 are well established; however, the effect on biofilm adhesion to food as well as the six emerging E. coli serotypes ("big six") have not been fully understood. Chlorine sanitization with 1-min 100 mg/L was applied against seven pathogenic E. coli (O111, O121:H19, O45:H2, O26:H11, O103:H11, O145, and O157:H7) biofilms on high-density polyethylene (HDPE) and stainless steel (SS) coupons, respectively. Using sponge cake as a food model, the adhesion behavior was evaluated by comparison of bacteria transfer rate before and after treatment. Besides, the metabolic profiles of biofilms were analyzed by nuclear magnetic resonance (NMR) spectrometer. A significant decrease in transfer rate (79% decline on SS and 33% decline on HDPE) was recorded as well as the distinctive pattern between SS and HDPE coupons was also noticed, with a low population (6-7 log CFU/coupon) attached and low survivals (0-3 log CFU/coupon) upon chlorine on SS, while high population (7-8 log CFU/coupon) attached and high survivals (5-7 log CFU/coupon) on HDPE. Moreover, O121:H19 and O26:H11 demonstrated the highest resistance to chlorine with the least metabolic status and pathways affected. O103:H11, O145, and O111 followed similar metabolic patterns on both surfaces. Distinct metabolic patterns were found in O45:H2 and O157:H7, where the former had more affected metabolic status and pathways on SS but less on HDPE, whereas the latter showed an opposite trend. Overall, a potential contamination source of STEC infection in flour products was demonstrated and metabolic changes induced by chlorine were revealed by NMR-based metabolomics, which provides insights to avoid "big six" biofilms contamination in food.

Porous chitosan derivative scaffolds affect proliferation and osteogenesis of mesenchymal stem cell via reducing intracellular ROS.

The proliferation and osteogenic differentiation of mesenchymal stem cells (MSCs) needed to be promoted for their use in stem cell-based therapy for large bone defects. This study aimed to prepare porous 2a-2 g scaffolds with antioxidant activity to reduce intracellular reactive oxygen species (ROS), which resulted in promoting the proliferation and osteogenic differentiation of MSCs. A series of novel chemically modified 2a-2 g scaffolds were fabricated by an acid-soluble/alkali-insoluble method. Besides, these 2a-2 g scaffolds had good biocompatibility, physicochemical properties and the ability to promote osteogenic differentiation of human adipose-derived stem cells (hADSCs). However, the proliferation ability of hADSCs on 2a-2f scaffolds was weakened. Interestingly, 2 g scaffold had a positive effect on hADSCs proliferation. These results indicated that the reduction of intracellular ROS was not conducive to hADSCs proliferation but beneficial to hADSCs osteogenic differentiation. Taken together, these significant results highlighted potential therapeutic benefit of 2 g scaffold in large bone defects.