This paper aims to investigate the biofilm formation and related genes expression of Escherichia coli isolated from pork. E. coli was isolated from pig slaughtering line, carcass surfaces and pork meat and identified using selective agar plates and specific polymerase chain reaction (PCR). After culture at 15 ℃ for 72 h, the biofilm forming abilities of the isolates were evaluated by means of microplate crystal violet staining. Furthermore, representative strains were selected to explore the changes of biofilm biomass, the composition of extracellular polymeric substances (EPS), and biofilm related-genes expression by means of reverse transcription real-time PCR in the process of biofilm formation. The results showed that a total of 31 strains of E. coli were isolated and identified in this study, among which 11 were obtained from pig slaughtering line, 4 from the surface of pig carcasses, and 16 from raw pork meat. The results of crystal violet staining showed that there were significant differences in the biofilm-forming ability among the 31 isolates, and 64.52% of them had weak biofilm-forming ability, one of which was then selected to study the biofilm formation process. The biofilm biomass of strain D_4-18 on the microplate increased continuously during 168 h incubation at 15 ℃. After incubation for 72 and 168 h, EPS were secreted by strain D_4-18, and the gene expression levels of papC、fimH、csgA genes were 0.095、0.933、0.435 copies/cm², respectively. With the increase of culture time, the contents of protein and polysaccharide in loose EPS (L-EPS) and the protein content in bound EPS (B-EPS) increased significantly (P < 0.01). After incubation for 168 h, the expression of the papC and fimH genes increased, while csgA gene expression did not change significantly, indicating that these genes play different regulatory roles in the biofilm formation of E. coli.

This study aimed to explore mixed-species biofilm formation (BF) between meat-derived Escherichia coli and Pseudomonas. The biofilm-forming capacity of isolates was evaluated by means of microplate crystal violet staining and the process of mixed-species biofilm formation between representative strains was investigated. The dynamic changes of biofilm amount, microstructure and the composition of extracellular polymeric substances (EPS) were examined. Absolute quantitative real-time PCR was used to analyze the difference in the expression of BF-related genes. The results showed that there were significant inter-strain differences in the biofilm-forming capacity of E. coli and Pseudomonas (P < 0.05), in which the strains with strong biofilm-forming capacity accounted for 25.81% and 23.08%, respectively. The results of plate counting showed that the BF processes of E. coli D_4-18 + Pseudomonas Y_2-2 10, and E. coli C_-13 + Pseudomonas Y_2-1 1 were different on stainless steel sheet, and the interaction between the two species changed with BF time. Similar changes in the microstructure were observed by confocal laser scanning microscope (CLSM). The secretion of protein and polysaccharide in EPS was inhibited during mixed-species BF; the amount of protein decreased by 31% and 48%, respectively, and the amount of polysaccharide decreased by 38% and 56%, respectively, at 72 and 168 h of culture compared with the total EPS of the two single-species biofilms. The results of gene expression per unit cell number showed that the expression of BF-related genes was significantly up-regulated in the biofilm compared with the planktonic cells (P < 0.05). Compared with the single-species biofilm, the papC and csgA genes of E. coli in the mixed biofilm were significantly up-regulated at 72 and 168 h (P < 0.01), while the phoR and cbrA genes of Pseudomonas were significantly down-regulated at 72 h (P < 0.01). The absolute amount of genes expression per unit area in mixed-species BF changed compared with that in single-species BF. This study can provide a scientific basis for understanding the mixed-species BF of meat-derived spoilage microorganisms and for the risk assessment and control of microbial contamination in meat production and processing.