Single- and Dual-Species Biofilm Formation by Shiga Toxin-Producing Escherichia coli and Salmonella, and Their Susceptibility to an Engineered Peptide WK2

Shiga toxin-producing Escherichia coli (STEC) and Salmonella enterica are important foodborne pathogens capable of forming both single- and multi-species biofilms. In this study, the mono- and dual-species biofilms were formed by STEC O113:H21 and Salmonella enterica serovar Choleraesuis 10708 on stainless steel in the presence of beef juice over 5 d at 22 °C. The dual-species biofilm mass was substantially (p < 0.05) greater than that produced by STEC O113:H21 or S. Choleraesuis 10708 alone. However, numbers (CFU/mL) of S. Choleraesuis 10708 or STEC O113:H21 cells in the dual-species biofilm were (p < 0.05) lower than their respective counts in single-species biofilms. In multi-species biofilms, the sensitivity of S. Choleraesuis 10708 to the antimicrobial peptide WK2 was reduced, but it was increased for STEC O113:H21. Visualization of the temporal and spatial development of dual-species biofilms using florescent protein labeling confirmed that WK2 reduced cell numbers within biofilms. Collectively, our results highlight the potential risk of cross-contamination by multi-species biofilms to food safety and suggest that WK2 may be developed as a novel antimicrobial or sanitizer for the control of biofilms on stainless steel.

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Activity of Bacteriophage and Complex Tannins against Biofilm-Forming Shiga Toxin-Producing Escherichia coli from Canada and South Africa

Antibiotics ◽

10.3390/antibiotics9050257 ◽

2020 ◽

Vol 9 (5) ◽

pp. 257

Author(s):

Emmanuel W. Bumunang ◽

Collins N. Ateba ◽

Kim Stanford ◽

Yan D. Niu ◽

Y. Wang ◽

...

Keyword(s):

Escherichia Coli ◽

South Africa ◽

Stainless Steel ◽

Foodborne Pathogens ◽

Shiga Toxin ◽

Biofilm Formation ◽

Condensed Tannin ◽

Plant Secondary Metabolites ◽

Brown Seaweed ◽

E Coli

Bacteriophages, natural killers of bacteria, and plant secondary metabolites, such as condensed tannins, are potential agents for the control of foodborne pathogens. The first objective of this study evaluated the efficacy of a bacteriophage SA21RB in reducing pre-formed biofilms on stainless-steel produced by two Shiga toxin-producing Escherichia coli (STEC) strains, one from South Africa and the other from Canada. The second objective examined the anti-bacterial and anti-biofilm activity of condensed tannin (CT) from purple prairie clover and phlorotannins (PT) from brown seaweed against these strains. For 24-h-old biofilms, (O113:H21; 6.2 log10 colony-forming units per square centimeter (CFU/cm2) and O154:H10; 5.4 log10 CFU/cm2), 3 h of exposure to phage (1013 plaque-forming units per milliliter (PFU/mL)) reduced (p ≤ 0.05) the number of viable cells attached to stainless-steel coupons by 2.5 and 2.1 log10 CFU/cm2 for O113:H21 and O154:H10, respectively. However, as biofilms matured, the ability of phage to control biofilm formation declined. In biofilms formed for 72 h (O113:H21; 5.4 log10 CFU/cm2 and O154:H10; 7 log10 CFU/cm2), reductions after the same duration of phage treatment were only 0.9 and 1.3 log10 CFU/cm2 for O113:H21 and O154:H10, respectively. Initial screening of CT and PT for anti-bacterial activity by a microplate assay indicated that both STEC strains were less sensitive (p ≤ 0.05) to CT than PT over a concentration range of 25–400 µg/mL. Based on the lower activity of CT (25–400 µg/mL), they were not further examined. Accordingly, PT (50 µg/mL) inhibited (p ≤ 0.05) biofilm formation for up to 24 h of incubation at 22 °C, but this inhibition progressively declined over 72 h for both O154:H10 and O113:H21. Scanning electron microscopy revealed that both SA21RB and PT eliminated 24 h biofilms, but that both strains were able to adhere and form biofilms on stainless-steel coupons at longer incubation times. These findings revealed that phage SA21RB is more effective at disrupting 24 than 72 h biofilms and that PT were able to inhibit biofilm formation of both E. coli O154:H10 and O113:H21 for up to 24 h.

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Effects of Beef Juice on Biofilm Formation by Shiga Toxin–Producing Escherichia coli on Stainless Steel

Foodborne Pathogens and Disease ◽

10.1089/fpd.2019.2716 ◽

2020 ◽

Vol 17 (4) ◽

pp. 235-242 ◽

Cited By ~ 2

Author(s):

Zhi Ma ◽

Kim Stanford ◽

Xiao M. Bie ◽

Yan D. Niu ◽

Tim A. McAllister

Keyword(s):

Escherichia Coli ◽

Stainless Steel ◽

Shiga Toxin ◽

Biofilm Formation

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Mixed Biofilm Formation by Shiga Toxin–Producing Escherichia coli and Salmonella enterica Serovar Typhimurium Enhanced Bacterial Resistance to Sanitization due to Extracellular Polymeric Substances†

Journal of Food Protection ◽

10.4315/0362-028x.jfp-13-077 ◽

2013 ◽

Vol 76 (9) ◽

pp. 1513-1522 ◽

Cited By ~ 48

Author(s):

RONG WANG ◽

NORASAK KALCHAYANAND ◽

JOHN W. SCHMIDT ◽

DAYNA M. HARHAY

Keyword(s):

Escherichia Coli ◽

Food Safety ◽

Foodborne Pathogens ◽

Shiga Toxin ◽

Biofilm Formation ◽

Extracellular Polymeric Substances ◽

Bacterial Resistance ◽

Salmonella Enterica Serovar Typhimurium ◽

E Coli ◽

Serovar Typhimurium

Shiga toxin–producing Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium are important foodborne pathogens capable of forming single-species biofilms or coexisting in multispecies biofilm communities. Bacterial biofilm cells are usually more resistant to sanitization than their planktonic counterparts, so these foodborne pathogens in biofilms pose a serious food safety concern. We investigated how the coexistence of E. coli O157:H7 and Salmonella Typhimurium strains would affect bacterial planktonic growth competition and mixed biofilm composition. Furthermore, we also investigated how mixed biofilm formation would affect bacterial resistance to common sanitizers. Salmonella Typhimurium strains were able to outcompete E. coli strains in the planktonic growth phase; however, mixed biofilm development was highly dependent upon companion strain properties in terms of the expression of bacterial extracellular polymeric substances (EPS), including curli fimbriae and exopolysaccharide cellulose. The EPS-producing strains with higher biofilm-forming abilities were able to establish themselves in mixed biofilms more efficiently. In comparison to single-strain biofilms, Salmonella or E. coli strains with negative EPS expression obtained significantly enhanced resistance to sanitization by forming mixed biofilms with an EPS-producing companion strain of the other species. These observations indicate that the bacterial EPS components not only enhance the sanitizer resistance of the EPS-producing strains but also render protections to their companion strains, regardless of species, in mixed biofilms. Our study highlights the potential risk of cross-contamination by multispecies biofilms in food safety and the need for increased attention to proper sanitization practices in food processing facilities.

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Ralstonia insidiosa serves as bridges in biofilm formation by foodborne pathogens Listeria monocytogenes, Salmonella enterica, and Enterohemorrhagic Escherichia coli

Food Control ◽

10.1016/j.foodcont.2016.01.004 ◽

2016 ◽

Vol 65 ◽

pp. 14-20 ◽

Cited By ~ 11

Author(s):

Nancy T. Liu ◽

Gary R. Bauchan ◽

Charlotte B. Francoeur ◽

Daniel R. Shelton ◽

Y. Martin Lo ◽

...

Keyword(s):

Escherichia Coli ◽

Listeria Monocytogenes ◽

Foodborne Pathogens ◽

Salmonella Enterica ◽

Biofilm Formation ◽

Enterohemorrhagic Escherichia Coli ◽

Ralstonia Insidiosa

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A brief overview of emergencies and dissemination of Shiga-toxin-producing E. coli (STEC) and Salmonella enterica serovar Typhimurium DT104 in humans and food producing animals

Archives of Veterinary Medicine ◽

10.46784/eavm.v13i2.239 ◽

2020 ◽

Vol 13 (2) ◽

pp. 5-15

Author(s):

Maja Velhner ◽

Branko Velebit ◽

Dalibor Todorović ◽

Miloš Pelić ◽

Suzana Vidaković Knežević ◽

...

Keyword(s):

Escherichia Coli ◽

Foodborne Pathogens ◽

Shiga Toxin ◽

Salmonella Enterica ◽

Salmonella Enterica Serovar Typhimurium ◽

Management Practice ◽

Phage Type ◽

Resistance Mechanisms ◽

Transmission Mechanisms ◽

Serovar Typhimurium

Shiga-toxin-producing Escherichia coli (STEC) and Salmonella enterica serovar Typhimurium (STDT104) are foodborne pathogens of public health significance. It is less known that Shiga-toxin-producing Escherichia coli (with cattle being the most probable natural reservoir) can be isolated from pigs, sheep and wildlife as well. The basic information about detection of Shiga-toxin-producing genes in STEC as well as the origin of Salmonella Typhimurium definite phage type 4 (STDT4) the virulence and resistance mechanisms including their distribution in the world is presented. Due to the foodborne transmission mechanisms we emphasize the role of veterinary scientist in Serbia in implementing good management practice on animal farms and in strengthening laboratory diagnostic capacities.

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Biofilm Formation by Shiga Toxin-Producing Escherichia coli on Stainless Steel Coupons as Affected by Temperature and Incubation Time

Microorganisms ◽

10.3390/microorganisms7040095 ◽

2019 ◽

Vol 7 (4) ◽

pp. 95 ◽

Cited By ~ 9

Author(s):

Zhi Ma ◽

Emmanuel W. Bumunang ◽

Kim Stanford ◽

Xiaomei Bie ◽

Yan D. Niu ◽

...

Keyword(s):

Escherichia Coli ◽

Stainless Steel ◽

Shiga Toxin ◽

Incubation Time ◽

Food Processing ◽

Biofilm Formation ◽

Crystal Violet Staining ◽

Plate Counts ◽

Air Liquid Interface ◽

Over Time

Forming biofilm is a strategy utilized by Shiga toxin-producing Escherichia coli (STEC) to survive and persist in food processing environments. We investigated the biofilm-forming potential of STEC strains from 10 clinically important serogroups on stainless steel at 22 °C or 13 °C after 24, 48, and 72 h of incubation. Results from crystal violet staining, plate counts, and scanning electron microscopy (SEM) identified a single isolate from each of the O113, O145, O91, O157, and O121 serogroups that was capable of forming strong or moderate biofilms on stainless steel at 22 °C. However, the biofilm-forming strength of these five strains was reduced when incubation time progressed. Moreover, we found that these strains formed a dense pellicle at the air-liquid interface on stainless steel, which suggests that oxygen was conducive to biofilm formation. At 13 °C, biofilm formation by these strains decreased (P < 0.05), but gradually increased over time. Overall, STEC biofilm formation was most prominent at 22 °C up to 24 h. The findings in this study identify the environmental conditions that may promote STEC biofilm formation in food processing facilities and suggest that the ability of specific strains to form biofilms contributes to their persistence within these environments.

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Influence of Bacterial Competitors on Salmonella enterica and Enterohemorrhagic Escherichia coli Growth in Microbiological Media and Attachment to Vegetable Seeds

Foods ◽

10.3390/foods10020285 ◽

2021 ◽

Vol 10 (2) ◽

pp. 285

Author(s):

Da Liu ◽

Ronald Walcott ◽

Kevin Mis Solval ◽

Jinru Chen

Keyword(s):

Escherichia Coli ◽

Foodborne Pathogens ◽

Salmonella Enterica ◽

Lactobacillus Rhamnosus ◽

Probiotic Bacterium ◽

Enterohemorrhagic Escherichia Coli ◽

Biocontrol Agents ◽

Human Pathogens ◽

Bacterial Strains ◽

Pathogen Populations

Interests in using biological agents for control of human pathogens on vegetable seeds are rising. This study evaluated whether probiotic bacterium Lactobacillus rhamnosus GG, bacterial strains previously used as biocontrol agents in plant science, as well as a selected plant pathogen could compete with foodborne human pathogens, such as Salmonella enterica and enterohemorrhagic Escherichia coli (EHEC), for growth in microbiological media and attachment to vegetable seeds; and to determine whether the metabolites in cell-free supernatants of competitive bacterial spent cultures could inhibit the growth of the two pathogens. The results suggest that the co-presence of competitive bacteria, especially L. rhamnosus GG, significantly (p < 0.05) inhibited the growth of Salmonella and EHEC. Cell-free supernatants of L. rhamnosus GG cultures significantly reduced the pathogen populations in microbiological media. Although not as effective as L. rhamnosus GG in inhibiting the growth of Salmonella and EHEC, the biocontrol agents were more effective in competing for attachment to vegetable seeds. The study observed the inhibition of human bacterial pathogens by competitive bacteria or their metabolites and the competitive attachment to sprout seeds among all bacteria involved. The results will help strategize interventions to produce vegetable seeds and seed sprouts free of foodborne pathogens.

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Transfer of Salmonella enterica Serovar Typhimurium from Beef to Tomato through Kitchen Equipment and the Efficacy of Intermediate Decontamination Procedures

Journal of Food Protection ◽

10.4315/0362-028x.jfp-15-531 ◽

2016 ◽

Vol 79 (7) ◽

pp. 1252-1258 ◽

Cited By ~ 6

Author(s):

E. GKANA ◽

A. LIANOU ◽

G.-J. E. NYCHAS

Keyword(s):

Foodborne Pathogens ◽

Salmonella Enterica ◽

Fruits And Vegetables ◽

Salmonella Enterica Serovar Typhimurium ◽

Tap Water ◽

Pathogenic Microorganisms ◽

Cross Contamination ◽

Raw Meat ◽

Serovar Typhimurium ◽

Cutting Boards

ABSTRACT It is well established that a high percentage of foodborne illness is caused by failure of consumers to prepare food in a hygienic manner. Indeed, a common practice in households is to use the same kitchen equipment for both raw meat and fresh produce. Such a practice may lead to cross-contamination of fruits and vegetables, which are mainly consumed without further processing, with pathogenic microorganisms originating from raw meat. The present study was performed to examine the transfer of the pathogenic bacterium Salmonella enterica serovar Typhimurium from inoculated beef fillets to tomatoes via contact with high-density polyethylene (PE), stainless steel (SS), and wooden (WD) surfaces and through cutting with SS knives. Furthermore, the following decontamination procedures were applied: (i) rinsing with tap water, (ii) scrubbing with tap water and liquid dish detergent, and (iii) using a commercial antibacterial spray. When surfaces and knives that came into contact with contaminated beef fillets were not cleaned prior to handling tomatoes, the lowest level of pathogen transfer to tomatoes was observed through PE surfaces. All of the decontamination procedures applied were more effective on knives than on surfaces, while among the surface materials tested, WD surfaces were the most difficult to decontaminate, followed by PE and SS surfaces. Mechanical cleaning with tap water and detergent was more efficient in decontaminating WD surfaces than using commercial disinfectant spray, followed by rinsing only with water. Specifically, reductions of 2.07 and 1.09 log CFU/cm2 were achieved by washing the WD surfaces with water and detergent and spraying the surfaces with an antibacterial product, respectively. Although the pathogen's populations on SS and PE surfaces, as well as on tomatoes, after both aforementioned treatments were under the detection limit, the surfaces were all positive after enrichment, and thus, the potential risk of cross-contamination cannot be overlooked. As demonstrated by the results of this study, washing or disinfection of kitchen equipment may not be sufficient to avoid cross-contamination of ready-to-eat foods with foodborne pathogens, depending on the decontamination treatment applied and the material of the surfaces treated. Therefore, separate cutting boards and knives should be used for processing raw meat and preparing ready-to-eat foods in order to enhance food safety.

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