Biological control of foodborne pathogens by lacticacid bacteria: A focus on juice processing industries

Abstract The use of lactic acid bacteria (LAB) in foods as biocontrol agents against foodborne pathogens has become increasingly known. Under the premise that controlling the adhesion of microorganisms to food contact surfaces is an essential step for meeting the goals of food processing, the aim of this work was to investigate the inhibitory and anti-biofilm effectiveness of Lactobacillus rhamnosus GG (ATCC 53103) and Lactobacillus casei (ATCC 393) against Escherichia coli O157:H7, Salmonella enterica and Listeria monocytogenes. Lactobacillus strains (108UFCCFU/ml) and pathogens (104UFCCFU/ml) were evaluated to monitor LAB anti-adhesive and antibiofilm effect, in two main scenarios: (i) co-adhesion and (ii) pathogen incorporation to stainless steel surfaces with a protective biofilm of Lactobacillus cells. In (i) the predominant effect was observed in L. rhamnosus against S. enterica and L. monocytogenes, whereas in (ii) both LAB significantly reduced the number of pathogenic adherent cells. The effect of pre-established LAB biofilms was more successful in displacing the three pathogens than when they were evaluated under co-adhesion. These findings show that both LAB can be considered good candidates to prevent or inhibit the adhesion and colonization of L. monocytogenes, S. enterica and E. coli O157:H7 on surfaces and conditions of relevance for juice processing industries, offering alternatives for improving the safety and quality of fruit-based products.

Resumen Existe un creciente interés en el uso de bacterias ácido lácticas (BAL) como agentes de biocontrol frente a patógenos de transmisión alimentaria. Bajo la premisa de que el control de la adhesión de microorganismos a superficies de contacto con alimentos es el paso esencial para evitar su contaminación, el objetivo de este trabajo fue investigar la efectividad inhibitoria y antibiofilm de Lactobacillus rhamnosus GG (ATCC 53103) y Lactobacillus casei (ATCC 393) frente a Escherichia coli O157:H7, Salmonella enterica y Listeria monocytogenes. A fin de cumplir con el objetivo propuesto, las cepas de Lactobacillus (108UFCUFC/ml) y los patógenos (104UFCUFC/ml) se ensayaron en 2 escenarios: (1) coadhesión, y (2) incorporación de los patógenos a las superficies de acero inoxidable con un biofilm preformado de Lactobacillus. En (1), el efecto predominante se observó con L. rhamnosus frente a S. enterica y L. monocytogenes, mientras que en (2), ambas BAL redujeron significativamente el número de células patógenas adheridas. En función de estos resultados, concluimos que el efecto de un biofilm preformado de ambas BAL fue más exitoso en el desplazamiento de los 3 patógenos que en coadhesión. Ambas BAL pueden considerarse buenas candidatas para mitigar la adhesión y colonización de L. monocytogenes, S. enterica y E. coli O157:H7 en superficies en condiciones de relevancia para la industria procesadora de jugos, y, de esta manera, ofrecer alternativas para mejorar la seguridad y calidad de los alimentos a base de frutas.

Biological control of foodborne pathogens by lactic acid bacteria: A focus on juice processing industries.

The use of lactic acid bacteria (LAB) in foods as biocontrol agents against foodborne pathogens has become increasingly known. Under the premise that controlling the adhesion of microorganisms to food contact surfaces is an essential step for meeting the goals of food processing, the aim of this work was to investigate the inhibitory and anti-biofilm effectiveness of Lactobacillus rhamnosus GG (ATCC 53103) and Lactobacillus casei (ATCC 393) against Escherichia coli O157:H7, Salmonella enterica and Listeria monocytogenes. Lactobacillus strains (108CFU/ml) and pathogens (104CFU/ml) were evaluated to monitor LAB anti-adhesive and antibiofilm effect, in two main scenarios: (i) co-adhesion and (ii) pathogen incorporation to stainless steel surfaces with a protective biofilm of Lactobacillus cells. In (i) the predominant effect was observed in L. rhamnosus against S. enterica and L. monocytogenes, whereas in (ii) both LAB significantly reduced the number of pathogenic adherent cells. The effect of pre-established LAB biofilms was more successful in displacing the three pathogens than when they were evaluated under co-adhesion. These findings show that both LAB can be considered good candidates to prevent or inhibit the adhesion and colonization of L. monocytogenes, S. enterica and E. coli O157:H7 on surfaces and conditions of relevance for juice processing industries, offering alternatives for improving the safety and quality of fruit-based products.

Application of natamycin and farnesol as bioprotection agents to inhibit biofilm formation of yeasts and foodborne bacterial pathogens in apple juice processing lines.

Biofilms serve as a reservoir for pathogenic and spoilage microorganisms, and their removal from different surfaces is a recurring problem in the beverage industry. This study aimed to investigate the effect of a combination of natamycin (NAT, 0.01 mmol/l) and farnesol (FAR, 0.6 mmol/l) against biofilms on ultrafiltration (UF) membranes and stainless steel (SS) surfaces using apple juice as food matrix. The co-adhesion of Rhodotorula mucilaginosa, Candida tropicalis, C. krusei and C. kefyr (mixed-yeast) with Listeria monocytogenes, Salmonella enterica or Escherichia coli O157:H7 (multi-species) in presence of NAT + FAR was evaluated for 2, 24, 48 h. In biofilms treated with NAT + FAR were observed by cell quantification and microscopy, inhibition of the filamentous yeast forms, disruption of the tri-dimensional structure and a high detachment of yeast cells. NAT + FAR affected the biofilms independently of the surfaces used and the presence (or not) of bacteria. L. monocytogenes was the most susceptible (p < 0.001) in multi-species biofilms, followed by E. coli O157:H7 on both surfaces (p < 0.001), whereas the growth of S. enterica was reduced (p < 0.05) in SS but not in UF-membranes (p > 0.05). Since the combination NAT + FAR affected the structure and viability of yeast species and foodborne pathogens in multi-species biofilms developed on UF-membranes and SS surfaces, the combination proposed could be considered a promising control agent to prevent biofilms in apple juice processing lines.

Microencapsulation of Lactobacillus casei and Lactobacillus rhamnosus in pectin and pectin-inulin microgel particles: Effect on bacterial survival under storage conditions.

The main objective of the research was to evaluate the performance of synbiotic delivery systems using pectin microgels on the protection of two probiotic strains (Lactobacillus casei ATCC 393 and Lactobacillus rhamnosus strain GG [ATCC 53103]) to simulated gastrointestinal digestion (GD) and storage conditions (4 ± 1 °C) in a 42 days trial. Microgel particles were prepared by ionotropic gelation method and three variables were evaluated: incubation time (24 and 48 h), free vs encapsulated cells, and presence or absence of prebiotic (commercial and Jerusalem artichoke inulin). Results demonstrated an encapsulation efficiency of 96 ± 4% into particles with a mean diameter between 56 and 118 µm. The viability of encapsulated cells after 42 days storage stayed above 7 log units, being encapsulated cells in pectin-inulin microgels more resistant to GD compared to non-encapsulated cells or without prebiotics. In all cases incubation time influenced the strains' survival.

Candida krusei isolated from fruit juices ultrafiltration membranes promotes colonization of Escherichia coli O157:H7 and Salmonella enterica on stainless steel surfaces.

To clarify the interactions between a common food spoilage yeast and two pathogenic bacteria involved in outbreaks associated with fruit juices, the present paper studies the effect of the interplay of Candida krusei, collected from UF membranes, with Escherichia coli O157:H7 and Salmonella enterica in the overall process of adhesion and colonization of abiotic surfaces. Two different cases were tested: a) co-adhesion by pathogenic bacteria and yeasts, and b) incorporation of bacteria to pre-adhered C. krusei cells. Cultures were made on stainless steel at 25°C using apple juice as culture medium. After 24 h of co-adhesion with C. krusei, both E. coli O157:H7 and S. enterica increased their counts 1.05 and 1.11 log CFU cm2, respectively. Similar increases were obtained when incorporating bacteria to pre-adhered cells of Candida. Nevertheless C. krusei counts decreased in both experimental conditions, in a) 0.40 log CFU cm2 and 0.55 log CFU cm2 when exposed to E. coli O157:H7 and S. enterica and in b) 0.18 and 0.68 log CFU cm2, respectively. This suggests that C. krusei, E. coli O157:H7, and S. enterica have a complex relationship involving physical and chemical interactions on food contact surfaces. This study supports the possibility that pathogen interactions with members of spoilage microbiota, such as C. krusei, might play an important role for the survival and dissemination of E. coli O157:H7 and Salmonella enterica in food-processing environments. Based on the data obtained from the present study, much more attention should be given to prevent the contamination of these pathogens in acidic drinks.

Role of hydrophobicity in adhesion of wild yeast isolated from the ultrafiltration membranes of an apple juice processing plant.

The role of cell surface hydrophobicity in the adhesion to stainless steel (SS) of 11 wild yeast strains isolated from the ultrafiltration membranes of an apple juice processing plant was investigated. The isolated yeasts belonged to four species: Candida krusei (5 isolates), Candida tropicalis (2 isolates), Kluyveromyces marxianus (3 isolates) and Rhodotorula mucilaginosa (1 isolate). Surface hydrophobicity was measured by the microbial adhesion to solvents method. Yeast cells and surfaces were incubated in apple juice and temporal measurements of the numbers of adherent cells were made. Ten isolates showed moderate to high hydrophobicity and 1 strain was hydrophilic. The hydrophobicity expressed by the yeast surfaces correlated positively with the rate of adhesion of each strain. These results indicated that cell surface hydrophobicity governs the initial attachment of the studied yeast strains to SS surfaces common to apple juice processing plants.