Genetic and phenotypic assessment of the antimicrobial activity of three potential probiotic lactobacilli against human enteropathogenic bacteria.

Introduction: Lactobacilli are avid producers of antimicrobial compounds responsible for their adaptation and survival in microbe-rich matrices. The bactericidal or bacteriostatic ability of lactic acid bacteria (LAB) can be exploited for the identification of novel antimicrobial compounds to be incorporated in functional foodstuffs or pharmaceutical supplements. In this study, the antimicrobial and antibiofilm properties of Lactiplantibacillus pentosus L33, Lactiplantibacillus plantarum L125 and Lacticaseibacillus paracasei SP5, previously isolated form fermented products, were examined, against clinical isolates of Staphylococcus aureus, Salmonella enterica subsp. enterica serovar Enteritidis and Escherichia coli. Methods: The ability of viable cells to inhibit pathogen colonization on HT-29 cell monolayers, as well as their co-aggregation capacity, were examined utilizing the competitive exclusion assay. The antimicrobial activity of cell-free culture supernatants (CFCS) was determined against planktonic cells and biofilms, using microbiological assays, confocal microscopy, and gene expression analysis of biofilm formation-related genes. Furthermore, in vitro analysis was supplemented with in silico prediction of bacteriocin clusters and of other loci involved in antimicrobial activity. Results: The three lactobacilli were able to limit the viability of planktonic cells of S. aureus and E. coli in suspension. Greater inhibition of biofilm formation was recorded after co-incubation of S. enterica with the CFCS of Lc. paracasei SP5. Predictions based on sequence revealed the ability of strains to produce single or two-peptide Class II bacteriocins, presenting sequence and structural conservation with functional bacteriocins. Discussion: The efficiency of the potentially probiotic bacteria to elicit antimicrobial effects presented a strain- and pathogen-specific pattern. Future studies, utilizing multi-omic approaches, will focus on the structural and functional characterization of molecules involved in the recorded phenotypes.

Assessment of the Physicochemical, Antioxidant, Microbial, and Sensory Attributes of Yogurt-Style Products Enriched with Probiotic-Fermented Aronia melanocarpa Berry Juice.

The aim() of this study was to create() various formulations of yogurt enriched with freeze()-dried adjuncts, namely() (i) probiotic Lactobacillus plantarum ATCC 14917 culture(), and (ii) L. plantarum ATCC 14917 fermented black chokeberry juice, along with a commercial() starter culture(). The goal was to enhance() functionality and optimize the nutritional() value() of the products. These new yogurt-style() formulations were subsequently() compared with commercially produced yogurt. All products demonstrated() favorable() physicochemical properties, and the probiotic strain() consistently() maintained viable() levels exceeding 7 log() cfu/g throughout() the entire() storage() period(). The fermented milk produced with the adjunct-free L. plantarum cells, as well as the yogurt produced with the proposed() lactobacilli-fermented chokeberry juice, exhibited the highest lactic acid() production() (1.44 g/100 g yogurt by the end of storage()). Levels of syneresis were observed at lower() values() in yogurt produced with freeze()-dried fermented chokeberry juice. Yogurts prepared() with the lactobacilli-fermented freeze()-dried chokeberry juice displayed elevated total() phenolic content() and antioxidant capacity() (25.74 µg GAE/g and 69.05 µmol TE/100 g, respectively()). Furthermore, sensory tests revealed a distinctive() fruity flavor() in samples incorporating fermented juice. The results demonstrate() that probiotic L. plantarum-fermented chokeberry juice enhances() both the antioxidant capacity() and the viability of beneficial() bacteria() in yogurt while it can be readily() applied and commercialized, especially in the form of a freeze()-dried formulation.

Assessment of Antimicrobial Activity of Pomegranate, Cranberry, and Black Chokeberry Extracts against Foodborne Pathogens.

Herbal and plant extracts are being applied to a wide range of foods against different types of foodborne pathogens. In the present study, ethanolic and aqueous extracts of different concentrations (5% v/v, 10% v/v, and 20% v/v) from cranberry (Vaccinium macrocarpon), black chokeberry (Aronia melanocarpa), and pomegranate (Punica granatum L.) plants were applied in five concentrations (62.5 to 1000 mg/mL) against foodborne strains of Staphylococcus aureus,Escherichia coli, and Streptococcus pyogenes. The results revealed that a low concentration of solvents (5% v/v) did not exhibit decreased antimicrobial activity in comparison with higher solvent concentrations (10% and 20% v/v). Additionally, both aqueous and ethanolic extracts were highly effective against pathogens even in their low concentrations (62.5 mg/mL and 125 mg/mL). Likewise, the extracts exhibited promising results (aqueous extracts of pomegranate, cranberry, and black chokeberry in a food-compatible concentration of 2% w/v) were applied to raw pork meatball production, and their antimicrobial activity was recorded versus Enterobacteriaceae, total mesophilic bacteria (TMB), lactic acid bacteria (LAB), Staphylococcus spp., Pseudomonas spp., and yeasts/molds. The outcome demonstrated that meatballs that contained aqueous extracts of pomegranate were more resistant to spoilage compared to all of the other samples, as it was preserved for more days. Likewise, these extracts of a plant origin could be used as natural preservatives in meat products, even in their low concentrations.

Assessment of the Immunomodulatory Properties of the Probiotic Strain Lactobacillus paracasei K5 in vitro and In Vivo.

Lactobacillus paracasei K5 is a lactic acid bacteria (LAB) strain that has been isolated from dairy products. Previous studies have established its probiotic potential in a series of in vitro tests, including molecular characterization, safety profiling, and tolerability of the gastrointestinal tract conditions. To characterize its beneficial actions on the host, we have shown previously that L. paracasei K5 adheres to Caco-2 cells and exerts anti-proliferative effects through the induction of apoptosis. In the present study, we focused on the immunomodulatory potential of this strain. We employed the dorsal-air-pouch mouse model of inflammation and recorded an eight-fold increase in the recruitment of immune cells in mice treated with the probiotic strain, compared to the control group. Analysis of the exudates revealed significant changes in the expression of pro-inflammatory mediators on site. Treatment of Caco-2 cells with L. paracasei K5 induced significant upregulation of cytokines interleukin-1α (IL-1α), ΙL-1ß, IL-6, tumor necrosis factor-alpha (TNF-α), the chemokine C-X-C motif ligand 2 (CXCL2), and the inflammation markers soluble intercellular adhesion molecule (sICAM) and metallopeptidase inhibitor-1 (TIMP-1). Transient induction of the Toll-like receptors (TLRs) 2, 4, 6, and 9 expression levels was recorded by real-time PCR analysis. These results highlight the immunomodulatory potential of this strain and further support its probiotic character.

Assessment of Ready-to-Use Freeze-dried Immobilized Biocatalysts as Innovative Starter Cultures in Sourdough Bread Making.

In the present study the effect of innovative biocatalysts as starter cultures in sourdough bread making was explored. The biocatalysts consisted of Lactobacillus paracasei K5 and Lactobacillus bulgaricus ATCC 11842 (in single and mixed form), immobilized on delignified wheat bran (DWB), and freeze dried without cryoprotectants. The parameters monitored were physicochemical characteristics, mold and rope spoilage appearance, volatile composition, and organoleptic characteristics. Results obtained showed that both biocatalysts exhibit good fermentative activity. However, the best results were achieved when freeze-dried immobilized L. paracasei K5 was applied as a single culture. In particular, the produced bread had a higher acidity (8.67 mL 0.1 N NaOH) and higher organic load (2.90 g/kg lactic acid and 1.11 g/kg acetic acid). This outcome was the main reason why this bread was preserved more regarding mold spoilage (14 days) and rope spoilage (12 days), respectively. In addition, the employment of freeze-dried immobilized L. paracasei K5 led to bread with better aromatic profile in terms of concentrations and number of volatile compounds produced as gas chromatography/mass spectrometry (GC/MS) analysis proved. Finally, no significant differences were observed through sensorial tests. Last but not least, it should be highlighted that the used microorganisms were cultured in cheese whey, minimizing the cost of the proposed biotechnological procedure.