Advances in Characterizing the Transport Systems of and Resistance to EntDD14, A Leaderless Two-Peptide Bacteriocin with Potent Inhibitory Activity.

Enterocin DD14 (EntDD14) is a two-peptide leaderless bacteriocin produced by the Enterococcus faecalis 14 strain previously isolated from meconium. This bacteriocin is mainly active against Gram-positive bacteria. Leaderless bacteriocins do not undergo post-translational modifications and are therefore immediately active after their synthesis. As a result, the cells that produce such bacteriocins have developed means of protection against them which often involve transport systems. In this and our previous work, we constructed different mutants deleted in the genes involved in the transport functions, thus covering all the supposed components of this transport system, using Listeria innocua ATCC 33090 as the indicator strain to assess the activity of externalized EntDD14. We also assessed the self-resistance of the WT and all its engineered derivative mutants against EntDD14, provided extracellularly, in order to evaluate their self-resistance. The results obtained highlight that the ABC transporter constituted by the DdG, H, I, and J proteins contributes to EntDD14 export as well as resistance to an external supply of EntDD14. Our results also have established the essential role of the DdE and DdF proteins as primary transporters dedicated to the externalization of EntDD14. Moreover, the in silico data showed that DdE and DdF appear to assemble in a formation that forms an essential channel for the exit of EntDD14. This channel DdEF may interact with the ABC transporter DdGHIJ in order to control the flow of bacteriocin across the membrane, although the nature of this interaction remains to be elucidated.

Metabolic Shift of an Isogenic Strain of Enterococcus faecalis 14, Deficient in Its Own Bacteriocin Synthesis, as Revealed by a Transcriptomic Analysis.

The production of antimicrobial molecules often involves complex biological pathways. This study aimed at understanding the metabolic and physiological networks of enterocin EntDD14-associated function, in the bacteriocinogenic strain, Enterococcus faecalis 14. A global and comparative transcriptomic study was carried out on E. faecalis 14 and its isogenic mutant Δbac, inactivated in genes coding for EntDD14. The in vitro ability to form biofilm on polystyrene plates was assessed by the crystal violet method, while the cytotoxicity on human colorectal adenocarcinoma Caco-2 cells was determined by the Cell Counting Kit-8. Transcriptomic data revealed that 71 genes were differentially expressed in both strains. As expected, genes coding for EntDD14 were downregulated in the Δbac mutant, whereas the other 69 genes were upregulated. Upregulated genes were associated with phage-related chromosomal islands, biofilm formation capability, resistance to environmental stresses, and metabolic reprogramming. Interestingly, the Δbac mutant showed an improved bacterial growth, a high capacity to form biofilm on inanimate surfaces and a very weak cytotoxicity level. These multiple metabolic rearrangements delineate a new line of defense to counterbalance the loss of EntDD14.

Abundance of Lactobacillus plantarum Strains with Beneficial Attributes in Blackberries (Rubus sp.), Fresh Figs (Ficus carica), and Prickly Pears (Opuntia ficus-indica) Grown and Harvested in Algeria.

This first study performed on traditional fruits consumed in North Africa reveals their richness in microorganisms with beneficial attributes like cholesterol lowering capabilities. Blackberries (Rubus sp.), fresh figs (Ficus carica), and prickly pears (Opuntia ficus-indica) are fruits largely and traditionally consumed in Kabylia, a beautiful northern Algerian region. Here, 85 lactic acid bacteria (LAB)-isolates were isolated and identified by MALDI-TOF mass spectrometry. The identified species belong to Lactobacillus and Leuconostoc genera. These 85 LAB-isolates were then assessed for their capabilities to grow under conditions mimicking the gastrointestinal tract, and the resulting data were statistically treated with principal component analysis (PCA). After which, only 26 LAB-isolates were selected and characterized for their genetic relatedness using random amplified polymorphic DNA (RAPD) method. Following the genetic relatedness assessment, only 10 LAB-strains, among which nine Lactobacillus plantarum and one Lactobacillus paracasei were studied for their pathoproperties and some probiotic features. Interestingly, all of these 10 LAB-strains were devoid of adverse effects, but capable to adhere to human epithelial colorectal adenocarcinoma Caco-2 cells. Of note, these 10 LAB-strains exhibited an important in vitro hypocholesteromia effect, in strain-dependent manner. Moreover, the Lactobacillus strains exhibited a high bile salt hydrolase (BSH) activity which was correlated with expression of bsh2, bsh3 and bsh4 genes.