Amensalistic activity of Lactobacillus spp., isolated from human samples

Abstract

Lactobacillus is a bacterial genus belonging to LAB (Lactic Acid Bacteria) and they are among the most common probiotics. Recent guidelines on probiotics, issued by the Italian Ministry of Health, state that, on the basis of the available literature, the amount sufficient to obtain a temporary colonization of the intestine by a probiotic strain is at least 10^9 living cells. A microorganism can be defined as a probiotic strain if it is of human origin, if it survive to the gastrointestinal tract, resisting the acidity of the stomach and the action of bile, and it should have immunostimulant activity. In addition, the strain should be able to adhere to the mucosa causing no toxicity, and to produce substances with antibacterial activity against some pathogens. The aim of the work was the isolation and identification of lactobacilli of human origin. It was also deepened the study of their amensalistic properties, with particular attention to the resistance to gastrointestinal transit and their antagonism against pathogenic microorganisms. Three hundred fifty-nine lactobacilli strains were isolated from swabs of healthy people and identified using molecular techniques based on the study of 16S rDNA. The identification of some strains was confirmed by further analysis DHPLC V1 and V3 of 16S rDNA. The strains were subjected to the evaluation of the resistance to bile salts and low pH, to the production of hydrogen peroxide and more particularly, it has been evaluated the ability to produce substances with antibacterial activity. Finally, the attention was focused on the characterization of an active supernatant produced by an oral strain. The isolation of the substance provided chromatographic procedures such as SEC (Size Exclusion Chromatography using Sephadex 50) and SPE (Reverse Phase Chromatography using C18 column). The results were shown that the active fraction has a low molecular weight and for its chemical-physical characteristics is not a common bacteriocin, for this reason are on going further chromatographic studies using columns with increasing polarity (C4, phenyl, cyano, and amino) . Future outlooks are focused on the identification of the molecule in question, by MALDI-TOF and ESI-TOF and then optimizing the whole process to standardize the entire method. In this way, the opportunity to bring to light new molecules will be possible, with the ultimate goal of being able to take advantage from these antibacterial substances.