Adhesion of Two Lactobacillus gasseri Probiotic Strains on Caco-2 Cells

Previous in vitro and in vivo studies showed that two human isolates of Lactobacillus gasseri, LF221 and K7 are able to survive the passage through the gastrointestinal tract and to colonise intestines of pigs at least temporarily. The aim of this study was to examine the adhesion ability of LF221 and K7 strains to Caco-2 cells. Adhesion of lactobacilli from early stationary growth phase was examined at two pH values of DMEM buffer (4.5 and 7). Lactobacillus rhamnosus GG, a widely used strain with clinical evidences of its efficiency, served as a positive control. The number of lactobacilli added to each well was found to be crucial in the adhesion assay. When added, lactobacilli were in range of 2.5 · 106 to 2.5 · 108 cfu/well, the linear correlation between the number of adhered cells (log cfu) and the number of added cells (log cfu) was found for all three strains (R2 > 0.99) at both pH values (4.5 and 7). At the highest concentration of added K7 and GG cells tested (app. 109 cfu/well), the efficiency of adhesion was reduced. pH value of the medium strongly affected the adhesion, which was promoted in acidic conditions (pH=4.5). The adhesion of K7 strain was slightly weaker compared to GG strain at both pH values, while at pH=4.5 the adhesion of LF221 strain was even better than GG adhesion, at least at lower concentration of lactobacilli. The direct comparison of these strains was possible by regression analysis. At lower concentration of lactobacilli (2.5 · 106), the best efficiency of adhesion (% of adhered bacteria) was observed for the strain LF221, reaching the values of 7.8 and 1.9 % at pH=4.5 and 7, respectively, while at higher lactobacilli concentration the ration of adhesion was higher for GG strain (3.3 % at pH=4.5). In conclusion, strains LF221 and K7 were demonstrated to be adhesive, especially in acidic conditions. The level of adhesion of K7 and GG strains positively correlates with the number of added lactobacilli only up to the certain point when the saturation of potential binding sites on Caco-2 cells probably occurs. As the adhesion to Caco-2 cell cultures alone does not guarantee the adhesion of examined strains in vivo, additional studies on experimental animals are in progress and human clinical studies are planned as well

Determination of labeled bacteria in probiotic food supplements and medicinal products (OTC) on Slovenian market

Na slovenskem trgu se število probiotičnih dopolnil oziroma zdravil hitro povečuje. Pomemben kriterij kakovosti teh izdelkov je število živih probiotičnih mikroorganizmov, ki zagotavljajo njihovo učinkovitost. Proizvajalci, distributerji ali prodajalci se redko odločajo za preiskave probiotičnih izdelkov v neodvisnih labratorijih, saj slovenska zakonodaja ne predpisuje preverjanja njihove sestave pred prodajo. Namen raziskave je bil zato pregledati ustreznost probiotičnih prehranskih dopolnil ter probiotičnih zdravil (OTC), naprodaj v slovenskih lekarnah. Od 20 probiotičnih prehranskih dopolnil je le pet izdelkov vsebovalo dovolj živih mikroorganizmov glede na deklaracijo, pri dveh izdelkih pa število ni bilo deklarirano. V šestih izdelkih z metod PCR nismo našli živih predstavnikov vseh navedenih vrst. Obe probiotični zdravili sta ustrezali tako po vrstni sestavi kot po številu probiotičnih bakterij. Rezultati kažejo na potrebo po boljši kontroli kakovosti tovrstnih izdelkov.There are an increasing number of probiotic food supplements or medicinal products on the Slovenian market. Probiotic microorganisms in these products have to be viable otherwise the effects may not be expected. Producers, distributors or sellers do not decide often for the analysis of probiotic products in independent laboratories since Slovenian legislation does not require control of their content before being sold. The aim of this study was to survey the suitability of probiotic food supplements and medicinal products (OTC) available in Slovenian pharmacies. Five food supplements out of 20 tested contained enough viable microorganisms while the number was not declared in two products. In six products the viable representatives of all declared species were not found by PCR method. Both probiotic medicinal products were suitable regarding the species composition and number of probiotic bacteria. Results indicate the need for better quality control of such products

Wide-inhibitory spectra bacteriocins produced by Lactobacillus gasseri K7

The aim of study was to determine the genetic characterization and classification of Lb. gasseri K7 bacteriocins, comparison with bacteriocins of the Lb. gasseri LF221 strain and other related strains. Bacteriocin-encoding genes were amplified by PCR, subjected to DNA sequencing, and BLAST sequence analysis was performed to search the database for homologous peptides. Lb. gasseri K7 produces two-peptide bacteriocins, named gassericin K7 A and gasserin K7 B. Their nucleotide sequences were deposited at GenBank, under accession numbers EF392861 for the gassericin K7 A and AY307382 for the gassericin K7 B. Analysis of geneclausters of bacteriocins in Lb. gasseri K7 strain revealed a 100 percent sequence identity with bacteriocins in LF221 strain. An active peptide of gassericin K7 B is homologous to the complementary peptide of gassericin T, and a complementary peptide of gassericin K7 B is homologous to the active peptide of gassercin T. Another surprising finding was that the sakacin T0beta peptide is partly homologous to the active peptide of gassercin K7 A, while the other sakacin T peptide (alfa) is partly homologous to the complementary peptide of gassericin K7 B. Gassericins of Lb. gasseri K7 strain were both classified as two-peptide bacteriocins. Human probiotic strains Lb. gasseri K7 and LF221 aredifferent isolates but with identical bacteriocin genes. They produce wide-inhibitory spectra bacteriocins that are new members of two-peptide bacteriocins with some homologies to other bacteriocins in this group. Described bacteriocins offer a great potential in applications in food industry, pharmacy and biomedicine