A highly acid-resistant novel strain of Lactobacillus johnsonii No. 1088 has antibacterial activity, including that against Helicobacter pylori, and inhibits gastrin-mediated acid production in mice

Abstract

A novel strain of Lactobacillus johnsonii No. 1088 was isolated from the gastric juice of a healthy Japanese male volunteer, and characterized for its effectiveness in the stomach environment. Lactobacillus johnsonii No. 1088 was found to have the strongest acid resistance among several lactobacilli examined (>10% of cells survived at pH 1.0 after 2 h), and such a high acid resistance property was a specific characteristic of this strain of L. johnsonii. When cultured with various virulent bacteria, L. johnsonii No. 1088 inhibited the growth of Helicobacter pylori, Escherichia coli O-157, Salmonella Typhimurium, and Clostridium difficile, in which case its effectiveness was more potent than that of a type strain of L. johnsonii, JCM2012. In addition to its effect in vitro, L. johnsonii No. 1088 inhibited the growth of H. pylori in human intestinal microbiota-associated mice in both its live and lyophilized forms. Moreover, L. johnsonii No. 1088 suppressed gastric acid secretion in mice via decreasing the number of gastrin-positive cells in the stomach. These results taken together suggest that L. johnsonii No. 1088 is a unique lactobacillus having properties beneficial for supporting H. pylori eradication by triple therapy including the use of a proton pump inhibitor (PPI) and also for prophylaxis of gastroesophageal reflux disease possibly caused after H. pylori eradication as a side effect of PPI.

Keywords: Gastroesophageal reflux disease; Helicobacter pylori; Lactobacillus johnsonii; lactic acid bacteria; probiotics.

Figure 1

Schematic drawing of time schedule of the experiment to examine the effect of proton pump inhibitor (PPI) and Lactobacillus johnsonii No. 1088 on the number of gastrin-positive cells and stomach weight.

Figure 2

Viability of Lactobacillus johnsonii No. 1088 and other lactobacilli at pH 1.0 (A), 1.5 (B), and 2.0 (C). Growing bacteria (L. johnsonii No. 1088, L. johnsonii La1, L. gasseri OLL2716, L. casei shirora, L. rhamnosus GG, L. acidophilus BF, and L. brevis KB290 for pH 1.0, 1.5, and 2.0; and type strains L. johnsonii JCM2012 and L. gasseri JCM1131 added for pH 1.0) were diluted at a density around 10⁷ CFU/mL in acidic buffer and then incubated at 37°C up to 120 min. Counts of living bacterial counts were made at various time points.

Figure 3

Antibacterial effect of Lactobacillus johnsonii No. 1088 and its type strain L. johnsonii JCM2012 under mixed-culture conditions. Lactobacillus johnsonii No. 1088 or JCM2012 (10⁶ CFU/mL) was co-cultured with various virulent bacteria (A, Helicobacter pylori No. 130 [10⁷ CFU/mL]; B, Escherichia coli O-157 [10⁷ CFU/mL]; C, Salmonella. Typhimurium LT2 [10⁷ CFU/mL]; D, Clostridium difficile JCM1296 [10⁷ CFU/mL] or E, Candida albicans TI3001 [10⁶ CFU/mL]), and numbers of viable bacteria were determined at the various time points indicated. Symbols are defined in the figure itself.

Figure 4

Anti-Helicobacter pylori effect of Lactobacillus johnsonii No. 1088 in human intestinal microbiota-associated mice. Mice associated with human intestinal microbiota were prepared by using germ-free mice and then infected with Helicobacter pylori No. 130 (10⁹ CFU/mice) as described in Materials and Methods. Helicobacter pylori-bearing mice were orally and daily administered growing live L. johnsonii No. 1088 (A and C) or a comparable number of lyophilized cells (B and D) for 2 or 4 weeks. In both mice treated with live or the lyophilized form of L. johnsonii No. 1088, the number of H. pylori in the stomach significantly decreased (A and B). Along with the decrease in the number of H. pylori, a steady number of L. johnsonii No. 1088 was detected in the stomach (C and D). Statistical significance was determined by use of Student's t-test (*P < 0.05, **P < 0.01, ^‡P < 0.0001 versus no treatment for comparable time periods).

Figure 5

Effect of proton pump inhibitor (PPI) and Lactobacillus johnsonii No. 1088 on the number of gastrin-positive cells and stomach weight. Germ-free Balb/c mice (4 weeks old) were orally administered 10⁹ CFU of live L. johnsonii No. 1088 (closed bars) or PBS (open bars). At 8 weeks of age, 4–5 mice in each group were administered s.c. with 200 μg of the PPI omeprozaol every 2 days (8-week PPI treatment group). At 12 weeks of age, the remaining 3–5 mice in each group were administered PPI as above (4-week PPI treatment group). At 16 weeks of age, all mice were sacrificed; and the number of gastrin-positive cells (A) and stomach weight (B) were analyzed as described in Materials and Methods. Statistical significance of difference between groups was determined by Tukey's honestly significant difference test and summarized in Table2, whereas selected results are indicated in the graph (**P < 0.01; ***P < 0.001 as indicated by the brackets).

Figure 6

Decrease in serum gastrin concentration after oral treatment of germ-free mice with heat-killed Lactobacillus johnsonii No. 1088 for 10 days. Various numbers of heat-killed L. johnsonii No. 1088 were orally administered for 10 days, and serum concentrations of gastrin were determined with ELISA. Administration with 10⁹ or 10¹⁰ CFU of heat-killed L. johnsonii No. 1088 significantly decreased the serum gastrin concentration. *P < 0.05 versus PBS (Dunnett's test).