Inhibition of Streptococcus pneumoniae adherence to human epithelial cells in vitro by the probiotic Lactobacillus rhamnosus GG

Abstract

Background: Colonization of the nasopharynx by Streptococcus pneumoniae is considered a prerequisite for pneumococcal infections such as pneumonia and otitis media. Probiotic bacteria can influence disease outcomes through various mechanisms, including inhibition of pathogen colonization. Here, we examine the effect of the probiotic Lactobacillus rhamnosus GG (LGG) on S. pneumoniae colonization of human epithelial cells using an in vitro model. We investigated the effects of LGG administered before, at the same time as, or after the addition of S. pneumoniae on the adherence of four pneumococcal isolates.

Results: LGG significantly inhibited the adherence of all the pneumococcal isolates tested. The magnitude of inhibition varied with LGG dose, time of administration, and the pneumococcal isolate used. Inhibition was most effective when a higher dose of LGG was administered prior to establishment of pneumococcal colonization. Mechanistic studies showed that LGG binds to epithelial cells but does not affect pneumococcal growth or viability. Administration of LGG did not lead to any significant changes in host cytokine responses.

Conclusions: These findings demonstrate that LGG can inhibit pneumococcal colonization of human epithelial cells in vitro and suggest that probiotics could be used clinically to prevent the establishment of pneumococcal carriage.

Figure 1

Time-course of pneumococcal adherence to epithelial cells. Cells were inoculated with 7.2 x 10⁵ CFU (95%CI 5.8 x 10⁵, 8.7 × 10⁵ CFU) and the number of adherent pneumococci measured over three hours. Mean ± SD for four pneumococcal isolates are depicted (n=2).

Figure 2

Effect of LGG on pneumococcal adherence to epithelial cells. Pneumococcal adherence was determined when incubated with medium alone (Pnc), or with medium containing 100 U/ml heparin (Heparin), or ~5 x 10⁶ CFU LGG (LGG low), or ~5 x 10⁷ CFU LGG (LGG high) added one hour before (A), concurrently (B), or one hour after adding pneumococci (C). One-way ANOVA revealed significant differences in adherence levels (P < 0.05) for all isolates except PMP812 (serotype 5) in the post-addition assay. For each isolate, Bonferroni’s post-test was used to compare heparin, LGG low, and LGG high to Pnc: *, P < 0.001; #, P < 0.05. Data are mean + SD (n≥3).

Figure 3

Effect of LGG on epithelial cytokine production. Concentrations of IL-6 and IL-8 in culture supernatants of untreated epithelial cells (epithelial) or epithelial cells incubated with pneumococci (Pnc), pneumococci with 100 U/ml heparin (Heparin); or~5 × 10⁶ CFU LGG (LGG low), or ~5 × 10⁷ CFU LGG (LGG high) added one hour prior to the addition of pneumococcal isolates PMP843 (serotype 19 F) (A and C) and PMP6 (serotype 5) (B and D) in the adherence assay. Data reported as mean + SD (n=3).