Effects of Lactobacillus crispatus as a candidate for oral probiotic bacteria on Haemophilus influenzae

Kaoru Sogabe-Ashigaki, DDS,a Susumu Imai, PhD,b Ayako Okada, DDS, PhD,b Khairul Matin, DDS, PhD,b Gaku Akimaru, DDS,a Tomohiko Terai, MSc,c Takekazu Okumura, PhD,c Nobuhiro Hanada, DDS, PhD,b and Hiroshi Kawahara, DDS, PhDa

aDepartment of Dental Anesthesiology, bDepartment of Translational Research, Tsurumi University School of Dental Medicine, Yokohama, and cYakult Central Institute, Food Research Department, Tokyo, Japan

Purpose: To verify the effects of five candidates for oral probiotics on Haemophilus influenzae and to more characterize the effects of Lactobacillus crispatus YIT 12319 (L.c1).
Materials and Methods: Cell free culture supernatants (CFCSs) were acquired from cultures of the clinical isolates (oral probiotic candidate bacteria). Two laboratory strains of H. influenzae (type b and non-typeable) were freshly cultured in Muller-Hinton broth medium. A paired competition assay was performed using each clinical isolate versus one of the two H. influenzae strains on culture plates. Antibacterial effects of all unprocessed CFCSs, neutralized CFCSs, and an ammonium sulfate precipitate of the CFCSs against H. influenzae were determined using a radial diffusion assay. The bactericidal effect of L.c1 CFCS was also verified using a colony formation assay. Furthermore, the organic acids produced by L.c1 were analyzed using HPLC.
Results: L.c1 displayed uninterrupted growth by suppressing the growth of both stains of H. influenzae equally, while the results for the rest of the clinical isolates were inconsistent. Only unprocessed CFCS of L.c1 clearly inhibited the growth of H. influenzae strains and showed potentially strong bactericidal effects. Reasonable amounts of lactic acid, acetic acid, succinic acid, and propionic acid were detected by HPLC.
Conclusion: The effects of L.c1 on H. influenzae strains observed in this study will serve as important supporting evidence for the potential use of lactobacillus species as probiotics. In addition, the growth appeared to be inhibited by the organic acids produced by L.c1.
(Asian Pac J Dent 2015; 15: 3-11.)
Key Words: Haemophilus influenzae, Lactobacillus crispatus, oral probiotics, organic acid