Research article

Influences of naturally occurring agents in combination with fluoride on gene expression and structural organization of Streptococcus mutans in biofilms

Jae-Gyu Jeon^1,3,5 , Marlise I Klein¹ , Jin Xiao¹ , Stacy Gregoire¹ , Pedro L Rosalen^1,4,5 and Hyun Koo^1,2,5

¹  Center for Oral Biology, University of Rochester Medical Center, Rochester, New York, USA

²  Department of Microbiology and Immunology, University of Rochester Medical Center, Rochester, New York, USA

³  Department of Preventive Dentistry, School of Dentistry and Institute of Oral Bioscience, Chonbuk National University, Jeonju, Republic of Korea

⁴  Department of Physiological Sciences, Piracicaba Dental School, University of Campinas (UNICAMP), Sao Paulo, Brazil

⁵  Natural Product Research in Oral Biology Group (NatPROB), Piracicaba Dental School, University of Campinas (UNICAMP), Sao Paulo, Brazil

author email corresponding author email

BMC Microbiology 2009, 9:228doi:10.1186/1471-2180-9-228

Published:	28 October 2009

Abstract

Background

The association of specific bioactive flavonoids and terpenoids with fluoride can modulate the development of cariogenic biofilms by simultaneously affecting the synthesis of exopolysaccharides (EPS) and acid production by Streptococcus mutans, which enhanced the cariostatic effectiveness of fluoride in vivo. In the present study, we further investigated whether the biological actions of combinations of myricetin (flavonoid), tt-farnesol (terpenoid) and fluoride can influence the expression of specific genes of S. mutans within biofilms and their structural organization using real-time PCR and confocal fluorescence microscopy.

Results

Twice-daily treatment (one-minute exposure) during biofilm formation affected the gene expression by S. mutans both at early (49-h) and later (97-h) stages of biofilm development. Biofilms treated with combination of agents displayed lower mRNA levels for gtfB and gtfD (associated with exopolysaccharides synthesis) and aguD (associated with S. mutans acid tolerance) than those treated with vehicle-control (p < 0.05). Furthermore, treatment with combination of agents markedly affected the structure-architecture of S. mutans biofilms by reducing the biovolume (biomass) and proportions of both EPS and bacterial cells across the biofilm depth, especially in the middle and outer layers (vs. vehicle-control, p < 0.05). The biofilms treated with combination of agents were also less acidogenic, and had reduced amounts of extracellular insoluble glucans and intracellular polysaccharides than vehicle-treated biofilms (p < 0.05).

Conclusion

The data show that the combination of naturally-occurring agents with fluoride effectively disrupted the expression of specific virulence genes, structural organization and accumulation of S. mutans biofilms, which may explain the enhanced cariostatic effect of our chemotherapeutic approach.