Bacterial Diversity in the Saliva and Plaque of Caries-free and Caries-active Filipino Adults
Candids Patrice A. Reyes1 and Leslie Michelle M. Dalmacio1
1Department of Biochemistry and Molecular Biology,
College of Medicine, University of the Philippines, Manila
2Department of Biology, College of Arts and Sciences,
University of the Philippines, Manila
corresponding author: This email address is being protected from spambots. You need JavaScript enabled to view it.
ABSTRACT
The oral cavity is inhabited by hundreds of bacterial species that play vital roles in maintaining oral health or in shifting to a diseased state such as dental caries. These microorganisms have been studied in the past using culture-dependent methods. However, due to the limitations of microbial identification through culture techniques, culture-independent techniques are currently being utilized to better understand the bacterial etiology of dental caries. In this study, molecular-based techniques were utilized to determine the bacterial profile of the saliva and supragingival dental plaque of caries-free and caries-active healthy Filipino adults. Bacterial DNA was amplified using primers for Eubacterial 16S rRNA gene (16S rDNA), diversity was assessed through denaturing gradient gel electrophoresis (DGGE), and the microorganisms were identified through sequencing of the 16S rDNA. Based on the DGGE profile and 16S rDNA sequences, the caries-free group exhibited a more diverse microflora compared with its caries-active counterpart. This suggests that part of the microflora may be inhibited or absent in a caries-active oral cavity. This is the first study that provides a baseline profile of the oral microbial diversity in caries-free and caries-active Filipino adults using culture-independent techniques. This profile may assist researchers and dental practitioners to gain better understanding of the microbial etiology of dental caries. It may also be used in future caries risk assessment and anti-caries vaccine development.
INTRODUCTION
Dental caries is a disease that involves the destruction of the calcified tissues of the teeth. It is caused by the interplay of factors involving various oral bacteria in the presence of fermentable carbohydrates surrounding the tooth over a period of time (Lamont & Jenkinson 2010; Lundeen & Roberson 1995). It is considered by the World Health Organization (2003) as one of the most important global oral health burdens . . . . . . . . . . . . . . .
REFERENCES
AAS JA, GRIFFEN AL, DARDIS SR, LEE AM, OLSEN I, DEWHIRST FE, LEYS EJ, PASTER BJ. 2008. Bacteria of dental caries in primary and permanent teeth in children and young adults. J Clin Microbiol 46(4): 1407-17.
ALLISON HE, HILLMAN JD. 1997. Cloning and Characterization of Prevotella melaninogenica Hemolysin. Infect Immun 65(7): 2765-71.
CAUFIELD PW, LI Y, A DASANAYAKE and D SAXENA. 2007. Diversity of Lactobacilli in the Oral Cavities of Young Women with Dental Caries. Caries Res 41(1): 2-8.
CHHOUR KL, NADKARNI MA, BYUN R, MARTIN FE, JACQUES NA, HUNTER N. 2005. Molecular Analysis of Microbial Diversity in Advanced Caries. J Clin Microbiol 43(2): 843-849.
CORBY PM, LYONS-WEILER J, BRETZ WA, HART TC, AAS JA, BOUMENNA T, GOSS J, CORBY AL, JUNIOR HM, WEYANT RJ, PASTER BJ. 2005. Microbial Risk Indicators of Eary Childhood Caries. J Clin Microbiol 43(11): 5753-59.
DAEP CA, LAMONT RJ, DEMUTH DR. 2008. Interaction of Porphyromonas gingivalis with Oral Streptococci Requires a Motif that Resembles the Eukaryotic Nuclear Receptor Box Protein-Protein Interaction Domain. Infect Immunol 76(7): 3273-80.
DOYLE JJ, DOYLE JL. 1987. Isolation of DNA from fresh plant tissue. Focus 12: 13-15.
GRICE, EA, HH KONG, S CONLAN, CB DEMING, J DAVIS, AC YOUNG, GG BOUFFARD, RW BLAKESLEY, PR MURRAY, ED GREEN, ML TURNER, JA SEGRE. 2009. Topographical and Temporal Diversity of the Human Skin Microbiome. Science. 324(5931): 1190-92.
GRICE EA, SNITKIN ES, YOCKEY LJ, BERMUDEZ DM. 2010. Longitudinal Shift in Diabetic Wound Microbiota Correlates with Prolonged Skin Defense Response. Proc Nat Acad Sci 107(33): 14799-804.
HANDELSMAN J. 2004. Metagenomics: Application of Genomics to Uncultured Microorganisms. Microbiol Mol Biol Rev 68(4): 669-685.
HARALDSSON, G. 2005. Oral commensal Prevotella species and Fusobacterium nucleatum: Identification and potential pathogenic role. (Academic Dissertation) University of Helsinki, Finland. 1-72.
HOOPER SJ, CREANAN SJ, LEWIS MAO, SPRATT DA, WADE WG, WILSON MJ. 2006. Viable Bacteria Present within Oral Squamous Cell Carcinoma Tissue. J Clin Microbiol 44(5): 1719-25.
HUTTER G, SCHLAGENHAUF U, VALENZA G, HORN M, BURGEMEISTER S, CLAUS H, VOGEL U. 2003. Molecular analysis of bacteria in periodontitis: evaluation of clone libraries, novel phylotypes and putative pathogens. Microbiol 67-75.
KANG MS, NA HS, OH JS. 2005. Coaggregation of Weissela cibaria isolates with Fusobacterium nucleatum and their Adhesiveness to Epithelial Cells. FEMS Microbiol Lett 323-329.
KIM-LY C, NADKARNI MA, BYUN R, MARTIN E, JACQUES N, HUNTER N. 2005. Molecular Analysis of Microbial Diversity in Advanced Caries. J Clin Microbiol 43(2): 843-849.
KURAMITSU HK, HE X, LUX R, ANDERSON MH, SHI W. 2007. Interspecies Interactions within Oral Microbial Communities. Microbiol Mol Biol Rev 71(4): 653-670.
LAMONT RJ, JENKINSON HF. 2010. Caries as an infectious disease. In Oral Microbiology at a Glance. Singapore: Wiley-Blackwell. p. 6-7.
LEE BR, BILLINGS RJ, KASTER A. 1986. Quantitative comparisons of potentially cariogenic microorganisms cultured from noncarious and carious root and coronal tooth surfaces. Infect Immunol 51(3): 765-770.
LI Y, KU CYS, XU J, SAXENA D, CAUFIELD PW. 2005. Survey of oral microbial diversity using PCR-based denaturing gradient gel electrophoresis. J Dental Res 84(6): 559-564.
LUNDEEN TF, ROBERSON T. 1995. Cariology: the lesion, etiology, prevention and control. In: The Art and Science of Operative Dentistry. Sturdevant CM. Third Edition. St. Louis, Missouri, USA: Mosby-Year Book, Inc. p. 60-126.
MADIGAN MT, MARTINKO JM, PARKER J. 2004. Brock Biology of Microorganisms. Tenth edition. USA: Prentice Hall. p. 614-631.
MARCOTTE H, LAVOIE MC. 1998. Oral Microbial Ecology and the Role of Salivary Immunoglobulin A. Microbiol Mol Biol Rev 62(1): 71-109.
MARSH PD. 2006. Dentla Plaque as a Biofilm and a Microbial Community – Implications for Health and Disease. BMC Oral Health 6(1).
MARTIN NC, PIRIE AA, FORD LV, CALLAGHAN CL, MCTURK K, LUCY D, SCRIMGER DG. 2006. The use of phosphate buffered saline for the recovery of cells and spermatozoa from swabs. Science and Justice. 43(3): 179-184.
MONSE B, MABUNGA S, SAROL J, HEINRICHWELTZIEN R. 2006. National Oral Health Survey Among the Public School Population in the Philippines. Pasig City, Philippines: Department of Education. 1-57p. Retrieved from: www.fitforschool.ph
MUNSON, MA, A BANERJEE, TF WATSON and WG WADE. 2004. Molecular analysis of the microflora associated with dental caries. J Clin Microbiol 42(7): 3023-29.
MUYZER G, DE WAAL EC, UITTERLINDEN AG. 1993. Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chainreaction-amplified genes coding for 16S rRNA. Appl Environ Microbiol 59: 695-700.
NADKARNI MA, CALDON CE, CHHOUR KL, FISHER IP, MARTIN E, JACQUES NA, N HUNTER. 2004. Carious Dentine Provides a Habitat for a Complex Array of Novel Prevotella-like Bacteria. J Clin Microbiol 42(1): 5238-44.
NAKATSU CH, TORSVIK V, ØVREÅS L. 2000 Soil community analysis using DGGE of 16S rDNA polymerase chain reaction products. Soil Sci Soc Am J 64(4): 1382-88.
[NMEDS] National Monitoring and Epidemiological Dental Survey. Department of Health, Philippines. 1998. Retrieved from www.whocollab.od.mah.se on 25 August 2010.
NEWMAN M, TAKEI H, CARRANZA F. 2002. Carranza’s Clinical Periodontology. USA: W.B. Saunders Company. 108-109p.
PASTER BJ, BOCHES SK, GALVIN JL, ERICSON RE, LAU CN, LEVANOS VA, SAHASRABUDHE A, DEWHIRST FE. 2001. Bacterial diversity in human subgingival plaque. J Bacteriol 183(12): 3770-83.
PHILIP K, TEOH WY, MUNIANDY S, YAAKOB H. 2008. Identification of major cultivable aerobic bacteria in the oral cavity of Malaysian subjects. Am J Biochem Biotechnol 4(4): 367-370.
PREZA D, OLSEN I, AAS J, WILLUMSEN T, GRINDE B, PASTER B. 2008. Bacterial profiles of root caries in elderly patients. J Clin Microbiol 46(6): 2015-21.
ROLPH HJ, LENNON A, RIGGIO MP, SAUNDERS WP, MACKENZIE D, COLDERO L, BAGG J. 2001. Molecular Identification of Microorganisms from Endodontic Infections. J Clin Microbiol 39(9): 3282-89.
SAMARANAYAKE L. 2006. Microbes of relevance to dentistry. In Essential Microbiology for Dentistry. Third Edition. USA: Elsevier. Churchill Livingstone. p. 113-177.
SIMON L. 2007. The role of Streptococcus mutans and oral ecology in the formation of dental caries. Lethbridge Undergraduate Research Journal. p. 2.
SUCHODOLSKI JS, RUAUX CG, STEINER JM, FETZ K, WILLIAMS DA. 2004. Application of molecular fingerprinting for qualitative assessment of smallintestinal bacterial diversity in dogs. J Clin Microbiol 42(10): 4702-08.
TANZER JM, LIVINGSTON J, THOMPSON A. 2001. The Microbiology of Primary Dental Caries in Humans. J Dental Educ 65(10): 1028-38.
TORTORA GJ, FUNKE BR, CASE CL. 2007. The Prokaryotes: Domains Bacteria and Archaea. In: Microbiology: An Introduction. Ninth Edition. Singapore: Pearson Education, Inc. p. 312-340.
TURNER ML, SEGRE JA. 2009. Science. 2009 May 29; 324(5931): 1190-1192.
VICKERMAN MM, BROSSARD KA, FUNK DB, JESIONOWSKI AM, GILL SR. 2007. Phylogenetic analysis of bacterial and archaeal species in symptomatic and symptomatic endodontic infections. J Med Microbiol 56(1): 110-118.
WATANABE K, KODAMA Y, HARAYAMA. 2001. Design and evaluation of PCR primers to amplify bacterial 16S ribosomal DNA fragments used for community fingerprinting. J Microbiological Methods 44(3): 253-262.
[WHO] World Health Organization. 2003. The World Oral Health Report. Retrieved on 1 July 2009 from www2.paho.org/hq/dmdocuments/2009/OH_st_WHO.pdf
YOO SY, PARK SJ, JEONG DK, KIM KW, LIM SH, LEE SH, CHOE SJ, CHANG YH, PARK I, KOOK JK. 2007. Isolation and characterization of the Mutans Streptococci from the dental plaques in Koreans. J Microbiol 45(3): 246-255.
ZAURA E, KEIJSER BJF, HUSE SM , CRIELAARD W. 2009. Defining the healthy “core microbiome” of oral microbial communities. BioMed Central Microbiol 9: 259.