Denaturing gradient gel electrophoresis as a diagnostic tool in periodontal microbiology.
Bacteria play an important role in the initiation and progression of periodontal diseases and are part of a biofilm, which can contain over 100 different species. The aim of the present study was to show the potential of denaturing gradient gel electrophoresis (DGGE) as a tool for the detection of clinically relevant species and to compare the results of detection by DGGE with those by PCR and culturing. Hybridization of the bands from the DGGE profiles with species-specific probes was developed to confirm the band positions in the marker obtained with reference strains. The sensitivities of DGGE compared to those of cultivation for the detection of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, and Tannerella forsythensis were 100, 100, 88, and 100%, respectively; and the sensitivities of DGGE compared to those of PCR were 100, 90, 88, and 96%, respectively. DGGE as a diagnostic tool could easily be extended to other species, as shown for Treponema denticola, which could be detected in 48% of the samples. Three different groups of A. actinomycetemcomitans serotypes could be distinguished by DGGE (i.e., a group comprising serotypes a, d, e, and f; a group comprising serotype b; and a group comprising serotype c). Amplicons from P. gingivalis and T. denticola migrated to the same position in the gel, and P. intermedia produced multiple bands. In the present study we show that the DGGE profiles represent clinically relevant species which can be detected by hybridization with species-specific probes. With DGGE, large numbers of samples can be analyzed for different species simultaneously, and DGGE may be a good alternative in periodontal microbial diagnostics.
Zijnge
Vincent
V
Department of Medical Microbiology, University Medical Center Groningen, Hanzeplein 1, P.O. Box 30.001, 9700 RB Groningen, The Netherlands. h.j.m.harmsen@med.umcg.nl.
Welling
Gjalt W
GW
Degener
John E
JE
van Winkelhoff
Arie Jan
AJ
Abbas
Frank
F
Harmsen
Hermie J M
HJ
D001419:Bacteria / Q000145:classification* / Q000302:isolation & purification*; D001431:Bacteriological Techniques / Q000379:methods*; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D005881:Gingiva / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D013045:Species Specificity