ARBITRARILY PRIMED POLYMERASE CHAIN-REACTION FINGERPRINTING AND CLONAL ANALYSIS OF ORAL FUSOBACTERIUM-NUCLEATUM ISOLATES

F. nucleatum is the most commonly isolated microorganism from subgingi val plaque, but the role of this microorganism in periodontal diseases remains undefined. Arbitrarily primed polymerase chain reaction (AP-P CR) was evaluated as a method for fingerprinting F. nucleatum isolates and for use in clonal analysis. Pulsed field gel electrophoresis was used to further differentiate F. nucleatum isolates, with identical AP -PCR patterns. Extremely heterogeneous AP-PCR fingerprints were observ ed among the 98 F. nucleatum isolates, with 36 different genotypes obs erved with primer C1 and 30 different genotypes detected with primer C 2. Combining the results of the AP-PCR genotype analysis from C1 and C 2 primer amplifications revealed that up to 7 different genotypes coul d be distinguished from isolates from the same oral cavity and that up to 4 different genotypes were observed within a single site. An inten se amplicon at approximately 450 bp generated in AP-PCR amplification with primer C2 was associated with F. nucleatum subsp. nucleatum (ATCC 25586) and with 15 F. nucleatum isolates from diseased sites and 2 is olates from healthy sites. Pulsed field gel electrophoresis confirmed the AP-PCR genotypes and demonstrated increased discriminatory power o ver AP-PCR. The results indicated that AP-PCR and pulsed field gel ele ctrophoresis provide a simple and sensitive means for differentiating oral F. nucleatum isolates and further demonstrate the heterogeneity o f this species. These techniques may serve as useful tools in the clon al and epidemiological analysis of F. nucleatum isolates, which may he lp define the role of these microorganisms in periodontal diseases.