Detection of identical ribotypes of Porphyromonas gingivalis in patients residing in the United States, Sudan, Romania and Norway

Abstract

Porphyromonas gingivalis has been isolated from periodontitis lesions in subjects from many geographical locations. The purpose of this investigation was to determine whether similar ribotypes of P. gingivalis could be detected among strains isolated in different countries. A total of 198 isolates of P. gingivalis were obtained from 52 periodontitis patients in Boston (130 isolates), Bergen, Norway (17 isolates), Khartoum, Sudan (26 isolates), and Bucharest, Romania (25 isolates). DNA was isolated from each strain, cut separately by the restriction endonucleases KpnI and PstI. The resulting preparations were subjected to electrophoresis in a 0.8% agarose gel using a Tris-acetate EDTA buffer. Uncut lambda and a 1000-bp fragment of 16S rRNA were included as internal standards in each lane. In addition, a HindIII digest of lambda was present in a separate lane in each run. The DNA fragments were transferred to a nylon membrane by downward capillary transfer. 16S rRNA bands were detected using a 1000-kb digoxigenin-labelled probe generated by a polymerase chain reaction. At the same time, a digoxigenin-labelled probe to lambda was employed to detect the internal and molecular weight standards. The bands were detected using antibody to digoxigenin conjugated to alkaline phosphatase and chemiluminescence. The positions of the bands relative to the internal standards were determined and normalized to correct for run-to-run variations, and the molecular weight of each band was determined by comparison with standards within each gel. The resulting data for the 2 enzymes were combined and subjected to cluster analysis using an average unweighted linkage sort. In some instances, isolates that appeared to be of identical ribotype using one endonuclease gave different ribotypes using the other. Strains of P. gingivalis within a subject were usually identical, except for 3 patients who harbored 2 different ribotypes/individual. All subsequent analyses employed a single ribotype strain for each subject. A total of 32 ribotypes were observed for isolates from distant countries. A total of 11.5% of the patients had isolates exhibiting the same ribotype: ribotype 7a. Identical ribotypes of P. gingivalis can be recovered from subgingival plaque samples of periodontitis patients in different countries.