No abstract

Eighty-nine strains of Haemophilus ducreyi from a chancroid epidemic in Orange County, California, were examined for plasmid content. Seventy-eight (88%) of these isolates were found to contain a plasmid of 3.2 megadaltons which conferred P-lactamase production. Restriction endonuclease digests indicated that this was the same plasmid that was found in some strains of ,-lactamase-producing Neisseria gonorrhoeae.Plasmids of 5.7 and 7.0 megadaltons (MD) mediating ampicillin resistance in Haemophilus ducreyi have been previously described (2,3,8). In addition, a 3.6-MD plasmid mediating 1-lactamase production has also been reported (7,15). In this study, we have demonstrated a 3.2-MD plasmid encoding for ,-lactamase production in H. ducreyi isolates from Orange County, California. This plasmid appears to be identical to the plasmid found in Neisseria gonorrhoeae previously linked epidemiologically to West Africa (1,12,14).A total of 89 strains of H. ducreyi from a chancroid epidemic in Orange County, California (4), were studied. Isolates were examined for P-lactamase activity with the chromogenic cephalosporin nitrocefin (11). MICs were determined by a modification of a method previously described (10), using Mueller-Hinton broth supplemented with 10% fetal bovine serum. Microdilution plates were incubated in candle extinction jars at 35°C. Isolates were screened for plasmid content by electrophoresing ethanol-precipitated DNA from cleared lysates through a 0.7% agarose gel, as previously described (9). Gels were stained with ethidium bromide and viewed with a UV transilluminator (Fotodyne model 3-4400; New Berlin, Wis.). Seventy-eight (88%) isolates were found to be ,-lactamase positive, and the MICs were determined to be .8 ,ug/ml for ampicillin with each of these strains. All 78 of the isolates that demonstrated Plactamase activity were found to contain a 3.2-MD plasmid when compared with plasmid standards of known molecular size (Fig. 1).Purified plasmid DNAs from both an Orange County H. ducreyi isolate and a strain of N. gonorrhoeae containing the previously described 3.2-MD plasmid were prepared with cesium chloride-ethidium bromide density gradients. These two types of DNA preparations were used to transform Escherichia coli HB101 to ampicillin resistance by the calcium chloride-heat shock method (5). Purified plasmid DNAs from HB101 containing H. ducreyi(pKC83) and HB101 containing N. gonorrhoeae(pGR167) were compared by digestion with several restriction endonucleases. The digestion patterns obtained with three enzymes (TaqI, AluI, and Hinfl) are shown in Fig. 2. The cleavage patterns and fragment sizes obtained for both plasmids were identical.Although the 3.2-and 4.4-MD plasmids described in N. gonorrhoeae are molecular derivatives of the 5.7-and 7.0-MD plasmnids found in H. ducreyi (2), it was not clear from * Corresponding author.other reports (7,15)