Evaluation of a fluorescent DNA hybridization assay for the detection ofNeisseria gonorrhoeae

This study evaluates a four-hour fluorescent DNA hybridization assay using both known bacterial isolates and clinical specimens. A biotinylated oligonucleotide probe from a sequence of the plasmid-encoded genecppB was used. Hybrids were detected by addition of a streptavidin-alkaline phosphatase conjugate, followed by incubation for 30 min in a fluorescent substrate for alkaline phosphatase. The level of detection of the fluorescent assay was 0.1 pg of cryptic plasmid DNA or 200 cfu of the plasmid-containing strain NG 34/85 ofNeisseria gonorrhoeae. A total of 119 reference strains ofNeisseria gonorrhoeae and other related bacteria were tested for reactivity with the probe. AllNeisseria gonorrhoeae strains, including eight plasmid-free strains, hybridized with the probe. Fluorescence ratios were 2.67 for plasmid-free strains and 3.85 for plasmid-containing strains. Of the heterologous microorganisms tested, only one of six strains ofNeisseria cinerea gave a fluorescence ratio above the 2.0 cut-off value for positivity with the probe at a cell density of 1 × 10⁴ cfu. The probe was also evaluated using clinical specimens from 100 patients attending a clinic for sexually transmitted diseases. The sensitivity of the assay was 100 % while the specificity was 97.5 %. Positive and negative predictive values were 91.2 % and 100 %, respectively. The fluorescent DNA hybridization assay for the detection ofNeisseria gonorrhoeae described here thus appears to be a highly specific and sensitive assay.