Abstract
OBJECTIVE: To develop and evaluate a nucleic acid amplification test for spectinomycin-resistant Neisseria gonorrhoeae (N. gonorrhoeae). METHODS: N. gonorrhoeae-specific primers NG1/NG2 and primers specific to the N. gonorrhoeae rpsE gene mutation (80_82 delTTA) were designed. Genomic nucleic acids of spectinomycin-sensitive and resistant N. gonorrhoeae, Escherichia coli, Pseudomonas aeruginosa, and Salmonella typhi were used as templates to be amplified by PCR and quantitative real-time PCR (qPCR). The sensitivity and specificity of the method were evaluated accordingly. RESULTS: The NG1/NG2 primers could effectively amplify specific fragments of N. gonorrhoeae, yielding negative results for the nucleic acid amplification test of the other types of bacteria tested. E64/E175R and E-87/E95R could effectively differentiate the wild type and mutant (80_82 delTTA) rpsE genes. In PCR reactions, the minimum limits of NG1/NG2, E64/E175R, and E87/E95R for the target genes were 414.8 copies, 414.8 copies, and 4.1 copies /μL, respectively, while those for qPCR reactions were 41.5, 41.5, and 4.1×10(－2) copies /μL, respectively. CONCLUSION: A nucleic acid amplification test for spectinomycin-resistant N. gonorrhoeae with high specificity and sensitivity was successfully established in this study, which is expected to provide support for the rapid diagnosis of N. gonorrhoeae infection and treatment decision-making in clinical settings.