Detection ofNeisseria Gonorrhoeae by polymerase chain reaction (PCR)

Abstract

Three different sets of primers were designed using FASTA homology search and PRIMERSELECT for the specific detection ofNeisseria gonorrhoeae using polymerase chain reaction (PCR). These primers amplified the highly conserved regions of genes for Open Reading Frame (ORF), Outer Membrane Protein (OMP) and 23S rRNA sequences ofN. gonorrhoeae. Each of the PCR primer set was evaluated using the DNA samples isolated from eight different positive isolates ofN. gonorrhoeae cultured from urethral swabs of patients visiting Maulana Azad Medical College and Safdarjung Hospital. Amplification products were analyzed on agarose gel electrophoresis. Two sets of PCR primers, designated as Ngu1/Ngu2 and Ngu5/Ngu6, specific for ORF and OMP gene respectively, amplified four regions of the gene which may help to differentiate the various strains ofN. gonorrhoeae infecting indigenous population. In contrast, a single, specific PCR product of 650 bp was visualized on agarose gel with primers Ngu3/Ngu4, amplifying the 23S rRNA gene. Under optimum conditions, as low as 25ng of DNA isolated from eight different clinical strains ofN. gonorrhoeae could be detected by PCR using Ngu3/Ngu4 set of primers. Our results suggested that Ngu3/Ngu4 could serve as good primers for the specific, reproducible and sensitive diagnosis ofNeisseria gonorrhoeae from clinical samples.