Abstract
Neisseria gonorrhoeae is a host-adapted human pathogen that causes sexually transmitted gonorrhea and remains to be a serious global public health challenge, especially in low- and middle-income regions. It is vital to devise a reliable, simple, cost-saving, and easy-to-use assay for detecting the N. gonorrhoeae agent. In the current study, we firstly report a novel approach, loop-mediated isothermal amplification linked with a polymer nanoparticle–based biosensor (LAMP-PNB), that was used for identifying N. gonorrhoeae in clinical samples. The results showed that the LAMP primers based on the orf1 gene were valid for development of the N. gonorrhoeae-LAMP-PNB assay. The detection system with optimal conditions could be performed at a fixed temperature of 64°C for 40 min. The whole process, including genomic DNA preparation (approximately 10 min), LAMP reaction (40 min), and PNB reporting (approximately 2 min), could be accomplished within 60 min. The limit of detection (LoD) of the N. gonorrhoeae-LAMP-PNB assay was 50 copies per test. The specificity of the current assay was 100%, and no cross-reactions to non–N. gonorrhoeae isolates were observed. These results confirmed that the N. gonorrhoeae-LAMP-PNB technique is a reliable, specific, sensitive, rapid, low-cost, and easy-to-use method for detecting gonococci isolates. More importantly, this assay has great potential to develop a point-of-care (POC) testing method in clinical practice, especially in resource-constrained regions.
Highlights
Neisseria gonorrhoeae, a host-adapted human pathogen, is the causative agent of gonorrhea, which belongs to the most frequent sexually transmitted infections that remain one of the major global public health concerns (Quillin and Seifert, 2018; Hook and Bernstein, 2019)
The loop-mediated isothermal amplification linked with the polymer nanoparticle–based biosensor (LAMP-PNB) technique was devised firstly for simple, specific, sensitive, rapid, and visual identification of N. gonorrhoeae by targeting the orf1 gene (Chaudhry and Saluja, 2002; Edwards et al, 2014), and it showed no homology with other microbial genomes in GenBank by Basic Local Alignment Search Tool (BLAST) searches
Using the PNB, the test line (TL) and control line (CL) were observed in the detection region, indicating positive results
Summary
A host-adapted human pathogen, is the causative agent of gonorrhea, which belongs to the most frequent sexually transmitted infections that remain one of the major global public health concerns (Quillin and Seifert, 2018; Hook and Bernstein, 2019). According to World Health Organization (WHO) estimations, there are around 87 million new infections worldwide each year (Rowley et al, 2019). The vast majority of gonococcal infections (>80 million) are in developing countries of Africa, Asia, and Latin America (World Health Organization, 2016; Rowley et al, 2019). Because the various clinical symptoms of gonorrhea are largely not specific, and most gonococcal infections are in resource-constrained regions, developing a specific, sensitive, rapid, and cost-saving assay for the accurate identification of N. gonorrhoeae isolates is necessary for reducing ongoing gonorrhea transmission
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