Visible colorimetric growth indicators of Neisseria gonorrhoeae for low-cost diagnostic applications

Abstract

N. gonorrhoeae is one of the most pressing antibiotic resistant threats of our time and low-cost diagnostics that can easily identify antibiotic resistance are desperately needed. However, N. gonorrhoeae responds so uniquely to growth conditions that it cannot be assumed gonorrhea will respond to common microbiological methods used for other pathogenic organisms. In this paper, we explore visual colorimetric indicators of N. gonorrhoeae growth that can be seen without a microscope or spectrophotometer. We evaluate growth media, pH indicators, resazurin-based dyes, and tetrazolium-based dyes for their use in simple colorimetric system. Overall, we identified Graver Wade media as the best at supporting robust gonococcal growth while also providing the least background when analyzing results of colorimetric tests. XTT, a tetrazolium-based dye, proved to show to brightest color change over time and not negatively impact the natural growth of N. gonorrhoeae. However, other dyes including PrestoBlue, MTT, and NBT are less expensive than XTT and work well when added after bacterial growth has already occurred. By identifying the specific use cases of these dyes, this research lays the groundwork for future development of a color-based antibiotic susceptibility low-cost test for N. gonorrhoeae.

Multiplexed paper-based assay for personalized antimicrobial susceptibility profiling of Carbapenem-resistant Enterobacterales performed in a rechargeable coffee mug

“…1b). PrestoBlue is a redox dye that starts as blue colored resazurin and is reduced to resorufin, a pink product, or further reduced to dihydroresorufin, a colorless product 18,19 . In contrast, XTT in solution starts colorless and is reduced to a vibrant orange, especially in the presence of additional electron acceptors such as phenazine methosulfate.…”

Activity-Based Diagnostics: Recent Advances in the Development of Probes for Use with Diverse Detection Modalities

“…74 Additionally, MTT and its analogs have been shown to be processed in a variety of bacterial infections, including sepsis, 75 UTIs, 76 and more recently Neisseria gonorrheae. 77 However, the clinical application of these dyes is limited due to the lack of specificity to any one particular indication, potentially confounding results in complex biological samples. 78 A limitation of colorimetric diagnostic assays is the amount of signal required to be readily viewed by the eye.…”

“…This method was recently employed to detect succinate dehydrogenase activity in sperm, as low enzymatic activity is a biomarker for low total motile sperm concentration (TMSC) . Additionally, MTT and its analogs have been shown to be processed in a variety of bacterial infections, including sepsis, UTIs, and more recently Neisseria gonorrheae . However, the clinical application of these dyes is limited due to the lack of specificity to any one particular indication, potentially confounding results in complex biological samples …”