TriTOX: A novel Trichomonas vaginalis assay platform for high-throughput screening of compound libraries

Abstract

Trichomonas vaginalis is a neglected urogenital parasitic protist that causes 170 million cases of trichomoniasis annually, making it the most prevalent non-viral, sexually transmitted disease. Trichomoniasis treatment relies on nitroheterocyclics, such as metronidazole. However, with increasing drug-resistance, there is an urgent need for novel anti-trichomonals. Little progress has been made to translate anti-trichomonal research into commercialised therapeutics, and the absence of a standardised compound-screening platform is the immediate stumbling block for drug-discovery. Herein, we describe a simple, cost-effective growth assay for T. vaginalis and the related Tritrichomonas foetus . Tracking changes in pH were a valid indicator of trichomonad growth ( T. vaginalis and T. foetus ), allowing development of a miniaturised, chromogenic growth assay based on the phenol red indicator in 96- and 384-well microtiter plate formats. The outputs of this assay can be quantitatively and qualitatively assessed, with consistent dynamic ranges based on Z′ values of 0.741 and 0.870 across medium- and high-throughput formats, respectively. We applied this high-throughput format within the largest pure-compound microbial metabolite screen (812 compounds) for T. vaginalis and identified 43 hit compounds. We compared these identified compounds to mammalian cell lines, and highlighted extensive overlaps between anti-trichomonal and anti-tumour activity. Lastly, observing nanomolar inhibition of T. vaginalis by fumagillin, and noting this compound has reported activity in other protists, we performed in silico analyses of the interaction of fumagillin with its molecular target methionine aminopeptidase 2 for T. vaginalis , Giardia lamblia and Entamoeba histolytica , highlighting potential for fumagillin as a broad-spectrum anti-protistal against microaerophilic protists. Together, this new platform will accelerate drug-discovery efforts, underpin drug-resistance screening in trichomonads, and contributing to a growing body of evidence highlighting the potential of microbial natural products as novel anti-protistals.

Novel Treatment Approaches to Combat Trichomoniasis, a Neglected and Sexually Transmitted Infection Caused by Trichomonas vaginalis: Translational Perspectives

“…In efforts to demonstrate the applicability of a colorimetric technique for detecting trichomonacidal activity, the authors identified a promising candidate from a vast library of 812 compounds. An inhibitor of methionine aminopeptidase 2, described as fumagillin, was one of the hit components identified via in vitro assay, with an IC 50 of 0.26 µM and with target action confirmed by in silico assays [44].…”

Fuligopyrones from the Fruiting Bodies of Myxomycete Fuligo septica Offer Short-Term Protection from Abiotic Stress Induced by UV Radiation

“…Fuligopyrone ( 1 ) and fuligopyrone B ( 2 ) were tested for biological activity against the bacteria Bacillus subtilis (ATCC 6633) and Staphylococcus aureus (ATCC 25923), the fungi Candida albicans (ATCC 10231) and Saccharomyces cerevisiae (ATCC 9763), the protozoan Tritrichomonas fetus (KV-1), the mammalian cell lines NS-1 murine myeloma and neonatal foreskin fibroblasts (NFF), and seeds of the plant Eragrostis tef ( Teff ), using previously described methods. , …”

Type-I Hemins and Free Porphyrins from a Western Australian Sponge Isabela sp.

“…The plates were incubated in anaerobic jars (Oxoid AG25) containing an Anaerogen satchel (Oxoid AN25) in a 37 °C (5% CO 2 ) incubator. At 48 h and 72 h, MIC end points were determined visually and absorbance was measured using Spectromax plate reader (Molecular Devices, Sunnyvale, CA, USA) at 570 nm [ 21 ].…”