Anti-Shigella activity and killing kinetics of compounds from MMV Pathogen and COVID Boxes: Identifying leads against Shigellosis

Abstract

Shigellosis is a leading diarrheal disease, posing critical public health challenges in low- and middle-income countries. Rising antibiotic resistance exacerbates treatment limitations. We screened 560 compounds from the MMV Pathogen and COVID Boxes for in vitro activity against Shigella flexneri , S. sonnei , and S. dysenteriae via resazurin-based microdilution. Cytotoxicity (RAW 264.7 cells) and killing kinetics against S. flexneri were assessed. From the primary screen, 13 compounds (9 from the Pathogen Box, 4 from the COVID Box) showed potent anti- Shigella activity against S. flexneri (MIC ≤10 μM). Among these, seven non-antibiotic Pathogen Box compounds exhibited low-micromolar MICs (0.31–5 μM) and low cytotoxicity (CC 50 >100 μM). The known antibiotic levofloxacin (MMV687798) showed broad-spectrum activity (MIC 0.15–2.5 μM). MMV1804312 (salinomycin) was the most active COVID Box hit (MIC 1.25 μM). Time-kill assays revealed rapid, concentration-dependent bactericidal activity (≥3-log reduction within 4–6 h) for Pathogen Box compounds MMV675997 and MMV021660, while MMV000858 displayed bacteriostatic action. This study identifies several novel, non-antibiotic leads from the MMV Pathogen Box with potent anti- Shigella activity and favorable cytotoxicity profiles, offering promising starting points for further development against drug-resistant shigellosis. • Thirteen anti-Shigella compounds identified from MMV libraries. • Seven novel non-antibiotic leads show potent anti-Shigella activity. • Selected leads exhibit low MICs (≤5 μM) and low cytotoxicity (CC 50 > 100 μM) • Six Pathogen Box compounds (including levofloxacin) show ≥3-log kill within 4–6 h.