Novel Anti-Campylobacter Compounds Identified Using High Throughput Screening of a Pre-selected Enriched Small Molecules Library

Abstract

Campylobacter is a leading cause of foodborne bacterial gastroenteritis worldwide and infections can be fatal. The emergence of antibiotic-resistant Campylobacter spp. necessitates the development of new antimicrobials. We identified novel anti-Campylobacter small molecule inhibitors using a high throughput growth inhibition assay. To expedite screening, we made use of a “bioactive” library of 4182 compounds that we have previously shown to be active against diverse microbes. Screening for growth inhibition of Campylobacter jejuni, identified 781 compounds that were either bactericidal or bacteriostatic at a concentration of 200 μM. Seventy nine of the bactericidal compounds were prioritized for secondary screening based on their physico-chemical properties. Based on the minimum inhibitory concentration against a diverse range of C. jejuni and a lack of effect on gut microbes, we selected 12 compounds. No resistance was observed to any of these 12 lead compounds when C. jejuni was cultured with lethal or sub-lethal concentrations suggesting that C. jejuni is less likely to develop resistance to these compounds. Top 12 compounds also possessed low cytotoxicity to human intestinal epithelial cells (Caco-2 cells) and no hemolytic activity against sheep red blood cells. Next, these 12 compounds were evaluated for ability to clear C. jejuni in vitro. A total of 10 compounds had an anti-C. jejuni effect in Caco-2 cells with some effective even at 25 μM concentrations. These novel 12 compounds belong to five established antimicrobial chemical classes; piperazines, aryl amines, piperidines, sulfonamide, and pyridazinone. Exploitation of analogs of these chemical classes may provide Campylobacter specific drugs that can be applied in both human and animal medicine.

Incorporation of Selected Antimicrobial Small Molecule Compounds into Tapioca Starch and the Effects on Thickness, Moisture, and Oxygen Mass Transfer, and Mechanical Properties of the Films

“…These tests determined the lowest concentrations of each compound to cause the death of human colon cells after a 72 h treatment with each of the compounds separately, using a sulphorodhamine assay. Details of these determinations are reported in a previous publication . The results showed that the values for JA‐144, TH‐4, and TH‐8 were 200 µM.…”

“…These microorganisms appeared not to develop antimicrobial resistance to these small molecules, when compared to other regularly used antimicrobial agents. These small molecule compounds were selected after a high throughput screening of a library of ≈4,200 compounds …”

Novel small molecule modulators of quorum sensing in avian pathogenic Escherichia coli (APEC)

“…Bioluminescence of SMs treated culture supernatants were compared to DMSO treated control. The Z-score was calculated to evaluate the quality of the bioluminescence screening [56]. Four independent experiments were conducted for the 69 compounds that inhibited ≥ 75% of the AI-2 mediated bioluminescence and the average inhibition percentage was calculated.…”

“…The selected 10 AI-2 inhibitors were evaluated for their toxicity to Caco-2 (ATCC® HTB-37™) using LDH assay [56]. Briefly, Caco-2 cells (1.4 × 10 5 cells/well) were grown in a 96-well plate in minimal essential medium (MEM) supplemented with 20% fetal bovine serum (FBS; Gibco), 1% non-essential amino acid (NEAA, Invitrogen Life Technologies) and 1mM sodium pyruvate, at 37°C in a humidified 5% CO 2 incubator for 48 h until a complete monolayer was formed.…”

Moderate-Throughput Identification and Comparison of Campylobacter-Infecting Bacteriophages

“…Taken together, these observations indicate that the development of novel antimicrobials for the reduction or treatment of human Campylobacter infections is necessary (7). Several avenues of reducing the incidence of human campylobacteriosis have been pursued at various levels of the infectious cycle (3).…”