Abstract
Purpose :
To determine the effects of glycyrrhizin (GLY) on virulence factors of multi-drug resistant systemic (MDR9) vs ocular (B1045) clinical isolates.
Methods :
Isolates were grown in 25ml of PTSB at 37oC for 18h and proteomes profiled using LC-MS/MS with label free quantitation by the WSU Proteomics Core. Mining factors included genes and proteins: Virulence and Quorum sensing (toxA, exoS, plcH, plcN, lasB, hcnC, aroE, rpoN, ccmg, sodB, phzS, 6cl5, lasI, lasR, rhlR); Antibiotic resistance (ampC, mexC); Biofilms-exopolysaccharides, proteins and proteinaceous surface appendages; adherence (pslA, pslB, pslC, pslD, pslE, pslF, pslG, pslH, pslI, pslJ, pslK, pslL, pslM, pslN, pslO, pelA, pelB, pelC, pelD, pelE, pelF, GacA, RetS, LadS, algL,T4P, CdrA, CupA, CupB, CupC, CupE, HptB, T3SS, T6SS).
Also, transfected human (HCET) and mouse (MCEC) corneal epithelial cells were seeded onto chambered slides and incubated overnight. Cell chambers were washed and media added without antibiotics. Each isolate was combined with GLY (0, 5 and 10 mg/mL) before application to the slides (10 MOI/cell). After 3 hr (37°C, aerobic conditions), slides were washed, stained (Wright-Giemsa), air dried, and cover slipped. Bacteria adherent to cells (n=100/group) were photographed, counted, averaged, and expressed as number bacteria/cell. Biofilm was tested by a colorimetric microtiter plate assay.
Results :
3,254 proteins were identified and 3,211 were quantified. Using a cutoff of 0.1 for false-discovery corrected p-values, 1,355 proteins were different between GLY treated vs untreated MDR9 and 5 between GLY treated vs untreated B1045. In MDR9 almost half of the proteome was affected whereas B1045 was minimally affected. Principal component analysis and hierarchical clustering distinguished treated from untreated MDR9 samples but yielded mixed results for B1045. However, GLY prevented adherence to both groups of cells and decreased biofilm.
Conclusions :
GLY had a greater effect on MDR9 vs B1045 at the proteomic level, but was similar for each at disrupting adherence, the first step in biofilm formation.
This is a 2020 ARVO Annual Meeting abstract.