In vitro antivirulence activity of baicalin against Clostridioides difficile

Abstract

Introduction. Clostridioides difficile is an enteric pathogen that causes a serious toxin-mediated colitis in humans. Bacterial exotoxins and sporulation are critical virulence components that contribute to pathogenesis, and disease transmission and relapse, respectively. Therefore, reducing toxin production and sporulation could significantly minimize C. difficile pathogenicity and disease outcome in affected individuals. Aim. This study investigated the efficacy of a natural flavone glycoside, baicalin, in reducing toxin synthesis, sporulation and spore germination in C. difficile in vitro. Methodology. Hypervirulent C. difficile isolates BAA 1870 or 1803 were cultured in brain heart infusion broth with or without the subinhibitory concentration (SIC) of baicalin, and incubated at 37 °C for 24 h under strictly anaerobic conditions. The supernatant was harvested after 24 h for determining C. difficile toxin production by ELISA. In addition, a similar experiment was performed wherein samples were harvested for assessing total viable counts, and heat-resistant spore counts at 72 h of incubation. Furthermore, C. difficile spore germination and spore outgrowth kinetics, with or without baicalin treatment, was measured in a plate reader by recording optical density at 600 nm. Finally, the effect of baicalin on C. difficile toxin, sporulation and virulence-associated genes was investigated using real-time quantitative PCR. Results. The SIC of baicalin significantly reduced toxin synthesis, sporulation and spore outgrowth when compared to control. In addition, C. difficile genes critical for pathogenesis were significantly down-regulated in the presence of baicalin. Conclusion. Our results suggest that baicalin could potentially be used to control C. difficile , and warrant future studies in vivo.

Abatement of the dysfunctional hypothalamic–pituitary–gonadal axis due to ciprofloxacin administration by selenium in male rats

“…The limited reports available on the influence of selenium on ciprofloxacin effectiveness in vitro demonstrated that sodium selenite markedly enhanced the sensitivity of Clostridium difficile to ciprofloxacin leading to reduced bacterial growth in samples compared with antibiotic alone. [ 28 ] The greater effect of ciprofloxacin loaded selenium‐lipid nanoparticle in exterminating bacterial cells than the control has been attributed to its strong antimicrobial and antioxidant activities in the treatment of lung infection or diseases. [ 29 ] Moreover, coadministration of selenium and ciprofloxacin elicited a greater preventive effect on chronic bacterial prostatitis than ciprofloxacin alone in rats.…”

Protective Effect of Baicalin against Clostridioides difficile Infection in Mice

“…Although the fecal C. difficile counts on day 6 were comparable across the challenged mice groups treated with or without BC (5a (iii)), there was a significant reduction in the fecal toxin-mediated Vero cell cytotoxicity in the CD+BC2 mouse group ( Figure 5 b(ii)). The reduced CDI severity in baicalin-treated mice could be attributed to the inhibitory effect of baicalin on C. difficile toxin production, as observed in our in vitro studies [ 31 ]. In addition, baicalin is known to possess anti-inflammatory and anti-diarrheal properties [ 34 , 35 , 36 ], which could also have contributed to the improved clinical outcome in BC-administered mice.…”

“…Previous research conducted in our laboratory revealed that sub-inhibitory concentration of baicalin reduced C. difficile toxin production and cytotoxicity in vitro. Additionally, baicalin inhibited C. difficile spore germination and outgrowth [ 31 ]. The results from the current study translate our previous findings in vivo by demonstrating a dose-dependent reduction of CDI severity in BC-supplemented mice.…”

“…In this method, 15 mg of fecal material was weighed and mixed thoroughly by vortexing with 200 µL of sterile PBS and subsequently centrifuged at 14,000× g at 4 °C for 10 min (samples were stored at −80 °C if not used immediately). The supernatants of the fecal slurry were subjected to Vero cell cytotoxicity assay [ 31 ]. Fecal slurry supernatants were serially diluted by 1:10 up to a dilution of 1:100,000,000 onto confluent Vero cell monolayers in 96-well microtiter plates.…”

“…Baicalin (5,6-dihydroxy-7-O-glucuronide flavone) is a flavone glycoside present in the plant Scutellaria baicalensis Georgi, known to possess antimicrobial, antioxidant, and anti-inflammatory properties [ 24 , 25 , 26 , 27 , 28 , 29 , 30 ]. Previously, our laboratory demonstrated the use of baicalin as a potential anti- C. difficile therapeutic agent owing to its inhibitory effect on C. difficile toxin production with minimal effects on the growth of selected beneficial microbiota in vitro [ 31 ]. As a logical next step, this study investigated the prophylactic and therapeutic effects of baicalin against CDI in an in vivo model by focusing on the clinical course and host microbiome changes in mice.…”