Protective Signatures of Roselle (Hibiscus sabdariffa L.) Calyx Fractions against Staphylococcus aureus in Drosophila Infection Model

Abstract

The rise of antibiotic-resistant Staphylococcus aureus-related clinical cases is an alarming chronicle for global communities. This research was conducted to examine the antistaphylococcal effect of roselle (Hibiscus sabdariffa L.) calyx fractions in the Drosophila model. In the infection experiment, wild-type and immunodeficient Drosophila were pricked with S. aureus and subsequently subjected to fly survivorship and colony-forming assays, in the presence or absence of roselle calyx fractions. The Involvement of immune stimulation in the host antibacterial protection was assessed in vitro using cell-based luciferase reporter assay and in vivo using RT-qPCR analysis on adult flies. A declining rate of fly survivorship and augmentation of bacterial growth were observable in S. aureus-infected wild-type flies but subject to improvement in the presence of roselle calyx fractions. Cell-based analysis revealed the absence of host immune stimulation via Drosophila Toll pathway and roselle calyx fractions-treated immune-deficient flies lacking for components in the Toll pathway were protected from infection-induced early death phenotype and harbored reduced number of S. aureus colonies. Overall, our data confirmed the in vivo anti-staphylococcal activity of roselle calyx fractions in Drosophila infection model and such protective signature was devoid of host immune stimulation.

Proof-of-Concept Preclinical Use of Drosophila melanogaster in the Initial Screening of Immunomodulators

“…These two compounds have been reported for their immunomodulatory activities in mammals [19][20][21]29,30] as well as their antibacterial activities [31][32][33]. We have shown that D. melanogaster can be used to screen for drug candidates with antibacterial activities [24][25][26][27]34,35]. In this study, we extend our study to investigate whether D. melanogaster can be used as an initial model organism to screen for immunostimulants and antibacterial drugs candidates before further examination in the mammalian animal models.…”

“…The bacterial inoculum was collected, washed carefully with phosphate-buffered saline (PBS), diluted equal to 0.5 McFarland density, and suspended in PBS before use. Fly infection experiment was performed on the thorax of D. melanogaster by using the pricking method, as described previously [24][25][26][27]. Briefly, adult male fruit flies that were 4-7 days old (30 flies per group of treatment) were pricked using a tungsten fine needle that had been previously dipped in the S. aureus culture (100-fold dilution of 0.5 McFarland density).…”

“…Briefly, adult male fruit flies that were 4-7 days old (30 flies per group of treatment) were pricked using a tungsten fine needle that had been previously dipped in the S. aureus culture (100-fold dilution of 0.5 McFarland density). All pricked flies were maintained at 29 • C and subjected to assessment on fly survivorship, as described previously [24][25][26][27].…”

Enhancement in Site-Specific Delivery of Chloramphenicol Using Bacterially Sensitive Microparticle Loaded Into Dissolving Microneedle: Potential For Enhanced Effectiveness Treatment of Cellulitis

“…The absence of these components thus prevents the production of AMPs, which is necessary for the activation of humoral innate immunity in response to SA infection in Drosophila. 51,52 This results in a condition that resembles immunodeficiency, facilitating the easy preparation of an infection model. Such mutant flies have been demonstrated to succumb to pathogens faster than their wild-type counterparts.…”

“…Based on Figure 7C, infected larvae without treatment showed larval death; only a few survived to become pupae and adult flies. According to several studies, the increased mortality rate of the dying host has been connected to bacterial load, 52 since giving CHL-loaded MPs, which was mixed on fly food, increased the survival rate of infected Drosophila larva. Because this finding is not significantly different than in healthy controls (p > 0.05), it is tempting to assume that this phenotype was caused by the prevention of the bacterial growth in vivo model.…”