ANTIBACTERIAL MECHANISM OF PLANT-DERIVED PHENOLICS AGAINST SALMONELLA ENTERICA SEROVAR TYPHIMURIUM

Salmonella enterica serovar Typhimurium (ST) remain one of the main bacterial pathogens responsible for illnesses, hospitalizations, and deaths in the USA. Its ubiquitous prevalence in nature, invasive pattern and increasing antibiotic resistance make it a public health threat, warranting the discovery of novel antimicrobials that can be implemented as either treatments or as forms of control. Plant-derived compounds have been proposed as potential antimicrobials that can be used against gram-negative pathogens, with phenolic acids being of interest for their prevalence in nature and bioactivity. This research studied the effects of gallic acid (GA), protocatechuic acid (PA) and vanillic acid (VA) against ST. Findings showed these compounds to be able to inhibit bacterial growth in vitro, while also showing a reduction in the expression of key virulence genes, without inducing resistance over multiple passages. Further studies using a human epithelial cell line for studying host-pathogen interactions, showed their capability to reduce the number of ST that were able to invade the host cells. Further studies were performed in cecal fluid to test their potency in more complex environments and assess their effects on the microbiome. When in cecal fluid, compounds showed a reduced inhibitory potency compared to in vitro, but still exerted antimicrobial pressure against ST. When analyzing relative abundance of other bacteria through 16S-rRNA gene sequencing, there was an overall decrease in the Protobacteria phylum, while no significant negative effect was seen for other phyla like that of the Firmicutes and Actinobacteria. Experiments to determine the mechanism of action against ST showed these phenolic acids to permeabilize the cell plasma membrane, in addition to reducing cell wall synthesis. Scanning electron microscopy showed treated bacteria to have dents at the polar ends of the cell, while others were found in a duplet formation, suggesting further disruption of specific bacterial functions associated to cell division and structure. These findings suggest that despite their similarities, these compounds are capable of exerting different types of antimicrobial pressure against ST that could better inform their future use as control measures against ST, and their potential use case based on the desired outcome

Antimicrobial and Antivirulence Impacts of Phenolics on Salmonella Enterica Serovar Typhimurium

Salmonella enterica serovar Typhimurium (ST) remains a major infectious agent in the USA, with an increasing antibiotic resistance pattern, which requires the development of novel antimicrobials capable of controlling ST. Polyphenolic compounds found in plant extracts are strong candidates as alternative antimicrobials, particularly phenolic acids such as gallic acid (GA), protocatechuic acid (PA) and vanillic acid (VA). This study evaluates the effectiveness of these compounds in inhibiting ST growth while determining changes to the outer membrane through fluorescent dye uptake and scanning electron microscopy (SEM), in addition to measuring alterations to virulence genes with qRT-PCR. Results showed antimicrobial potential for all compounds, significantly inhibiting the detectable growth of ST. Fluorescent spectrophotometry and microscopy detected an increase in relative fluorescent intensity (RFI) and red-colored bacteria over time, suggesting membrane permeabilization. SEM revealed severe morphological defects at the polar ends of bacteria treated with GA and PA, while VA-treated bacteria were found to be mid-division. Relative gene expression showed significant downregulation in master regulator hilA and invH after GA and PA treatments, while fliC was upregulated in VA. Results suggest that GA, PA and VA have antimicrobial potential that warrants further research into their mechanism of action and the interactions that lead to ST death.https://doi.org/10.3390/antibiotics910066