Small interfering RNAs targeting agrA and sarA attenuate pathogenesis of Staphylococcus aureus in Caenorhabditis elegans.

Terissa Alexas Thompson,Paul Dean Brown

doi:10.3855/jidc.13664

Abstract

The use of small interfering RNA (siRNA) gene silencing is a promising therapeutic option as it does not impose selective pressure on bacteria that is often associated with the development of resistance. The study assessed the effect of siRNA targeted to sarA and agrA in S. aureus and the relationship between the transcriptional response, biofilm formation and pathogenicity. siRNAs designed against agrA and sarA were electroporated into methicillin-resistant and methicillin-susceptible S. aureus strains. mRNA levels, growth kinetics, biofilm formation and minimal inhibitory concentration were measured. Efficacy of siRNA in bacteria was assessed using survival assays in a C. elegans model. Differences in gene expression before and after siRNA treatment were anaysed using the paired t-test, while the log rank test was used to assess the significance of any difference among survival rates of nematodes. Biofilm formation decreased significantly in siRNA treated strains and growth rates of siRNA treated strains were significantly higher compared to untreated strains. We observed significant decreases in the transcriptional response in siRNA treated strains, with concomitant significant increases in the lifespan of C. elegans worms exposed to siRNA-treated versus untreated strains. siRNA targeted to agrA and sarA lowered mRNA transcription and pathogenicity of S. aureus.

Highlights

The use of small interfering RNA gene silencing is a promising therapeutic option as it does not impose selective pressure on bacteria that is often associated with the development of resistance
The growth kinetics of strains exposed to small interfering RNA (siRNA) treatment showed shorter lag phases and longer exponential phases
Effect on gene expression The extent of incorporation of labelling of cytoplasmic contents siRNA in the various strains are given in Table 3. mRNA production in HA-MRSA+siRNAagrA and HAMRSA+siRNAagrA and sarA decreased when compared to untreated HA-MRSA (Figure 2, top), with the lowering in the latter treatment group being significant (p < 0.05)

Summary

Introduction

The use of small interfering RNA (siRNA) gene silencing is a promising therapeutic option as it does not impose selective pressure on bacteria that is often associated with the development of resistance. The study assessed the effect of siRNA targeted to sarA and agrA in S. aureus and the relationship between the transcriptional response, biofilm formation and pathogenicity. Conclusions: siRNA targeted to agrA and sarA lowered mRNA transcription and pathogenicity of S. aureus. Knowledge of the signal transduction systems involved in bacterial pathogenesis have provided insights regarding gene expression at different stages of infection [5]. The accessory gene regulator (agr) operon is a TCSTS in S. aureus and is one of the most well studied systems of bacterial human pathogenesis. S. aureus occupies numerous niches within its host, a feature that is tightly regulated by the selective expression of virulence genes [7], and the agr operon is central to this regulation [8,9]. P3 activation leads to RNAIII production that results in the up-regulation of secreted toxins to favour infection and the down-regulation of adhesins and surface proteins to facilitate the switch from colonization to invasion [5,12]

Methods

Results

Conclusion