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A Fluorescence-based Method to Study Bacterial Gene Regulation in Infected Tissues
Published on: February 19, 2019
An induced mutation of ABC-transporter component VraF(K84E) contributes to vancomycin resistance and virulence in
Ruobing Cao1, Huimin Su2, Zichun Wei1
1Institute of Advanced Technology, University of Science and Technology of China, Hefei, China.
Abstract:
Staphylococcus aureus is a notorious pathogen responsible for various severe diseases. Due to the emergence of drug-resistant strains, the prevention and treatment of S. aureus infections have become increasingly challenging. Vancomycin is considered to be one of the last-resort drugs for treating most methicillin-resistant S. aureus (MRSA), so it is of great significance to further reveal the mechanism of vancomycin resistance. VraFG is one of the few important ABC (ATP-binding cassette) transporters in S. aureus that can form TCS (two-component systems)/ABC transporter modules. ABC transporters can couple the energy released from ATP hydrolysis to translocate solutes across the cell membrane. In this study, we obtained a strain with decreased vancomycin susceptibility after serial passaging and selection. Subsequently, whole-genome sequencing was performed on this laboratory-derived strain MWA2 and a novel single point mutation was discovered in vraF gene, leading to decreased sensitivity to vancomycin and daptomycin. Furthermore, the mutation reduces autolysis of S. aureus and downregulates the expression of lytM, isaA, and atlA. Additionally, we observed that the mutant has a less net negative surface charge than wild-type strain. We also noted an increase in the expression of the dlt operon and mprF gene, which are associated with cell surface charge and serve to hinder the binding of cationic peptides by promoting electrostatic repulsion. Moreover, this mutation has been shown to enhance hemolytic activity, expand subcutaneous abscesses, reflecting an increased virulence. This study confirms the impact of a point mutation of VraF on S. aureus antibiotic resistance and virulence, contributing to a broader understanding of ABC transporter function and providing new targets for treating S. aureus infections.
Insights
A novel mutation in the Staphylococcus aureus VraF gene reduces susceptibility to vancomycin and daptomycin. This VraF mutation also increases virulence, offering new therapeutic targets for drug-resistant S. aureus infections.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Staphylococcus aureus infections pose a significant threat due to rising antibiotic resistance.
- Vancomycin is a critical last-resort antibiotic for treating methicillin-resistant S. aureus (MRSA).
- Understanding vancomycin resistance mechanisms is crucial for developing effective treatments.
Purpose of the Study:
- To investigate the genetic basis of decreased vancomycin susceptibility in Staphylococcus aureus.
- To elucidate the role of ABC transporters, specifically VraFG, in antibiotic resistance.
- To explore the impact of identified mutations on S. aureus virulence and cell surface properties.
Main Methods:
- Serial passaging and selection to generate antibiotic-resistant strains.
- Whole-genome sequencing to identify genetic mutations.
- Gene expression analysis (lytM, isaA, atlA, dlt operon, mprF).
- Assessment of cell surface charge, autolysis, and hemolytic activity.
Main Results:
- A single point mutation in the vraF gene was identified in a vancomycin- and daptomycin-tolerant S. aureus strain.
- The vraF mutation reduced bacterial autolysis and downregulated key autolytic enzyme genes.
- The mutation altered cell surface charge, increased expression of dlt operon and mprF, enhanced hemolytic activity, and expanded abscess formation, indicating increased virulence.
Conclusions:
- A point mutation in VraF significantly impacts Staphylococcus aureus antibiotic resistance and virulence.
- This study highlights the role of ABC transporters in mediating resistance and virulence.
- The findings provide potential new targets for combating S. aureus infections.
