Evolution of resistance mechanisms and biological characteristics of rifampicin-resistant Staphylococcus aureus strains selected in vitro

Abstract

Background We aimed to determine the evolutionary pathways of rifampicin resistance in Staphylococcus aureus, and the impact of resistance mutations in the rpoB gene on fitness. Methods Three clinical strains and one reference strain were used to select for rifampicin-resistant S. aureus variants. The mutations responsible for rifampicin resistance in all of the selected isolates in vitro were investigated by polymerase chain reaction (PCR) and DNA sequencing. To compare the fitness cost of rpoB mutations against their corresponding original isolates, we performed bacterial growth curve assays, static biofilm assays, in vitro competition experiments and an infection model of Galleria mellonella larvae. Results We obtained four rifampicin-resistant S. aureus isolates that showed high levels of resistance to rifampicin with a minimal inhibitory concentration (MIC) of 128 mg/L, and all isolates had a mutation at position 481 (H481F/Y) in RpoB. A broth microdilution assay indicated that mutation of H481F/Y did not affect susceptibility to common antibacterial drugs but slightly increased the vancomycin MIC. To identify the pathways involved in the development of rifampicin resistance, 32 variants (eight mutants for each strain) and four original isolates were selected for gene sequencing. Different generations of isolates were found to harbor various mutations sites. Compared with the corresponding original isolates, an in vitro fitness assay of the variant isolates showed that growth and virulence were reduced, with a statistically significantly decreased fitness, whereas the capacity for biofilm formation was elevated. Conclusions Our findings suggested that the acquisition of rifampicin resistance in S. aureus was dynamic and was associated with a significant fitness cost.

Antimicrobial and Transconjugants Characteristics of Sul3 Positive Escherichia Coli Isolated from Animals in Nanning, Guangxi Province

“…And compared with other studies [29][30][31], the recombinant plasmid had no effect on the growth performance of the strain, just like the wild plasmid in this study, but the wild plasmids reduced the competitiveness of host bacteria in vitro to a greater extent. Although the types and quantities of drugresistant genes studied are different, this also indicates that the adaptation cost of wild plasmids will bring greater adaptive cost to the recipient bacteria due to multiple drug-resistant genes or other unknown genes.…”

“…In vitro competition experiments refer to previous descriptions [36]. First, two kinds of bacteria were cultured to 0.5 McFarland, then took 100 µL for each mixed in a 1:1 ratio and added to 10 mL LB broth, and incubated for 16 h at 37℃, 220 r/min.…”

Evaluation of Staphylococcal Bacteriophage Sb-1 as an Adjunctive Agent to Antibiotics Against Rifampin-Resistant Staphylococcus aureus Biofilms

“…With increased rifampin use, rifampinresistant staphylococcal strains are increasing worldwide, representing an important concern. For example, in China, the rifampin resistance in MRSA isolates increased from 15.5 to 50.2% within 4 years (2004)(2005)(2006)(2007)(2008) (Wang C. et al, 2019). Against rifampin-resistant mutants, rifampin has no biofilm activity in vitro or in vivo (Croes et al, 2010).…”

Risk Factors for Mortality in Patients with Elizabethkingia Infection and the Clinical Impact of the Antimicrobial Susceptibility Patterns of Elizabethkingia Species

“…RNA polymerase mutations not only induce rifampin resistance but also cause other alterations, especially virulence-related factors. For instance, Staphylococcus aureus, one of the biofilm-forming bacteria like Elizabethkingia, which gained RNA polymerase mutations showing a high level of rifampin MIC values, presented the elevated capacity for biofilm formation [43].…”