RESUMEN
Background: Currently, 1-2% of all prosthetic joint surgeries are followed by an infection. These infections cause approximately 4% of deaths in the first year after surgery, while the 5-year mortality rate is up to 21%. Prosthetic joint infections are mainly caused by Staphylococcus aureus or Staphylococcus epidermis strains. Both species share the capability of biofilm formation and methicillin resistance. The formation of biofilm helps bacterial cells to withstand critical environmental conditions. Due to their tolerance against antibacterial substances, biofilms are a significant problem in modern medicine. Alternatives for the use of methicillin as a therapeutic are not yet widespread. The use of omega-3 fatty acids, such as docosahexaenoic acid, may help against prosthetic joint infections and lower mortality rates. The aim of this study is to evaluate if docosahexaenoic acid offers a safe anti-biofilm activity against Staphylococcus aureus and MRSA without enhancing icaADBC-dependent biofilm formation or additional stress responses, therefore enhancing antibiotic tolerance and resistance. Methods: In this study, we examined the gene expression of biofilm-associated genes and regulators. We performed RT-qPCR after RNA extraction of Staphylococcus aureus ATCC 29213 and one clinical MRSA strain. We compared gene expression of icaADBC, SarA, SigB, and agrAC under the influence of 1.25 mg /L and 0.625 mg/L of docosahexaenoic acid to their controls. Results: We found a higher expression of regulatory genes such as SarA, SigB, agrA, and agrC at 1.25 mg/L of docosahexaenoic acid in ATCC 29213 and a lower increase in gene expression levels in clinical MRSA isolates. icaADBC was not affected in both strains at both concentration levels by docosahexaenoic acid. Conclusions: Docosahexaenoic acid does not enhance icaADBC-dependent biofilm formation while still reducing bacterial CFU in biofilms. Docosahexaenoic acid can be considered an option as a therapeutic substance against biofilm formation and may be a good alternative in reducing the risk of MRSA formation.
RESUMEN
Bacterial antibiotic resistance and biofilm formation are mechanisms usually involved in the pathogeny of implant-related infections. Worldwide, antibiotic susceptibility tests are usually carried out using nutrient-rich media. Clinical routine laboratories and even research centers use for example EUCAST or CLSI for guidelines. In this study, we investigated the effect of different nutrient media on the antibiotic susceptibility and icaADBC gene expression of bacteria in biofilm. As media, Müller-Hinton Bouillon (MHB), Tryptic Soy Broth (TSB) and human synovial fluid (SF) diluted 1:4 in phosphate buffered saline (PBS), each also supplemented with 1% glucose, were used. The influence of different nutrient media on the antibiotic susceptibility of coagulase-negative staphylococci (CoNS) was evaluated by counting of colony-forming units (CFU) and by checking the metabolic activity of the bacteria. We used reverse transcriptase and real-time qPCR to investigate the influence of nutrient media on the biofilm gene expression. We used two-way analysis of variance (ANOVA). p < 0.05 was considered to be statistically significant. Significant differences in growth and antibiotic susceptibility were detected in all strains tested among the different media used. The nutrient media showed influence on the cell viability of all bacteria after antibiotic treatment. IcaADBC gene expression was significantly influenced by glucose and all nutrient media. The results highlight the influence of glucose on the antibiotic susceptibility, growth and gene expression of all strains tested. For all strains, a significant difference in bacterial recovery, viability and gene expression were found when compared to biofilm grown in SF.