English (United Kingdom)

https://curated-unify.zendy.io/wp-json/zendy-region/v1/featured_content/oa?rat=en

https://curated-unify.zendy.io/wp-json/zendy-region/v1/highlighted_journal/

Global: Zendy Plus annual

Presents the access of premium content as premium feature

Premium Content

Presents the keyphrase highlighting as premium feature

Keyphrase Highlighting

Presents the summarisation as premium feature

Summarisation

Insights

Presents the pdf analysis as premium feature

PDF Analysis

Presents the zaia usage as premium feature

ZAIA

Global: Zendy Plus monthly

Global: Zendy Tools annual

Global: Zendy Tools monthly

Global: Zendy Free Plan

Background Daptomycin (DAP) is a key first-line agent for the treatment of vancomycin-resistant enterococcal infections. Resistance to DAP in enterococci is regulated by the liaFSR three-component regulatory system that consists of a histidine kinase sensor (LiaS), a response regulator (LiaR) and a transmembrane protein of unknown function (LiaF). Previous studies indicate that deletion of isoleucine in position 177 of LiaF results in DAP tolerance and is sufficient to change membrane architecture. Here, we dissect the role of LiaF in DAP resistance Methods We generated three liaF mutants in OG1RF, a DAP-susceptible laboratory strain of Efs (DAP MIC = 2 µg/mL): (i) a non-polar, C-terminal truncation of liaF (OG1RFliaF∆152), (ii) a null liaF mutant with a premature stop-codon (OG1RFliaF*11), and (iii) an isoleucine deletion at position 177 (OG1RFliaF177). We determined DAP MIC by Etest and characterized the localization of anionic phospholipids microdomains using 10-nonyl-acridine-orange (NAO). The expression of the liaXYZ (the main target of LiaR) and liaFSR clusters were evaluated by qRT-PCR and relative expression ratios (Log2 fold change) were calculated by normalizing to gyrB expression. We assessed activation of LiaFSR by evaluating surface exposure of LiaX by ELISA. We used the bacterial adenylate cyclase two-hybrid system (BACTH) to evaluate the protein-protein interaction between LiaF and LiaS. Results Full deletion of liaF or the C-terminal truncation of LiaF did not have any effect on DAP MICs, membrane architecture or a significant increase in LiaX surface exposure compared with parental strain OG1RF. In contrast, deletion of the codon encoding isoleucine in position 177 of LiaF caused a major increase (8-fold) in LiaX exposure and redistribution of anionic phospholipid microdomains away from the septum without changes in the actual DAP MIC. Transcriptional analyses indicated upregulation (&gt;2 log2-fold) in the liaXYZ gene cluster indicating activation of the stress response. We also observed a positive interaction between LiaF and LiaS. Conclusion LiaF is likely a key activator of the LiaFSR stress response and the critical regulatory domain appears to be located in a stretch of four isoleucines toward the C-terminal of the protein. Disclosures All authors: No reported disclosures.

open forum infectious diseases

599. LiaF is an Activator of the LiaR-Mediated Response Against Daptomycin and Antimicrobial Peptides in Multidrug-Resistant Enterococcus faecalis (Efs)