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HexSDF Is Required for Synthesis of a Novel Glycolipid That Mediates Daptomycin and Bacitracin Resistance in C. difficile
Journal article   Open access   Peer reviewed

HexSDF Is Required for Synthesis of a Novel Glycolipid That Mediates Daptomycin and Bacitracin Resistance in C. difficile

Anthony G Pannullo, Ziqiang Guan, Howard Goldfine and Craig D Ellermeier
mBio, Vol.14(2), e0339722
04/25/2023
DOI: 10.1128/mbio.03397-22
PMCID: PMC10128005
PMID: 36786594
url
https://doi.org/10.1128/mbio.03397-22View
Published (Version of record) Open Access

Abstract

Clostridioides difficile is a Gram-positive opportunistic pathogen responsible for 250,000 hospital-associated infections, 12,000 hospital-associated deaths, and $1 billion in medical costs in the United States each year. There has been recent interest in using a daptomycin analog, surotomycin, to treat C. difficile infections. Daptomycin interacts with phosphatidylglycerol and lipid II to disrupt the membrane and halt peptidoglycan synthesis. C. difficile has an unusual lipid membrane composition, as it has no phosphatidylserine or phosphatidylethanolamine, and ~50% of its membrane is composed of glycolipids, including the unique C. difficile lipid aminohexosyl-hexosyldiradylglycerol (HNHDRG). We identified a two-component system (TCS), HexRK, that is required for C. difficile resistance to daptomycin. Using transcriptome sequencing (RNA-seq), we found that HexRK regulates expression of , a three-gene operon of unknown function. Based on bioinformatic predictions, encodes a monogalactosyldiacylglycerol synthase, encodes a polysaccharide deacetylase, and encodes an MprF-like flippase. Deletion of leads to a 4-fold decrease in daptomycin MIC, and that deletion of leads to an 8- to 16-fold decrease in daptomycin MIC. The Δ mutant is also 4-fold less resistant to bacitracin but no other cell wall-active antibiotics. Our data indicate that in the absence of HexSDF, the phospholipid membrane composition is altered. In wild-type (WT) C. difficile, the unique glycolipid HNHDRG makes up ~17% of the lipids in the membrane. However, in a Δ mutant, HNHDRG is completely absent. While it is unclear how HNHDRG contributes to daptomycin resistance, the requirement for bacitracin resistance suggests it has a general role in cell membrane biogenesis. Clostridioides difficile is a major cause of hospital-acquired diarrhea and represents an urgent concern due to the prevalence of antibiotic resistance and the rate of recurrent infections. Little is understood about C. difficile membrane lipids, but a unique glycolipid, HNHDRG, has been previously identified in C. difficile and, currently, has not been identified in other organisms. Here, we show that HexSDF and HexRK are required for synthesis of HNHDRG and that production of HNHDRG impacts resistance to daptomycin and bacitracin.
 Gene Expression Signal Transduction glycolipid synthesis glycolipids two-component regulatory system cell envelope membrane biogenesis

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