HexSDF and HexRK are Required for Clostridioides difficile Synthesis, Which Impacts Resistance to Daptomycin and Bacitracin

The 2-component system, HexRK, and 3-gene operon, hexSDF, are required for the synthesis of the Clostridioides difficile (C difficile) lipid aminohexosyl-hexosyldiradylglycerol (HNHDRG), which impacts resistance to daptomycin and bacitracin, according to a study published in mBio.

“C 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,” wrote Anthony Pannullo, Department of Microbiology and Immunology, Carver College of Medicine, University of Iowa in Iowa City, Iowa, and coauthors. “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,” they added.

Transcriptome sequencing (RNA-seq) discovered that HexRK regulates the expression of hexSDF, a three-gene operon with an unknown function. Bioinformatic predictions suggest that the hexS gene encodes a monogalactosyldiacylglycerol synthase, hexD encodes a polysaccharide deacetylase, and hexF encodes an MprF-like flippase. When hexRK is deleted, the daptomycin minimum inhibitory concentration (MIC) decreases 4-fold. Similarly, deleting hexSDF leads to an 8- to 16-fold decrease in daptomycin MIC and a 4-fold decrease in resistance to bacitracin.

The absence of HexSDF alters the composition of the phospholipid membrane. In wild-type (WT) C difficile, HNHDRG glycolipid makes up approximately 17% of the lipids in the membrane. However, this glycolipid is absent in a ΔhexSDF mutant. Although it is unclear how HNHDRG contributes to daptomycin resistance, its requirement for bacitracin resistance suggests that it plays a vital role in cell membrane biogenesis.

“Here, we show that HexSDF and HexRK are required for synthesis of HNHDRG and that production of HNHDRG impacts resistance to daptomycin and bacitracin,” wrote the study authors.

Reference

Pannullo AG, Guan Z, Goldfine H, Ellermeier CD. HexSDF is required for synthesis of a novel glycolipid that mediates daptomycin and bacitracin resistance in C. difficile. mBio. 2023;14(2):e0339722. doi:10.1128/mbio.03397-22