The Clostridioides difficile S-Layer Protein A (SlpA) serves as a general phage receptor

Abstract

Therapeutic bacteriophages (phages) are being considered as alternatives in the fight against Clostridioides difficile infections. To be efficient, phages should have a wide host range, but the lack of knowledge about the cell receptor used by C. difficile phages hampers the rational design of phage cocktails. Recent reports suggested that the C. difficile surface layer protein A (SlpA) is an important phage receptor, but clear and unambiguous experimental evidence is lacking. Here, using the epidemic R20291 strain and its FM2.5 mutant derivative lacking a functional S-layer, we show that the absence of SlpA renders cells completely resistant to infection by phiCD38-2, phiCD111 and phiCD146, which normally infect the parental strain. Complementation assays with 12 different S-layer Cassette Types (SLCTs) expressed from a plasmid revealed that SLCT-6 also allowed infection by phiCD111, and SLCT-11 enabled infection by phiCD38-2 and phiCD146. Of note, expression of SLCTs 1, 6, 8, 9, 10 or 12 conferred susceptibility to infection by 5 myophages that normally do not infect the R20291 strain, namely phiMMP02, phiMMP03, phiMMP04, phiCD506 and phiCD508. Adsorption assays showed that >50% adsorption was required for productive phage infection. Altogether, our data suggest that many phages use SlpA as their receptor and most importantly, morphologically distinct phages of the Siphoviridae and Myoviridae families target SlpA despite major differences in their tail structures. Our study therefore represents an important breakthrough in our understanding of the molecular interaction between C. difficile and its phages.

Pathogenicity and virulence of Clostridioides difficile

“…To date, no strictly lytic C. difficile phage have been identified, and all of the characterized temperate phage are double-strand DNA viruses, either contractile myoviruses or non-contractile siphoviruses [ 211 ]. Additionally, few C. difficile cell surface receptors have been identified, however the S-layer seems to be a common target [ 128 ; 212 ; 213 ; 214 ; 215 ]. Given the high degree of sequence variability seen in the S-layer [ 123 ], it is likely that a cocktail of phage would be required in an effective CDI therapeutic.…”