A multivalent mRNA-LNP vaccine protects against Clostridioides difficile infection

Clostridioides difficile infection (CDI) is an urgent public health threat with limited therapeutic options. In this work, we developed a messenger RNA (mRNA)-lipid nanoparticle (LNP) vaccine targeting C. difficile toxins and virulence factors. This multivalent vaccine elicited robust and long-lived systemic and mucosal antigen-specific humoral and cellular immune responses across animal models and independent of changes to the intestinal microbiota. Vaccination protected mice from lethal CDI in both primary and recurrent infection models, and inclusion of non-toxin cellular and spore antigens improved decolonization of toxigenic C. difficile from the gastrointestinal tract. Our studies demonstrate mRNA-LNP vaccine technology as a promising platform for the development of novel C. difficile therapeutics with potential for limiting acute disease and promoting bacterial decolonization., mRNA design and production
The amino acid sequence of the TcdA and TcdB receptor binding domain (RBD) containing the combined repetitive oligopeptides (CROP) motifs were obtained from GenBank accession numbers P16154.2 and P18177.3 respectively. The amino acid sequences of PPEP-1 and CdeM was obtained from GenBank accession number Q183R7, and WP_009893169.1, respectively.  The putative N-glycosylation sites were disrupted by substituting the asparagine residue at the predicted N-glycolysation site with a glutamine (N to Q) to prevent posttranslational epitopes masking in eukaryotic cells. The sequences underwent codon optimization and GC enrichment using our proprietary algorithm to improve expression and reduce potential immunogenicity of the in vitro transcribed mRNA. The codon-optimized sequences were gene synthesized by Genscript with an optimized and modified IL-2 secretion signal, and cloned into our proprietary in vitro transcription template containing an optimized T7 promoter, ..., , # Data from: A multivalent mRNA-LNP vaccine protects against Clostridioides difficile infection

## Description of the data and file structure

We have submitted our raw data for each figure as it appears in the paper (adn4955_SourceData_FigureX.csv). File names indicate the specific figure from the manuscript (ie. FigureX indicates a main figure, where X equals 1-4; FigureSX indicates a supplemental figure, where X equals 1-16). Sheets in each file represent panels in an indicated figure (ie. adn4955_SourceData_Figure1.csv includes sheets labeled \"Figure 1b\", \"Figure 1c\", \"Figure 1d\", \"Figure 1e\", \"Figure 1f\", and \"Figure 1g\"). We have also submitted core genome alignments (adn4955_SourceData_Figure1A_CoreGeneAlignment), and a zip file included with Shovill v1.1.0 used for phylogenomic analysis as described in the Materials and Methods. 

#### Variable definitions for immune response to vaccination datasets

* *Antigen-specific IgG titers:* Concentration of IgG antibodies specific to *...