Described are non-toxigenic Clostridium difficile strains and spores. Also described are vaccines comprising the Clostridium difficile spores. Further described are methods of preventing or treating a Clostridium difficile infection in a subject in need thereof.
1. A non-toxigenic Clostridium difficile strain comprising:
a) an immunogenic protein comprising
i) a glucosyltransferase domain of Clostridium difficile toxin TcdB;
ii) a cysteine proteinase domain of Clostridium difficile toxin TcdB; and
iii) a receptor binding domain of Clostridium difficile toxin TcdA,
wherein the glucosyltransferase domain of Clostridium difficile toxin TcdB comprises a W102A amino acid substitution and a D288N amino acid substitution when compared to SEQ ID NO.: 5.
2. The non-toxigenic Clostridium difficile strain of claim 1 , wherein the immunogenic protein comprises the amino acid sequence of SEQ ID NO.: 4.
3. The non-toxigenic Clostridium difficile strain of claim 2 , wherein the strain is non-toxigenic Clostridium difficile strain 138 (NTCD_Tcd138).
4. The non-toxigenic Clostridium difficile strain of claim 1 , wherein the Clostridium difficile form spores.
5. A non-toxigenic Clostridium difficile strain comprising:
a) an immunogenic protein comprising
i) a glucosyltransferase domain of Clostridium difficile toxin TcdB;
ii) a cysteine proteinase domain of Clostridium difficile toxin TcdB;
iii) a receptor binding domain of Clostridium difficile toxin TcdA; and
iv) a receptor binding domain of Clostridium difficile toxin TcdB,
wherein the glucosyltransferase domain of Clostridium difficile toxin TcdB comprises a W102A amino acid substitution and a D288N amino acid substitution and the cysteine proteinase domain of Clostridium difficile toxin TcdB comprises a C698A amino acid substitution when compared to SEQ ID NO.: 5.
6. The non-toxigenic Clostridium difficile strain of claim 5 , wherein the immunogenic protein comprises the amino acid sequence of SEQ ID NO.: 3.
7. The non-toxigenic Clostridium difficile strain of claim 6 , wherein the strain is non-toxigenic Clostridium difficile strain 169 (NTCD_Tcd169).
8. The non-toxigenic Clostridium difficile strain of claim 5 , wherein the Clostridium difficile form spores.
9. A vaccine comprising the Clostridium difficile spores of claim 4 and a pharmaceutically acceptable excipient or carrier.
10. A vaccine comprising the Clostridium difficile spores of claim 8 and a pharmaceutically acceptable excipient or carrier.
11. A method of treating or preventing Clostridium difficile bacterial infection in a subject in need thereof, the method comprising administering the vaccine of claim 9 .
12. The method of claim 11 , wherein the vaccine is administered orally.
13. The method of claim 11 , wherein the vaccine increases levels of anti-TcdA and anti-TcdB IgG antibodies in the subject.
14. The method of claim 11 , wherein the Clostridium difficile bacterial infection is caused by a hyper-virulent strain of Clostridium difficile.
15. The non-toxigenic Clostridium difficile strain of claim 1 , wherein the glucosyltransferase domain of Clostridium difficile toxin TcdB is positioned immediately upstream of the cysteine proteinase domain of Clostridium difficile toxin TcdB, wherein the amino acid sequence of the linked glucosyltransferase domain of Clostridium difficile toxin TcdB and the cysteine proteinase domain of Clostridium difficile toxin TcdB is set forth in SEQ ID NO.: 8; and wherein the receptor binding domain of Clostridium difficile toxin TcdA comprises the amino acid sequence of SEQ ID NO.: 6.
16. The non-toxigenic Clostridium difficile strain of claim 5 , wherein the glucosyltransferase domain of Clostridium difficile toxin TcdB is positioned immediately upstream of the cysteine proteinase domain of Clostridium difficile toxin TcdB, wherein the amino acid sequence of the linked glucosyltransferase domain of Clostridium difficile toxin TcdB and the cysteine proteinase domain of Clostridium difficile toxin TcdB is set forth in SEQ ID NO.: 9; wherein the receptor binding domain of Clostridium difficile toxin TcdA comprises the amino acid sequence of SEQ ID NO.: 6; and wherein the receptor binding domain of Clostridium difficile toxin TcdB comprises the amino acid sequence of SEQ ID NO.: 7.