Orally administered beta-lactamase enzymes represent a novel strategy to prevent colonization by Clostridium difficile.
OBJECTIVE
Antibiotics that are excreted into the intestinal tract and that disrupt the indigenous microbiota may promote infection by Clostridium difficile. We previously demonstrated that oral administration of a proteolysis-resistant, recombinant class A beta-lactamase inactivates ampicillin or piperacillin excreted into the small intestine during parenteral treatment. We hypothesized that oral administration of this beta-lactamase in conjunction with parenteral ampicillin or piperacillin would preserve the colonic microbiota, thus preventing the overgrowth of and toxin production by C. difficile in mice.


METHODS
Subcutaneous ampicillin, subcutaneous piperacillin or either of these plus oral beta-lactamase or either of these plus tazobactam-inactivated oral beta-lactamase were administered to mice 24 and 12 h prior to harvest of caecal contents. Contents were inoculated with one of four strains of C. difficile, and growth and toxin production were assessed after 24 h of incubation under anaerobic conditions. To assess changes in stool microbiota, denaturing gradient gel electrophoresis (DGGE) of PCR-amplified ribosomal RNA genes was performed.


RESULTS
Mice treated with ampicillin, piperacillin or either of these plus tazobactam-inactivated oral beta-lactamase developed high-density colonization with C. difficile, whereas those treated with ampicillin or piperacillin plus the beta-lactamase did not. DGGE demonstrated that antibiotic treatment resulted in significant alteration of the indigenous stool microbiota, whereas antibiotic plus beta-lactamase treatment did not.


CONCLUSIONS
Administration of oral recombinant beta-lactamase preserved the colonic microbiota of mice during parenteral beta-lactam antibiotic treatment and prevented the overgrowth of and toxin production by C. difficile in caecal contents. Oral beta-lactamase therapy may represent a novel approach towards preventing C. difficile infections in healthcare settings.
Stiefel
Usha
U
Research Service, Louis Stokes Cleveland Veterans Affairs Medical Center, Cleveland, OH 44106, USA. stiefel@medscape.com
Nerandzic
Michelle M
MM
Koski
Pertti
P
Donskey
Curtis J
CJ
D000284:Administration, Oral; D000667:Ampicillin / Q000009:adverse effects*; D000818:Animals; D001426:Bacterial Proteins / Q000008:administration & dosage*; D001427:Bacterial Toxins / Q000096:biosynthesis; D016360:Clostridioides difficile / Q000254:growth & development*; D016172:DNA Fingerprinting; D004269:DNA, Bacterial / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004761:Enterocolitis, Pseudomembranous / Q000517:prevention & control*; D005260:Female; D005766:Gastrointestinal Contents / Q000382:microbiology; D051379:Mice; D009691:Nucleic Acid Denaturation; D010878:Piperacillin / Q000009:adverse effects*; D012335:RNA, Ribosomal / Q000235:genetics; D001618:beta-Lactamases / Q000008:administration & dosage*
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004269:DNA, Bacterial; D012335:RNA, Ribosomal; D000667:Ampicillin; D001618:beta-Lactamases; D010878:Piperacillin