Degree Type


Date of Award


Degree Name

Doctor of Philosophy


Veterinary Microbiology and Preventive Medicine

First Advisor

Ricardo F. Rosenbusch


The phospholipase C (PLPC) gene from Clostridium hemolyticum was cloned using the polymerase chain reaction. An open reading frame which encodes a 399-amino acid protein, containing a 27-amino acid signal sequence, was identified. The molecular weight of the active protein was 42,869 daltons. A 16-amino acid N-terminal sequence determined by Edman degradation exactly matched the putative amino acid sequence of the gene product. Comparison of the nucleotide and amino acid sequences with Genebank databases demonstrated that the beta toxin of C. hemolyticum exhibits high homology with other bacterial PLPCs. The N-terminal portion of the beta toxin contains the zinc-binding domain common to other bacterial PLPCs, and the C-terminal domain of the beta toxin protein shows considerable homology with the C-terminal domains of C. perfringens alpha toxin and C. novyi type A PLPC, and the N-terminal domain of bovine arachidonate 5-lipoxygenase.;The beta toxin of C. hemolyticum was purified by preparative isoelectric focusing and used to develop toxin neutralizing and non-neutralizing monoclonal antibodies. Guinea pigs passively immunized with toxin-neutralizing monoclonal antibodies were protected from a 100 LD50 spore challenge. The toxin-neutralizing ability of the monoclonal antibodies, as measured in in vitro and in vivo toxin neutralization assays, correlated to the protective effects in guinea pigs. Protection of guinea pigs injected with varying doses of immunoaffinity purified beta toxin was correlated to the presence of anti-beta toxin antibodies in the serum.



Digital Repository @ Iowa State University,

Copyright Owner

Paul Joseph Hauer



Proquest ID


File Format


File Size

84 pages