Immunization against clostridium difficile disease

Abstract

The invention provides active and passive immunization methods for preventing and treating Clostridium difficile infection, which involve percutaneous administration of C. difficile toxin-neutralizing polyclonal immune globulin, C. difficile toxoids, or combinations thereof. Also provided by the invention are C. difficile toxoids, C. difficile toxin-neutralizing polyclonal immune globulin, and methods of identifying subjects that produce C. difficile toxin-neutralizing polyclonal immune globulin.

What is claimed is: 
     
         1 . A method of producing a  Clostridium difficile  toxoid, said method comprising the steps of:
 providing a  Clostridium difficile  bacterium;   culturing said bacterium in medium lacking animal products to generate a culture;   purifying clostridial toxin A and clostridial toxin B from said culture to generate a mixture of purified toxins A and B; and   inactivating said purified toxins A and B by incubation in formaldehyde at a temperature of 25° C. or less to generate said clostridial toxoid.   
     
     
         2 . The method of  claim 1 , wherein said purified toxins A and B are incubated in formaldehyde at a temperature of about 15° C. or less, or a temperature of about 5° C. or less. 
     
     
         3 . The method of  claim 1 , wherein said purified toxins A and B are present in said mixture at a ratio of about 0.1:1 to 10:1 or a ratio of about 2:1. 
     
     
         4 . A  Clostridium difficile  toxoid produced by the method of  claim 1 . 
     
     
         5 . A vaccine composition comprising the  Clostridium difficile  toxoid of  claim 4 . 
     
     
         6 . The vaccine composition of  claim 5 , further comprising 0.012-0.020% formaldehyde. 
     
     
         7 . The vaccine composition of  claim 5 , further comprising an adjuvant. 
     
     
         8 . The vaccine composition of  claim 7 , wherein the adjuvant is an aluminum compound. 
     
     
         9 . A method of producing human immune globulin directed against a clostridium, said method comprising the steps of:
 administering a clostridial toxin or toxoid to a human; and   isolating said immune globulin against a clostridium from said human.   
     
     
         10 . The method of  claim 9 , wherein the clostridium is  Clostridium difficile.    
     
     
         11 . The method of  claim 9 , wherein said clostridial toxoid comprises toxin A and toxin B. 
     
     
         12 .  Clostridium difficile  immune globulin produced by administering a  Clostridium difficile  toxin or toxoid to a human, and isolating said  Clostridium difficile  immune globulin from said human 
     
     
         13 . A method of identifying a human producing a  Clostridium difficile  immune globulin, said method comprising the steps of:
 obtaining a blood sample from a human vaccinated with a  Clostridium difficile  toxoid;   determining the level of antibodies to  Clostridium difficile  Toxins A and B in said blood sample by an enzyme-linked immunosorbent assay; and   determining the level of in vitro cytotoxicity neutralization activity against  Clostridium difficile  Toxins A and B in said blood sample, wherein detection of increased levels of antibodies to  Clostridium difficile  Toxins A and B in said blood sample, and detection of in vitro cytotoxicity neutralization activity against  Clostridium difficile  Toxins A and B in said blood sample, indicates identification of a human producing a  Clostridium difficile  immune globulin useful for treatment or prevention of  Clostridium difficile  infection.