FEMSMicrobiology Letters97 (1992)77-82 © 1992Federation of EuropeanMicrobiological Societies0378-1097/92/$05.01) Publishedby Elsevier FEMSLE 05067 Comparison of the nucleotide sequence and development of a PCR test for the epsilon toxin gene of Clostridium perfringens type B and type D Helen L. Havard, Sophie E.C. Hunter, and Richard W. Titball Chemical and BiologicalDefence E.~tablishment, Porton Down. Salisbury UK Received 16June 1992 Accepted 8 July 1992 Key words: Clostridium perfringens; Epsilon toxin; DNA sequence 1. SUMMARY The sequence of the epsilon toxin gene of Clostridium perfringens type D was determined and compared with that of the previously re- ported type B sequence. It showed two nu- cleotide changes in the open reading frame, giv- ing rise to one amino acid substitution. The pro- moter sequences were not homologous, and dif- ferent putative -35 and -10 regions have been identified in each. The sequence information was used to develop PCR primers which were specific for the epsilon toxin gene. The utility of this system for identifying type B or D strains of C. perfringens was demonstrated. Correspondence to: H.L. Harvard, Chemical and Biological Defence Establishment. Porton Down, Salisbury,Wiltshire SP4 0JQ, UK. 2. INTRODUCTION The epsilon toxin of Clostrtdium perfringens is produced by type B and type D strains. Type B strains produce the major lethal toxins alpha, beta, and epsilon, and are responsible for lamb dysentery, and sheep and goat enterotoxaemia. Type D strains produce alpha and epsilon major toxins, and cause enterotoxaemia in sheep and pulpy kidney disease in lambs [1]. The epsilon toxin has been purified from some type D strains, and shown experimentally to increase vascular permeability in the brain [2], cause kidney dam- age [3] and contraction of the ileum [4], and to possess cardiovascular and pressor activities [5]. Little is known about the mode of action of epsilon toxin, and no enzymatic activity has yet been assigned to it, but the importance of certain amino acid residues for toxicity has been demon- strated [6-10].