A phoE‐lacZ hybrid gene encoding the N‐terminal 300 amino acid residues of pre‐PhoE protein, fused to an almost complete beta‐galactosidase molecule was constructed in vitro. Cell fractionation experiments suggested that the hybrid gene product is transported to the outer membrane. However, by using immuno‐cytochemical labelling on ultra‐thin cryosections it was shown that the hybrid protein accumulated in the cytoplasm. Thus, it appears that: (i) data on the localization of hybrid proteins merely based on cell fractionation experiments are not reliable, and (ii) either the C‐terminal 15% of PhoE protein contain information which is essential for transport, or PhoE‐LacZ hybrid proteins can never be transported out of the cytoplasm. The implications of these results for current models on the translocation of outer membrane proteins are discussed.

Several gene banks of Clostridium perfringens in E. coli were constructed. Using a mixture of synthetic 29-mer DNA probes clones were selected containing inserts from the C. perfringens gene coding for the enterotoxin. This has allowed sequencing of the complete gene and its flanking regions. The decuded amino acid sequence (320 a.a.) was found to differ at several sites from the sequence published previously by others. Two 40-mer DNA-probes were used to detect the toxin gene in C. perfringens strains isolated from the faeces of different non-symptomatic animals. Only 6% of the strains were found to possess the gene.

Four synthetic oligonucleotides encoding different parts of the Clostridium perfringens enterotoxin gene were used to test the enterotoxigenicity of C. perfringens strains isolated from confirmed outbreaks of food poisoning. Of the 245 strains isolated from food and feces originating from 186 separate outbreaks, 145 (59%) gave hybridization reactions with each of the four DNA probes used, while 104 strains did not hybridize with any of the probes. There was no correlation between serotype and the presence of the enterotoxin gene, although the C. perfiringens enterotoxin gene was rarely detected among nontypable strains (17%). Results show that DNA hybridization is a suitable method for the identification of C. perfringens strains which have the potential to produce enterotoxin.

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