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Pyruvoyl-dependent histidine decarboxylases are produced as proenzymes that mature by cleavage followed by formation of the pyruvoyl prosthetic group. The histidine decarboxylation pathway of Streptococcus thermophilus CHCC1524 that consists of the pyruvoyl-dependent histidine decarboxylase HdcA and the histidine/histamine exchanger HdcP was functionally expressed in Lactococcus lactis. The operon encoding the pathway contains in addition to the hdcA and hdcP genes a third gene hdcB. Expression of different combinations of the genes in L. lactis and Escherichia coli followed by analysis of the protein products demonstrated the involvement of HdcB in the cleavage of the HdcA proenzyme. The HdcA proenzyme and HdcB protein were purified to homogeneity and cleavage and activation of the histidine decarboxylase activity was demonstrated in vitro. Substoichiometric amounts of HdcB were required to cleave HdcA showing that HdcB functions as an enzyme. In agreement, expression levels of HdcB in the cells were low relative to those of HdcA. The turnover number of HdcB in vitro was extremely low (0.05 min(-1)) which was due to a very slow association/dissociation of the enzyme/substrate complex. In fact, HdcB was shown to co-purify both with the HdcA S82A mutant that mimics the proenzyme and with the mature HdcA complex.

Scanning surveillance by the Veterinary Laboratories Agency revealed the emergence of suspected botulism in ruminants in 2003, presented as flaccid paralysis. From 2003 to 2009, 168 cattle and 19 sheep incidents were recorded, with mortality between 5 and 80 per cent. All sheep incidents and 95 per cent of cattle incidents had proximity to broiler litter. From July 2006, the gut contents collected from 74 affected cattle and 10 affected sheep were tested for Clostridium botulinum toxins using mice bioassays and for organisms by culture. Type D toxin was identified in 32 per cent of cattle and 18 per cent of sheep samples. C botulinum type D organisms were identified in 40 per cent of cattle and 30 per cent of sheep samples, but broth from one sample reacted with C and D antisera. Type C botulism has previously been reported more commonly than type D in the UK and has been associated with the use of poultry litter as fertiliser, bedding or feed. The almost exclusive association with C botulinum type D toxins or organisms in the gut contents in this survey suggests a change in the source or epidemiology of botulism in the UK. The source of C botulinum type D was uncertain. Broilers may carry C botulinum type D in their gut flora subclinically. The emergence of a new type D strain, or changes in broiler husbandry and nutrition, medication and other enteric infections may have affected colonisation with C botulinum. Further investigation of poultry and farm environments for sources of type D awaits the development of tests for C botulinum toxins that do not require the use of mice.