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Characterization of a haemolytic phospholipase A₂ activity in clinical isolates of Campylobacter concisus

Taghrid S. Istivan¹, Peter J. Coloe¹, Benjamin N. Fry¹, Peter Ward² and Stuart C. Smith³

¹Department of Biotechnology and Environmental Biology, RMIT University, PO Box 71, Bundoora, Victoria 3083, Australia ²Microbiology Department, Royal Children's Hospital, Melbourne, Australia ³School of Health Sciences, Deakin University, Melbourne, Australia

Correspondence Taghrid S. Istivan taghrid.istivan{at}rmit.edu.au

Received November 26, 2003
Accepted February 10, 2004

A membrane-bound, haemolytic phospholipase A₂ (PLA₂) activity was detected in clinical strains of Campylobacter concisus isolated from children with gastroenteritis. The clinical strains were assigned into two molecular groups (genomospecies) based on PCR amplification of their 23S rDNA. This calcium-dependent, heat-stable, haemolytic PLA₂ activity was detected in strains from both genomospecies. A crude haemolysin extract (CHE) was initially prepared from cellular outer-membrane proteins of these isolates and was further fractionated by ultrafiltration. The haemolytic activity of the extracted fraction (R30) was retained by ultrafiltration using a 30 kDa molecular mass cut-off filter, and was designated haemolysin extract (HE). Both CHE and HE had PLA₂ activity and caused stable vacuolating and cytolytic effects on Chinese hamster ovary cells in tissue culture. Primers for the conserved region of pldA gene (phospholipase A gene) from Campylobacter coli amplified a gene region of 460 bp in all tested isolates, confirming the presence of a homologous PLA gene sequence in C. concisus. The detection of haemolytic PLA₂ activity in C. concisus indicates the presence of a potential virulence factor in this species and supports the hypothesis that C. concisus is a possible opportunistic pathogen.

Abbreviations: CHE, crude haemolytic extract; CHO, Chinese hamster ovary; HE, haemolytic extract; LPS, lipopolysaccharides; OMP, outer-membrane protein; OMPLA, outer-membrane phospholipase A; PC, phosphatidylcholine; PL, phospholipase.

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