Preparation of Organisms

The ability of normal and Crohn's disease neutrophils to kill Candida albicans has been studied using neutrophils isolated from peripheral blood and suspended in phosphate buffered saline at 5x 106 cells per ml. C albicans was grown to a stationary phase in broth culture and suspended in phosphate buffered saline at 107 organisms/ml. Neutrophils and Candida were then incubated together at 37°C in a shaking water bath in the presence of fresh serum. At 30 and 60 minutes samples were withdrawn, neutrophils lysed, and Candida survival assessed by colony counting. Results were compared with control suspensions of Candida incubated with serum alone. After 30 and 60 minutes in the presence of autologous serum normal neutrophils had killed significandy more Candida than Crohn's disease neutrophils (mean (SD) 61-0 (16.7)% v 40 5 (16-2)% at 30 minutes, p<00001; 83*2 (7)% v 70-8 )16)% at 60 minutes, p<0005). The results did not alter significantly when nonnal neutrophils were incubated with Candida in the presence of Crohn's disease serum instead of normal serum. When Crohn's disease neutrophils were incubated with Candida in the presence of normal serum instead of autologous serum there was some improvement in candidacidal ability at 30 minutes (48.9 (20.6)% v 40 5 (16.2)%, p<003) but not at 60 minutes. Phagocytosis, measured using a radiometric assay, was normal. Neutrophils from patients with Crohn's disease have an impaired ability to kill this granuloma provoking organism. It is not due to serum inhibitors or defective phagocytosis. Departments of Surgery and Gastroenterology, The General Hospital, Birmingham F T Curran D J Youngs RNAllan M R B Keighley Correspondence to: Mr F T Curran, Queen Elizabeth Hospital, Edgbaston, Birmingham B 15. Accepted for publication 19 March 1990 The inactivation of invading micro-organisms is the most important function of the neutrophil. Although various abnormalities of neutrophil function have been reported in Crohn's disease,' their capacity to kill granuloma provoking organisms has not been evaluated. This study was designed to investigate the ability of Crohn's disease neutrophils to kill Candida albicans. The organism was chosen for study because it is present in the gastrointestinal tract of 14-47% of the healthy population2 3 and causes chronic inflammatory diseases, such as chronic mucocutaneous candidiasis, and leads to granuloma formation.3 It may also cause enteritis4 and allergic colitis.56 It is a convenient organism to culture compared to other organisms responsible for causing chronic inflammatory conditions. Whereas C albicans can be cultured in two to three days, mycobacteria, for example, require four to 12 weeks. It is a relatively safe organism to handle and is the species most resistant to killing by neutrophils.7 Phagocytosis was studied using a sensitive radiometric assay to determine whether impairment of candidacidal activity could be a result of this.