Antibacterial activity in the blood cells of the solitary ascidian, Ciona intestinalis, in vitro

The antibacterial properties of the blood cells of the solitary ascidian, Ciona intestinalis, were investigated in vitro. The results show that blood cell lysate supernatants have potent antibacterial activity against a wide range of bacteria. Activity was strongest against Gram positive bacteria, where typically there was a 95-99% reduction in bacterial viability within 1 h. By contrast, Gram negative bacteria were less susceptible to-the antibacterial effects, with the viability of these bacteria generally reduced by approximately 20-40% after 3 h. Activity against both types of bacteria was further found to be heat-stable (even after heating to 100 degrees C for 10 min or freezing to -20 degrees C for up to 1 month), Ca2+- but not Mg2+-dependent, and active at very low titres. Comparison of the antibacterial vigour of lysate supernatants made from separated blood cell populations revealed that activity resides mainly in the morula cells with some, albeit weaker, activity also detectable in the hyaline amoebocytes. No activity was present in the plasma. Preliminary investigations into the mode of action of the active factors indicate that the response is not due to lysozyme, is not directly lytic in character, and is not attributable to agglutination. Instead, as preliminary characterization by gel filtration on Sephadex G-75 revealed 4 main protein peaks, with antibacterial activity confined to peak 3 (molecular weight range of 8-15 kDa), the phenomenon seems to be due to the action of one or more low molecular weight proteins. The location, action, and release of these factors appear to differ from those of the cecropins and defensins described in other animal groups. (C) 1995 Wiley-Liss, Inc.