Invasion of fish epithelial cells by Photobacterium damselae subsp piscicida: evidence for receptor specificity, and effect of capsule and serum

Photobacterium damselae subsp. piscicida is a fish pathogen which causes se rious disease in commercial warmwater fish species. Because information on the initial stages of the infection is scarce, an investigation of the inva sion ability of this pathogen was undertaken utilizing a fish epithelial ce ll line (epithelioma papillosum carpio, EPC), a virulent capsulated strain of P. damselae (MT1415), an avirulent non-capsulated strain of P. damselae (EPOY-8803-II) and Escherichia coli HB101 as a non-invasive control. P. dam selae was found to be able to adhere to and invade fish epithelial cells an d remain inside them for 6-9 h. There were no significant differences in in vasiveness between the capsulated and non-capsulated strains. A kinetics st udy demonstrated that P. damselae invasiveness was more efficient at low m. o.i., reaching saturation at higher m.o.i., suggesting internalization may be receptor-mediated. Invasion efficiency (IE) was significantly higher tha n in the control E. coli HB101. Engulfment of bacteria was possibly by an e ndocytic process and was unaffected by killing the bacteria with UV light. However, heat-killed bacteria had significantly reduced invasion capability . Ultrastructural studies showed that inside the epithelial cells, the bact eria remained within large vacuoles for a few hours and no evidence of intr acellular replication was found, by either fluorescence or electron microsc opic studies. Normal sea bass serum slightly reduced the invasion capabilit y of the MT1415 strain, but heat-inactivated normal serum had no effect. On the other hand, heat-inactivated fish antiserum raised against the same st rain reduced the percentage of invaded epithelial cells by 50%. As for othe r pathogens, an intracellular phase of P. damselae may be a mechanism to de lay or avoid phagocytosis and host immune responses, favouring the spread o f infection.