A heightened immune response, in which immune responses are primed by repeated exposure to a pathogen, is an important characteristic of vertebrate adaptive immunity. In the present study, we examined whether invertebrate animals also exhibit a primed immune response. The LD50 of Gram-negative enterohemorrhagic Escherichia coli O157:H7 Sakai in silkworms was increased 100-fold by pre-injection of heat-killed Sakai cells. Silkworms pre-injected with heat-killed cells of a Gram-positive bacterium, Staphylococcus aureus, did not have resistance to Sakai. Silkworms preinjected with enterohemorrhagic E. coli peptidoglycans, cell surface components of bacteria, were resistant to Sakai infection. Silkworms preinjected with S. aureus peptidoglycans, however, were not resistant to Sakai. Silkworms preinjected with heat-killed Sakai cells showed persistent resistance to Sakai infection even after pupation. Repeated injection of heat-killed Sakai cells into the silkworms induced earlier and greater production of antimicrobial peptides than a single injection of heat-killed Sakai cells. These findings suggest that silkworm recognition of Gram-negative peptidoglycans leads to a primed immune reaction and increased resistance to a second round of bacterial infection.
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