Invertebrates rely on their innate immune responses to protect themselves from pathogens, one of which is melanization of bacteria mediated by the activation of phenoloxidase (PO). Furthermore, invertebrate hemolymph, even that of healthy individuals, has been shown to contain bacterial species. The mechanisms that prevent these bacteria from proliferating and becoming deleterious to the host are, however, poorly understood. Here, we show that knocking down the activity of the inactive precursor of PO [prophenoloxidase (proPO)] by RNA interference resulted in a significant increase in the bacterial load of kuruma shrimp, Marsupenaeus japonicus, even in the absence of a bacterial or viral challenge. Silencing of proPO also led to a sharp increase in shrimp mortality. In addition, the hemolymph of proPO‐depleted shrimp had significantly lower hemocyte counts and PO activity than control samples. Microarray analysis after proPO silencing also showed a decrease in the expression of a few antimicrobial peptides, but no effect on the expression of the genes involved in the clotting system. Treatment with antibiotics prior to and after proPO dsRNA injection, to counteract the loss of proPO, resulted in a significant increase in shrimp survival. Our results therefore show that the absence of proPO renders the shrimp incapable of controlling bacteria present in the hemolymph, and that proPO is therefore essential for its survival.

Background: Lysozyme is ubiquitous in many organisms and functions primarily in bacterial lysis. Results: The absence of lysozyme in shrimp resulted in the increase of bacteria in the hemolymph leading to mortality. Conclusion: Lysozyme is essential to shrimp survival particularly in regulating bacterial communities in the hemolymph. Significance: A clear understanding of the immune effectors in shrimp, such as AMPs, is vital in ensuring resistance against bacterial pathogens.

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