Enzymes manage biofilms on crab surfaces aiding in feeding and antifouling

Abstract

Biofilms are sticky exopolymer matrices with embedded microorganisms that form on virtually all submerged surfaces. Depending on the context, biofilms are beneficial or detrimental to macroorganisms. Two groups of decapod crustaceans, sand fiddler crabs (Uca pugilator) and blue crabs (Callinectes sapidus), were investigated to test whether externally secreted enzymes were present and if they were used to remove or prevent biofilm formation in the environment and on their surfaces. Three hydrolytic enzymes were of interest because of their ability to degrade biofilm components: amylase, trypsin and lysozyme. The role of these enzymes was investigated in two contexts, removal of biofilms from the environment for food detection and processing and prevention of biofilm formation on the macroorganism surface itself. To test this, enzyme activity was measured on feeding appendages of fiddler crabs, which feed on biofilms, and on egg masses externally brooded by blue crabs. All three enzyme activities were found on feeding appendages of fiddler crabs and in egg masses of blue crabs. For the context of removal of biofilms from the environment, the combined data of the enzymes present in fiddler crab saliva and the high diatom extraction efficiency (60–90% removal in less than 9s), suggests a role in separation of organic content from sand particles. Secondly, the data of enzymes present on fiddler crab legs (the main location of their chemoreceptors) and that leg exposure to a human amylase inhibitor significantly decreased fiddler crab feeding responses, suggests a role of the enzymes on fiddler crab legs in generating phagostimulants. For the context of prevention of microfouling, it was found that the same enzymes are present in blue crab egg mass fluids and that the activity levels increase as the eggs mature, particularly amylase, which increased dramatically in clutches with late stage embryos. These findings of enzymes present in egg mass fluid throughout maturation inform a large body of work on pheromones and hatching cues for blue crabs. Detachment of brooded eggs and incubation with enzymes at levels found in egg masses resulted in survival and hatching while detachment without addition resulted in death. These findings suggest that the same suite of enzymes is important to biochemically manage biofilms in two ecologically relevant contexts. Biochemical management of biofilms may be more prevalent in macroorganisms than previously thought.