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Reactive oxygen species (ROS), a group of unstable and highly reactive molecules or free radicals, are productions of molecular oxygen through cell metabolism. In invertebrates, especially in crustaceans, hemolymph plays an extremely important role in innate immune regulation and inflammatory responses, in which various microbes were modified by ROS. Even so, the information about ROS in relation to hemolymph microbiota and innate immunity is still limited in mud crab. To investigate the underlying mechanism, the ROS-excessive mud crabs (Scylla paramamosain) caused by knocking down catalase (SpCAT) were used to demonstrate the effects of abnormal levels of ROS on the changes in the hemolymph microbiota. Our results showed that SpCAT silence significantly increase ROS production and decrease the load of the bacteria in the hemolymph in mud crab. In the siCAT group, the dominant phyla were Proteobacteria, Actinobacteria, Acidonacteria, Bacteroidetes, Firmicutes, Gemmatimonadetes, Chloroflexi, Planctomycetes, Rokubacteria and Verrucommicrobia. Among them, the relative abundance of Firmicutes and Cyanobacteria was significantly increased upon the elevated ROS production. Some bacterial genera such as Bacteroides, Parabacteroides, Faecalibacterium and Romboutsia also increased with higher level of ROS. Moreover, the results showed that excessive ROS could promote the evident autophagy of the hemocytes. The result of this study demonstrated that ROS plays a vital role in modulating the homeostasis of the microbiota, and SpCAT is also crucially important in the ROS-mediated hemolymph microbiota homeostasis and the autophagy of the hemocytes in mud crab.
•SpCAT is involved in regulating hemolymph microbiota in mud crab.•ROS-mediated hemolymph homeostasis is crucially important for mud crab.•SpCAT could regulate autophagy of hemocytes in mud crab.•The bacterial genera Parabacteroides could be a key strain affecting ROS production in mud crab.