The New Antimicrobial Peptide SpHyastatin from the Mud Crab Scylla paramamosain with Multiple Antimicrobial Mechanisms and High Effect on Bacterial Infection

Abstract

SpHyastatin was first identified as a new cationic antimicrobial peptide in hemocytes of the mud crab Scylla paramamosain. Based on the amino acid sequences deduced, it was predicted that this peptide was composed of two different functional domains, a proline-rich domain (PRD) and a cysteine-rich domain (CRD). The recombinant product of SpHyastatin displayed potent antimicrobial activities against the human pathogen Staphylococcus aureus and the aquatic animal pathogens Aeromonas hydrophila and Pseudomonas fluorescens. Compared with the CRD of SpHyastatin, the PRD presented better antimicrobial and chitin binding activities, but both regions were essential for allowing SpHyastatin complete antimicrobial activity. The binding properties of SpHyastatin to different microbial surface molecules suggested that this might be an initial and crucial step for performing its antimicrobial activities. Evaluated using propidium iodide uptake assays and scanning electron microscopy images, the antimicrobial mechanism of SpHyastatin was found to be prone to disrupt cell membrane integrity. Interestingly, SpHyastatin exerted its role specifically on the surface of S. aureus and Pichia pastoris whereas it directly killed P. fluorescens through simultaneous targeting the membrane and the cytoplasm, indicating that SpHyastatin could use different antimicrobial mechanisms to kill different species of microbes. As expected, the recombinant SpHyastatin increased the survival rate of crabs challenged with Vibrio parahaemolyticus. In addition, SpHyastatin could modulate some V. parahaemolyticus-responsive genes in S. paramamosain.

Analysis of the antimicrobial mechanism of porcine beta defensin 2 against E. coli by electron microscopy and differentially expressed genes

“…The locations of pBD2 in bacteria cells were further confirmed by immune-gold TEM, which demonstrated that pBD2 was a cell-penetrating peptide. Antimicrobial peptide magainin 2, chicken cathelicidin-2, and SpHyastatin among others have been reported to pass through the cell membrane and are located in the cytoplasm of bacteria 34 – 38 . Wang et al .…”

“…Immuno-fluorescence analysis was performed according to methods published with some modifications 38 . E .…”

A Brief Review on the Antioxidants and Antimicrobial Peptides Revealed in Mud Crabs from the Genus of Scylla

“…Amino acid sequences of SCY2 were 65.08% similar to the previous scygonadin found in seminal plasma of S. serrata. In a more recent finding achieved by Shan et al [45], the new cationic AMP, SpHyastatin, in the hemocytes of mud crab, S. paramamosain, managed to be discovered. Interestingly, the recombinant product of SpHyastatin tends to exert different mechanisms to kill different bacteria such as Staphylococcus aureus, Aeromonas hydrophila, and Pseudomonas fluorescens.…”

“…Recently, the antibacterial activities of mud crab from the genus Scylla have been intensively studied and reported in many researches [42,47,[51][52][53]. Furthermore, [45] in supporting the antimicrobial activity in mud crab from earlier studies, Hoq et al [52] have partially purified the antimicrobial protein from the induced hemolymph of S. serrata. In this case, hemolymph refers to a fluid that circulates in the interior parts of crabs [53], with copper containing hemocyanins as the major protein component of hemolymph in crustaceans.…”

Enhanced Immune Protection of Mud Crab Scylla paramamosain in Response to the Secondary Challenge by Vibrio parahaemolyticus

“…Antimicrobial peptides (AMPs) have been reported to be important components of the host immune defense system against pathogen infection. In crab species, even in S. paramamosain, several AMPs have been discovered and characterized (45)(46)(47). In addition to AMPs, classical immunerelated processes, such as the TLR signaling pathway, are also reported to be involved in the immunity of S. paramamosain against Gram-negative and -positive bacteria (48)(49)(50).…”