Characteristics delineation of piscidin 5 like from Larimichthys crocea with evidence for the potent antiparasitic activity.

Several researches reported that piscidin members of teleosts owned strong antiparasitic activity. Cryptocaryon irritans, a type of ectoparasite, could infect most of the marine teleosts. Larimichthys crocea could severely suffer from marine white spot disease caused by C. irritans, and their mortality rate was significantly high. Concentrating on this problem, we have done many related works. Piscidin 5 like (termed Lc-P5L) was another piscidin member isolated from a comparative transcriptome of C. irritans-immuned L. crocea. In the paper, quantitative Real-time PCR (qRT-PCR) showed Lc-P5L was upregulated in examined tissues, including gill, head kidney, muscle, liver, spleen and intestine after challenged by C. irritans, the significant upregulation time was in accordance to key developmental stages of C. irritans, which implied different infection stages could result in host immune response. Furthermore, using microscope techniques, we observed theronts or trophonts became weakly motile, cilia became detached, cells were out of shape, membranes eventually lysed in different cell positions and cytoplasmic contents leaked. Laser confocal scanning microscope (LCSM) observed theronts macronucleus grew swell and depolymerized after treated by recombinant Lc-P5L (rLc-P5L). Data suggested rLc-P5L was significantly lethal to C. irritans, and the death state of the parasite incubated with rLc-P5L was remarkably similar to other piscidin members or other antiparasitic peptides (APPs). Thus, these data provided new insights into L. crocea immunity against C. irritans and potential of rLc-P5L as a therapeutic agent against pathogen invasion.

Excavating differentially expressed antimicrobial peptides from transcriptome of Larimichthys crocea liver in response to Cryptocaryon irritans.

Larimichthys crocea, the special marine economy fish, owns the largest annual yield for a single species in China. One of the most significant factors affecting large yellow croaker culture is the diseases, especially the threat of marine white spot disease which caused by a protozoan Cryptocaryon irritans. Antimicrobial peptides (AMPs) have been demonstrated to be active against bacterium, fungi and parasites, showing their potential usefulness in aquaculture as substitutes for antibiotics. Many researches have been carried out about the AMPs concentrating on the activity resist on C. irritans, and piscidin-like of L. crocea owning widely antibacterial spectrum and strong activity against C. irritans was screened in our team. In the paper, taking advantage of the large yellow croaker hepatic comparison transcriptome in response to C. irritans at 3d post infection, seven kinds of AMPs have been excavated from the differently expressed genes, including LEAP2 like, LEAP-2A, hepcidin, hepcidin-like, piscidin-5-like, piscidin-5-like type 4 and bactericidal permeability increasing protein (BPI). Hepcidin, hepcidin-like, piscidin-5-like, piscidin-5-like type4 and BPI were up-regulated to protect large yellow croaker from being damaged by C. irritans infection; while LEAP2 like and LEAP-2A were down-regulated, they might be as a negative-feedback regulation factor or some other regulatory mechanisms to adjust the immune response in the process of C. irritans infection. The differential expression changes were verified with quantitative real-time PCR (qRT-PCR) to illustrate the reliability of the sequenced data. Hearteningly, piscidin-5-like type 4 was a novel type which was high similar to other piscidin-5-like types. Interestingly, the infection may well cause alternative splicing of LEAP-2A mRNA, which was a surprised phenomenon and finding after C. irritans infection, but more further study was needed to be conducted. Therefore, the data showed that these AMPs were involved in the immune response to the C. irritans infection. In all, these results implied that the immune response of AMPs to C. irritans infection was a complex and sophisticated regulatory process.