Abstract
In this study, we investigated the hemocytes’ immune response to white spot syndrome virus (WSSV) or Vibrio alginolyticus infection at the protein level. The differential proteomes from crab hemocytes infected with WSSV or V. alginolyticus were analyzed using the isobaric tags for relative and absolute quantitation approach immediately after infection. Using this approach, we identified 1,799 proteins by their by LC–MS/MS spectra and sequencing data. These included 157 upregulated proteins and 164 downregulated proteins after WSSV infection. Similarly, 243 proteins were determined to be differentially expressed during V. alginolyticus infection, of these, 121 were upregulated and 122 were downregulated after infection. Interestingly, among these differentially expressed proteins, 106 were up- or downregulated significantly in both WSSV and V. alginolyticus infection. Six genes, β-actin, myosin-9, anti-lipopolysaccharide factor isoform 4, anti-lipopolysaccharide factor 4, transketolase-like protein 2-like isoform 1, and sarcoplasmic calcium-binding protein 1 were chosen for further study. The expression of these genes all showed a trend of upregulation at 24 h post-WSSV or V. alginolyticus infection except for myosin-9 in response to WSSV. To confirm the protective effects of the six genes, crabs were injected with specific dsRNAs before WSSV or V. alginolyticus challenge. The results showed that the knockdown of these genes led to an increase in the morbidity and mortality (P < 0.01) rate, and a decrease in infection time in WSSV-infected crabs. During the first 84 h, knockdown of these genes also led to an increase in the morbidity rates in V. alginolyticus -infected crabs, and results of four genes showed a higher mortality rate than that of the control after they were knocked down. This is the first report of the proteome response in crab hemocytes during WSSV or V. alginolyticus infection. These findings will contribute to our understanding of the immune response to WSSV and V. alginolyticus infection in crabs.
Highlights
The mud crab Scylla paramamosain, one of a number of commercially important crustaceans, is widely distributed along the coast line of southern China and the broader Pacific area
In an effort to analyze the functional distribution of the identified proteins, the known proteins from S. paramamosain hemocytes were used as a reference in the Cluster of Orthologous Groups of proteins (COG) database (Figure 1C)
V. alginolyticus infection would damage the mitochondria of hemocytes and cause hemocyte death in crabs [19]
Summary
The mud crab Scylla paramamosain, one of a number of commercially important crustaceans, is widely distributed along the coast line of southern China and the broader Pacific area. Several bacterial and viral pathogens have been reported to infect crabs [1]. Bacterial infections are the main cause of crab disease and constraint in the crab farming industry in China [2, 3]. Vibrio alginolyticus is an important bacterial pathogen that would cause huge economic losses and has caused many cases of natural infection in the Chinese mitten crab [4]. White spot syndrome has become the most hazardous and devastating disease in shrimp culture [5]. White spot syndrome virus (WSSV) can infect several species of crab both in the natural and experimental setting [6,7,8]
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