Abstract
Background Crassostrea ariakensis Gould is a representative bivalve species and an economically important oyster in China, but suffers severe mortalities in recent years that are caused by rickettsia-like organism (RLO). Prevention and control of this disease is a priority for the development of oyster aquaculture. It has been proven that mammalian HMGB (high mobility group box) can be released extracellularly and acts as an important pro-inflammatory cytokine and late mediator of inflammatory reactions. In vertebrates, HMGB’s antibody (anti-HMGB) has been shown to confer significant protection against certain local and systemic inflammatory diseases. Therefore, we investigated the functions of Ca-HMGB (oyster HMGB) and anti-CaHMGB (Ca-HMGB’s antibody) in oyster RLO/LPS (RLO or LPS)-induced disease or inflammation.Methodology/Principal FindingsSequencing analysis revealed Ca-HMGB shares conserved structures with mammalians. Tissue-specific expression indicates that Ca-HMGB has higher relative expression in hemocytes. Significant continuous up-regulation of Ca-HMGB was detected when the hemocytes were stimulated with RLO/LPS. Recombinant Ca-HMGB protein significantly up-regulated the expression levels of some cytokines. Indirect immunofluorescence study revealed that Ca-HMGB localized both in the hemocyte nucleus and cytoplasm before RLO challenge, but mainly in the cytoplasm 12 h after challenge. Western blot analysis demonstrated Ca-HMGB was released extracellularly 4–12 h after RLO challenge. Anti-CaHMGB was added to the RLO/LPS-challenged hemocyte monolayer and real-time RT-PCR showed that administration of anti-CaHMGB dramatically reduced the rate of RLO/LPS-induced up-regulation of LITAF at 4–12 h after treatment. Flow cytometry analysis indicated that administration of anti-CaHMGB reduced RLO/LPS-induced hemocyte apoptosis and necrosis rates.Conclusions/SignificanceCa-HMGB can be released extracellularly and its subcellular localization varies when stimulated with RLO. Ca-HMGB is involved in oyster immune reactions and functions as a pro-inflammatory cytokine. Anti-CaHMGB can significantly suppress RLO/LPS-induced inflammatory responses and hemocyte necrosis and apoptosis, suggesting that Ca-HMGB is a potential target to prevent and control RLO/LPS-induced disease or inflammation.
Highlights
Mollusca is one of the most diverse groups of invertebrates with more than 100,000 living species, mostly located in marine environments [1]
The results showed that the rickettsia-like organism (RLO)/LPS challenge could up-regulate the expression level of LITAF in the hemocyte, while administration of anti-CaHMGB can reduce RLO or LPS (RLO/LPS)-induced up-regulation of LITAF
The results showed that administration of anti-CaHMGB can reduce the RLO/LPSinduced hemocyte cell apoptosis and necrosis rates and increase the cell survival rate
Summary
Mollusca is one of the most diverse groups of invertebrates with more than 100,000 living species, mostly located in marine environments [1]. Oyster culture has suffered from severe mortality caused by the pathogen rickettsia-like organism (RLO) [17,18]. Rickettsias are Gram-negative bacteria, generally described as obligate intracellular pathogens that multiply only within host cells [19] This prokaryote has been reported in many aquatic species including fishes [20,21], crustaceans [22,23] and mollusks [24,25]. Crassostrea ariakensis Gould is a representative bivalve species and an economically important oyster in China, but suffers severe mortalities in recent years that are caused by rickettsia-like organism (RLO). We investigated the functions of Ca-HMGB (oyster HMGB) and anti-CaHMGB (Ca-HMGB’s antibody) in oyster RLO/LPS (RLO or LPS)-induced disease or inflammation
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