Abstract
Infection with Edwardsiella tarda, a Gram-negative bacterium, causes high morbidity and mortality in both marine and freshwater fish. Outer membrane vesicles (OMVs) released from Gram-negative bacteria are known to play important roles in bacterial pathogenesis and host immune responses, but no such roles for E. tarda OMVs have yet been described. In the present study, we investigated the proteomic composition of OMVs and the immunostimulatory effect of OMVs in a natural host, as well as the efficacy of OMVs when used as a vaccine against E. tarda infection. A total of 74 proteins, from diverse subcellular fractions, were identified in OMVs. These included a variety of important virulence factors, such as hemolysin, OmpA, porin, GAPDH, EseB, EseC, EseD, EvpC, EvpP, lipoprotein, flagellin, and fimbrial protein. When OMVs were administrated to olive flounder, significant induction of mRNAs encoding IL-1β, IL-6, TNFα, and IFNγ was observed, compared with the levels seen in fish injected with formalin-killed E. tarda. In a vaccine trial, olive flounder given OMVs were more effectively protected (p<0.0001) than were control fish. Investigation of OMVs may be useful not only for understanding the pathogenesis of E. tarda but also in development of an effective vaccine against edwardsiellosis.
Highlights
Outer membrane vesicles (OMVs) are spherical blebs of average diameter 10–300 nm that are naturally released from Gramnegative bacteria into the environment [1]
The budding mechanisms are unclear, it has been shown that OMVs are continuously produced during growth of various Gram-negative bacteria including Escherichia coli, Helicobacter pylori, Neisseria meningitidis, Pseudoaltermonas antarctica, Pseudomonas aeruginosa, Shigella flexneri, and Vibrio cholerae [2,3,4,5,6,7]
Transmission electron microscopy (TEM) examination of OMVs Numerous ovoid-to-round-shaped blebs were evident on the surface of ED45 cells when thin sections were stained to show OMVs (Figure 1-A)
Summary
Outer membrane vesicles (OMVs) are spherical blebs of average diameter 10–300 nm that are naturally released from Gramnegative bacteria into the environment [1]. The budding mechanisms are unclear, it has been shown that OMVs are continuously produced during growth of various Gram-negative bacteria including Escherichia coli, Helicobacter pylori, Neisseria meningitidis, Pseudoaltermonas antarctica, Pseudomonas aeruginosa, Shigella flexneri, and Vibrio cholerae [2,3,4,5,6,7] Such vesicles are known to contain lipopolysaccharide (LPS), lipoproteins, outer membrane, periplasmic, and cytoplasmic proteins, DNA, and RNA [1,8,9,10], and have been suggested to be involved in exclusion of competing bacteria, conveyance of proteins or genetic material to other bacteria, and presentation of virulence factors to the host [1]. Edwardsiella tarda is the causative agent of edwardsiellosis in a variety of cultured freshwater and marine fish, including channel catfish Ictalurus punctatus, olive flounder Paralichthys olivaceus, Japanese eel Anguilla japonica, red sea bream Pagrus major, mullet Mugil cephalus, and turbot Scophthalmus maximus [11,12,13,14,15,16]. Typical clinical symptoms of E. tarda infection in olive flounder are exophthalmia, enlargement of the spleen, malodorous ascites, and rectal hernia [17]
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