COBIA (Rachycentron canadum)

subsp. damselae and may be useful in controlling or treating P. damselae subsp. damselaeinfections in aquaculture and clinical therapy.77. Chuang, W.H., Liu, P.C., Hung, C.Y. & Lee, K.K. (2014). Purification, characterization andmolecular cloning of alpha-2-macroglobulin in cobia, Rachycentron canadum. Fish & ShellfishImmunology, 41(2), 346-355.Alpha-2-macroglobulin (α-2-M) is a broad spectrum protease inhibitor which is abundant in theplasma of vertebrates and several invertebrates. The α-2-M was purified from cobia (Rachycentroncanadum) plasma by a four-step procedure: poly ethylene glycol fractionation, affinitychromatography, hydrophobic interaction chromatography and ion exchange chromatography onFast Protein liquid chromatography system in the present study. It migrated as one protein bandwith a molecular mass of about 360 kDa in the native state, whereas in SDS-PAGE it was about 180kDa under non-reducing condition. This result revealed that the native protein was a dimer. Inaddition, it was cleaved into two different fragments of molecular mass about 93 and 87 kDa whenreduced by dithiothreitol (DTT). The anti-protease activity of the purified α-2-M was apparentlydecreased as temperature elevated above 50 °C. The α-2-M exhibited highest protease inhibitoryactivity at pH 9. The results indicate that the α-2-M is a heat-labile and alkaline protease inhibitor.The purified α-2-M exhibited more than 50 protease inhibitory activity against extracellularproducts (ECP) of Vibrio alginolytius isolated from diseased cobia. It seems that the proteaseactivities in ECP may be affected by the plasma α-2-M. The protease inhibitory activities of cobiaplasma or purified α-2-M were decreased when incubated with 10 mM methylamine for 30 min.The α-2-M cDNA consisted of 4611 bp with an open reading frame of 4374 bp had been clonedfrom cobia liver. This sequence contained thioester domain (GCGEQ) and thirteen predicted N-linked glycosylation sites. In addition, the amino acid sequence of thioester domain and genes ofadjacent regions of cobia α-2-M were further compared with sequences of known fish species inGenBank. The unweighted pair group method using arithmetic average (UPGMA) was employed toconstruct the phylogenetic trees of α-2-M among different fish species (freshwater fish, sea waterfish and primitive fish), and all these fish species were then clustered into three groups. The cobiaα-2-M was closer to that of sea water fish than that of freshwater fish compared basing on itssimilarity of amino acid sequence and phylogenetic analysis of the partial gene.78. Coriolano, M.C., Silva, C.D.C., Melo, C.M.L., Souza B.R., Santos, A.J.G., Pereira, V.R.A.,Coelho, L.C. & Breitenbach, B. (2012). Immunomodulatory response of mice splenocytes inducedby RcaL, a lectin isolated from cobia fish (Rachycentron canadum) serum. Applied Biochemistryand Biotechnology, 168(5), 1335-1348.This work reports the isolation of a serum lectin from cobia fish (Rachycentron canadum) namedRcaL. Immunomodulatory activity on mice splenocyte experimental cultures through cytotoxicassays and cytokine production were also performed. RcaL was obtained through precipitation withammonium sulphate and affinity chromatography on a Concanavalin A-Sepharose 4B column. Theammonium sulphate fraction F3 showed the highest specific hemagglutinating activity and wasapplied to affinity chromatography. The lectin was eluted with methyl-α-D-mannopyranoside. RcaLshowed highest affinity for methyl-α-D-mannopyranoside and D-mannose; eluted fractions of RcaLagglutinated rabbit erythrocytes (titre, 128 -1 ) retained 66 % of chromatographed lectin activity, andthe obtained purification factor was 1.14. Under reducing conditions, a polypeptide band of 19.2kDa was revealed in sodium dodecyl sulphate polyacrylamide gel electrophoresis (PAGE). PAGEconfirmed RcaL as an acidic protein revealed in a single band. Cytotoxic and immunomodulatoryassays with RcaL in mice splenocyte cultures showed that the lectin was not cytotoxic and inducedhigher interferon gamma and nitric oxide production in splenocyte cultures. Purified RcaL inducedpreferential Th1 response, suggesting that it acts as an immunomodulatory compound.79. Dawson, C.E. (1969). Records of the barnacle Conchoderma virgatum from two Gulf of Mexicofishes. Proceedings of the Louisiana Academy of Science, 32: 58-62.Direct attachment of C. virgatum to the halfbeak Hyporhamphus unifasciatus and to the copepodLernaeolophus sultanus parasitizing a cobia, Rachycentron canadum, is reported from Mississippiwaters. A list of the fishes known to be associated with this barnacle is provided.80. Deardorff, T.L. & Overstreet, R. M. (1980). Taxonomy and biology of North American speciesof Goezia (Nematoda: Anisakidae) from fishes, including three new species. Proceedings of theHelminthological Society of Washington, 47(2), 192-217.23