Calnexin is a 90-kDa integral membrane protein of the endoplasmic reticulum (ER). Calnexin binds Ca2+ and may function as a chaperone in the transition of proteins from the ER to the outer cellular membrane. We have purified human calnexin in association with the human interferon-gamma receptor and cloned calnexin cDNA from placenta. Fragments of calnexin have been prepared as glutathione S-transferase fusion proteins and analyzed for their abilities to bind 45Ca2+ and ruthenium red. A subdomain containing four internal repeats binds Ca2+ with the highest affinity. This sequence is highly conserved when compared to calreticulin (a luminal ER protein), an Onchocerca surface antigen, and yeast and plant calnexin homologues. Consequently, this sequence represents a conserved motif for the high-affinity binding of Ca2+, which is clearly distinct from the "E-F hand" motif. An adjacent subdomain, also highly conserved and containing four internal repeats, fails to bind Ca2+. The carboxyl-terminal, cytosolic domain is highly charged and binds Ca2+ with moderate affinity, presumably by electrostatic interactions. The calnexin amino-terminal domain (residues 1-253) also binds Ca2+, in contrast to the amino-terminal domain of calreticulin, which is relatively less acidic. We have also determined the cDNA sequences of mouse and rat calnexins. Comparison of the known mammalian calnexin sequences reveals very high conservation of sequence identity (93-98%), suggesting that calnexin performs important cellular functions. The gene for human calnexin is located on the distal end of the long arm of human chromosome 5, at 5q35.

ABSTRACT. A culture of Vibrio harveyi, isolated from diseased Penaeus vannamei, was pathogenic in penaeid shrimp larvae when used in a bath at 105 cells n~l-' for 2 h. The resultant disease had characteristics of Bolitas negricans, as observed in Ecuadorian hatcheries, namely the development of bioluminescence, reduced feeding and retarded development, sluggish swimming, reduced escape mechanisms, degeneration of hepatopancreatic tissue with resultant formation of necrotic bundles, and increased mortality. Koch's Postulates were confirmed by reisolation and identification of the organism. Histopathology showed the presence of distlnctive melanotic tissue aggregates within the hepatopancreas, with irnmunohistochemistry confirming the presence of large numbers of v harveyi in the intestine and hepatopancreas.These results indicate a suitable infection protocol, which can be used to test the pathogenicity of putative pathogens of penaeid shrimp larvae.

Wild Litopenaeus Õannamei females in different stages of sexual maturation were sampled, Ž . including spent females and their nauplii, for determination of the lipid content, lipid class LC Ž . composition, fatty acid FA composition, vitamin C content and vitamin E content. Free FA Ž . Ž . Ž . Ž . FFA , triacylglycerol TAG , phosphatidylcholine PC and sterol esters SE were the dominant Ž . LC in the midgut gland. TAG and phospholipids PL , mainly PC and phosphatidylethanolamine Ž .Ž . Ž . PE , were the dominant ovarian LC. Neutral lipids NL prevailed over polar lipids POL in midgut gland lipids, while ovarian lipids displayed an inverse relationship. An increase in ovarian Ž . Ž . TL was observed from stage 0 immature to stage 1 early maturing . Later, from stage 1 to stage Ž . 2 mid maturing , a decrease in midgut gland TL was observed. TAG was most responsible for these changes in TL. Lipids were preferentially transferred to the nauplii, which contained relatively high TAG and PC levels. In both midgut gland and ovaries, 16:00, 18:00, 16:1n y 7, Ž . Ž . 18:1n y 9, arachidonic acid ARA; 20:4n y 6 , eicosapentaenoic acid EPA; 20:5n y 3 and Ž . docosahexaenoic acid DHA; 22:6n y 3 were the principal FA. All tissues and nauplii displayed n y 3 ) n y 6 and EPA ) DHA relationships, and contained high proportions of n y 3 highly Ž . unsaturated FA n y 3 HUFA . During sexual maturation, the sum of poly-unsaturated FA Ž . PUFA decreased in the ovaries due to the decrease in n y 6 PUFA such as ARA. The sum of Ž . mono-unsaturated FA MUFA , on the other hand, increased in the ovaries. AA levels were high ) Corresponding author. Laboratory of Aquaculture and Artemia Reference Center, Ghent University, Rozier 44, B-9000 Gent, Belgium. Tel.: q32-9-264-37-54; fax: q32-9-264-41-93.Ž . E-mail address: r.wouters@inve.be R. Wouters . Wouters et al.r Aquaculture 198 2001 307-323 308 in immature, maturing and mature ovaries. They were low in the ovaries of spent females and nauplii. Vitamin E levels were low in immature ovaries, increased substantially during ovarian maturation, and then decreased again upon spawning. High vitamin E levels were retained in the nauplii. The findings of this study, combined with those reported in related studies, suggest the importance of n y 3 HUFA for larval development, of vitamin C for egg development and hatching, and of vitamin E for ovarian maturation and larval development. These nutrients cannot be synthesised de novo by shrimp, and should be included at high levels in the broodstock diet. q

Several bacteriological surveys were performed from 1994 to 1996 at different Litopenaeus vannamei hatcheries (in Ecuador) and shrimp farms (in Mexico). Samples were taken from routine productions of healthy and diseased L. vannamei larvae, postlarvae, and their culture environment and from healthy and diseased juveniles and broodstock. In Ecuador, the dominant bacterial flora associated with shrimp larvae showing symptoms of zoea 2 syndrome, mysis mold syndrome, and bolitas syndrome has been determined. Strains were characterized by Biolog metabolic fingerprinting and identified by comparison to a database of 850 Vibrio type and reference strains. A selection of strains was further genotypically fine typed by AFLP. Vibrio alginolyticus is predominantly present in all larval stages and is associated with healthy nauplius and zoea stages. AFLP genetic fingerprinting shows high genetic heterogeneity amongV. alginolyticus strains, and the results suggest that putative probiotic and pathogenic strains each have specific genotypes. V. alginolyticus was found to be associated with larvae with the zoea 2 syndrome and the mysis mold syndrome, while different Vibrio species (V. alginolyticus and V. harveyi) are associated with the bolitas syndrome. V. harveyi is associated with diseased postlarvae, juveniles, and broodstock. The identities of the strains identified as V. harveyi by the Biolog system could not be unambiguously confirmed by AFLP genomic fingerprinting.Vibrio strain STD3-988 and one unidentified strain (STD3-959) are suspected pathogens of only juvenile and adult stages. V. parahaemolyticus, Photobacterium damselae, and V. mimicus are associated with juvenile and adult stages.