H. Nelis

Variation of ascorbic acid content in different live food organisms.

Abstract Ascorbic acid (AA) is an essential nutrient both in particle and live aquafeeds. In order to better assess the needs for this nutrient during larviculture the AA content of algae, rotifers and Artemia was studied with respect to their suitability at startfeeding. In general, the microalgae evaluated were rich in AA (1000–4000 μg AA/g DW), but showed a considerable variability among the different species: e.g. the concentration in Isochrysis and Chlorella reached values 3-to 4-fold the percentage of Tetraselmis (0.11 % of DW). Brachionus routinely cultured on Chlorella contained 2300 μg AA/g DW. Cysts of various batches and strains of Artemia differed considerably in ascorbic acid-2-sulphate (AAS) concentration (296–517 μg AA/g DW). The amount of AA available in the freshly-hatched nauplii reflected exactly the AAS reserve present in the cysts, what evidences the complete conversion of AAS to free AA during completion of embryonic development into nauplii. Boosting techniques both for Brachionus and Artemia using ascorbyl palmitate (AP) as the vitamin C source were established. The addition of 20% AP in the diet of Brachionus enhanced their AA content 10-fold over 3 days of culture. Supplementation of the enrichment emulsion for Artemia with 20% AP increased the AA content up to 2000 μg/g DW after 24 h enrichment. This lipophilic derivative of AA appeared to be a stable form of vitamin C for enhancing AA levels in the live diets during culture and/or enrichment. This bioencapsulation method provides a tool for hatcheries to build up high AA concentrations in the live prey administered to first feeding larvae of aquaculture organisms in case of specific requirements (e.g. with respect to handling stress, deformities). A survey of commercial hatcheries indicated that a wide range of products is used for the cultivation and boosting of rotifers, which consequently affect their AA levels. In general, the AA content in the algae and, consequently, the algal enrichment of Brachionus tended to score lower in the hatcheries than under lab conditions.

An oral challenge for turbot larvae with Vibrio anguillarum

M. Chair 1, M. Dehasque 1, S. Van Poucke 2, H. Nelis 2, P. Sorgeloos ~* and A.P. De Leenheer 3 1 Laboratory of Aquaculture and Artemia Reference Center, University of Ghent, Rozier 44, B-9000 Ghent, Belgium 2Laboratory for Pharmaceutical Microbiology, University of Ghent, Hare/bekestraat 72, B-9000 Ghent, Belgium 3Laboratory of Medical Biochemistry and Chemical Analysis, University of Ghent, Harelbekestraat 72, B-9000 Ghent, Belgium

Retention mechanisms of tetracyclines on a C8 reversed-phase material

Abstract The influence of eluent parameters on the retention of tetracyclines on a C8 reversed-phase column has been investigated. Optimal performance was obtained using eluents containing organic acids at relatively low pH, where the solutes mainly exist in their cationic forms. In contrast to earlier studies, which suggested that ionpair formation was the separation mechanism for tetracyclines, a more complex model is presented. Retention behaviour in the present system was not consistent with one single mechanism but was obviously caused by a mixture of phenomena. Essentially, several ionic interactions are likely to predominate over purely hydrophobic effects.

The effect of supplemental ascorbic acid in enriched live food for Clarias gariepinus larvae at startfeeding

Abstract The effect of three dietary ascorbic acid (AA) concentrations, each applied via two feed types, on production characteristics and physiological condition of African catfish ( Clarias gariepinus ) larvae has been assessed in two 10-day culture trials. Three treatments received only Artemia nauplii enriched with an experimental emulsion containing 0, 10, or 20% ascorbyl palmitate (AP) and yielding 530, 1200 and 1600 μg AA g −1 DW Artemia , respectively; the other three treatments were fed the same Artemia diets which were partially substituted by an artificial diet containing no vitamin C (ratio 20:80). No differences in survival could be observed; however, from day 6 onwards the 20%-AP group showed significantly better growth compared to the 0%- and 10%-AP treatments. For the cofeeding series, the same positive, but not significant, influence of vitamin C on dry weight was found. Moreover, the animals receiving the highest vitamin C supplementation displayed a considerably lower stress sensitivity than those of the 0%- and the 10%-AP groups, for both the 100%- and the 20%- Artemia series. These differences had occurred by day 2, which might be indicative of the importance of AA in early development. A second trial, which was a repetition of the first one, revealed the same tendencies; however, growth differences were smaller, probably due to the higher incorporation levels of AA obtained in the live diet (530, 1700 and 2300 μg AA g −1 DW) and in the catfish larvae. Growth results of both experiments were supported with data from the ultrastructural evaluation of the hepatocytes; i.e. a more organized cell compartmentation and better-structured cell organelles in the 20%-AP group of the Artemia series compared to the control are indicative of a more active metabolism. The slow growth in the cofeeding series was documented by the poor condition of the hepatocytes. In a third experiment it was verified that the growth effect of the 20%-AP boosted Artemia diet was the result of the extra AA incorporation and not of the concomitant palmitic acid (PA), which was set free after hydrolysis of AP in the Artemia nauplii, and which could possibly be used as a supplemental energy source. The three treatments were fed Artemia nauplii enriched with 0% AP, 12% PA and 20% AP, respectively. Growth and stress resistance of the latter group were significantly higher compared to the control and the PA-supplemented fish. To our knowledge this is the first evidence for the positive role of high dietary vitamin C levels (more than 1500 μg AA g −1 DW) on larval development of an aquaculture species, and more specifically of C. gariepinus .

Effect of diet on the photoperiodic induction of diapause in three species of predatory mite, Amblyseius potentillae, A. cucumeris and Typhlodromus pyri

The predatory mitesAmblyseius potentillae, A. cucumeris andTyphlodromus pyri entered diapause in response to a short-day photoperiodic regime, when they were reared on pollen of the ice plant,Dorotheanthus bellidiformis. With pollen of the broad bean,Vicia faba, as food, however, diapause was virtually absent inA. potentillae andA. cucumeris under the same short-day regime, but full diapause was found inT. pyri. The importance of carotenoids for the photoperiodic response in these predatory mites is discussed.

Evaluation of vitamin C-enriched Artemia nauplii for larvae of the giant freshwater prawn

The effect of high levels of ascorbic acid (AA) delivered through enriched live food has been verified through the successful culture of larval giant freshwater prawn, Macrobrachium rosenbergii. Two successive feeding trials were set up using a control (550 Μg AA g−1 DW) and two different AA-enrichment levels in Artemia (1300 and 2750 Μg AA g−1 DW). Under standard culture conditions, no differences in growth nor survival could be observed demonstrating that the nutritional requirements are below 550 Μg AA g−1 DW, which is the normal level occurring in freshly-hatched Artemia. However, a significantly positive effect could be demonstrated on the physiological condition of the postlarvae, measured by means of a salinity stress test, when vitamin C-boosted live food was administered. Since the AA levels in the predator larvae are linked with the enrichment levels in the live prey, it may be assumed that a positive influence on stress resistance was caused by feeding vitamin C-enriched Artemia. It is expected that under suboptimal conditions, supplementation of high vitamin C levels might also enhance production characteristics.

Liquid chromatographic determination of efficacy of incorporation of trimethoprim and sulfamethoxazole in brine shrimp (Artemia spp.) used for prophylactic chemotherapy of fish.

The brine shrimp Artemia, an excellent live food source in aquaculture, has been studied as a carrier to deliver selected chemotherapeutic agents to fish for prophylactic treatment of infectious diseases. To monitor the efficiency of incorporation of trimethoprim and sulfamethoxazole in Artemia franciscana, a sensitive and specific analytical method was developed. It is based on homogenization of Artemia nauplii in methanol, extraction of lipids with hexane, solid-phase cleanup on C18 cartridges, and reversed-phase liquid chromatography with detection at 210 nm. The method is sensitive (detection limit, on the order of 3 micrograms/g with a sample quantity of 30 mg [dry weight]) and reproducible (coefficients of variation, 2.2 and 1.8% for trimethoprim and sulfamethoxazole at levels of 79.6 and 257 micrograms/g of body weight, respectively). Preliminary quantitative data indicated excellent uptake and persistence of both therapeutic agents in A. franciscana, with levels of 115 micrograms/g for trimethoprim and 277 micrograms/g for sulfamethoxazole.

Influence of dietary vitamin C dosage on turbot (Scophthalmus maximus) and European sea bass (Dicentrarchus labrax) nursery stages

Abstract Stable forms of vitamin C were verified as dietary sources of ascorbic acid (AA) for the nursery stages of European sea bass and turbot. In a first experiment, various concentrations of ascorbyl palmitate (AP) and one level of AA 2-polyphosphate (ApP) were evaluated in a semi-purified diet for European sea bass, Dicentrarchus labrax . A significantly lower AA concentration was detected in the fry fed the AP-supplemented diets compared to the ApP-fed group, providing evidence that AP is an inferior source of vitamin C for young stages of sea bass. ApP was subsequently selected for the determination of the AA requirements of both sea bass and turbot ( Scophthalmus maximus ) fry. The AA levels incorporated in the body tissues reflected the levels of ApP in the diet, providing evidence of the bioavailability of this stable phosphate ester. Reduced production results were only obtained in the AA-free treatment, indicating that a level of 20 mg AA/kg is sufficient for sea bass and turbot during nursery rearing. This was supported with data from the ultrastructural evaluation of the hepatocytes. Results of challenge tests with pathogenic Vibrio strains remained inconclusive and did not provide evidence for an immunostimulative effect of high AA levels during the early post-weaning phase of these species.

Communications: Bioencapsulation of the Antibacterial Drug Sarafloxacin in Nauplii of the Brine Shrimp Artemia franciscana

Abstract Nauplii of the brine shrimp Artemia franciscana were enriched with the antibacterial drug sarafloxacin hydrochloride to determine if levels effective against four pathogenic strains of Vibrio spp. would accumulate in brine shrimp. Three vibrio strains were Vibrio anguillarum; the fourth was V. vulnifcus. Concentrations of sarafloxacin ranging from 1 to 40% (weight volume) were incorporated into the oil phase of Super Selco brine shrimp diets. Enriched nauplii and controls were sampled at 2-h intervals for 24 h. Antibiotic sensitivity testing was evaluated with a modification of the Kirby–Bauer disk diffusion method. A microbiological assay with Escherichia coli was used to determine the concentration of sarafloxacin per sample of nauplii. The results indicate that sarafloxacin is rapidly taken up by larval brine shrimp and can be detected at 2 h of enrichment by microbiological assay. The efficacy of sarafloxacin appears to depend on the strain of pathogen present.

Reversed-phase ion-pair chromatography of anthracyclines

Abstract A new reversed-phase liquid chromatographic separation of anthracyclines on a spherical octadecylsilica material is described. The effect of variation of the content of organic modifier, pH, counter-ion concentration and ionic strength on retention and selectivity has been investigated. There is evidence that, in acidic eluents not containing a counter-ion, mixed retention mechanisms are operative. Solvophobic interactions appear to predominate in water-rich eluents, whereas high organic modifier concentrations favour polar-type retention mechanisms, possibly involving silanol groups. An eluent containing sodium dodecyl sulphate afforded improved efficiency and resolution and was used to resolve eight anthracycline derivatives in one isocratic run.