Anisakids are parasitic nematodes of fish worldwide, producing economic and human health concerns. It is thus important to ascertain their in vitro life cycle in laboratory studies. Here we describe the in vitro development of third-stage larvae (L3) of Hysterothylacium aduncum isolated from blue whiting Micromesistius poutassou, to the hatching of L3 from eggs obtained from H. aduncum worms grown in GLIT medium (a modified mixture of Yaeger's LIT [Liver Infusion Tryptose] and Grace's media) at pH 4.0, 13°C and with 5% CO 2 in air. Under these conditions, L3 recovered from fish developed to mature adults (3.4 to 6.2 cm in length), with oviposition starting from Day 26 in culture. Fertilized eggs (mean size 64 × 52 µm) had a thick, rugose eggshell and were larger than unfertilized ones (mean size 49 × 42 µm), whose eggshells were thin and smooth. Eggs laid during the first and second week of oviposition, and maintained in 2.8% NaCl solution at 13°C, developed to L3. Under these maintenance conditions, between 20.6 and 52.5% of the eggs laid during the first week developed into larvae. Motile larvae, enclosed in a sheath, hatched from between 2 and 11% of these eggs. The larvae started to hatch 23 d after deposition. These larvae were 144 to 215 µm in length, enclosed in a 237 to 305 µm-long sheath. This GLIT culture medium may help to study the biology of this and other anisakids.KEY WORDS: Fish parasite · Hysterothylacium aduncum · Nematoda · Anisakidae · In vitro culture · Egg development · Hatching
Resale or republication not permitted without written consent of the publisherDis Aquat Org 58: [41][42][43][44][45] 2004 mesistius poutassou (blue whiting), family Gadidae, purchased from the fish market of Granada (southern Spain). Blue whiting, on the Atlantic and Mediterranean Spanish coasts, are frequently parasitized by H. aduncum (Ruiz-Valero et al. 1992, Valero et al. 2000. The worms, found free in the host body cavity, were collected with the help of a needle with a blunt tip, placed on a Petri dish and washed in 0.9% NaCl solution several times. The worms were observed individually under an inverted microscope, and those that showed any kind of internal or external damage were discarded. They were then identified according to morphological features (Hartwich 1975, Petter & Maillard 1988, Petter & Cabaret 1995.Prior to cultivation, each larva was placed in an antibiotic-antifungal solution (80 mg gentamicin sulfate, 0.625 mg amphotericin B, 10 000 IU sodium penicillin G, 10 mg streptomycin sulfate and 4.5 ml Hanks' solution, for a final solution volume of 10 ml) and axenized as described elsewhere (Iglesias et al. 1997, Iglesias et al. 2001. Worms were cultured in sterile polystyrene 30 ml flasks. The culture medium (10 ml) was placed into each flask with 8 to 10 parasites. The culture flasks were then placed in an incubator at 13°C and 5% CO 2 in humid air, and the culture medium was renewed once a week. The worms were observed daily for motility, moulting and survival. The culture medium (GLI...