Materials and methods
Gravid adult female sea lice were removed from farmed Atlantic salmon and larvae were hatched and reared in the laboratory at 10°C. Infective copepodids were used to infect 20 Atlantic salmon. Single fish were sacrificed every 24h and all attached larval stages removed, perforated to allow fixative penetration and fixed for transmission electron microscopy according to the methods of Eisenman & Alfert (1982) modified by omission of the pre-fixation step. Chalimus II larvae, present from day 10 to day 18 post-infection, and thereby providing the longest larval moulting sequence for examination, were chosen as the subject for this study.
Specimens for TEM were post-fixed in 1% OsO4, dehydrated through a graded acetone series and embedded in Spurr resin. The resin was polymerised at 70°C. 80nm sections were cut on a Reichert Ultracut E and stained with uranyl acetate and lead citrate according to the methods of Hayat (1989) and Reynolds (1963) respectively. The uranyl acetate method was modified by the use of methanol rather than water as the solvent. Grids were observed using a Philips 301 TEM running at 80 KV. Observations, except where stated, concern the dorsal cephalothoracic cuticle.