A scanning electron microscopy study of Anisakis physeteris molecularly identified: from third stage larvae from fish to fourth stage larvae obtained in vitro

The development of the fourth larval stage (L4) of Anisakis physeteris was studied using scanning electron microscopy (SEM), comparing it with third larval stage (L3) recently obtained from the host fish, blue whiting (Micromesistius poutassou), from the western Mediterranean Sea (east coast of Spain, zone FAO 37.1.1). After molting to L4, samples of the parasite were examined at different times in order to observe their development. Following collection of the L4, a small portion was taken from the middle of the larva for molecular identification, confirming in all cases that it was A. physeteris. The anterior and posterior sections of the larvae were prepared for morphological study by SEM. The development of a row of denticles on each of the three prominent lips, almost reaching the buccal commisures, was observed in the L4. Pores of unknown function were found in the upper external part of each lip. Clearly developed cephalic papillae, amphids, and deirids were also observed in L4, while, although present in L3, these were beneath the cuticle. Phasmids were detected in L4 but not in L3. The L4 tail finished in a conical lobe with a blunt point, absent in L3. In the oldest L4, some preanal papillae were observed beneath the cuticle in males, while, in females, the vulva could be seen by light microscopy, apparently still covered by the cuticle.