44 



ONTOGENY AND SYSTEMATICS OF FISHES-AHLSTROM SYMPOSIUM 



introduced into the freeze dryer, where water subUmes, leaving 

 the specimen dry and intact. Critical point drying, on the other 

 hand, requires dehydration through a graded series of alcohols 

 (20% for 24 h. then 10-20 min each in 50%, 70%, 80%, 90%, 

 95%, and two changes of absolute ethanol). The ethanol is then 

 replaced with either freon or acetone depending on whether 

 freon or carbon dioxide critical point dryers are used. The steps 

 of dehydration and transfer can be done in small specimen 

 holders to minimize handling and possible surface damage. Af- 

 ter dehydration, specimens must be mounted on SEM studs 

 with any of several available adhesives and tapes. The dried 

 specimens are particularly delicate and should be handled with 

 a small camel-hair brush to avoid damage to the surface. They 

 are then oriented onto the stud under a dissecting microscope. 

 Before coating, no further preparation is necessary with larvae, 

 but eggs have only a small area of electrical contact with the 

 stud. It is therefore advisable to use a conductive adhesive (such 

 as silver paint) to make a more complete electrical connection 

 and prevent charging, which decreases image quality. This paint 

 should be allowed to become tacky prior to positioning the eggs, 

 or it may cover portions of the egg itself Finally, specimens are 

 coated with a thin conductive layer, typically of gold or gold- 

 palladium, by either vacuum evaporation or ion sputtering, prior 

 to viewing on the SEM. At most facilities, trained SEM tech- 

 nicians are available; their advice and assistance are invaluable 

 and should be sought. 



Results and Discussion 



Shrinkage and other artifacts will vary depending upon the 

 type of material, preservation, and method of dehydration. For 

 fresh material preserved in a mixture of formalin, glutaralde- 

 hyde, and acrolein, Stehr and Hawkes ( 1979) observed a shrink- 

 age of approximately 10% in the eggs of Platichthys stellatus 

 and Oncorhynchus gorbuscha; the latter had been punctured 

 prior to dehydration. In the present study, eggs of Maurolicus 

 muellen initially preserved in 5% buffered formalin showed 

 varying degrees of shrinkage and collapse depending upon sub- 

 sequent treatment. The least shrinkage (12%, Fig. 18B) was 

 noted in material which was freeze dried, whereas post-fixation 

 and dehydration through freon 1 1 3 associated with critical point 

 drying resulted in shrinkage of up to 67% of the original diameter 

 (Fig. 18D). Eggs of this species show a hexagonal sculpturing; 

 under the light microscope the sculpturing is hyaline and difficult 

 to interpret (Fig. 18A). Eggs prepared by freeze drying clearly 

 show the surface sculpturing; note particularly the ridges, which 

 are more clearly defined (Fig. 188). For comparison, an egg 

 which had partially collapsed during dehydration is shown (Fig. 

 18D). The obvious differences in shrinkage point out the im- 

 portance of specifying method, initial size, and shrinkage values, 

 particularly for comparative or taxonomic studies. 



Eggs from other species are shown to give an idea of the range 

 of chorion structures which may be observed. The hexagonal 

 pattern on M. muellen overlies a highly porous surface structure 



Fig. 18. (A) Egg of Maurolicus muellen from off South Africa taken under the compound light microscope with transmitted, polarized light. 

 Note the emphasis of the points on the hyaUne chorion, which represent the intersections of ridges. Bar = 100 ^m. (B) Egg of A/, muellen under 

 the scanning electron microscope. Note the areas between what one would interpret as points on Figure 18A. which are now seen as polygonal 

 facets or ridges. Bar = 500 nm. (C) Individual facet of the egg of At. muellen. Note the porous and diaphanous nature of the egg surface. Bar = 

 50 Mm. (D) Egg of A/, muelleri post-fixed in osmium tetroxide and critical point dried. The shrinkage of this specimen is approximately 65%. 

 Note the differences in morphology of the ridges and surface of the egg. Bar = 100 /jm. (E) E^of Pleuronichlhys coenosus. The facets are relatively 

 small by comparison with M. muellen and the pattern units are more regularly hexagonal. Bar = 100 Mm. (F) Detail of two hexagons from the 

 egg of P. coenosus. Note the morphological differences between both the ridges and chorion surface as compared to M. muellen. Bar = 10 Mm. 



Fig. 19. (A) Egg of Alherinopsis californiensis. The filaments are single, terminate in loose ends, and are distributed over the entire egg surface. 

 Bar = 1 ,000 Mm. (B) Egg of .-itherinops affiiUs. The egg of this species is characterized by filaments which are looped, with no free ends (Curless, 

 1979). This differentiates it from the egg of ,-1. californiensis, as do filament length, abundance, and basal morphology. Closed-loop filaments have 

 also been noted in .Aniennanus caudimaculatus eggs by Pietsch and Grobecker ( 1 980). Bar = 1 ,000 Mm. (C) Chorion of Paracaltionymus costatus 

 collected off South Africa. The surface features are irregular and cover the entire egg surface. This differs from species of Callionymus. which 

 have hexagonal patterns. Bar = 10 Mm. (D) Chorion surface of Mugil cephalus. These structures are irregular and cover the entire egg surface. 

 Note the superficial similarity to Paracallionymus. Bar = 10 Mm. (E) Chorion surface of an advanced ovarian egg of Coryphaenoides filifer. Note 

 that the surface "blebs" are arranged in hexagonal patterns and may be the precursors of a hexagonal pattern typical on eggs in this family. The 

 pelagic egg of this species has not been described. Bar = 10 Mm. (F) Chorion surface of an advanced ovarian egg of Coryphaenoides acrolepis. 

 The hexagonal ridges are better developed than in Fig. I9E. There are holes under the ndges between the intersections, which might indicate that 

 this species, whose egg is also undescribed, may have the hexagonal network supported on "stills" as described for eggs of Coelorhynchus spp. 

 (Robertson. 1981; Sanzo, 1933a). Bar = 10 Mm. 



Fig. 20. (A) Spines on the chorion surface o( Oxyporhamphus microplerus. These are distributed over the entire surface of the egg. Bar = 100 

 Mm. (B) Chorion surface from Scomhereso.x saurus collected off South Africa. The tufts are characterized by a relatively complex basal morphology 

 and depending upon method of fixation, may resemble small bundles of hairs or, as here, simply coalesced tufts. Bar = 10 Mm. (C) Micropyle 

 and associated pores of the egg of Laclona diaphana from the Eastern Tropical Pacific. The pores shown here are restricted to this region around 

 the micropyle and appear to penetrate the outer layer of the chorion. Bar = 50 Mm. (D) Secondary, smaller pit structures on the remainder of the 

 egg of Laclona diaphana. I refer to these depressions as "pits" because closer examination does not reveal penetration through any layer of the 

 chorion, as opposed to the pores surrounding the micropyle in 20C. Bar = 1 Mm. (E) Head region of a larval Sebasles melanops shortly after 

 parturition. Polygonal epidermal cells may be noted on some parts of the body. Bar = 100 Mm. (F) Epidermis on the dorsal surface, just posterior 

 to the head, on an embryonic S. melanops approximately 28 days post fertilization. Note the distinct microndges and cell borders characteristic 

 of developing teleost epidermis. Bar = 10 Mm. 



