42 



ONTOGENY AND SYSTEMATICS OF FISHES-AHLSTROM SYMPOSIUM 



nell, 1976; Theilacker, 1978). These techniques will suffice for 

 the examination of soft tissue morphology given adequately 

 fixed specimens. To avoid their loss, small specimens may be 

 prestained with borax-carmine before embedding and section- 

 ing; this stain can be washed out before subsequent histological 

 staining (Engen, 1968). 



Plastic embedding (Bennett et al., 1976) is advantageous for 

 examination of small delicate structures, for precise records of 

 specimen orientation and section plane, and for the resolution 

 of fine cellular detail. Glycol methacrylate (Bennett etal., 1976), 

 epoxy resins (Humason, 1979), and other low viscosity plastics 

 (Hulet, 1978; L. R. White resin, London Resin Company Lim- 

 ited) are useful embedding media. Small specimens that can 

 become indistinguishable or even lost in paraffin blocks can be 

 easily observed in the plastic block during sectioning. As whole 

 mounts, specimens can be examined, measured, and meristic 

 characters counted before sectioning (Hulet, 1978). Techniques 

 developed by Ruddell (in press) reduce swelling of tissues, an 

 artifact sometimes encountered with glycol methacrylate 

 embedding. While the spectrum of histological and histochem- 

 ical stains applicable to plastic sections is somewhat limited, 

 toluidine blue counter stained with acid fuchsin has staining 

 reactions analogous to the more commonly used hematoxylin 

 and eosin. Other stain combinations also are applicable to larval 

 tissue embedded in glycol methacrylate (for examples see Go- 

 voni, 1980; Govoni et al., 1984): alkali blue 68 counter stained 

 with neutral red reveals fine cellular structure; VanGiesen's 

 picric acid counter stained with acid fuchsin reveals collagenous 

 fibers, the anlagen of actinotrichia; periodic acid-Schiff reagent 

 reacts strongly with acid mucopolysaccharides, including chon- 

 dromucin, and can be used to reveal cartilaginous precursors of 

 cartilage (endochondral) bone; alizarin red S reacts with Ca + + 

 ions and can reveal both calcified cartilage and bone. 



Examples of Application 



Histological preparations may serve the systematist in two 

 ways: by clarifying tissue composition and by resolving struc- 

 ture, thereby allowing for the determination of ontogenetic pres- 

 ence or absence of tissues and by offering comparisons of tissue 

 organization among taxa. 



An example of the first use is in the identification of cartilage 

 and bone. The literature is replete with errors that result from 

 the naive interpretation of alcian blue and alizarin red S reac- 

 tions with cartilage and bone tissue in whole mounts. Alcian 

 blue reacts histochemically with the sulfate and carboxyl groups 

 of mucopolysaccharides (Pearse, 1968) including chondromu- 

 cin, the ground substance of cartilage, but it may also react with 

 developing bone matrices, which are rich in mucopolysaccha- 

 rides as well (Belanger, 1973). An alcian blue reaction, therefore, 

 may indicate cartilage when developing membrane (dermal) bone 

 is present. The reaction of alizarin red S with calcium ions 

 (Pearse, 1968) may indicate calcified cartilage as well as true 

 bone. While the clearing and staining of skeletal elements re- 

 mains a powerful tool (Potthoff, this volume), histological prep- 

 arations can clarify the identity of cartilage and bone tissue in 

 extremely small specimens wherein their identity may not be 

 clear in whole mounts. 



To date, comparisons of larval fish characters revealed by 

 histological techniques have not been extensive and examples 



of application are few. Comparative histological sections of elo- 

 pomorph and clupeomorph larvae illustrate the unique char- 

 acter of the elopomorph leptocephalus (Smith, this volume). 

 The unique configuration of organs and tissues is apparently 

 inclusive of anguilliform, elopiform, and notocanthiform lep- 

 tocephali. Inasmuch as Hulet (1978) also found peculiarities in 

 the kidney structure of the eel leptocephalus that may be unique 

 among vertebrates, the kidney structure of anguilliform lepto- 

 cephali should be compared with that of other elopomorph 

 leptocephali. Transient, hyaline plates occur in the basal end of 

 the outer integumentary epithelium of some clupeiform larvae 

 (Jones et al., 1966; Lasker and Threadgold, 1968; O'Connell, 

 1981a; Fig. 17 A), but this feature was not mentioned in the 

 integumentary descriptions of anguilliforms (Hulet, 1978) and 

 pleuronectiforms (Wellings and Brown, 1969; Roberts et al., 

 1973), nor is it apparent in the perciform Leiostomus xanthunis 

 (Fig. 1 7B). These plates presumably function as osmotic barriers 

 (O'Connell, 1981a), but their systematic presence or absence is 

 not completely established and remains unexplained. The or- 

 ganization of axial musculature is another histological difference 

 among higher taxa. The two-layered musculature of clupeiform 

 larvae is aligned in opposing directions within myotomal seg- 

 ments (Blaxter, 1969b; O'Connell, 1981a; Fig. 17C), whereas 

 in perciform larvae the orientation of axial muscle fibers is 

 closely parallel (O'Connell, 1981a; Fig. 1 7D); this difference may 

 have a functional basis related to gross body form and swimming 

 postures (O'Connell, 1981a). 



An example of the use of histological preparations to compare 

 microanatomical characters is the differences exhibited in elon- 

 gate dorsal fin rays. Elongate dorsal fin rays are features of many 

 unrelated taxa offish larvae (Moser, 1981), but the microana- 

 tomical structure of these homologous derivatives differs among 

 taxa (Govoni et al., 1984). A major difference is the bilateral, 

 paired, collagenous supporting elements of the carapid elongate 

 ray, as in Echiodon dawsoni (Fig. 1 7E), and the singular supports 

 of elongate rays of the bregmacerotid Bregmaceros atlanticus 

 (Fig. 17F) and the serranid Liopropoma (Kotthaus, 1970). 

 Monophyly in carapids has been inferred, in part, from the 

 distinctiveness of this synapomorphy, the elongate first dorsal 

 ray of their highly specialized larvae (OIney and Markle, 1979; 

 Markle and OIney, 1980; Gordon et al., this volume). 



The often remarkable similiarity of cells and tissues, even 

 among phyla (Andrew, 1959), and the development of tissues 

 from the undifferentiated to the complex, may limit the use of 

 a histological approach to systematics. Yet, the unusual diversity 

 that characterizes ontogenetic patterns of fishes (Wourms and 

 Whitt, 1981), and some apparent contrasts in tissue organiza- 

 tion and composition that correlate with current supraordinal 

 classification, make histological comparisons tenable. The pre- 

 ceding examples of tissue and microanatomical dissimilarities 

 may serve to illustrate the kinds of comparisons that may prove 

 useful in inferring relationships as more information becomes 

 available. Histological techniques may provide a potentially 

 useful tool to the systematist; more comparative work is clearly 

 warranted. 



National Marine Fisheries Service, Southeast Fisheries 

 Center, Beaufort Laboratory, Beaufort. North 

 Carolina 28516. 



