Comparative Anatomy and Phylogeny of the Cloacae of Salamanders (Amphibia: Caudata)

JOURNAL OF MORPHOLOGY 211:63-72 (1992)

Comparative Anatomy and Phylogeny of the Cloacae of Salamanders (Amphibia: Caudata). Ill. Amphiumidae

DAVID M. SEVER Department of Biology, Saint Mary's College, Notre Dame, Indiana 46556

ABSTRACT Histological descriptions of female and male cloacae are pre- sented for the three species of Amphiuma. Little interspecific variation occurs in either females or males. Females possess numerous spermathecae around the walls of the posterior cloacal tube and anterior cloacal chamber as well as rudimentary vent glands, of uncertain homology, in the caudal end of the cloacal chamber. Ventral glands such as those known from female salamanders in other families are absent, likely a secondary loss in the amphiumid clade. In a female A. tridactylum with large ovarian follicles (5- to 8-mm diameter), the lips of the cloacal orifice are highly cornified, and stored sperm in the spermathecae appear in organized bundles, either free in the lumen or with the sperm heads seemingly buried in the spermathecal epithelium. Males possess some gland clusters present in other members of the suborder Salamandroidea (dorsal pelvic glands, vent glands, Kingsbury's glands, anterior ventral glands) but lack others (lateral pelvic glands, posterior ventral glands), resulting in the most plesiomorphic cloacal anatomy in the suborder. However, males possess an autapomorphic gland cluster, called the pit glands, in the posterior wall of the cloacal chamber. The pit glands consist of shallow infoldings of the cloacal epithelium unlike other cloacal glands, which are relatively tubular and elongate. The "pits" contain columnar eosinophilic cells that give positive staining reactions for acid mucopolysaccharides and proteins.

The salamander family Amphiumidae con- found that amphiumids possess the most ple- sists of three species of Amphiuma found in siomorphic cloacal anatomy in the suborder the southeastern United States. Amphiumas Salamandroidea. This paper presents histo- are elongate salamanders with reduced limbs, logical descriptions of amphiumid cloacae in and adults retain one external gill slit, a order to extend the observations of Wilson paedomorphic character (Duellman and ('41) and Kreeger ('42) and to provide the Trueb, '86). Individuals of two species, Amph- anatomical basis for the character states used iuma means and A. tridactylum, are large, by Sever ('91). often in excess of 70-cm snout-vent length MATERIALS METHODS (SVL), and are aquatic, while A. pholeter is a AND dwarf species, 20- to 25-cm SVL, that usually SVL refers to the distance from the tip of is found burrowing in mucky banks (Conant, the snout to the posterior end of the vent. '75). Specimens, as far as known, were initially Davison (1895) described the gross anat- preserved in 10% formalin, and some were omy of the cloacae of male and female Amph- stored in isopropanol or ethanol of varying iuma means and was the first to provide concentrations before cloacae were excised anatomical evidence for internal fertilization for histological preparation. Specimens used in the family. Kreeger ('42) provided a histo- are in the collections of the University of logical description of the cloaca of female A. Michigan Museum of Zoology (UMMZ), Flor- tridactylum, but the only such account for ida Museum of Natural History (FM), or in male amphiumids is an abstract by Wilson my possession (DMS). The following speci- ('41) on the same species. Sever ('91) listed mens were examined histologically: Amphi- presencelabsence data for 23 cloacal characters in amphiumids and other salamanders, Address reprint requests to Dr. David M. Sever, Department of and his cladistic analysis of these characters Biology, Saint Mary's College, Notre Dame, IN 46556.

G 1992 WILEY-LISS, INC. 64 D.M. SEVER uma means: female, DMS 3005, 232 mm, image reconstruction. Right lateral views June 22,1977, Alachua County, Florida, and were reconstructed by computer software, male, UMMZ 187365, 520 mm, April 14, with the sections rotated 25” clockwise. The 1987, Marion County, Florida; A. pholeter: cloacal displays were stretched appropriately female, UMMZ 186277, 238 mm, July 31, in the anteroposterior direction to help allevi- 1985, and male, FM 62441,160 mm, May 12, ate geometric distortions caused by skipping 1985, Levy County, Florida; and A. tridacty- sections between those digitized. Each digi- lum: females, DMS 16111, 526 mm, DMS tized section of cloacal glands was repeated 16110, 528 mm, and UMMZ 187364, 575 once so that gaps between displayed sections mm, collection dates unknown, Orleans Par- were more nearly closed. Finally, alternate ish, Louisiana, and males, UMMZ 187363, digitized sections of the cloacal walls were 421 mm, and UMMZ 187366, 481 mm, no displayed to improve clarity. other data, and DMS 4103,725 mm, May 29, Following Sever (’78),the cloaca is divided 1974 (preserved 17 November 1980), Or- into the cloacal tube and the cloacal chamber. leans Parish, Louisiana. The cloacal tube is cylindrical and extends Following dehydration in ethanol, the ex- caudally to the cavity dorsal to the vent, the cised cloacal region was cleared in toluene or cloacal chamber. Relative length of the cloa- Histosol (National Diagnostics, Inc., Man- cal tube (cloacal tube length/total cloacal ville, NJ), embedded in paraffin, and 10-pm length = CTLITCL elsewhere) may have sagittal (male Amphiuma means) or trans- phyletic significance (Sever, ’91). To deter- verse (remaining specimens) sections cut with mine lengths, the number of transverse sec- a rotary microtome. Sections from each spe- tions of the cloacal tube and cloacal chamber cies were stained with hematoxylin-eosin were counted. The most anterior section of (H&E) (for general cytology), and additional the cloacal tube was considered the one imme- sections from some specimens were treated diately posterior to the junction of the Wolf- with diagnostic stains using procedures de- fian ducts with the hindgut. scribed by Humason (’79). Slides from female A. pholeter and one female A. tridacty- RESULTS lum (UMMZ 187364) were stained with the Female cloacal anatomy ninhydrin-Schiff reactiodfast green (pro- One female Amphiuma tridactylum teins), periodic acid-Schiff reagent/ fast green (UMMZ 187364) has sperm in the spermath- FCF (PAS, neutral carbohydrates), and al- ecae and enlarged ovarian follicles (5-8 mm cian blue/nuclear fast red at pH 2.5 (acid in diameter). The other A. tridactylum and mucopolysaccharides). The cloacal sections the A. means lack sperm in their spermathe- of male A. pholeter were not subjected to cae and possess small ovarian follicles (<1 diagnostic staining, but some from male A. mm in diameter). Spermathecae are well de- means were stained with alcian blue. Sec- veloped but devoid of sperm in the A. pholeter tions from one male A. tridactylum (UMMZ examined. Three-dimensional reconstruc- 187363) were treated with PAS and alcian tions through the cloaca of A. pholeter are blue, and sections from another (DMS 4103) shown in Figure 1, and transverse sections were treated with PAS, alcian blue, and nin- through cloacae of A. pholeter and A. tridac- hydrin Schiff. No tests for lipids were done tylum are illustrated in Figure 2. since some reagents used for paraffin embed- ding remove lipids. Secretory products with a Cloaca1 conformation and anatomy of the stringy appearance were considered “fibrous” linings while spheroidal secretory droplets were The CTL/TCL quotient is 0.41 in A. means called “granular,” if uniformly small, or and 0.49 in A. pholeter. Some anterior and/or “globular,” if composed of coalesced gran- posterior sections are lacking in the Amphi- ules. uma tridactylum examined; thus the CTL/ Three-dimensional reconstructions of cloa- TCL could not be determined for that spe- cae of female and male Amphiuma pholeter cies. were done using PC3D software (Jandel Sci- Posterior to the junction of the urogenital entific, Corte Madera, California) and a Jan- ducts, the cloacal tube has rugose walls lined del digitizing tablet with a Zenith ZF-248 with glandular, alcian blue+, pseudostrati- microcomputer. Every sixth (female) or tenth fied epithelium. Some epithelial cells in the (male) transverse section was digitized for cloacal tube possess cilia. Cilia do not occur AMPHIUMA CLOACAE 65 Ct with enlarged ovarian follicles and spermatozoa in the spermathecae is especially thick and cornified (Fig. 2G).

Cloacal glands Spermathecae and vent glands occur in all specimens examined. In the Amphiuma tridactylum with large ovarian follicles, sperma- A tozoa occur in short, tubular glands in the A posterior half of the cloacal tube and the anterior two-fifths of the cloacal chamber, defining these glands as spermathecae. Glands similar in appearance and location, but devoid of sperm, occur in these regions in the other female amphiumids examined. An- teriorly, the first spermathecae appear dorso- medially, and more posteriorly they spread around the dorsal and lateral walls of the cloaca (Figs. lB, 2A-D). In one A. tridacty- 8 lum (DMS 16111) and in the A. means examined, some glands appear around the inferior border of the cloacal tube as well (the “ven- B tral group” of spermathecae described by Kreeger, ’42).The most posterior spermathe- Fig. 1. Three-dimensiond reconstructions of the clo- cae are dorsomedial and, in A. pholeter, pass acal cavities and cloacal glands of female Amphiuma pholeter. Lateral view, with the anterior end (toward the into portions of the cloacal chamber where right) rotated clockwise 25”. A. Cloacal tube and cloacal the lining has become epidermal (Fig. 2D). chamber. B. Relationships of spermathecae and vent Spermathecal cytoplasm is scant and the glands to the cloacal cavities. The distance between sections of the cloacal walls represents 120 pm. Cc, cloacal cells squamous in Amphiuma means and in chamber; Ct, cloacal tube; St, spermathecae; Vg, vent the A. tridactylum lacking stored sperm. The glands. spermathecal epithelium is eosinophilic and cuboidal in A. pholeter and in the A. tridacty- dorsally beyond the midpoint of the cloacal lum containing sperm. In A. pholeter, the tube, while cilia remain ventrally until the spermathecae contain some small granules anterior end of the cloacal chamber. that are PAS+ and ninhydrin Schiff-t. The A dorsomedial fold is prominent in the apical cytoplasm of spermathecae is PAS+ in cloacal tube, but at the anterior end of the the individual of A. tridactylum with sperm. cloacal chamber, the dorsal apex narrows, As illustrated by Kreeger (’42)and Sever and the dorsolateral wall becomes less rugose (’871, the sperm in the spermathecae are (Figs. lA, 2A-C). Ventrally, three pairs of organized into bundles with sperm heads usu- rugae continue from the cloacal tube into the ally oriented in the same direction. Some anterior end of the cloacal chamber. The groups of sperm appear free in the lumen infolding of epithelium at the anterior end of while others seem to have their heads embed- the cloacal orifice is epidermal, consisting of ded in the spermathecal cytoplasm. Light stratified cuboidal cells with a superficial microscopy does not provide the necessary cornified layer (Fig. 2B,C). The epidermal resolution to determine whether sperm- lining gradually spreads dorsad, replacing the epithelial contacts are coincidental, or if em- pseudostratified lining. In Amphiuma phol- bedded sperm are receiving nourishment or eter, the entire caudal three-fourths of the undergoing degradation. cloacal chamber is lined with epidermis (Fig. A separate group of glands occurs at the 2D,E), but in A. means and A. tridactylum, posterior end of the cloaca in female Amphi- the dorsal end of the cloacal chamber retains uma (Figs. lB, 2E-F). These glands do not the pseudostratified lining until the caudal contain sperm in the A. tridactylum with angle of the vent. The lining of the cloacal sperm in the spermathecae and are cytologi- orifice (cloacal “lips”) in the A. tridactylum cally different from spermathecae in all spec- 66 D.M. SEVER

Fig. 2. Transverse sections through the cloaca of fe- fied epidermis around the cloacal orifice. Bv, blood ves- male Amphiumapholeter (A-E) and A. tridactylum (F). sels; Ce, cornified epidermis of the cloacal lips; Ep, A. Middle of cloacal tube. B. Posterior end of the cloacal epidermal lining of the cloacal chamber; Ng, neural tube. C. Anterior end of the cloacal chamber. D. Start of ganglion. Other labels as for Figure 1. Scale bar in lower the posterior three-fourths of the cloacal chamber. E. right corner = 250 bm in A, B, C, D, 200 km in E, and Posterior fourth of the cloacal chamber. F. Highly corni- 500 bm in F. imens. This posterior group of glands is simi- glands secrete only onto cornified epithelium lar in appearance and position to the vent (Fig. 2E), but in A. means and A. tridacty- glands found of male amphiumids (as de- lum, they secrete onto simple columnar epi- scribed below) and males of many other mem- thelium as well as the cornified linings of the bers of the suborder Salamandroidea (Sever, cloaca. In the A. tridactylum containing '91). Hence, these are considered female vent stored sperm, the cells of vent glands are glands. columnar and basophilic, and the apical cyto- Vent glands occur in the caudal half of the plasm is strongly PAS+, although the lu- cloacal chamber, and some secrete onto the mina are devoid of secretory product. In the skin posterior to the cloacal orifice (Fig. 1B). other specimens, vent gland cytoplasm is In Amphiuma pholeter, these short, tubular scant, and the glands seem inactive. In A. AMPHIUMA CLOACAE 67

Fig. 3. Three-dimensional reconstruction of the clo- tween sections of the cloacal wall represents 200 pm. Av, aca of male Amphiuma pholeter. Lateral view, with the anterior ventral glands; Dp, dorsal pelvic glands; Kg, anterior end (toward the right) rotated clockwise 25". A. Kingsbury's glands; Pi, pit glands. Other labels as for Dorsal pelvic glands. B. Anterior ventral glands. C. Kings- Figure 1. bury's and vent glands. D. Pit glands. The distance be- pholeter, however, small PAS+ granules oc- pseudostratified throughout the cloaca ex- cur in the vent gland cytoplasm. cept on the medial edges of the cloacal orifice where the lining is epidermal and continuous Male cloacal anatomy with that of the skin. The male cloacal anatomy of all three spe- Anteriorly, the cloacal tube is laterally nar- cies of Amphiuma is similar. Three-dimen- rowed dorsally and widened ventrally. In the sional reconstructions through the cloaca of anterior cloacal tube, a middorsal fold is male A. pholeter are shown in Figure 3, and prominent and 4-5 pairs of rugae occur ven- transverse sections through the cloaca of this trally (Figs. 3,4A-B). Posteriorly in the cloa- individual are illustrated in Figures 4 and 5. cal tube, the floor narrows and invaginates The cloacal glands in one individual of A. toward the cloacal orifice. Rugae continue tridactylum (UMMZ 187366) possess scant into the anterior half of the cloacal chamber cytoplasm, small lumina, and no apparent (Figs. 3, 4C-D). The rugae are ciliated in secretory activity, but those of the other spec- individuals with active cloacal glands, project imens examined appear hypertrophied and medially into the cloacal cavity, and are ob- contain secretory material. liquely slanted with the anterior end superior to the posterior end. Cloaca1 conformation and anatomy of the At the midpoint of the cloacal tube, the linings dorsal portion of the cloacal tube widens, and The CTL/TCL quotient is 0.35 in Amphi- the roof in the posterior half of the cloacal umapholeter and 0.16 in one A. tridactylum tube and anterior end of the cloacal chamber (UMMZ 187366). Sagittal sections were cut bears unciliated papillae (Figs. 3, 4B-C). In forA.means, and, thus, a CTL/TCL quotient the anterior end of the cloacal chamber, the could not be determined. The epithelium is dorsal portion narrows and a dorsal invagina- 68 D.M. SEVER

Fig. 4. Transverse sections through the cloaca of male the cloacal chamber. D. Anterior fourth of the cloacal Amphiumapholeter. A. Anterior end of the cloacal tube. chamber. Labels same as for Figures 1and 3. Scale bar in B. Posterior end of the cloacal tube. C. Anterior end of lower right corner = 250 Km. tion occurs into the roof, resulting in the extend beyond the posterior end of the cloa- dorsal portion becoming a narrow slit lined cal orifice (Figs. 3D, 5D). with unciliated papillae (Figs. 3, 4D). Thus, the anterior half of the cloacal chamber con- Cloaca1 glands sists of a narrow cavity with papillae dorsally The pit glands are eosinophilic with colum- and rugae ventrally. nar cells (Fig. 5). In the apical cytoplasm, an In the posterior half of the cloacal cham- alcian blue+ fibrous material and numerous ber, the ventral portion becomes less rugose, granules that are PAS+ and ninhydrin and the lining of the dorsal portion becomes Schiff+ occur. The luminal secretion product glandular and infolded, forming the pit glands appears as a mixture of alcian blue+ fibrous (Figs. 3D, 5A). All cloacal glands are exo- and PAS+ globular material. The other crine, apparently formed by invaginations of eosinophilic glands are the dorsal pelvic the epithelial surface of the cloacal lining, glands and the vent glands. Lateral pelvic but cloacal glands other than the pit glands glands such as those described from males of are relatively elongate, tubular structures some other species in the Salamandroidea with orifices smaller than the diameter of the (Sever, '91) are not present. tubules. The pit glands are simply shallow Dorsal pelvic glands secrete onto the uncil- infoldings of the surface, without definitive iated papillae of the roof of the posterior tubular structure or rounded orifices (Fig. 5). cloacal tube and anterior cloacal chamber The most anterior pit glands are middorsal (Figs. 3A, 4C,D, 5A). Distal ends of glands (Figs. 3D, 5A). Posteriorly, pit glands extend radiate from the cloaca, so that sections ventrally until they occupy the entire lining through tubules occur anterior as well as of the cloaca chamber except for the area just posterior to the area of secretion (Figs. 4A,B, superior to the cloacal orifice (Figs. 3D, 5B,C). 5B). The epithelium generally is cuboidal and, The caudal end of the cloacal chamber is like that of pit glands, contains alcian blue+ abrupt (Fig. 3). Distal portions of pit glands fibrous material and granules that are PAS+ AMPHIUMA CLOACAE 69

Fig. 5. Transverse sections through the cloaca of male Posterior end of the cloacal chamber. D. Posterior to the Amphiuma, continued. A. Midpoint of the cloacal cham- cloacal orifice. Labels same as for Figures 1 and 3. Scale ber. B. Posterior three fourths of the cloacal chamber. C. bar in lower right corner = 250 pm. and ninhydrin Schiff+. The lumen contains a anterior end of the cloacal tube and, after the mixture of these granules in a matrix of the roof flattens and widens, some Kingsbury’s fibrous material, and the amount of secre- glands secrete onto the roof of the cloacal tory product in the lumen is greater than in tube medial to dorsal pelvic glands (Figs. 3, pit glands. 4A,B). Dorsal pelvic glands replace medial The vent glands are short and tubular with Kingsbury’s glands in the posterior end of distal ends that pass posteriorly. Vent glands the cloacal tube. Kingsbury’s glands have a secrete into the caudal end of the cloacal fibrous secretion that is alcian blue+. This orifice and onto the skin surrounding the material is scant in lumina but abundant in posterior end of the vent (Figs. 3C, 5C,D). apices of the columnarlcuboidal epithelial Vent glands replace in position the most cau- cells. dal anterior ventral glands. Vent glands are Anterior ventral glands secrete onto the not numerous, but are convoluted so that surfaces of the rugae that line the lateral two sections of the same gland may appear in walls of the posterior cloacal tube and ante- the same transverse section. The epithelium rior half of the cloacal chamber (Figs. 3B, 4, is columnar, although somewhat festooned, 5A,B). After the inferior lining of the cloacal and contains granules that are PAS+ and chamber becomes smooth midway through ninhydrin Schiff+ . Luminal material, how- the cloacal chamber, some anterior ventral ever, generally is fibrous and alcian blue+. glands secrete into this area, but ventral Like that of pit glands, luminal product is glands are replaced more posteriorly by vent less evident than in dorsal pelvic glands. glands (Fig. 5B,C). Distal ends of anterior Basophilic glands are Kingsbury’s glands ventral glands generally pass cranially (Fig. and the anterior ventral glands. Kingsbury’s 3B). Histochemically, the secretion is like glands are short tubules that secrete along that of Kingsbury’s glands, a fibrous sub- the borders of the dorsomedial fold in the stance that is alcian blue+. Unlike Kings- 70 D.M. SEVER bury’s glands, the lumina of anterior ventral ventral portions of the cloacal tube and ante- glands contain copious amounts of secretory rior cloacal chamber. Kreeger reported a small material. Epithelial cells generally are group of glands at the posterior end of the squamous, and the luminal borders also are cloaca, which she thought were nonfunc- alcian blue + . tional and possibly rudimentary “abdominal” DISCUSSION glands. My results confirm Kreeger’s observations on spermathecae, and the glands she The ancestral condition in all salamanders called “abdominal glands” are here referred except the Sirenidae (which lack cloacal to as female vent glands. glands) is the possession of at least one type As regards the spermathecae, those of fe- of cloacal gland (Sever, ’91). In the clade male amphiumids are similar in appearance leading to extant salamanders in the subor- and distribution to those found in most other der Salamandroidea, parts of the ancestral members of the Salamandroidea with the cloacal gland mass are modified for produc- notable exception of the Plethodontidae, in tion of spermatophores (males) or storage of which all spermathecae unite with the cloa- sperm (females). Other regions of cloacal cal wall by means of a common tube, an glands in both sexes may secrete mating pher- autapomorphic trait. Female amphiumids are omones, representing, perhaps, retention of nearly unique, however, in the absence of the ancestral function for cloacal glands anterior ventral glands and the presence of (Sever, ’91). Homologies of cloacal gland divisions in different groups of salamanders are vent glands. determined by comparative morphology (e.g., The only groups of salamanders in which position of glands in relation to regions of the ventral glands are absent in females are the cloacal cavities, cytology, staining reactions) Sirenidae (in which all cloacal glands are and/or functions of glands in sperm storage, absent), the Rhyacotritoninae of the Dicamp- spermatophore formation, or pheromone pro- todontidae, and the Amphiumidae. In addi- duction. tion, females of some species in each of the Although more work needs to be done on subfamilies of the Plethodontidae lack ven- the functions of amphiumid cloacal glands, tral glands. The absence of ventral glands is my results provide the morphological basis considered the ancestral condition for Si- for recognition of homologous and autapo- renidae and a secondary loss when it occurs morphous cloacal structures in amphiumids. in other species (Sever, ’91). Previous studies, especially on males, lacked Ventral glands in female salamanders usu- histological detail, failed to recognize the com- ally are limited to the anterior end of the parative value of some characters, or misin- cloacal chamber, so the female vent glands of terpreted homologies. Amphiuma are not equivalent in position to Davison (1895) gave brief descriptions of ventral glands. Female amphiumid vent the gross anatomy of the cloacae of male and glands are not found in the area where “dor- female Amphiuma means and, after finding sal glands” have been described from female sperm in a “viscid substance” exuding from ambystomatid and plethodontid salamanders the vent, concluded that fertilization was in- either. Pleurodeles waltlii and Tylototriton ternal, by cloacal apposition. His other find- uerrucosus of the Salamandridae possess ings were somewhat fanciful, as he described some small, tubular glands in the posterior the cloacal walls as lined with “membranous angle of the vent, although Sever (’91) could laminae” (= rugae?) that connect to a series not ascertain whether these are homologous of “capillary tubes” around the cloacal orifice to vent glands of female amphiumids. that “induce” spermatozoa into the female These divisions (ventral glandslvent cloaca following apposition (Davison, 1895). glands) are derived from the same ancestral In A. tridactylum, Baker (’37) found that sex cloacal gland mass (Sever, ’91). Thus, the easily could be determined by examining the anterior ventral portion of the cloacal gland cloaca. The inside of the female’s cloaca is mass was lost in female amphiumids rather darkly pigmented with soft, somewhat folded than the posterior ventral portion as in the walls, while males possess a white cloaca ancestors to other salamander families in the with a “small rather deep pocket in the dor- Salamandroidea. sal roof. . . .” (Baker, ’37, p. 208). Female vent glands are similar in position Kreeger (’42) described the cloacal histol- and some aspects of cytology to the male vent ogy of female Amphiuma tridactylum. She glands in amphiumids and in other males of found spermathecae around the dorsal and the Salamandroidea. The male vent glands AMPHIUMA CLOACAE 71 show considerable variation among the many Unfortunately, Wilson never published his species in the suborder that have been exam- completed study, and I could not locate his ined, and a role for the glands in mating slides at Tulane University, where Wilson pheromone production has been postulated did the work. I assume that one of the groups (Sever, ’88a). In some female salamanders, of pelvic glands Wilson found are glands I ventral glands have been implicated in mat- recognize as Kingsbury’s glands, and the ing pheromone production (Sever, ’88b). glands he called “abdominal glands,” I refer Since female amphiumids lack ventral glands, to as vent glands. The homologies of Kings- perhaps their vent glands assume the role of bury’s glands and vent glands with glands of pheromone production, which is, as noted the same name in other salamanders are above, the hypothesized function for vent clear (Sever, ’911, but the glands I believe glands in males. The vent glands in female Wilson was referring to as “pit glands” are amphiumids, however, may be vestigial struc- unique in amphiumids. Studies correlating tures with no function in mating. The func- gland secretion with spermatophore struc- tion of vent glands in female amphiumids ture in amphiumids are necessary to see if awaits further study, but their occurrence the pit glands are involved in spermatophore could represent retention of a symplesiomor- formation or if they have another function, phic character for both sexes in a vestigial such as pheromone production. form. The autapomorphic pit glands do not eluci- As far as cloacal conformation and linings date phyletic relations of amphiumids with are concerned, female amphiumids show few other salamander families. The basal posi- specializations. Female amphiumids possess tion of the Amphiumidae in their suborder relatively long cloacal tubes (CTL/ resulting from analysis of cloacal characters TCL > 0.05) and cilia on the cloacal linings, is due to the lack of a definitive posterior both considered ancestral characters by Sever ventral gland cluster like that known from (’91). The extent of epidermis into the cloaca other male salamanders in the Salamandroi- shows two states in female amphiumids. The dea (Sever, ’91). The place that the largest cluster of posterior ventral glands occurs in ancestral condition is that a completely epi- other salamanders is occupied largely by pit dermal lining of the cloacal walls occurs only glands in amphiumas. in the posterior half of the cloacal chamber In their phylogeny incorporating data from (Amphiuma means, A. tridactylum), and in comparisons of ribosomal RNA segments, the derived condition, such a lining extends Larson and Wilson (’89) found that the Pleth- into the anterior half of the cloacal chamber odontidae is the sister group of all other (A.pholeter). Sever (’91) regarded the condi- salamander families and that among the taxa tion in female A. pholeter as an indepen- examined the tree topology is: Amphiuma dently derived character within the Amphiu- (Andrias (Rhyacotriton (Notophthalmus midae. This character also occurs in one genus (Necturus (Ambystoma))))).Since among in the Salamandridae, Euproctus, which has these genera, only Andrias possesses exter- a specialized cloacal tube that again is most nal fertilization (and reduction of cloacal likely independently derived. The derived con- glands to just the anterior ventral gland clus- dition, however, is a synapomorphy for Rhya- ter), this tree implies that internal fertiliza- cotritoninae-Plethodontidae-Proteidae tion evolved at least twice, or a reversal to (Sever, ’91). external fertilization occurred in Andrias. Male amphiumid cloacal anatomy previ- Morphological data do not support such a ously had not been described in detail. Baker dichotomy between plethodontids and other (’37), examining Amphiuma tridactylum, salamanders nor the placement of Andrias noted that males have cloacal glands in “oval, (Sever, ’91). It is interesting, however, that slightly rough patches having a white back- Larson and Wilson (’89) found Amphiuma ground with dotted lines of pigment” which the basal taxon in its clade, since the cloacal are more numerous at the base and along the characters reveal that amphiumids have the sides of the oval patches. Wilson (’41), in an most plesiomorphic cloacal anatomy of any of intriguing abstract four sentences long, stated the Salamandroidea. that he found five types of glands in male A. tridactylum, “anterior pelvic, middle pelvic, ACKNOWLEDGMENTS cloacal, abdominal and pit glands.” He re- I thank P. Moler for the male Amphiuma ported that he had studied both the develop- means and both A. pholeter, and I thank H. ment and seasonal variation of these glands. Dundee for two of the A. tridactylum. The 72 D.M. SEVER female A. means was collected with the aid of Larson, A., and A.C. Wilson. (1989) Patterns of riboso- A. Braswell, D. Forester and S. Perrill. This mal RNA evolution in salamanders. Mol. Biol. Evol. 6t131-154. work was supported by NSF grants BSR Sever, D.M. (1978) Male cloacal glands of Plethodon 8715341 and BSR 9024918. cinereus and Plethodon dorsalis (Amphibia: Plethodon- tidae). Herpetologica 34t1-20. LITERATURE CITED Sever, D.M. (1987) Hemidactylium scutatum and the Baker, L.C. (1937) Mating habits and life history of phylogeny of cloacal anatomy in female salamanders. Amphiuma tridactylum: Cuvier and effect of pituitary Herpetologica 43:105-116. injections. J. Tenn. Acad. Sci. 205-91. Sever, D.M. (1988a) Male Rhyacotriton olympicus (Di- Conant, R. (1975)A Field Guide to Reptiles and Amphib- camptodontidae: Urodela) has a unique cloacal vent ians of Eastern and Central North America. Boston: gland. Herpetologica 44r274-280. Houghton-Mifflin. Sever, D.M. (1988b) The ventral gland in female Eurycea Davison, A. (1895) A contribution to the anatomy and bislineata (Amphibia: Plethodontidae). Copeia 1988: phylogeny ofAmphiuma means (Gardner).J. Morphol. llt375-410. 572-579. Duellman, W.E., and L. Trueb. (1986) Biology ofAmphib- Sever, D.M. (1991) Comparative anatomy and phylogeny ians. New York: McGraw-Hill. of the cloacae of salamanders (Amphibia: Caudata). I. Humason, G.L. (1979) Animal Tissue Techniques. 4th Evolution at the family level. Herpetologica 47t165- Ed. San Francisco: W.H. Freeman. 193. Kreeger, F.R. (1942). The cloaca of the female Amphi- Wilson, F.H. (1941) The cloaca in the male Amphiuma uma tridactylum. Copeia 1942:240-245. tridactylum. Anat. Rec. (Suppl. Abst.) 81:63.