ON THE ROLES OF PHYLOGENY AND STOCHASTICITY

IN THE EVOLUTION OF PERENNIBRANCHIATE TROGLOBITIC

THOMAS R. JONES' Museum of Zoology, University of Michigan, Ann Arbor, MI 48109 USA;

313-936-0134; Bitnet USERGFTV@UMICHUM; Internet [email protected]

and

D. BRUCE THOMPSON Department of Zoology, Arizona State University, Tempe, AZ 85287 USA

602-263-1556

Key Words: , troglobitic, paedomorphosis, adaptation, speciation,

perennibranchiate

Suggested running head: Troglobitic salamander evolution

1 Manuscript corresponding author

1 ABSTRACT

Many troglobitic salamanders exhibit paedomorphic morphologies including perennibranchiation, a morphology that has been viewed as a specific adaptation to cave environments. We present a reexamination of that assumption in a phylogenetic context. We suggest that the perennibranchiate condition in troglobitic salamanders is not an adaptation to cave life that has evolved repeatedly and independently, but rather is a consequence of historical, i.e., phylogenetic, processes. Perennibranchiation appears to be a synapomorphy of the clade of hemidactyliine salamanders exclusive of

Hemidactylium, and as such has not evolved independently in the obligately troglobitic forms. Further, all known proteid salamanders are perennibranchiate, and therefore that morphology cannot be a specific adaptation in Proteus anquinus. We submit that perennibranchiation in troglobitic salamanders is a result of stochastic "entrapment" of epigean species that were polymorphic (or fixed) for a perennibranchiate condition.

Cave-dwelling, perennibranchiate Ambystoma tiqrinum mavortium in south-central

New Mexico are an example of a possible intermediate stage in the evolution of obligately troglobitic salamanders. Finally, if a genetic propensity for a perennibranchiate polymorphism exists within a lineage, ecological isolation in caves might be a sufficient condition for further morphological divergence and speciation within that lineage.

2 Recently, there has been a renewed awareness of the important union of ecological and adaptive inference with phylogenetic systematics. While much of the discussion has been necessarily methodological (Brooks, 1985;

Coddington, 1988; Brooks and McLennan, 1991), few empirical studies have been directed at particular groups of organisms (e.g., Dunham and Miles, 1985;

McLennan et al., 1988; Pearson et al., 1988; Donoghue, 1989; Mooi et al., 1989). This body of work correctly suggests that many commonly held assumptions about adaptation and character evolution should be reexamined in a historical context. Herein we present a case study of the evolution of a perennibranchiate morphology in troglobitic salamanders in such a context. A variety of paedomorphic features may comprise the morphology of troglobitic salamanders (see Wake, 1966), the most notable of which are represented in non-metamorphosing, branchiate adults'. The evolution of perennibranchiate troglobitic salamanders often is discussed implicitly as a deterministic process in which metamorphosing, epigean (surface-dwelling) salamander ancestors enter caves, and descendants subsequently evolve a suite of morphological characteristics, including a perennibranchiate condition, that provide them with a selective advantage in the cave environment. Alternatively, we suggest that a perennibranchiate condition in troglobitic salamanders is a consequence of historical, i.e., phylogenetic, processes rather than an adaptation to cave life, and that the commonness of a perennibranchiate condition in troglobitic salamanders is a result of stochastic "entrapment" of epigean species that are already polymorphic (or fixed) for a perennibranchiate condition. We also provide an example of perennibranchiate, cave-dwelling tiger salamanders (Ambvstoma tiqrinum mavortium), that could represent an intermediate stage in the evolution of obligately troglobitic salamanders. We frame our remarks with respect to

1 In the absence of definitive data describing the developmental evolution of a paedomorphic, perennibranchiate morphology, i.e., neoteny or progenesis (sensu Gould, 1977; Alberch et al., 1979), and to facilitate discussion, we use "perennibranchiate" or "mature branchiate" to refer specifically to sexually mature, permanently gilled salamanders ("perennibranchiation" describes the condition), whether that morphology is considered facultative or obligate.

3 West-Eberhard's (1986, 1989) reviews of the evolutionary significance of alternative phenotypes, and close with a comment regarding general attempts to understand the evolution of perennibranchiate salamanders.

TROGLOBITIC TAXA AND HYPOTHESES EXPLAINING PERENNIBRANCHIATE MORPHOLOGY

There are six genera of true troglobitic salamanders in two families

(Brandon, 1971; see Barr and Holsinger, 1985; Holsinger, 1988 for reviews and definitions) (Table 1.). The tribe Hemidactyliini, family , includes about ten troglobitic species all North American endemics ( of some groups is unclear, and is likely to change in the near future; P.T.

Chippindale and D.M. Hillis, pers. comm.), while the European proteid, Proteus anquinus, is the only form known elsewhere (Brandon, 1971; Sweet, 1984; Barr and Holsinger, 1985). In North America troglobitic salamanders are found in the Valley and Ridge, adjacent Appalachian Plateau, and Dougherty Plain physiographic provinces (in the southeastern USA), the Ozark Mountains of Arkansas, Missouri, Oklahoma and Kansas, and the Edwards Plateau region,

Texas. Perhaps the most obvious feature of troglobitic salamanders is that nearly all species are perennibranchiate; only Gyrinophilus subterraneus and Typhlotriton spelaeus metamorphose.

Hypotheses explaining evolution of troglobitic organisms fall generally into two catagories, in which epigean forms either 1) actively enter subterranean systems, either to escape harsh surface environments or to exploit unoccupied niches, or 2) enter caves passively as a result of physical forces beyond their control (see Culver, 1982). Hypotheses in the former category emphasizing escape have been advanced for most troglobitic salamanders (Wake, 1966; Brandon, 1971; Duellman and Trueb, 1986) and terrestrial invertebrates (summaries in Culver, 1982; Holsinger, 1988;

Holsinger and Culver, 1988). Mitchell and Smith (1972) extended this theme, suggesting that cave colonizers survived inhospitable climates that removed their epigean relatives. Howarth's (1973, 1981) adaptive-shift theory

4 suggests terrestrial invertebrates actively enter new, ecologically "empty" caves (e.g., lava tubes). In the second category, Sweet (1982, 1984) suggested failure of springs following erosion of their water-bearing strata led to underground retreat of epigean, spring-dwelling salamanders along the edge of the Edwards Plateau, Texas. Similar explanations include entry into caves via springs or stream capture, especially in the case of aquatic invertebrates (Holsinger, 1988; Holsinger and Culver, 1988).

A perennibranchiate condition in cave salamanders is framed typically as an adaptive response to cave conditions, although hypothesized selective pressures may vary. For example, Hecht and Edwards (1976:669) said, "Proteus, Typhlomolge, Haideotriton, and other troglobitic salamanders resemble each other in many morphological characters as a result of the severe orthoselective pressures of the cave environment. These resemblances are without question the result of convergent evolution." Here we summarize various hypotheses offered to explain the evolution of perennibranchiates, explicitly in troglobitic salamanders. Wake (1966:82) postulated that within caves, perennibranchiate plethodontids might have had a selective advantage over transforming individuals resulting in eventual fixation of the larval morphology. Dent (1968:303) suggested that selective pressures such as food abundance (in aquatic vs. terrestrial cave environments) likely "brought about the rise of neoteny [sic] in caves." Brandon (1971) also suggested a perennibranchiate morphology was a likely evolutionary response to food requirements and availability, in which morphological features of perennibranchs (e.g., neuromasts, spatulate snouts) allow more efficient feeding in permanently dark, aquatic habitats; this idea also has been emphasized by Culver (1982). In addition, perennibranchs avoid energy expenditures necessary for metamorphosis, which might be important in caves where food is less abundant relative to surface habitats (Brandon, 1971). According to Wilbur and Collins (1973), within caves a selective disadvantage may exist for metamorphosing salamanders that leave stable and relatively productive aquatic habitats to enter unproductive terrestrial environments.

5 Bruce (1976) provided life history data on Eurycea neotenes (as currently recognized, an epigean and troglobitic species) suggesting perennibranchiation evolved through lowering size and age at maturity in response to high juvenile mortality and environmentally determined minimum metamorphic size. Although Bruce (1976) did not implicate cave conditions, Sweet (1977:374) argued against Bruce's suggestions, saying perennibranchiate E. neotenes apparently evolved as a response to "selective disadvantages of metamorphosis" in subsurface aquatic habitats where branchiate could feed more effectively. Bruce (1979:1000) later modified and extended Brandon's (1971) ideas regarding perennibranchiate Gyrinophilus palleucus, suggesting the prey resource base was the "relevant environmental variable" influencing shifts in life histories. Specifically, in its epigean close relative G. porphyriticus, metamorphosis occurs at a body size nearly optimal for switching from invertebrate to salamander prey, an option generally unavailable to cave- dwelling G. palleucus for which salamander prey are typically scarce.

Consequently, larval morphology and utilization of invertebrate prey in caves might have been prolonged (e.g., G. subterraneus) or fixed (e.g., G. palleucus) (Bruce, 1979). A common theme in these discussions is that a perennibranchiate morphology is a deterministic outcome of cave colonization, representing an adaptation to a particular set of ecological circumstances unique to subterranean environments. Bruce's (1976) hypothesis is an exception, although he also postulated an independent origin of the perennibranchiate condition in E. neotenes (see below). This idea has become accepted generally in the speleological and herpetological literatures

(Porter, 1972:371,373; Culver, 1982:34-35; Jackson, 1982:131; Duellman and Trueb, 1986:192-194). A notable exception to this general assumption comes from Mitchell and Smith (1972), who believed perennibranchiation evolved in the ancestors of Typhlomolqe and Texas Eurycea prior to cave colonization. They suggested a perennibranchiate condition was likely an important, if not necessary, "'preadaptation' for a potential cave colonizing urodele," and thus, surmised that epigean, ancestral Eurycea/Typhlomolqe occupied central

6 Texas for enough time to allow "sufficient time to permit the evolution of neoteny [sic] prior to the time of cave accessibility" (Mitchell and Smith, 1972:358). Although we concur with the basic idea that the evolution of perennibranchiation likely precedes cave colonization, we do so for different reasons.

SALAMANDERS IN CAVES

Caves in temperate regions are typically stable, cool and humid (Culver,

1982), and commonly enter caves wherever they are accessible. Although not troglobites, metamorphosed adult plethodontids commonly occupy caves, often well beyond the twilight zone, for refuge, feeding, or reproduction. Some species are known primarily from caves, or are quite common cave inhabitants, frequently breeding deep within. Examples include

Chiropterotrition magnipes, C. mosaueri, Eurycea lucifuqa and Gyrinophilus porphyriticus in the New World (e.g., Brandon and Rutherford, 1967; Cooper and Cooper, 1968; Holsinger and Culver, 1988; Martin, 1958; Peck and Richardson,

1976), and Hydromantes spp. in Europe (Thorn, 1968; Steward, 1969). Additional facultative cave dwellers that may be more irregular in their use of caves (some neotropical forms are poorly known) include Chiropterotrition multidentatus, Desmoqnathus monticola, Eurycea bislineata, E. longicauda,

Hydromantes shastae, Nyctanolis pernix, Plethodon albaqula, P. caddoensis, P. dorsalis, P. glutinosis, P. petraeus, P. richmondi, P. serratus, P. wehrlei,

Pseudoeurycea bellii, and P. scandens (Brandon and Rutherford, 1967; Conant, 1975; Cooper, 1961; Elias and Wake, 1983; Fowler, 1951; Gorman and Camp, 1953; Martin, 1958; Myers, 1958; Reddell and Mitchell, 1971; Stebbins, 1986; Wynn et al., 1988; P. Chippindale, pers. comm.; S.E. Trauth, pers. comm.; pers. obs.).

Among metamorphosing Ambystoma, A. jeffersonianum has been found in large numbers in West Virginia caves (Cooper, 1961), A. texanum have been collected in Texas caves (P. Chippindale, pers. comm.), and A. tiqrinum are often common in caves. For example, metamorphosed adult and subadult A. t. nebulosum occur

7 in about 90% of ca. 50 caves surveyed along the Mogollon Plateau in Arizona

(Thompson, unpubl.), while A. t. mavortium have been found in Oklahoma caves (Black, 1969; Bozeman, 1987). Although reproduction has been reported in

Oklahoma caves (Black, 1969), perennibranchs were not. Metamorphosed A. t. mavortium also have been recorded in over 40 caves in south-central New Mexico (D. Belski, pers. comm.).

As we reviewed above, the evolution of perennibranchiation in troglobitic salamanders is usually considered an adaptation to the unique cave environment. Also, both the colonization of caves and the evolution of perennibranchiates are linked frequently to escape from adverse terrestrial or surface conditions (e.g., Brandon, 1971; Sprules, 1974). Although an escape hypothesis at first might appear logical considering the climatic history of the temperate world, there is no need to postulate escape when many amphibians readily enter caves for a variety of reasons (see above). In addition, as Barr and Holsinger (1985) noted for evolution of perennibranchiate troglobitic salamanders, conditions in the relatively mesic eastern USA indicate the need for a more satisfactory explanation especially in light of the species-rich epigean eastern salamander fauna.

Although we will not argue with the idea that selective pressures within caves are unique and likely influence morphologies among a wide variety of organisms, we suggest that adaptive explanations for the evolution of the perennibranchiate condition in troglobitic salamanders are unnecessary. We present evidence below that phylogenetic constraints within the lineages of troglobitic salamanders and their close relatives dictated the morphologies available to potential cave colonists.

DISCUSSION

Two significant pieces of evidence argue against a selectionist hypothesis for the repeated and independent evolution of perennibranchiate, troglobitic salamanders. First, we will discuss the phylogenetic distribution

8 of perennibranchiates. Second, we will discuss the singular exception to the adaptationist prediction.

Phylogenetic Distribution of Troglobitic and Perennibranchiate Salamanders

To analyze evolutionary mechanisms, such as a repeated, developmental shift to perennibranchiation, requires an analysis of evolutionary patterns

(Fink, 1982), and a "phylogenetic context, however crude, is a fundamental prerequisite for a hypothesis of heterochrony since any such hypothesis is a comparative statement (Fink, 1982:285). In salamanders, perennibranchiation is phylogenetically widespread, occurring in some taxa within almost all families. While some families are completely perennibranchiate, or exhibit only partial metamorphosis, others include both obligate and facultative perennibranchs (summaries in Duellman and Trueb, 1986; Larson, 1991). It is important to note that generalized morphologies commonly referred to as

"paedomorphic" (i.e., perennibranchiate) include many specific features that are not phylogenetic homologues (Larson, 1991). Disregarding salamander families in which complete metamorphosis is unknown, perennibranchiates are most widely distributed within the Ambystomatidae and Plethodontidae, both of which include facultative and obligate perennibranchs (e.g., see Shaffer, 1984; Duellman and Trueb, 1986; Larson, 1991), although a biphasic life history (i.e., aquatic larva and metamorphosed form) is considered the

"typical" condition. We will review two groups of perennibranchiate salamanders: Proteidae and Plethodontidae, because they contain all known troglobitic salamanders. We follow the hypotheses of salamander phylogenetic relationships postulated by Larson (1991; see also Larson [1984] and references therein), but also recognize that many questions remain regarding salamander systematics.

Proteidae includes two extant genera, Proteus and Necturus. Metamorphosis does not occur in this family; all extant and extinct representatives are paedomorphs, presumably perennibranchiate (Estes, 1981;

9 Duellman and Trueb, 1986). All Necturus are epigean (North America) and the

European troglobite, Proteus, is the only other extant representative. The family Plethodontidae contains two subfamilies, Desmognathinae and

Plethodontinae (see Duellman and Trueb, 1986) (Fig. 1). Most desmognathines have a biphasic life history, with free-living aquatic larvae that undergo metamorphosis (some species have lost the free-living larval stage); perennibranchiates are absent. Plethodontinae contains three tribes:

Hemidactyliini, Plethodontini and Bolitoglossini. The latter two exhibit direct development, i.e., no free-living larvae. The primitive hemidactyliine morphology and biphasic life history is represented in Hemidactylium. The remaining hemidactyliine genera comprise a separate clade, sharing a uniquely derived set of hyobranchial characters (Lombard and Wake, 1976; 1977), and contain several facultative and obligate perennibranchs, including the troglobitic species (Table 1., Fig. 1). In addition to the highly specialized troglobitic hemidactyliine plethodontids, perennibranchiation and "trends" toward that morphology (i.e., extended larval periods or precocial gonadal maturation) occur elsewhere within the genera Eurvcea and Gyrinophilus (Table 1.). Two epigean species, E. tynerensis and E. multiplicata are obligately and facultatively perennibranchiate, respectively (Dundee, 1965a; 1965b; S.E. Trauth pers. comm.); perennibranchiate E. multiplicata can also reside in caves (Dundee,

1965a). A single epigean perennibranchiate G. porphyriticus has been reported from a spring-fed pond in a non-karst region of northern Georgia (Martof and

Humphries, 1955; R.L. Humphries, pers. comm. [To verify sexual maturity, we examined the testes histologically; mature spermatozoa were abundant within seminiferous tubules.]). Elsewhere, cave-dwelling perennibranchiate G. porphyriticus have also been reported (Holsinger and Culver, 1988).

Additionally, Besharse and Holsinger (1977) reported a "larval" G. subterraneus with sperm producing testes. Bruce (1979) suggested that perennibranchiates may have evolved through phyletic extension of the larval period (neoteny). Data for just such an extension are available for

10 Gyrinophilus (Brandon, 1971; Bruce 1978; 1979; 1980; Holsinger and Culver,

1988), and some populations of E. bislineata. In the latter (= E. aquatica

[Rose and Bush, 1963; Conant and Collins, 1991]) gonadal maturation may occur just prior to metamorphosis in third or fourth year larvae (Rose and Bush,

1963; Jones, unpubl.); see also Bruce (1982a,b; 1985; 1988) for comparative data.

Perennibranchiation is not randomly distributed among plethodontids, instead perennibranchiate lineages share a distinct phylogenetic history, i.e., they belong to a monophyletic assemblage of hemidactyliine genera. The hypothesis of repeated, independent evolution of the perennibranchiate condition in this group (e.g., Larson, 1984) is unwarranted. (We acknowledge that the absolute test of this hypothesis requires a more robust cladistic analysis of the Hemidactyliini than is available.) Within Plethodontidae, the character "perennibranchiation" is restricted to particular clades, and absent from other lineages (Fig. 1), irrespective of environmental cues that may (or may not) be required to initiate the ontogenetic shift towards expression of a perennibranchiate morphology in an individual or a lineage. The restricted phylogenetic distribution of perennibranchiates within these taxa argues strongly for a significant genetic component in the evolution of perennibranchiation in troglobitic salamanders (clearly in Proteidae no other morphology is known). The historical and therefore genetic basis of perennibranchiation is central to our discussion of perennibranchiate troglobitic salamanders.

Finally, when adaptive explanations are viewed in a phylogenetic context the exceptions to adaptive predictions are among "the strongest 'falsifier[s]' of adaptation scenarios" (Brooks and McLennan, 1991:84). Typhlotriton spelaeus provides such an exception (Fig. 3). Typhlotriton spelaeus is a highly specialized and obligately troglobitic salamander, it has a typical biphasic life history, and is widespread and relatively abundant (see Brandon,

1971 regarding erroneous reports of mature branchiate populations). Thus, T. spelaeus falsifies hypotheses predicting cave salamanders should be

11 perennibranchiate because terrestrial forms would be selected against, and clearly the failure to metamorphose is not a prerequisite for successful utilization of the subterranean niche (see also, Smith, 1948). In addition, another obligate troglobite, G. subterraneus, metamorphoses (Besharse and

Holsinger, 1977), although larval life may be relatively long and at least one perennibranchiate individual has been reported (see above). Thus, perhaps a more interesting question than why are most cave salamanders perennibranchiate, is why are there so few metamorphosing troglobites?

An Alternative Hypothesis and an Empirical Example

The phylogenetic data clearly suggest a common historical distribution of perennibranchiation in epigean and troglobitic plethodontids. Similarly, all proteids are perennibranchiate. Thus, the most parsimonious explanation for similarity of form within each of those families is their respective histories, i.e., phylogenetic constraint. Consequently, we reject the idea that evolution of perennibranchiate troglobitic salamanders is the result of deterministic processes, e.g., escape from adverse surface conditions and subsequent selection to maintain larval (= aquatic) traits. Rather, we suggest that perennibranchiate troglobites evolved through a series of stochastic events in which epigean forms polymorphic (or fixed) for the expression of the perennibranchiate condition entered caves, perhaps for no other reason than convenient shelter, and opportunistically bred therein.

Those immigrant breeders produced a percentage of offspring that failed to metamorphose and were therefore confined to the aquatic cave habitat. Under favorable conditions new cave populations persisted such that natural selection could later mold larval features already present in perennibranchiate adults, to produce the extreme morphologies present in some species. Below we present an empirical example illustrating the conditions sufficient for establishing a perennibranchiate, cave-dwelling salamander population.

12 Elsewhere we reported the occurrence of small, breeding populations of perennibranchiate Ambystoma tigrinum mavortium in two caves (Antelope Gyp and Flat Rock) in south-central New Mexico (Thompson and Jones, 1992; see also

Peerman, 1992), in an area of abundant gypsum caves referred to as "GYPKAP"

(Peerman and Belski, 1991). Despite the common association of A. tigrinum and caves elsewhere, these are the only reported cases of cave-dwelling perennibranchs. The cave pools in which salamanders were found are isolated, having no direct connection with surface waters. There is no evidence that A. tigrinum breeds in any other caves in that area, although metamorphosed adults are common in virtually all nearby caves (Peerman and Belski, 1991; D. Belski, pers. comm.). A. t. mavortium is widespread throughout much of New Mexico, south and east of the Rocky Mountain axis (Stebbins, 1986; Jones and Collins, 1992). In southern New Mexico mature branchiates are common in most permanent habitats (Jones and Collins, 1992; Jones, unpublished), and the cave branchiates did not appear to differ morphologically from others we have observed. In this regard, and phylogenetically, A. tigrinum in Antelope Gyp and Flat Rock caves provide an example of the set of circumstances we have outlined above.

Within the genus Ambystoma perennibranchiates occur in two lineages

(reviewed in Sprules, 1974a): A. talpoideum and A. gracile, and the A. tigrinum/A. mexicanum species group (Fig. 2). The first two species are facultatively perennibranchiate (Sprules, 1974b; Semlitsch, 1985) and the third lineage comprises nine or more facultatively and obligately perennibranchiate species (Shaffer, 1984). According to Kraus (1988) and

Shaffer et al. (1991) the A. tigrinum/A. mexicanum species group is likely monophyletic. Dicamptodon, considered the sister group of Ambystoma (see Larson, 1991; Shaffer et al., 1991), includes species polymorphic for metamorphosis and mature branchiates, and an obligately perennibranchiate species (Nussbaum, 1976). Thus, when perennibranchiation is mapped on a cladogram of ambystomatids we see that it is the ancestral condition, and in Ambystoma is restricted to two clades (Fig. 2). As in Plethodontidae there is

13 a common historical basis for the distribution of perennibranchiates, and it is unnecessary to postulate the multiple and independent evolution of this trait (contra Shaffer, 1984).

The mature branchiate morphology is a facultative trait in Ambystoma tiqrinum, and in the western USA is common in several taxa: A. t. mavortium

(Rose and Armentrout, 1976), A. t. diaboli (Gehlbach, 1966), A. t. nebulosum (Sexton and Bizer, 1971; Collins, 1981), and A. t. stebbinsi (Collins et al., 1988; Jones et al., 1988); additionally, some Mexican populations of A. tiqrinum cf. velasci are facultatively or obligately perennibranchiate

(Shaffer, 1984). Although perennibranchiate A. tiqrinum often can be induced to metamorphose in the laboratory (reviewed in Dent, 1968), the metamorphs are often debilitated and may die. Some authors have suggested that some perennibranchiate salamanders might be less able to metamorphose beyond an undefined age threshhold (Lipchina, 1929 [cited in Dent, 1968]; Snyder, 1956), although data are not conclusive. In A. t. stebbinsi, where mature branchiates comprise about 90% of the adult population (Collins et al., 1988), death usually follows the onset of metamorphosis in the laboratory (Jones et al., 1988). Thus, in Ambystoma once an becomes perennibranchiate it might be difficult (although certainly not impossible) to metamorphose later.

We will discuss the significance of this below.

Considering this example, how might a perennibranchiate condition and a troglobitic existence become coupled? If individual metamorphosed tiger salamanders reproduced in suitable cave habitat, the net effect of any metamorphic constraints on their offspring (i.e., a genetic predisposition, irrespective of environmental cues) would be to confine those individuals to the cave. Assuming the perennibranchiate morphology has a heritable genetic component (which is likely, given its phyletic distribution), increasing numbers of offspring in successive generations would also be perennibranchiate. Metamorphosing offspring could easily exit the cave and thus contribute little or no genetic input to subsequent generations, i.e., they would likely breed in surface habitats that are relatively common in the

14 immediate area (pers. obs.). As Mitchell and Reddell (1965:24-25) pointed out, the extent to which the gene pool of the evolving troglobite is diluted by epigeal invaders.. .would affect greatly the length of time required for a high degree of adaptation." The same could be said to affect instead the time to fixation of a perennibranchiate condition in an otherwise polymorphic population. A number of geomorphic processes could later act to isolate the new cave occupants such as stream capture or erosion of cave openings beyond the aquifer bearing sediments, as suggested by Sweet (1982, 1984) and Holsinger (1988).

Our hypothesis can explain mature branchiate morphologies in any troglobitic salamander. In particular, consider Proteus anquinus. Obviously since all known fossil and extant proteids are perennibranchiate (Estes, 1981), a perennibranchiate morphology in P. anquinus cannot be considered an adaptation to a subterranean habitat. Ancestral, epigean Proteus might have been washed into caves or may have entered through captured surface waters, and later became isolated. Thus, its history can incorporate aspects of our hypothesis (i.e., inability to escape the aquatic cave habitat) with others regarding capture of epigean aquatic systems (Sweet, 1982; 1984; Holsinger,

1988; Holsinger and Culver, 1988).

THE EVOLUTION OF PERENNIBRANCHIATE MORPHOLOGY

It is beyond the scope of this paper to address adequately the complex issues regarding the evolution of mature branchiates. In order to understand the evolution of a character, or suite of characters, such as this we have to identify homologies and homoplasies among perennibranchiate morphologies (i.e., are all perennibranchs a result of the same set of developmental processes?--probably not; see Larson, 1991), establish robust phylogenies of groups in which mature branchiates occur, and dissect historical patterns from ecological/evolutionary patterns (e.g., Fink, 1982). Treating perennibranchiation as an adaptive response to cave-dwelling (or any other set

15 of conditions) confuses the important distinction between conditions that maintain a perennibranchiate condition with those leading to its origin (see Brooks and McLennan, 1991). We suggest in the case of the New Mexico A. tiqrinum and for troglobitic salamanders in general, that a genetic propensity for a perennibranchiate morphology already existed (i.e., phylogenetic constraint) and simply provided a means by which salamanders initially became ecologically isolated in caves, setting the stage for further morphological divergence and speciation (sensu West-Eberhard, 1986; 1989).

Intraspecific polymorphic morphologies, or life history patterns, operating at an ecological level, could be precursors to speciation (Gould,

1977; Stanley, 1979; West-Eberhard, 1986; 1989). It is this type of variation, existing in a unique evolutionary setting, that could have led to the evolution of many troglobitic salamanders. We suggest that perennibranchiate A. tiqrinum in Antelope Gyp and Flat Rock caves provide evidence for the type of evolutionary opportunity for speciation that might have produced most troglobitic salamanders; i.e. a pre-existing polymorphism

(perennibranchiation) becomes fixed in cave-dwelling populations because of stochastic events and subsequent ecological segregation. We do not necessarily suggest that these particular A. tiqrinum populations are speciating, nor do we know if they could persist long enough to become truly troglobitic and genetically isolated (the geomorphology of gypsum caves, i.e., their high solubility and low mechanical strength, suggests they are transient and unlikely to persist for long periods of geological time [White, 1988]). Nonetheless, the circumstances we have observed could be instructive evolutionarily, suggesting an alternative approach to studies of the evolution of troglobitic salamanders and of perennibranchiate morphologies in general. Perhaps renewed efforts to search for genetic and developmental bases of perennibranchiation (e.g., Jones, 1989; Routman, 1990; Harris et al., 1991), in a strong phylogenetic context, combined with ecological tests of the hypotheses will provide answers.

16 ACKNOWLEDGEMENTS

We are especially grateful to D. Belski for bringing the Ambystoma populations to our attention, and to the GYPKAP survey teams and members of the Pecos Valley Grotto of the National Speleological Society for general information and hospitality in the field. We appreciate the field assistance of M. Safford and E. Earle. R.L. Humphries kindly supplied us with the perennibranchiate G. porphyriticus from his personal collection (RLH A739; now UMMZ 194070) and pertinent field data. We thank S.E. Trauth who performed the histological work on extremely short notice. We appreciate discussions and correspondence with P.T. Chippindale, A. Kluge, S. Meagher, D.L. Pearson, C.W.

Seyle, S.E. Trauth, D.B. Wake, and K.E. Zerba. TRJ benefitted from additional discussions with J.P. Collins, P.J. Fernandez, J.R. Holomuzki, K.E. Zerba regarding the evolution of paedomorphosis. P.T. Chippindale, F. Kraus, R.

Olson, M.E. Pfrender, D.B. Wake and K.E. Zerba reviewed earlier drafts.

Bureau of Land Management, Roswell Resource Area provided permits to enter the cave and remove biological samples. Locality data for the NM caves are available from the authors, the National Speleological Society, Huntsville,

Alabama, USA, or U.S. Bureau of Land Management, Roswell New Mexico Field

Office.

17 LITERATURE CITED

ALBERCH, P., S.J. GOULD, G.F. OSTER, AND D.B. WAKE. 1979. Size and shape in ontogeny and phylogeny. Paleobiology 5:296-317.

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25 Life History Perermibranchiate taxa

Taxa' metamorphose facultative obligate epigean troglobitic perennibranchiate perennibranchiate

Gyrinophilus palleucus - +2 - +

G. porphyriticus + +I- - ± -

G. subterraneus -I- - - N/A N/A

Haideotriton (1) - - -I- . ±

Hemidactylium (1) + - - N/A N/A

Pseudotriton (2) + - - N/A N/A

Stereochilus (1) + - - N/A N/A

Typhlomolge (2) - - + - -4-

Typhlotriton (1) + - N/A N/A

Eurycea bislineatd + - - N/A N/A

E. longicaida ± - N/A N/A

E. lucifuga + - N/A N/A

.5 E. multiplicata + + - ±

E. nana - - ± ±

4 E. neotenes - - +2 + +

E. quadridigitata -I- - - N/A N/A

E. tridentOra ± - +

5 E. tynerensis - - -I- +

Proteus (1) - - -I- - +

Species are listed separately for genera in which life history traits vary. Number of species in parentheses. 2 Occasional naturally occurring metamorphosed individuals have been reported (Simmons, 1976; Sweet, 1977; Yeatman and Miller, 1985; Brandon et al, 1986). 3 One naturally occurring peretunbranch has been reported (see text). 4 Taxon may comprise multiple cryptic or incompletely delineated species (e.g., Jacobs, 1987; P.T. Chippendale, unpubl.) 5 Cave dwelling populations are known, but are they are not true troglobites. Table Legend

TABLE 1. DISTRIBUTION OF LIFE HISTORY CHARACTERISTICS AMONG HEMIDACTYLIINE AND KNOWN TROGLOBITIC SALAMANDERS. PAEDOMORPHIC TAXA ARE LISTED ACCORDING TO

WHETHER THEY INCLUDE EPIGEAN OR TROGLOBITIC POPULATIONS. INDIVIDUAL SPECIES

ARE NOT LISTED FOR THOSE GENERA IN WHICH ALL SPECIES EXHIBIT THE SAME LIFE HISTORY CHARACTERISTICS; OTHER SPECIES ARE LISTED SEPARATELY. A PLUS (+)

DENOTES THE TRAIT IS EXHIBITED, A MINUS (-) INDICATES IT IS NOT.

Figure Legends

FIG. 1.--Phylogenetic relationships among salamanders in the family Plethodontidae (modified from Lombard and Wake, 1986; Larson, 1984; 1991). P at arrow = the evolutionary appearance of paedomorphosis. See Table 1 for distribution of life history characteristics.

FIG. 2.--Phylogenetic relationships among salamanders of the genus Ambystoma (modified from Kraus, 1988; Shaffer et al., 1991; and Jones et al., In press); sister group is Dicamptodon. The perennibranchiate condition is indicated by bold type, and in Ambystoma is primitive with respect to the outgroup. An open circle indicates loss of the perennibranchiate condition in a clade.

FIG. 3.--Hypothetical cladogram in which pattern of distribution of paedomorphosis (X1) does not match movement into a new environment (i.e., caves), thus falsifying the hypothesis of a functional fit between X1 and the environment (modified from Brooks and McLennan, 1991:84-85). Ancestral state,

X0 = metamorphosis.

27 Desmognathinae

Hemidactyliini Hemidactylium

Eurycea

Gyrinophilus

Pseudotriton Haideotriton

Stereochilus Typhlomolge Typhlotriton

Bolitoglossini

Plethodontini annulatum

cingulatum

barbouri

texanum (W.)

texanum (E.) mabeei jeffersonianurn

opacum laterale

macrodactylum "Mexican radiation" tigrinurn gracile maculatum talpoideum

Outgroup \c c Aezc` _,Ao e21‘ sci\k ' Ao`ti\t‘ e2S\ twv ' g ' , (ov 'e ksov ec\q ec\v es` c 6` e9\ge2''‘