AN INTEGRATED APPROACH TO EVOLUTIONARYSTUDIES OF

DAVIDB. WAKE*

CONTENTS der Caudata. Studying organisms from one Introduction ...... 163 perspective can inform and direct investiga- Phylogenetics ...... 163 tions that have other goals, and an integra- Hierarchical Approaches ...... 166 tive approach to organismal evolution can Ontogeny and Phylogeny ...... 168 result. As evolutionary biologists, our ultimate Recognition of Species ...... 170 goal is to understand how biological diversifi- Species Diversity ...... 172 cation occurs, and as herpetologists our pri- A Plea for Conservation ...... 174 mary focus is the whole organism, as exempli- Acknowledgments ...... 175 fied by and reptiles. The examples Literature Cited ...... 175 I use are mainly from my own laboratory, be- cause I can see most clearly in these cases how Abstract: This essay is an examination of the ways integration can be achieved. Although this in which seemingly disparate studies focused on a paper is topical in nature, I attempt to provide single, rather small taxon, the Order Caudata, can connections between topics. provide a foundation for development of programs in evolutionary biology. Topics addressed include Any evolutionary study focused at the the use of individual characters as contrasted with complexly integrated systems of characters in phy- organismal level requires a framework of logenetic analysis, the nature of integration and its phylogenetics, with clearly stated hypotheses impact on specializations and adaptation, and on of relationships underlying all data analyses. the conceptualization and recognition of species. The At the same time, however, one must frankly possibility that salamanders and other amphibians acknowledge that the Caudata have been and might play a role as bio-indicators in environmental remain a controversial group; we have yet to studies is raised. achieve a robust phylogenetic hypothesis for the group as a whole, or even for the genera INTRODUCTION within the larger families. Thus, I envision phylogenetics as an on-going process with re- Y GOAL IS TO SHOW HOW STUDIES BROADLY ciprocal illumination occurring as we learn M focused on the organismal level can be more of the nature of characters, character used to develop a framework for an under- complexes, and novel data. standing of the evolutionary biology of a ma- jor taxon-in this case, the salamanders, Or- PHYLOGENETICS

is * Department of Integrative Biology and Museum of There growing acceptance that the living Vertebrate Zoology, University of California, Berke- amphibians form a monophyletic group, the ley, California 94720, USA. Lissamphibia, and that the Order Caudata

163 164 DAVID B. WAKE also is a monophyletic group (Trueb and characters used by Duellman and Trueb Cloutier 1991; Duellman and Trueb 1985). I (1985), and when these recoded characters will not deal with these issues further. were placed on the trees of Duellman and Trueb (based mainly on morphology) and of Progress has been made in understanding Larson (based on DNA sequences), the fit to phylogenetic relationships at high taxonomic the former was somewhat more parsimonious levels, but one of the most vexing problems than to the latter; Larson argues that the facing a biologist is the difficul- difference is not statistically significant. ties encountered in generating even a rela- Therefore,Larson’s hypothesis is equallyvalid. tively robust phylogenetic hypothesis concern- Hillis (1991) has attempted to combine some ing the relationships of salamander families. of the morphological data and sequence data The hypothesis of Duellman and Trueb (1985) available at the time he wrote his paper, but serves as a convenient point of departure, for as yet the result is not robust and has low it displays the traditional arrangement of a internal consistency. basal position for the Sirenidae, a mono- phyletic group including Hynobiidae and A major problem is the reconciliation of Cryptobranchidae that is also relatively basal, these different data sets. Larson (1991) has and a deeply nested position for the Pletho- made an initial effort along these lines and his dontidae. Thus, the taxa with external fertili- results demonstrate anew that any hypoth- zation (Hynobiidae and Cryptobranchidae, esis of relationships of the salamander fami- and presumably Sirenidae) are separated from lies requires extensive homoplasy for nucleo- a monophyletic group that practices internal tide, morphological, and other kinds of fertilization. characters. We need a fresh look at the nature of mor- New studies of morphological characters phological and other characters and their and of DNA sequences challenge this orthodoxy in several important respects. Cloutier (in interrelationship, and new characters. We prep.), usingmorphologicaltraits, not only fails must give attention to the ways in which char- to find a monophyletic Hynobiidae + Crypto- acters are generated in development and the branchidae, but also believes that the family degree to which characters are developmen- Hynobiidae is paraphyletic. He does agree with tally and functionally linked. There is so much earlier authors in recognizing the Pletho- homoplasy in the morphological characters of salamanders that cladistic hypotheses that use dontidae as being deeply nested. In contrast, Larson (1991) has studied the nucleotide states that cannot be directly observedin taxa are suspect. I especially distrust characters sequences of ribosomal RNA and concludes that the Hynobiidae + Cryptobranchidae is that, on the basis of a parsimony analysis monophyletic but not basal, and that the alone, are considered to be lost and thus used , Amphiumidae, and Rhyaco- as a synapomorphy low in a tree-yet reap- tritonidae (a family recognized on morpho- pear (attributed to evolutionary reversal) logical grounds by Good and Wake [in press]) higher in the tree. In general, I mistrust char- are relatively basal. According to Larson, the acters whose state must be inferred rather remaining families form a monophyletic group than directly observed. and the Dicamptodontidae and Ambystomat- We lack critical information for various taxa. idae are deeply nested sister taxa; this For example, it is amazing that we still do not challenges the findings of Edwards (1976). know whether sirenids have external or inter- Larson has re-evaluated the morphological nal fertilization. On the one hand, the absence EVOLUTIONARY STUDIES OF SALAMANDERS 165 of cloacal glands suggests that no spermato- treated as four primary taxa. (Presch [19891 phore is produced and that sperm storage does has questioned this procedure, which I defend not occur. On the other, the fact that eggs are on the grounds ofthe demonstrable homoplasy laid individually suggests that internal fer- in the group; I reject Presch?s reanalysis ofour tilization might occur. data, which involves recoding of data and use of groups of genera as primary taxa.) By ac- My focus has been the Family Pletho- cepting these four primary taxa, we are ac- dontidae, which includes over 60%of the ap- cepting that projectile tongues evolved inde- proximately 390 living species (Frost [19851 pendently at least three times in the family, and subsequent descriptions). Homoplasy is and because we think that attached projectile extensive in the family (Wake 1966, and in tongues are functionally and phylogenetically press; Wake and Larson 1987) and has led to independent of free projectile tongues, projec- problems in developing a robust phylogenetic tile tongues of some sort must have evolved at hypothesis. Since the publication of my com- least five times, assuming that our four pri- parativeosteological study ofthe family(Wake mary taxa are monophyletic. We examined 1966), however, there has been relative taxo- the 15 dichotomous trees that could be gener- nomic stability. The monophyletic nature of ated for four groups, and found that two are the family is unquestioned, as is the mono- more parsimonious than the alternatives. Our phyletic status of the two major sister taxa, choice requires that the free projectile tongue the subfamilies Desmognathinae (3 genera evolved three times and the attached projec- and 14 species) and Plethodontinae (15 gen- tile tongue evolved three times in the family. era and about 235 species). The species of the It will be difficult with any rearrangement of Plethodontinae (still about 60% of living taxa to reduce this more than by one; Lombard species!) were placed in three tribes: Pletho- and Wake (1986) treated Batrachoseps and dontini, Hemidactyliini (Dubois [ 19841 has the supergenus Bolitoglossa as sister taxa, argued that the correct name should be but if the sister taxon of Bolitoglossa is Hydro- Mycetoglossini), and Bolitoglossini. Of these, mantes the number of evolutions of free the Bolitoglossini is an unquestioned mono- tongues could be reduced by one. phyletic group and the Plethodontini is very likely a monophyletic group, but evidence for At present we have too little information to the monophyly of the third tribe is weaker. choose among the various alternatives, but data have been building that support certain Due to the extensive homoplasy and the relationships. For example, details of the or- relative dearth ofcharacters that can be scored ganization of the nervous system (such as the at the level of the tribes, I have argued that complete loss of the lateral line system, which one might treat the feeding system (which is not a necessary condition associated with involves many characters that could be sepa- direct development [Wake et al. 19871) sup- rately coded) as an integrated and somewhat port the concept ofthe monophyly of the Pletho- self-organized system that could be interpreted dontini + Bolitoglossini. as one large, complex character. I have exam- ined the implications of analysis of that sys- I would like to see the development of a tem for understanding the phylogeny of the comprehensive data matrix (including DNA family (Roth and Wake 1985). Nevertheless, sequence data) at the level of genera or spe- Lombard and I(1986)also attempted acladis- cies groups for the family, but to accomplish tic analysis usingall data then available to us. this will require sufficient insight into char- The two subfamilies and three tribes were acter ontogeny and evolution to deal, at least 166 DAVID B. WAKE

in part, with the extensive homoplasy in the triton bromeliacia, richardi, and family. When such is available, it will doubt- all species of ). lessly continue to show substantial homoplasy, but by looking at different subsets of the ma- Ecologists generally accept that body size is trix perhaps we will come to understand the single most important variable in an indi- whether the integrated complex approach I vidual from the perspective of population or have espoused is useful in phylogenetic analy- community ecology. Thus, one expects that sis. I look forward to data sets based on nu- the evolutionary dynamics operative at the cleotide sequences, for they will be helpful in population and community levels of organiza- sorting out the morphological homoplasy. tion must have animportant role in determin- Eventually we will have to arrive at an under- ing size at first reproduction, maximum body standing of relationships based on all avail- size, and the minimum size at which indepen- able data, but how that will be accomplished dence can occur. Furthermore, many investi- is as yet unclear. gators have shown that the size of a female is strongly related to the numbers of eggs pro- duced, with each species having a relatively HIERARCHICAL APPROACHES narrow and predictable relationship between eggsize andeggnumber(Houck 1977;Kaplan There is increasing interest in hierarchical and Salthe 1979). approaches in evolutionary biology (for ex- ample, Eldredge 1985; Vrba 1989).Much dis- Genome size in salamanders is huge (Ses- cussion about hierarchies has focused on the sions and Kezer 1991; Sessions and Larsen question of whether or not selection occurs at 1987; Morescalchi 1990; Olmo 1983) in rela- the level of species, but my interest is in the tion to other amphibians and especially in interactions taking place between levels in the relation to other vertebrates. In fact, the larg- molecular-organismal-populationalheir- est genome size in salamanders is exceeded archy. I can best illustrate this by focusing on among vertebrates only by the lungfishes the phenomenon of miniaturization in sala- (Morescalchi 1990).An empirical relationship manders, for a great deal of useful informa- exists between genome size (amount of DNA tion has accumulated in recent years concern- per haploid nucleus) and cell size. Some ing miniaturization and its effects (for recent researchers believe that genome size increases reviews see Hanken, in press; Roth et al. 1990; in a ratchet-like manner, tending to increase Wake, in press). unless countered by other factors (the“selfish DNA” hypothesis). While there is some evidence Miniaturization is found in several fami- that genome sizes can both increase and de- lies of salamanders, but it is particularly evi- crease within lineages of salamanders (Ses- dent in the Plethodontidae, in which it has sions and Larson 19871, there might be some- evolved repeatedly. I consider any species that thing of a ratchet effect, since all of the becomes sexually mature at body sizes of about bolitoglossines have relatively large genome 30 mm to be miniaturized, and by this criterion sizes, and in different genera (for example, there are numerous miniaturized pletho- Hydromantes and Bolitoglossa) the genomes dontids (some examples are: Desmognathus have become enormous. aeneus, D. wrighti, Hemidactylium scutatum, Eurycea quadridigitata, Plethodon websteri, In some miniaturized species, such as Batrachoseps attenuatus, Bolitoglossa rufes- Desmognathus aeneus, no special problem cens, Chiropterotriton dimidiata, Dendro- arises with miniaturization, for these small EVOLUTIONARY STUDIES OF SALAMANDERS 167 salamanders also have smallcells. As genome receptors in the eye (contrasted with about sizes increase, however, problems arise. This 450,000 in a frog such as Rum),but there are is perhaps most evident in members of the 30,000 retinal ganglion cells; thus, the whole Mexican genus Thorius (Figure l),studied retina is a functional fovea (Roth, Rottluff et extensively by Hanken (1982, 1983u, 1984) al. 1988). The main integration center in am- and Roth et al. (1990). In such cases hier- phibian brains, the optic tectum.(which con- archical interactions have profound impact on tains the cells that are directly related to vi- organismal-level phenomena, such as struc- sual function), has only about 35,000 cells ture of the skull and of the limbs, and espe- (Roth et al. 19901, as compared to about cially of the central nervous system. 500,000 cells in that of Ram. Most species of Thoriusinhabit small spaces Although the Thorius brains are extraordi- under bark or surface cover, and their eyes narily tiny, visual acuity and distance percep- protrude only slightly beyond the limits of the tion remain at effective levels. A number of head. Since the cells are large, however (be- compensatory changes in the brain have been cause the genome is large), the eye must be identified by Roth et al. (1990). The visual large in relation to the head in order to be centers are relatively enlarged at the expense image-recording. Thorius has the most highly of other areas, such as the forebrain. Cell- specialized projectile tongues (Lombard and packing approaches perfect packing density- Wake 19771, and they are dependent on rela- apparently at the expense of glial cells, which tively high visual acuity for feeding, as are all are extraordinarily reduced in number. Glial plethodontids (Roth 1987). The eyes impinge cells are generally thought to play critical roles strongly on the space for the brain, however, in brain function, andinothervertebrates they which lies between the eyes in the platytrabic are about as abundant as neurons. This trade- (that is, broad and flat, lying between two off does not occur even in humans, with our cranial trabeculae) brain case. The impinging greatly expanded brains and trillions of neu- eyes lead to deformation of the forebrain and rons. The close packing and large size of the to posterior displacement (illustrated by Roth neurons in these tiny has many im- et al. 1990).There are only about 25,000photo- plications for such phenomena as cell migra-

FIGURE1. Thorius pennatulus (from near Orizaba, Mexico) and Pseudoeurycea bellii (from near Putla de Guerrero, Oaxaca, Mexico), showing the smallest and largest species of tropical salamanders (family Plethodontidae, tribe Bolitoglossini). Scale is in centimeters. 168 DAVID B. WAKE tion (which seems to be strongly impeded) and ONTOGENY AND PHYLOGENY may well contribute to the general impact of paedomorphosis in bolitoglossines. The existence of the axolotl (Ambystoma mexicanum)and the widely knownlarval state Exactly what these tradeoffs and compro- of such curious organisms as Necturus and mises mean to these species functionally is Proteus force salamander biologists to accept unclear, but one might expect the effects to be the fact that paedomorphosis has played an most strongly felt at hatching. It turns out important role in salamander evolution. What that the hatchlings of Bolitoglossa, which we has been recognized recently is that we have have studied in the laboratory at Berkeley, focused too much attention on the phenome- are very altricial, and they do not move much non of larval reproduction, and this has for some days. Occasionally we find, however, diverted us from appreciating how profound a one-month-old climbing to the top of the influence of paedomorphosis has been in its container(anorma1behavior for larger ani- direct-developingsalamandersthat lack alar- mals). For some weeks they do not eat, and val stage. These direct-developing organisms perhaps they are incapable ofusing their eyes &splay ontogenetic repatterning, an exten- at first. Other general effects might be pre- sive reordering of morphogenetic events in dicted. For example, the forebrain of bolito- time and space that can lead to novelty (Wake glossine salamanders is relatively small, and and Roth 1989).Ontogenetic repatterning has Sever et al. (1990) have shown that these been of profound influence in amniote evolu- animals also have poorly developed cloacal tion and I believe that it is important in direct- glands. I predict that tropical bolitoglossines, developing amphibians as well. in particular, will turn out to be less oriented to olfactory cues in daily and courtship behav- There is a tendency to use the terms “neo- ior than are North American species such as teny” and “paedomorphosis” for ecologically Plethodon cinereus and Desmognathus ochro- labile conditions. I prefer to restrict the terms phaeus, which use olfactory cues for many to phylogenetic contexts, but this may be a functions (David and Jaeger 1981;Horne and losing battle. The present state of affairs is a Jaeger 1988;Houck and Reagan 1990;Jaeger muddle, with some biologists even calling late- 1990;Mathis 1990). I would also expect them metamorphosing larvae that reproduce in a to show relatively little territoriality as a con- single reproductive season paedomorphic. I sequence. Further studies are needed to test recommend using the terms “larval repro- these predictions. duction,” or “perennibranchiate,” for larvae that reproduce in normally metamorphosing I have used miniaturization to show how species, and “delayed metamorphosis” for evolution at one hierarchical level can have those individuals that do not metamorphose an effect on another, in both upward and down- at the expected time. I will discuss here the ward directions. The organism is in essence phylogenetic phenomena and will restrict “squeezedn by the combination of downward paedomorphosis to this level (compare with effects from the population level imposing hilly, in press). To illustrate my point, I will miniaturization and upward effects from in- focus on the bolitoglossine plethodontid creased genome size imposing large cells. salamanders, for they have been studied most Something has to give, and does. It presents extensively in regard to ontogenetic phe- us with an excellent opportunity to explore nomena (for example, Alberch and Alberch the role of certain morphological features in 1981;Hanken l982,1983a, 1984;Wake 1966, behavior and ecology. 1989). EVOLUTIONARY STUDIES OF SALAMANDERS 169

Recently, it was discovered that the bolito- parative osteology has been used to demon- glossines differ from other salamanders in strate paedomorphic effects. The absence of many aspects of central nervous system late-developing bones, such as the prefrontal morphology (Roth, Nishikawa et al. 1988; and septomaxillary in many tropical sala- Wake et al. 19881, and we believe that much manders, is probably the result of this phe- of this derives from paedomorphosis (Roth et nomenon (Wake 1966; Wake and Elias 1983). al., in press). For example, in the bolito- Such paedomorphic evolution leads to pro- glossines-which differ from other pletho- found difficulties (because of extensive homo- dontids in having higher to much-higher plasy) in using such characters in phyloge- genome sizes and thus different cellular char- netic reconstruction (Wake 1989,and in press). acteristics (larger cells with longer cell cyclesbthere is a striking difference in the Homoplasy is very common in salamanders; organization of the motor nuclei that control paedomorphosis is only one of the reasons. feeding. In Plethodon and other outgroups Some years ago (Wake 1970), I pointed out (virtually all other ), these motor that structural opisthocoely (a bony, cartilage- nuclei have a row of relatively undifferentiated covered cap on the anterior end of a vertebral cells in a medial column and a row of differen- centrum, fitting into abony, cartilage-covered tiated cells in a lateral column. In bolito- depression on the posterior end of a more an- glossines there is a different organization; cells terior vertebra) arises from two very different arise medially from the ependymal layer, as developmental patterns in salamanders. One in other vertebrates, but they fail to migrate is the result of an extension of the normal pat- fully and they never fully differentiate (Wake tern of development of ancestral taxa, while et al. 1988;Nishikawaet al. 1991).As aresult, the other results from a novel pattern of devel- there is only a single, rather ragged column of opment in highly derived, miniaturized taxa. undifferentiated cells. These cells are retarded, Regardless of the developmental pathway, that is, they are paedomorphic in relation to there is only a limited structural response, so the homologous cells in less-derived taxa. This we find a true convergence (Wake, in press). phenomenon is evident in other parts of the A final example illustrates the complexity brain as well, which in salamanders in gen- of the evolutionary pathways leading to the eral and bolitoglossines in particular has a diversification of structure in the premaxil- very simple, embryonic appearance, as noted lary bone of plethodontids. To oversimplify, by many authors (Roth et al., in press). We the bone is either paired or fused, but the ways believe that the paedomorphic cells in the in which this happens varies (Wake 1966, motor nuclei of bolitoglossines are a manifes- 1989; Wake and Larson 1987). Larval pletho- tation of a general ontogenetic repatterning dontids all have a single bone, although the in these species, and that there are some im- ancestral condition must have been paired. portant functional implications. For example, The ancestral state might result from meta- among salamanders, it is only the bolito- morphosis or from continued development in glossines that fail to lunge forward during direct-developing taxa. Conversely, the lar- feeding (Larsen et al. 1989); we believe that val state might be retained (by either paedo- this is connected to ontogenetic repatterning morphosis or peramorphosis). There has been (Wake et al. 1988; Nishikawa et al. 1991). extensive convergence, and possibly reversal, Ofcourse, paedomorphosis is evident at sev- and this single apparently simple character eral levels oforganization, from the cellular to reveals the complexities that arise from knowl- the organ to the organismal. Historically, com- edge of ontogeny. 170 DAVID B. WAKE

RECOGNITION OF SPECIES species because we had inadequate geographic sampling (we did examine 29 populations, Studies of protein variation are revealing pre- which also says something about the need for viously undetected diversity in the genetics of farmore extensive and in-depth sampling than species and genera of salamanders. Salaman- has been characteristic of previous work). ders live in a chemical world, and mate-recog- Subsequently, additional sampling was done nition signals are largely not visible to us. (Good and Wake, in press). We now conclude Thus, our impression is that they are as mo- that there are four distinct species and that notonous genetically as some of them appear the most recent divergence is several million to be morphologically. This is very mislead- years old. There are some color differences, ing. I will present just a few examples and use and some subtle osteological differences, but them to raise questions concerning the crite- there are no detectable differences in mor- ria we should use to recognize species-level phometry, and there are only small ecological taxa in salamanders. differences (some species mature in 4% to 5 years, but others in 5% to 6 years). In 1987, David Good, Gloria Wurst, and I presented data that showed that the Olympic The more general message from this study salamander, Rhyacotriton, is really far more relates to criteria for recognition of species. complicated that it appears. This group of My position is rather close to that of Frost and small, stream- and spring-dwelling sala- Hillis (1990).In the case of Rhyacotriton, we manders occurs from Washington State to used the criterionofgenetic cohesion, together northern California, west of the Cascade with a criterion of genetic and geographic dis- Mountains. Itwas discoveredlate(Gaige 1917) tinctiveness. When we plotted Nei genetic and attracted little attention for many years, distance against geographic distance, we found because it was thought to be some kind of a straight line going through the origin for all ambystomatid (Dunn 1920). Stebbins and comparisons within species, but the regres- Lowe (1951)examined geographic variation sion line rises along the y-axis between spe- in morphology and coloration, and concluded cies. For us this was a very useful factor in that two subspecies should be recognized, one designating species limits and borders. It sug- in the north and along the coast andone in the gests that the world of salamanders is very south and inland to the Cascade Mountains, large indeed, and that isolation by distance is but the degree of differentiation was thought a phenomenon that might generally apply. to be relatively slight. The zone of inter- gradation between these two was mapped as Highton’s (1989)taxonomic revision of the being about as extensive as the range ofeither. Plethodon glutinosus complex has proven to be controversial, but if the same criteria we We undertook our study of protein varia- used with Rhyacotriton are applied-rather tion because I had detected some geographic than his apparently (see below) arbitrarylevel variation in osteology (Wake 1981a) and of Nei genetic distance-results very similar wanted toreject the possibility that there were to his are obtained((;;ood,in prep.). When geo- different species involved. I had had exten- graphic distances are plotted against genetic sive experience with the species and thought distances, the regression lines are nearly flat that it was clearly a single species. We were and intercept the Y-axiswell above the origin, shocked to discover that there was extensive or the comparisons form polygons positioned subdivision within the genus, and we only re- relatively high on the y-axis. This indicates frained from breaking the group into different that the taxa identified by Highton are dis- EVOLUTIONARY STUDIES OF SALAMANDERS 17 1

Crete units by this criterion as well as by the ethological studies. Based on laboratory court- criterion he reported (discrete clusters in a ship experiments, an ethological isolation in- UPGMA phenogram; see also Highton [19901). dex was devised. Amultiple-correlation analy- sis showed interactions between genetic, What is not emphasized by Highton is that geographic, and ethological isolation; when ge- he and his group, in extensive field and labo- netic distance is held constant there is a sig- ratory work, have focused on the borders be- nificant relationship between geographic and tween taxa. For example, Duncan andHighton ethological distance. When geographic dis- (1979) examined in detail the secondary con- tance is held constant, however, the relation tact between Plethodon ouachitae and P. between genetic distance and level of etho- fourchensis, and showed that there is a very logical isolation is not significant. Tilley and sharp gene frequency cline between the two, his colleagues concluded that ethological iso- occurring in a restricted geographic zone. One lation and genetic divergence reflect the could take the position that these kinds of gradual divergence of allopatric populations. interactions illustrate that reproductive isola- tion has not yet evolved, and that these are Somewhat surprising, then, is their finding semi-species. Even if that is the case, how- that the relationship of geographic to genetic ever, I support Highton in recognizing such distance is not significant when ethological groups as species-taxa. While I would empha- isolation is held constant. A clue is provided size pattern over magnitude of genetic differ- by the published plot of genetic against geo- entiation (and hence largely agree with Frost graphic distance. There are two dramatic out- and Hillis [19901), I also believe that the rule liers, in which genetic distance is much higher of thumb level of genetic distance (D = 0.15) than expected. These spots both involve the that Highton has adopted is informed by his northernmost population (Mount Rogers, Vir- extensive and in-depth knowledge of interac- ginia). One reported value of genetic distance tions on the borders of the ranges of geneti- is shockingly high: D = 0.643 for the compari- cally cohesive groups. Furthermore, his appar- son of Mount Rogers with Mount Mitchell, ently arbitrary approach using level of North Carolina, a geographic separation of differentiation is not arbitrary when coupled only about 100 km. Furthermore, the small- with geographic analysis that elucidates the est genetic distance of Mount Rogers to any of relation to history of the taxa. Certainly there the other populations is 0.384,a relatively high is no disagreement with the recognition ofcryp- level of differentiation in such a small geo- tic, sympatric species; the controversy arises graphic region (compare with the results re- solely in respect to the recognition of allopat- ported by Highton 1989). The authors noted ric and parapatric forms (see also Larson 1984). that genetic &stances for comparisons with the Mount Rogers population show no ten- Another study that relates directly to the dency to increase with geographic distance. controversy over recognition of species-taxa is From data presented, one might conclude that based on an experimental approach to repro- the northernmost population is most appro- ductive isolation (Tilley et al. 1990). Desrno- priately placed in a separate species-taxon, gnathus ochrophaeus is a small salamander but the situation is more complex than pre- that is widespread in northeastern North sented (Tilley, pers. comm.). Perhaps it would America, but becomes fragmented geo- have been useful to tag this outlying popula- graphically in the southern portions of its tion throughout the study and indicate that it range. In this study, salamanders were se- is a “candidate-species” (that is, it might be lected from ll populations for allozymic and assignable to D. ochrophaeus [sensu stricto], 172 DAVID B. WAKE while the others might be best considered D. species,” Ensatina eschscholtzii, has shown carolinensis). For example, it would be useful that there are two morphologically and ge- to see how the Mount Rogers population ranks netically dfferentiated species living in sym- as an out-group, for then one could appropri- patry in southern California (Wake andYanev ately examine the break-up of what appears 1986; Wake et al. 1986, 1989), but the geo- to be a separate historical entity. graphic scale ofthe studies conducted was too coarse to permit detection of species borders. To return briefly to the Rhyacotriton ex- Frost and Hillis (1990) argued that the two ample, in that study the critical factor in de- groups in southern California should be rec- termining species status is at what point one ognized taxonomically as separate species, and moves from a pattern of gene flow within a that other species in the complex would be group, or among recently separated groups, to recognized in the future (see also Collins 1991). a hierarchical pattern that indicates phyloge- What they did not know was that large, un- netic independence. I once argued that allo- published data sets fill in many of the geo- patric populations should not be given species graphic gaps in the previous studies. What we status on grounds of allozyme differentiation are seeking, before disrupting the current long- alone “in the absence of significant morpho- stable , is some evidence of mono- logical differentiation” (Wake 1981b). Find- phyly for the separate segments of the ring ings since that time, and in particular, my andof a hierarchical structure within the com- experience with Rhyacotriton, have led me to plex. To date, it appears that genetic distance modify that view. In the case ofRhyacotriton, builds largely as a function of isolation and we see an instance in which only trivial mor- distance throughout the ring, and that the phological differentiation has occurred and in geographic gaps in distribution correspond which morphometric analysis shows that more or less to what one would predict for the populations are more similar to genetically measured genetic distances, based on regions distinct nearby populations than to geo- ofcontinuous distribution in the ring. Further- graphically more-remote but genetically less- more, data from nucleotide sequences of mito- distinct populations. This and other studies chondrial DNA shows that the classical in- lead me to conclude that it is the pattern of terpretation of Stebbins (1949), gradual genetic differentiation and the degree to which divergence in allopatry, cannot be rejected genetically cohesive units can be recognized (Moritz et al., in press). I agree with Frost and that are the critical elements in recognition of Hillis that we should refer to the components cryptic and sibling species. In general, I agree of this group as the E. eschscholtzii complex with many recent authors in arguing that it is until the taxonomy is settled. more useful to finely divide taxa so as not to obscure information. We lose more informa- tion by “lumping“ than by “splitting,”and this SPECIES DIVERSITY is especially important when we are concerned with maintaining biological diversity and Recognition of species relates directly to spe- studying its implications for evolutionary and cies diversity. Species ofsalamanders continue biogeographic analyses. to be discovered at a high rate. Some are subdi- visions of taxa formerly considered to be single It is not simple, even with extensive data species (see examples above), but others rep- bases, to make decisions concerning the tax- resent new discoveries. In particular, in the onomy of differentiated and fragmented plethodontid tribe Bolitoglossini, centered in groups. My own work on the celebrated “ring- Middle America, there is very high diversity EVOLUTIONARY STUDIES OF SALAMANDERS 173 as measured in relative number ofspecies and For a number of years my colleagues and I in terms of numbers of co-occurring species have attempted to work out the patterns of and the degree of ecological segregation. relationships of the tropical salamanders, and we have also attempted to discern the main According to my most recent calculation, features of the local and regional distribution plethodontids constitute about 65% of the of taxa(Wake andLynch 1976;Wake andElias species of salamanders. Other large groups 1983; Elias 1984; Larson 1983; Wake 1987). include the families Hynobiidae (as currently We have focused our attention on studies of recognized), with a large number of species in local elevational transects, for since the time far eastern Asia into Central Asia and as far of Schmidt (1936),it has been recognized that as the Ural Mountains; Ambystomatidae, elevational zonation of the tropical species is which are exclusively North American in dis- much more marked than is that of extratropi- tribution; and the Salamandridae. The last is cal species. In nearly every instance in which the only family that approaches a cosmopoli- we have conducted detailed local studies along tan distribution, in the sense that it occurs in elevational transects, we have uncoverednov- all major centers of distribution of salaman- elties, which suggests that we still have agreat ders except the New World tropics. The deal to learn-even at the level of species rec- Plethodontidae is represented by a few spe- ognition-as far as tropical salamanders are cies in the central Mediterranean region of concerned. Europe, but is otherwise exclusively New World in distribution. The plethodontid sub- Generalities are beginning to emerge. family Desmognathinae and a number of gen- Whereas in the temperate parts of the globe era of the subfamily Plethodontinae (includ- salamander diversity is related to ing groups with larval development and with diversity, life-history diversity, and larval direct development) occur in eastern North adaptations, in the tropics the picture is very America, long considered to be the area of on- different. First, all of the species are strictly gin and center of diversification of the family terrestrial, and second, all of the species uti- (Dunn 1926). Only one supergenus of the lize the same feeding mechanisms based on a plethodontine tribe Bolitoglossini is found in highly projectile tongue. Relatively few spe- the New World tropics, but this group is of cies are microsympatric-I believe the great- great interest because of its impressive diver- est number is six, in Guatemala, but this sity (Figure 1). It now includes about two- number might be greater in the Tapanti for- thirds of the plethodontids and over 45% of all est reserve in Costa Rica (Wake and Lynch species of salamanders. 1976; Wake 1987; Wake et al., in press). The main factors separating species are altitude Despite the relatively great numbers of and microhabitat. There has been extensive tropical salamanders, these salamanders are homoplasy based on microhabitat speciali- fairly obscure, even to the scientific com- zation, and species in different genera have munity. A recent example of the failure to independently evolved adaptations for climb- appreciate this diversity is the account of ing and for fossorial life. Arbreality is espe- salamanders (Halliday and Verrell 1986) in cially common. In the lowlands, where compe- “The Encyclopedia of Reptiles and Amphib- tition with other vertebrates is expected to be ians,” which almost completely ignores the most intense, one rarely encounters a tropical species and gives no indication that salamander on the soil surface. Rather, the nearly half of the salamanders in the world species are almost universally arboreal or are tropical! fossorial. 174 DAVID B. WAKE

At the present time I know ofapproximately the public can more easily relate to such sym- 25 well-differentiated species of tropical sala- bols. But the megafauna has no more intrinsic manders that await description, and there are worth than these remarkable tropical sala- a number of additional cryptic species that manders. We who are informed have a special have been detected by electrophoretic analy- obligation to work in the political and social sis (for example, Hanken 19833). Some of the arena at local, regional, national, and inter- recently described species have been among national levels to check the destructive forces the most spectacular salamanders yet discov- that are permanently altering the evolution- ered. Destruction of tropical , how- ary potential not only of salamanders, and ever, is proceeding at such a rapid rate that other amphibians and reptiles, but also of we are at risk of losing many species before Homo sapiens. they are even discovered. A good example is At the First World Congress of Herpetol- the newly described genus and species of scan- ogy, many of us first became aware of the fact sorial salamander from Chiapas, Mexice that many amphibians are in danger of be- niger (Figure 2kwhich is capable coming extinct and that popula- of leaping and bounding along vertical rock tions are in decline in many parts of the world. faces and on tree trunks (Wake and Johnson I had personally been aware of the decline of 1989). We fear that it may already be extinct. frog populations in California and of the de- Within the past ten years two other genera of cline of populations of frogs and salamanders equally interesting tropical salamanders have in Costa Rica, but the congress focused atten- been discovered, Nyctanolis and Bradytriton. tion on what was apparently a general phe- nomenon. I returned to the United States to A PLEA FOR CONSERVATION work with colleagues on the Board on Biology of the National Research Council to convene a The world’s biota faces a crisis of unprece- workshop to examine the issue. The meeting dented proportions. has resulted in an understanding that there was accelerated to an alarming rate. Up to now, con- indeed a problem, and elicited a great deal of servationists have appropriately stressed the attention in the news media. A number of rec- megafauna, such as Kemp’s ridley turtle, for ommendations resulted (Blaustein and Wake 199 11, and there has been substantial progress (Wake 1991). The most noteworthy develop- ment has been the establishment of a Task Force on Declining Amphibian Populations by the Species Survival Commission of the Inter- national Union for Conservation of Nature and Natural Resources (IUCN). Perhaps amore lasting outcome has been a growing aware- ness that amphibians in general, because of a whole complex of physiological, morphologi- cal, ecological, behavioral, and life historical factors, may have extraordinary potential as indicators of environmental degradation and FIGURE2. Inahtriton niger (Wake and Johnson 1989), a remarkable,newly discovered salamander from Chiapas, even of global climate change. We may yet Mexico, that was placed in a new genus and species. come to see how apparently esoteric studies of This taxon may already be extinct. Scale is 25 mm simple salamanders have global impact. EVOLUTIONARY STUDIES OF SALAMANDERS 175

ACKNOWLEDGMENTS Gaige, H. 1917. Description of a new salamander from Washington. Occas. Papers Mus. Zool. Univ. Michi- I thank the organizers of the First World Con- gan, 40: 1-3. Good, D. A., andD. B. Wake. In press. Geographicvaria- gress of Herpetology for the invitation to tion and speciation in the torrent salamanders of the present a plenary lecture. I thank Richard genus Rhyactotriton (Caudata: Rhyacotritonidae). Cloutier, David Hillis, Allan Larson, Steve Univ. California Publ. Zool. %illy, and Linda Trueb for providing infor- Good, D. A., G. Z. Wurst, andD. B. Wake. 1987.Patterns of geographic variation in allozymes of the Olympic mation on work in progress. My research has salamander Rhyacotriton olympicus (Caudata: been sponsored by a succession of grants from Dicamptodontidae). Fieldiana: Zool., new ser., 32: the U.S. National Science Foundation (the 1-15. Halliday, T. R., and P. Verrell. 1986. Salamanders and present grant is BSR 90-19810). newts, p. 18-31. In T. Halliday and K. Adler (eds.), The Encyclopedia of Reptiles andhphibians. Facts LITERATURE CITED on File, New York. Hanken, J. 1982. Appendicular skeletal morphology in minute salamanders, genus Thorius (Amphibia: Alberch, P., and J. Alberch. 1981. Heterochronic mecha- Plethodontidae): growth regulation, adult size deter- nisms of morphological diversification and evolution- mination and natural variation. Jour. Morphol., 174: ary change in the neotropical salamander, Bolito- 57-77. glossa occidentalis (Amphibia:Plethodontidae). Jour. -. 1983a. Miniaturization and its effects on cranial Morphol., 167: 249-264. morphology in plethodontid salamanders, genus Blaustein, A., and D. B. Wake. 1991. Declining amphib- Thorius (Amphibia: Plethodontidae). 11. The fate of ian populations: a global phenomenon? Trends Ecol. the brain and sense organs and their role in skull Evol., 5: 203-204. morphogenesis and evolution. Jour. Morphol., 177: Collins, J.T. 1991.Viewpoint:anewtaxonomicarrange- 255-268. ment for some North American amphibians and rep- 1983b. Genetic variation in a dwarfed lineage, the tiles. Herpetol. Rev., 22: 42-43. -. Mexican salamander genus Thorius (Amphibia: David, R. S., andR. G. Jaeger. 1981. Prey location through Plethodontidae); taxonomic, ecologic, and evolution- chemical cues by a terrestrial salamander. Copeia, ary implications. Copeia, 1983: 1051-1073. 1981: 435-440. Dubois, A. 1984. Miscellanea nomenclatorica batra- -. 1984. Miniaturization and its effects on cranial mor- chologica (V). Alytes, 3: 111-116. phology in plethodontid salamanders, genus Thorius Duellman, W. E., andL. Trueb. "1986"( 1985). Biology of (Amphibia:Plethodontidae). I. Osteological variation. Amphibians. McGraw-Hill Book Co., New York. xix, Biol. Jour. Linnean SOC.London, 23: 55-75. 670 pages. -. In press. Adaptation of bone growth to miniaturi- Duncan, R., andR. Highton. 1979.Genetic relationships zation of body size, chapter 4. In B. K. Hall (ed.), ofthe eastern large Plethodon ofthe OuachitaMoun- Bone,Vol. 7:BoneGrowth-B. CRC Press,BocaRaton, tains. Copeia, 1979: 95-110. F1 orida . Dunn, E. R. 1920. Notes on two Pacific coast Amby- Highton, R. 1989. Biochemical evolution in the slimy stomidae. Proc. New England Zool. Club, 7: 55-59. salamanders of the Plethodon glutinosus complex in -. 1926.TheSalamande1-softheFamily Plethodontidae. the eastern United States. Part I. Geographic pro- Smith College, Northampton, Massachusetts. vii, 441 tein Variation. Illinois Biol. Monog., 57: 1-78. pages (reprinted: SOC.Study Amphib. Reptiles, 1972). -. 1990. Taxonomic treatment of genetically differenti- Edwards, J. L. 1976. Spinal nerves and their bearing on ated populations. Herpetologica, 46: 114-121. salamander phylogeny. Jour. Morphol., 148: 305-328. Hillis, D. M. 1991.The phylogeny of amphibians: current Eldredge, N. 1985. Unfinished Synthesis; Biological knowledge and the role of cytogenetics, p. 7-31.h D. Hierarchies and Modern Evolutionary Thought. M. Green and S. K. Sessions (eds.),Amphibian Cyto- Oxford Univ. Press, Oxford. vii, 237 pages. genetics and Evolution. Academic Press, New York. Elias, P. 1984. Salamanders of the northwestern high- Horne, E. A., and R. G. Jaeger. 1988. Territorial phero- lands ofGuatemala. Nat. Hist. Mus. Los Angeles Co., mones of female red-backed salamanders. Ethology, Contrib. Sci., 348: 1-20. 78: 143-152. Frost, D. (ed.). 1985. Amphibian Species of the World. Houck, L. D. 1977. Life history patterns and reproduc- Allen Press, Lawrence, Kansas. v, 732 pages. tive biology of neotropical salamanders, p. 43-72. In Frost, D. R., and D. M. Hillis. 1990. Species in concept D. H. Taylor and S. I. Guttman (eds.), The Reproduc- and practice: herpetological applications. Herpe- tive Biology of Amphibians. Plenum, New York. tologica, 46: 87-104. Houck, L. D., and N. L. Reagan. 1990. Male courtship 176 DAVID B. WAKE

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