Proc. Nat. Acad. Sci. USA Vol. 71, No. 9, pp. 3517-3521, September 1974

Three and Three Islands: Parallel Evolution in ( Sea/Indian Qcean/ecology/land ) HAMPTON L. CARSON Department of Genetics, University of Hawaii, Honolulu, Hawaii 96822 Communicated by Th. Dobzhansky, May 9, 1974

ABSTRACT Most organisms are evolutionary conserva- status of the that he studies and decide which are the tives; they may subdivide their niches but tend to remain which promise of be- within them. Yet the fossil record shows many cases of unalterable "conservatives" and gives breakthrough to a new mode of life. How may such evolu- coming the rare "liberal" that starts in a new direction. tionary innovation be recognized at this time level? Three In many world-wide groups of organisms, the most unusual species of Drosophila have accomplished an innovation in and deviant members are found on islands. An island need not that they breed as obligate commensals on tropical land be literally defined; caves and isolated mountain peaks qualify crabs. This could be dismissed as a curious aberration were it not for the fact that the three flies concerned repre- in the present context (2). The peculiarities of island life are sent three different phyletic lines of the family. Although often dismissed as evolutionary cul-de-sacs, interesting only Drosophila is abundant on the continents, these three as oddities arising in an isolated and often discordant eco- parallel innovative evolutions have occurred on islands. system. In my view, these unusual evolutionary events de- The proposal is made that the genetic systems of many for the conservative groups of organisms carry variability that mand special attention as possible prototype systems would permit them to evolve in a novel direction. The evolutionary synthesis of novelties. realization of this capacity, however, is possible only under special environmental conditions. The biology of Drosophila New information that has come to light on certain members Evolutionary innovation of the family Drosophilidae may suggest ways to approach As they occur on the continents, the members of a family of the problem of evolutionary innovation. Probably more is organisms tend to be conservative in their . Once known about the comparative genetic systems of Drosophila a basic ecological niche has been conquered, evolution leading than any other closely knit group of higher organisms. Less in a drastically different direction is rare. Related species widely recognized is the fact that such genetic studies can tend to repeat the old theme with only minor variations. now be interpreted in the light of an increasingly sophisticated Thus, most contemporary species belonging to a single family knowledge of the systematics and ecology of these flies. Ap- have a certain dull sameness about their ecologies, even in proximately 2500 species of the family have been described, the presence of sprightly morphological differences. and data on the life histories of many species have accumu- Most differences between species do indeed have ecological lated. Most species are local endemics; only a few have come meaning, but this lies often in the development of delicate and to be associated with man on a world-wide basis (3, 4). recondite methods of packing species into an ever increasingly Drosophila are small, highly vagile insects that subsist on subdivided niche and avoiding competition. Although such , especially yeast and bacteria. The common microadaptational developments are biologically important, substrates supporting the relevant microorganisms in nature they rarely provide clues as to how large-scale evolutionary are fruit, fungus, flowers, bark, roots, slime flux (infected sap innovations are accomplished. Yet, when confronted with the flows), fallen leaves, and broken stems. As decay begins, fossil record, one sees abundant evidence of biological break- Drosophila is usually one of the first insects to arrive. The throughs from one basic adaptive norm to another. adults feed both on the carbohydrates and proteins released Everyone knows, for example, the famous diagram showing from the substrates, and subsequently they consume the "adaptative radiation" in the mammals (1). A small terres- microorganisms themselves. Drosophila flies are usually trial generalist sits in the center of the picture, surrounded opportunistic as feeding adults; they visit many types of by a circle of representatives of the different families; there ephemeral carbohydrate sources before the latter dessicate. are burrowing, flying, aquatic, and arboreal forms. Surely Oviposition by gravid females tends to be much more precise, the characters that adapt these organisms, now so useful in and breeding sites are often visited by only one or a few the systematic diagnosis of higher-order differences, each species. must have had its inception in some real, ancient Mendelian population. Is our ingenuity, technical skill, and scientific Drosophila and land crabs imagination not great enough to decipher such beginnings? Departing drastically from the above pattern are three species Or are they so infrequent as to preclude their being recog- of drosophilid flies from insular areas; two of these are from nized at the present time level? Is the microevolutionist in- the and one is from the Indian Ocean. Each escapably condemned to remain mired in trivia? stems from a different phyletic line, yet each has evolved, What sort of change, at the present time level, might indeed in closely parallel fashion, into a truly novel ecological associa- constitute the microevolutionary inception of innovative tion for the family. They have adopted an intimate associa- evolution? Each ecological geneticist must survey the adaptive tion of obligate with land crabs. 3517 Downloaded by guest on October 1, 2021 35183518Zoology:Zoology:CarsonCarson~~~~~~~~~~~~Proc.Nat. Acad. Sci. USA 71 (1974)

TABLE 1. Comparison of three species of Drosophila The decapod Crustacea are primarily aquatic . Nevertheless, within several families, groups of species have Drosophila Lissocephala evolved that have at least partially conquered terrestrial Drosophila endobranchia Powelli habitats, especially in the tropical parts of the world. Because carcinophila Carson and Carson and the adults live out much of their lives above the tide Wheeler Wheeler high Species Wheeler level, these animals are commonly referred to as land crabs, Subgenus Drosophila Sturtevant although all of them apparently retain aquatic larval stages. Species re,'pleta quinaria- Separate Crabs belonging to two genera of the family , group (mercatorum virilis section subfamily and , are especially adapted for ter- complex) (aberrant restrial life; these two are the most important hosts for the form) obligatory Drosophila commensals. The two genera are Distribution Nreo'tropical Holarctic , south1- closely similar and together include forms that have of group east Asia, global Micronesia pan-tropical oceanic distributions, being largely confined to Distribution Islands -of West Cayman' Christmas small, isolated, but richly vegetated islands. Descriptions of of species Indies and Islands and Island, these three cases of Drosophila commensalism have recently Caribbean Sea Indian Oc3ean been published (5, 6, 8). The three are compared and the (widespread) basic facts a-re summarized in Table 1. What follows is a brief Sympatry All three species allopatric account of the biological relationships observed as the three Group breeding Fungus and/ Fungus flies and their crabs have been studied on three islands. or bark ecology breeding? Drosophila carcinophila- on Cat Cay. North Cat Cay is a breeding? small Crab hosts Gecarcinus8 Gecarcinus Gecarcoidea island in south of the Bimini group, about km east ruricola ruricola; humei; Bii,rgus 80 of the Florida coast (northernmost closed circle, Gecarcinus latro; Geo- Fig. 1). Cat Cay is separated from neighboring Gun Cay by a lateralis grapsus crrini- strait, which is a'bout 400 m wide. Both islands are less than pes; Cardi- 3 km long and rise no more than, 10 mn above sea level. De- soma carnifex spite their narrowness (150-200 in), both support sub- Adult flies Have three pairs of dorsocentral Normal dorso- stantial broadleaf coppices growing on the rich black lime- bristles centrals stone soil. This is the habitat of , a large Normal Posterior Normal semi-fossorial land crab. These crabs are especially abundant scutellars scutellars scutellars on Gun which is uninhabited a small turned Cay, except for light- up- G. ruricola rare or on right and house. is absent Bimini, and one may crossed conjecture that this is due primarily to the fact that the Size 2.5 mm 3.8 mm 2.0 mm crab is considered a culinary delicacy. It is, therefore, (length) relentlessly hunted down in the populated parts of its range. Feeding Wholly on : rarely if ever Mostly off the Although it was apparently once common throughout the leave crab; will not come to host; come Greater and Lesser , it remains today principally on' baited traps readily to small, isolated islands. baits The Black Crab, as it is commonly known in the area, is a 4 long filaments; 2 short fila- 12 short fila- strictly terrestrial ', being largely confined to dense placement on ments; ce- ments; place- forest, where it excavates shallow under limestone crab in un- mented on ment at base rocks or tree roots. Despite its common name, it shows exten- known site carapace, of 2nd anten- sive color polymorphim. The crabs apparently leave the near eyes na forests the brief when Larvae All instars re- First inistar in First and third onlyduring spawning season, they migrate as main in ex- nephric instars in to the hard coastal limestone, known the "iron shore." ternal nephric groove; 2nd nephric This is a spectacular mass migration, which occurs only once groove instar in groove; 2nd a year; the crabs remain at the shore briefly and then migrate branchial instar in back to the forests. In , G. ruricola has been taken chamber; branchial as high as 1500 m above sea level. 3rd instar chamber Of 17 crabs caught on Cat Cay, 15 showed infections with near (Gecarcoidea) Drosophila carcinophila. The is small and very active and mandibles or all instars is found around on the often in in branchial running carapace, moving chamber around the mouthparts where it feeds actively. In the labora- (Geograpsus tory, the adult flies are reluctant to leave the crabs; if held in and Birgus) isolation on laboratory food for a few days, they will feed, Puparium 3rd instar Larvae leave the crab and but when released back into a terrarium with the crabs, they secretes ce- pupate in soil are drawn back as if by a magnet. ment; pupariunr The site of oviposition of D. carcinophila on G. ruricola is affixed to 3rd not known, but first-instar larvae of laboratory-infected crabs maxilliped are found embedded in a curious pair of felt-like pads that Duration of 4 weeks 4 months 4 weeks lie in a groove under the third maxillipeds (Fig. 2, arrow).- life cycle (or more) (estimated) This pad is continually moistened by fluid from the openingr at 250 (egg of the green gland, which lies just above it. This fluid reenters to adult) the haemocoel at the base of the carapace groove that sup- Downloaded by guest on October 1, 2021 Proc. Nat. Acad. Sci. USA 71 (1974) Parallel Evolution in Drosophila 3519

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FIG. 2. Anterior (facial) view of Gecarcinus ruricola. The crab's left third maxilliped (to the right as viewed) has been re- moved, exposing the felt-like nephric pad (arrow) in which Droso- phila larvae live. The nephrostome (opening of the green gland) is at the top (dorsal end) of the pad. Photograph by H. D. Stalker. (Gecarcinus FIG. 1. Distribution of the Black Land Crab between YucatAn and Cuba (lowermost solid square, Fig. 1).- ruricola) in the Caribbean Area. 0, No Drosophila infections; 0, forest infection with D. carcinophila; *, infection with D. endobranchia. G. ruricola survives in remote areas where the original still stands. In coastal areas it is accompanied by a -close rela- tive, G. lateralis. This is a sand-burrowing species, which is A of microorganisms occurs ports the pad. copious growth the more abundant of the two species throughout the Carib- turn support all in the interstices of the pad, and these in bean area. three larval instars. The larvae do not leave this groove until Ninety G. ruricola and 54 . lateralis were exa'mined. Sixty- is formed; the latter is tightly cemented to the the puparium one percent of the former and 30% of the latter showed in- maxilliped. inner surface of the third fections with a wholly new species of Drosophila. No speci- groove appears serve a nephric function. The felted to mens of D. carcinophila were found. may be removed by microbial ac- Thus, nitrogenous wastes This new species, D. endobranchia, is a robust, sluggish fly tion before the fluid is reabsorbed, as an apparent water- which, like D. carcinophila, rides on the crab most of the time conservation system. The niche of the Drosophila larvae, (Fig. 3). Its highly modified have a leathery chorion therefore, appears to be that of a -laden urinal. with two rigid, blunt filaments extending directly forward. The fly larvae do not appear to be in a relation of mutualism They are attached to the exposed dorsal surface of the cepha- to the crab because some crabs have no infections. In fact, lothorax, especially around the eyes. from Gun Cay, not one case among 51 crabs examined nearby First instar larvae move briefly to the pads of the nephric of Drosophila infection was discovered. grooves, but soon migrate into the gill chambers, where they from the area When a third instar larva is removed under remain for an extended period, feeding on microorganisms the third maxilliped and placed on a dry microscope slide, it around the gill tissue. Later they move out to a more or less exudes a collagenous material. This forms a cement that fixed position around the mouth of the crab. Here they be- fastens the newly formed puparium to the inner surface of the come literal messmates; as many as nine larvae are arranged third maxilliped. Adult flies emerge within ten days, com- in a halo around the mandibles. When undisturbed, they can pleting the life cycle. extend far out, their ends are anchored at D. carcinophila remains tightly although posterior The distinctive puparia of the base of the mouthparts by an inflated, bursa-like posterior glued to the host even in museum specimens. Examinations end. When feeding ends, the larvae become short and thick showed puparia on 43% of 430 preserved specimens of G. and fall to the ground where they pupate. ruricola. From these data, a detailed determination of the These details suggest that D. endobrartchia is not closely geographical distribution of the fly was possible. Only a few related to D. carcinophila. Furthermore, anatomical studies were available from the islands in the specimens, however, suggest that it has affinities to the virilis or quinaria sections south and western portion of the Caribbean Sea. of the quite removed from the repleta show that Drosophila subgenus Drosophila, Cytological and anatomical studies group. group of the subgenus carcinophila belongs to the repleta D. endobraichia has been found only on Grand and Little Drosophila. This large species-group is neotropical in origin, Caoman and at Guantenamo Bay, Cuba (see Fig. 1). Al- although several species have become world-wide associates though D. carcinophila is found elsewhere in Cuba, there is no of man. The native members of the group inhabit deserts, evidence that the two flies are sympatric. The distribution of where they breed on cactus. Why the novel associations of D. D. endobranchia indicates that it may have evolved as an such a group is not clear, carcinophila should evolve from inhabitant of an ancient mountain axis, now the deeply sub- although several other repleta-group species are endemic to merged Cayman Ridge, which once connected the Sierra cactus on the Caribbean islands. Maestra of southeastern Cuba with YucatAn (7). The Cay- Drosophila endobranchia on Grand Cayman. During the mans are apparently vestiges of this ancient connection. In course of the above investigation, an excursion was made to any event, the two species of crab fly in the Caribbean Sea Grand Cayman Island, which lies in the western Caribbean, have widely different evolutionary histories. Downloaded by guest on October 1, 2021 3520 Zoology: Carson Proc. Nat. Acad. Sci. USA 71 (1974)

FIG. 3. Two specimens of D. endobranchia near the eye of Gecarcinus ruricola. Three eggs are seen on the rim above the eye. Photo- graph by H. D. Stalker. Lissocephala powelli on Christmas Iskand. Christmas is a adult leaves the crab readily and feeds on any kind of fruit, single isolated island on the fringe of the Java trench in the fallen flower, or other substance. They are attracted in large Indian Ocean. The island is oceanic, with a basaltic base numbers to fruit baits artificially placed in the forest. capped by limestone. It rises to about 360 m above sea level This species deposits its 12-filamented eggs at the base of and supports a high tropical rainforest, characteristic of the the second antenna of the crab, and the larvae make their Malay Archipelago. way under the third maxillipeds into the nephric pad region. The island supports large populations of five species of Larvae of all instars may be found there; they are often only land crabs; these same species are widespread on islands in the barely concealed by the hairs at the distal ends of the maxil- equatorial areas of the Pacific Basin. The most abundant is the lipeds. Thus, the larval site resembles D. carcinophila more Red Crab, Gecarcoidea humei, which is found almost every- than D. endobranchia. Nevertheless, examinations show that where on the island where forest remains. about 25% of the larvae (probably second instar) have made Despite the generic distinction, this animal is closely similar their way back into the branchial chamber, in the same man- in habits and ecology to Gecarcinus ruricola of the Caribbean ner as D. endobranchia. Like that species, third instar larvae Sea. Thus, it prefers to make shallow burrows in black lime- do not remain in the branchial chamber but return to the posi- stone soil, feeds on vegetable detritus, and stages spectacular tion under the maxillipeds. When mature, the larvae drop off annual seaward movements for spawning. Of 83 of these crabs and pupate in the soil. Emergence occurs after about 5 or 6 examined, over 95% were found infected with a small crab days. drosophilid, quite unlike either of those from the Caribbean Lissocephala powelli is one of the commonest Drosophila area. The species has been assigned to Lissocephala, one of flies on ; its ubiquity matches that of the most primitive genera of the family Drosophilidae. Gecarcoidea. In addition to the latter, several other species of Lissocephala powelli adults are exceedingly active flies. land crabs are found on the island. Of these, the Robber Crab, With wings constantly in motion, they move rapidly over the Birgus latro, serves as an important host for Lissocephala surface of the crabs. Unlike the Caribbean flies, however, the powelli. Oviposition on this crab takes place at the base of Downloaded by guest on October 1, 2021 Proc. Nat. Acad. Sci. USA 71 (1974) Parallel Evolution in Drosophila 3521

the second antenna, as in Gecarcoidea. In the absence of any sizes, need be invoked. Indeed, it is possible that it could be external nephric pad (Birgus is a terrestrial pagurid, or hermit accomplished by recombination and selection of polygenes crab), the larvae move directly to the gills and the entire and regulatory genes rather than single rare mutations. larval life cycle is spent there. In each of the cases, innovative evolution appears to be con- L. powelli also oviposits on two other species of land crab, nected with insular, not continental, conditions. This means carnifex and Geograpsus crinipes. On Christmas that the population sizes of the ancestral stocks might be Island, these two are less frequent than Gecarcoidea or Birgus, judged to be of small or at least moderate size. being confined to low moist areas near the seacoast. Of 34 These considerations lead to the conclusion that the capac- Cardisoma examined, half showed substantial numbers of ity for innovative evolution may indeed be widespread in Lissocephala eggs, up to 50 per crab. Despite this, very few natural populations. If, as the evidence indicates, response can larvae were found and these were almost exclusively first in- occur widely in small populations, it should be possible to stars. Accordingly, there is considerable question as to whether study the process of evolutionary innovation at the micro- the life cycle can be completed on Cardisoma. Geograpsus evolutionary level. In the context of the present article, it crinipes is rather rare, but stages in the development of would be interesting to assay the response of a species that is Lissocephala were found on four of them. living in the presence of an unoccupied innovative niche. For Rather extensive examinations of these same species of land example, in large areas of the Pacific, land crabs exist in the crabs from islands in the north Pacific Ocean area (Western presence of a variety of species of Drosophila, none of which Carolines and Marshalls) showed no infection with any dip- appears to have made the relevant breakthrough. How many teran associates, and so far the experience at Christmas Island of these are in actual ecological contact needs to be deter- is unique. Crabs of the genus Cardisoma, furthermore, are mined. Can we find evidence for the beginnings of adaptive abundant not only in the Pacific but in the Caribbean area as changes in some of these species? well. During the study of the Gecarcinus association there, Because vast populations do not appear to be necessary, it large numbers of were examined and all would seem possible, furthermore, to study natural selection proved negative. for innovation under laboratory conditions. For example, knowing that the land crab niche is a suitable one for Droso- Discussion phila, experimental exposure of a population of a species In three geographically isolated instances, drosophilid flies deemed competent to adapt in this manner could be arranged have evolved an obligatory commensal association with land in population cages. crabs. In each case, the fly belongs to a distinctly different Most studies of population genetics and evolution have phyletic line of the family. Although these are basically cases dealt with situations in equilibrium or with trivial adaptive of parallel evolution, each association shows unique details. changes. The considerations in this paper lead us to believe The external nephric groove and pad of the gecarcinid crabs that the genetic systems of many organisms are not doomed obviously offers itself as an open and available niche to to change only in trivial ways but, even at this time level, can evolving drosophilids. show innovation if a relevant ecological stimulus impinges on When viewed from the perspective of the ecology of droso- them. philids on a world-wide basis, the evolution of obligatory This work was supported by a grant from the National Science commensalism appears as a remarkable innovation. Other Foundation. than these three crab flies, Drosophila has little use for animals 1. Hegner, R. W. (1942) College Zoology (The MacMillan Com- or animal protein. Males of the genus Amiota are curiously pany, New York), p. 815. attracted to human eyes as one walks through, the woods, 2. Hedberg, 0. (1971) Adaptive Aspects of Insular Evolution, ed. although it seems unlikely that this is much of a clue to their Stern, W. L. (Washington State Univ. Press, Pullman, Wash.), pp. 16-23. ecology. The only other clear animal parasites in this large 3. Wheeler, M. R. & Hamilton, N. (1972) Univ. Texas Publ. family are the species of the Hawaiian scaptomyzoid genus 7213, 257-268. Titanochaeta (9). These flies oviposit into the egg clusters of 4. Carson, H. L. (1971) in "The Ecology of Drosophila Breeding certain endemic Hawaiian spiders and their larvae consume Sites" (Univ. of Hawaii Press, Honolulu), Harold L. Lyon the a such ten Arboretum Lecture No. 2, pp. 1-28. spider eggs. Only single lineage, numbering 5. Carson, H. L. (1967) Amer. Midl. Nat. 78, 324-343. species, is known. 6. Carson, H. L. & Wheeler, M. R. (1968) Ann. Ent. Soc. Amer. That this capacity for innovative evolution was latent in 61, 675-678. three separate phyletic lines suggests that the genetic basis of 7. Schuchert, C. (1935) in Historical Geology of the Antillean- innovation must be widely distributed in natural populations. Caribbean Region (Wiley & Sons, New York), p. 811. 8. Carson, H. L. & Wheeler, M. R. (1973) PacificInsects 15, 199- A corollary of this is that when a truly novel unoccupied niche 208. appears in the environment of a species, no special source of 9. Hardy, D. E. (1965) Drosophilidae of Hawaii (University specific mutational response, available only in vast population of Hawaii Press, Honolulu), p. 814. Downloaded by guest on October 1, 2021