SZAQYSTEMATICS Anopheles (Nyssorhynchus) dominicanus sp. n. (Diptera: Culicidae) from Dominican Amber

1 THOMAS J. ZAVORTINK AND GEORGE O. POINAR, JR.

Department of Biology, University of San Francisco, San Francisco, CA 94117Ð1080

Ann. Entomol. Soc. Am. 93(6): 1230Ð1235 (2000) ABSTRACT The female and egg of Anopheles dominicanus sp. n. are described from Dominican amber. The species is placed tentatively into the subgenus Nyssorhynchus on the basis of the following wing characters: a preapical pale costal spot; an accessory sector pale spot that is not joined to a sector pale spot; pale scales at the apex of vein R4ϩ5; basal, median, and apical pale spots on vein A; and the termination of vein Sc at the proximal end of the subcostal pale spot. This newly described mosquito from Dominican amber establishes an ancient lineage of Anopheles in the New World.

KEY WORDS Anopheles dominicanus, new species, Culicidae, Diptera, mosquitoes, Dominican amber

AS A RESULT of their small size and fragile constitution, Materials and Methods fossil mosquitoes are rare (see Poinar et al. [2000] for The amber with the new anopheline is clear yellow, a list and critical examination of known fossil Culici- with numerous small cloudy droplets, and with many dae). The current study describes the female and egg minute irregular or ovoid particles, some of which are of a new species of spotted-wing Anopheles in amber joined into chains. It includes the adult female Anoph- from the Dominican Republic. This is the Þrst de- eles; two adjacent eggs laid by it; three long, slender, scription of a mosquito from Dominican amber and nonculicid insect legs; a plant bud; and a small, dark, the Þrst fossil anopheline from the New World. hemispheric object covered with fungal hyphae. The The fossil described here is unquestionably an piece of amber is irregularly oval in outline, Ϸ9.5 by Anopheles. Its subgeneric placement is less certain, 11.0 mm, with a portion of its perimeter polished ßat. however, in part because the subgeneric classiÞcation The piece has been polished to a thickness varying of Anopheles is based to a great extent on male char- from Ϸ1.5 to 2.0 mm, and its weight is 0.18 g. The acters, and in part because many taxonomic characters amber has been examined with both stereoscopic of femalesÑdevelopment of the buccopharyngeal ar- (with magniÞcations up to 140 diameters) and com- mature, presence or absence of pruinose markings on pound (with magniÞcations up to 250 diameters) mi- the thorax, presence or absence of scales or scale-tufts croscopes using both transmitted and reßected light. on various parts of the body, and ornamentation of the Measurements were made at various magniÞcations legsÑare missing, not discernable, or not known with with an ocular reticule and the original line drawings certainty. We have had to rely heavily on character- were prepared with the aid of the reticule. Photo- istics of the wing and egg in our attempts to relate the graphs were taken with a Nikon Orthophot micro- fossil Anopheles to modern species. The fossil appears scope. The drawing of the wing (Fig. 2) is diagram- to be most similar to species in the subgenus Nysso- matic and intended only to show the positions of the rhynchus, so we place it there tentatively. pale spots. Morphological terminology follows Mc- The amber containing the mosquito originated from Alpine et al. (1981) in large part, Wilkerson and Pey- the northern mountain ranges of the Dominican Re- ton (1990) for wing spots, and Hinton (1968) and public. Amber mines there are in the El Mamey For- Linley (1992) for egg characters. mation (Upper Eocene), which is a shale-sandstone interspersed with a conglomerate of well-rounded pebbles (Eberle et al. 1980). The exact age of Domin- Results ican amber is unknown, and estimates based on var- ious analyses of microfossils and the amber provide a The fossil Anopheles is damaged, and many charac- range from 15Ð20 million years (Iturralde-Vincent and ters used in anopheline taxonomy are not present. The MacPhee 1996) to 30Ð45 million years (Cepek in proboscis and palpi extend to the edge of the amber, Schlee 1990). and the distal parts are missing. Only the right foreleg is complete, and both midlegs and hindlegs are broken off and missing. Various parts of the mosquitoÑthe 1 Department of Entomology, Oregon State University, Corvallis, head, antennal pedicels, antepronota, proepisterna, all OR 97331Ð2907. coxae, halters, and cerciÑare ßattened, apparently

0013-8746/00/1230Ð1235$02.00/0 ᭧ 2000 Entomological Society of America November 2000 ZAVORTINK AND POINAR:NEW Anopheles FROM DOMINICAN AMBER 1231 having collapsed during fossilization. The thorax is without scales; ßagellomere 5 about six times longer cracked and distorted. The abdomen is ßattened, the than broad; median ßagellomeres with eight bristles in cuticle of the tergites and sternites is wrinkled, and the basal whorl and 1Ð3 short setae distally; each of ßag- pleural membrane is extruded on the left side of seg- ellomeres 10Ð13 slightly longer than preceding one; ments IIÐV. The head, thorax, and abdomen are largely apical ßagellomere about 11 times longer than broad, denuded. The presence or absence of scales on the pointed. Thorax. Scutal integument dark brown; thorax and abdomen has been particularly difÞcult to darker areas not evident. Long rows of large setal determine, and any pruinose pattern that may have alveoli in acrostichal, dorsocentral, and supraalar ar- been present on the scutum or pleuron has been oblit- eas; additional large setal alveoli along lateral and erated during fossilization. One of the eggs is cracked posterior edges of scutal fossa. Left humeral area with and shriveled; both eggs are unsclerotized and visible three long, narrow, dark erect scales; no other scales only in lateral aspect. evident. Scutellum rounded; six large setal alveoli vis- ible along right half of posterior border, the lateral alveoli closer together; without smaller setae or alveoli Anopheles dominicanus Zavortink & Poinar, sp. n. dorsad of row of large alveoli; scales not evident. Pleu- (Figs. 1–5) ral integument dark brown. Left antepronotum with Diagnosis. A spotted-wing Anopheles, the wing pat- several setal alveoli and six long, narrow, dark erect tern most similar to that of subgenus Nyssorhynchus, scales above, at least Þve setae or alveoli below; pro- the following wing characters diagnostic alone or in episternum with one long seta; spiracular setae two; combination: accessory sector pale spot absent on prealar knob with several setal alveoli; upper katepis- veins C and Sc, present on vein R; veins Rs, R2ϩ3, and ternum with two setal alveoli; lower katepisternum R3 primarily dark scaled; vein R2 dark at apex; vein with one long and one shorter seta; upper R4ϩ5 with short median pale spot; vein M1 without mesepimeron with at least one long and one shorter pale scales beyond fork; vein CuA dark at base and its seta. Legs. Scales not evident on coxae. Integument of branches primarily dark scaled; basal pale fringe spot coxae and trochanters dark brown. Femur of foreleg absent. Flagellomeres uniformly brown, relatively not swollen basally. Foreleg partly denuded, appar- long and slender, ßagellomere 5 about six times longer ently dark scaled, apex of tibia possibly with narrow than broad, ßagellomere 13 about 11 times longer than pale ring. Tarsal claws of foreleg small, subequal, sim- broad. Antepronotum with long, narrow, erect scales. ple. Wing (Fig. 2). Both wings present; basal portion Pleuron with one strong proepisternal seta, two spi- of costa and radius of both partly to completely de- racular setae, and two lower katepisternal setae. Fe- nuded. Fringe missing from behind M1 to before A on mur of foreleg not swollen basally. Egg apparently right wing and from behind M2 to before A on left without deÞnite polygonal or blister-like outer chori- wing. Scales on costa in subbasal dark spot with six onic cells; ßoats dorsolateral, broad, Ϸ0.5 length of striations, those on vein R in subbasal dark spot with egg; frill deep, extending to ends of egg; lobed tuber- four striations. Costa with basal, humeral, sector, sub- cles apparently present. costal, and preapical pale spots; humeral pale spot 0.7 Female(Figs. 1Ð3). Length of scutum and scutel- length of subbasal dark spot; sector pale spot 0.2 length lum: 0.86 mm. Forefemur: 1.58 mm. Wing: 2.80 mm. of sector dark spot; subcostal pale spot 0.33 preapical Abdomen: 2.2 mm. Head. Flattened dorsoventrally, dark spot. Stem vein denuded, but probably bore dark the left compound eye extruded and ruptured; largely scales as vein is darkly pigmented. Vein R-R1 denuded denuded. Integument dark brown. Interocular space basally, with sector, accessory sector, subcostal, and apparently concave, its width equal to width of 3Ð4 preapical pale spots. Vein Rs dark scaled at base; with facets. Frontal tuft missing. Dorsal surface of head distinct basal spur with one scale. Vein Rs-R2ϩ3 with with one long, dark erect scale left of center on vertex, small pale spots at base of R4ϩ5, over fork into R2 and one long, slender, dark erect scale caudolaterally on R3, before apex on R2, and at apex of R3. Vein R4ϩ5 left side, and two long setae and one long, slender, dark without basal spur; with scales of dorsal surface erect scale caudolaterally on right side. Clypeus prom- spreading; small pale spots at base and apex and longer inent, bare. Proboscis and palpi extending to edge of pale spot basad of level of forks in R2ϩ3 and M. Stem amber, distal portions missing. Proboscis dark scaled, of vein M relatively densely scaled. Vein M with small scales decumbent; only one basal bristle evident. One pale spot at crossveins, over fork, and at apex of M2. mandibular and both maxillary stylets present, free of Vein CuA-CuA1 with scales of dorsal surface spread- labial groove, recurved; apex of mandibular stylet ob- ing; CuA with long pale spot on stem, shorter pale spot scured; apex of maxillary stylet with 10 or 11 teeth. at fork, small pale spots on CuA1 at crossvein m-cu and Pharynx not visible. Palpus dark scaled; palpomeres 1 at apices of CuA1 and CuA2. Vein A with small pale and 2 shaggy, the scales outstanding, palpomere 3 spots at base and apex and longer pale spot from basal largely denuded, the few scales remaining not as out- 0.25Ð0.55. Fringe with large pale spot associated with standing as those on palpomere 2; distal part of pal- preapical pale spot of costa and small pale spots at pomere 3 and palpomeres 4 and 5 missing. Both an- apices of veins R3,R4ϩ5,M2, and probably also CuA1 tennae present; pedicels brown; ßagellomeres and CuA2 (the costa is pale at the ends of CuA1 and uniformly brown; ßagellomere 1 about 1.4 length of CuA2); basal pale fringe spot absent. Alula bare. Upper ßagellomere 2, with numerous bristles and scales; ßag- calypter with marginal bristles. Halter. Flattened. In- ellomeres beyond one long and slender, apparently tegument of stem pale, of knob brown. Knob with 1232 ANNALS OF THE ENTOMOLOGICAL SOCIETY OF AMERICA Vol. 93, no. 6

Fig. 1. Anopheles dominicanus sp. n. female and eggs (arrows). numerous dark scales. Abdomen. Attached scales not Deck not visible, but obviously present, apparently evident; amber adjacent to abdomen with numerous narrow, judged to be Ϸ0.3 width of egg. Lobed tu- loose long setae and a few loose dark scales, one of bercles apparently present, anterior end with at least these (Fig. 3) to left of segment III particularly large, two, posterior end with at least one. Micropylar ap- spatulate, truncate. Integument of tergites brown. paratus not visible. Tergites I, VII, VIII with long setae at side. Few details Type Material. HOLOTYPE female, El Mamey For- of sternites discernable; sternites VI-VIII with long mation, northern mountain ranges, Dominican Re- setae. Genitalia. Postgenital lobe with two setae. Sper- public. Accession number D-7Ð6A deposited in the matheca not visible. Cerci large, ßattened; with long Poinar amber collection maintained at Oregon State setae; scales not evident. University, Corvallis, OR 97331. Egg. (Figs. 1, 4, and 5). Length 0.43 and 0.48 mm. Anterior end appearing truncate, probably actually Discussion blunt, posterior end upturned, tapered. Upper (mor- phologically ventral) surface slightly concave, lower The wings of An. dominicanus are relatively well surface more strongly convex. Egg slightly deeper preserved and provide the most convincing evidence anteriorly than posteriorly. Lateral surface of un- that the fossil should be placed in the subgenus Nys- cracked egg appearing slightly wrinkled and tubercu- sorhynchus. The presence of a preapical pale costal late, but deÞnite polygonal or blister-like outer cho- spot rather than an apical one argues for placement of rionic cells not evident. Floats present, one pair, the species in Nyssorhynchus rather than Cellia (Root dorsolateral, broad, Ϸ0.5 length of egg, extending closer to anterior end of egg than to posterior end, concave above. Float ribs curved or wavy, apparently Ϸ22Ð24. Upper surface with deep frill with columnar walls; frill extending from ßoats to both ends of egg.

Fig. 2. Anopheles dominicanus sp. n. wing. Abbreviations for wing spots: ASP, accessory sector pale; BP, basal pale; HP, humeral pale; PD, preapical dark; PP, preapical pale; SBD, subbasal dark; SCP, subcostal pale; SD, sector dark; SP, sector Fig. 3. Anopheles dominicanus sp. n. scale from caudola- pale. Scale ϭ 1. 0 mm. teral scale tuft. November 2000 ZAVORTINK AND POINAR:NEW Anopheles FROM DOMINICAN AMBER 1233

sessed caudolateral scale tufts in life. The occurrence of such scale tufts in An. dominicanus is consistent with its placement in Nyssorhynchus. The ßagellomeres of An. dominicanus are elongate, the Þfth about six times longer than broad and the last about 11 times longer than broad. We are not aware of any extant species of Anopheles with ßagellomeres as long. Although we are not absolutely certain, the egg of An. dominicanus appears to lack polygonal outer cho- rionic cells and appears to have lobed tubercles at both ends. We have compared the egg of An. dominicanus to published scanning electron micrographs of species of Nyssorhynchus and Kerteszia. Except for the pres- ence of lobed tubercles, the egg of An. dominicanus is Fig. 4. Anopheles dominicanus sp. n. egg. Anterior end at not too unlike that of some species in the subgenus left. Nyssorhynchus (as, for example, An. aquasalis Curry [Linley et al. 1993] and An. nuneztovari Gabaldon [Linley et al. 1996]). Lobed tubercles do not occur on 1926, Wilkerson and Peyton 1990), and the presence the eggs of species of Nyssorhynchus in the Albimanus of an accessory sector pale spot argues for placement and Argyritarsis sections (P. Lounibos, personal com- of the species in Nyssorhynchus rather than in subge- munication), but they are present in at least An. parvus nus Anopheles (Wilkerson and Peyton 1990). The (Chagas) of the Myzorhynchella section (Forattini et presence of a sector pale spot that is not joined to an al. 1998). Except for the absence of conspicuous po- accessory sector pale spot and the presence of basal, lygonal outer chorionic cells and the presence of a median, and apical pale spots on vein A argue for greater number of ßoat ribs, the egg of An. dominica- placement of the species in Nyssorhynchus rather than nus also resembles those of species of Kerteszia (Fo- in Lophopodomyia (Wilkerson and Peyton 1990). The rattini and Marucci 1993, Forattini et al. 1997). We termination of the subcostal vein at the proximal end believe that if the egg of An. dominicanus had poly- of the subcostal pale spot, the presence of pale scales gonal outer chorionic cells as conspicuous as those of at the apex of vein R4ϩ5 associated with an isolated pale fringe spot, and the presence of basal, median, species of Kerteszia, we would be able to discern them. and apical pale spots on vein A suggest placement of The subgenus Nyssorhynchus includes 29 extant spe- the species in Nyssorhynchus rather than in Kerteszia cies, distributed from southern Florida and southern (Wilkerson and Peyton 1990, Peyton et al. 1992). The Texas in the United States south through Central only signiÞcant departure of the wing pattern of An. America to Chile, Argentina, and Uruguay (Faran dominicanus from that of Nyssorhynchus is the dark- 1980, Linthicum 1988). Only three speciesÑAn. albi- scaled base of vein CuA. manus Wiedemann, An. aquasalis, and An. argyritarsis No attached scales are evident on the abdomen of Robineau-DesvoidyÑextend from the North or South An. dominicanus, but among the loose scales embed- American continents onto islands in the Caribbean, ded in the amber beside the abdomen is one partic- and only one of these, An. albimanus, occurs in the ularly large, dark, spatulate, and truncate scale (Fig. Greater Antilles, including the island of Hispaniola. 3). This is the kind of scale found in the tufts of The extant species of Nyssorhynchus are placed into outstanding scales that occur at the caudolateral cor- three sections, the Albimanus section, the Argyritarsis ners of the abdominal segments of some species of section, and the Myzorhynchella section. Anopheles subgenera Anopheles, Cellia, and Nyssorhynchus. We dominicanus appears to be closest to the Myzorhynch- take this scale as evidence that An. dominicanus pos- ella section. Anopheles ? rottensis Statz, 1944 from the upper Oligocene of Germany is the only other described fossil mosquito placed, at least tentatively, in the genus Anopheles. Relatively few characteristics of this spe- cies are known because the only specimen is pre- served in yellowbrown slate. This specimen, shown in a photograph accompanying the original description, does not have the habitus of a male anopheline mos- quito, but Statz (1944) states that there are no scales on the abdomen, and he thanks E. Martini for exam- ining the specimen and judging its systematic position. Statz describes the wing veins, particularly on the anterior edge of the wing, as having small, yellow- Fig. 5. Anopheles dominicanus sp. n. egg. Anterior end at brown scales, so whether the species is an anopheline left. Scale ϭ 0. 1 mm. or not, it is different from An. dominicanus. 1234 ANNALS OF THE ENTOMOLOGICAL SOCIETY OF AMERICA Vol. 93, no. 6

Capasso (1991) concluded that Anopheles arose in eles eggs display several specializations that, because the Old World, citing the absence of New World fossils they do not occur in any other mosquitoes, cannot be as evidence, and argued that the genus did not enter primitive characters. Although the discovery of An. the New World earlier than the late Tertiary. The dominicanus sheds no light on the phylogeny of discovery of An. dominicanus now provides evidence Anopheles, it does provide evidence that several of the that Anopheles was present in the New World in the specialized features of Anopheles eggsÑthe ßoats, frill, middle Tertiary. Quite in contrast to CapassoÕs view, and lobed tuberclesÑhad evolved by the middle Ter- mosquito systematists have considered the Neotropi- tiary. cal anopheline fauna to be old, citing its uniqueness as The adults of most species of Anopheles are crep- evidence of its antiquity. Christophers (1933) be- uscular or nocturnal and rest in protected sites during lieved that “there were already subgenus Anopheles- the day. The specimen described here may have en- like forms, as well as some earlier type from which countered the resin in which it became embedded Nyssorhynchus arose” present in South America before while seeking a resting site on a tree trunk. Bates its isolation from other continents, which was judged (1949) reported that in the forests near Villavicencio, at that time to be the middle Eocene on the basis of Colombia, adults of the mosquito Orthopodomyia fas- the history of mammals. Belkin (1962) considered the cipes (Coquillett) rest on tree trunks where their var- fact that the majority of the more generalized and iegated markings make them difÞcult to distinguish. annectant forms in the tribe Anophelini occur around He presumed this was a case of protective coloration. the American Mediterranean Region (the interconti- Although the markings of An. dominicanus differ nental area of the New World and adjacent portions greatly from those of Or. fascipes, the spotted wings of of the North and South American continents) as pos- the fossil species could also have made the mosquito sible evidence that the initial differentiation of the difÞcult to distinguish as it rested on tree trunks. The tribe took place in that region. More recently, Har- immatures of most species of Nyssorhynchus are found bach and Kitching (1998) believed the basal position in fresh ground waters exposed to the sun or partially of the Neotropical genus Chagasia in the anopheline shaded, but those of some species are found in brack- clade of their phylogenetic analysis suggested a pos- ish coastal waters or in deeply shaded sites. SpeciÞc sible New World origin for anopheline mosquitoes. habitats include lakes, ponds, swamps, ditches, seeps, Most systematists who have commented on the po- animal tracks, ßooded areas, and stream margins. Al- sition of Anopheles within the Culicidae have consid- though no species breeds in phytotelm habitats reg- ered it to be primitive. Howard et al. (1915) believed ularly, the immatures of An. albimanus have some- anophelines were the lowest of the true mosquitoes, times been found in tree holes. although they did consider some features of both the eggs and larvae to be specializations. Edwards (1923) stated: “Although no fossil Anopheles has yet been Acknowledgments found, there can be no doubt from its morphology that this is also an old genus, most probably older than any We are indebted to Bruce A. Harrison, Phil Lounibos, E. L. Culicine form. . . .” Ross (1951) stated that Anopheles Peyton, Maria Anice Mureb Sallum, and Richard C. Wilker- was little changed from the original ancestor of mos- son for sharing their knowledge of anopheline mosquitoes with us. We also thank Bruno Pfammatter for translating the quitoes and was a living fossil at least 70 million years description of Anopheles ? rottensis. old. Wood and Borkent (1989), in their phylogenetic analysis and classiÞcation of the Nematocera, stated that Anopheles was probably the most primitive mem- References Cited ber of the family Culicidae. The phylogenetic analysis of the Culicidae by Harbach and Kitching (1998) also Bates, M. 1949. The natural history of mosquitoes. 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