PROC. ENTOMOL. SOC. WASH. 113(4), 2011, pp. 497–507

USING GENITALIA CHARACTERS AND MITOCHONDRIAL COI SEQUENCES TO PLACE “LEUCOCHLAENA” HIPPARIS (DRUCE) IN SPODOPTERA GUENE´ E (: )

MICHAEL G. POGUE

Systematic Entomology Laboratory, PSI, Agricultural Research Service, U.S. Department of Agriculture, c/o Smithsonian Institution, P.O. Box 37012, NMNH, MRC-168, Washington, DC 20013-7012, U.S.A. (e-mail: [email protected])

Abstract.—The species “Leucochlaena” hipparis (Druce) belongs in the Spodoptera based on morphological characters of the male and female genitalia, as well as maximum parsimony and maximum likelihood analysis of the 658 bp “bar- code” region of cytochrome oxidae (COI). Adult and male and female genitalia are illustrated. Cladograms are included to illustrate the placement of “Leucochlaena” hipparis within Spodoptera. Key Words: DNA barcode region, morphology, DOI: 10.4289/0013-8797.113.4.497

The DNA barcode is a 658 base pair (Lafontaine and Poole 2010). No phy- region of mitochondrial DNA (COI) that logenetic analysis was done. The bar- has been used to find cryptic species coding region has been sequenced for within what was thought to be a single specimens of “Leucochlaena” hipparis species (Burns and Janzen 2001, 2005; (Druce) and several species of Spodoptera Hebert et al. 2004; Hajibabaei et al. 2006; Guene´e, but the potential placement Smith et al. 2006; Burns et al. 2007, of “L.” hipparis in Spodoptera has not 2008). Other applications of DNA bar- been assessed through analysis of those codes have been used in pest identification data. (Ball and Armstrong 2006) and associ- Druce (1889) originally described ation of larvae with adults. The generic S. hipparis in the genus Heliophobus classification of the New World Acon- Boisduval, 1829. Hampson (1906) de- tiinae (Lepidoptera: Noctuidae) was re- scribed the genus Leucochlaena that in- cently revised comparing male genitalic cluded five species, one of which was characters with COI barcodes. The COI S. hipparis. Hampson (1906) probably barcodes were analyzed using phenetic included S. hipparis in Leucochlaena clustering and produced a neighbor-joining because of the widely bipectinate male tree that was similar to the generic con- antenna and the forewing pattern that is cept of the male genitalic morphology similar to other species of Leucochlaena, but S. hipparis shows no structural affini- ties with Leucochlaena. Leucochlaena * Edited by John W. Brown; accepted by Robert currently includes 10 species, all from the R. Kula Old World (Poole 1989). 498 PROCEEDINGS OF THE ENTOMOLOGICAL SOCIETY OF WASHINGTON

Todd and Poole (1980) correctly rec- (Hall 1999). Phylogenetic relationships ognized that S. hipparis did indeed be- were constructed under the maximum long in Spodoptera and that it shared the parsimony (MP) (Figs. 6–7) and maxi- bipectinate male antenna with several mum likelihood (ML) (Fig. 8) methods other species of Spodoptera, including implemented in PAUP 4.0b10 (Swofford the Galapagos Islands endemic Spo- 1999). Outgroup taxa were chosen based doptera roseae Schaus. All other spe- on the current understanding of Spodoptera cies of Spodoptera in which the male phylogenetic relationships (Pogue 2002). has bipectinate antennae are Old World Maximum parsimony analysis with equally (Pogue 2002). Pogue (2002) did not in- weighted characters was computed via a clude S. hipparis in Spodoptera based on full heuristic search using tree bisection- the lack of a tuft of fine hairlike scales on reconnection (TBR) branch swapping with the female 8th abdominal segment and the random taxon addition replicates (mini- unusual male genitalia. mally 1000). If analyses produced more than one most parsimonious tree, a strict MATERIALS AND METHODS consensus was performed. Clade support Images of adult and genitalia was assessed using the non-parametric were taken with a Visionary Digital Im- bootstrap (Felsenstein 1985) under the aging System using a Nikon D1X camera same search conditions described above and modified K2 long-distance lens and for MP and ML. a pulsed xenon flash. Genitalia dissec- For the ML phylogenetic analysis, nu- tions were prepared following Pogue cleotide substitution model parame- (2002) except specimens were mounted ters were inferred using MrModeltest v in Euparal and stained with mercuro- 2.3 (Nylander 2004). The Akaike In- chrome. Terms used in the descriptions formation Criterion (AIC) was used to of male genitalia follow Forbes (1954) select the best-fit nucleotide substitu- and those for the female follow Klots tion model (Posada and Buckley 2004). (1970). Forewing length was measured Maximum likelihood analysis was per- using a calibrated ocular micrometer formed using the general time reversible from the juncture of the thorax to the model with a proportion of sites invariant apex, including the fringe. and gamma distributed rates (GTR+I+G; Barcode sequences from the Barcode of nst=6, rates=invgamma). Maximum like- Life Data Systems (BOLD) (Ratnasingham lihood analysis was conducted by full heu- and Hebert 2010) (Table 1) were used ristic search using TBR branch swapping to produce the results in Figs. 6–8. For with five random taxon addition replicates Lepidoptera, it is standard to submit a and starting tree obtained by stepwise leg to BOLD for DNA barcoding analysis. addition. The two specimens of S. hipparis used in this study have the following locality data: RESULTS AND DISCUSSION New Mexico, Valencia Co., Tome, 31 May Although Todd and Poole (1980) 2006, elevation: 1493.54 m, R.W. Holland. stated that S. hipparis and Spodoptera Sequences are deposited with BOLD exigua (Hu¨bner) lacked the large scale (http://www.boldsystems.org). tuft associated with the 8th abdominal Since there was an absence of indels segment in the female, I have found that in the cytochrome oxidase I (COI) se- both of these species have this character quences analyzed, sequence data were state. This scale tuft is present in all spe- aligned by eye using the program BioEdit cies of Spodoptera and was overlooked VOLUME 113, NUMBER 4 499

Table 1. DNA samples examined.

Barcode ID Sample ID Species Phylogeny ID County Locality State/Province Region/County

LNCNW065-06 06-NCNW-0065 grata E_grata United States Alleghany Co. LNCB525-07 07-NCNW-0209 partita G_partita United States North Carolina Ashe Co. RDNMD637-06 CNCNoctuoidea12969 Leucochlaena L_hipparis 1 United States New Mexico Valencia Co. hipparis RDNMD638-06 CNCNoctuoidea12970 Leucochlaena L_hipparis 2 United States New Mexico Valencia Co. hipparis LOFLB202-06 06-FLOR-1142 Spodoptera S_albula 1 United States Highlands Co. albula LOFLB437-06 06-FLOR-1377 Spodoptera S_albula 2 United States Florida Highlands Co albula BLPDC921-09 08-SRNP-107876 Spodoptera S_androgea 1 Costa Rica Alajuela Area de Conservacion androgea Guanacaste BLPDD477-09 08-SRNP-108372 Spodoptera S_androgea 2 Costa Rica Guanacaste Area de Conservacion androgea Guanacaste BLPDD478-09 08-SRNP-108373 Spodoptera S_androgea 3 Costa Rica Guanacaste Area de Conservacion androgea Guanacaste BLPDD685-09 08-SRNP-108580 Spodoptera S_androgea 4 Costa Rica Guanacaste Area de Conservacion androgea Guanacaste BLPDD686-09 08-SRNP-108581 Spodoptera S_androgea 5 Costa Rica Guanacaste Area de Conservacion androgea Guanacaste BLPAA820-06 06-SRNP-100819 Spodoptera S_dolichos 1 Costa Rica Alajuela Area de Conservacion dolichos Guanacaste BLPAA926-06 06-SRNP-100925 Spodoptera S_dolichos 2 Costa Rica Alajuela Area de Conservacion dolichos Guanacaste BLPAA927-06 06-SRNP-100926 Spodoptera S_dolichos 3 Costa Rica Alajuela Area de Conservacion dolichos Guanacaste BLPAB037-06 06-SRNP-100976 Spodoptera S_dolichos 4 Costa Rica Alajuela Area de Conservacion dolichos Guanacaste BLPAB238-06 06-SRNP-101177 Spodoptera S_dolichos 5 Costa Rica Alajuela Area de Conservacion dolichos Guanacaste BLPCK258-08 08-SRNP-100351 Spodoptera S_eridania 1 Costa Rica Alajuela Area de Conservacion eridania Guanacaste BLPDA469-09 08-SRNP-105544 Spodoptera S_eridania 2 Costa Rica Alajuela Area de Conservacion eridania Guanacaste LOFLA396-06 06-FLOR-0396 Spodoptera S_exigua 1 United States Florida Putnam Co. exigua LOFLB270-06 06-FLOR-1210 Spodoptera S_exigua 2 United States Florida Highlands Co exigua LOFLC241-06 06-FLOR-2121 Spodoptera S_exigua 3 United States Florida Highlands Co exigua LOFLC242-06 06-FLOR-2122 Spodoptera S_exigua 4 United States Florida Highlands Co exigua RDNMD436-06 CNCNoctuoidea12768 Spodoptera S_exigua 5 United States California San Diego Co exigua BLPDE131-09 08-SRNP-108966 Spodoptera S_frugiperda 1 Costa Rica Guanacaste Area de Conservacion frugiperda Guanacaste BLPAB120-06 06-SRNP-101059 Spodoptera S_frugiperda 2 Costa Rica Alajuela Area de Conservacion frugiperda Guanacaste BLPAG434-07 06-SRNP-106073 Spodoptera S_frugiperda 3 Costa Rica Guanacaste Area de Conservacion frugiperda Guanacaste BLPBA709-07 06-SRNP-107852 Spodoptera S_frugiperda 4 Costa Rica Guanacaste Area de Conservacion frugiperda Guanacaste BLPCC401-08 07-SRNP-106886 Spodoptera S_frugiperda 5 Costa Rica Guanacaste Area de Conservacion frugiperda Guanacaste BLPCA212-08 07-SRNP-104817 Spodoptera S_albula 6 Costa Rica Guanacaste Area de Conservacion Janzen187 Guanacaste 500 PROCEEDINGS OF THE ENTOMOLOGICAL SOCIETY OF WASHINGTON

Table 1. Continued.

Barcode ID Sample ID Species Phylogeny ID County Locality State/Province Region/County

BLPCA843-08 07-SRNP-105448 Spodoptera S_albula 5 Costa Rica Guanacaste Area de Conservacion Janzen187 Guanacaste BLPBB657-07 06-SRNP-108740 Spodoptera S_frugiperda 6 Costa Rica Guanacaste Area de Conservacion Janzen24 Guanacaste BLPAA382-06 06-SRNP-100381 Spodoptera S_latifascia 1 Costa Rica Guanacaste Area de Conservacion latifascia Guanacaste BLPAA386-06 06-SRNP-100385 Spodoptera S_latifascia 2 Costa Rica Guanacaste Area de Conservacion latifascia Guanacaste BLPDE113-09 08-SRNP-108948 Spodoptera S_latifascia 3 Costa Rica Guanacaste Area de Conservacion latifascia Guanacaste BLPDE114-09 08-SRNP-108949 Spodoptera S_latifascia 4 Costa Rica Guanacaste Area de Conservacion latifascia Guanacaste BLPDE115-09 08-SRNP-108950 Spodoptera S_latifascia 5 Costa Rica Guanacaste Area de Conservacion latifascia Guanacaste MJMSL004-10 10MA-0004 Spodoptera S_ornithogalli United States Barnstable Co. ornithogalli RDNMF642-08 NOC14728 Spodoptera S_praefica 1 United States California Modoc Co. praefica RDNMF644-08 NOC14730 Spodoptera S_praefica 2 Canada Alberta Lethbridge praefica RDNMF645-08 NOC14731 Spodoptera S_praefica 3 United States California Plumas Co. praefica LOCRE458-10 10-CRBS-983 Spodoptera S_albula 3 Costa Rica San Jose Puriscal sunia LOCRE491-10 10-CRBS-1016 Spodoptera S_albula 4 Costa Rica San Jose Puriscal sunia in S. hipparis by Pogue (2002). In ad- K = 10; AIC = 4578.2075) model with dition to this scale tuft, all species of estimated base frequencies as follows: Spodoptera, including S. hipparis,have A = 0.2902, C = 0.1394, G = 0.1527, and in the male genitalia a basal sclerite that T = 0.4177. The rate matrix parameters extends from the apex of the sacculus estimated were as follows: R(a) [A-C] = and attaches to the base of the clasper 2390268.75, R(b) [A-G] = 2581590.50, proper (Fig. 3). R(c) [A-T] = 3177736.00, R(d) [C-G] = Data were generated from 658 nucle- 0.00, R(e) [C-T] = 13365338.00, and otides of COI spanning mitochondrial R(f) [G-T] = 1.00. The proportion of positions 1516 to 2173 following the invariable sites (PINVAR) was 0.6085, numbering scheme of the Drosophila and the alpha shape parameter (a)ofthe yakuba Burla genome sequence (Clary gamma (G) distribution was 1.1684. The and Wolstenholme 1985). The number treeshowninFig.8wasproducedusing of parsimony informative sites was 121. the ML method under the GTR+I+G Parsimony analysis using PAUP found substitution model. two equally parsimonious trees (length Both the MP (Figs. 6–7) and ML (Fig. 284 steps). The consistency index (CI) 8) methods produced similar results and was 0.662, and the retention index (RI) support the inclusion of “Leucochlaena” was 0.877. One of these trees is shown in hipparis in Spodoptera. Fig. 6. A strict consensus tree of these The amount of sequence divergence two trees is shown in Fig. 7. between S. hipparis and its nearest neigh- MrModeltest (Nylander 2004) se- bor, S. exigua, is 9.656 – 9.688%. The lected the GTR+I+G (-lnL = 2279.1040; amount of sequence divergence between VOLUME 113, NUMBER 4 501

S. hipparis and the outgroup Elaphria Spodoptera hipparis; Todd and Poole 1980: grata Hu¨bner is 18.231% and the distance 733. between the outgroup Diagnosis.—The male of this species Guene´e is 17.247%. is easily recognized by the bipectinate Given the presence of a scale tuft and antennae with extremely long setae (Fig. 1). homologous basal sclerite, as well as The only other New World Spodoptera results of the phylogenetic analyses, “Leu- species with male bipectinate antennae cochlaena” hipparis is here considered is S. roseae; however, in S. hipparis the a species of Spodoptera as Todd and setae are more than 33 as long as in S. Poole (1980) suggested. roseae. Both sexes can be distinguished from all other species of Spodoptera by Spodoptera hipparis (Druce) the light brown hind wings in S. hipparis, (Figs. 1–5) whereas all other Spodoptera have white Heliophobus hipparis Druce 1889: 272. hind wings. The male genitalia of S. hip- Leucochlaena hipparis form pallens Draudt paris are also distinct with the lack of 1924: 178. coremata; all other species of Spodoptera Leucochlaena hipparis form colossa Draudt have coremata. The uncus has a unique 1924: 178. shape, being very deep with a broad apex.

Figs. 1–5. Adults and genitalia of S. hipparis. 1, Male; 2, Female; 3, Male genital capsule; 4, Male aedeagus; 5, Female genitalia. Abbreviations: amp = ampulla, bs = basal sclerite, clp = clasper proper, sac = sacculus. 502 PROCEEDINGS OF THE ENTOMOLOGICAL SOCIETY OF WASHINGTON

Fig. 6. Phylogenetic relationships inferred through maximum parsimony analysis (length = 284; CI = 0.662, RI = 0.877). The numbers above branches represent non-parametric bootstrap support.

The dorsobasal sclerotized patch at the in S. exigua the ventral plate is a large base of the vesica where it emerges from quadrate plate. The signum is absent in the aedeagus is a thin, granulose ribbon S. hipparis, whereas it is an elongate, with a row of minute spines along its spinulate sclerotized ribbon in S. exigua. dorsal edge. A large spinelike cornutus Redescription.—Male. Head: Labial emerges from the apex of the vesica. palpus brown with a ventral fringe of The ventral plate of the ostium bursae in long, hairlike scales. Bulbous part of frons the female genitalia of S. hipparis is nar- with short cream-colored to brown scales; row,withalength53 the height, whereas remainder of head and vertex covered VOLUME 113, NUMBER 4 503

Fig. 7. Strict consensus of two trees generated through maximum parsimony analysis (length = 284; CI = 0.662, RI = 0.877). The numbers above branches represent non-parametric bootstrap support. with long, thin, light brown to pale gray ish brown; tibia with ventral tuft of long scales tipped with white giving bushy hairlike scales, with medial and apical appearance to head. Antenna bipectinate; band of cream-colored scales, epiphysis scape with short, flat cream-colored scales almost as long as tibia, tarsi gray with and thin, long brown scales along dorsal cream-colored apical bands; middle leg margin. Thorax: Covered with narrow, with gray femur, tibia cream colored with brown to pale gray scales. Foreleg gray- ventral tuft of long hairlike scales, tarsi 504 PROCEEDINGS OF THE ENTOMOLOGICAL SOCIETY OF WASHINGTON

Fig. 8. Phylogenetic relationships inferred through maximum likelihood analysis under the GTR+G+I substitution model. The numbers above branches represent non-parametric bootstrap support. gray with cream-colored apical bands; wing; antemedial line black edged with hind leg cream colored with ventral tuft of white proximally; claviform spot black long hairlike scales, tarsi gray with cream- and contiguous with antemedial line; or- colored apical bands that are broader than bicular spot white to pale rufous with on middle tarsi. Forewing (Fig. 1): Length rufous center, outlined proximally and 9.6–13.7 mm; ground color pale gray to distally with black; reniform spot less brown; basal line indistinct, a series of conspicuous than orbicular, black, some- short black dashes just distad to base of times with gray scales, outlined in white VOLUME 113, NUMBER 4 505 then black proximally and distally; small Papillae anales trapezoidal with ventral ovate spot with rufous center either out- apex slightly produced, long setae along lined in black or white between orbicular dorso-ventral margin; base of apophyses and reniform spots below M vein; post- anteriores wide, tear-drop shaped; apoph- medial line broadly crenulate, white to yses posteriors short; ventral plate of light brown bordered with black; sub- ostium bursae small, rectangular, length terminal line sinuate, white to light brown 53 height; ductus bursae short, mem- bordered proximally by varying amount branous; appendix bursae adjacent to of scattered black scales, more obscure ductus bursae and not differentiated from than postmedial line; terminal line a se- corpus bursae; corpus bursae elongate, ries of black, somewhat triangular shaped slightly constricted medially; signum dots between veins; fringe gray to light absent. brown interspersed with black scales. Type material.—Heliophobus hipparis Hind wing: Light brown, faint discal spot Druce. The syntypes are in The Natural present; fringe cream colored to light History Museum, London (BMNH). Type brown. Abdomen: Cream colored vari- localities: Mexico: Districto Federal, ously speckled with brown and gray Mexico City; Durango, Milpas, 5900 ft.; scales; genital tuft cream colored con- Veracruz, Jalapa. sisting of long, hairlike scales. Genitlalia Material examined (all from USNM (Figs. 3–4): Uncus curved, deep, apex unless otherwise noted).—MEXICO: broad with round apex and minute medial AGUASCALIENTE. Agauscaliente, tooth; scaphium a wide tube, sclerotized 3-5 Aug. 1963 (4m), P.J. Spangler. ventrally and membranous dorsally; tegu- DISTRICTO FEDERAL. Lomas de men lacking pair of projecting arms on Chapultepec, 7 Apr. 1939 (1m), 9 Apr. upper third; costa of valve a sclerotized, 1939 (1m), 17 May (1f), 27 May 1941 setose, tubular shape extending 0.453 (1m), 17 June 1939 (1m), 19 June 1940 length of valve, ending just prior to (1f), 8 July 1940 (1m), 13 July 1940 clasper; costal process absent; basal scler- (2m),21July1939(1m),23–24July ite well developed, rodlike, extending 1939 (2m), 5–6 Aug. 1939 (4m), 19 from apex of sacculus to base of ampulla- Aug. 1939 (1f); San Jacinto, 8 Oct. 1931 clasper proper; ampulla setose, swollen, (1m); Mexico City, (1m), R. Muller. dorso-basal lobe of clasper proper; clasper DURANGO. 10 W El Salto, collected proper a rounded plate, perpendicular to between 9 June–8 Aug. 1964 (;400m, valve; sacculus sclerotized, 0.453 length 5f), J.E.H. Martin & W.C. McGuffin, of valve, apex round; clavus absent; juxta 21 Apr. 1961 (1m), Howden & Martin an ovate plate; vinculum U-shaped; ae- (CNC). MEXICO. La Marquesa, 5-9 July deagus slightly curved, apex with fish- 1965 (23m , 1f), 13 July 1965 (14m, 1f), scalelike plates that extend onto base of 17 July 1965 (4m), 20 July 1965 (1m), vesica giving granular appearance; vesica Flint & Ortiz; Chapingo, 12 Mar. 1960 straight tube, curved ventrally, with lat- (2m), 15 Mar. 1960 (1m), H. Velasco; eral scobinate band from base; apical Toluco, 8 July 1969 (1f), D. Kritch cornutus a large, teardrop-shaped spine (CNC). MICHOACAN. P.N. Morelos, with pointed apex. nr. Morelia, 14 July 1966 (1f), Flint & Female (Fig. 2). Similar to male ex- Ortiz. VERACRUZ. La Joya, Rt. 140, cept antenna filiform. Forewing: Length Km.307,21July1965(2m),Flint& 9.9–11.8 mm. Abdomen: Fine dense scales Ortiz; Veracruz, (2m). U.S.A.: ARIZONA. on 8th segment white. Genitalia (Fig. 5): Apache Co., White Mts., near McNary 506 PROCEEDINGS OF THE ENTOMOLOGICAL SOCIETY OF WASHINGTON

P.O., 1–15 Sep. 1923 (1m), O.C. Poling. Literature Cited Cochise Co., Sierra Vista, 24 Apr. 1966 Ball,S.L.andK.F.Armstrong.2006.DNA (3m), R.F. Sternitsky (CNC). Graham barcodes for pest identification: a test Co., White Mts., San Carlos Indian Res- case with the tussock moths (Lepidoptera: ervation, 15–30 June 1925 (1m), O.C. Lymantriidae). Canadian Journal of Forest Poling. Navajo Co., White Mts., near Research 36: 337–350. Boisduval, J. B. A. D. de 1829. Europaeorum Rice, 7000 ft., 1–15 July 1925 (1m), O.C. Lepidopterorum Index Methodicus. De Plassan, Poling. NEW MEXICO. Burnalillo Co., Paris. 103 pp. Cedar Crest, 7500 ft., 11 Oct. 1976 (1m) Burns, J. M. and D. H. Janzen. 2001. Biodiversity of (CNC). Valencia Co., Tome, 4900 ft., 31 pyrrhopygine skipper butterflies (Hesperiidae) May 2006, R.W. Holland (CNC). intheAreadeConservacio´n Guanacaste, Costa Rica. Journal of the Lepidopterists’ Distribution.—Collections in the U.S. Society 55: 15–43. are from eastern Arizona and central Burns, J. M. and D. H. Janzen. 2005. Pan- New Mexico, and in Mexico they are neotropical genus Venada (Hesperiidae: Pyr- from Aguascalientes, Districto Federal, ginae) is not monotypic: Four new species Durango, Michoacan, and Veracruz. occur on one volcano in the Area de Con- servacio´n Guanacaste, Costa Rica. Journal of Discussion.—The overall coloration of the Lepidopterists’ Society 59: 19–34. the thorax and forewings varies from pale Burns, J. M., D. H. Janzen, M. Hajibabaei, gray to brown. The females tend to have W. Hallwachs, and P. D. N. Hebert. 2007. a darker grayish brown forewing, and the DNA barcodes of closely related (but mor- pattern is not as distinct as in the male. phologically and ecologically distinct) spe- cies of skipper butterflies (Hesperiidae) can differ by only one to three nucleotides. ACKNOWLEDGMENTS Journal of the Lepidopterists’ Society 61: I thank Don Lafontaine, Canadian Na- 138–153. tional Collection, Ottawa, Ontario, Canada Burns,J.M.,D.H.Janzen,M.Hajibabaei, W. Hallwachs, and P. D. N. Hebert. 2008. DNA for showing me the DNA barcoding re- barcodes and cryptic species of skipper but- sults of specimens in the CNC and sug- terflies in the genus Perichares in Area de gesting that I write this paper. He also Conservacio´n Guanacaste, Costa Rica. Pro- sent me locality data from the CNC. Paul ceedings of the National Academy of Sci- ences of the United States of America 105: Hebert and his staff at the Canadian 6350–6355. Centre for DNA Barcoding, Biodiversity Clary, D. O. and D. R. Wolstenholme. 1985. The Institute of Ontario, University of Guelph, mitochondrial DNA molecule of Drosophila Guelph, Canada, provided the data and yakuba. Nucleotide sequence, gene organiza- information from the Barcode of Life tion and geneticcode. Journal of Molecular Evolution 22: 252–271. Data (BOLD) system. Gary Ouellette of Draudt, M. 1925. 2. Family: Noctuidae [part], the USDA, Systematic Entomology Lab, pp. 173–180. In A. Seitz, ed. The Macrolepi- produced the phylogenetic analysis. doptera of the World. 7. Volume: Noctuiformes. Charyn Micheli (formerly USDA Sys- Alfred Kernen, Stuttgart. 412 pp., 64 plates. tematic Entomology Lab [SEL], currently [Signature 23, published February 19, 1925]. Druce, H. 1889. Lepidoptera-Heterocera, pp. Smithsonian Institution Department of 265–272. In F. D. Godman and O. Salvin, Entomology) prepared the illustrations. eds. Biologia Centrali-Americana. Insecta. Don Lafontaine, as well as Thomas Lepidoptera-Heterocera. Vol. 1. Taylor and J. Henry and Matthew L. Buffington of Francis, London. 490 pp. [Signature 2 ll, published May 1889]. SEL, reviewed the manuscript and gave Felsenstein, J. 1985. Confidence limits on phy- helpful suggestions. USDA is an equal logenies: an approach using the bootstrap. opportunity provider and employer. Evolution 39: 783–791. VOLUME 113, NUMBER 4 507

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