Taxonomy and biology of two seed-parasitic gracillariid moths (Lepidoptera, Gracillariidae), with description of a new species

Abstract A new species and new record of gracillariid moths from China are reported: Conopomorpha flueggella Li, sp. n. and Epicephala relictella Kuznetzov, 1979. Specimens were collected on flowers or leaves of Flueggea suffruticosa (Pall.) Baill. (Euphorbiaceae) at night, and reared from fruits in captivity. Larvae of both species feed on the seeds of Flueggea suffruticosa, but they can be differentiated externally by the position of the red pattern on the thorax and abdomen. Morphology of the eggs, larvae, pupae and the life history of the two species are described and compared. Images of the life history and figures of the genital structures are provided.


Introduction
Most species of Gracillariidae are leaf-miners, although some are stem-, fruit-or peel-miners or feed on fl ower buds (Fletcher 1920, Huang et al. 1997Vargas and Landry 2005, Grafton-Cardwell et al. 2008, Kawahara et al. 2009). An obligate pollination mutualism exists between Epicephala moths and Euphorbiaceae (or Phyllanthaceae) trees. Th e seedparasitic habit of Epicephala, as in the pollination mutualisms of fi g wasps, yucca moths and senita moths, is unique in Gracillariidae (Riley 1892, Fleming and Holland 1998, Bai and Li 2008, Weiblen 2002, Kato et al. 2003, Pellmyr 2003, Kawakita and Kato 2004aand b, Kawakita 2010. Currently, the biology and hostplants of most Epicephala species remain unknown according to our recent study. Previous studies in the Euphorbiaceae-Epicephala mutualism have revealed a high degree of specifi city between pollinating moths and plants, although the relationships between hostplants and insects are not always an exact one-to-one relationship (Kawakita and Kato 2006). We found a high degree of diversity in the biology of adults and larvae, which enables us to understand more about the obligate pollination mutualism between the Epicephala moths and the Euphorbiaceae trees.
In this paper, we describe the morphology and biology of adults and larvae of Conopomorpha fl ueggella Li, sp. n. and Epicephala relictella Kuznetzov, 1979. Both species are seed-parasites of Flueggea suff ruticosa in the Baxian Mountain State Nature Reserves in Tianjin, China. Th e hostplant of E. relictella is here recorded for the fi rst time, and the biology of the two gracillariid species is described and compared. Images of the adults and genitalia are provided.

Material and methods
Field studies were conducted from 2007 to 2009 in the Baxian Mountain State Nature Reserves (40°7'24"-40°13'53"N, 117°30'35"-117°36'24"E) in Tianjin, China (Fig. 1), at an altitude ranging from 500 to 800 m. Th e area covers 5360 hm 2 , with 1583 hm 2 as the core region. It is characteristic of warm temperate deciduous broad-leaved forest, and belongs to the warm and humid continental monsoon climate. Th e annual average rainfall amounts to 968.5 mm, and the annual average temperature is 8-10 °C (Li et al. 2009).
Flueggea suff ruticosa (Pall.) Baill. (Fig. 3) occurs in scrubby slopes, forest margins and at road sides ( Fig. 2) at an altitude of 500 to 2500 m. It is distributed in China (except in Gansu, Qinghai, Xinjiang and Tibet), Japan, Korea, Mongolia and Russia (Li 1994). Flueggea suff ruticosa (Pall.) Baill. forms typically 1-3 m tall shrubs, is dioecious, and the infl orescences are axillary and cymose. Th e male fl owers have 3-18 clusters, 5 sepals, free fi laments, and 5 stamens. Th e female fl owers have 3 styles, erect to spreading horizontally, free or connate at base, and bifi d; the ovary is 3-celled, each cell having 2 ovules. Th e fruit is an oblate capsule, reddish brown when ripe (Fig. 4). Th e fl owering period lasts from May to August, and the fruiting period from June to November in Baxian Mountain.
Th e biology of Conopomorpha fl ueggella Li, sp. n. and Epicephala relictella Kuznetzov was observed and studied during August-October 2007 and May-October of 2008 and 2009. Life history observations were made during fl owering and fruiting seasons. Th e developing and mature fruits were collected from diff erent individuals and dissected to examine the feeding habit with a light microscope. In addition, the developing fruits were collected in a cylindrical box (10 cm × 10 cm 2 ) to rear mature larvae and braconid wasps, and the behaviors of the mature larvae were observed.
Specimens examined in this study were collected on fl owers or leaves of Flueggea suff ruticosa at night, and reared from fruits in captivity, and a few specimens were collected by using light traps. Genitalia dissection and mounting follow Li and Zheng (1996). Photographs of F. suff ruticosa and moths were taken primarily in the fi eld using Canon G10 and Canon S3 IS digital cameras. Photographs of adult specimens were taken with a Nikon D300 digital camera. Dissections of genitalia were conducted under an Olympus SZ11 stereo zoom microscope. Figures of genitalia were prepared using an Olympus C-7070 digital camera attached to an Olympus BX51 microscope.
Th e type specimens are deposited in the Insect Collection, College of Life Sciences, Nankai University, Tianjin, China.

Taxonomic history
Meyrick (1885) established the genus Conopomorpha based on the type species C. cyanospila Meyrick, 1885. Th e wing pattern suggests it may be closely related to Epicephala Meyrick, 1880, but the complete separation of the sacculus and costa as well as the simple ovipositor distinguishes Conopomorpha from Epicephala. Conopomorpha currently consists of 13 species worldwide: eight species in the Australian Region, three in the Oriental and Afrotropical regions respectively, and one in the Palearctic Region De Prins 2005, 2011). Prior to this study, three species, C. litchiella Bradley, 1986, C. sinensis Bradley, 1986and C. cramerella (Snellen, 1904, were recorded in China. Th ey are important fruit pests on litchi, longan and cacao in Fujian, Guangdong, Hainan, Hong Kong, Taiwan (Hwang et al. 1989, Huang et al. 1997, Robinson et al. 2001, Shapiro et al. 2008).
Meyrick (1880) described Epicephala based on the type species E. colymbetella Meyrick, 1880. Species of this genus are extremely similar and diffi cult to distinguish. Th ey usually have a fi ne or indistinct, curved, transverse silvery-metallic line before the apical area and a small round black dot in the apical area. Th e general structure is close to the genus Caloptilia Hübner, 1825, diff ering from it in the venation and in the peculiar resting posture: the Epicephala adult rests with its head appressed horizontally, the hind-part raises considerably and is seemingly supported by the hind legs, the fore and mid legs extended laterally and appressed to the horizontal surface (Meyrick 1880); the hind tibiae are bristly above, which also distinguishes the genus from Caloptilia. Th e wing pattern of Epicephala shows some resemblance to the genus Stomphastis Meyrick, 1912, but from this Epicephala can be separated by the wing venation and the very peculiar shape of the apo-and antapophyses in the female genitalia (Vári 1961).
Epicephala includes 40 described species: 18 in the Oriental Region, 15 in the Australian Region, six in the Afrotropical Region, and one in the Palearctic Region (Russian Far East). Two species were recorded to occur in China prior to this study: E. venenata Meyrick, 1935 and E. albifrons (Stainton, 1859) De Prins 2005, Kendrick 2005). Epicephala venenata occurs only in Taiwan, and the hostplant is unknown. Epicephala albifrons is widely distributed in Hong Kong, India, Indonesia, Sri Lanka, Th ailand and Vietnam. Th e larvae of E. albifrons are known to feed on Phyllanthus niruri Linn. (Euphorbiaceae).
In 2007, we discovered Conopomorpha fl ueggella Li, sp. n. and Epicephala relictella Kuznetzov, 1979 in the Baxian Mountain State Nature Reserves in Tianjin, China, whose larvae feed on Flueggea suff ruticosa (Pall.) Baill. (Euphorbiaceae). Epicephala relictella is the only species of the genus distributed in the Palearctic Region, and is newly recorded for China. Its hostplant and biology were unknown previously. Diagnosis. Th is species is similar to Conopomorpha litchiella, but distinguishable by the uniformly greyish brown to dark brown forewing with three pairs of stripes (more conspicuous when moths alive); the valva without protuberance on ventral margin distally and the saccus long linguiform in the male genitalia; the corpus bursae shorter than twice the length of the ductus bursae in the female genitalia; and the larva red-coloured. In C. litchiella, the forewing is whitish yellow in distal portion; the valva has one large and one small protuberance on ventral margin distally, and the saccus is very short and small; the corpus bursae is twice as long as the ductus bursae; and the larva is yellowish green.

Conopomorpha fl ueggella
Description. Adult (Figs 5-6). Wing expanse 8.0-15.5 mm. Head grey to greyish brown, frons greyish white. Compound eye dark brown. Labial palpus white, second segment with outer surface and distal tuft of ventral surface fuscous, third segment porrect or obliquely upward. Maxillary palpus greyish brown to dark brown. Antenna with scape greyish brown, fl agellum brown to dark brown ringed with greyish white basally. Th orax and tegula dark brown. Forewing narrow, costal and dorsal margins nearly parallel; ground color greyish brown to dark brown; costal and dorsal margins with three oblique greyish white stripes respectively, fi rst costal stripe from near middle extending obliquely to end of cell; dorsal margin with black speck at basal 1/3; bluish grey fascia with metallic refl ection extending from near costal 5/6 to dorsum and along termen, respectively, between them set a large black spot; cilia pale greyish brown except fuscous apically. Hindwing and cilia greyish brown. Fore and mid legs brown; hind leg greyish white, distal half of tibia dark fuscous on outer surface. Abdomen grey, with fi rst two segments shining white; ventral surface with fi ve pairs of dark brown stripes along lateral sides.
Pupa (Fig. 8). 4.0-6.0 mm, fusiform. Greenish yellow in early pupal stage, changing gradually to yellowish brown, blackish brown before eclosion. A corniform cocoon breaker on forehead. Forelegs to third abdominal segment, midlegs to fourth abdominal segment, hindlegs to seventh or eighth abdominal segment, wings to fi fth abdominal segment, antenna to or slightly exceeding end of abdomen.
Host plant. Euphorbiaceae: Flueggea suff ruticosa (Pall.) Baill. Life history. Conopomorpha fl ueggella has two generations annually in Tianjin, China (Table 1). Th e larvae feed on the seeds of Flueggea suff ruticosa (Fig. 10). Mature larvae quit the fruits before they are ripe and pupate on leaves or leaf litter. Th e pupal stage lasts from 9 to 12 days. Adults of the second generation hibernate.
Adults occur from May to the fi rst ten days of June, and from the last ten days of June to the fi rst ten days of August. Adults can emerge during the whole day, but the peak occurs in the morning. Th e mating occurs usually in the morning. At night, the moths are actively drinking nectar and ovipositing. Adults come sometimes at light. A parasitic Ichneumonid species was reared from pupae collected on leaves of F. suff ruticosa in the fi eld.

Epicephala relictella
Redescription. Adult (Figs 15-16). Wing expanse 9.0-13.0 mm. Head white, tufted. Labial palpus white except outer surface grey. Antenna with scape pale grey dorsally, white ventrally; fl agellum dark brown dorsally, copper-colored ventrally. Th orax white. Tegula and forewing greyish brown; white stripes at costal 2/5, 3/5, 4/5 and near apex as well as at dorsal 2/5 and 3/5 respectively, concentrated obliquely outward to 2/3 length and outside of cell, outmost one shortest, between fi rst three stripes sometimes with short white strigulae; thin bluish white fascia with metallic refl ection extending from costal 5/6 to dorsum; large black spot near apex; dorsal margin white tinged with ocherous yellow, longitudinally forming a broad band; termen dark brown; cilia white except fuscous distally from costal 5/6 along termen to before tornus, greyish brown along dorsal margin. Hindwing and cilia greyish brown. Fore and mid legs brown, hind leg greyish white, tibiae and tarsi with white rings. Abdomen greying brown on dorsal surface except fi rst two segments grey; vental surface grey, with fi ve pairs of oblique dark brown stripes along lateral sides.
Larva (Fig. 17). Young instar larva very similar to that of Conopomorpha fl ueggella. Mature larva 5.0-6.5 mm; head capsule brownish yellow, median 2/3 of each segment on thorax and abdomen dark red, anterior and posterior ends white; thoracic segments slightly blue, abdominal segments with blue spots. Body with sparse white setae. Th ree pairs abdominal legs on segment 3, 4 and 5 respectively; anal leg protruded backward.
Pupa (Fig. 18). 4.0-5.5 mm, fusiform. Greenish yellow in early pupal stage, changing gradually to dark brown. A corniform coccon breaker on forehead. Forelegs to third abdominal segment, midlegs to fourth abdominal segment, hindlegs to eighth abdominal segment, wings to fi fth abdominal segment, antenna obviously exceeding end of abdomen.
Life history. Epicephala relictella has one generation annually in Tianjin, China (Table 2). Th e larvae feed on the seeds of Flueggea suff ruticosa (Fig. 20). Th e larval stage is completed within one fruit. When completing larval development, the mature larvae quit the fruits and pupate on the leaves, and overwinter under leaf litter or stones.
Adults appear from June to July. Th ey can emerge during the whole day, but the peak occurs in the morning. Th e moths are most active at night, drinking nectar and ovipositing. During the daytime they rest on leaves or branches. Adult longevity is 3-10 days, but adults generally live for 5-7 days. Adults hardly come to light.

Discussion
Calybites securinella (Ermolaev, 1986) was the only species in Gracillariidae known to be associated with Flueggea suff ruticosa. It occurs in Russia (Primorye) and Korea (Ermolaev 1986, Kawahara et al. 2010. Associations of the gracillariid moths with fruits or seeds of diff erent euphorb genera, including Flueggea, are known for Epicephala (Kawakita and Kato 2009). Conopomorpha fl ueggella is another seed-feeder, similar to several species of the genus that feed on fruits or seeds of other plant families (Bradley 1986). Similar to Conopomorpha fl ueggella, Epicephala relictella also feeds on the seeds of Flueggea suff ruticosa. Th ey are very similar in morphology and biology, and hard to distinguish. Table 3 compares the life histories of the two species.
• egg, -larva, □ pupa, (□) pupa through the winter, + adult F: First ten days, M: Middle ten days, L: Last ten days. Most gracillariid species are leaf-miners, and the seed-parasitic habit is infrequent. Epicephala is noteworthy for its obligate pollination habit, which involves a mutualistic relationship with trees of Euphorbiaceae (Kawakita 2010). However, Epicephala relictella is not pollinating its host.