A revision of the genus Chlorocryptus Cameron (Hymenoptera, Ichneumonidae), with the first record of the genus from Japan

The genus Chlorocryptus is revised and two species are recognized: Chlorocryptus purpuratus (Smith, 1852) and C. coreanus (Szépligeti, 1916). Cryptaulax metallicus Szépligeti, 1916, which has been hitherto listed in catalogs as Chlorocryptus fuscipennis Townes et al., 1961, is shown as a new synonym of Chlorocryptus purpuratus. Chlorocryptus purpuratus and C. coreanus are redescribed and illustrated in detail and a key is provided. Chlorocryptus purpuratus is recorded from Japan and Vietnam for the first time and is considered to have recently been introduced into Japan.


Introduction
Chlorocryptus  is a small genus presently comprising three species: C. purpuratus (Smith, 1852), C. coreanus , and C. fuscipennis . They are easily sighted because of their large body size (10-15 mm) and metallic blue, purple, or green luster throughout the body. Chlorocryptus purpuratus is widely distributed in the Oriental Region to northern China and is well known as a common parasitoid of limacodid moths, which are injurious to leaves of various trees, especially oil palms (Gauld 1987, Mariau 1999. Chlorocryptus purpuratus and C. coreanus are well recognized, particularly in China , He and Chen 2006, He et al. 1996, 2004, Sheng et al. 2013. However, difficulties in identification or taxonomic confusion still exist in other countries (Gauld 1987, Jonathan 2006. In contrast, Cryptaulax metallicus , for which Townes et al. (1961) proposed a new name Chlorocryptus fuscipen-Chlorocryptus purpuratus from Japan as an alien species to provide the basis for monitoring the future expansion of the distribution range of this species.

Material and methods
Collection and field survey of Chlorocryptus purpuratus in Osaka and adjacent area were conducted by the second author (RM). Two immature specimens were collected from a cocoon of Parasa consocia Walker (Lepidoptera, Limacodidae). Cocoons of the moth are easily found clustered on the ground at the base of certain trees such as Cerasus × yedoensis (Matsum.) A.N.Vassiljeva, Celtis sinensis Pers., and Salix spp. in the study area. RM also conducted successful rearing experiments by supplying C. purpuratus females with Parasa consocia cocoon.
Morphological observations of Chlorocryptus specimens collected by RM and other colleagues and those from museum collections were conducted by the first author (TY). Specimen depositories are abbreviated as follows: Natural History Museum, London (BMNH), Hungarian Natural History Museum, Budapest (HNHM); Muséum national d'Histoire naturelle, Paris (MNHN); Osaka Museum of Natural History, Osaka (OMNH); Systematic Entomology, Graduate School of Agriculture, Hokkaido University, Sapporo (SEHU), Taiwan Agricultural Research Institute in Taichung (TARI). A part of the specimens were dissected for detailed observations. Genitalia and other body parts were detached from the specimens that were previously macerated in hot water, and the detached body parts were further dissected in 80% ethanol. The dissected body parts were heated in 10% KOH solution at 60 °C for a sufficient period of time depending on the condition of the material to remove muscles from the exoskeleton. Cleaned parts were then washed in diluted acetic acid and distilled water and observed in 80% ethanol or pure glycerol.
Larval exuviae of the parasitoid were extracted from the host cocoon and their cephalic structures and spiracles were observed. They were macerated and stretched in 10% KOH solution and processed as body parts of adult specimens. One 5th instar larva collected in the field was killed to obtain DNA barcode sequences and subsequently treated as the exuviae in morphological observations. Stereomicroscopes (Nikon SMZ-10 and Olympus SZ40), a light-microscope (Olympus BX40), and a field emission scanning electron microscope (SEM) (JEOL JSM-6301F, Tokyo, Japan) were used for morphological observations. Dissections and observations were carried out under the Olympus stereomicroscope, line drawings were made with a drawing tube attached to the Nikon stereomicroscope, and images of the wings and genitalia were taken using a Nikon Coolpix 4500 digital camera attached to the Olympus stereomicroscope. Images of larval exuviae were taken using the same digital camera attached to the light-microscope. Specimens for SEM analysis were not coated and were observed with an ac-celerating voltage of 1.0 kV. Digital images were edited using Adobe Photoshop Elements® 6.0.
Terminology used for adult morphology follows Gauld (1991) and Harris (1979), whereas descriptions of immature stages follow Short (1978). OOL and POL refer to ocello-ocular line and postocellar line, respectively. Nervellar index is defined as in Gauld and Mitchell (1981), i.e., the relative length of the first abscissa of Cu1 to the length of cu-a of the hind wing (Fig. 22).
DNA barcoding was carried out by RM. The DNA barcode sequences of cytochrome c oxidase I (COI) were obtained from an adult and a larva collected in Japan. Amplification of COI was carried out using the primer set LCO1490 and HC02198 designed by Folmer et al. (1994). The PCR protocol consisted of an initial 5-min denaturation at 94 °C, followed by 40 cycles at 94 °C for 15 s, 46 °C for 15 s, and 72 °C for 15 s, and ending with a 10-min extension step at 72 °C. Gene regions were sequenced using the same primer set and a BigDye® Terminator ver.  (Fig. 7), there densely covered with mixture of one type of sensilla chaetica and one type of sensilla basiconica (Figs 41,42); apex round, not truncated, possessing a bundle of thick sensilla, which are widened apically, and whose sockets interrupted on one side (Figs 39,40). Male flagellum not flattened ventrally; with 5 or 6 tyloids which can be on 13th-20th flagellomeres; apex of flagellum with thick sensilla as in female but fewer.
Wings infumate in various degrees . Fore wing with areolet small, quadrangular, 0.3-0.4 times as long as 2m-cu; 2m-cu joining behind middle of areola; cu-a a little distad of Rs&M. M+Cu of hind wing straight on apical 0.6; nervellar index less than 2.8; 2A curving toward anal margin.

Chlorocryptus purpuratus (Smith)
Description. Adult. ♀. Head 2.0-2.1 times as wide as long in dorsal view (Fig. 4). Temple flat and narrow, occupying only 0.2-0.3 of length of head in dorsal view (Fig. 4). POL/OOL=0.4-0.8. Vertex and gena densely punctate with shallow and small punctures; punctures smaller on gena than on vertex. Face 1.4-1.5 times as wide as high, distinctly convex medially, densely punctate with fine punctures and with some sublateral transverse rugae (Figs 2, 5). Clypeus with median obtuse protuberance on apical margin (Fig. 3); punctures sparser than on face and irregular with mixture of different sized punctures. Malar space 0.9-1.2 times as wide as mandible width. Mandible with acute teeth, its lower tooth about half length of upper one (Fig. 6). Antenna with 31-32 flagellomeres; apical third of flagellum weakly thickened and weakly flattened ventrally, weakly tapered to apex; first flagellomere 3.4-4.3 times as long as apical width, a little longer than 2nd flagellomere; fifth flagellomere 1.5-2.1 times as long as apical width (Fig. 7).
Epomia turns horizontally toward mesal line of pronotum at midway to upper edge of pronotum ( Fig. 9). Prepectus with a short vertical carina opposite lower corner of pronotum. Mesoscutum densely punctate, with longitudinal to oblique rugae on median lobe, and with moderate-sized glabrous area on lateral lobe (Fig. 8 Legs slender. Hind femur 6.0-7.3 times as long as maximum width. Hind tibia 9.3-10.7 times as long as apical width. Fore wing about 10-15 mm long. Nervellar index 1.8-2.8 (Figs 22-25).
First metasomal tergite 1.8-2.1 times as long as apical width, with postpetiole 0.6-0.9 times as long as apical width; area between median longitudinal carinae distinctly raised at proximal margin of postpetiole (Figs 11, 12). Ovipositor sheath 0.8-1.0 times as long as hind tibia.
Body with metallic luster in blue, purple or green. Flagellum entirely black, without white band. Fore and middle tibiae and all tarsi dark brown to black. Ovipositor sheath and ovipositor black. Wings infumate in variable degrees (Figs 22-25). Veins dark brown to black, strongly pigmented all over.
♂. Similar to female except as follows: face 1.2-1.3 times as wide as long; malar space 0.8-1.0 times as long as basal width of mandible; flagellum with 35-37 flagellomeres, with 5 or 6 tyloids which can be on 13th-20th flagellomeres; 1st flagellomere 2.7-3.3 times as long as apical width; 1st metasomal segment 1.6-2.1 times as long as apical width; hind femur 7.3-8.0 times as long as median depth; hind tibia 10.3-12.7 times as long as apical width; anterior side of fore femur, fore tibia and dorsal stripe of middle tibia light brown. Subgenital plate with posterior margin evenly convex and with postero-lateral corner gently rounded (Figs 33-35).
Second instar larva. Prognathus. Narrow epistomal spur present. Two pairs of setae on labium of 1st instar remaining but upper most one situated just below salivary orifice present only as a hole. Spiracle with atrium differentiated.
Third and fourth instar larvae. Epistomal spur more developed than in 2nd instar and stipital sclerite present.
Biology. Several limacodid moths have been recorded as hosts. Some of the moths are serious pests of palm trees in the Oriental Region (Gauld 1987, Mariau 1999; hence, C. purpuratus is of economic importance as their natural enemy. The host records are summarized in Table 1. In this study, we observed females collected in Osaka ovipositing into cocoons of Parasa consocia Walker (Fig.  1). They spent more than 30 min to penetrate the moth cocoons and to accomplish the oviposition.
Remarks. Although this species shows a wide range of variation in density of the surface sculpture and setae, wing color (Figs 22-25), and shape of the subgenital plate (Figs 33-35), these variations are continuous. The specimens from Sulawesi, including the lectotype of Cryptaulax metallicus Szépligeti, have extensively dark  wings (Fig. 25); however, we considered this an extreme end of the variation. Jonathan (2006) treated Chlorocryptus coeruleus Cameron and Chlorocryptus metallicus Cameron as distinct species. However, given his descriptions, they fall within the variation of Chlorocryptus purpuratus.
Despite these large intraspecific variations, this species is clearly distinct from Chlorocryptus coreanus with the above given diagnostic characters.
The distribution of Chlorocryptus purpuratus extends further south than that of C. coreanus although the distributions of both species largely overlap in China (Figs 58,  59). We do not know what determines such a difference in the distribution and whether the species compete or segregate where they co-exist. The host and habitat preferences might be different between the species, which may also be of practical interest in the biological control of the notorious limacodids on various trees. This is the first record of Chlorocryptus purpuratus from Japan and Vietnam. Considering that C. purpuratus has never before been collected in Japan, most probably the Japanese populations have been introduced recently. Its potential hosts, limacodid moths, are abundant and some of them have been well studied as pests in Japan, including their natural enemies (Kaji 1979, Koike 1985, 1998, Komeda and Hisamatsu 2006, Minamikawa 1962, Oda and Uezumi 1978, Togashi 1981, Togashi and Ishikawa 1994, 1995, Toyomura 1970, Yuasa 1934.
In 2012 and 2013, alongside the adults, we also collected two larvae in Parasa consocia cocoons in Osaka. This indicates that C. purpuratus has already been established in this area for two or three years.
One of the limacodid moths, Parasa lepida (Cramer), which had been originally distributed in South Asia, Southeast Asia, and southern China, invaded western Japan and became established in the late 1970's Hattori 1981, Higa andKishimoto 1984). It is probable that cocoons of Parasa lepida or other Limacodidae containing C. purpuratus were introduced accidentally in Japan with imported nursery trees.
The holotype of Neodontocryptus hyalina Saxena could not be located during the course of this study.
The holotype of the species was deposited in the V. K. Gupta collection, which was originally housed at the University of Delhi. Later, his collection was transferred to Florida, first to the University of Florida (Yu et al. 2012) and finally to FSCA (Virendra K. Gupta, personal communication). Therefore, we expected that the holotype of N. hyalina would be at FSCA; however, only one paratype was located, whereas the holotype was not found (Jim Wily, personal communication).
The lectotypes of Cryptaulax cyaneus Szépligeti and C. metallicus Szépligeti were designated by Townes et al. (1961). Five paralectotypes (two females and three males) of C. cyaneus and one paralectotype (female) of C. metallicus are preserved at HNHM together with their lectotypes (Gellért Puskás, personal communication). One female paralectotype of C. metallicusis is preserved in the Uchida collection at SEHU, which he obtained probably after visiting HNHM during his study trip to Europe and the United States from 1937 to 1939 (Uchida 1940).
Epomia ends at midway to upper edge of pronotum without turning toward mesal line of pronotum (Fig.  19). Prepectus without a short vertical carina opposite lower corner of pronotum. Mesoscutum and scutellum lacunose, without rugae, and with only narrow glabrous stripe on lateral lobe (Fig. 18) Legs slender. Hind femur 6.0-7.7 times as long as maximum width. Hind tibia 8.5-9.5 times as long as apical width.
Body with metallic luster in blue, purple or green. Flagellum black with white band on 6th-9th flagellomeres. Fore and middle tibiae and all tarsi are dark brown to black. Ovipositor sheath and ovipositor black. Wings narrowly infumate apically (Fig. 26).
♂. Similar to female except as follows: face 1.0-1.2 times as wide as long; flagellum with 37-39 flagellomeres, with 5 or 6 tyloids which can be on 14th-20th flagellomeres; 1st flagellomere 2.8-3.3 times as long as Nepal apical width; 1st metasomal segment 2.0-2.4 times as long as apical width; hind femur 7.0-8.5 times as long as median depth; hind tibia 9.3-10.0 times as long as apical width; stripe on anterior face of fore femur, fore tibia and dorsal stripe of middle tibia light brown. Subgenital plate with postero-lateral corner more angulated than in C. purpuratus, with posterior margin convex, with small concavity on middle of convexity (Figs 36, 37), but one aberrant specimen from Korea without concavity (Fig.  38); usually with white spot near base of apodeme. Wings narrowly infumate in specimens from Korea and North China, but Taiwanese specimens with extensively brown wings (Fig. 27).
In the description of Cryptaulax coreanus, Szépligeti (1916) did not originally designate the holotype nor mentioned the number of specimens he examined. The Szépligeti's collection at HNHM contains only one specimen of the species, which Townes et al. (1961) treated as the holotype fixed by monotypy.
Cryptus trirrhogmaniformis Sonan was described based on two male syntypes. One was deposited at TARI and the other was probably donated to Uchida at that time and now is located at SEHU. The TARI specimen was treated as "type" by Townes et al. (1961), which is considered a lectotype designation as it is evident that the authors cited the specimen to serve as the name-bearing type (ICZN 74.5).

Key to species
1 Temple swollen and wide, occupying 0.3-0.4 of length of head in dorsal view (Fig. 15). Face without transverse rugae ( Fig. 13). Female flagellum with white annulus around mid-length and with apical third ventrally weakly flattened but not widened. Mesoscutum punctate, without rugae, and glabrous area on lateral lobe narrow (Fig. 18) (Smith).