Taxonomy and phylogeny of the genus Gastrocentrum Gorham (Coleoptera, Cleridae, Tillinae), with the description of five new species

Abstract The genus Gastrocentrum Gorham, 1876 is revised to include nine species. Five new species are described in this genus: G. magnumsp. nov. (NE India), G. regularesp. nov. (Cameron Highlands, Malaysia), G. xiaodongisp. nov. (Gyirong, Xizang, China), G. zayuensesp. nov. (Zayü, Xizang, China), and G. gaoligongensesp. nov. (Fugong, Yunnan, China). Gastrocentrum nitidum Schenkling, 1916 is transferred to the genus Tillus as a new combination. All the species in this genus are described (except G. brevicolle), and a key is provided for their identification. Illustrations of male genitalia, female reproductive organs, and other important structures are provided. An interspecific phylogeny-estimate of Gastrocentrum is presented based on morphological data, with two main clades recognized: a clade containing G. unicolor and G. laterimaculatum, and a clade containing the remaining six species (the latter a polytomy consisting of G. magnumsp. nov., G. dux, and G. regularesp. nov., and a well-supported sub-clade representing the remaining species). Additionally, the taxonomic and phylogenetic importance of female reproductive organs is discussed.


Introduction
of reproductive organs were dyed with Chlorazol Black. Habitus images were captured using a Nikon D7000 digital camera with a Tamron SP 90mm lens. Terminalia images were captured by a Canon 450D digital camera fitted to a Nikon SMZ-1500 stereoscopic dissecting microscope.
Morphological terminology follows the works of Ekis (1977) and Opitz (2010) in general. For the convenience of taxonomic description and phylogenetic analysis, elytral asetiferous punctations are classified into primary asetiferous punctation (PAP) and accessory asetiferous punctation (AAP). PAP refers to the major ten rows of punctations which are also present in many other genera of Tillinae, such as Tillus, Cladiscus, Diplopherusa, and Diplocladus (Fig. 10B, C). AAP is the additional punctation that presents on interspaces among PAP rows and, in Gastrocentrum specifically, AAP presents on interspaces between 1 st -2 nd , 3 rd -4 th , and 5 th -6 th PAP rows (Fig. 10B, a). In some species such as G. zayuense, PAP decreases in quantity and PAP rows are less than ten rows.
The term microtrichia on the inner surface of elytra was adopted from Gorb (2001). A new term interphallic plate was introduced to depict a plate that inserted at the membranous part of phallus where situated between the two phallic plates (Fig. 11C, H, ipp).

Phylogenetic analysis
Phylogenetic analysis was made using PAUP 4.0a (build 167) (Swofford 2002). Twenty-two morphological characters for eight ingroup and two outgroup taxa were compiled and analyzed. Gastrocentrum brevicolle was not included. Both the two species of Isocymatodera (I. kolbei and I. atricolor) were selected as outgroups. Exheuristic maximum parsimony analyses were performed. Characters were unordered and of equal weight. Branch support was determined for parsimony analyses using bootstrap with 1000 replicates in PAUP*. A bootstrap consensus tree and a list of character changes were obtained by PAUP*, and unambiguous character were mapped onto the tree by Illustrator 21.0.0.
Diagnosis. The genus Gastrocentrum was included in the Philocalus genus group close to the genus Isocymatodera (Gerstmeier 2005;Gerstmeier and Weiss 2009). Both genera have the claw with one inner denticle, which is similar to and only very slightly smaller than the apical portion of the claw (Fig. 20F). The genus Gastrocentrum can be differentiated from Isocymatodera by antennae broadly expanded laterally from 7 th or 8 th antennomeres onwards, the connecting membrane of male aedeagus specialized in both dorsal and ventral surface, the female vagina devoid of sclerites. While in Isocymatodera, the antennae are expanded laterally from 3 rd or 4 th antennomeres onwards, the connecting membrane of male aedeagus is only specialized in dorsal surface, and the female vagina possesses sclerites.
Redescription. General appearance: body length 9-29 mm; oblong, somewhat robust; all the species except G. laterimaculatum uniformly dark brown (Figs 1-9); vested with long, yellow setae all over the body. Head: hypognathous, moderately large, including eyes slightly broader than pronotum; eyes sizable, emarginate, coarsely faceted, ocular notch small, distance of eyes as long as or only slightly greater than transverse diameter of eyes; gula broad, gular sutures parallel or slightly converging in anterior; antenna comprised of eleven antennomeres, broadly expanded laterally from 7 th or 8 th -11 th , the expanded antennomeres triangular except the last one cultriform, all the expanded antennomeres clothed with fine and dense pubescence; labrum emarginate, mandibles stout with inner dens, terminal segment of maxillary palpi digitiform, that of labial palpi broadly securiform. Thorax: pronotum long campaniform, constricted posteriorly, anterior transverse depression feeble, surface punctate, faintly wrinkled, clothed with long, yellow hairs; pro-intercoxal process thin. Elytra: oblong, sides parallel, anterior ridge present from humerus to scutellum; inner surface with a wedge-shaped protuberance at lateral middle of each elytron (Fig. 20A, B), leaning to the lateral side of first ventrite of abdomen in resting position (except for two species: G. xiaodongi and G. zayuense), and with a microtrichia field on anterio-lateral area (Fig. 20A, mt; similar to the structure found in Tenebrionidae, Gorb, 2001: 125, fig. 8.1, EAL); elytra have two types of punctations: asetiferous and setiferous punctations, the former comprised of primary asetiferous punctation (PAP) and accessory asetiferous punctation (AAP); each elytron possesses ten rows of PAP in general, the fifth row situate just before the humerus (Fig. 10C, D eral elytron (in G. zayuense and G. gaoligongense, Fig. 10E-H); AAP may present on the interspaces between 1 st -2 nd , 3 rd -4 th , and 5 th -6 th PAP rows and similar with PAP in size, making these two types of punctations more or less indistinguishable (Fig. 10A, B); setiferous punctations minute, bearing setae, densely dispersed over the whole elytral surface, not in rows. Legs: tibia without longitudinal ridge, tibial spur formula 1-2-2 (but in G. laterimaculatum 0-2-2), protibia without tooth at outer apex (except in G. unicolor and G. laterimaculatum where protibia possess a blunt tooth at outer apex); tarsi formula 5-5-5, first to fourth tarsomeres of all legs more or less bilobed and bearing evident pulvilli; claw with one inner denticle, which similar to and only very slightly smaller than apical portion of claw (Fig. 20F). Abdomen: abdomen longer than broad, parallel in front and tapering in rear; first to fifth abdominal ventrites each with a pair of short longitudinal ridges (Fig. 20D) and a pair of less pigmented circles (Fig. 20C) in lateral; sixth ventrite partly or totally slid under the fifth, so in parts of specimens only five ventrites visible; intercoxal process of the first ventrite keel-like, with or without longitudinal groove; first ventrite strongly ridged behind metacoxae. Male genitalia: pygidium subquadrate; sixth ventrite subtriangular to semicircle, posterior margin somewhat rounded, secondary sexual modifications slight (Figs 11F,G,13G,15H,17G,19G); central parts of sixth ventrite membranous, shape of membranous region different among species; spicular fork well developed, plates slender, apodemes not fused centrally, longitudinal intraspicular plate present (Figs 11D,13E,15F,17E,19E); tegmen tubiform, sclerotized from dorsal midline to lateral sides, barely sclerotized and unpigmented in ventral middle, tegmen lobed distally, parameres bent to ventral direction, tip simple or hooked, phallobasic apodeme present (Fig. 15B, C); phallus comprised of two thin phallic plates devoid of dentations, an interphallic plate present on the membrane between the two phallic plates (Fig. 11C, H, ipp), phallus apex simple, knot-like, phallic struts long and slender; connecting membrane between tegmen and phallus well sclerotized and thickened except the dorsal midline and ventral midline, forming a nearly whole sheath covering the phallus, which surface densely equipped with microhooks ( Fig. 11A-C). Female reproductive organs: pygidium subquadrate; sixth ventrite sub-triangular, disc membranous (Figs 12H, 14B, 16C, E, 18C), spiculum ventrale present; ovipositor as long as abdomen, moderately sclerotized, light yellow, semi-transparent (Figs 12F, 14C, D, 16A, F, 18A, ovp), with proctigeral bacculi in dorsal surface (Fig. 16F, pgb) and ventral and oblique bacculi in ventral surface (Fig. 14C, vtb, olb); vagina and alimentary canal partially enclosed in ovipositor, unenclosed part of vagina as long as or slightly longer than ovipositor, tubular or saccular; bursa copulatrix clearly defined and positioned distally (Figs 12F,16A,18A,bc) with the exception of the species G. gaoligongense, where bursa copulatrix is a mere swollen continuation of the vagina (Fig. 22E); spermatheca attached to the base of bursa copulatrix, boundary between spermathecal duct and spermathecal capsule somewhat obscure (Figs 16G,21), spermathecal duct slender (Fig. 16A) or sometimes inflated in distal and continuous with spermatheca (Figs 16G, 18A), spermathecal capsule moderately to minimally sclerotized, both spermathecal duct and spermathecal capsule with spiral micro-texture, distal part of spermathecal capsule strongly bent, angled no more than 90° (Fig. 16A, G), spermathecal gland duct inserted at the outer edge of the angle (Fig. 21, spgd); in ground-plan, spermathecal gland have three taillike endings: one located distally, opposite to its opening to spermatheca (top tail, Figs 12E, 14C, spgtt), the other two situated laterally (lateral tail, Figs 12E, 14C, spglt), any of which may be missing in different species and sometimes can be extremely long (Fig. 14C, spglt).
Variation. The tegmen apices of G. unicolor are simple, unhooked, unspecialized (Fig. 11A). However, specimens from Sulawesi with tegmen apices slightly more prominent than those from other regions. Phallus apex is normally knot-like with the two phallic plates convergent at a point before the tip (Fig. 11H), but in the holotype of G. pauper, edges of the two phallic plates are almost parallel to the tip (Fig. 11B). The apical tip of phallus is longer than broad, with length/width ratio varied in a range of 2.0~3.0; usually teardrop-shaped with length/width ratio 2.0~2.5, but oblong in specimens from Sulawesi with length/width ratio approximate to 3.0 (Fig. 11H).
Both of the two female specimens examined from Hainan has spermatheca tubiform (Fig. 12B), which is different from those from other localities with spermatheca inflated distally (Fig. 12A, C, D). However, given its same external structure and lacking male specimens, we consider the specimens from Hainan as the same species with G. unicolor.
Discussion. Gahan (1910) proposed that G. pauper was a junior synonym of G. unicolor without explanation, which treatment was afterward followed by Schenkling (1912), Chapin (1924), Corporaal (1950), and temporarily by Mawdsley (1999) and the present paper. In our research, we have only examined specimens from SE Asia and determined they are identical with G. pauper. However, additional materials from India or Sri Lanka need to be compared with those from SE Asia thoroughly, which will lead to the confident assignment of the synonymy. Diagnosis. This species is the only one in this genus that has elytral pattern and can be separated from other species without difficulty. Its elytra is yellow-brown, with a pair of large semicircular dark spots in lateral sides which is extended to the lower sides of humeri; elytral asetiferous punctations larger than other species, with punctation diameter greater than interspace between 2 nd -3 rd PAP rows; antennae broadly extended laterally from 8 th antennomere onwards.
Supplementary description. Elytral wedge-shaped protuberance present on inner surface; elytral asetiferous punctations somewhat irregular, arranged in more than ten rows, PAP in ten rows, AAP on interspaces between 1 st -2 nd , 3 rd -4 th , and 5 th -6 th PAP rows, AAP setting in two rows on each interspace; AAP almost same size as PAP, which is more sizable than punctations in other species, with punctation diameter greater than interspace between 2 nd -3 rd PAP rows; both PAP and AAP beginning to decrease in size postmedially to apical third, and completely vanished at apical fifth; elytral inner surface with a wedge-shaped protuberance at lateral middle; tibiae spur formula 0-2-2 (other species in this genus 1-2-2); protibia with a blunt tooth at outer apex; 1 st -2 nd abdominal ventrites dark brown, 3 rd -5 th yellow, 6 th light yellow to transparent; first abdominal ventrite strongly ridged behind coxae, intercoxal process raised, triangular, slightly longer than broad, grooved longitudinally.
Distribution. Malaysia (Peninsular Malaysia, Sabah).  Diagnosis. Earlier researchers identified one of the paratypes of this new species as G. dux. The new species can be separated from G. dux by: asetiferous punctations on elytra continuing to the tip (Fig. 10A); intercoxal process of first abdominal ventrite grooved longitudinally (Fig. 20F); female pygidium with anterior margin notched in a shallow triangular shape (Fig. 14A), and lateral tails of spermathecal gland extremely long, ca. 2 × the length of ovipositor (Fig. 14C, spglt). Gastrocentrum unicolor is sym-patric with the new species in India, but G. magnum can be separated from it by much larger body size and five expanded terminal antennomeres (Fig. 13H).
Variation. The female paratype collected from Hainan has the spermatheca faintly bifurcated distally. This individual variation was also observed in specimens of G. zayuense collected from the same locality ( Fig. 18D-J).
Etymology. This new species, together with G. dux, have the largest body size in this genus. The specific epithet comes from the Latin adjective magnus (=large). ( Fig. 2). Diagnosis. The specimen examined can be separated with G. magnum by elytral asetiferous punctations stop by apical fifth, not continuing to the tip (Fig. 10B), intercoxal process of first abdominal ventrite not grooved, female pygidium with a semi-circle membranous region proximally, reaching half length of pygidium, lateral tail of spermathecal gland much shorter, only slightly longer than spermatheca (Fig. 14D, spglt).

Gastrocentrum dux
Description. General appearance: length 23-29 mm, robust, dark brown. Head: including eyes feebly broader than pronotum; eyes moderately large, distance between eyes nearly as long as the transverse diameter of eye; gular suture slightly convergent in anterior; antennae expanded laterally from 7 th antennomere onwards; vertex and frons densely punctate, with a very faint ridge along midline, postgenae rugose. Pronotum: oblong, length/width ratio ca. 1.4, constricted posteriorly; surface finely and densely punctate, clothed with light yellow hairs. Elytra: oblong, sides subparallel, length/ width ratio ca. 2.31, vested with light yellow setae; wedge-shaped protuberance present on inner surface; PAP in ten rows, AAP on interspaces between 1 st -2 nd , 3 rd -4 th , and 5 th -6 th PAP rows; AAP present in two very incomplete rows, number of AAP less than that in G. magnum; AAP faintly smaller than PAP; interspace between 2 nd -3 rd PAP rows greater than punctation diameter; elytral punctations decreasing in size postmedially, and completely vanished at apical fifth (Fig. 10B). Legs: outer apex of protibia very faintly extending outwards, not forming a distinct tooth. Abdomen: intercoxal process of the first ventrite flat, not grooved. Male genitalia: not studied. Female reproductive organs: pygidium slightly wider than long, posterior margin rounded, a semi-circle membranous region present proximally, reaching to half length of the pygidium; sixth ventrite trapezoidal, wider than long; bursa copulatrix clearly defined; spermathecal gland only with one lateral tail, which slightly longer than spermatheca in fully stretched condition (Fig. 14D, spglt); spermatheca boot-shaped (Fig. 14D, sp).
Note on type specimen. Mawdsley (1999) claimed that the type specimen of G. dux was deposited in the Hope Department of Entomology, University Museum, Oxford, United Kingdom, but it was not located during a visit to that museum in 2011 by the first author. Westwood (1852) indicated that the type specimen was from "Mus. Melly", but efforts to locate it in Melly's collection in the Natural History Museum, Geneva, yielded no results either. The whereabouts of the type specimen remains unknown.
Discussion. The Australian type locality of this species is doubted by the Australian entomologist and clerid worker Justin Bartlett who, after viewing the Cleridae holdings of all major museum, and several agricultural and private collections from all Australian states, is yet to find a single Gastrocentrum specimen, and therefore does not believe G. dux to be an Australian species. He also doubts that the locality label of the specimen examined in this manuscript represents an actual collecting event, but rather was labelled after it was identified as G. dux, with the associated type locality of 'Australie' (pers. comm. J Bartlett). He also pointed out that another apparently Australian specimen from Melly's collection, a longicorn Hephaestion acraetus Newman, is in fact a Chilean species (see Saunders 1850), providing a precedent for erroneously labelled specimens from Melly's collection. Despite this, no more practical specimen-based evidence for or against this argument has been found. Hence, we can only describe this species based on the specimen mentioned above at the moment, as we can only take the label at face value and assume it to represent an actual collecting label.
We found a Tenebrionidae beetle with the same Swan River type locality also originating from Melly's collection and described by Westwood: Prophanes aculeatus Westwood, 1849. It is presently treated as a valid species, with an eastern, not western, Australian distribution (Westwood 1849;Carter 1913;Matthews 1992). Gastrocentrum dux may have a similar historical story and its correct occurrence could be in other areas of Australia or in other regions of the world, but this hypothesis needs to be proved by further specimens.
Distribution. Malaysia (Peninsular Malaysia). Etymology. Refer to the highly regular elytral asetiferous punctations of this species. Diagnosis. This new species is different from G. regulare sp. nov. by: antennae expanded laterally from 8 th antennomere onwards; elytral asetiferous punctations stop by middle (Fig. 10D); elytral inner surface without wedge-shaped protuberance; intercoxal process of first abdominal ventrite not grooved (Fig. 20C, ip); spermathecal capsule thicker, rounded distally (Fig. 16F, sp). The new species also looks similar to G. zayuense sp. nov. and G. gaoligongense sp. nov. at first glance, but it differs from the latter two species by: elytral asetiferous punctations somewhat larger and reaching lateral margins (Fig. 10D), female antennae expanded laterally from 8 th antennomere onwards (Fig. 16H). It also differs from G. gaoligongense sp. nov. by elytral inner surface without a wedge-shaped protuberance.
Distribution. China (Xizang, Gyirong), Nepal. Etymology. We are pleased to dedicate this species to its collector and our friend, Mr Yang Xiaodong.
Variation. All examined specimens are from exactly same locality, they vary individually in the number of punctations on one elytron from zero to 27, and spermatheca apex being simple or feebly bifurcate distally.
Distribution. China (Xizang, Zayü). Ecology. Habitat is shown in Fig. 24. The specimens were collected on the tree trunk at night.
Etymology. The new species is named after its type locality.  Diagnosis. The new species differs from G. zayuense sp. nov. by: elytra with a pair of wedge-shaped protuberance on inner surface; tegmen apices bulged on ventral surface (Fig. 19b); female bursa copulatrix not differentiated, merely a swollen continuation of the vagina, distal part of spermathecal capsule, long and slender, length/width ratio > 5 (Fig. 22E).

Gastrocentrum brevicolle
Note. This species was not studied because specimens were unavailable. Gerstmeier (2005) stated that the position of this species in the genus Gastrocentrum is doubtful because its pronotum was not elongated, but rather spherical.

Figure 23.
A preliminary phylogenetic analysis of Gastrocentrum, showing 50% majority-rule consensus MP tree. Only unambiguous characters are shown. Black circles represent characters having a CI of 1.000 while each state is derived only once, whereas white circles represent characters having a CI less than 1.000 while each state is derived more than once. Bootstrap support values are given at nodes. Female spermathecal glands are illustrated with top tail orientated to left side, but it is not known in G. laterimaculatum.

Significance of female reproductive organs
Female reproductive organs are inferred to have taxonomic and phylogenetic importance in genus Gastrocentrum. In certain Oriental genera of Tillinae, the vagina is equipped with a pair of sclerites or a joint sclerite, such as in Tillus (Kolibáč 1989: 17, figs 46, 47), Isocymatodera (Fig. 22A, C, D) and Cladiscus (unpublished data), but this sclerite is absent in all the species of Gastrocentrum.
In the present study, we find that almost all the species in Gastrocentrum have a clearly defined bursa copulatrix (Figs 12, 14, 16, 18), the only exception being G. gaoligongense (Fig. 22E). This disproves the idea of Opitz (2010: 51) that the presence or absence of a bursa copulatrix is consistent within stable genera.
The morphology of the spermathecal gland was rarely extensively studied previously in Cleridae. In Gastrocentrum, we find that this structure was phylogenetically significant at the infra-generic level (Fig. 23). The three-tailed spermathecal gland occurring in G. unicolor and the outgroup Isocymatodera was supposed to be plesiomorphic in Gastrocentrum (Fig. 23). In G. magnum and G. dux one lateral tail was lost, while in G. regulare, G. zayuense, G. xiaodongi, and G. gaoligongense, two lateral tails were lost. The absence of the top tail was autapomorphic for G. dux.
The shape of the spermathecal capsule was believed by Opitz (2010: 51) to be consistent within stable genera, however, we found that this structure was different among several Gastrocentrum species and thus was of some value for species-level identification, for example, the spermathecal capsules were tapered in G. regulare (Fig. 16A), barrel-shaped in G. xiaodongi (Fig. 16G), short in G. zayuense (Fig. 18A), and long and slender in G. gaoligongense (Fig. 22E).