Factors Affecting Thanatosis in the Braconid Parasitoid Wasp Heterospilus prosopidis.

Thanatosis, also called death feigning, is often an antipredator behavior. In insects, it has been reported from species of various orders, but knowledge of this behavior in Hymenoptera is insufficient. This study examined the effects of sex, age (0 or 2 days old), temperature (18 or 25 °C), and background color (white, green, or brown) on thanatosis in the braconid parasitoid wasp Heterospilus prosopidis. Thanatosis was more frequent in 0-d-old individuals and in females at 18 °C. The duration of thanatosis was longer in females, but this effect of sex was weaker at 18 °C and in 0-d-old individuals. The background color affected neither the frequency nor duration. These results were compared with reports for other insects and predictions based on the life history of this species, and are discussed from an ecological perspective.

Unexpected Species Identities and Interspecific Relationships in a Subterranean Beetle Lineage, the Pterostichus macrogenys Species Group (Coleoptera, Carabidae), Revealed by Fine-Scale Field Sampling and Detailed Morphological Comparisons.

An endemic subterranean Japanese carabid beetle lineage, the Pterostichus macrogenys species group, was recently revealed to have marked regional differentiation. Studies of such features reveal insect species diversity and provide insight into the mechanisms driving species diversity. We examined specimens of this species group collected from the southern Tohoku District of Honshu, Japan, where its diversity has not yet been fully elucidated, using fine-scale field sampling and detailed comparative morphological analysis of male genitalia. In total, 103 specimens from 13 localities were classified into one new (P. monolineatus sp. n.) and eight known species. In four of the known species, we observed disjunct distributions, which have not previously been reported in this species group and may be more common than previously recognized. Species coexistence was observed at four sites, with two species of different body sizes coexisting at three sites and three species coexisting at the remaining site. The three coexisting species included one large and two small species, the latter of which have male genitalia of a different size. This newly discovered coexistence pattern implies separate effects of differential body and genital size in species coexistence, which has rarely been reported in insects.

Taxonomic and nomenclatural changes in three species of Pterostichus Bonelli (Coleoptera: Carabidae) from the Far East.

The ground beetle genus Pterostichus Bonelli has diversified in regions including the Far East, but taxonomic issues remain even at the species level. This study presents taxonomic and nomenclatural changes in three species of Pterostichus from the Far East: P. (Petrophilus) eximius Morawitz, P. (Lenapterus) wellschmiedi Kirschenhofer, and P. (L.) subrugosus Straneo. The analyses are based on comparative studies of the endophallus of male genitalia, which is taxonomically useful in Carabidae but has not been examined in these species. The first species, P. eximius, has been treated as a monotypic species that is widely distributed in Eastern Siberia, the Russian Far East, and adjacent regions, but our results revealed that populations from Rishiri-tô and the Sakhalin islands are distinct from the nominotypical population from Transbaikalia. The name for the Sakhalin population, which was synonymized with P. eximius, is reinstated as subspecies P. eximius sachalinensis stat. nov., and the Rishiri-tô population is described as Pterostichus eximius rishiridakensis ssp. nov. Conspecificity (syn. nov.) was confirmed for P. wellschmiedi, which was described from southern Sakhalin, and P. marginatus Matsumura, which was described earlier from southern Sakhalin. No conspicuous differences in the endophallus structure were found between P. subrugosus, which was described from Hokkaidô, and P. marginatus, although differences are recognized in the elytral sculptures and in the metallic luster of the dorsal surface. The taxon was thus downgraded to subspecies P. marginatus subrugosus stat. nov.

Taxonomic studies of the ground beetle subgenus Falcinebria Ledoux & Roux, 2005 (Coleoptera, Carabidae, Nebria) from Honshû, Japan.

Nebria (Falcinebria) reflexa Bates (Carabidae), a Japanese endemic flightless ground beetle, is geographically polytypic and was thought to be composed of four subspecies (including nominotypical subspecies). Populations from Honshû recognized as three subspecies were taxonomically revised based primarily on the shape of the endophallus, a membranous inner sac everted from the aedeagus. Three known taxa, Nebria reflexa, Nebria niohozana Bates, and Nebria uenoi Nakane, are redefined based on endophallus morphology, and the latter two are described as distinct species rather than subspecies of N. reflexa. Seven new species are described: N. sagittata sp. nov., N. iidesana sp. nov., N. furcata sp. nov., N. pisciformis sp. nov., N. kuragadakensis sp. nov., N. dichotoma sp. nov., and N. chugokuensis sp. nov. Comparative morphology, primarily of the endophallus, indicated that N. furcata, N. pisciformis, N. kuragadakensis, and N. uenoi form basal lineages, and the remaining six species form a clade in which N. niohozana and N. dichotoma are sister taxa. Species phylogeny and known distributions suggest that the initial diversification of these species occurred in the western Chûbu and eastern Kinki regions of Japan.

Learning-induced host preference in male parasitoid wasps as a potential driver of ecological speciation.

Ecological speciation via a host shift plays an important role in the diversification of phytophagous and parasitic insects. However, it is not clear how separation is maintained among initial populations in which genetic separation mechanisms are not fully established. Learning-induced host preference in females can contribute to host fidelity and result in a barrier to gene flow in the initial populations. However, the role of males, which also affects gene flow, has been largely unexplored. We examined host preference through induced learning in males, which can contribute to initial population divergence, in the parasitoid wasp, Anisopteromalus calandrae, and two host species, Callosobruchus chinensis and C. maculatus. Behavioural experiments were performed using a four-chamber olfactometer. When wasps were conditioned during the preimaginal and early adult stage without mating experience, no preference was induced regardless of the host species. However, when wasps were conditioned at the early adult stage using the odour of the rearing host and mate reward, preference was induced in both host species. These results demonstrate mate reward learning-induced host preference in males. Interestingly, when the rearing host species and the mating conditioning host species were interchanged, preference was induced only in males conditioned to C. maculatus at mating. Thus, depending on the host species, preimaginal and early adult learning is vital to preference induction. Our results suggest that behavioural change via learning in males plays a more important role in the ecological speciation of phytophagous and parasitic insects than previously recognized.

Taxonomic notes on the ground beetles in the genus Trephionus Bates, 1883 from central Honshu, Japan (Coleoptera, Carabidae, Sphodrini, Synuchina).

Trephionus Bates, 1883, a Japanese endemic genus in the subtribe Synuchina (Coleoptera, Carabidae, Sphodrini), is revised taxonomically based mainly on the shape of the endophallus, a membranous inner sac everted from the aedeagus of the male genitalia. Three known species from central Honshu, T. kinoshitai Habu, 1954; T. shibataianus Habu, 1978; and T. babai Habu, 1978, are re-defined based on this genital character, and five new species are described from the region: T. cylindriphallus Sasakawa, sp. n., T. niumontanus Sasakawa, sp. n., T. inexpectatus Sasakawa & Itô, sp. n., T. abiba Sasakawa & Itô, sp. n., and T. bifidilobatus Sasakawa & Itô, sp. n. The observed interspecies differences in endophallus morphology are discussed in terms of the species-level phylogeny and genus-level taxonomy of Trephionus.

Notes on the Reproductive Ecology and Description of the Preimaginal Morphology of Elaphrus sugai Nakane, the Most Endangered Species of Elaphrus Fabricius (Coleoptera: Carabidae) Ground Beetle Worldwide.

Elucidating the basic life-history of endangered species is the first important step in the conservation of such species. This study examined the reproductive ecology and the preimaginal morphology of the endangered ground beetle Elaphrus sugai Nakane (Coleoptera: Carabidae); currently, the Watarase wetland of the central Kanto Plain, Japan is the only confirmed locality of this beetle species. Laboratory rearing of reproductive adults collected in early April revealed that females can lay more than 131 eggs. Eggs were laid in mud, without an egg chamber. Larvae reached adulthood when fed a diet of mealworms, indicating that E. sugai larvae are insect larvae feeders. An earthworm diet, the optimal diet for larvae of a congeneric species (E. punctatus Motschulsky), was lethal to E. sugai larvae. The egg stage was 3-4 days in duration under a 16L8D cycle (22°C). The duration from hatching to adult eclosion was 23-42 days at various temperatures simulating those of the reproductive period. Larval morphology was similar to that of consubgeneric species described previously. The pupa is unusual, in that the setae on the abdominal tergites are long (twice as long as those of the abdominal segment) and have somewhat "coiled" apices. Finally, the current endangered status of E. sugai was compared to that of E. viridis Horn, which has been regarded as the most endangered species of the genus worldwide.

Two new species of the ground beetle subgenus Sadonebria Ledoux & Roux, 2005 (Coleoptera, Carabidae, Nebria) from Japan and first description of larvae of the subgenus.

Sadonebria Ledoux & Roux, 2005 is one of the more diverse subgenera of the genus Nebria Latreille, 1802 in East Asia, and its taxonomy remains unrevised at the subgeneric and specific levels. In this paper, two new species of this subgenus are described from Japan. Nebria quinquelobata sp. n. is described from Mt. Myôkô and is externally similar to Nebria saeviens Bates, 1883, to which specimens of this new species previously had been assigned. Nebria yatsugatakensis sp. n. is described from the Yatsugatake Mountains and is externally similar to locally adjacent species that had been recognized as Nebria sadona Bates, 1883 and were recently revealed as separate species. Both new species are distinguished by morphological (the shape of the endophallus) and morphometric (geometric morphometrics of the pronotum and aedeagus) features. For Nebria yatsugatakensis, the morphology of all larval instars is described based on specimens reared from eggs laid by collected adults. These results, together with previous studies of the species-level taxonomy of Sadonebria and larval morphology of other Nebria subgenera, suggest (i) the utility of geometric morphometrics in species-level taxonomy; (ii) the importance of larval secondary setae in the subgeneric taxonomy of the genus Nebria; and (iii) the presence of further cryptic species in Sadonebria.

A novel technique for identifying the instar of field-collected insect larvae.

Many field studies of insects have focused on the adult stage alone, likely because immature stages are unknown in most insect species. Molecular species identification (e.g., DNA barcoding) has helped ascertain the immature stages of many insects, but larval developmental stages (instars) cannot be identified. The identification of the growth stages of collected individuals is indispensable from both ecological and taxonomic perspectives. Using a larval-adult body size relationship across species, I present a novel technique for identifying the instar of field-collected insect larvae that are identified by molecular species identification technique. This method is based on the assumption that classification functions derived from discriminant analyses, performed with larval instar as a response variable and adult and larval body sizes as explanatory variables, can be used to determine the instar of a given larval specimen that was not included in the original data set, even at the species level. This size relationship has been demonstrated in larval instars for many insects (Dyar's rule), but no attempt has been made to include the adult stage. Analysis of a test data set derived from the beetle family Carabidae (Coleoptera) showed that classification functions obtained from data sets derived from related species had a correct classification rate of 81-100%. Given that no reliable method has been established to identify the instar of field-collected insect larvae, these values may have sufficient accuracy as an analytical method for field-collected samples. The chief advantage of this technique is that the instar can be identified even when only one specimen is available per species if classification functions are determined for groups to which the focal species belongs. Similar classification functions should be created for other insect groups. By using those functions together with molecular species identification, future studies could include larval stages as well as adults.

Different patterns of oviposition learning in two closely related ectoparasitoid wasps with contrasting reproductive strategies.

Many parasitoid wasps learn host-associated cues and use them in subsequent host-searching behavior. This associative learning, namely "oviposition learning," has been investigated in many studies. However, few studies have compared multiple species, and no comparative study has previously been conducted on ectoparasitoid species. We compared the effects of oviposition learning on host preference and offspring sex ratio in two closely related ectoparasitoid wasps with contrasting reproductive strategies, Anisopteromalus calandrae (r-strategist) and its sibling species (K-strategist). Using two bruchine hosts, Callosobruchus chinensis and Callosobruchus maculatus larvae infesting the cowpea Vigna unguiculata, oviposition choice experiments were performed at high and low host densities. In both species, no conspicuous effect on the offspring sex ratio was detected, but effects on host preference were found to differ between the species. In A. calandrae, the effects were detected only at high host density, suggesting that oviposition learning plays a role in host discrimination from a short distance but not from a long distance. In the sibling species, those effects were not detected in any of the cases, suggesting the absence of oviposition learning. These results are compatible with those of previous comparative studies of endoparasitoid wasps in that few lifetime oviposition experiences and/or low reward per foraging decision result in low or absent oviposition learning ability. This finding may indicate that ecological traits contributing to learning ability are similar between endoparasitoid and ectoparasitoid wasps. Thus, our species comparison of ectoparasitoids provides another model system for investigating learning and memory dynamics in parasitoid wasps.

Field observations of climbing behavior and seed predation by adult ground beetles (Coleoptera: Carabidae) in a lowland area of the temperate zone.

Granivory is a specialized food habit in the predominantly carnivorous beetle family Carabidae. Most studies of carabid granivory have been conducted under laboratory conditions; thus, our knowledge of the feeding ecology of granivorous carabids in the field is insufficient. I conducted field observations of climbing behavior and seed predation by adult carabids in a lowland area of eastern Japan, from early October to late November in 2008. This is the first systematic field observation of the feeding ecology of granivorous carabids in the temperate zone. In total, 176 carabid individuals of 11 species were observed, with 108 individuals feeding on plant seeds/flowers. Each carabid species was primarily observed feeding on a particular plant species. Frequently observed combinations were: Amara gigantea Motschulsky on Humulus scandens (Loureiro) Merrill (Moraceae) seed, Amara lucens Baliani on Artemisia indica Willdenow (Asteraceae) flower, and Amara macronota (Solsky) and Harpalus (Pseudoophonus) spp. on Digitaria ciliaris (Retzius) Koeler (Poaceae) seed. In all but one species, the sex ratio of individuals observed feeding was female-biased. In Am. gigantea and Am. macronota, a larger proportion of females than males ate seeds. In the three Amara species, copulations on plants, with the female feeding on its seeds/flowers, were often observed. These observations may indicate that, whereas females climb onto plants to feed on seeds, males climb to seek females for copulation rather than forage. Because granivorous carabids play important roles as weed-control agents in temperate agro-ecosystems, the present results would provide valuable basic information for future studies on this subject.

Sperm bundle and reproductive organs of carabid beetles tribe Pterostichini (Coleoptera: Carabidae).

The morphological characteristics of sperm and reproductive organs may offer clues as to how reproductive systems have evolved. In this paper, the morphologies of the sperm and male reproductive organs of carabid beetles in the tribe Pterostichini (Coleoptera: Carabidae) are described, and the morphological associations among characters are examined. All species form sperm bundles in which the head of the sperm was embedded in a rod-shaped structure, i.e., spermatodesm. The spermatodesm shape (left-handed spiral, right-handed spiral, or without conspicuous spiral structure) and the condition of the sperm on the spermatodesm surface (with the tail free-moving or forming a thin, sheetlike structure) vary among species. In all species, the spiral directions of the convoluted seminal vesicles and vasa deferentia are the same on both sides of the body; that is, they show an asymmetric structure. The species in which the sperm bundle and the seminal vesicles both have a spiral structure could be classified into two types, with significant differences in sperm-bundle length between the two types. The species with a sperm-bundle spiral and seminal-vesicle spiral of almost the same diameter have longer sperm bundles than the species with a sperm-bundle spiral and seminal-vesicle tube of almost the same diameter. In the former type, the spiral directions of the sperm bundles and seminal vesicles are inevitably the same, whereas they differ in some species with the later type. Therefore, increased sperm bundle length appears to have been facilitated by the concordance of the sperm bundle's coiling direction with the coiling direction of the seminal vesicle.

Morphological association between spermatophores and male genitalia in carabid beetles of the tribe Pterostichini (Coleoptera: Carabidae).

A morphological association between genitalia and ejaculates could provide insight into the function and evolution of genitalia. In this study, the morphologies of the ejaculates and male genitalia of 15 species of Pterostichini and two species of Platynini (Coleoptera: Carabidae) are described. All the species examined formed a spermatophore, the morphology of which could be classified into three types based on its relative volume in the female vaginal cavity and the presence or absence of a pluglike conformation. Male genital morphology could be divided into two types by the direction of the endophallus and gonopore. Species with a strongly bent endophallus invariably formed a pluglike spermatophore. The results suggest that the peculiar shape of endophallus found in some species of Pterostichini may function in forming the pluglike structure of the spermatophore.

Phylogenetic studies of the subgenus Nialoe (s. lat.) (Coleoptera, Carabidae, Genus Pterostichus), part 1: Homology of the component parts of male genitalia and higher phylogeny.

A phylogenetic hypothesis of the subgenus Nialoe (s. lat.) of genus Pterostichus is proposed based on a cladistic analysis of seventeen morphological characters. Through comparative studies of the sclerites on the endophallus of male genitalia, it became apparent that the arrangement of these sclerites reflects seven different origins. The results show that Nialoe (s. lat.) is monophyletic and composed of four clades, while some traditional taxa are para- or polyphyletic, and are defined only by symplesiomorphies. Pterostichus (Nialoe) mosaicus sp. nov., which is indispensable for tracing the homology of sclerites in this subgenus, is described.

Phylogenetic studies of the subgenus Nialoe (s. lat.) (Coleoptera, Carabidae, Genus Pterostichus), part 2: The Asymmetricus species group.

Pterostichus (Nialoe) asymmetricus Bates and its allies are revised based on the membranous parts of their genitalia. Four new taxa, P. (N.) ovaliphallus sp. nov., P. (N.) fujimurai ibukiyamanus ssp. nov., P. (N.) shotaroi kiimontanus ssp. nov., and P. (N.) basilobatus sp. nov. are described. A cladistic analysis based on 31 morphological characters reveals that Daisenilaoe (s. str.) Nakane and Straneo is included as one clade within Nialoe (s. str.). The speciation and dispersal process of this species group are also discussed.