Deficiency of Trex1 leads to spontaneous development of type 1 diabetes.

BACKGROUND: Type 1 diabetes is believed to be an autoimmune condition, characterized by destruction of insulin-producing cells, due to the detrimental inflammation in pancreas. Growing evidences have indicated the important role of type I interferon in the development of type 1 diabetes. METHODS: Trex1-deficient rats were generated by using CRISPR-Cas9. The fasting blood glucose level of rat was measured by a Roche Accuchek blood glucose monitor. The levels of insulin, islet autoantibodies, and interferon-ß were measured using enzyme-linked immunosorbent assay. The inflammatory genes were detected by quantitative PCR and RNA-seq. Hematein-eosin staining was used to detect the pathological changes in pancreas, eye and kidney. The pathological features of kidney were also detected by Masson trichrome and periodic acid-Schiff staining. The distribution of islet cells, immune cells or ssDNA in pancreas was analyzed by immunofluorescent staining. RESULTS: In this study, we established a Trex1-deletion Sprague Dawley rat model, and unexpectedly, we found that the Trex1-/- rats spontaneously develop type 1 diabetes. Similar to human diabetes, the hyperglycemia in rats is accompanied by diabetic complications such as diabetic nephropathy and cataract. Mechanistical investigation revealed the accumulation of ssDNA and the excessive production of proinflammatory cytokines, including IFN-ß, in Trex1 null pancreas. These are likely contributing to the inflammation in pancreas and eventually leading to the decline of pancreatic ß cells. CONCLUSIONS: Our study links the DNA-induced chronic inflammation to the pathogenesis of type 1 diabetes, and also provides an animal model for type 1 diabetes studies.

Rosmarinic acid ameliorates autoimmune responses through suppression of intracellular nucleic acid-mediated type I interferon expression.

Recognition of intracellular nucleic acids is a vital step for host to mount prompt immune responses against microbial pathogens. However, inappropriate response to self-nucleic acids leads to sustained type I interferon (IFN) production, which is implicated in the development of several autoimmune diseases, such as Aicardi-Goutières syndrome (AGS). Therefore, effective confinement of intracellular nucleic acid-induced IFN expression is a potential strategy for the treatment of such autoimmune diseases. In this study, we found that rosmarinic acid (RA), a natural compound isolated from rosemary, inhibits intracellular nucleic acid-stimulated IFN expression. Mechanistic investigation revealed that RA binds to both G3BP1 and cGAS, and impairs cGAS activation through disrupting the binding of DNA with cGAS. More importantly, we showed that RA could effectively attenuate the expression of IFN-stimulated genes (ISGs) in the well-established cell models for AGS. Thus, our study provides a promising compound for the treatment of autoimmune responses induced by aberrant nucleic acid-sensing.

Pollinator sharing, copollination, and speciation by host shifting among six closely related dioecious fig species.

The obligate pollination mutualism between figs (Ficus, Moraceae) and pollinator wasps (Agaonidae, Hymenoptera) is a classic example of cospeciation. However, examples of phylogenetic incongruencies between figs and their pollinators suggest that pollinators may speciate by host shifting. To investigate the mechanism of speciation by host shifting, we examined the phylogenetic relationships and population genetic structures of six closely related fig species and their pollinators from southern China and Taiwan-Ryukyu islands using various molecular markers. The results revealed 1) an extraordinary case of pollinator sharing, in which five distinct fig species share a single pollinator species in southern China; 2) two types of copollination, namely, sympatric copollination by pollinator duplication or pollinator migration, and allopatric copollination by host migration and new pollinator acquisition; 3) fig species from southern China have colonized Taiwan repeatedly and one of these events has been followed by host shifting, reestablishment of host specificity, and pollinator speciation, in order. Based on our results, we propose a model for pollinator speciation by host shifting in which the reestablishment of host-specificity plays a central role in the speciation process. These findings provide important insights into understanding the mechanisms underlying pollinator speciation and host specificity in obligate pollination mutualism.

Three nuclear protein-coding genes corroborate a recent phylogenomic model of the Branchiopoda (Crustacea) and provide estimates of the divergence times of the major branchiopodan taxa.

The class Branchiopoda (Crustacea) shows great diversity in morphology and lifestyle among its constituent higher-level taxa: Anostraca, Notostraca, Laevicaudata, Spinicaudata, Cyclestherida and Cladocera. The phylogenetic relationships among these taxa have long been controversial. We sequenced three orthologous nuclear genes that encode the catalytic subunit of DNA polymerase delta and the largest and second-largest subunits of RNA polymerase II in the expectation that the amino acid sequences encoded by these genes might be effective in clarifying branchiopod phylogeny and estimating the times of divergence of the major branchiopodan taxa. The results of phylogenetic analyses based on these amino acid sequences support the monophyly of Branchiopoda and provide strong molecular evidence in support of the following phylogenetic relationships: (Anostraca, (Notostraca, (Laevicaudata, (Spinicaudata, (Cyclestherida, Cladocera))))). Within Cladocera, comparison of the nucleotide sequences of these same genes shows Ctenopoda to be the sister group of Haplopoda + Anomopoda. Three statistical tests based on the present amino acid sequence data-the approximately unbiased test, Kishino-Hasegawa test and weighted Shimodaira-Hasegawa test-tend to refute most of the previous molecular phylogenetic studies on Branchiopoda, which have placed Notostraca differently than here; however, our results corroborate those of one recent phylogenomic study, thus confirming the effectiveness of these three genes to investigate relationships among branchiopod higher taxa. Divergence time estimates calibrated on the basis of fossil evidence suggest that the first divergence of extant branchiopods occurred about 534 Ma during the early Cambrian period and that diversification within the extant branchiopod lineages started in or after the late Permian.

Evolutionary history of carabid ground beetles with special reference to morphological variations of the hind-wings.

Most beetles belonging to the subfamily Carabinae of the family Carabidae (so-called carabid ground beetles) cannot fly, because their hind-wings are highly degenerated. However, about half of the species in the subtribe Calosomina within the same subfamily can fly. From extensive morphological examinations of the hind-wings of Carabinae species in conjunction with DNA molecular phylogenetic trees, the process and possible causes of hind-wing degeneration in the Carabinae are discussed.

Invasion of the assassin bug Agriosphodrus dohrni (Hemiptera: Reduviidae) to Japan: Source estimation inferred from mitochondrial and nuclear gene sequences.

A large-sized assassin bug Agriosphodrus dohrni (Signoret), has been recorded from India, Vietnam, China and Japan. It is one of the potential biological control agents against some important agricultural and forest pests. This species is speculated to have invaded Japan from its native range in China about 60 years ago. We used three mitochondrial gene fragments (COI, Cytb, and ND5) and one nuclear gene fragment (EF-1α) to clarify the invasion history of A. dohrni and assess the effects of geographic events and associated ecological adaptation on the distribution pattern. The native populations of A. dohrni in China are divided into three distinct groups, which might be molded by the Early Pleistocene glaciation event and diverged during the Calabrian Stage. However, consistent with the hypothesis of a recent invasion, extremely low level of genetic variation was detected in the Japanese populations, with only two haplotypes for the combined mitochondrial genes. Both the splits network and the ML/BI phylogenetic trees revealed that haplotypes of Japan were more closely-related to those from eastern China. Therefore, we postulate that there has been only one introduction event, probably from somewhere around the Nanjing (NJ) and Lin'an (LA) populations of eastern China.

Silent evolution.

Phylogenetic analyses using mitochondrial DNA sequences of several kinds of beetles have shown that their evolution included a silent stage in which no morphological changes took place. We thus propose a new category of evolutionary process called "silent evolution".

Genome-wide sequence data suggest the possibility of pollinator sharing by host shift in dioecious figs (Moraceae, Ficus).

The obligate mutualism of figs and fig-pollinating wasps has been one of the classic models used for testing theories of co-evolution and cospeciation due to the high species-specificity of these relationships. To investigate the species-specificity between figs and fig pollinators and to further understand the speciation process in obligate mutualisms, we examined the genetic differentiation and phylogenetic relationships of four closely related fig-pollinating wasp species (Blastophaga nipponica, Blastophaga taiwanensis, Blastophaga tannoensis and Blastophaga yeni) in Japan and Taiwan using genome-wide sequence data, including mitochondrial DNA sequences. In addition, population structure was analysed for the fig wasps and their host species using microsatellite data. The results suggest that the three Taiwanese fig wasp species are a single panmictic population that pollinates three dioecious fig species, which are sympatrically distributed, have large differences in morphology and ecology and are also genetically differentiated. Our results illustrate the first case of pollinator sharing by host shift in the subgenus Ficus. On the other hand, there are strict genetic codivergences between allopatric populations of the two host-pollinator pairs. The possible processes that produce these pollinator-sharing events are discussed based on the level and pattern of genetic differentiation in these figs and fig wasps.

Relationship between tyrosine phosphorylation and protein expression of insulin receptor and insulin resistance in gestational diabetes mellitus.

The relationship between tyrosine phosphorylation (TP) and protein expression of insulin receptor (InsR) and insulin resistance (IR) in patients with gestational diabetes mellitus (GDM) was investigated. The InsR expression and TP in skeleton muscle tissue were determined by Western blotting and immunoprecipitation in women with GDM (GDM group, n=22), normal pregnant women (normal pregnancy group, n=22) and normal non-pregnant women (normal non-pregnant group, n=13). Fasting plasma glucose (FPG) and fasting insulin (FINS) were measured by oxidase assay and immunoradioassay. The results showed that the levels of FPG (5.61±0.78 mmol/L), FINS (15.42±5.13 mU/L) and Homeostasis model assessment-IR (HOMA-IR) (1.21±0.52) in GDM group were significantly higher than those in normal pregnancy group (4.43±0.46 mmol/L, 10.56±3.07 mU/L and 0.80±0.31 respectively) (P<0.01). The levels of FINS and HOMA-IR in normal pregnancy group were significantly higher than those in normal non-pregnant group (7.56±2.31 mU/L and 0.47±0.26 respectively) (P<0.01). There was no significant difference in the InsR expression level among the three groups (P>0.05). TP of InsR with insulin stimulation was significantly decreased in GDM group (0.20±0.05) as compared with normal pregnancy group (0.26±0.06) (P<0.01). TP of InsR with insulin stimulation in normal pregnancy group was lower than that in normal non-pregnant group (0.31±0.06) (P<0.01). TP of InsR with insulin stimulation was negatively related with HOMA-IR in GDM group (r=-0.525, P<0.01). There was no correlation between the protein expression of InsR and HOMA-IR in GDM group (r=-0.236, P>0.05). It was suggested that there is no significant correlation between the protein expression of InsR in skeletal muscle and IR in GDM, but changes in TP of InsR are associated with IR in GDM.

Molecular phylogeny of Myriapoda provides insights into evolutionary patterns of the mode in post-embryonic development.

Myriapoda, a subphylum of Arthropoda, comprises four classes, Chilopoda, Diplopoda, Pauropoda, and Symphyla. While recent molecular evidence has shown that Myriapoda is monophyletic, the internal phylogeny, which is pivotal for understanding the evolutionary history of myriapods, remains unresolved. Here we report the results of phylogenetic analyses and estimations of divergence time and ancestral state of myriapods. Phylogenetic analyses were performed based on three nuclear protein-coding genes determined from 19 myriapods representing the four classes (17 orders) and 11 outgroup species. The results revealed that Symphyla whose phylogenetic position has long been debated is the sister lineage to all other myriapods, and that the interordinal relationships within classes were consistent with traditional classifications. Ancestral state estimation based on the tree topology suggests that myriapods evolved from an ancestral state that was characterized by a hemianamorphic mode of post-embryonic development and had a relatively low number of body segments and legs.

Molecular phylogenetic analyses support the monophyly of Hexapoda and suggest the paraphyly of Entognatha.

BACKGROUND: Molecular phylogenetic analyses have revealed that Hexapoda and Crustacea form a common clade (the Pancrustacea), which is now widely accepted among zoologists; however, the origin of Hexapoda remains unresolved. The main problems are the unclear relationships among the basal hexapod lineages, Protura (proturans), Collembola (springtails), Diplura (diplurans), and Ectognatha (bristletails, silverfishes, and all winged insects). Mitogenomic analyses have challenged hexapod monophyly and suggested the reciprocal paraphyly of Hexapoda and Crustacea, whereas studies based on nuclear molecular data support the monophyletic origin of hexapods. Additionally, there are significant discrepancies with respect to these issues between the results of morphological and molecular studies. To investigate these problems, we performed phylogenetic analyses of Pancrustacea based on the protein sequences of three orthologous nuclear genes encoding the catalytic subunit of DNA polymerase delta and the largest and second largest subunits of RNA polymerase II from 64 species of arthropods, including representatives of all hexapod orders. RESULTS: Phylogenetic analyses were conducted based on the inferred amino acid (aa) sequences (~3400 aa in total) of the three genes using the maximum likelihood (ML) method and Bayesian inference. Analyses were also performed with additional datasets generated by excluding long-branch taxa or by using different outgroups. These analyses all yielded essentially the same results. All hexapods were clustered into a common clade, with Branchiopoda as its sister lineage, whereas Crustacea was paraphyletic. Within Hexapoda, the lineages Ectognatha, Palaeoptera, Neoptera, Polyneoptera, and Holometabola were each confirmed to be monophyletic with robust support, but monophyly was not supported for Entognatha (Protura + Collembola + Diplura), Ellipura (Protura + Collembola), or Nonoculata (Protura + Diplura). Instead, our results showed that Protura is the sister lineage to all other hexapods and that Diplura or Diplura + Collembola is closely related to Ectognatha. CONCLUSION: This is the first study to include all hexapod orders in a phylogenetic analysis using multiple nuclear protein-coding genes to investigate the phylogeny of Hexapoda, with an emphasis on Entognatha. The results strongly support the monophyletic origin of hexapods but reject the monophyly of Entognatha, Ellipura, and Nonoculata. Our results provided the first molecular evidence in support of Protura as the sister group to other hexapods. These findings are expected to provide additional insights into the origin of hexapods and the processes involved in the adaptation of insects to life on land.

Phylogenetic analyses suggest a hybrid origin of the figs (Moraceae: Ficus) that are endemic to the Ogasawara (Bonin) Islands, Japan.

The Ogasawara Islands are oceanic islands and harbor a unique endemic flora. There are three fig species (Ficus boninsimae, F. nishimurae and F. iidaiana) endemic to the Ogasawara Islands, and these species have been considered to be closely related to Ficus erecta, and to have diverged within the islands. However, this hypothesis remains uncertain. To investigate this issue, we assessed the phylogenetic relationships of the Ogasawara figs and their close relatives occurring in Japan, Taiwan and South China based on six plastid genome regions, nuclear ITS region and two nuclear genes. The plastid genome-based tree indicated a close relationship between the Ogasawara figs and F. erecta, whereas some of the nuclear gene-based trees suggested this relationship was not so close. In addition, the phylogenetic analyses of the pollinating wasps associated with these fig species based on the nuclear 28S rRNA and mitochondrial cytB genes suggested that the fig-pollinating wasps of F. erecta are not sister to those of the Ogasawara figs These results suggest the occurrence of an early hybridization event(s) in the lineage leading to the Ogasawara figs.

Ecology of hemiepiphytism in fig species is based on evolutionary correlation of hydraulics and carbon economy.

Woody hemiepiphytic species (Hs) are important components of tropical rain forests, and they have been hypothesized to differ from non-hemiepiphytic tree species (NHs) in adaptations relating to water relations and carbon economy; but few studies have been conducted comparing ecophysiological traits between the two growth forms especially in an evolutionary context. Using common-garden plants of the genus Ficus, functional traits related to plant hydraulics and carbon economy were compared for seven NHs and seven Hs in their adult terrestrial "tree-like" growth phase. We used phylogenetically independent contrasts to test the hypothesis that differences in water availability selected for contrasting suites of traits in Hs and NHs, driving evolutionary correlations among functional traits including hydraulic conductivity and photosynthetic traits. Species of the two growth forms differed in functional traits; Hs had substantially lower xylem hydraulic conductivity and stomatal conductance, and higher instantaneous photosynthetic water use efficiency. Leaf morphological and structural traits also differed strikingly between the two growth forms. The Hs had significantly smaller leaves, higher leaf mass per area (LMA), and smaller xylem vessel lumen diameters. Across all the species, hydraulic conductivity was positively correlated with leaf gas exchange indicating high degrees of hydraulic-photosynthetic coordination. More importantly, these correlations were supported by correlations implemented on phylogenetic independent contrasts, suggesting that most trait correlations arose through repeated convergent evolution rather than as a result of chance events in the deep nodes of the lineage. Vatiation in xylem hydraulic conductivity was also centrally associated with a suite of other functional traits related to carbon economy and growth, such as LMA, water use efficiency, leaf nutrient concentration, and photosynthetic nutrient use efficiency, indicating important physiological constraints or trade-offs among functional traits. Shifts in this trait cluster apparently related to the adaptation to drought-prone canopy growth during the early life cycle of Hs and clearly affected ecophysiology of the later terrestrial stage of these species. Evolutionary flexibility in hydraulics and associated traits might be one basis for the hyper-diversification of Ficus species in tropical rain forests.

Phylogenetic relationships among insect orders based on three nuclear protein-coding gene sequences.

Many attempts to resolve the phylogenetic relationships of higher groups of insects have been made based on both morphological and molecular evidence; nonetheless, most of the interordinal relationships of insects remain unclear or are controversial. As a new approach, in this study we sequenced three nuclear genes encoding the catalytic subunit of DNA polymerase delta and the two largest subunits of RNA polymerase II from all insect orders. The predicted amino acid sequences (In total, approx. 3500 amino acid sites) of these proteins were subjected to phylogenetic analyses based on the maximum likelihood and Bayesian analysis methods with various models. The resulting trees strongly support the monophyly of Palaeoptera, Neoptera, Polyneoptera, and Holometabola, while within Polyneoptera, the groupings of Isoptera/"Blattaria"/Mantodea (Superorder Dictyoptera), Dictyoptera/Zoraptera, Dermaptera/Plecoptera, Mantophasmatodea/Grylloblattodea, and Embioptera/Phasmatodea are supported. Although Paraneoptera is not supported as a monophyletic group, the grouping of Phthiraptera/Psocoptera is robustly supported. The interordinal relationships within Holometabola are well resolved and strongly supported that the order Hymenoptera is the sister lineage to all other holometabolous insects. The other orders of Holometabola are separated into two large groups, and the interordinal relationships of each group are (((Siphonaptera, Mecoptera), Diptera), (Trichoptera, Lepidoptera)) and ((Coleoptera, Strepsiptera), (Neuroptera, Raphidioptera, Megaloptera)). The sister relationship between Strepsiptera and Diptera are significantly rejected by all the statistical tests (AU, KH and wSH), while the affinity between Hymenoptera and Mecopterida are significantly rejected only by AU and KH tests. Our results show that the use of amino acid sequences of these three nuclear genes is an effective approach for resolving the relationships of higher groups of insects.

Molecular phylogenies of figs and fig-pollinating wasps in the Ryukyu and Bonin (Ogasawara) islands, Japan.

The interaction between figs (Ficus, Moraceae) and fig-pollinating wasps (Chalcidoidea, Agaonidae) is one of the most specific mutualisms, and thus is a model system for studying coevolution and cospeciation. In this study we focused on figs and their associated fig-wasps found in the Ryukyu and Bonin (Ogasawara) Islands, Japan, because it has been suggested that breakdown in the specificity may occur in islands or at edge of a species' distribution. We collected 136 samples of 15 native fig species and 95 samples of 13 associated fig-wasps from all major islands in the Ryukyu Islands, including two fig species and one fig-wasp species endemic to the Bonin Islands. We performed molecular phylogenetic analyses using plastid DNA and nuclear ITS sequences for the figs and nuclear 28S rRNA and mitochondrial COI genes for the fig-wasps to investigate the interspecific phylogenies and intraspecific variation within the mutualism. Our phylogenetic analyses using multiple samples per species show the single clade of each fig (except the Bonin endemic species) and fig-pollinating wasp species. Fig species belonging to the same subgenera formed well-supported clades in both plastid and ITS trees, except for the subgenus Urostigma. Likewise, fig wasps emerging from host fig species belonging to the same subgenera formed mostly well supported clades in both 28S and COI trees. Host specificity between the figs and fig-wasps functions strictly in these islands. There was very little sequence variation within species, and that no major geographic structure was found. The two Bonin endemic species (F. boninsimae and F. nishimurae) or their common ancestor and the associated fig-wasps (Blastophaga sp.) are apparently derived from F. erecta and its associated fig-wasps (B. nipponica), respectively, and probably migrated from the Ryukyu Islands.

Ancient divergence of animal protein tyrosine kinase genes demonstrated by a gene family tree including choanoflagellate genes.

Animal-specific gene families involved in cell-cell communication and developmental control comprise many subfamilies with distinct domain structures and functions. They diverged by subfamily-generating duplications and domain shufflings before the parazoan-eumetazoan split. Here, we have cloned 40 PTK cDNAs from choanoflagellates, Monosiga ovata, Stephanoeca diplocostata and Codosiga gracilis, the closest relatives to animals. A phylogeny-based analysis of PTKs revealed that 40 out of 47 subfamilies analyzed have unique domain structures and are possibly generated independently in animal and choanoflagellate lineages by domain shufflings. Seven cytoplasmic subfamilies showed divergence before the animal-choanoflagellate split originated by both duplications and shufflings.

Multiple receptor-like kinase cDNAs from liverwort Marchantia polymorpha and two charophycean green algae, Closterium ehrenbergii and Nitella axillaris: Extensive gene duplications and gene shufflings in the early evolution of streptophytes.

Plant receptor-like kinases (RLKs) comprise a large family with more than several hundred members in vascular plants. The RLK family is thought to have diverged specifically in the plant kingdom, and no family member has been identified in other lineages except for animals and Plasmodium, both of which have RLK related families of small size. To know the time of divergence of RLK family members by gene duplications and domain shufflings, comprehensive isolations of RLK cDNAs were performed from a nonvascular plant, liverwort Marchantia polymorpha and two charophycean green algae, Closterium ehrenbergii, and Nitella axillaris, thought to be the closest relatives to land plants. We obtained twenty-nine, fourteen, and thirteen RLK related cDNAs from M. polymorpha, C. ehrenbergii, and N. axillaris, respectively. The amino acid sequences of these RLKs were compared with those of vascular plants, and phylogenetic trees were inferred by GAMT, a genetic algorithm-based maximum likelihood (ML) method that outputs multiple trees, together with best one. The inferred ML trees revealed ancient gene duplications generating subfamilies with different domain organizations, which occurred extensively at least before the divergence of vascular and nonvascular plants. Rather it remains possible that the extensive gene duplications occurred during the early evolution of streptophytes. Multicellular-specific somatic embryogenesis receptor kinase (SERK) involved in somatic embryogenesis was found in a unicellular alga C. ehrenbergii, suggesting the evolution of SERK by gene recruitment of a unicellular gene.

Establishment of hybrid-derived offspring populations in the Ohomopterus ground beetles through unidirectional hybridization.

An approach to deduce the mechanism of stabilization of the hybrid-derived populations in the Ohomopterus ground beetles has been made by comparative studies on the phylogenetic trees of the mitochondrial and nuclear DNA. A phylogenetic tree based on the internal transcribed spacer (ITS) of nuclear ribosomal gene roughly reflects the relations of morphological species group, while mitochondrial NADH dehydrogenase subunit 5 (ND5) gene shows a considerable different topology on the tree; there exist several geographically-linked lineages, most of which consist of more than one species. These results suggest that the replacement of mitochondria has occurred widely in the Ohomopterus species. In most cases, hybridization is unidirectional, i.e., the species A (♂) hybridized with another species B (♀) and not vice versa, with accompanied replacement of mitochondria of A by those of B. The results also suggest that partial or complete occupation of the distribution territory by a hybrid-derived morphological species. The morphological appearance of the resultant hybrid-derivatives are recognized as that of the original species A. Emergence of a morphological new species from a hybrid-derived population has been exemplified.

Evolutionary history of Calosomina ground beetles (Coleoptera, Carabidae, Carabinae) of the world as deduced from sequence comparisons of the mitochondrial ND 5 gene.

We deduced the phylogenetic relationships of 54 individuals representing 27 species of the Calosomina (Coleoptera, Carabidae) from various regions of the world from the mitochondrial NADH dehydrogenase subunit 5 (ND 5) gene sequences. The results suggest that these Calosomina radiated into 17 lineages within a short time about 30 million years ago (Mya). Most of the lineages are composed of a single genus containing only one or a few species. In some cases, several species classified into the same genus (e.g., Calosoma maximowiczi, Calos. inquisitor and Calos. frigidum) appear separately in independent lineages, while in others a series of species classified into different genera fall into one lineage (e.g., Chrysostigma calidum, Blaptosoma chihuahua, Microcallisthenes wilkesi and Callisthenes spp.). Based on this molecular phylogeny and morphological data, the probable evolutionary history and mode of morphological differentiation of the Calosomina are discussed.

Phylogeny and evolution of Digitulati ground beetles (Coleoptera, Carabidae) inferred from mitochondrial ND5 gene sequences.

Genealogical trees have been constructed using mitochondrial ND5 gene sequences of 87 specimens consisting of 32 species which have been believed to belong to the division Digitulati (one of the lineages of the subtribe Carabina) of the world. There have been recognized six lineages, which are well separated from each other. Each lineage contains the following genus: (1) the lineage A: Ohomopterus from Japan; (2) the lineage B: Isiocarabus from eastern Eurasian Continent; (3) the lineage C: Carabus from China which are further subdivided into three sublineages; (4) the lineage D: Carabus from USA; (5) the lineage E: Carabus from the Eurasian Continent, Japan and North America; and (6) the lineage F: Eucarabus from the Eurasian Continent. Additionally, the genus Acrocarabus which had been treated as a constituent of the division Archicarabomorphi has been recognized to be the 7th lineage of the division Digitulati from the ND5 genealogical analysis as well as morphology. These lineages are assumed to have radiated within a short period and are largely linked to their geographic distribution.