Evaluation of the host specificity of Eimeria uekii and Eimeria raichoi for Japanese rock ptarmigans by oocyst transfer to taxonomically related birds.

Eimeria spp. are protozoan parasites that are commonly found in a broad range of vertebrate hosts. These parasites generally exhibit strict host specificity, but some Eimeria spp. can infect groups of closely related species such as species within a genus or family. Compared with Eimeria spp. that infect livestock, limited information is available about such infections in wild animals including data on host specificity, virulence, and prevalence. The Japanese rock ptarmigan, Lagopus muta japonica, is an endangered bird belonging to the family Phasianidae, order Galliformes, and inhabits only alpine areas of Japan. In conservation efforts for these birds, two Eimeria spp., E. uekii and E. raichoi, were frequently detected. Here, we examined cross-transmission of the parasites to other bird species to characterize their infectivity as well as the development of experimental bird models to contribute to conservation programs by the oocyst transfer. Consequently, among the examined eight bird species (chickens, Japanese pheasants, turkeys, chukar partridges, quails, helmeted guineafowls and ducks), only turkeys (family Phasianidae, order Galliformes) could be infected with E. raichoi. However, the number of oocysts per feces was relatively low, and few parasites in the intestinal mucosa could be found by histopathological analyses. These results might indicate that E. uekii and E. raichoi are highly adapted to Japanese rock ptarmigans that inhabit the alpine zone although further studies are anticipated.

Predator presence and recent climatic warming raise body temperatures of island lizards.

In ectothermic predator-prey relationships, evasion of predation by prey depends on physiological and behavioural responses relating to the thermal biology of both predator and prey. On Japan's Izu Islands, we investigated a prey lizard's physiological and thermal responses to the presence of a snake predator over geologic time in addition to recent climatic warming. Foraging lizard body temperatures increased by 1.3 °C from 1981 to 2019 overall, yet were 2.9 °C warmer on snake islands relative to snake-free islands. We also detected snake predator-induced selection on hind leg length, which in turn is a major determinant for sprint speed only in lizard populations exposed to predation by snakes. Accordingly, we found that warmer prey body temperatures result in faster sprint speeds by the prey at temperatures suboptimal for the snake predator, and therefore contribute to escaping predation. Given recent climatic change, further warming could irrevocably alter this and other ectothermic predator-prey relationships.

Spatial optimization of invasive species control informed by management practices.

Optimization of spatial resource allocation is crucial for the successful control of invasive species under a limited budget but requires labor-intensive surveys to estimate population parameters. In this study, we devised a novel framework for the spatially explicit optimization of capture effort allocation using state-space population models from past capture records. We applied it to a control program for invasive snapping turtles to determine effort allocation strategies that minimize the population density over the whole area. We found that spatially heterogeneous density dependence and capture pressure limit the abundance of snapping turtles. Optimal effort allocation effectively improved the control effect, but the degree of improvement varied substantially depending on the total effort. The degree of improvement by the spatial optimization of allocation effort was only 3.21% when the total effort was maintained at the 2016 level. However, when the total effort was increased by two, four, and eight times, spatial optimization resulted in improvements of 4.65%, 8.33%, and 20.35%, respectively. To achieve the management goal for snapping turtles in our study area, increasing the current total effort by more than four times was necessary, in addition to optimizing the spatial effort. The snapping turtle population is expected to reach the target density one year after the optimal management strategy is implemented, and this rapid response can be explained by high population growth rate coupled with density-dependent feedback regulation. Our results demonstrated that combining a state-space model with optimization makes it possible to adaptively improve the management of invasive species and decision-making. The method used in this study, based on removal records from an invasive management program, can be easily applied to monitoring data for wildlife and pest control management using traps in a variety of ecosystems.

Embryonic developmental process governing the conspicuousness of body stripes and blue tail coloration in the lizard Plestiodon latiscutatus.

The combination of body stripes and vivid blue tail color has independently evolved in different lizard families. To understand how and when lizards developed this coloration, we microscopically compared the embryonic development of pigment cells in two island populations of Plestiodon latiscutatus that exhibit either striped and blue tailed or inconspicuously striped and blue tailed juveniles, based on the newly determined 12 normal developmental stages of embryos from shortly after egg laying to just before hatching. We focus on the role of the melanophores in the body stripe and the role of iridophore morphotypes in the extent of blue tail coloration. The melanophore density in the black background region was higher in lizards with vivid stripes than in lizards with drab stripes. Iridophores started to fill the dermal space that was not yet occupied by melanophores, which resulted in a higher iridophore density in stripes than in the inter-stripe regions. We also discovered that iridophores with thin platelets reflecting blue structural coloration appeared at a specific region in the tail at stage 11. The position of the tail where iridophores emerged coincided with the boundary area separating anterior brown and green colored tail from posterior blue colored tail after hatching.

Phylogeny mandalas for illustrating the Tree of Life.

A circular phylogeny with photos or drawings of species is named a phylogeny mandala. This is one of the ways for illustrating the Tree of Life, and is suitable to show visually how the biodiversity has developed in the course of evolution as clarified by the molecular phylogenetics. To demonstrate the recent progress of molecular phylogenetics, six phylogeny mandalas for various taxonomic groups of life were presented; i.e., (1) Eukaryota, (2) Metazoa, (3) Hexapoda, (4) Tetrapoda, (5) Eutheria, and (6) Primates.

Phylogeny mandalas of birds using the lithographs of John Gould's folio bird books.

The phylogeny mandala, which is a circular phylogeny with photos or drawings of species, is a suitable way to show visually how the biodiversity has developed in the course of evolution as clarified by the molecular phylogenetics. In this article, in order to demonstrate the recent progress of avian molecular phylogenetics, six phylogeny mandalas of various taxonomic groups of birds are presented with the lithographs of John Gould's folio bird books; i.e., (1) whole Aves, (2) Passeriformes, (3) Paradisaeidae in Corvoidea (Passeriformes), (4) Meliphagoidea (Passeriformes), (5) Trochili in Apodiformes, and (6) Galliformes.

Was the universal common ancestry proved?

The question of whether or not all life on Earth shares a single common ancestor has been a central problem of evolutionary biology since Darwin. Although the theory of universal common ancestry (UCA) has gathered a compelling list of circumstantial evidence, as given in ref. 2, there has been no attempt to test statistically the UCA hypothesis among the three domains of life (eubacteria, archaebacteria and eukaryotes) by using molecular sequences. Theobald recently challenged this problem with a formal statistical test, and concluded that the UCA hypothesis holds. Although his attempt is the first step towards establishing the UCA theory with a solid statistical basis, we think that the test of Theobald is not sufficient enough to reject the alternative hypothesis of the separate origins of life, despite the Akaike information criterion (AIC) of model selection giving a clear distinction between the competing hypotheses.

Polymorphism and evolution of ribosomal DNA in tea (Camellia sinensis, Theaceae).

Ribosomal DNA (rDNA) repeats often exhibit a high level of within-species homogeneity, but intra-individual polymorphism of rDNA has been found in Camellia species. In order to reveal the evolutionary pattern of rDNA repeats in tea [Camellia sinensis (L.) O. Kuntze], we identified the 45S rDNA loci, estimated their copy number, and cloned partial regions of them from different PCR products and from digested genomic DNA. The results show that there are 3 loci (6 sites) containing 45S rDNA in the tea genome; they are located at the ends of the short arms of 6 chromosomes and consist of a maximum of ∼6500 repeat units. On one hand, 164 sequences of the 26S rDNA cloned from PCR products contained 147 haplotypes, consisting of 62% pseudogenes, 24% putative functional genes, and 14% PCR-mediated recombinants, suggesting that the 45S rDNA of tea maintains an extremely high level of polymorphism and divergence; on the other hand, rDNA fragments cloned directly from genomic DNA exhibited a very high level of homogeneity: only one of 33 rDNA fragments was from a pseudogene. These results demonstrate that although over 60% of the 26S rDNA sequences identified in PCR products belong to pseudogenes, most 45S rDNA repeats are functional genes and have undergone concerted evolution. In this study, strong PCR bias and PCR-mediated recombination greatly increased the apparent proportion of pseudogenes in PCR products. Phylogenetic analysis and genetic divergence values for 26S rDNA sequences obtained in this study show that many pseudogenes have originated independently from functional genes at different times, and despite thus escaping from concerted evolution, they have failed to be eliminated from the tea genome over a long period, some of them having even produced addition copies by rapid expansion. Importantly, our study suggests that in order to determine the true pattern of evolution of rDNA it is necessary to combine data from more than one method rather than relying only on sequences from PCR products.

Nursing care process for releasing psychiatric inpatients from long-term seclusion in Japan: modified grounded theory approach.

Based on a modified grounded theory approach, in this study, we sought to elucidate the nursing care process used to guide psychiatric inpatients in long-term seclusion towards release from seclusion. Participant observations and interviews were conducted with a total of 18 nurses from three long-term psychiatric wards at two institutions from September 2011 to November 2012, to collect data on the nursing care they provided for psychiatric patients in long-term seclusion. Consequently, four categories and 15 concepts were extracted. The nurses viewed "a mature therapeutic environment that utilizes flexible apportionment of care" as the foundation (i.e. the core category) in guiding psychiatric inpatients towards release from long-term seclusion. The results revealed a care structure in which nurses in such a treatment environment provided care by flexible apportionment of three types of care: care aimed at avoiding mental and physical exhaustion, standardized care that does not confer a disadvantage to patients, and immediately responding to prevent problematic behaviors.

The Plasmodium apicoplast genome: conserved structure and close relationship of P. ovale to rodent malaria parasites.

Apicoplast, a nonphotosynthetic plastid derived from secondary symbiotic origin, is essential for the survival of malaria parasites of the genus Plasmodium. Elucidation of the evolution of the apicoplast genome in Plasmodium species is important to better understand the functions of the organelle. However, the complete apicoplast genome is available for only the most virulent human malaria parasite, Plasmodium falciparum. Here, we obtained the near-complete apicoplast genome sequences from eight Plasmodium species that infect a wide variety of vertebrate hosts and performed structural and phylogenetic analyses. We found that gene repertoire, gene arrangement, and other structural attributes were highly conserved. Phylogenetic reconstruction using 30 protein-coding genes of the apicoplast genome inferred, for the first time, a close relationship between P. ovale and rodent parasites. This close relatedness was robustly supported using multiple evolutionary assumptions and models. The finding suggests that an ancestral host switch occurred between rodent and human Plasmodium parasites.

The effect of pioneer carrion beetles on the emission of volatile organic compounds and carrion insect community assembly.

Mechanisms of carrion insect succession have been interpreted separately from interspecific interactions between early and later colonists or from changes in volatile organic compounds perceived by insects resulting from the progression of decomposition. To link these perspectives, we examined through laboratory and field experiments whether the modification of volatile organic compounds by early colonists could be a mechanism of succession. In the laboratory experiment, we used Necrophila japonica (Coleoptera, Staphylinidae) as an early colonist and examined its effects on the emissions of important volatile attractants for carrion insects, dimethyl disulfide (DMDS) and dimethyl trisulfide (DMTS) from carcasses. We collected DMDS and DMTS, using the static headspace method, under the following conditions: (i) rat carcass, (ii) rat carcass with artificial damage to the abdomen, (iii) rat carcass fed on by 10 Ne. japonica individuals, and (iv) 10 Ne. japonica individuals, and analyzed the collected gases using a gas chromatograph-mass spectrometer. After 12 and 30 h, carcasses fed on by Ne. japonica emitted higher concentrations of DMDS and DMTS than in other conditions. In the field experiment, we examined the effects of DMDS and DMTS on the attraction of carrion insects using traps baited with a mixture of DMDS and DMTS, hexane (odors unrelated to carcasses), or an empty microtube. Traps baited with DMDS and DMTS attracted more necrophagous species and individuals than traps not baited with this combination. These results showed that accelerated emissions of DMDS and DMTS from carcasses caused by early colonists may contribute to community assembly during carrion insect succession.

Domestication relaxed selective constraints on the yak mitochondrial genome.

Wild yaks roaming in the high-altitude Qinghai-Tibetan plateau have to maintain high metabolic efficiency. However, domestic yaks do not require such high efficiency because of their limited activity. Hence, domestication may have caused the relaxation of selective constraints on the yak mitochondrial genome because mitochondrial mutations are extremely sensitive to energy-related selective pressures. We have tested this hypothesis by analyzing the mitochondrial genomes of 51 domestic yaks and 21 wild yaks. The results show that the ratio of nonsynonymous/synonymous substitutions in mitochondrial protein-coding genes is significantly higher in domestic yak lineages than those of wild yaks. This genetic difference suggests that the relaxation of selective constraints following the domestication in addition to bottlenecks has allowed faster accumulation of nonsilent substitutions in the yak mitochondrial genome, despite its short domestication history.

Phylogenomic datasets provide both precision and accuracy in estimating the timescale of placental mammal phylogeny.

The fossil record suggests a rapid radiation of placental mammals following the Cretaceous-Paleogene (K-Pg) mass extinction 65 million years ago (Ma); nevertheless, molecular time estimates, while highly variable, are generally much older. Early molecular studies suffer from inadequate dating methods, reliance on the molecular clock, and simplistic and over-confident interpretations of the fossil record. More recent studies have used Bayesian dating methods that circumvent those issues, but the use of limited data has led to large estimation uncertainties, precluding a decisive conclusion on the timing of mammalian diversifications. Here we use a powerful Bayesian method to analyse 36 nuclear genomes and 274 mitochondrial genomes (20.6 million base pairs), combined with robust but flexible fossil calibrations. Our posterior time estimates suggest that marsupials diverged from eutherians 168-178 Ma, and crown Marsupialia diverged 64-84 Ma. Placentalia diverged 88-90 Ma, and present-day placental orders (except Primates and Xenarthra) originated in a ∼20 Myr window (45-65 Ma) after the K-Pg extinction. Therefore we reject a pre K-Pg model of placental ordinal diversification. We suggest other infamous instances of mismatch between molecular and palaeontological divergence time estimates will be resolved with this same approach.

Angiogenesis inhibitors identified by cell-based high-throughput screening: synthesis, structure-activity relationships and biological evaluation of 3-[(E)-styryl]benzamides that specifically inhibit endothelial cell proliferation.

Proliferation of endothelial cells is critical for angiogenesis. We report orally available, in vivo active antiangiogenic agents which specifically inhibit endothelial cell proliferation. After identifying human umbilical vein endothelial cell (HUVEC) proliferation inhibitors from a cell-based high-throughput screening (HTS), we eliminated those compounds which showed cytotoxicity against HCT116 and vascular endothelial growth factor receptor 2 (VEGFR-2) inhibitory activity. Evaluations in human Calu-6 xenograft model delivered lead compound 1. Following extensive lead optimization and alteration of the scaffold we discovered 32f and 32g, which both inhibited the proliferation and tube formation of HUVEC without showing inhibitory activity against any of 25 kinases or cytotoxicity against either normal fibroblasts or 40 cancer cell lines. Upon oral administration, 32f and 32g had good pharmacokinetic profiles and potent antitumor activity and decreased microvessel density (MVD) in Calu-6 xenograft model. Combination therapy with a VEGFR inhibitor enhanced the in vivo efficacy. These results suggest that 32f and 32g may have potential for use in cancer treatment.

Some problems in proving the existence of the universal common ancestor of life on Earth.

Although overwhelming circumstantial evidence supports the existence of the universal common ancestor of all extant life on Earth, it is still an open question whether the universal common ancestor existed or not. Theobald (Nature 465, 219-222 (2010)) recently challenged this problem with a formal statistical test applied to aligned sequences of conservative proteins sampled from all domains of life and concluded that the universal common ancestor hypothesis holds. However, we point out that there is a fundamental flaw in Theobald's method which used aligned sequences. We show that the alignment gives a strong bias for the common ancestor hypothesis, and we provide an example that Theobald's method supports a common ancestor hypothesis for two apparently unrelated families of protein-encoding sequences (cytb and nd2 of mitochondria). This arouses suspicion about the effectiveness of the "formal" test.

The position of gnetales among seed plants: overcoming pitfalls of chloroplast phylogenomics.

The phylogenetic position of Gnetales is one of the most contentious issues in the seed plant systematics. To elucidate the Gnetales position, an improved amino acid substitution matrix was estimated based on 64 chloroplast (cp) genomes and was applied to cp genome data including all three lineages of Gnetales in maximum likelihood analyses of proteins. Although the initial analysis strongly supported the sister relation of Gnetales with Cryptomeria (Cupressophyta or non-Pinaceae conifers) (the "Gnecup" hypothesis), the support seems to be caused by a long-branch attraction (LBA) artifact. Indeed, by removing fastest evolving proteins that are most likely associated with the LBA, the support drastically declined. Furthermore, another analysis of partial genome data with dense taxon sampling of conifers showed that, in psbC, rpl2, and rps7 proteins, there are many parallel amino acid substitutions between the lineages leading to Gnetales and to Cryptomeria, and by further excluding these three genes, the sister relation of Gnetales with Pinaceae (the "Gnepine" hypothesis) became supported. Overall, our analyses indicate that the LBA and parallel substitutions cause a seriously biased inference of phylogenetic position of Gnetales with the cp genome data.

The reconstituted 'humanized liver' in TK-NOG mice is mature and functional.

To overcome the limitations of existing models, we developed a novel experimental in vivo platform for replacing mouse liver with functioning human liver tissue. To do this, a herpes simplex virus type 1 thymidine kinase (HSVtk) transgene was expressed within the liver of highly immunodeficient NOG mice (TK-NOG). Mouse liver cells expressing this transgene were ablated after a brief exposure to a non-toxic dose of ganciclovir (GCV), and transplanted human liver cells are stably maintained within the liver (humanized TK-NOG) without exogenous drug. The reconstituted liver was shown to be a mature and functioning "human organ" that had zonal position-specific enzyme expression and a global gene expression pattern representative of mature human liver; and could generate a human-specific profile of drug metabolism. The 'humanized liver' could be stably maintained in these mice with a high level of synthetic function for a prolonged period (8 months). This novel in vivo system provides an optimized platform for studying human liver physiology, including drug metabolism, toxicology, or liver regeneration.

Development of nuclear SSRs for the insular shrub Clerodendrum izuinsulare (Verbenaceae) and the widespread C. trichotomum.

PREMISE OF THE STUDY: Nuclear microsatellite primers were developed in the insular shrub Clerodendrum izuinsulare and the closely related widespread C. trichotomum to provide molecular tools to undertake a comparative study of the reproductive systems and genetic structures of the two Clerodendrum species. METHODS AND RESULTS: Using an improved protocol for isolating codominant compound microsatellite markers, 19 primer sets were developed in C. izuinsulare and C. trichotomum. The primers amplified dinucleotide repeats with one to nine alleles per locus in C. izuinsulare and one to 15 alleles per locus in C. trichotomum. Tests of cross-amplification showed that four to 16 loci could be amplified using these markers in five other species of Clerodendrum that were studied. CONCLUSIONS: The microsatellite markers described here will be useful for comparative study of the reproductive systems and genetic structures of the narrow endemic C. izuinsulare and the widespread C. trichotomum.

Population genetic structure of raccoons as a consequence of multiple introductions and range expansion in the Boso Peninsula, Japan.

The raccoon (Procyon lotor) is an invasive carnivore that invaded various areas of the world. Although controlling feral raccoon populations is important to reduce serious threats to local ecosystems, raccoons are not under rigid population control in Europe and Japan. We examined the D-loop and nuclear microsatellite regions to identify spatially explicit and feasible management units for effective population control and further range expansion retardation. Through the identification of five mitochondrial DNA haplotypes and three nuclear genetic groups, we identified at least three independent introductions, range expansion, and subsequent genetic admixture in the Boso Peninsula. The management unit considered that two were appropriate because two populations have already genetic exchange. Furthermore, when taking management, we think that it is important to monitor DNA at the same time as capture measures for feasible management. This makes it possible to determine whether there is a invasion that has a significant impact on population growth from out of the unit, and enables adaptive management.

Distribution of Eimeria uekii and Eimeria raichoi in cage protection environments for the conservation of Japanese rock ptarmigans (Lagopus muta japonica) in the Japanese Alps.

Japanese rock ptarmigans, Lagopus muta japonica, are classified as an endangered species in Japan and are found only in the Japanese Alps. The number of birds has decreased in the last half century and cage protection projects have been undertaken as in situ conservation strategies (one of the projects for the recovery plan of Japanese rock ptarmigan) in the mountains. During the period with cage protections, some chicks died and two Eimeria spp., E. uekii and E. raichoi, were identified in the chicks. Here, we examined the soil within the cages and in the surrounding environment to assess potential sources of infection between July to August 2020. We found high numbers of oocysts in the cages, especially at the back sides where the ptarmigan family frequently congregated, but soils in other areas outside the cages were less contaminated or not contaminated at all. The time required for more than 50% of the oocysts to sporulate at 15, 20 and 25 °C for E. uekii was 20, 11, and 5 h, respectively, and 72, 48 and 18 h, respectively, for E. raichoi. Our results cast some doubt that coprophagia by chicks is the source of infection because chicks consumed fresh cecal feces (approximately within 1 h) as far as we know, and instead, the protected chicks might be directly or indirectly infected by oocysts in soils or the environment.