Poorly understood but highly diverse microbial communities exist within anoxic and oxygen-depleted marine sediments. These communities often harbour single-celled eukaryotes that form symbiotic associations with different prokaryotes. During low tides in South-western British Columbia, Canada, vast areas of marine sand become exposed, forming tidal pools. Oxygen-depleted sediments within these pools are distinctively black at only 2-3 cm depth; these layers contain a rich variety of microorganisms, many of which are undescribed. We discovered and characterized a novel (uncultivated) lineage of heterotrophic euglenozoan within these environments using light microscopy, scanning and transmission electron microscopy, serial sectioning and ultrastructural reconstruction, and molecular phylogenetic analyses of small subunit rDNA sequences.Bihospites bacati n. gen. et sp. is a biflagellated microbial eukaryote that lives within low-oxygen intertidal sands and dies within a few hours of exposure to atmospheric oxygen. The cells are enveloped by two different prokaryotic episymbionts: (1) rod-shaped bacteria and (2) longitudinal strings of spherical bacteria, capable of ejecting an internal, tightly wound thread. Ultrastructural data showed that B. bacati possesses all of the euglenozoan synapomorphies. Moreover, phylogenetic analyses of SSU rDNA sequences demonstrated that B. bacati groups strongly with the Symbiontida: a newly established subclade within the Euglenozoa that includes Calkinsia aureus and other unidentified organisms living in low-oxygen sediments. B. bacati also possessed novel features, such as a compact C-shaped rod apparatus encircling the nucleus, a cytostomal funnel and a distinctive cell surface organization reminiscent of the pellicle strips in phagotrophic euglenids.B. bacati n. gen. et sp. Molecular phylogenetic analyses demonstrated that this species belongs to the Euglenozoa and currently branches as the earliest diverging member of the Symbiontida. This is concordant with ultrastructural features of B. bacati that are intermediate between C. aureus and phagotrophic euglenids, indicating that the most recent ancestor of the Symbiontida descended from phagotrophic euglenids. Additionally, the extrusive episymbionts in B. bacati are strikingly similar to so-called "epixenosomes", prokaryotes previously described in a ciliate species and identified as members of the Verrucomicrobia. These parallel symbioses increase the comparative context for understanding the origin(s) of extrusive organelles in eukaryotes and underscores how little we know about the symbiotic communities of marine benthic environments.We characterized the ultrastructure and molecular phylogenetic position of  Euglena and Phacus), but the majority of the species are heterotrophic (osmotrophs or phagotrophs). Photosynthetic euglenids evolved from phagotrophic ancestors with a complex feeding apparatus and a large number of pellicle strips that facilitate a characteristic peristaltic cell movement called "euglenoid movement". This combination of characters allows phagotrophic euglenids to engulf large prey cells, such as eukaryotic algae, which eventually led to the acquisition of chloroplasts via secondary endosymbiosis .Both resin-embedded cells used for TEM and cells on gold sputter-coated SEM stubs have been deposited in the Beaty Biodiversity Research Centre (Marine Invertebrate Collection) at the University of British Columbia, Vancouver, Canada.Figs Tidal sand-flat at Centennial Beach, Vancouver, British Columbia, Canada .Marine sand, black layer 2-3 cm deep.bacati, ornamented with pearls. The etymology for the specific epithet reflects the presence of distinct longitudinal rows of spherical-shaped episymbionts, reminiscent of pearl necklaces.Specific epithet, Latin LSID for article: urn:lsid:zoobank.org:pub:40211D82-B95C-494A-B8D0-7E061E80DD18Bihospites: urn:lsid:zoobank.org:act:794D6C7B-BFB1-45C7-8DDA-32D44F3B0E50LSID for the genus B. bacati: urn:lsid:zoobank.org:act:E1549565-5434-4F85-B936-7D0C485596B8LSID for the species ar: accessory rod; CGS: congregated globule structure; Cyt: cytostome; Db: dorsal basal body; Db': dorsal pro-basal body; Df: dorsal flagellum; DL: dorsal lamina; DMt: dorsal microtubules; DR: dorsal root; E: extrusome; epi: epixenosome; ER: endoplasmic reticulum; FP: flagellar pocket; IR: intermediate root; LM: light microscope; MtD: mitochondrion-derived organelle; N: nucleus; Nu: nucleolus; PR: paraxial rod; r: rod; S: strips; SF: striated fibre; SEM: scanning electron microscope; TEM: transmission electron microscope; tz: transition zone; Vb: ventral basal body; Vb': ventral pro-basal-body; Vf: ventral flagellum; VL: ventral lamina; VR: ventral root; vt: vestibulum.SAB collected the sediment samples from Boundary Bay; generated the LM, SEM, and SSU rDNA sequence data; and wrote the first draft of the paper. NY generated the TEM data and helped with the phylogenetic analyses and interpretation of the TEM data. MH carried out the sampling, identification and LM work of the German material and helped with the identification of the Canadian material. BSL funded and supervised the collection and interpretation of the ultrastructural and molecular phylogenetic data and contributed to writing the paper. All authors have read, edited and approved the final manuscript.


Dr. Luis D. Giavedoni was not included in the author byline. He should be listed as the sixth author and affiliated with Department of Virology and Immunology and Southwest National Primate Research Center, Southwest Foundation for Biomedical Research, San Antonio, Texas, United States of America.The author contributions should also be amended to the following: Conceived and designed the experiments: TRK JBB RDH JP PD. Performed the Experiments: RDH JC JP AC CT AC TRK LDG. Analyzed the data: RDH JC TRK AC LDG CT AC JBB PD. Contributed reagents/materials/ analysis tools: AC TRK JBB. Wrote the paper: JC JBB PD.


Escherichia coli (E. coli). To gain an insight into the genes and pathways involved in hepatic APR of dairy cows we performed a global gene expression analysis of liver tissue sampled at different time points before and after intra-mammary (IM) exposure to E. coli lipopolysaccharide (LPS) treatment.Liver plays a profound role in the acute phase response (APR) observed in the early phase of acute bovine mastitis caused by Approximately 20% target transcripts were differentially expressed and eight co-expression clusters were identified. Each cluster had a unique time-dependent expression profile and consisted of genes involved in different biological processes. Our findings suggest that APR in the liver is triggered by the activation of signaling pathways that are involved with common and hepatic-specific transcription factors and pro-inflammatory cytokines. These mediators in turn stimulated or repressed the expression of genes encoding acute phase proteins (APP), collectins, complement components, chemokines, cell adhesion molecules and key metabolic enzymes during the APR. Hormones, anti-inflammatory and other hypothalamus-pituitary-adrenal axis (HPAA) linked mediators also seemed to participate in APR.E. coli mastitis, and that the bovine hepatic APR follows the same pattern as other mammals when they are challenged with LPS. Our work presents the first insight into the dynamic changes in gene expression in the liver that influences the induction, kinetics and clinical outcome of the APR in dairy cows.Performing global gene expression analysis on liver tissue from IM LPS treated cows verified that the liver plays a major role in the APR of  Escherichia coli (E. coli) is a common disease in lactating dairy cows. During infection, lipopolysaccharide (LPS) released from the cell wall of E. coli rapidly induces a sophisticated inflammatory response. Locally it is characterized by recruiting leukocytes especially neutrophils to the injured mammary tissue, as well as activation of macrophages in the mammary gland to produce pro-inflammatory cytokines  (the probe set is Bt.21216.1.S1_at), were significant.Differential expression of individual genes was assessed using linear modeling and empirical Bayes methods  as impleTo identify co-expression clusters, the 4610 DE transcripts were ordered by the hierarchical clustering. Centered Pearson correlation method was used as the distance measure. The optimum cluster number was determined using the ratio of the sums of squares between and within cluster  see Fig.There are 1789 UniGenes possessing Gene Ontology (GO) among all DE transcripts in contrast to 6843 UniGenes from the whole array. Within each co-expression cluster, overrepresentation of gene sets defined by the Biological Process (BP) of GO was tested using Fisher's exact test.E. coli: Escherichia coli; EGF: epidermal growth factor; GCRMA: GeneChip robust multi-array analysis; GH: growth factor; GO: gene ontology; HPAA: hypothalamus-pituitary-adrenal axis; IM: intra-mammary; i.p.: intraperitoneal; IU: international units; i.v.: intravenous; LIMMA: linear models for microarray analysis; LPS: lipopolysaccharide; MAS: microarray suite; MCP: monocyte chemotactic protein; MIP: macrophage inflammatory protein; NAP: neutrophil-activating protein; PG: prostaglandin; PRL: prolactin; PRLR: prolactin receptor; Q-PCR: quantitative polymerase chain reaction; RMA: robust multi-array analysis; SCC: somatic cell countAMPK: adenosine monophosphate-activated protein kinase; APP: acute phase proteins; APR: acute phase response; ATP: adenosine 5'-triphosphate; BP: biological process; CAM: cell-adhesion molecules; CC: complement components; cRNA: complementary ribonucleic acid; DE: differentially expressed; LJ and PS analyzed the microarray data and drafted the manuscript. CR carried out the animal challenge, KLI performed the liver biopsy and LV performed the Q-PCR. PS, CR and KLI designed the experimental plan. LJ, PS, CR and KLI contributed to the interpretation of results and iterative refinement of the manuscript. LJ is responsible for depositing microarray data in NCBI GEO.  LJ and PS contributed equally to this work.


The Author Contributions are incorrect. They should read as follows: Conceived and designed the experiments: GA. Performed the experiments: AAM GA. Analyzed the data: AAM GA. Wrote the paper: GA GB.


The Author Corrections are incomplete. The full Author Contributions are: Conceived of the study: CL BD. Performed the numerics: CL. Wrote the paper: CL BD.


However, in order to fully utilize the scale-independent properties of CGR, it should be accessible through scale-independent user interface instead of static images. Here we describe a web server and Perl library for generating zoomable CGR images utilizing Google Maps API, which is also easily searchable for specific motifs. The web server is freely accessible at  Moreover, other web-pages or web-database sites can utilize our service to add CGR and  website .CGR: Chaos Game Representation; FCGR: Frequency matrices extracted from CGR; G-language GAE: G-language Genome Analysis Environment; IFS: Iterated Function System; REST: Representational State Transfer; ZUI: Zoomable User Interface.The authors declare that they have no competing interests.KA designed and developed the software, and drafted the manuscript. KO participated in the implementation of the software. MT supervised the project. All authors have read and approved the final manuscript.


There were errors in the Author Contributions. The correct contributions are: Conceived and designed the experiments: GH DRS. Performed the experiments: GH SY NS KJD TS. Analyzed the data: GH DRS. Wrote the paper: GH DRS. Made the figures: GH KJD.


Correction to: British Journal of Cancer (2006) 94, 1351. doi:10.1038/sj.bjc.6603068Owing to an author error, the title of the above letter was incorrect. The title should be:‘Mobile phone use and risk of acoustic neuroma: results of the Interphone case–control study in five North European countries’.


Daphnia magna (Crustacea:Cladocera) to isolate variable number of tandem repeat (VNTR) markers for linkage mapping, Quantitative Trait Loci (QTL), and functional studies.Expressed sequence tag (EST) databases provide a valuable source of genetic data in organisms whose genome sequence information is not yet compiled. We used a published EST database for the waterflea Seventy-four polymorphic markers were isolated and characterised. Analyses of repeat structure, putative gene function and polymorphism indicated that intragenic tandem repeats are not distributed randomly in the mRNA sequences; instead, dinucleotides are more frequent in non-coding regions, whereas trinucleotides (and longer motifs involving multiple-of-three nucleotide repeats) are preferentially situated in coding regions. We also observed differential distribution of repeat motifs across putative genetic functions. This indicates differential selective constraints and possible functional significance of VNTR polymorphism in at least some genes.Databases of VNTR markers situated in genes whose putative function can be inferred from homology searches will be a valuable resource for the genetic study of functional variation and selection. Daphnia (Crustacea:Cladocera) are small planktonic crustaceans found in standing freshwater bodies around the world. They have a long history as model organisms for evolutionary, ecological and ecotoxicological research. Recently, the genus has been the focus of a major sequencing effort, and the full genome sequence of Daphnia pulex is now available : Concentration of MgCl2 used in the PCR buffer. An asterisk following MgCl2 concentration indicates hot start PCR.Click here for fileDescription of the 74 polymorphic VNTR loci. EST locus: locus containing the VNTR; Size: size of the repeated motif; Sequence: repeat consensus sequence. N: number of repeats, ; A: number of alleles. *: total number of alleles in loci with more than one repeated motif. He: Gene diversity.Click here for fileResults of homology searches and putative functions of EST loci. TM: transmembrane regions. Function categories: DEF: defense; MET: metabolism; OTH: other; SIG: signaling and gene expression regulation; SUR: surface and integumental proteins; UNK: unknown function. No hit: no significant homolog found .Click here for fileDistribution of allele sizes of polymorphic loci in each sampled location.Click here for file


The author contributions are incorrect. The correct contributions are: Performed the experiments: ZA RC LC ADB EAAF ON CT CU HK HPS


Hemophagocytic syndrome is a rare clinicopathological condition characterized by the activation of the mononuclear phagocyte system, resulting in hemophagocytosis in the reticuloendothelial systems. The pathogenesis of HPS remains unclear.We report the case of a 20-year-old soldier suffering from HPS. Because of long history fever and no reasons being found, his left groin lymph node and left neck lymph node biopsy were done with two weeks interval. We found a marked increase in plasma cells in left neck lymph node during the course of the disease.Our result provides a new thought for the researchers to understand the mechanisms responsible for the phagocytosis in HPS. Hemophagocytic syndrome (HPS) is a rare entity characterized by the dysfunction of cytotoxic T cells (CTL) and natural killer (NK) cells, and the activation of the mononuclear phagocyte system ,2. Its c9/L, red blood cell 3.42 × 1012/L, hemoglobin 107 g/L, platelet count 156 × 109/L, fibrinogen 0.67 g/L, triglyceride 13.9 mmol/L (Table A 20-year-old enlisted man presented with a 1-month history of discontinuous fever, panic, and hypodynamia, and was admitted to People's Liberation Army 152 hospital in May 2008. He had no other complaints. The patient had previously been healthy, with no history of infectious diseases or allergies to food or drugs. Physical examination showed a temperature of 37.3°C and no other remarkable signs. Laboratory measurements and examinations revealed no abnormalities. On admission, the patient was administered broad spectrum antibiotics and antiviral drugs. However, his fever failed to resolve and his temperature rose to between 38.5°C and 40.9°C. More than 2 weeks later, lymphadenopathy of the left neck, armpit, and left groin was detected by ultrasound. Left groin lymph node and bone marrow puncture biopsies were performed. Histopathological examination of the groin lymph node showed reactive hyperplasia and revealed one histiocyte engulfing few blood cells Figure. . The bonLeft neck lymph node biopsy and bone marrow puncture smear were performed. The results were characteristic of HPS Figure. . The gerHPS is a life-threatening condition characterized by the activation of the MPS. It can be divided into genetic HPS and secondary HPS . GeneticAlthough progress has been made in understanding the pathophysiology of HPS, the exact mechanisms responsible for the phagocytosis are unknown in HPS. Researchers found out that there have the RBC antibodies in the EBV-HPS and anti-RBC antibodies were involved in the phagocytosis of RBCs by macrophages . Also seThis case report showed plasma cells increased markedly in the left neck node during the progression of HPS. The result maybe provide one way to study the pathophysiology of the HPS.HPS: hemophagocytic syndrome; CTL: cytotoxic T cells; MPS: mononuclear phagocyte system; HAV: hepatitis A virus; HBV: hepatitis B virus; HCV: hepatitis C virus; HIV: human immunodeficiency virus; CP: chlamydia pneumoniae; MP: mycoplasma; RSV: respiratory syncytial virus; Adv: adenovirus; WBC: white blood cell count; RBC: red blood cell; HGB: hemoglobin; PLT: platelet count; PT: prothrombin time; NR: international normalized ratio; TT: thrombin time; APTT: activated partial thromboplastin time; FIG: fibrinogen; TP: total protein; ALB: albumin; GLOB: globulin; DB: direct bilirubin; IB: indirect bilirubin; γ-GT: γ-glutamyl transpeptidase; CK: creatine kinase; CK-MB: creatine kinase-MB; HBDH: hydroxybutyrate dehydrogenase; LDH: lactate dehydrogenase; TG: triglyceride; β2-MG: β2-microglolulin; CRP: C-reactive protein; ALP: alkaline phosphatase.Written informed consent was obtained from the patient' older brother for publication of this case report and accompanying images. A copy of the written consent is available for review by the Editor-in-Chief of this Journal.The authors declare that they have no competing interests.L CY and M YY interpreted the data, operated on the patient and wrote the manuscript. L CG performed the histological examination. H XX reviewed the paper. All authors read and approved the final manuscript.


Axiothella rubrocincta. Employing immunocytochemistry in combination with confocal laserscanning microscopy, we broaden the database on capitellid organogenesis, thereby incorporating classical histological data in our analysis. Besides assessing possible shared features with the echiurans, we also discuss the variability of neural and muscular characters within the Capitellida.Numerous phylogenetic analyses on polychaete annelids suggest a taxon Capitellida that comprises the three families Maldanidae, Arenicolidae and Capitellidae. Recent molecular studies support the position of the Echiura, traditionally ranked as a separate phylum, within the capitellids. In order to test the robustness of this molecular-based hypothesis we take a different approach using comparative analyses of nervous and muscle system development in the maldanid Axiothella, consists of cerebral commissures that give rise to simple circumesophageal connectives with fused ventral and dorsal roots and a single ventral neurite bundle. From the latter arise segmental neurites that innervate the peripheral bodywall. Since there is no observable regular pattern, and individual neurites are lost during ontogeny, their exact arrangement remains elusive. The pharynx is encircled by a prominent stomatogastric nerve ring, with a pair of anterior and lateral proboscis neurites directly connecting it to the central nervous system. One pair of ventral and one pair of dorsal longitudinal muscles form the earliest rudiments of the bodywall musculature in late larval stages, while a continuous layer of circular muscles is lacking throughout ontogeny.The scaffold of the adult central nervous system, which is already established in early developmental stages of Comparative neurodevelopmental analysis of capitellid and echiuran species reveals several common characters, including simple circumesophageal connectives, a single fused ventral nerve strand, and a stomatogastric ring nerve, that support a close relationship of both taxa, thus corroborating recent molecular phylogenetic analyses. The data on myogenesis show that four longitudinal muscle bands most likely represent an ancestral character not only for the Capitellida, but for the Annelida in general. Whether or not circular muscles are part of the annelid groundpattern remains uncertain. Axiothella rubrocincta not only offers an opportunity to assess the ingroup variability of neural characters within the Capitellida but also allows to compare neurogenesis and nervous system organization with those data that recently have become available for echiurans. This serves as an independent test of the molecular data which propose the placement of the echiurans within the Capitellida.The Maldanidae, also referred to as 'bamboo worms', comprise a group of deposit-feeding polychaete annelids that live in tubes composed of bottom material. They are usually considered related to the Arenicolidae and Capitellidae, and these three families are grouped together in the taxon Capitellida . Recent Axiothella may also aid in casting light on the ancestral state of muscular systems in annelids and lophotrochozoans as a whole. The musculature of the Capitellida comprises a closed outer layer of circular fibers similar to that of clitellate oligochaetes . The primary antibodies, polyclonal rabbit anti-serotonin , polyclonal rabbit anti-FMRFamide , and monoclonal mouse anti-acetylated α-tubulin , all in block-PTA, were either applied separately or in a mixed cocktail for 24 h. Subsequently, the specimens were rinsed in block-PTA with three changes over 6 h and incubated in a mixture of 4'6-diamidino-2-phenyl-indole [DAPI ], secondary fluorochrome-conjugated antibodies  and, for F-actin visualization, Alexa Fluor 488 phalloidin  in block-PTA overnight. Finally, the specimens were washed three times in PB without NaN3 and were directly mounted in Fluoromount G  on glass slides. A minimum of 10 immunolabeled specimens per developmental stage was analyzed for each antibody. Approximately 65 image stacks of optical sections were recorded as Z-wide-projections with 0.1-0.5 μm step size using a Leica DM IRE2 fluorescence microscope equipped with a Leica TCS SP 2 confocal laserscanning unit . Setae are visible in the tubulin scans due to autofluorescence. Images were processed with Adobe Photoshop CS3 to adjust contrast and brightness and were arranged into figure plates using Adobe Illustrator CS3 . The three-dimensional computer reconstructions were generated with the imaging software Imaris v. 5.5.3  using surface rendering algorhithms.The following steps were all performed at 4°C. Antibody staining was preceded by tissue permeabilization for 1 h in 0.1 M PB with 0.1% NaN1s-7s: setigers; a: anus; ac: apical cilia; apn: anterior proboscis neurite; as: anal sphincter; a-str: accessory stomatogastric nerve ring; bun: nerves of the buccal epithelium; cc: circumesophageal connective; cg: cerebral ganglion; cm-rs: circular muscle of the retractor sheath; dl-cg: dorsal lobe of the cerebral ganglion; dlm: dorsal longitudinal muscle bundle; d-sp: dorsal serotonergic perikarya; d-vlm: diagonal ventral longitudinal muscle; e: esophagus; ge: glandular epidermis; h: head; i: intestine; lm: longitudinal muscle fiber; lm-rs: longitudinal muscle of the retractor sheath; lpn: lateral proboscis neurite; ma: main strand; me: median strand; nn: nuchal neurite; nt: neurotroch; pc: posterior cilia; per: peristomium; ph: pharynx; pn: protonephridium; pnn: peripheral network of neurites; prn: prostomial nerves; pro: prostomium; pt: prototroch; py: pygidium; s: setae; sm: setal muscles; stp: neuronal stomatogastric projection; str: stomatogastric nerve ring; s-vlm: straight ventral longitudinal muscle; tt: telotroch; vl-cg: ventral lobe of the cerebral ganglion; vlm: ventral longitudinal muscle bundle; vnb: ventral neurite bundle; v-sp: ventral serotonergic perikarya.NB performed research, analyzed data and drafted the manuscript. AW designed and coordinated research and contributed to writing of the manuscript. Both authors conceived the study, read, and approved the final version of the manuscript.


Due to the addition of author Dao-Fu Dai, the author contributions should now read: Conceived and designed the experiments: WL. Performed the experiments: LCE JFM PRT. Analyzed the data: LCE PRT MJE DFD WL. Contributed reagents/materials/analysis tools: NSW GSM PSR WL. Wrote the paper: LCE.


Because the author Aruna Panda was added to the author byline after publication, the Author Contributions section does not correctly reflect the new authorship. The new Author Contributions should read:Conceived and designed the experiments: GN WK ZY DP SR CW DS. Performed the experiments: WK ZY DP LD SR CW ES HS HW GR AP. Analyzed the data: GN WK ZY DP MN SR CW DS. Wrote the paper: GN WK ZY DP SR CW DS AP.


Objective: To investigate clinical experience with the apparent malignant transformation of benign liver cell adenomas.Design: Retrospective review of personal experience and literature.Setting: University hospital and affiliated community hospitals.Patients: All patients diagnosed with liver cell adenomas over a 30-year period.Interventions: Liver resection and/or tumor biopsy.Main Outcome Measures: Gender, age, drug associations, alpha-fetoprotein levels, response to treatment, and survival.Results: Thirteen patients from personal experience and 26 patients from the reports of others had liver cell adenomas that were not resected. Five of these patients subsequently developed hepatocellular carcinoma.Conclusions: Malignant transformation of a liver cell adenoma is a rare phenomenon, but it does occur. Alphafetoprotein levels may be more helpful in diagnosis than expected from previous reports. Solitary benign adenomas should be resected whenever possible. Patients with diffuse multiple tumors should be observed closely over a long period.


There were errors in the Author Contributions. The correct contributions are: Conceived and designed the experiments: TK KH ZPF PR. Performed the experiments: TK KH WK NR GS. Analyzed the data: TK KH ZPF PR. Wrote the paper: TK KH ZPF PR.


With the advent of functional magnetic resonance imaging (fMRI) in awake animals it is possible to resolve patterns of neuronal activity across the entire brain with high spatial and temporal resolution. Synchronized changes in neuronal activity across multiple brain areas can be viewed as functional neuroanatomical circuits coordinating the thoughts, memories and emotions for particular behaviors. To this end, fMRI in conscious rats combined with 3D computational analysis was used to identifying the putative distributed neural circuit involved in aggressive motivation and how this circuit is affected by drugs that block aggressive behavior.1a receptor antagonist SRX251 or the selective serotonin reuptake inhibitor fluoxetine, drugs that block aggressive behavior, both caused a general suppression of the distributed neural circuit involved in aggressive motivation. However, the effect of SRX251, but not fluoxetine, was specific to aggression as brain activation in response to a novel sexually receptive female was unaffected.To trigger aggressive motivation, male rats were presented with their female cage mate plus a novel male intruder in the bore of the magnet during image acquisition. As expected, brain areas previously identified as critical in the organization and expression of aggressive behavior were activated, e.g., lateral hypothalamus, medial basal amygdala. Unexpected was the intense activation of the forebrain cortex and anterior thalamic nuclei. Oral administration of a selective vasopressin VThe putative neural circuit of aggressive motivation identified with fMRI includes neural substrates contributing to emotional expression , emotional experience  and the anterior thalamic nuclei that bridge the motor and cognitive components of aggressive responding. Drugs that block vasopressin neurotransmission or enhance serotonin activity suppress activity in this putative neural circuit of aggressive motivation, particularly the anterior thalamic nuclei. Aggression is a normal component of mammalian behavior . For ani1a and 5HT1b receptor agonists  arginine vasopressin, blocks aggressive behavior -1for each subject (i) was also calculated.Anatomy images for each subject were obtained at a resolution of 2562 with a depth of 1200 μm, many small brain areas  cannot be resolved; or, if they could be resolved they would be represented by one or two voxels only . Consequently, small detailed regions are not included in the analysis or are grouped into larger "minor volumes" of similar anatomical classification. For example, in these studies we have the basal nucleus of the amygdala listed as a minor volume. This area is a composition of the basomedial anterior part, basomedial posterior part, basolateral anterior part and basolateral posterior part with a composite voxel size of 54. In this study, 12 brain slices were collected extending from the tip of the forebrain to the end of the cerebrum stopping at the midbrain just rostral to the cerebellum. Within these rostral/caudal boundaries we delineated 83 minor volumes. In addition, we grouped brain areas into "major volumes" . The volume of activation (number of significant voxels) can be visualized in these 3D major and minor anatomical groupings  is 486 μmt tests were performed on each voxel  of each subject within their original coordinate system. The baseline threshold was set at 2%. The t test statistics used a 95% confidence level, two-tailed distributions, and heteroscedastic variance assumptions. As a result of the multiple t test analyses performed, a false-positive detection controlling mechanism was introduced -1, mapped it within a voxel of subject (i). A tri-linear interpolation of the subject's voxel values (percentage change) determined the statistical contribution of subject (i) to the composite  location. The use of [Ti]-1 ensured that the full volume set of the composite was populated with subject contributions. The average value from all subjects within the group determined the composite value. The BOLD response maps of the composite were somewhat broader in their spatial coverage than in an individual subject; so only average number of activated pixels that has highest composite percent change values in particular ROI was displayed in composite map. Activated composite pixels are calculated as follows:A statistical composite was created for each group of subjects. The individual analyses were summed within groups. The composite statistics were built using the inverse transformation matrices. Each composite pixel location , premultiplied by [The composite percent change for the time history graphs for each region was based on the weighted average of each subject, as follows:N is number of subjects.where The percent change in BOLD signal for time history graphs see Figs , 7, 8 weACA: anterior cingulated; ACB: accumbens; AH: anterior hypothalamus; AL: agranular insular cortex; ALP: anterior lobe pituitary; AM: anteromedial nucleus thalamus; AON: anterior olfactory nucleus; AUD: auditory cortex; AV: anteroventral nucleus thalamus; BNST: bed nucleus stria terminalis; CA1: hippocampus; CA3: hippocampus; COA: cortical nucleus amygdale; DG: dentate gyrus; DS: dorsal striatum; ENT: entorhinal cortex; IC: inferior colliculi; ILA: infralimbic cortex; IPN: interpeduncular nucleus; LHA: lateral hypothalamus; LP: lateral posterior nucleus thalamus; LS: lateral septum; MEA: medial amygdale; MM: mammillary nuclei; MO: motor cortex; MRN: mesencephalic reticular nucleus; ORB: orbital cortex; OT: olfactory tubercle; PAG: periaqueductal gray; PIR: piriform cortex; PL: prelimbic cortex; PTL: parietal cortex; PVN: paraventricular nucleus hypothalamus; PVT: paraventricular nucleus thalamus; RE nucleus reunions thalamus; RSP: retrosplenial cortex; RPH: raphe; SC: superior colliculi; SN: substantia nigra; SSP: primary somatosensory cortex; VMH: ventromedial hypothalamus; VP: ventral posterior nucleus thalamus; VTA: ventral tegmental areaCraig Ferris and Mathew Brevard have a financial interest in Insight MRI the bioengineering company that manufactures the animal imaging equipment used in these studies. Craig Ferris and Neal Simon have a financial interest in Azevan Pharmaceuticals the company developing SRX251 an orally active V1a receptor antagonist used in these studies.Each author has made a significant contribution to these studies either in the acquisition of data , software development, data analysis , or study design and manuscript preparation . All authors read and approved the final manuscript.


Escherichia coli, allowing for the estimation of energy generation and cell synthesis, based on intracellular fluxes and that may be used to characterize phenotypic growth.It is quite important to simulate the metabolic changes of a cell in response to the change in culture environment and/or specific gene knockouts particularly for the purpose of application in industry. If this could be done, the cell design can be made without conducting exhaustive experiments, and one can screen out the promising candidates, proceeded by experimental verification of a select few of particular interest. Although several models have so far been proposed, most of them focus on the specific metabolic pathways. It is preferred to model the whole of the main metabolic pathways in E. coli. Once intracellular fluxes were computed by this model, the specific ATP production rate, the specific CO2 production rate, and the specific NADPH production rate could be estimated. The specific ATP production rate thus computed was used for the estimation of the specific growth rate. The CO2 production rate could be used to estimate cell yield, and the specific NADPH production rate could be used to determine the flux of the oxidative PP pathway. The batch and continuous cultivations were simulated where the changing patterns of extracellular and intra-cellular metabolite concentrations were compared with experimental data. Moreover, the effects of the knockout of such pathways as Ppc, Pck and Pyk on the metabolism were simulated. It was shown to be difficult for the cell to grow in Ppc mutant due to low concentration of OAA, while Pck mutant does not necessarily show this phenomenon. The slower growth rate of the Ppc mutant was properly estimated by taking into account the lower specific ATP production rate. In the case of Pyk mutant, the enzyme level regulation was made clear such that Pyk knockout caused PEP concentration to be up-regulated and activated Ppc, which caused the increase in MAL concentration and backed up reduced PYR through Mez, resulting in the phenotypic growth characteristics similar to the wild type.In the present study, we considered the simulation of the main metabolic pathways such as glycolysis, TCA cycle, pentose phosphate (PP) pathway, and the anapleorotic pathways using enzymatic reaction models of E. coli for understanding metabolic changes inside the cell in response to specific pathway gene knockouts, considering the whole main metabolic pathways. The comparison of the simulation result with the experimental data indicates that the present model could simulate the effect of the specific gene knockouts to the changes in the metabolisms to some extent.It was shown to be useful to simulate the main metabolism of  One of the most challenging goals of metabolic engineering is to design the cell metabolism based on the analysis of metabolic regulation. For this, it is strongly desired to develop a mathematical model which can describe the dynamic behaviour of the cell in response to the changes in the culture environment and/or specific genetic modifications. Although an attempt has been made to develop a platform for the whole cell model , the totSaccharomyces cerevisiae  glucose, 0.4 g of [1-13C] glucose, and 3.2 g of naturally labeled glucose per liter. Biomass samples for GC-MS analysis were taken after one residence time. Sample preparation, analytical procedures for GC-MS analysis, and flux computation are given elsewhere  denotes the concentration, and the abbreviations for the metabolites are explained in Nomenclature. ll grows . As showThe estimation for the cell growth rate is by far important. In particular, it is critical for the prediction of the specific gene knockout mutant. The most typical equation for the cell growth rate is the Monod equation:S is the substrate concentration and S ≡ [ex Glc] in the present case. The drawback of this equation is that the cell will keep growing at μm as far as S ≫ Ks where Ks is usually quite small in the case of the utilization of glucose as the substrate. This does not reflect the fermentation characteristics in practice. Therefore, we introduced another term such aswhere X is the cell concentration and Xm is the final value of X in the batch culture, and n is the model parameter. Xm may be set as S(0)YS/X where S (0) is the initial substrate concentration, and YS/X is the yield coefficient. Although this equation can be used to simulate the dynamic behavior in batch culture, Eq. (2b) may not be able express the cell growth rate for the mutants. We, therefore, introduce another idea by taking into account the effect of ATP generation on the cell growth rate based on experimental observation.where 2 can contribute in generating ATP via oxidative phosphorylation. The pathways involved in electron transfer and oxidative phosphorylation have variable stoichiometry due to the use of different dehydrogenases and cytochromes. Namely, the NADH dehydrogenase NDH-I encoded by nuo gene transports 2 H+/e-, while NDH-II encoded by ndh gene transports 0 H+/e- /[NAD] ratio increases as given by Eq.10) of Table 0 of Tablm = 8 mM) and Icl  compete for ICIT. This branch point is under both gene level regulation by aceBAK operon and the enzyme level regulation by the phosphorylation/dephosphorylation of ICDH. Here, we considered the rate equation for ICDH as given by Eq.(15) of Table For citrate synthase reaction, Eq.14) in Table 4 in TablAs for the glyoxylate pathway, Eq. 16) of Table 6 of TablFor G6PDH and 6PGDH reactions in the PP pathway, Eqs.(23) and (24) of Table 2 production rate Once the fluxes were determined, the specific CO2 production rate was computed by the product of net production of CO2 and biomass concentration, i.e. and the total COvNADPH may also be estimated byMoreover, the specific NADPH production rate μ and vNADPH computed based on the flux estimates obtained from 13C-labeling experiments [It should be noted that the flux determination in the PP pathway is not easy for the enzyme-based kinetic equations. In particular, it is not easy to accurately predict the flux of G6PDH at the branch point of G6P. Figure eriments ,15, indikNADPH value from Figure kNADPH after μ was computed. Then, the flux of G6PDH may be estimated.Once we identified As stated in introduction, it is not easy, even if it is not impossible, to express gene-level regulation by mathematical equations. Here, we alternatively introduced several rules for the simulation based on the experimental observations and the roles of global regulators such as given in Additional file Metabolites2KG: 2-Keto-D-gluconate; 6PG: 6-Phosphogluconolactone; ACE: Acetate; AcP: Acetyl phosphate; AcCoA: Acetyl-CoA; ASP: Aspartate; ADP: Adenosine diphosphate; ATP: Adenosine-5'-triphosphate; AMP: Adenosine monophosphate; DHAP: Dihydroxyacetone phosphate; E4P: Erythrose 4-phosphate; F6P: Fructose 6-phosphate; FDP: Fructose 1,6-bisphosphate; FUM: Fumarate; G6P: Glucose-6-phosphate; GAP: Glyceraldehyde 3-phosphate; ICIT: Isocitrate; NAD/NADH: Nicotinamide adenine dinucleotide; NADP/NADPH: Nicotinamide adenine dinucleotide phosphate; MAL: Malate; OAA: Oxaloacetate; P: Phosphate; PEP: Phosphoenolpyruvate; PYR: Pyruvate; R5P: Ribulose 5-phosphate; Ru5P: Ribose 5-phosphate; S7P: Sedoheptulose 7-phosphate; SUC: Succinate; X5P: Xylulose 5-phosphateProtien (enzyme)Glc,/EIICBGlc: Enzyme for glucose; Eno: Enolase; Fum: Fumarase; G6PDH: Glucose-6-phosphate dehydrogenase; GAPDH: Glyceraldehyde 3-phosphate dehydrogenase; ICDH: Isocitrate dehydrogenase; HPr: Histidine containing protien; Icl: Isocitrate lyase; MDH: Malate dehydrogenase; Mez: Malic enzyme; Ms: Malate synthase; Pck: Phosphoenolpyruvate carboxykinase; PDH: Pyruvate dehydrogenase; Pfk/Pfk-1: Phosphofructokinase-1; Pgi: Phosphoglucose isomerase/Glucosephosphate isomerase; Pgk: Phosphoglycerate kinase; Pgm: Phosphoglycerate mutase; Ppc: PEP carboxylase; Pps: Phosphoenolpyruvate synthase; Pta: Phosphotransacetylase; PTS: Phosphotransferase system; Pyk: Pyruvate kinase; Rpe: Ribulose phosphate 3-epimerase; Rpi: Ribulose 5-phosphate 3-isomerase; SDH: Succinate dehydrogenase; Tal: Transaldolase; TktA: TransketolaseI; TktB: TransketolaseII2KGDH: 2-Keto-D-gluconate Dehydrogenase; Ack: Acetate kinase; Acs: Acetyl coenzyme A synthetase; AcP: Acetyl phosphate; Aldo: Aldolase; CS: Citrate synthase; EI: Cytoplasmatic protein (enzyme1); EII, EIIB, EIIC,: Carbohydrate specific (enzyme II); EIIAGeneaceBAK: operon which encodes the metabolic and regulatory enzymes of the glyoxylate bypass; cra: Catabolite repressor/activator; crr: Catabolite repressor; fadR: Fatty acid metabolism regulator; icdA: Isocitrate dehydrogenase A; iclR: Transcriptional repressor IclR; pckA: Phosphoenolpyruvate carboxykinase gene; pfkA, pfkB: Phosphofructokinase gene; ppsA: Phosphoenolpyruvate synthase gene; ptsG, ptsHI: Pts gene; pykA, pykF: Pyruvate kinase geneThe authors declare that they have no competing interests.TAAK made modeling, experiment and computer simulation, AAM made some TCA cycle modeling and simulation, AMK supervised computer simulation, JM made formulation, KS made modeling and manuscripts preparation. All authors read and approved the final manuscripts.Effects of initial metabolite concentrations on the fermentation characteristics.Click here for fileSimulation result of wild type and Ppc mutant in batch culture.Click here for fileTable S1 - Ruled used for the simulation.Click here for fileSimulation result of wild type and Ppc mutant in continuous culture.Click here for fileSimulation result of wild type and Pck mutant in continuous culture.Click here for fileSimulation result of wild type and Pyk mutant in batch culture.Click here for fileSimulation result of wild type and Pyk mutant in continuous culture.Click here for fileComputational procedure for the simulation of fitting experimental data.Click here for file


There were errors in the Author Contributions. The correct contributions are: Conceived and designed the experiments: ACC KHAC LHW. Performed the experiments: ACC ELN LHW. Analyzed the data: ACC ELN KHBM KHAC LHW. Contributed reagents/materials/analysis tools: KHAC LHW. Wrote the paper: ACC KHAC LHW. Technical advice and discussion: ACC ELN KHBM KHAC LHW.


The EQ-5D has been extensively used to assess patient utility in trials of new treatments within the cardiovascular field. The aims of this study were to review evidence of the validity and reliability of the EQ-5D, and to summarise utility scores based on the use of the EQ-5D in clinical trials and in studies of patients with cardiovascular disease.A structured literature search was conducted using keywords related to cardiovascular disease and EQ-5D. Original research studies of patients with cardiovascular disease that reported EQ-5D results and its measurement properties were included.Of 147 identified papers, 66 met the selection criteria, with 10 studies reporting evidence on validity or reliability and 60 reporting EQ-5D responses (VAS or self-classification). Mean EQ-5D index-based scores ranged from 0.24 (SD 0.39) to 0.90 (SD 0.16), while VAS scores ranged from 37 (SD 21) to 89 (no SD reported). Stratification of EQ-5D index scores by disease severity revealed that scores decreased from a mean of 0.78 (SD 0.18) to 0.51 (SD 0.21) for mild to severe disease in heart failure patients and from 0.80 (SD 0.05) to 0.45 (SD 0.22) for mild to severe disease in angina patients.The published evidence generally supports the validity and reliability of the EQ-5D as an outcome measure within the cardiovascular area. This review provides utility estimates across a range of cardiovascular subgroups and treatments that may be useful for future modelling of utilities and QALYs in economic evaluations within the cardiovascular area. Cardiovascular disease (CVD) imposes a great burden on societies around the world, with an estimated 16.7 million - or 29.2% of total global deaths - resulting from various forms of CVD.Increasingly over time, clinical trials within the cardiovascular field have included HRQoL measures. Such measures, alongside clinical measures of functionality, can help evaluate the physical, mental and emotional implications of CVD as well as the effects of surgical and medical treatments. Commonly used functional classification systems within the cardiovascular field are the New York Heart Association (NYHA) functional classification system for heart failure patients and the Canadian Cardiovascular Society (CCS) grading scale for angina pectoris,5. HRQoLAmong the available generic measures, the EQ-5D has gained widespread use due to its simplicity to administer, score and interpret. It also imposes minimal burden on the respondent as it is a brief, simple measure for patients to understand and to complete. The index-based score is generated by applying societal preference weights to the health state classification completed by the patient that consists of five dimensions , each with three levels of response or severity . The ability to convert self classification responses into a single index score makes the EQ-5D practical for clinical and economic evaluation. The indThe principle aims of this paper were: to synthesise the evidence on the validity and reliability of the EQ-5D in studies within the cardiovascular field; to summarise the EQ-5D based scores reported in studies within the CVD field; and to attempt to stratify mean utility scores according to level of disease severity.http://www.euroqol.org was also used to identify unique references, including working papers and conference proceedings that may not have been captured in the initial literature search. Only full-text published papers were included for analysis.A computerised search of the current published literature was performed using MEDLINE and EMBASE for the period January 1988 to October 2008. The search strategy combined exploded or medical subject headings relating to the CVD field and the EQ-5D as follows: ('cardiovascular'/exp OR 'cardiovascular') OR ('cardiac'/exp OR 'cardiac') OR ('cardiology'/exp OR 'cardiology') AND 'euroqol' OR 'EQ 5D' OR 'EQ5D'. The EuroQol website The inclusion criteria required that the paper was original research, and that it reported EQ-5D scores specific to cardiovascular disease or reported psychometric properties of the EQ-5D in a population with cardiovascular disease. Studies that only reported EQ-5D index or VAS scores graphically in terms of change over time were excluded from the analysis. When multiple studies used the same dataset, EQ-5D scores were only reported from one article to avoid double counting. No language restrictions were imposed. Study abstracts that potentially met the inclusion criteria were identified, and full-text articles were retrieved for further review. A standard data abstraction form was developed to facilitate the structured review, which included study design, patient characteristics, intervention information, published source of index-based preference weights and EQ-5D scores as well as details of any other clinical measures; disease-specific quality of life and generic HRQoL instruments. A summary of the results of the literature search is provided in figure I2 statistic [I2 statistic uses the sum of the squared differences of each study from the pooled estimate and the degrees of freedom of the test to provide a measure of the percent of total variation across studies due to heterogeneity between studies. A meta-analysis yielding a value of I2 above 75% suggests a high level of heterogeneity between the studies. Psychometric properties were summarised according to the type of property assessed , the comparison performed, and the statistical test result.Initially, studies that reported EQ-5D index-based scores and/or VAS scores were sorted into cardiovascular subgroups  that were informed by the latest WHO International Classification of Diseases (ICD-10: I00-I99 - diseases of the circulatory system)  explored construct validity (convergent and discriminative), typically in terms of correlations with other disease-specific HRQoL measures as well as reliability and responsiveness . Both the CCS and NYHA scales range from class I (mild symptoms) to class IV (severe symptoms) and CCS can also be graded as 0 for no symptoms  rely on generic measures such as the EQ-5D for the calculation of QALYs. Increased use of the EQ-5D may also support the view that patient reported outcomes and quality of life are becoming more widely accepted as routine measures in clinical studies, with the EQ-5D being an internationally recognised generic measure of HRQoL. This summary of EQ-5D index and VAS scores in the cardiovascular field complements other published reports describing the use of the EQ-5D in the cancer and asthma/COPD literature and of utility scores associated with various conditions -50.The review found that the majority of studies that included the EQ-5D were within IHD I20 - I25) and cerebrovascular disease (I60 - I69), subgroups, reflecting the relative prevalence of these diseases worldwide. Stratification by disease severity  was possible for IHD patients and heart failure patients and illustrated a positive relationship with the EQ-5D when moving from severe to mild disease severity  as well as non-disease-related factors such as other co-morbidities and demographic characteristics. Furthermore, no Overall, this study illustrated the difficulty in attempting to adequately deal with statistical heterogeneity based on aggregated data from published studies . This woHRQoL measures such as the EQ-5D can be useful tools to clinicians in terms of evaluating the impact of cardiovascular disease on patients and can help to inform decision making and resource allocation. The use of the EQ-5D in CVD studies has increased in recent years and published studies provide evidence of its validity and reliability. The variation in EQ-5D index and VAS scores reported here largely reflect systematic differences in terms of disease stage, treatment and patient characteristics. In the future, as more studies of CVD present EQ-5D scores according to disease severity, it may be possible to calculate pooled mean estimates that can be useful in modelling of CVD-related health outcomes in economic evaluations.AAA: Abdominal aortic aneurysm; ACS: Acute coronary syndromes; ACT: Anticoagulation therapy; AF: Atrial fibrillation; AH-Drug: Anti-hypertensive drug therapy; AHF: Advanced heart failure; AMI: Acute myocardial infarction; Amp.: Amputation; Angio: Coronary Angiography; ASA: American Society of Anaesthesiologists; ASCOT-AHD: Anglo-Scandinavian cardiac outcomes trial - anti-hypertensive drug treatment; Asym./Sym.: Asymptomatic/Symptomatic; AUT: Austria; AVR: Aortic valve replacement; BI: Barthel Index; BOTH-CABG/PCI/MM: Patients who are suitable for both CABG and PCI and receive CABG/PCI/MM; CABG: Coronary artery bypass graft; CABG-80: Coronary artery bypass surgery in octogenarians; CABG-CABG/PCI/MM: Patients who are suitable for CABG and receive CABG/PCI/MM; CABG-CPB: CABG using heart lung machine; CAD: Coronary artery disease; C-Arrest: Cardiac arrest; CCR: Comprehensive cardiac rehabilitation; CCS: Canadian Cardiovascular Society; CCU: Coronary care unit; CES-D: Centre for Epidemiological Studies - Depression Scale; CHD: Coronary heart disease; CHD-PHARM/Control: Community pharmacy-led medicines management programme/control treatment for patients with CHD; CML: Case method learning supported lipid-lowering strategy; COMM - CVD/NOCVD: Community dwelling-based elderly patients with/without CVD; CR: Cardiac resynchronisation; CR-Home/Hosp: Home/Hospital-based cardiac rehabilitation; C-REHAB: Cardiac rehabilitation; CS: Conservative strategy; CVA: Cerebrovascular Accident; CVD: Cardiovascular disease; Duplex US: Duplex ultrasonography; Echo: Echocardiography; EHS-CR: Euro Heart Survey on coronary revascularisation; Endo: Endovascular AAA surgery; ES: Effect size; Exercise-Qol: Long-term effects of exercise training on quality of life; F-u: Follow-up; GRS: Guyatt's responsiveness statistic; HeartMed: Lifestyle advice intervention by community pharmacists for heart failure patients; HF: Heart failure; HOSP - CVD: Hospital-based elderly patients with CVD; HRQoL: Health-related quality of life; HUI2/3: Health Utilities Index mark 2/3; IC: Intermittent claudication; ICC: Intra-class correlation; ICD: Implantable cardioverter defibrillator; ICP: Integrated care pathway; IHD: Ischaemic heart disease; IQR: Inter-quartile range; IS: Interventional strategy; IV: Intravenous; KCCQ: Kansas City Cardiomyopathy Questionnaire; LV: Left ventricular; MacNew: MacNew Heart Disease Quality of Life Questionnaire; MCS: Mental component summary; MDT: Multi-disciplinary team; MEDMAN: Community pharmacy-led medicines management services; MEPS: Medical expenditure panel survey; MI: Myocardial infarction; MI - Self-help: Home-based self-help rehabilitation package for MI patients; MIDCAB: Minimally invasive direct CABG; MM: Medical management; MR Angio: Magnetic resonance angiography; MRI: Magnetic resonance imaging; MT: Medical therapy; MVPS: Mitral valve prolapse syndrome; NYHA: New York Heart Association; OP-CABG: Off-pump CABG; Open: Open AAA surgery; PAOD: Peripheral arterial occlusive disease; PCI: Percutaneous coronary intervention; PCI-BMS: PCI with bare-metal stents; PCI-CABG/MM/PCI: Patients who are suitable for PCI and receive CABG/PCI/MM; PCI-DES: PCI with drug-eluting stents; PCS: Physical component summary; PER: Peripheral endovascular revascularisation; Pre-op: Pre-operation; Proxy: HRQol questionnaire completed by spouse/family member; PSM: Patient self-management; P-PTCA: Primary PTCA; P-Stent: Primary stent placement; PTCA: Percutaneous transluminal coronary angioplasty; QLMI: Quality of Life after MI questionnaire; QoL: Quality of life; RCT: Randomised controlled trial; REV/NO REV: Eligible/Ineligible for Revascularisation; SAH: Subarachnoid haemorrhage; SCOPE-Drug/Control: Study on cognition and prognosis in the elderly - Drug/Control treatment; SD: Standard deviation; SES: Socioeconomic status; SF-36: Short-form 36-item health survey questionnaire; SF-6D: Short-form 6D; SF-12: Short-form 12-item health survey questionnaire; SPECT: Single photon emission computed tomography; SRM: Standardised response means; Stroke-4Y: Four years post-stroke; TIA: Trans-ischaemic attack; Trans.: Heart Transplantation; Tx: Treatment; UC: Usual care; VAD: Ventricular assist device; VAS: Visual analogue scale; VascuQol: Vascular Quality of Life Questionnaire; -ve/+ve: Deterioration/Improvement in condition; WHO-ICD: World Health Organisation - International Classification of Diseases; W-list: Waiting-list.The authors declare that they have no competing interests.MD participated in the design of the study, carried out the systematic literature review, conducted any data analysis and drafted the manuscript. KG provided support in the statistical analysis and helped to draft the manuscript. LS participated in the design of the study, provided support in the statistical analysis and helped to draft the manuscript. MB conceived of the study, participated in the design of the study and helped to draft the manuscript. All authors read and approved the final manuscript.Tables. Table 1: Description of studies that have used the EQ-5D as an outcome measure in clinical and observational studies of patients with cardiovascular disease. Table 2: Summary of studies examining validity and reliability of EQ-5D in cardiovascular disease (n = 10). Table 3: Summary of EQ-5D utility scores reported in cardiovascular studies. Table 4: Canadian Cardiovascular Society (CCS) and New York Heart Association (NYHA) classification systems [ systems -95.Click here for file


The initials for author A. David Redish appear incorrectly in the Author Contributions. The correct initials are ADR. The Author Contributions should read: "Conceived and designed the experiments: ZKN DDR. Performed the experiments: ZKN ADR. Analyzed the data: ZKN ADR. Wrote the paper: ZKN ADR."


Although homeobox genes have been the subject of many studies, little is known about the main amino acid changes that occurred early in the evolution of genes belonging to different classes.In this study, we report a method for the fast and efficient retrieval of sequences belonging to the ANTP (HOXL and NKL) and PRD classes. Furthermore, we look for diagnostic amino acid residues that can be used to distinguish HOXL, NKL and PRD genes.The reported protein features will facilitate the robust classification of homeobox genes from newly sequenced bilaterian genomes. Nevertheless, in non-bilaterian genomes our findings must be cautiously applied. In principle, as long as a good manually curated data set is available the approach here described can be applied to non-bilaterian organisms as well. Our results help focus experimental studies onto investigating the biochemical functions of key homeodomain residues in different gene classes. Genes that belong to the homeobox family are characterized by the ability to code for a protein that contains a recognizable, although very variable, 'homeodomain', usually 60 amino acids in length ,2. Many Here, we report amino acid patterns typical of bilaterian HOXL, NKL, and PRD genes that can be used to quickly and efficiently retrieve amino acid sequences belonging to these classes and subclasses, among hundreds of other homeodomain sequences. Retrieving a given class or subclass of sequences from many animal genomes may be thus an easier task than previously thought. However, we show that these typical amino acid patterns should be cautiously used when sequences come from non-bilaterian animals.Since phylogenetic analysis was one of the primary sources of evidence used to establish the different classes and sub-classes, it is likely that most of homeobox gene classes and subclasses represent monophyletic lineages. Hence, it is conceivable that amino acid changes important for protein function in a given lineage may be revealed as fixed differences between classes and subclasses. Previous attempts have been made to classify genes within HOX families -9 but noWe have used the hand-curated data set of Holland et al. , the PROIn order to find amino acid patterns that distinguish genes from the ANTP subclasses, as well as genes from the PRD class, from genes belonging to other classes, a fast word discovery program  encode this amino acid pattern. None of the human NKL genes encode this amino acid pattern.The pattern LE [AGKNR]E  was buiDrosophila melanogaster AbdB, Drosophila melanogaster btn, Strigamia maritima Hox3b, Gallus gallus HMD2 (PROSITE annotation), Danio rerio HXABA (PROSITE annotation), and Salmo salar HXB2 (PROSITE annotation). These sequences do not form a closely related subgroup of sequences. For instance, the Drosophila melanogaster Abd-B and btn genes belong to two different HOXL gene families. Moreover, other genes belonging to these families do show the HOXL 1 amino acid pattern. The Fugu rubripes HXDBB (PROSITE annotation) and the Gallus gallus HXB8 (PROSITE annotation) sequences are incomplete, and thus it is not possible to determine whether they show this amino acid pattern. In addition to the 194 known HOXL genes showing the HOXL 1 pattern, 178/288 'unclassified' sequences also had this pattern .The HOXL 2 pattern is not as specific as the HOXL 1 pattern, since it is observed in about 46% of the genes belonging to the POU class and in a few NKL (0.5%) and PRD (8%) genes. All 16 PRD genes showing the HOXL 2 pattern belong to the Pax4/6 gene family, thus it is likely that this is a case of convergent evolution. The human POU homeodomains showing HOXL 2 pattern are POU1F1, POU3F1, POU3F2, POU3F3, and POU3F4. The latter four genes are closely related, although the POU1F1 gene is distantly related to those genes  [^DGHMPTVWY] [^CDGKMNPQR] [FL] [^CFILPTWY] [AEFHKQRV] [ADEGKNSTW] [CHKMPQR] [FHY]P (positions 16–26) PRD pattern, use a negatively charged amino acid at position 27 (DE) and a less polar or non-polar amino acid (^RKDENQ) at position 30. About 80% of all animal bilaterian PRD genes follow this description.ANTP  genes show the [HQ] [IV] [AKLT] (positions 44–46) ANTP 1 pattern. In contrast to PRD genes, ANTP genes do not use a negatively charged amino acid at position 27 (DE). Furthermore, at position 30, 99.8% of all bilaterian ANTP genes use a polar amino acid (RKDENQ). 97% of all ANTP bilaterian genes follow this description (data not shown).HOXL genes are characterized by the presence of the HOXL 1 pattern. 96% of all bilaterian HOXL genes show the [KT] [IV]WFQNRR [AMV]K [DEHKLMQWY] [KR] [KR] (positions 46–58) HOXL 1 pattern.NKL genes show the NKL pattern (positions 41–45) or a NKL derived pattern. About 98% of the bilaterian NKL genes follow this pattern. If the NKL pattern is generalized to include the derived NKL patterns then many hits are observed in other homeodomain classes (data not shown). This is why several NKL-related patterns are reported.The above definitions suggest that the region in between amino acid positions 16–30 and 44–58 are the most important for function specificity of genes belonging to different classes and sub-classes. The former region corresponds to the end of Helix 1, the inter Helix 1–2 region and to the first positions of Helix 2 . The regThe situation observed for NKL genes suggest that specific amino acid patterns may exist for groups of genes within major classes, as previously suggested . It alsoIt is tempting to use these features to classify homeobox containing genes. Nevertheless, given the very old age of the gene families considered, the possibility of convergent evolution must be considered when analyzing a given amino acid sequence. Therefore, for the purpose of gene classification, the features here described should not be decisive but rather used as additional evidence. It should be noted, that the reconstruction of gene genealogies is a hard problem, namely when proteins belonging to the same family share some of the same protein interaction partners, thus facing a similar selective environment . Any additional piece of evidence that may shed light on the correct classification of genes should therefore be used.In this work we report a method for the fast retrieval of bilaterian ANTP and PRD sequences. Given the availability of a sufficiently large curated data set this method can be applied to any group of proteins. Furthermore, we report some of the main amino acid changes that occurred early in the HOXL, NKL and PRD lineages. These features can be used for the classification of gene sequences, although, as shown, convergent evolution must be considered as an explanation for the presence of a given pattern in a sequence. The possibility that the region of the protein that allows distinguishing the different classes, also allows the distinction of different families within classes, has important practical and evolutionary consequences and must be explored in more detail.NAF, JV and CPV conceived the design of the study. NAF developed the software needed for the large-scale analyses, while JV and CPV collected, aligned the sequence data, and performed the phylogenetic analyses. JV drafted the manuscript. All authors participated in the results discussion and helped writing the final version of the manuscript. All authors read and approved the final manuscript.Gi numbers of the 251 non-bilaterian non-redundant sequences used for the analysis presented in Table Nematostella vectensis: 32263856; 32263873; 74039490; 74039492; 82621559; 82621587; 82621611; 82621663; 83356315; 156363224; 156363226; 156387518; 156397205Ephydatia fluviatilis: 438584Acropora millepora: 117581722; 117581724; 117581726; 117581728; Anemonia erythraea: 158936936; Ephydatia fluviatilis: 438585; 510584; Halichondria bowerbanki: 33641771; Nematostella vectensis: 32816231; 78190373; 82621509; 82621525; 82621533; 82621555; 82621557; 82621585; 82621591; 82621609; 82621625; 82621657; 82621665; 82621667; 110339021; 110339029; 110339121; 156353243; 156363798; 156367335; 156372762; 156397943; 168693291;Suberites domuncula: 34786940; 49659003; Sycon raphanus: 11066243Anthopleura japonica: 144369330; 144369363; Hydra vulgaris: 2331219; 7635735; Nematostella vectensis: 82570553; 82621543; 82621677; 110339077; 110339133; 156376845; 156402818; 156406963; 156407174Nematostella vectensis: 82621563; 82621631; 82621655; 156366927; 156402173Nematostella vectensis: 82621579; 82621647; 82621651; 110339063; 156388033; 156400908; 156407394Nematostella vectensis: 110339027Baikalospongia intermédia: 62238212; Ephydatia fluviatilis: 1438870; Ephydatia muelleri: 3184520; Hydra vulgaris: 7635740; Nematostella vectensis: 82621517; 82621615; 82621619; 82621623; 82621635; 82621653; 156375827; 156375891; 156376334; 156401296; 156402692; Podocoryne carnea: 28188799; Potamolepis sp.: 62238208; Suberites domuncula: 49659001Anthopleura japónica: 144369334; Cladonema californicum: 9652040; Hydra vulgaris: 3021450; 3021452; 6503072; Nematostella vectensis: 38569883; 78190377; 82395396; 82395402; 82395404; 82395406; 82395408; 82395412; 82395414; 82570541; 82570557; 82570559; 82570563; 110339183; 110339213; 110339215; 110339225; 113120207; 156377162; 156615306;Tripedalia cystophora: 33391193Acropora formosa: 228960; Acropora millepora: 7335704; 7595811; 7595813; 13506878; 117581730; 117581732;Anthopleura japónica: 144369323; 144369326; 144369366; Aurelia aurita: 50841484; Cassiopea xamachana: 4894653; 4894655; 4894659; Cladonema radiatum: 47155918; 47155920; 47155922; Eleutheria dichotoma: 1147626; 91982983; 91982989; Hydra littoralis: 2102728; Hydra magnipapillata: 630481; 630482; 89242120; 144369369; 144369375; Hydra viridis: 7120; 7124; 7130; 83763566; Hydra vulgaris: 4433647; 4838455; 7635731; 7635733; 7635737; 7635742; 9945022; Hydractinia symbiolongicarpus: 2980868; 83272159; Nematostella vectensis: 5081328; 32816237; 74039494; 78190375; 82395398; 82395410; 82570519; 82570521; 82570527; 82570529; 82570537; 82570539; 82570545; 82570549; 82570555; 82570561; 82621507; 82621511; 82621513; 82621515; 82621519; 82621523; 82621527; 82621531; 82621535; 82621539; 82621541; 82621547; 82621549; 82621551; 82621565; 82621567; 82621571; 82621573; 82621575; 82621589; 82621595; 82621601; 82621603; 82621605; 82621613; 82621621; 82621627; 82621629; 82621633; 82621637; 82621643; 82621645; 82621671; 82621675; 83356317; 99030986; 110338989; 110339023; 110339061; 110339099; 110339101; 110339115; 110339171; 110339191; 110339211; 110339247; 113120203; 156356964; 156358580; 156361301; 156361303; 156364678; 156368221; 156371265; 156371439; 156372678; 156374167; 156377158; 156377160; 156377164; 156389076; 156390886; 156393340; 156395806; 156396978; 156398319; 156407912; Podocoryne carnea: 976094; 6118056; 6465862; 7649930; 9964019; 47155914; 62002543; Sarsia sp.: 9988771; Scolionema suvaense: 158936914; Suberites domuncula: 49659005; Trichoplax adhaerens: 38565482


The Open Rheumatology Journal, 2018, 12: 1-10The type of Article has been revised as follows:Review ArticleThe type of article originally provided was:Research ArticleThe revised headings of declarations are mentioned below:CONSENT FOR PUBLICATION:Not applicable.The original headings of declarations provided were:ETHICS APPROVAL AND CONSENT TO PARTICIPATENot applicable.HUMAN AND ANIMAL RIGHTSNo Animals/Humans were used for studies that are base of this research.


Correction to: Trials (2017) 18:22https://doi.org/10.1186/s13063-017-1906-7In the original publication  Figures There were some minor typographical errors on pages 2 and 4 and in the backmatter.Page 2 should have read: ‘There is a clear knowledge gap with regard to effective strategies aimed at recruiters’ ([The backmatter: Trial status: The first recruitment review took place in September 2015, and the last review in August 2016. The first 6-month follow-up took place in March 2016, with the last one in February 2017.’Incorrect name tagging:Last Name: SalmanFirst Name: Rustam Al-ShahiCorrect author name tagging:Last Name: Al-Shahi SalmanFirst Name: RustamFollowing publication of the first correction article , we have


Nature Communications8: Article number: 14475; DOI: 10.1038/ncomms14475 (2017); Published: 02152017, Updated: 06252018The original HTML version of this Article omitted the article number; it should have been ‘14475’. This has now been corrected in the HTML version of the Article. The PDF version was correct from the time of publication.


Retirement preparation program: evaluation of resultsPsychology: Research and ReviewIncorrect version:Retirement preparation program: evaluation of resultsIt should read:The original publication (Psicologia: Reflexão e CríticaPsychology: Research and Review,


This article has been corrected: The correct Grant Support information is given below:GRANT SUPPORTThis work was funded by National Natural Science Foundation of China grant 81472606, Chinese Ministry of Science and Technology grant (973 grant) 2011CB812401, and the Beijing Municipal Government.https://doi.org/10.18632/oncotarget.6461Original article: Oncotarget. 2016; 7:3884-3896.


This article has been corrected: The correct Acknowledgments information is given below:ACKNOWLEDGMENTSThe authors thank Tamar Ziv and Arie Admon  for the proteomic analysis. The authors also thank Antonella Di Pizio and Masha Y. Niv  for performing the structural analysis on predicted NF1 cleavage sites. The authors also thank The De Botton Protein Profiling institute of the Nancy and Stephen Grand Israel National Center for Personalized Medicine, Weizmann Institute of Science for identifying the proteolytic product of NF1. The work was supported by Minerva Foundation Grant.https://doi.org/10.18632/oncotarget.25805Original article: Oncotarget. 2018; 9:31264-31277.


Nature Communications8: Article number: 14907; DOI: 10.1038/ncomms14907 (2017); Published: 04072017; Updated: 05252018The original version of this Article contained an error in the spelling of the author Aysegul Gezer, which was incorrectly given as Aysegul Geyser. This has now been corrected in both the PDF and HTML versions of the Article.


AbstractHierodulapatellifera  occurs several Mediterranean countries, such as Italy. These arrivals could come from many factors: new pets or commercial human transport.Originally from Asia, Hierodulapatellifera  is here reported for the first time in France. A well settled and probably widespread population of this species is here discussed as its adaptability to the Mediterranean climate. Some considerations on the potential impacts on the local ecosystems and its future spreading in Europe as an invasive species are given.The presence of  Hierodulatenuidentata Saussure, 1869 in Italy, in the Po valley . Amongst these groups, insects are widely represented. In parallel, entomologists use, since 1999, the forum hosted on insecte.org to discuss their sigthings and upload them into a gallery of photographs where they are validated by the community. A third widely-used website for Citizen-Science is inaturalist.org. This website, which is also available as a mobile application, has been used to collect collateral information and monitor the presence of o valley .Mantodea male, located in Marseille, was put online with the app INPN Espèces and its identity discussed within the community. Sightings of alien Mantodea were then also highlighted on the forum insecte.org. The results of this research are presented here, highlighting the presence of a new Asian mantid species for France since at least 2013: Hierodulapatellifera .On 1 September 2019, a photo of a In addition, several species of mantids seem to invade Europe from Caucasus  and AsiaRecently, Citizen Science in France relies heavily on the application INPN Espèces. The sightings of insects are based on the website insecte.org and the World’s fauna and flora observations are organised on the online database inaturalist.org.These three methods of publishing sightings in France are presented here:INPN EspècesA genuine tool for raising awareness and knowledge, building on the French biodiversity, INPN Espèces allows everyone to discover, from their mobile phone, the diversity of the species around us and to participate in the biodiversity inventory in France. It is the only app in France that allows us to investigate all the French Fauna and Flora species (mainland and overseas) through their characteristics, distribution or conservation status. Thanks to a data entry eased by images and geolocalisation from one’s mobile, anyone can, with one click, improve the knowledge on the surrounding wildlife.To take part in the country’s species surveying by transmitting our records to experts, only a few clicks are needed to share the sightings. The first identification by the observer is then checked by experts of each group involved in the project.insecte.org (‘Le Monde des insectes’)This website is a gathering places for all insect lovers, whatever their level of knowledge, their approaches and their objectives. A discussion forum is organised according to the different orders of insects. When images in a post are validated collectively, the photos are placed in a gallery that serves as a reference library. Once a year, data are sent to the French national species inventory (INPN).inaturalist.orgOne of the world’s most popular online wildlife databases, iNaturalist helps everyone identify the plants and animals around us. It allows connection to a community of over 750,000 scientists and naturalists who can help us learn more about nature. Moreover, by recording and sharing our sightings, people will create research quality data for scientists who are working to better understand and protect nature. iNaturalist is a joint initiative by the California Academy of Sciences and the National Geographic Society.Mantodea used here follow the latest update  was published, from the 9th district of Marseille  , one with a juvenile at Arles  and, finally, one with a female travelling on a boat in the Black Sea.In parallel, research was initiated on the forum of the website insecte.org, as well as on inaturalist.org. Alien Specimens from Mouriès:Mantodea was observed in an olive grove. It was compared with a female Irisoratoria  on a photo published on insecte.org. During the many exchanges between naturalists, its identity was discussed between Sphodromantisviridis  and Hierodulatranscaucasica Brunner von Wattenwyl, 1878. On 28 September 2015, a new Mantodea female was discovered in the same locality. It seemed to be the same species. It was kept and put in breeding to observe an ootheca and to photograph it. It had been stated that the specimens observed were Hierodulatranscaucasica. The different people involved in the discussion were contacted. Detailed photographs were taken of the second captured female. The exact location of the captures was also noted. Oothecae and an old female were observed on olive trees in early November 2019 . It turns out that it was a female. Its identity was not fixed at the end of the discussions.On 27 July 2019, a nymph Specimen on the Black Sea:M.religiosa. Discussions then orientated for the identification towards H.transcaucasica. A synonymy with Hierodulatenuidentata Saussure, 1869, is suggested on the basis of recent references.On 22 September 2011, an adult female was photographed on a boat on the Black Sea. It was first mistakenly identified as Specimen from Marseille:H.patellifera. It was compared with a male of H.tenuidentata from the Greek island of Skopelos. Some morphological criteria, such as yellowish callous spots of the anterior coxae, are characteristic and can be observed on the photographed specimens. After examining the photographs of females from Mouriès and the nymph (then adult) from Arles, it is possible to identify them as H.patellifera.The male from Marseille was studied morphologically. It is H.tenuidentata.On the other hand, the morphological criteria of the female on the boat on the Black Sea and its geographical location indicate that it is Serville, 18394A66F50F-FA2C-5B96-BDB4-D5106A1E3CF2Type status:Other material. Occurrence: recordedBy: Mattias Perez; individualCount: 1; sex: female; lifeStage: adult; Location: continent: Europe; country: France; stateProvince: Bouches-du-Rhone; county: Mouries; verbatimElevation: 30 m; verbatimLatitude: 43.69858; verbatimLongitude: 4.85217; verbatimCoordinateSystem: decimal degrees; Identification: identifiedBy: Nicolas Moulin; Event: samplingProtocol: hand catching; eventDate: 2013-11-03; eventRemarks: on oliveType status:Other material. Occurrence: recordedBy: Mattias Perez; individualCount: 1; sex: female; lifeStage: adult; Location: continent: Europe; country: France; stateProvince: Bouches-du-Rhone; county: Mouries; verbatimElevation: 30 m; verbatimLatitude: 43.69858; verbatimLongitude: 4.85217; verbatimCoordinateSystem: decimal degrees; Identification: identifiedBy: Nicolas Moulin; Event: samplingProtocol: hand catching; eventDate: 2015-09-28; eventRemarks: on oliveType status:Other material. Occurrence: recordedBy: Gauillaume Paulus; individualCount: 1; sex: female; lifeStage: nymph; Location: continent: Europe; country: France; stateProvince: Bouches-du-Rhone; county: Arles; verbatimElevation: 5 m; verbatimLatitude: 43.673112; verbatimLongitude: 4.633544; verbatimCoordinateSystem: decimal degrees; Identification: identifiedBy: Nicolas Moulin; Event: samplingProtocol: hand catching; eventDate: 2019-07-26; eventRemarks: in a gardenType status:Other material. Occurrence: recordedBy: Didier Aurelle; individualCount: 1; sex: male; lifeStage: adult; Location: continent: Europe; country: France; stateProvince: Bouches-du-Rhone; county: Marseille; verbatimElevation: 45 m; verbatimLatitude: 43.2565769; verbatimLongitude: 5.4097269; verbatimCoordinateSystem: decimal degrees; Identification: identifiedBy: Nicolas Moulin; Event: samplingProtocol: hand catching; eventDate: 2019-09-01; eventRemarks: on a balcony, 6th floorHierodulapatellifera, described from Java  will be updated with all these observations.Hierodulapatellifera occurs in a French department, Bouches-du-Rhône, on the basis of these records. The arrival would be prior to 2013. Several generations and a nymph have been observed. The first individuals could have arrived by Cargo from Asia or could have been voluntarily released, because it is a commonly reared pet. This would not be a continuation of colonisation with the known populations of H.tenuidentata in Northern Italy, in the Po Valley . The hypothesis of displacement by boat is valid because, a few years ago, a female specimen of H.tenuidentata was observed aboard a ship on the Black Sea. Its place of arrival in the boat could not really be located between Ukraine, Turkey, Romania, Bulgaria, Albania and the Danube Estuary. It is, however, well known that this species occurs in these countries  and is represented by one species: Sphodromantisviridis (Finally, the species of viridis .Mantodea must be in charge to identify them.Today, thanks to citizen science, it is important and possible to monitor such invasive alien species. However, the problem of the specific attribution of these specimens through photos is important. Then, some specialists of Vespavelutina (These invasive species could destabilise existing ecological niches by creating competition and ecological pressure, like the Asian Hornet, velutina . At the


Correction to: BMC Genomics (2019) 20:389DOI: 10.1186/s12864-019-5782-2Following the publication of this article , the autIn this Correction article the incorrect and correct author name are shown. The original publication of this article has been corrected.Rona DoyleOriginally the author name was published as:Ronan DoyleThe correct author name is:The publisher apologizes to the authors and readers for any inconvenience caused by this error.


Objectives: To analyze how older adults conceptualize these intergenerational relationships. Methods: In this qualitative study, in-depth interviews were carried out with 316 older adults, aged 65-102, from three different nationalities who lived at home. Verbatim transcripts were examined. Results: Data analysis generated six themes representing intergenerational relationships: affection and reward; interest and integration; grandparent-grandchild interaction quality; privacy and boundaries definition; provision of support; and obligation of providing childcare, on two dimensions of ambivalence concerning their intergenerational relationships . Conclusions: The empirical findings from this research indicate how ambivalence in intergenerational relationships is experienced by older adults and stress the contradictory expectations of older adults with grandchildren. Keywords: Ambivalence; conflict; intergenerational relationships; older adults; support.


AbstractClavicornaltica is a genus of very small flea beetles living in the leaf litter layer of Asian forests, easily sampled with Winkler extraction. The genus is presumably very rich in species, but their taxonomy is hampered by their small size and morphological uniformity.Clavicornalticabelalongensis n. sp., was discovered and taxonomically treated by the course participants. We also present the first DNA barcodes for the genus.On a ‘taxon expedition’-style field course at Kuala Belalong Field Studies Centre in Brunei Darussalam (Borneo), a new species,  We expect this concept may (i) improve the public appreciation for taxonomy and (ii) yield valid taxonomic output , discovered during the same field course, will appear elsewhere .In this paper, we provide partial results from a taxon expedition to the Ulu Temburong rainforest in Brunei, namely the taxonomic treatment of a new species of the leaf litter chrysomelid Callophylum sp., Dacroydes sp., and Horsfieldia sp., 70-80% canopy coverage, few vines, several decaying logs, and a distinct Dicuspiditermes termite presence.We worked in the vicinity of the Kuala Belalong Field Studies Centre (KBFSC) in the Ulu Temburong National Park in the Temburong district of Brunei. We collected specimens at the start of the ‘Ashton Trail’ just west of KBFSC, adjacent to the ‘Earthwatch Plot’ . The areClavicornaltica specimens removed. Our Winkler sampling yielded 12 specimens of Clavicornaltica which we provisionally assigned to four morphospecies, of which one was represented by sufficient individuals to warrant taxonomic description.On 27.09.2018 and 1.10.2018, we carried out the following field workshop. Field course participants collected leaf litter from two microhabitats, sampled separately (~75 l each): (i) open forest floor and (ii) the area between buttress roots. Leaf litter was then passed through a 1-cm mesh-width beetle sieve and subsequently extracted in a Winkler apparatus, the yield of which was preserved in 96% ethanol. This was then sorted under a dissection microscope and all Morphological examinations were carried out in the field lab with a Nikon SMZ445 dissection microscope with 10× eye pieces (magnification up to 35×), a Leica ICC50 HD compound microscope and basic dissection materials. Photographs were made either with a smartphone through the eyepiece of a microscope or on a translucent white acrylic sheet with a Nikon D800e fitted with a Laowa 25 mm ultra-macro lens . Images were processed in Adobe Lightroom and subsequently stacked in Adobe Photoshop CS6. Genitalia and other dissected parts were embedded in polyvinylpyrrolidone . Lengthswww.boldsystems.com).One individual (TxExBr0004w-1) was analysed genetically as follows. DNA was isolated from a sample of abdominal soft tissue using the DNeasy Blood & Tissue Kit (Qiagen) and the extracted DNA was then purified with AMPureXP beads (Beckman Coulter). Around 10 ng of DNA were used in a PCR to generate amplicons for the COI barcoding region, using general primers ; institutionID: UBD; institutionCode: IBER-UBD; collectionCode: Zoology; basisOfRecord: PreservedSpecimenType status:Paratype. Occurrence: recordedBy: Taxon Expeditions field course participants; individualID: TxExBr0004w-2; individualCount: 1; sex: female; lifeStage: adult; preparations: card-mounted; disposition: in collection; Taxon: scientificName: Clavicornalticabelalongensis; kingdom: Animalia; phylum: Arthropoda; class: Hexapoda; order: Coleoptera; family: Chrysomelidae; genus: Clavicornaltica; specificEpithet: belalongensis; taxonRank: species; scientificNameAuthorship: Schilthuizen et al., 2019; nomenclaturalCode: ICZN; Location: locationID: TxExBr0004w; continent: Asia; island: Borneo; country: Brunei Darussalam; stateProvince: Temburong; locality: Kuala Belalong Field Studies Centre; verbatimLocality: Kuala Belalong Field Studies Centre, along Ashton trail; verbatimElevation: 120 m; verbatimLatitude: 4.5472; verbatimLongitude: 115.1571; Event: samplingProtocol: Winkler sampling; samplingEffort: 150 l of forest leaf litter; eventDate: 2018-09-27; habitat: Lowland dipterocarp forest; Record Level: type: PhysicalObject; bibliographicCitation: Clavicornalticabelalongensis (UBDM.3.01172); institutionID: UBD; institutionCode: IBER-UBD; collectionCode: Zoology; basisOfRecord: PreservedSpecimenType status:Paratype. Occurrence: recordedBy: Taxon Expeditions field course participants; individualID: TxExBr0014w-3; individualCount: 1; sex: female; lifeStage: adult; preparations: card-mounted; disposition: in collection; Taxon: scientificName: Clavicornalticabelalongensis; kingdom: Animalia; phylum: Arthropoda; class: Hexapoda; order: Coleoptera; family: Chrysomelidae; genus: Clavicornaltica; specificEpithet: belalongensis; taxonRank: species; scientificNameAuthorship: Schilthuizen et al., 2019; nomenclaturalCode: ICZN; Location: locationID: TxExBr0014w-3; continent: Asia; island: Borneo; country: Brunei Darussalam; stateProvince: Temburong; locality: Kuala Belalong Field Studies Centre; verbatimLocality: Kuala Belalong Field Studies Centre, along Ashton trail; verbatimElevation: 120 m; decimalLatitude: 4.5472; decimalLongitude: 115.1571; Event: samplingProtocol: Winkler sampling; samplingEffort: 150 l of forest leaf litter; eventDate: 2018-09-27; habitat: Lowland dipterocarp forest; Record Level: type: PhysicalObject; bibliographicCitation: Clavicornalticabelalongensis (UBDM.3.01173); institutionID: UBD; institutionCode: IBER-UBD; collectionCode: Zoology; basisOfRecord: PreservedSpecimenType status:Paratype. Occurrence: recordedBy: Taxon Expeditions field course participants; individualID: TxExBr0004w-4; individualCount: 1; sex: female; lifeStage: adult; preparations: card-mounted; disposition: in collection; Taxon: scientificName: Clavicornalticabelalongensis; kingdom: Animalia; phylum: Arthropoda; class: Hexapoda; order: Coleoptera; family: Chrysomelidae; genus: Clavicornaltica; specificEpithet: belalongensis; taxonRank: species; scientificNameAuthorship: Schilthuizen et al., 2019; nomenclaturalCode: ICZN; Location: locationID: TxExBr0004w; continent: Asia; island: Borneo; country: Brunei Darussalam; stateProvince: Temburong; locality: Kuala Belalong Field Studies Centre; verbatimLocality: Kuala Belalong Field Studies Centre, along Ashton trail; verbatimElevation: 120 m; decimalLatitude: 4.5472; decimalLongitude: 115.1571; Event: samplingProtocol: Winkler sampling; samplingEffort: 150 l of forest leaf litter; eventDate: 2018-09-27; habitat: Lowland dipterocarp forest; Record Level: type: PhysicalObject; bibliographicCitation: Clavicornalticabelalongensis (UBDM.3.01174); institutionID: UBD; institutionCode: IBER-UBD; collectionCode: Zoology; basisOfRecord: PreservedSpecimenType status:Paratype. Occurrence: recordedBy: Taxon Expeditions field course participants; individualID: TxExBr0004w-5; individualCount: 1; sex: female; lifeStage: adult; preparations: card-mounted; disposition: in collection; Taxon: scientificName: Clavicornalticabelalongensis; kingdom: Animalia; phylum: Arthropoda; class: Hexapoda; order: Coleoptera; family: Chrysomelidae; genus: Clavicornaltica; specificEpithet: belalongensis; taxonRank: species; scientificNameAuthorship: Schilthuizen et al., 2019; nomenclaturalCode: ICZN; Location: locationID: TxExBr0004w; continent: Asia; island: Borneo; country: Brunei Darussalam; stateProvince: Temburong; locality: Kuala Belalong Field Studies Centre; verbatimLocality: Kuala Belalong Field Studies Centre, along Ashton trail; verbatimElevation: 120 m; decimalLatitude: 4.5472; decimalLongitude: 115.157; Event: samplingProtocol: Winkler sampling; samplingEffort: 150 l of forest leaf litter; eventDate: 2018-09-27; habitat: Lowland dipterocarp forest; Record Level: type: PhysicalObject; bibliographicCitation: Clavicornalticabelalongensis (UBDM.3.01176); institutionID: UBD; institutionCode: IBER-UBD; collectionCode: Zoology; basisOfRecord: PreservedSpecimenType status:Paratype. Occurrence: recordedBy: Taxon Expeditions field course participants; individualID: TxExBr0004w-8; individualCount: 1; sex: female; lifeStage: adult; preparations: card-mounted; disposition: in collection; Taxon: scientificName: Clavicornalticabelalongensis; kingdom: Animalia; phylum: Arthropoda; class: Hexapoda; order: Coleoptera; family: Chrysomelidae; genus: Clavicornaltica; specificEpithet: belalongensis; taxonRank: species; scientificNameAuthorship: Schilthuizen et al., 2019; nomenclaturalCode: ICZN; Location: locationID: TxExBr0004w; continent: Asia; island: Borneo; country: Brunei Darussalam; stateProvince: Temburong; locality: Kuala Belalong Field Studies Centre; verbatimLocality: Kuala Belalong Field Studies Centre, along Ashton trail; verbatimElevation: 120 m; decimalLatitude: 4.5472; decimalLongitude: 115.1571; Event: samplingProtocol: Winkler sampling; samplingEffort: 150 l of forest leaf litter; eventDate: 2018-10-01; habitat: Lowland dipterocarp forest; Record Level: type: PhysicalObject; bibliographicCitation: Clavicornalticabelalongensis (UBDM.3.01175); institutionID: UBD; institutionCode: IBER-UBD; collectionCode: Zoology; basisOfRecord: PreservedSpecimenBody orange-red, small, nearly hemispherical, 1.15-1.30 mm long and 0.9-1.1 mm wide (i.e. ca. 1.25 times as long as wide) , densely beset with confluent double punctuation; tubercles and midfrontal sulcus poorly developed and inconspicuous. Eyes convex, each eye consisting of 26-30 ommatidia, ca. 1/5 the width of the head measured across the eyes in dorsal view. Antennae: clava long and moderately robust.Pronotum: Very weakly shagreened and punctuated; punctures sparse and minute, of similar strength to the subordinate punctuation on the elytra; pronotal surface therefore shiny. Lateral margin with a callosity that stretches from the anterior to the posterior corner. Lateral setiferous pore at 2/3 of the length of the margin, seta as long as the clava of the antenna, pore removed from the margin by a distance similar to the width of antennomere II. Posterior setiferous pore placed directly at the margin, the seta length similar to antennomeres IX+X.Hind wings: Absent.Elytra: Shiny, punctate in 9 longitudinal rows, scutellar row ¼ the length of the other rows, consisting of ca. 6 punctures. Punctures in all rows deeply impressed along their entire length (puncture width is similar to their interspaces). In between, the major punctures are irregularly scattered and there are much smaller subordinate punctures. A rudiment of a 10th row exists in the final 1/3 flanking the elytral margin. A fine groove runs along the entire margin continuing to the apex; apex itself slightly drawn out. The internal edge of epipleura carries a short row of punctures, alongside the 4th and 5th visible sternite.Legs: Tibia and tarsus orange, femur dark orange and robust. Metafemur robust, oval, covered in reticulate microsculpture. External edge of metatibia bearing two parallel rows of 8-10 minute stiff setae placed along the terminal one-fifth and flanking the basis of metatarsomere I. Internal side of metatibia bearing ca. 10 thin setae that are placed along the terminal half of the tibia and increase to about 2.5× the length of the external setae, then decrease in length towards the apex. The metatibia carries a long terminal spine of about the same length as metatarsomere I. No serrations or microteeth are visible on the spine.Mesosternum: Processus rounded, with a distinct margin, central area somewhat convex.Abdomen: Carina on the first visible abdominal sternite sharp and narrow, not broadened anteriorly or posteriorly, running along the length of the sternite. In reduced form, this carina is carried on to the four posterior sternites, which therefore, in lateral view, offer a slightly serrated aspect. The surface of the sternites carries a rough microsculpture of confluent punctures. Tergum IX (last visible tergite) with three longitudinal median ridges, the central one of which is much weaker than the two outermost. Subapically, tergum IX has a horizontal row of 8 serrations.Female genitalia: Spermatheca consisting of a pear-shaped receptacle, ca. 90 µm in length, with crosswise annulations Clavicornalticabelalongensis sp. n. differs from all other known Clavicornaltica are (i) the pear-shaped spermathecal receptacle that is distinctly separated from the pump and (ii) the medially keeled abdominal sternites. Furthermore, the new species can be separated from other oriental Clavicornaltica in the following respects: C.fortepunctata Scherer, 1974 (Vietnam) is more elongate  is black, the pronotum is impunctate and it is also much larger (1.9 mm)  have impunctate elytra  is larger (>1.5 mm)  is reddish-black and the elytral punctuation is reduced  has a widened first protarsomere and an anteriorly widened carina on the 1st abdominal segment  has a wide plate on the underside of the 1st abdominal sternite; Clavicornalticatrautneri Medvedev, 1993 is much larger (2.1 mm)  has a posteriorly narrowed pronotum  has an anteriorly widened carina on the 1st abdominal segment  has the spermathecal pump fused with the receptacle and a widened carina on the first abdominal segment  has the spermathecal pump fused with the receptacle  has the mesosternal processus flat, not convex  has the spermathecal pump wider than the receptacle  has vaginal palpi very short and the spermathecal pump wider than the receptacle  is 1.6 times as long as wide and has the carina on the 1st abdominal sternite widened posteriorly  is larger and more elongate; its head is not shagreened  has a spherical spermathecal receptacle, the carina on the 1st abdominal sternite is widened anteriorly and is flanked by rows of strong punctures, the metafemur is more elongated and the vaginal palpi are diverging, not parallel  has a much more elongate habitus  has impunctate elytra and a black body  ; Clavicoe elytra ; Clavico>1.5 mm) ; Clavico reduced ; Clavico segment ; Clavico(2.1 mm) ; Clavicopronotum ; Clavico segment ; Clavico segment ; Clavicoceptacle ; Clavicot convex ; Clavicoceptacle ; Clavicoceptacle ; Clavicoteriorly ; Clavicoagreened ; Clavicoparallel ; Clavico habitus ; finallyack body .Clavicornalticabelalongensis Schilthuizen et al., 2019, provided the full citation of this publication appears in the bibliography or elsewhere in the referring work.The species is named after the Belalong river; the new species was recorded in the close vicinity of the river’s left bank. Following Article 51C of the Code , the speKnown only from a location near the confluence of the Belalong and Temburong rivers, at 120 m elevation . However, as pointed out by Clavicornaltica, which currently stands at 26 species, has barely scratched the surface of the true diversity. Given the morphological inaccessibility of this diversity, we expect DNA barcoding to be of some use. We hope the sequence we present here for C.belalongensis will be a starting point for further building up a database of DNA barcodes for this genus.These and previous results (ors e.g. , the speors e.g.  suggesteDespite the present single-species description based on limited material from a single locality, we believe that taxonomic work is best carried out in the context of larger revisions. However, we think that concise treatments of single species such as we present here, have value . As reso


Nature Communications8:15223 doi: ; DOI: 10.1038/ncomms15223 (2017); Published 05122017; Updated 05252018The originally published version of this Article contained an error in Fig. 1. In panel d, the uppermost western blot was inadvertently inverted during typesetting of the figure. This has now been corrected in both the PDF and HTML versions of the Article.