Tbx5 and Tbx4, two genes expressed in forelimb and hindlimb-forming regions respectively, play crucial roles in the initiation of limb outgrowth. Evolution of regulatory elements that activate Tbx5 in rostral LPM was essential for the acquisition of forelimbs in vertebrates. We identified such a regulatory element for Tbx5 and demonstrated Hox genes are essential, direct regulators. While the importance of Hox genes in regulating embryonic development is clear, Hox targets and the ways in which each protein executes its specific function are not known. We reveal how nested Hox expression along the rostro-caudal axis restricts Tbx5 expression to forelimb. We demonstrate that Hoxc9, which is expressed in caudal LPM where Tbx5 is not expressed, can form a repressive complex on the Tbx5 forelimb regulatory element. This repressive capacity is limited to Hox proteins expressed in caudal LPM and carried out by two separate protein domains in Hoxc9. Forelimb-restricted expression of Tbx5 and ultimately forelimb formation is therefore achieved through co-option of two characteristics of Hox genes; their colinear expression along the body axis and the functional specificity of different paralogs. Active complexes can be formed by Hox PG proteins present throughout the rostral-caudal LPM while restriction of Tbx5 expression is achieved by superimposing a dominant repressive (Hoxc9) complex that determines the caudal boundary of Tbx5 expression. Our results reveal the regulatory mechanism that ensures emergence of the forelimbs at the correct position along the body. Acquisition of this regulatory element would have been critical for the evolution of limbs in vertebrates and modulation of the factors we have identified can be molecular drivers of the diversity in limb morphology.Tight control over gene expression is essential for precision in embryonic development and acquisition of the regulatory elements responsible is the predominant driver for evolution of new structures.  Tbx5. Activation of Tbx5 in the forelimb-forming region of the developing embryos is essential for forelimbs to form and disruption of human TBX5 causes limb abnormalities. We show that activation of Tbx5 in a restricted territory is achieved through a combination of activation inputs that are present broadly throughout the embryo flank and dominant, repressive inputs present only in more caudal regions of the flank. The sum of these inputs yields restricted activation in the rostral, forelimb-forming flank. Our results explain how the regulatory switches that were harnessed for the acquisition of limbs during evolution operate and how they can be turned off during the evolution of limblessness in species such as the snake.The acquisition of limbs during vertebrate evolution was a very successful innovation that enabled this group of species to diversify and colonise land. It has become clear recently that the primary driver behind the evolution of new structures, such as limbs, is the acquisition of novel regulatory elements that control when and where genes are activated rather than the proteins encoded by the genes themselves acquiring novel functions. We have identified the regulatory element from a gene,  Tbx5 in nascent forelimbs and Tbx4 in hindlimbs Fgf10 in the limb mesenchyme Fgf8 expression in the apical ectodermal ridge (AER) and Fgf8 produced from the AER, in turn, maintains Fgf10 expression in mesenchyme to establish a positive feedback loop of Fgf signalling that maintains limb growth. Mutations in human TBX5 cause Holt-Oram Syndrome (HOS OMIM142900), a disorder characterised by upper limb and heart abnormalities TBX4 cause Small Patella Syndrome (SPS OMIM 147891), a disorder characterised by knee, pelvis and toe defects Tbx5 is the earliest marker of presumptive forelimb mesenchyme and because activation of this factor within a defined region of the LPM ultimately dictates the position at which the forelimbs will arise, identifying the factors that control activation of this Tbx5 expression domain will reveal the mechanisms employed that allowed the acquisition of limbs in vertebrates and that dictate forelimb position in the embryo.Forelimbs and hindlimbs are derivatives of the lateral plate mesoderm (LPM) that arise at fixed positions along the vertebrate body axis. Limb formation is initiated by limb induction signals from axial tissues Tbx5 is initially expressed in the forelimb-forming region of LPM prior to the emergence of a bud and it is subsequently restricted to the forelimb region as development proceeds. Tbx5 is essential for forelimb formation and this exclusive requirement is limited to a short time window when limb bud initiation occurs Tbx4, the paralog of Tbx5, is able to rescue forelimb formation following conditional deletion of Tbx5Tbx4/5 gene represented by AmphiTbx4/5 of the limbless cephalochordate, amphioxus, can fully compensate for the loss of Tbx5 in the mouse Tbx5 expression in the LPM was a critical step in the acquisition of limbs during vertebrate evolution.Hox genes are conserved homeodomain-containing transcription factors that are arranged in clusters in the genome. The chromosomal organization of the genes in the complex reflects their expression pattern along the rostro-caudal body axis to determine positional identity Tbx5 regulatory element sufficient for early forelimb expression Tbx5. However, since the ability to activate Tbx5 is not strictly restricted to Hox genes expressed only at forelimb level, the mechanism by which a rostro-caudal Hox code establishes forelimb-restriction of Tbx5 remained unknown. Here, we demonstrate how Hox paralogous group members act cooperatively to restrict expression of Tbx5 in the LPM, which ultimately determines the positions the forelimbs will emerge from the flank of the embryo. We show that mutations of a single Hox binding site in the Tbx5 forelimb regulatory element cause expanded reporter gene expression in caudal LPM. Rostral restriction in Tbx5 expression through repression in the caudal LPM is mediated by Hoxc8/9/10 genes and this repressive function is limited to Hox genes that are expressed in Tbx5-negative caudal LPM. We further map the Hoxc9 protein domains required to confer transcriptional repression that distinguishes these paralogs from other Hox proteins expressed throughout the flank of the embryo. Our results demonstrate how a nested, combinatorial code of Hox protein transcriptional activation and repression along the rostro-caudal embryo axis restricts Tbx5 expression to the forelimb and ultimately determines forelimb position.We have previously identified a Tbx5 gene that recapitulates the dramatic forelimb-restricted expression of this gene Tbx5 expression, we generated a series of constructs in which each individual Hbs site1-6 is mutated and tested their ability to activate a LacZ reporter gene in transgenic mice. While the Tbx5 int2(361) reporter construct drove forelimb-restricted expression of LacZ (Tbx5 int2(565)) . To distinguish the requirement for Hbs2 and 6bp3′, we next mutated either of these sites )  that cont2(565)) . We haveess Tbx5 . These rse sites . Mutatioreporter , while mreporter  equivalereporter . These rThese results demonstrate that the binding sites within this regulatory element can be divided into 2 distinct functional groups. Hbs1 and 3-6 act as ‘on’ switches important for the amplitude of activation, whereas Hbs2 determines spatial resolution by hosting repressive complexes that restrict the domain of activation. This element can therefore have a binary function, serving as a site for the formation of transcriptional activation or repression complexes.Hox genes can activate this regulatory element Hox genes in chick and mouse embryos at stages when Tbx5 is first expressed in the forelimb-forming region. Tbx5 is first expressed at the level of somites 13–20 in chick and somites 4–11 in mouse embryos. As previously reported Hox4 and Hox5 paralogs overlap with that of Tbx5 in both mouse and chick embryos   construct  reporter, LacZ expression is repressed in the forelimb region   LacZ repficiency ), β-gal ficiency  indicatireporter  demonstrTbx5 expression, we co-electroporated Hoxc9 with a Tbx5 int2(361) reporter in which Hbs2 is mutated  showed clear repression of the reporter demonstrating that within the limits of this assay the entire N-terminus or the domain overlapping the junction between N1 and N2 is required for repressive activity  can reduce LacZ expression , suggestTbx5 as an assay, we have been able to distinguish the opposing transcriptional activities of different Hox paralogous group proteins. Hoxc9, as well as Hoxc8 and Hoxc10, that are normally expressed in the LPM caudal to the forelimb, can repress Tbx5 to restrict its expression to the forelimb level region of the LPM  is required for its repression in caudal LPM (Our results demonstrate that five of the six Hox binding sites (namely Hbs1 and Hbs3-6) within the udal LPM . One posudal LPM , suggesthttp://www.genomatix.de) is the presence of a 6-bp sequence named Pbx1-Meis1 complexes site located 3′ of Hbs2 (6bp3′). Pbx is a Hox co-factor that can attenuate Hox-mediated gene transcription by recruiting histone deacetylases (HDACs) Tbx5 forelimb regulatory element.An alternative model is that the transcriptional activity of the Hox complex bound at Hbs2 is determined by a co-factor(s). The sequence of Hbs2 is identical to the sequences of Hbs1 and Hbs3, therefore we compared the sequences surrounding these Hox binding sites. One distinguishing feature of Hbs2 identified using Mat Inspector  in the protein. The hexapeptide of AbdA represses dpp expression by inhibiting the function of a glutamine (Q)-rich C-terminal activation domain Tbx5 transcription, we suggest the model that Hoxc9 supresses Tbx5 expression by interaction with co-repressor(s) (in vitro translated proteins or nuclear extract from LPM (data not shown). Other potential collaborators are Smad proteins. In the Drosophila haltere, a Mad/Med/Shn complex works in combination with Ubx to repress Sal expression Tbx5 expression . Other candidate repressors are engrailed (En) and sloppy paired (Slp) since, in Drosophila, they form a complex with Hox, Exd and Hth to repress transcription Engrailed1 and Engrailed2, are expressed in LPM at pre-limb bud stages As it is unlikely that Hox protein itself directly represses essor(s) . One canTbx5 expression or that they share similar 3D structure domains in spite of their distinct amino acid sequences.We have shown that Hoxc8 and Hoxc10 have transcriptional repression ability similar to Hoxc9 . To gainTbx5 forelimb regulatory element reveals a direct link between patterning of the rostro-caudal axis of the embryo by Hox genes and the programme that controls positioning of the forelimb forming territory. A clear correlation between Hox expression and establishment of the forelimb territory of the LPM has previously been suggested Tbx5Hoxc9 is reduced Tbx5 (and the subsequent forelimb programme) being essential for emergence of an ectopic wing bud from this region. In the limbless python, Hoxc8 expression is rostrally expanded to the anterior limit of the trunk Tbx5-negative caudal LPM at pre-limb bud stages in chick and mouse , however, we did not detect expression of Hoxc9 in the forelimb-forming region, in contrast to the strong staining in caudal tissues. We therefore conclude that Hoxc9 is not present in the forelimb-forming region at stages when Tbx5 expression is first initiated. Later expression of Hoxc9 is not sufficient to cause detectable repression of the domain of Tbx5 already activated by Hox4/5 paralogous genes.A previous study has demonstrated the presence of and a function for Tbx5 expression, our study does not exclude the possibility that other caudally-expressed Hox genes have a similar repressive ability. We favour a model in which other caudally-expressed Hox paralogs have redundant functions in repression of Tbx5. Hoxc cluster null mice have no defects in the limb skeleton Tbx5 in these mutants have not been reported and we predict that the ectopic expansion of Tbx5 in caudal LPM would not cause any skeletal defects. Further analysis will be required to uncover the requirement of caudally–restricted Hox paralogs, such as Hox8, Hox9 and Hox10 for Tbx5 repression in caudal LPM.While we have shown that Hoxc8, Hoxc9 and Hoxc10 can repress Tbx5 in the LPM, a similar Hox protein code is present in axial tissues  that do not express Tbx5. The activity of the forelimb regulatory element of Tbx5 is restricted to LPM and this LPM restriction is maintained following mutation of Hbs2 that leads to caudal expansion in expression. One possible explanation for LPM restriction is the presence of unknown repressors in axial tissues or alternatively additional factors, which are active exclusively in LPM, are required for Tbx5 expression. Odd-skipped related (Osr) genes are candidates as they are expressed in LPM, but excluded from axial tissues such as neural tube and somites Tbx5 forelimb regulatory element to test if reporter activity was lost. The activity of the element was unaffected, however, suggesting Osr genes are not required for Tbx5 LPM expression  complex that ultimately determines the caudal boundary of Tbx5 expression. Thus, the regulation of Tbx5 expression in the LPM represents an excellent system to understand the interactions between neighbouring Hox binding sites and how the consequent output is integrated.Our analysis of the http://www.genomatix.de). Transgenic embryos were generated by the Procedural Service section, NIMR by standard pronuclear microinjection techniques. Mouse embryos were staged according to LacZ transgene were identified by PCR using specific primers . Sequences surrounding putative Hox binding sites and the mutations induced are as followings, binding sites are shown in bold; Hbs1, ATTATTGGAAC; mut Hbs1, ATGCTTGGAAC; Hbs2, ATTATCGACTCTCA; mut Hbs2, ACGATCGACTCTCA; mut 6bp3′, ATTATCGACTGCAA; mut Hbs2+6bp3′, ACGATCGACGCTTA; Hbs3, TAATTCAGA; mut Hbs3, TCGTTCAGA; Hbs4, TTATTAAGGCC; mut Hbs4, TTGGCAAGGCC; Hbs5, TTATCTTGCCAT; mut Hbs5, TCGTCTTGCCAT; Hbs6, TTATTTTG; mut Hbs6, TCGTTTTG.For reporter analysis in chick and mouse, we used the BGZA reporter vector in situ hybridizations were carried out essentially as previously described Pitx1, Tbx5 and mouse Hox genes have been described previously Whole mount Fertilized chick embryos  were incubated at 38°C and staged according to Hamburger Hamilton (HH) In vitro translated proteins were produced using a TnT Coupled Reticulocyte Lysate System (Promega). Proteins were labelled with 35S-Methionine (PerkinElmer) to verify and quantify translation. LPM strips adjacent to somites 5–10  and lateral to somite 14 to its caudal extreme  were dissected from E9 mouse embryos. Nuclear extracts were prepared using the NE-PER Nuclear and Cytoplasmic Extraction Kit (Pierce) following manufacturers instructions.32P by incubating with T4 polynucleotide kinase (NEB) for 30 minutes. 2 µl of in vitro translated protein or nuclear extract were blocked with 200 ng poly-dIdC, 2 µg of poly-dGdC or 2 µg of poly-dAdT in binding buffer  in a total volume of 22 µl for 15 minutes on ice. For super-shift, 2 µl of the antibody recognising flag epitope  was added to the binding reaction and incubated for a further 15 minutes. Then, 1 µl of 32P -labelled double-stranded oligonucleotides were mixed and incubated for 30 minutes. The protein∶DNA hybrids were resolved on 6% PAGE in 0.5xTBE.Double-strand oligonucleotides were labelled with Figure S1Tbx5 forelimb regulatory element. A–C. The Tbx5 int2(361) LacZ reporter construct was electroporated with pCAβ-dsRed-Express in the presumptive forelimb region of HH14 chick embryos. 24 hours later, the efficiency of electroporation was assessed by dsRed expression (A–C) and embryos were stained for LacZ to analyze the enhancer activity (A′–C′). Co-electroporation of pcDNA-mHoxc8 (B and B′) and pcDNA-mHoxc10 (C and C′) reduced LacZ expression.Hoxc8 and Hoxc10 can repress activity of the (TIF)Click here for additional data file.Figure S2LacZ reporter expression. A, Schematic representation of the Tbx5 intron2 regulatory element. A putative smad binding site and Hox binding sites are shown as green and blue boxes, respectively. B–C, E9.5 embryos were stained for β-galactosidase for a mutated construct of Tbx5 int2(565) reporter.Disruption of a putative smad binding site does not expand (TIF)Click here for additional data file.Figure S3LacZ reporter expression. A, Schematic representation of the Tbx5 intron2 regulatory element. A putative Osr binding site (purple box) locates proximal to Hbs5 (blue box). B–C, E9.5 embryos were stained for β-galactosidase for mutated constructs of Tbx5 int2(565) reporter.Disruption of a putative Osr binding site does not affect (TIF)Click here for additional data file.


Objective: The purpose of the study was to identify technical item flaws in the multiple choice questions submitted for the final exams for the years 2009, 2010 and 2011.Methods: This descriptive analytical study was carried out in Islamic International Medical College (IIMC). The Data was collected from the MCQ’s submitted by the faculty for the final exams for the year 2009, 2010 and 2011. The data was compiled and evaluated by a three member assessment committee. The data was analyzed for frequency and percentages the categorical data was analyzed by chi-square test.Results: Overall percentage of flawed item was 67% for the year 2009 of which 21% were for testwiseness and 40% were for irrelevant difficulty. In year 2010 the total item flaws were 36% and 11% testwiseness and 22% were for irrelevant difficulty. The year 2011 data showed decreased overall flaws of 21%. The flaws of testwisness were 7%, irrelevant difficulty were 11%.Conclusion: Technical item flaws are frequently encountered during MCQ construction, and the identification of flaws leads to improved quality of the single best MCQ’s. Altering the mode of examination to single best question type represents a major challenge for a faculty in any medical college. With change in the curriculum, the modality of assessment also changes. The new system of examination focuses on application, problem solving and integration of the different concepts taught. In Islamic International Medical College the mode of examination based on one best question is being practiced since 2009.1 Properly made MCQs leads to impartial testing of the student that can measure knowledge, comprehension, application and analysis.2,3MCQs or single best questions are difficult and time-consuming to construct, even for those who have been formally trained in the construction of MCQs.4 Questions that aim to assess really important topics cannot do so unless they are well-structured i.e. avoiding flaws that benefit the testwise examinee; those students who answer questions alone on their test taking skills and not on their amount of expertise on the subject that is being covered.5 Also avoiding irrelevant difficulty are prerequisites that must be met in order for test questions to generate valid scores.5Characteristics of effective MCQs can be described in terms of the overall item, the stem, and the options. The stem generally consist of a clinical case presentation and a lead-in question, followed by a series of choices, typically one correct/best answer and four distractors.3,4 Flawed MCQs interfere with accurate and meaningful interpretation of test scores and negatively impact student pass rates. One aspect where many MCQs fail is in having effective distractors.5 Teachers often spend a great deal of time constructing the stem and much less time on developing plausible options to the correct answer.5Outlines regarding effective item-writing have been documented; however manipulations of these principles are very common in medical education with resultant flawed item questions.4,5 Flaws related to irrelevant difficulty make the question difficult for reasons unrelated to the trait that is the focus of assessment. The increased test and item difficulty associated with the use of flawed items lead to artificial difficulty to the test scores.6Two types of technical item flaws: testwiseness and irrelevant difficulty are described in literature. Flaws related to testwiseness make it easier for some students to answer the question correctly, based on their test-taking skills alone.The purpose of this study was to examine structural concerns which are important for the formation of high-quality test questions. Thus the main objective of this study was to identify common technical flaws in assessment items encountered during the paper setting of examinations of 2009 to 2011.This descriptive study was conducted at Riphah University Rawalpindi after the completion of assessment for the year 2011. There were no human subjects involved in the study. Therefore it was exempted for obtaining an ethical approval certificate.The assessment data for the years 2009, 2010 and 2011 was collected and reviewed. These items were reviewed by a three member assessment committee. The original single best choice questions that had been submitted to the assessment committee for the purpose of exams were grouped according to the year and were then analyzed for technical item flaws.Inclusion criteria: All questions submitted for the years 2009, 2010 & 2011. During analysis intrinsic structure of the question was checked for technical accuracy. Items were classified as 'flawed' if they contained one of the flaws. Frequently observed flaws were grouped into:1- Issues Related to TestwisenessGrammatical Cues and errorsLogical cuesoften, sometimes in MCQ) terms.5Use of absolute  is 46.2% which is a study conducted on nursing curriculum.9 However, the evaluation of the data from 2011 showed that overall the total item flaws were 21%. This is substantially less as compared to data seen from the year 2009 and 2010.While much has been written in context of developing a good MCQ, there is very little actual data concerning the analysis of a MCQ. The results in this study show that the frequency of item flaws encountered in the year 2009 were 67% which is comparable to flaws encountered in a study conducted by Ellsworth et al in psychology test banks.4,5In the present, study a number of violations were found that help students correctly answer questions based on cues given in the stem or the options, rather than knowledge. Item writing flaws (IWFs) such as longest correct option, logical cues, word repeats, use of ‘‘all of the above,’’ and use of absolute terms make MCQs easier by providing helpful cues to students as to what is the correct answer. MCQ’s with heterogenous options apparently increase the difficulty of a question and deal with miscellaneous facts.10 Though that study was analysis of data from module exams that were conducted during the academic year, the proportion of the flaws related to testwiseness steadily decreased in 2010 and 2011.This study analyzed these item flaws encountered during the analysis of item submitted across three years. The frequency of flaws related to testwiseness were 21% and those which were correlated to irrelevant difficulty were 40% for the year 2009. This percentage of item flaws is comparable to a study by Danish in 2010 which had similar percentage of item flaws.11 Research in other disciplines has shown that training improves the quality of MCQs developed by teaching faculty.12 Despite the fact that in the year 2009, the faculty had been trained for the MCQ writing, lack of practice was a recognizable factor contributing to the high number of errors in items analyzed for the year 2009 and 2010. As the faculty preceded from 2009 into 2010 and then 2011, the level of MCQ construction and item writing skills of the faculty improved. Moreover, relevant feedback regarding removal of errors give good results in improving the quality of items prepared by the faculty members.The identification of these item writing flaws also highlights the fact that the faculty members preparing these items should be trained in a faculty development workshop.It is also suggested to the faculty members that in order to combat the issues of irrelevant difficulty, while preparing for lecture the relevant MCQ be prepared at the same time. Most often important assessment are written and assembled at the last moment. While the faculty members take lecturing seriously few make an effort to prepare the assessment. Furthermore, by the time items are collected from half a dozen or more lecturers who may have been involved in teaching there is inadequate time or opportunity to review before being submitted to the assessment committee.For planning an effective assessment it is emphasized that the items prepared by the faculty members should be carefully analyzed before they are put in an evaluation paper. The current research points in detail the types of mistake that are mainly committed in the construction of MCQs. It also gives guidelines for authors of the MCQ items about the common error committed during the preparation of MCQ’s.If an MCQ is going to be used to assess higher order cognitive skills, there needs to be a process in place where adequate instruction and feedback is given to the item authors. The results verify that with repetition and practice the standard of MCQ for assessment paper’s can be improved. To ensure better quality of MCQ it is suggested that the items before being submitted to the assessment committee should be evaluated at inter departmental level and then submitted to the finalizing committee. This will lead to better written items and save time as well.Technical item flaws are frequently encountered during MCQ construction, and the identification of these flaws leads to improved quality of the single best MCQ’s. In order to rectify these flaws the faculty should be trained in item writing skills. While on the spot training can be done at the time of assessment but better results can be obtained if the faculty is trained prior to the final exams.


Eyelid involvement in sarcoidosis is very rare. A search of the medical literature indicates one previous report of sarcoidosis with destructive eyelid lesions. We describe the case of a 50-year-old woman with severe systemic sarcoidosis, which included her eyelids. To our knowledge, the case presented herein represents the first to show the full-thickness histopathology of destructive eyelid lesions in sarcoidosis. Sarcoidosis is a multisystem inflammatory disease of unknown etiology characterized histopathologically by noncaseating granulomas. The condition has a female predilection with a peak incidence in the 3rd and 5th decades of life. Pulmonary involvement is the most common manifestation. Common extrapulmonary manifestations include the skin, central nervous system, liver, kidney, and musculoskeletal system. Furthermore, sarcoidosis can affect the heart, peripheral nervous system, salivary glands, eye, and ocular adnexa. Ocular involvement occurs in 25%–60% of patients with systemic sarcoidosis , 2. WhilA 50-year-old female with severe systemic sarcoidosis, including pulmonary, skin, and joint disease, presented for evaluation of ocular involvement. The patient had noted eyelid nodules for over 5 years; however, she recently developed eyelid deformities, tearing, and eye irritation.On examination, she had notching of her upper and lower eyelids and extensive scarring of the posterior lamella leading to entropion and focal trichiasis . Slit laThe patient subsequently underwent full-thickness wedge resections in both upper eyelids and her right lower eyelid resulting in symptomatic improvement and elimination of the localized areas of trichiasis. Histopathologic evaluation of all three full-thickness wedge resections showed typical “naked,” noncaseating granulomas composed of epitheliod histiocytes and multinucleated giant cells . Serial  Early reports have limited discussion of the incidence of eyelid involvement in sarcoidosis , 5. Eyelhttp://www.ncbi.nlm.nih.gov/pubmed/, accessed 10 March 2013) of “eyelid and sarcoid” indicates one previous case [A Pubmed literature search (ous case  of involous case . The autous case . To our


Here, we report from the Afrotropical greater honeyguide the first detailed observations of honeyguides killing host chicks with their specially adapted bill hooks, based on repeated video recordings . Adult greater honeyguides puncture host eggs when they lay their own, but in about half of host nests at least one host egg survived, precipitating chick killing by the honeyguide hatchling. Hosts always hatched after honeyguide chicks, and were killed within hours. Despite being blind and in total darkness, honeyguides attacked host young with sustained biting, grasping and shaking motions. Attack time of 1–5 min was sufficient to cause host death, which took from 9 min to over 7 h from first attack. Honeyguides also bit unhatched eggs and human hands, but only rarely bit the host parents feeding them.The most virulent avian brood parasites obligately kill host young soon after hatching, thus ensuring their monopoly of host parental care. While the host eviction behaviour of cuckoos (Cuculidae) is well documented, the host killing behaviour of honeyguide (Indicatoridae) chicks has been witnessed only once, 60 years ago, and never  While mthe nest . Second otropics  and the otropics , the parotropics . The repotropics  .Prodotiscus show bill hooks and host young have never been found alongside them [Nestlings of only five of the 17 honeyguide species have ever been observed, but since all including the basal genus ide them , it is aIndicator indicator is remarkable for its interactions with other species. It shows a unique mutualism with human honey-gatherers whom it guides to bees' nests [The Afrotropical greater honeyguide s' nests , as wells' nests ; (ii) th2.ca 5000 ha area centred on 16°45′′ S, 26°54′′ E. The habitat is mixed miombo woodland, seasonally flooded depressions and tobacco cultivation. Here, the little bee-eater Merops pusillus [ca 0.5 m long and 0.2–0.5 m underground) typically dug into the roofs of aardvark (Orycteropus afer) holes, or sometimes earth banks or bushpig (Potamochoerus larvatus) diggings. Clutch size is three to six eggs.We conducted fieldwork in the Choma region of southern Zambia during September–November 2008–2010, in a pusillus  is the cWe filmed chick behaviour by inserting an infrared camera at the end of the bee-eaters' access tunnels. Cables led to the surface where a digital recorder was concealed nearby. Bee-eaters ignored this equipment. Killing behaviour was filmed at five nests of three host species , involving four honeyguide chicks and six host chicks. In two nests, we had moved the honeyguide from another nest, matched in developmental stage, but attacked by ants or predators; foster nests are excluded from mass analyses. Data on honeyguides' impact on hosts all refer to the commonest host species, the little bee-eater.3.n = 194) of host eggs in parasitised nests were punctured. In 30 of 55 parasitised nests (55%), at least one (range one to four) host egg was unpunctured and could have hatched; proportions were similar for other host species. Hence, egg puncturing only partially removed the scope for subsequent chick killing.Of 172 little bee-eater nests followed, 113  were visited by a honeyguide, as revealed by a honeyguide egg and/or punctured host eggs. However, of these visited nests, 64 (56.6%) were deserted by hosts at this early stage. Therefore, a parasitic egg was incubated by hosts in only 49 (28.5%) of all host nests. Honeyguide oviposition occurred any time from the start of host egg-laying to late into the host incubation period. Greater honeyguides did not remove host eggs, but  to hatch, compared with 18–20 days for hosts [ca 2–4) days old when the first host chick hatched. At hatching, honeyguides weighed ca 3 g, possessed fully developed bill hooks (a) and immediately attempted to bite our fingers when handled. Honeyguides had attained substantially greater weight by the time host chicks hatched and were killed: 6.06–11.08 g  for the first-hatched host and 10.22–13.29 g  for the last-hatched. Hosts were attacked soon after hatching , when little bee-eaters weighed less than 2 g .Honeyguides always hatched before host young, despite sometimes asynchronous laying . Honeygll hooks a and imma–c and electronic supplementary material, videos S1–S4). They typically held onto hosts for a period of time  during which they repeatedly opened and closed their mandibles, and often shook the host chick, either from side to side or up and down by rocking on their hindlegs . Honeyguides reached out haphazardly rather than targeting particular body regions. They most commonly bit the back (n = 20 bites), head (14) and abdomen (14), and occasionally the neck (5), wings (4) and legs (3). Biting rarely caused open wounds, but rather haemorrhaging underneath the skin and heavy bruising (figure 2b). As previously noted [figure 2c). Host chicks sustained an average of 177 s of direct attacks , but from the time of first attack, hosts took from 9 min to over 7 h to die. Hosts did not attempt to avoid attacks and after a first bout of attack soon ceased any attempts to beg for food. Host corpses usually remained and decomposed in the nest, but in one nest a parent removed two of three corpses (figure 2f).Honeyguide chicks attacked host chicks using their bills (figure 2d). Only once did a honeyguide bite a host parent (d), despite being frequently brooded by them. A host chick was once attacked while host parents attended the nest (e).Honeyguides occasionally bit unhatched host eggs, but failed to puncture their shells d. Only ot parent d, despitthe nest e.e), corresponding to potentially late host encounters in large and asynchronously hatching host clutches. Hooks do not appear to be shed (cf. [ca one week before fledging (which occurs at about 30 days).The bill hook is retained for at least 14 days e, corres4.Virulence in the greater honeyguide results from a succession of three specialised adaptations which together ensure the death of all the host young: (i) egg puncturing by the laying female , which wHost killing is carried out by blind hatchlings in total darkness, and the proximal cues used by honeyguides to start and stop biting remain unclear. Honeyguides sometimes lay alongside host young for tens of minutes before suddenly starting to attack, locating the host chick by repeatedly biting into mid-air while rapidly moving the head from side to side. Attacks often ceased when host chicks lay maimed and largely immobile, suggesting that movement may be a more important cue for attack than body heat. We only once witnessed a honeyguide biting a host parent, in spite of regularly being brooded by them; perhaps either the parents' food calls, or their feathered texture, serve as cues to prevent honeyguides from biting the hand that feeds them.The degree of virulence shown by brood parasites should be determined by a trade-off between the costs and benefits of chick-killing, just as pathogen virulence is shaped by trade-offs . What arSecond, the fitness costs versus benefits of honeyguide chicks′ sole occupancy of the nest remain to be quantified. When parasitizing host species that they are easily able to outcompete in sibling competition, such as little bee-eaters, parasites may do better to reduce virulence and co-opt host nestling begging to their own advantage . We migh


The removal of non-coding sequences, introns, from the mRNA precursors is an essential step in eukaryotic gene expression. U12-type introns are a minor subgroup of introns, distinct from the major or U2-type introns. U12-type introns are present in most eukaryotes but only account for less than 0.5% of all introns in any given genome. They are processed by a specific U12-dependent spliceosome, which is similar to, but distinct from, the major spliceosome. U12-type introns are spliced somewhat less efficiently than the major introns, and it is believed that this limits the expression of the genes containing such introns. Recent findings on the role of U12-dependent splicing in development and human disease have shown that it can also affect multiple cellular processes not directly related to the functions of the host genes of U12-type introns. At the same time, advances in understanding the regulation and phylogenetic distribution of the minor spliceosome are starting to shed light on how the U12-type introns and the minor spliceosome may have evolved. © 2012 John Wiley & Sons, Ltd. U12-type introns were initially described as a handful of unusual introns containing non-consensus AT–AC termini and a high degree of conservation at the 5′ splice site (5′ss).The significance of having two parallel intron types and two machineries for their removal was initially perplexing, especially since the U12-type introns are present at very low frequencies. The function and significance of the U12-dependent spliceosome are still far from being fully understood, but there are several lines of evidence indicating that U12-type splicing has essential functions. These are related to the conservation of U12-type introns in distantly related organisms and their presence in specific types of genes, as well as to the slow kinetics of their removal, which may regulate the expression of the genes containing them. Here, we review the current knowledge on U12-type introns and the function of the U12-dependent spliceosome. In particular, we discuss the recent findings on the regulation of the U12-dependent splicing, its function in development and disease, and the impact of these findings on understanding the evolution of the two spliceosomes.Although the U12-type introns were first identified on the basis of terminal AT–AC dinucleotides  it was soon realized that these termini are not exclusively present in U12-type introns.6Naturally occurring U12-type introns mostly belong to either the AT–AC or GT–AG subtypes. A few natural examples of U12-type introns with other combinations of terminal residues have been reported, and non-canonical termini have also been shown to support U12-dependent splicing experimentally, although often with reduced efficiency.17Caenorhabditis elegans and SaccharomycescerevisiaeArabidopsis thaliana, while in Drosophilamelanogaster only 19 U12-type introns have been described.A.thaliana.22Currently, U12-type introns have been identified in all major eukaryotic taxa, including plants, fungi, and animals, as well as a few deep-branching single-celled eukaryotes, but are nonetheless absent in many species, including such common model organisms as The genes containing U12-type introns are enriched in certain functional classes and pathways as described originally by Burge et al.65 and U2AF35, respectively, together forming the spliceosomal commitment (or E) complex. During the formation of the pre-spliceosome, or A complex, U2 snRNP replaces SF1 at the BPS. At later stages, the U4/U6.U5 tri-snRNP stably associates with the spliceosome (B complex). Rearrangements in RNA and protein interactions lead to the formation of the catalytically active spliceosome (B* complex) that catalyzes the first transesterification reaction, followed by the second transesterification in the C complex  and additional non-snRNP protein components. complex .The U12-dependent spliceosome contains four specific snRNPs, U11, U12, U4atac, and U6atac, each of which contains a specific snRNA component that is equivalent to but distinct from its U2-type counterpart, i.e., U1, U2, U4, and U6, respectively. U5 snRNP is shared between the two spliceosomes. Although the sequences of the snRNAs with equivalent function are quite divergent in the two spliceosomes, they share a common overall secondary structure . U4atac,The overall assembly pathways of the two spliceosomes are similar, and the main difference is the absence of a separate commitment complex in the minor spliceosome. Instead, the preformed U11/U12 di-snRNP binds the intron as a unit, and the 5′ss and BPS are recognized in a cooperative manner within the A complex, although U11/5′ss basepairing still precedes the formation of stable U12/BPS basepairing.4935, Urp,45Interaction of the U12 snRNA with the BPS is similar to the U2/BPS basepairing in the major spliceosome, resulting in the exclusion of the branch point (BP) adenosine from the U12/BPS helix.In the major spliceosome the U1 and U2 snRNPs recognize the 5′ss and BPS independently, and non-snRNP proteins are required for their association to each other.The formation of the catalytically active spliceosome is thought to follow a pathway similar to that of the major spliceosome. After initial recognition of the splice sites by the U11/U12 di-snRNP, the U4atac/U6atac.U5 tri-snRNP associates with pre-spliceosome to form complex BAlthough the formation of the catalytic core appears to be somewhat more flexible in the U12-dependent spliceosome,in vitro splicing experiments documented a splicing rate for U12-type introns that was three- to fivefold slower than that of U2-type introns.in vivo experiments, where approximately twofold higher levels of unspliced U12-type introns were detected in the steady-state transcript pools isolated from insect and mammalian cells.72Both intron types are spliced co-transcriptionally within the nucleus Ref  and mighin situ hybridization studies, mostly with zebrafish, as well as cell fractionation and reverse transcription polymerase chain reaction (RT-PCR) analyses. The publication contradicted many of the earlier studies on minor spliceosome component localization and function  can be extremely long, such that it can take an hour for an entire intron to be transcribed. Therefore, initial splice site definition typically takes place over the considerably shorter exons, with 3′ss-recognizing factors in the upstream intron interacting with the 5′ss-recognizing factors in the downstream intron  contain overlapping U12 and U2-type introns or ‘twintrons’.dUrp mRNA isoform is apparently destroyed by NMD,prospero alternative splicing results in distinct developmentally regulated protein isoforms with amino-acid differences in the homeodomain region.86Given that AS is extremely prevalent in humans, and the extensive interactions of the minor spliceosomes with other splicing factors, it is somewhat surprising that AS events involving U12-type introns appear to be almost entirely absent. This may be because of the more rigid sequence and distance constraints on the U12-type 5′ss, BPS, and 3′ss. AS events involving the use of an alternative 3′ss have been observed, but it is not entirely clear whether they are the result of true regulated AS or of splicing errors, especially since the selection of the U12-type 3′ss has been shown to be prone to errors.D. melanogaster,A. thaliana also led to abnormal growth of the plant, but only after the bolting stage.prospero transcripts in D. melanogaster. However, it is likely that another, even more significant factor affecting developmental processes is the slower kinetics of minor splicing, which can limit not only the expression of the genes containing themD. melanogaster, likely underlying the developmental arrest.A. thaliana was also associated with downregulation of a specific set of genes related to gibberellic acid metabolism.Knockdown of protein components specific to the minor spliceosome leads to reduced proliferation of cultured cells,LKB1 and SEDL genes cause Peutz-Jeghers syndrome and spondyloepiphyseal dysplasia tarda, respectively,From a medical point of view, it would be interesting to know to what extent the activity of the minor spliceosome contributes to regulating the expression of genes in different human tissues. This question remains mostly unanswered. Some protein components specific to the minor spliceosome (31K and 65K) have been observed to be differentially expressed in different tissues,D. melanogaster carrying mutations in U6atac snRNA.D. melanogaster.A more complex class of diseases are those arising from defects in minor spliceosome components of which the TALS/MOPD1  is a recent example. This severe developmental disorder is caused by recessive mutations in the U4atac gene Refs . Unlike It is clear that the U12-dependent spliceosome is crucial for the viability and development of many multicellular organisms. However, intriguing questions about the minor spliceosome remain: Where did it originate? Why is it not present in all eukaryotes, and do its function and importance vary in different evolutionary lineages? We do not currently have clear answers to these questions, but recent advances in large-scale sequencing and bioinformatics are paving the way toward deeper understanding of the significance of the minor spliceosome.D. melanogaster has only 19 U12-type introns, almost all of them have orthologous U12-type introns in human.A. thaliana.67The two spliceosomes have been proposed to originate from group II self-splicing introns, but it is not clear whether they arose simultaneously or sequentially, and whether this took place in one ancestral eukaryote or separate lineages that later fused.Diptera, which typically only have 15–20 U12-type introns in total in their genomes, several fold less than in most other insects.21Despite the high overall conservation of U12-type introns, they are entirely absent from many eukaryotes whose relatives nonetheless have them, suggesting repeated loss of U12-dependent splicing during evolution. The loss seems to have been most prevalent among diverse eukaryotic microbes, but some animals, including some but not all nematodes, have also lost their U12-type introns Refs . The losdUrp twintron).In most cases, the mechanism of U12-type intron loss involves a simple deletion of the intronic sequence, apparently through the homologous recombination of reverse transcribed mRNAs.Diptera, where both the overall numbers and interspecies conservation of U12-type introns are low.Drosophila dUrpMany splicing factors are regulated through a feedback mechanism, in which they bind to regulatory elements within their own pre-mRNA, resulting in alternatively spliced, unstable mRNA isoforms. Interestingly, all canonical SR and hnRNP proteins, as well as a number of core spliceosomal factors, are regulated in this manner, and in many cases the regulatory elements are also evolutionarily conserved.Historically, insight into the function and significance of the U12-dependent spliceosome has come primarily from the biochemical characterization of splicing factors and spliceosome assembly, and has recently been complemented by the increasing number of bioinformatic analyses. However, our understanding of the significance of minor splicing is far from complete in both regards. The function of many of the U12-specific components is still unknown, and more components might still remain undetected. Importantly, although there are many independent lines of evidence indicating that U12-type introns are important for the regulation of gene expression, definitive large-scale evidence on the connection between the activity of the minor spliceosome and gene regulation is still lacking. Are there more regulatory elements that respond to U12-type factors? In what other ways is the minor spliceosome regulated, and how does this correlate with the expression of U12-type intron-containing genes in different tissues? Could there be U12-type introns with more degenerate splice sites, and could they be involved in more extensive alternative splicing? An exciting way of studying these questions is now offered by the rapidly developing high-throughput sequencing and cross-linking methods, and will hopefully deepen our understanding on the significance of minor splicing in the coming years.


The as-produced graphene was characterized by X-ray photoelectron spectroscopy, ultraviolet-visible spectroscopy, scanning electron microscopy, and high-resolution transmission electron microscopy (HR-TEM). In particular, the HR-TEM demonstrated the layered crystallites of graphene with fringe spacing of about 0.32 nm in individual sheets and the ultrafine facetted structure of about 20 to 50 nm of Au particles in graphene composite. Scanning helium ion microscopy (HIM) technique was employed to demonstrate direct write deposition on graphene by lettering with gaps down to 7 nm within the chamber of the microscope. Bare graphene and graphene-gold nanocomposites were further characterized in terms of their composition and optical and electrical properties.Graphene nanosheets were prepared using a modified Hummer's method, and Au-graphene nanocomposites were fabricated by  Graphene, structurally known as a monatomic layer of allotropic-carbon atoms in a hexagonal honeycomb two-dimensional lattice system, has always been a potential candidate for various applications due to its remarkable structural, physical, and electronic properties -9. The z4- ions in graphene suspensions to fabricate gold (Au) nanoparticles-graphene hybrid assemblies. In yet another report, Goncalves et al. [). These graphene/Au nanocomposites act as potential substrates for surface-enhanced Raman scattering. Min et al. [The composites of metal nanoparticles on graphene sheets also provide a new way to develop catalytic, magnetic, and optoelectronic materials. Moreover, adhesion of such metal nanoparticles to the graphene prevents their aggregation in dry state . Recentls et al.  demonstrn et al.  have alsBut in comparison to these reports, the work presented here highlights a simpler route to obtain stable Au nanoparticles-graphene nanocomposites. It also demonstrates the capability of direct labeling on nanocomposite by use of scanning helium ion microscopy (HIM).in situ ion lithography, nano-patterning, device prototyping, fabrication of quantum dots, beam-induced chemistry, and milling at nanoscale [The demonstration of imaging by helium (He) ions is relatively a new technique to characterize the surfaces at sub-nanoscale with extraordinary additional advantages of anoscale ,16. Suchanoscale -22 that 4) was purchased from Aldrich . Sodium borohydride (NaBH4) was acquired from Merck . Inorganic transparent electrodes of SnO2:F-coated glass  were cleaned in a soap solution, 30% HCl solution, double-distilled water, acetone, and trichloroethylene (in that order) prior to use. Deionized water (resistivity ≈ 18.2 MΩ cm) obtained through Milli-Q system, nitric acid (HNO3) (Merck), sulfuric acid (H2SO4) (Merck), and toluene were used as solvents.Hydrogen tetrachloroaurate (HAuCl2SO4:HNO3 in a 3:1 volume ratio (12 ml H2SO4 and 4 ml HNO3) and 2 g graphite powder was made in a flask and refluxed at 40°C for 16 h. The resulting solution was washed with deionized water till the pH was reduced to 5 or 6. As a result, a black colored solution of acid-functionalized graphene was obtained.For acid functionalization of graphene, a solution with Hin situ in graphene suspension by the reduction of gold(III) complex by NaBH4. A concentrated aqueous solution of 0.4 M NaBH4 was first mixed with acid-functionalized graphene suspension in toluene. With continuous stirring, 30 mM of HAuCl4 was then introduced into this suspension. After continuously stirred for 1 h, the resulting Au-graphene composites were collected by centrifugation and washed with water for three times.To fabricate Au-graphene nanostructures, Au nanoparticles were synthesized I-V measurements of films were carried out on Keithley 238 high-current electrometer characterization system. Absorbance (A) spectra were recorded in the 200- to 800-nm wavelength range in a Perkin Elmer Lambda 25 spectrophotometer . X-ray photoelectron spectroscopy (XPS) spectra were recorded for the as-synthesized graphene samples using a Perkin Elmer 1257 model PHI, Maple Grove, Minnesota, 55311 U.S.A operating at a base pressure of 3.8 × 10-8 Torr at 300 K with a non-monochromatized AlKα line at 1,486.6 eV, an analyzer pass energy of 60 eV kept for core level spectra and a hemispherical sector analyzer capable of 25-meV resolution. The overall instrumental resolution was about 0.3 eV. The core level spectra were deconvoluted using a non-linear iterative least squares Gaussian fitting procedure. For all fitting doublets, the FWHMs were fixed accordingly.Fourier transform infrared spectra for the films were recorded in reflection mode with a Perkin Elmer GX2000 OPTICA spectrophotometer at 28°C, RH ≈ 50% to 53%. 2 F 30 STWIN FEI, Achtseweg Noord 5 5651 GG Eindhoven, Netherlands (operated at the electron accelerating voltage of 300 kV). HR-TEM specimens were prepared by dispersing the graphene films on copper grid of 3.05 mm in diameter having a 200-mesh pore size. Further, the surface topography of graphene and graphene-Au composite films was analyzed by HIM . The He ion capability of the microscope was used to perform the experiments of nanoscale patterning on the surfaces of graphene.Surface morphology of the graphene sheets was studied employing a variable pressure scanning electron microscopy (SEM), model: Zeiss EVO MA10 Carl Zeiss SMT AG, Germany. Nanostructural imaging at high magnifications was carried out using HR-TEM model: FEI-Tecnai Gn→π* transitions of C=O bonds [-1 to 545 nm at 0.2 and 0.4 g l-1. The quenching in the peak intensity was also observed which is clearly visible in the inset of Figure The successful synthesis of graphene and Au nanoparticles decorated graphene was confirmed by ultraviolet-visible (UV-Vis) spectroscopy Figure . The UV-=O bonds . When Au5/2) and Au(4f7/2) [+ state. Deconvolution was performed on C(1s), O(1s) and Au(4f) XPS core spectra are shown in Figure 0 (81.8 and 85.3 eV) exhibit the metallic nature of Au, while Au+ state (83.7 and 87.4 eV) probably due to the interaction with the negatively charged graphene around Au induces a positive charge. The contribution of various spices of core level spectra is listed in Table The formation of stabilized Au-graphene nanocomposite was further confirmed by the XPS spectra as shown in Figure u4f7/2)  due to tu(4f7/2)  for Ag/gu(4f7/2) ,27. Inteu(4f7/2) , and the  due to rmed on Cs, O(1s) c-axis growth of carbon lattice also justify a distinct orientation relationship and therefore a crystallographic compatibility between the carbon as matrix and the Au as second phase.Crumpled, folded, layers of bare graphene can be seen in the SEM image shown in Figure in situ examination of the microstructure and direct write of arbitrary patterns on the given nanomaterial. HIM is showing the capability to create smaller structures than possible with other technique [in situ nano-scaled structural buildings elucidates a new field which is so far relatively unexplored in fabrication and process control of fundamentally important nano-objects like graphene. In the present work, the text  was created by deposition of carbon. This pattern, in the form of a bitmap, was opened up in Orion software Carl Zeiss NTS LLC, 1 Corporation Way, Peabody MA 01960, U.S.A. Subsequently, the user defined the size of the overall pattern, and the pixel size was scaled accordingly. The ion dose per pixel was also variable, being set in proportion to the gray level in the bitmap (up to 256 levels). Lettering with gaps down to 7 nm was observed. In this process, the direction of scanning was also user selectable. In another set of experiments, HIM was used to study the distribution of Au in the matrix of graphene (Figure Nano-patterning by use of a high-resolution microscope is a fast developing method which facilitates echnique -17. In te Figure . We haveI-V characteristics of Au-graphene and functionalized graphene were recorded in the following configuration: SnO2:F/Au-graphene/aluminum as shown in Figure 2:F-coated glass substrates. The AuNPs decoration was having a beneficial effect on the electronic conductivity of graphene. Both the films show ohmic contact with the substrate in 0 to +1 V potential region as can be seen from the linear variation of current with applied bias. For the Au-graphene composite, the conductivity was determined to be 0.49 S cm-1 which is much higher than blank functionalized graphene film where the value was estimated to be 0.07 S cm-1 which validates the role of interaction with AuNPs in the enhancement of conductivity. In the literature, the value of pristine graphene has been reported to be 0.2 S cm-1 [The I-V characteristics between bare graphene and its Au incorporated nanocomposites has been noticed. An important in situ direct write deposition on nanosheets of graphene has been demonstrated by employing He ions inside the chamber of the microscope.A simple modified Hummer's method was used to fabricate graphene and graphene-Au nanocomposites. A significant change in The authors declare that they have no competing interests.SB carried out the graphene preparation and interpretation of results. MD did the lectrical measurements. AGJ carried out XPS measurements and interpretations. APS assisted in synthesis of material. AKS initiated the idea of working on present topic and subsequently compiled the data.


Over time, a large number of question and answer (Q&A) pairs with high quality devoted by human intelligence have been accumulated as a comprehensive knowledge base. Unlike the search engines, which return long lists of results, searching in the CQA services can obtain the correct answers to the question queries by automatically finding similar questions that have already been answered by other users. Hence, it greatly improves the efficiency of the online information retrieval. However, given a question query, finding the similar and well-answered questions is a non-trivial task. The main challenge is the word mismatch between question query (query) and candidate question for retrieval (question). To investigate this problem, in this study, we capture the word semantic similarity between query and question by introducing the topic modeling approach. We then propose an unsupervised machine-learning approach to finding similar questions on CQA Q&A archives. The experimental results show that our proposed approach significantly outperforms the state-of-the-art methods.With the blooming of Web 2.0, Community Question Answering (CQA) services such as Yahoo! Answers ( With the proliferation and growth of Web 2.0, CQA services have become the integral part of information and knowledge acquisition. It provides a main platform for information seekers to post their specific questions in a wide range of topics and obtain answers, comments and other interactions, such as voting and rating, provided by other users. CQA services provide a real space for online communications. Either the topics or the questions and answers are posted by human, which we called user generated content (UGC). Hence the quality of Q&A pairs in CQA repositories is high. Over time, a large amount of Q&A pairs with high quality devoted by human intelligence have been accumulated as comprehensive knowledge base.http://answers.yahoo.com/), which is launched on December 13, 2005. It greatly facilitates the users to acquire knowledge and information One of the most popular CQA services is Yahoo! Answers  methods query and the question in topic space and automatically gather the similar questions from the Q&A repository.To address the above problems, we propose a topic-based semantic similarity computing approach to finding similar questions in CQA archives. We first utilize the topic model The remainder of the paper is organized as follows. Section Related Work introduces the related work. Section Background on Latent Dirichlet Allocation gives a briefly introduction about the Latent Dirichlet Allocation (LDA). Section Topic Clustering Approach presents the topic-based clustering approach to finding similar questions. Section Experiments gives the experimental results and error analysis, followed by the conclusion and future work in Section http://trec.nist.gov/) QA track, the QA systems need to capture the similarity between the questions and the candidate documents, and then return the relevant documents. In the application of the interactive QA, users input their questions in natural languages; the system then searches the candidate documents online and returns the answer list by computing the similarity between the users' questions and candidate answers.Text similarity computing is widely applied in question answering (QA). In the TREC  overlap method The syntactic-based methods focus on the similarity of syntactic structure. They consider the similarities of both the lexical and the syntactic structure. For example, they used tree kernel methods to calculate the common sub-trees between two questions query and questions, it can only capture the lexical level similarity.Furthermore, questions by reformulating them to a topic and focus structure. They then utilized the MDL based tree cut model to select relevant words. At last, they used the similar topics to finding similar questions. However, as the CQA questions are more complex, it is hard to identify the question topics.The existing topic-based similar question finding approach To investigate the above problem, in this study, we employ the Latent Dirichlet Allocation (LDA) model to identifying the question topics. We then propose a topic clustering based approach to finding similar questions, which can effectively measure the topic level semantic similarity between two questions.Latent Dirichlet Allocation (LDA) is first proposed by While using LDA on text data, a latent topic set is contained on the corpus. It means that each document can be represented as a mixture topic set. The processing of the whole text corpus is transferred to the processing of the topic sets. Hence, it reduces the dimension of corpus by text space mapping.Given corpus Choose Choose For each of Choose a topic Choose a word The following shows the joint distribution of LDA is a generative probability model. In text processing, the LDA model can randomly generate topic sequences. Hence, the representation of each document can be transferred to a topic sequence. Here, in Based on the above descriptions, we then make the following assumptions:Assumption A.Each question Assumption B.There exist a global function We use Next, we will introduce the features used for mining the similar questions from Yahoo! Answers Q&A repository. Besides the question topic related information, we also consider the question content based factors to finding similar questions. It means that in our proposed approach, we also capture the string matching features. Moreover, we explore the linguistic analysis technique to mining the semantic similarity between the questions.In this section, we plan to enhance the topic modeling based approach to finding similar questions by employing three filtering factors. They are question Levenshtein distance factor, part-of-speech (POS) sequence factor and word overlapping factor. We then utilize a unique function to combine the three factors for question similarity computing as follows:The minimum number of edit operations necessary to transform one string into another.Insert, Delete and Replace.The three edit operations are Where, Finally, we empirically set the According to the above descriptions, we utilize the LDA model to gather the similar questions in topic space. We then propose a similarity filtering approach to enhancing the result of similar question exploring. In this section, we will give an overall conclusion about our proposed topic clustering based approach to finding similar questions. We summarize the following steps:Preprocessing: Removing stop words and stemming.LDA modeling: Transferring the Q&A repository into the corresponding topic vectors.Topic guided clustering: Based on the three factors in Similar question filtering: Selecting and reranking the similar questions for each of the clusters by using http://developer.yahoo.com/answers/). It covers a wide range of topics, including buying and selling, internet, etc. For each question, we extracted the question title, question content and chosen answers as the experimental data from the returned content by the API. From this data collection, we randomly select 10,000 questions as the queries and 200 as development set to tune the involved parameters. As the questions which are extremely short may represent ambiguous intent, we filtered out the questions which contain less than three terms through the random selection process. The experimental data is available at http://pan.baidu.com/share/link?shareid=343582&uk=2903372971.We collected a total number of 1,123,034 questions from Yahoo! Answers using the Yahoo! Answers API : BoW based similar question clustering approach which only use the term frequency (TF) and inverse document frequency (IDF) as features (baseline 1).BoW+Cluster+Filter (BCF): BoW based similar question clustering with our proposed similar filtering approach (baseline 2).updatedBoW+Cluster (upBC): Updated BoW based similar question clustering approach which use our proposed featuresSee  (baselinSyntacticTreeMatching (STM): The state-of-the-art approach to finding similar questions in CQA repository which is proposed by LDA+Cluster (LDAC): Our proposed LDA model based similar question clustering approach with our proposed features.LDA+Cluster+Filter(LDACF): Our proposed LDA based similar question clustering approach which is also integrating the proposed similar filtering approach.p@1) to evaluate the performance of the comparing systems. p@1). Here, all the scores are the real values in the two evaluating measurements. Furthermore, we also capture the percentage of AP improvements. Here, the t-test works for testing the statistical significance on finding similar question result which contains a large number of questions. As the t-test works for the non-normal data only if the sample size is large, the t-test used in our experimental data set is rational.For baseline 4, we run the original syntactic tree matching system to finding similar questions in our data set. We use average precision (AP) and precision at position one  for similar question clustering. This is because that our proposed features not only take the lexical information into consideration, but also consider the topic distribution and weight for the questions. Hence, the similarity modeled by our proposed features is enhanced by combining the topic space similarity between the questions.Second, to see the results of the LDACF approach outperforms the BoW based methods. It demonstrates that, the topic modeling approach can better represent the Q&A data than the BoW methods and further better captures the similarity between the questions. This is because that our proposed approach captures the similarity of the questions not only through the lexical and linguistic information, but also mapping the Q&A text into the topic space. We thus measure the semantic similarity of the questions in the topic level.Third, we can observe that our proposed STM and LDACF, we can see that our proposed LDACF outperforms the STM, which is the state-of-the-art approach in finding similar questions in the CQA archives. This is because the STM approach only considers the syntactic tree structures. It employs the tree kernel function to measure the question similarity. However, it neither introduces the fuzzy matching scheme, nor considers the semantic similarity in topic level. In our proposed approach, we capture the questions' semantic similarity in the topic space. And we also take the advantages of the lexical and linguistic analysis techniques.Fourth, to comparing with the Furthermore, to analyzing the experimental results, we found that there exist the clusters that contain only one question. We remove these clusters in the similar question clustering result and evaluate the proposed approach again. LDACF is less than BC. It demonstrates that the topic guided clustering approach is better than the BoW based clustering approach in error cases handling. It is because those topics can be seen as a higher level lexical semantics. Hence, the clustering results of LDACF is more accuracy and robust than the BC.From BC and LDACF in the refined data set. For evaluation, we also employ the average precision (AP) and precision at position one (p@1). We also compare the performance between the http://homepages.inf.ed.ac.uk/gcong/qa/annotation_result.txt. The second data set is collected from Twitter (https://twitter.com/) by using the Twitter API (https://dev.twitter.com/docs/using-search). We collected a total number of 13,683,354 questions from Twitter as our experimental data set. . It covers a wide range of topics, including famous people, internet, makeup etc. From the data set, we randomly select 200 questions as queries and 100 questions as development set to tune the involved parameters. For preprocessing, we filtered out the non-English characters and the urls.To check the effectiveness of our proposed approach, we also test the performance on another two data sets. The first data set is used to evaluate the performance of question retrieval in To obtain the ground truth, we pooled top 20 relevance questions by utilizing various approaches, including vector space model, okapi BM25 model, language model, translation model, translation based language model For the evaluation, we utilize the average precision (AP) and precision at position one (p@1) on the above two data sets. From In this section, we plan to verify the influence of the topic number on the final results of finding similar questions. We then run our proposed system in the various topic numbers. From In the paper, we proposed an unsupervised machine learning approach to finding similar questions in Yahoo! Answers Q&A repository. Although our experiment results outperform the baselines significantly, the final results also need to be further improved. Hence, we analyze the experiment results and we then conclude the following errors which influence the performance of finding similar questions.Although we employed LDA model to capture the topic information, there are a large amount of new entities in our Q&A repository. As we have no entity recognition module, our approach fails to deal with the specific name entities. For example, the similar questions “Where can I find a cheap canon 60D camera?” and “Where is the cheapest online source to buy canon 500d?”, our approach cannot distinguish the “canon 60D” and “canon 500d” as both of the questions' topic is “canon”.For the diversity representation of the user questions, there exist the questions with ambiguous meanings. However, the question content and the chosen answers can be used to clarify the question meaning. For example, “Blackberry?” and “How do i fix it?” are two questions which share no common words. Through the analysis of the question content, we found that both of the two questions are asking for the “camera light” problem. Hence, how to mine the question similarity between two questions that share none or fewer common words is a non-trivial task.In order to illustrate the topic distribution in our experiment data set, we list several topic words with high probability as follows. From In this section, we will discuss the limitations of this study. We would like to thank the anonymous reviewer for the comment of which we should test our approach on a diverse range of data sets. To check the effectiveness of our proposed approach, we will test our proposed approach on more available data sets in future work. Meanwhile, we would like to thank the anonymous reviewer for the comment on feature fusing. In future work, we will consider to automatically fusing the features in finding similar questions. Inspired by We also noticed that the precision of our proposed approach is still low. In our future work, we plan to employ the name entity recognition scheme so that we can accurately identify topic related terms. We will utilize more semantic resources, such as phrase based paraphrasing and translation based synonym extracting, to capture the lexical semantic similarity. Specifically, we will try to reformulate the original query question into the semantic similar questions using the approach which is proposed by Zhang et al. In this paper, we proposed a topic guided clustering approach to finding similar questions in CQA archives. We utilized the LDA model to map the Q&A data set into topic space and took the advantages of the topic modeling as guided information to cluster the questions which share the same or similar topics. We verified that the LDA guided clustering approach significantly outperformed the state-of-the-art approach of finding similar questions in CQA archives as well as other baselines.Our proposed topic clustering approach can adapt to other applications. In future work, we plan to use our topic clustering for finding correlated salient object segments


Graphene, a 2D crystal bonded by π and σ orbitals, possesses excellent electronic properties that are promising for next-generation optoelectronic device applications. For these a precise understanding of quasiparticle behaviour near the Dirac point (DP) is indispensable because the vanishing density of states (DOS) near the DP enhances many-body effects, such as excitonic effects and the Anderson orthogonality catastrophe (AOC) which occur through the interactions of many conduction electrons with holes. These effects renormalize band dispersion and DOS, and therefore affect device performance. For this reason, we have studied the impact of the excitonic effects and the AOC on graphene device performance by using X-ray absorption spectromicroscopy on an actual graphene transistor in operation. Our work shows that the excitonic effect and the AOC are tunable by gate bias or metal contacts, both of which alter the Fermi energy, and are orbital-specific. Graphene, a honeycomb network consisting of carbon atoms bonded by π- and σ-orbitals, is a model two-dimensional 2D) compound in physics1D compoun45123FMany-body effects, such as electron-electron interactions, are prominent in graphene owing to a limited density of states (DOS) near the DPAn orbital-specific investigation is vital for the study of the many-body effects in graphene because they critically depend on the molecular orbitals involved. X-ray absorption spectroscopy (XAS) is a suitable tool for this as it can provide an element- and orbital-specific probe of electronic structure affected by dynamical many-body effects during the absorption processes, including excitonic effects2022242526F.Here we present the results of microscopic XAS (μ-XAS) studies on an actual graphene transistor in operation. We demonstrate the orbital-specificity and tunability of excitonic effects and the onset of the AOC using gate bias or metal contacts, both of which alter Eg) was applied using a SiO2 thin film as a capacitor and a Si substrate as a backgate. Under our experimental conditions, a negative gate bias means positive charging of graphene and therefore hole doping. The μ-XAS spectra were acquired by using a photoemission electron microscope (PEEM) whose lateral resolution was 22 nm in the Auger electron yield modeIn the graphene device used in this work, bilayer graphene see  and Ni tG* (~307 eV)G* peak shows a change in only peak intensity. The π* peak position is redshifted by increasing the gate bias, as shown in 2020212221F22iE is the self-energy of the core hole, which lowers the peak positionFF. The redshift induced by increasing the gate bias is in fact corroborated by a recent theoretical calculation of X-ray absorption spectra of graphene into which positive charges are doped20212223242526G* ~7 eVG* peA(ω) follows a power law23α is governed by the AOC as well as the excitonic effect222323Aα) is thus given in terms of the DOS at EF asFN and V0 are the DOS at the Fermi level and the screened Coulomb interaction, respectivelyF is situated near the DP because of the vanishing of FNF away from the DP owing to the increase of FNF moves away from the DP thus accounts for the reduction of the π* peak intensity.The π* peak intensity is reduced as the gate bias is increased . This ch22G* peak by the gate bias application is examined. As shown in G* peak position is unchanged when varying the gate bias. According to a previous theoretical work20G* peak are to delocalized conduction band statesG* peak position is related with the absence of the excitonic effect for the σG* orbital. It is therefore the ground-state unoccupied DOS (UDOS) without the influence of the core hole which is reflected in the line shape of the σG* peak, in contrast to the π* peak whose shape is determined by the excited-state UDOS with the influence of the core holeTo scrutinize the orbital-specificity of the many-body effects, the change of the σG* peak intensity. The intensity is enhanced by increasing the gate bias, in contrast to the depression of the π* peak intensity , in the order of 10 V/μm. On the other hand, our previous scanning photoelectron microscopy on graphene/SiO2/Si in contact with Ni electrodes shows that the electric field near the contact is in the order of 0.1 V/μmG* intensity. We have thus shown here that the contact of graphene with a metal produces an orbital-specific modulation of the many-body effects in a way similar to the application of a gate bias.The orbital-specificity of the many-body effects due to the metal contact can be seen by considering the σF, as explained above. The variation of the π* peak position at the interface therefore indicates that the value of EF at the interface varies with gate bias. This demonstrates that Fermi-level pinning, which has been controversialAn important observation is that the π* peak position at the graphene-Ni interface is not pinned for different gate biases see . This beF]−1/2FN is increased by moving EF away from the DP.Also worth noting is how the redshift of the π* peak near the interface, that is, in the charge transfer region, varies with gate bias. As can be seen in F through application of a gate bias or contact with a metal, by using μ-XAS on an actual graphene transistor in operation. Since the many-body effects are involved dynamically in photoabsorption processes, our work is important for an understanding device characteristics, such as output characteristics of lasers and the energy distribution of emitted photons which is related to the  DOS and band structure.In conclusion, we have demonstrated that graphene possesses orbital-specific many-body effects, which vary with E2 thin film (90 nm thick) used as a capacitor, a p-Si(100) substrate used as the backgate and Ni thin film electrodes in contact with the graphene. The bilayer graphene was exfoliated from Kish graphite and transferred onto the SiO2 thin film−5 Pa and were subjected to a lift-off process in warm acetone. The gate bias was applied between the graphene and the Si backgate. The graphene in this work was positively charged so it could be doped with holes by applying a negative gate bias.In this work, the device consisted of bilayer graphene, a SiOThe μ-XAS spectra were measured by the Auger electron yield method using a PEEM  installed at BL17SU at SPring-8H.F. assisted with sample preparation, performed μ-XAS experiments, participated in designing the study, and participated in writing of the manuscript; M.K. performed μ-XAS experiments, participated in designing the study, and participated in writing of the manuscript; K.N. undertook sample preparation, participated in designing the study, and participated in writing of the manuscript; R.S. performed μ-XAS experiments; T.O. performed μ-XAS experiments; T.I. participated in designing the study, and participated in writing of the manuscript; A.T. oversaw sample preparation, participated in writing the manuscript; M.S. assisted with writing the manuscript; T.K. participated in writing the manuscript and discussed about the content of the manuscript.Supplementary information


RH) instantaneously, by applying two ultrasonic transducers and an array of four temperature sensors. Measurement ranges are: gas velocity of 0–12 m/s with an error of ±0.13 m/s, temperature 0–100 °C with an error of ±0.07 °C and relative humidity 0–100% with accuracy better than 2 % RH above 50 °C. Main advantage over conventional humidity sensors is the high sensitivity at high RH at temperatures exceeding 50 °C, with accuracy increasing with increasing temperature. The sensors are non-intrusive and resist highly humid environments.An acoustic relative humidity sensor for air-steam mixtures in duct flow is designed and tested. Theory, construction, calibration, considerations on dynamic response and results are presented. The measurement device is capable of measuring line averaged values of gas velocity, temperature and relative humidity ( Some tyCO2 concentration and to a lesser extent absolute pressure. By simultaneous measurement of speed of sound and air temperature, relative humidity can be calculated for given pressure and CO2 concentration.One option to measure relative humidity is by means of acoustic techniques. From kinetic gas theory it follows that the speed of sound in air depends on the composition and condition of the air . The mosAcoustic sensors are non-intrusive, in contrast to other conventional humidity sensors like wet and dry bulb sensors or capacitive sensors. This ensures no pressure loss for in-line applications, high life expectancy and insensitivity to contamination. Another advantage over conventional sensors is the high temperature range. Most important advantage is the high sensitivity and the increase of sensitivity with increasing temperature.et al. [et al. [i.e., with a measuring volume in the order of one cubic mm. The acoustic technique of the present paper, on the other hand, yields a chordal beam average of humidity in the desired portion of the duct. Another type of acoustic sensor was developed by Zipser et al. [et al. uses an ultrasonic sensor for temperature measurement with a correction for humidity [Recently, many studies for humidity sensors have been published, particularly acoustic sensors. However, most of these recent developments in acoustic humidity sensors are based on Surface Acoustic Wave (SAW) sensors, which works with another principle, see the work of Wu et al. , for exa [et al. . The abor et al. , which hhumidity .In the present study, the design and tests of a high accuracy in-line acoustic relative humidity sensor for flowing air-steam mixtures in a duct flow are presented. This includes theory, construction, calibration, considerations on dynamic response and results.2.B, is given, can be calculated from [T is in K:The speed of sound in a gas for which the second virial coefficient, ted from ,7. In thγ and B must be known to calculate the speed of sound in the gas mixture. By measuring the speed of sound at constant temperature, T, and pressure, p, determined from measurements of air, the composition of air at constant T and p uniquely depends on the speed of sound.For each constituent of a gas mixture, The constituents of standard dry air according to ISO norm 2533 are listed in N2, O2, Ar, CO2 and Ne and the amount of water vapor have a significant effect on the molar mass of air. If the composition is assumed to be constant except for the amount of water vapor, the mole fraction of water can be determined from the speed of sound.Only the concentrations of B of a mixture of gases to calculate humidity, RH, is examined in [The use of the second virial coefficient mined in . Much moia} are determined by calibration in reference air of known temperature, T, known humidity, RH and known speed of sound at the measurement frequency, c. From 2, the mole fraction of water vapor, wx, is determined. Relative humidity is then calculated with the aid of:The coefficients {The saturated vapor pressure of water is calculated from, for example, an Antoine relation :(4)psv=A = 8.07131, B = 1730.63, C = 233.426, valid for 1–100 ° C, T in °C and svp in Pa.Coefficients are c increases with increasing temperature and with increasing RH.The relation between the speed of sound, temperature and relative humidity according to mt the transit time averaged in ts. L is the total length of the acoustic trajectory in m between transducers Tr1 and Tr2, see t1 is the transit time in downstream direction and t2 the transit time in upstream direction in s.The speed of sound is determined by measuring the ultrasonic transit time of the acoustic signal on a trajectory. The transit time is influenced by the air-steam flow velocity, which is taken into account by averaging the speed of sound in upstream and downstream direction:sL. sL is the part of the acoustic trajectory where the ultrasonic waves have a component in the direction of the gas flow  and four SMT temperature sensors are mounted in the duct as shown schematically in Hz. Gas flow in the duct is measured at constant temperature and flow. In the heat exchanger test rig, fast response times are of minor importance.An overview of the device without insulation is shown in 4.Temperature sensors are calibrated from 0 to 100 °C with an insulated Julabo MP open bath circulator and a reference thermometer. Accuracy of each SMT sensor is 0.14 °C. To take possible temperature gradients into account, temperatures are averaged over the height and weighed by the corresponding mass velocity . Accuracc is known and transit times t1 and t2 should be equal. Lt is found to be 502.8 ± 0.1 smm. L is determined by the design of the measurement device and is found to be 260.0 ± 0.1 mm.The length of the acoustic trajectory is calibrated by measuring the transit times at no flow conditions for given temperature and relative humidity in an insulated reference box. m/s to 0.13 m/s accurate in a wind tunnel with a reference flow meter [The average gas flow velocity is calibrated over a range of 0 to 12 ow meter .RH and a psychrometer better than 3 %RH accurate. Calibration is performed at ambient pressure. A field of 40 measurements is assessed: temperatures from 20.0 to 90.0 °C in steps of 10.0 °C at relative humidities of 10 to 90 % in steps of 20 %. This results in coefficients of Relative humidity measurements are calibrated in a Weiss SB22-300 climate chamber with a Michell S4000 cooled mirror optical dewpoint hygrometer, accurate to ±1 %5.s. With temperature fluctuations in time, the relative humidity measurement will show a delay. Various methods to minimize this delay are considered:Use smaller temperature sensors, with smaller time constants.Damp the thermal fluctuations by installation of a large thermal mass at the duct inlet. This overcomes erroneous measurement of relative humidity due to temperature fluctuations, but cancels out the advantage of the fast response of the acoustics.Install temperature sensors with different time constants and use the derivative of the fastest temperature sensor to correct the slowest temperature sensor.One of the major advantages of acoustic sensors in general is the fast response time, because the response time mainly depends on the speed of sound and typical transit times of the acoustic trajectory. However, if the goal of the acoustic sensor is to measure relative humidity in a duct, a separate temperature measurement in the duct is needed. Response times are dependent on the slowest measurement, in this case the temperature sensors which have time constants of about 5 These strategies are now under consideration to further improve the acoustic relative humidity sensor.6.6.1.The measurement device is preliminarily tested at the test section inlet of a wind tunnel for condensing heat exchangers at the Department of Mechanical Engineering, Eindhoven University of Technology, see . A flow Heat flow rate from the gas is:The energy balance is given by:The acoustic flow measurements are compared with the volumetric flow rate of dry air measured by a gasrotormeter positioned upstream the acoustic measurement device, see RH at 80 °C and 2–6 m/s for the points shown in Energy balance measurement accuracy improved by replacing capacitive humidity sensors at the gas flow inlet by the acoustic measurement device, see 7.m/s, 0–100 °C and 0–100% relative humidity at ambient pressure. Main advantage over conventional humidity sensors is the high sensitivity at high RH at temperatures exceeding 50 °C, with accuracy increasing with increasing temperature. The sensors are non-intrusive and resist highly humid environments. Accuracy for line averaged flow velocity is 0.13 m/s, average temperature 0.07 °C after calibration. With this temperature measurement accuracy, intrinsic accuracy of relative humidity is better than 2 %RH above 50 °C and within 1 %RH from 70 to 100 °C. The practical accuracy in relative humidity at constant temperature solely depends on the the humidity calibration with the cooled mirror optical dewpoint hygrometer and a psychrometer, which is typically 1 to 3 %RH best practice, respectively [In this study, an in-line acoustic relative humidity sensor for air-steam mixtures in duct flow has been designed. The measurement device is capable of measuring line averaged gas velocity, temperature and humidity instantaneously by applying two ultrasonic transducers and an array of four SMT temperature sensors. Measurement range is gas velocity of 0–12