A deleterious Laves phase forms in the solidified structure of Inconel 718 (IN718) alloy during laser cladding. However, effective removal methods have not yet been identified. In this study, we first added the IN718 alloy cladding layers with a trace amount of vanadium . Then, we studied the solidification structure of cladding layers using a confocal laser scanning microscope and scanning electron microscopy. The microstructure and Laves phase morphology were investigated. The distribution of niobium (Nb) was observed by experiment as well. We found that V is evenly distributed in dendrites and interdendritic zones. A more refined dendrite structure, reduced second dendrite arm spacing and lower volume fraction of Laves phase were observed in the solidification structure. The results of linear energy-dispersive X-ray spectroscopy (EDS) indicate that the concentration of Nb decreases with an increasing of the distance from the Laves phase. The V-containing sample displayed a relatively slower decreasing tendency. The IN718 alloy sample was harder with the addition of V. In addition, the porosity of the sample decreased compared with the blank sample. The presented findings outline a new method to inhibit the Nb segregation in IN718 alloy during laser cladding, providing reference significance for improving the performance of IN718 alloy samples during actual processing. Laser cladding  is a surAccording to Han et al., IN718 alloy with Mo can reduce the solubility of Nb in the dendrite arm and Laves phases. The addition of Mo transforms the Laves phase morphology from eutectiform to granular and lessens the area of segregation zone around the Laves phase . The effVanadium has many desirable physical and chemical properties . It was The addition of V appears to positively influence IN718 alloy microstructures. Due to the lack of information of the influence of V on the solidification structure of IN718 alloy during laser cladding, this was our goal in this study. The previous work ,10 on thThe materials in this experiment included spherical IN718 alloy powder a prepare2). An OLYMPUS-OLS4100 Confocal Laser Scanning Microscope  and a Zeiss ULTRA PLUS Scanning Electron Microscope  with a X-Max 50 Energy Dispersive Spectrum  were used to characterize the microstructure and chemical composition. The average diameter of equiaxed dendrites (DED), secondary dendrite spacing (SDS), average size of columnar dendrites’ primary dendrite spacing (PDS) and volume fraction of Laves phase (LPVF) measurements were counted and calculated by Image-Pro Plus6.0 software  using secondary electron micrographs of the etched alloys captured by SEM. Three locations were selected along the height of the sample to measure the DED, SDS, and PDS  from both macroscopic and microscopic aspects. In addition, the results were compared with the samples  without the addition of V. The chemical composition comparison between the two alloys is illustrated in The addition of a trace amount of other powder into the original powder may change several parameters and mechanical properties of cladding layers. Generally, a more refined secondary dendrite spacing is desirable. According to Ahmadetal et al. , SDS depSolidification structures with smaller secondary dendrite spacing limit the diffusion range of Nb Hence, decreasing the area ratio of the element segregation regions was more effective to achieve homogenization after heat treatment in No.1 alloy. In summary, the addition of V decreases the porosity in cladding layers and leads to a certain degree of dendrite refinement. Refined solidification and low porosity can enhance the performance of cladding layer.In this study, we performed a surface scan analysis on V-containing cladding layers. The results of linear EDS analysis indicated that Nb is heavily enriched in Laves phase. The Nb concentration decreases with increasing distance from the Laves phase. This tendency was observed in both No.1 and No.2 alloys. The difference was that the V-containing sample displayed a slower decreasing tendency. In addition, the average concentration of Nb in the analysis zone of No.1 alloy was less than that of the No.2 alloy, which is consistent with the results in As is stated before, the existence of Laves phase can drastically degrade the performance of IN718 alloy. Under higher magnifications (2000×), a clear island-like segregation zone can be defined, in which the Laves phase particles are distributed . The addHardness is an important performance index used to measure the hardness degree of metal materials. Meanwhile, hardness has been widely used as a preliminary evaluation of the wear resistance of alloy . TherefoThe discussion above indicates that the addition of V can inhibit element segregation and change the morphology of the Laves phase, which is similar to adding other alloying elements. However, the decreased of porosity and increased micro hardness in the IN718 alloy sample fabricated by laser cladding was not mentioned in previous studies. Furthermore, a new phase precipitated in the interdendritic region of IN718 alloy with the addition of B or Co, such as B-bearing phase and Mo-depleted gray phase, and the influence on performance is unknown. The addition of V can provide more refined microstructure, without substantially affecting the element distributions and phase compositions.In this study, the influence of V on IN718 alloy solidification structure during laser cladding was investigated using experiments. Based on the obtained results our main conclusions were drawn:(1) With the addition of V, the porosity of cladding layer decrease by 89%, and the average secondary dendrite arm spacing decreased from 3.8 to 2.6 μm, as did the average size of dendrite.(2) The Nb concentration in the Laves phase of the V-containing sample decreased compared with blank sample. The results of linear EDS indicate that Nb concentration decreases with the increase in the distance from the Laves phase. However, the V-containing sample displayed a slower decreasing tendency.(3) The morphology of the Laves phase in the V-containing cladding layer was modified dramatically, which changed from rod-like shape to a particle-like feature. The volume fraction of the Laves phase decreased by 80% with the addition of a trace amount of V.(4) The hardness of the V-containing alloy was higher compared with the blank alloy. However, the distribution of micro hardness was uneven.Consequently, the addition of V positively influence the microstructure and element segregation of IN718 alloy cladding layers. This investigation provides a new and effective method for inhibiting the formation of Laves phase and enhancing the performance of IN718 samples fabricated by laser cladding.


This study was designed to assess the analytical performance of verified a: bias%/TEa) on the Y-axis and the normalized imprecision (CVa: mean CV%/TEa) on the X-axis.The coefficient of variation (CV) was calculated according to the mean and standard deviation (SD) derived from the internal quality control for 20 consecutive days. The data were measured on an Architect c16000 analyser with reagents from four manufacturers. Commercial reference materials were used to estimate the bias. Total allowable error (TEa) was based on the CLIA 1988 guidelines. Sigma metrics were calculated in terms of CV, percent bias and TEa. Normalized method decisions charts were built by plotting the normalized bias . All reagents performed well for the aspartate aminotransferase and uric acid assays with a sigma of 5 or higher. Abbott achieved the best performance for the urea assay, evidenced by the sigma of 2.83 higher than all reagents, which were below 1-sigma.Sigma metrics analysis system is helpful for clarifying the performance of candidate non-verified reagents in clinical laboratory. Our study suggests that the quality of non-verified reagents should be assessed strictly. Quality control (QC) is the essence and core of a laboratory, assuring the quality of results that are critical for clinical diagnosis and patient care. To produce reliable results, the quality of testing is usually monitored with Levey-Jennings charts to provide a good review of analytical precision in particular, but establishing the internal quality control (IQC) program for analytical phase is not sufficient  and make them become an objective and integrated assessment of the quality of analytical tests. Sigma metrics analysis provides a standardized scale for comparing the quality of testing results by evaluating the performance of testing procedure or methods  or International Organization for Standardization (ISO) 15189 are well accepted in many laboratories, no global consensus currently exists on which allowable total error to choose for assessing acceptability. Sigma is a metric that measures the performance of a process quantitatively. In statistics, six sigma represents an ideal situation with the possibility of producing defects in a process of only 3.4 defects per million  to compare their performance on Sigma scale. Our goal was to provide a practical and instructional approach in assessing the performance of verified and non-verified reagents. This is of great significance to the quality control of clinical laboratories in China.We conducted a prospective study in the clinical laboratory in Shengjing Hospital, using an Architect c16000 chemistry analyser. The study was conducted over the period of one month from July 2017 to August 2017. Reagents for biochemical analytes were provided by four manufacturers, including Abbott , Leadman , Mindray  and Zybio . The reagents of Abbott and Leadman had been verified on Abbott Architect c16000. They were regarded as verified reagents. The reagents of Mindray and Zybio had not been verified on Abbott instruments. They were non-verified reagents. All tests were performed on the same instrument using the same reagent probe to ensure the comparability of results. The same lots of reagents were used during the study period. As the mandatory and representative analytes for enzyme activity and metabolism, alanine aminotransferase , aspartate aminotransferase , urea and uric acid (UA) were tested with the reagents from manufacturers of Abbott , Leadman  and Mindray  except Zybio , simultaneously to acquire the internal quality control (IQC) data and external quality control (EQC) data. No Zybio reagent was available for UA assay.The internal QC data were analysed against the recommended calibration intervals in the reference manual. For reagents of Abbott, ALT and AST assays were calibrated with water. Urea and UA assays were calibrated with Multiconstituent Calibrator . For the reagents of Leadman, multi-biochemical calibrator  was used for calibration. ANDOX CAL2351  was used for calibration of the reagents of Mindray and Zybio.Internal QC materials, Lyphochek Assayed Chemistry Control   were tested once daily, using reagents from four manufacturers concurrently. The coefficients of variation (CV) were calculated with the mean values of internal QC data during 20 consecutive days.Bias data were acquired from the commercial reference materials with target values, provided by National Institute of Metrology  for UA (GBW09157) and Medical device laboratory  for urea (GBW(E)090547), ALT and AST (GBW(E)090593). The relative bias was calculated with the average results of five repeated tests and the target values.According to the Clinical Laboratory Improvement Amendments of 1988 (CLIA 1988), the recommended allowable error (TEa) values for analytes are presented in IQC(%) = (standard deviation × 100) / laboratory mean(IQC). Percent bias was calculated as Bias(%) =  × 100 / . Sigma metrics were calculated from CV, percent bias and TEa for the analytes using the formula Sigma metrics = (TEa - Bias) / CVIQC , and CV were analysed with SPSS version 21.0  (a = bias / TEa) on the Y-axis and the normalized imprecision  on the X-axis, considering the different TEa .The Sigma metrics values calculated for each control level and calculated from the mean calculated CV are shown in a exceeded the maximum scale on the X axis.To provide a quick visual decision about the acceptability of verified and non-verified reagents for biochemical analytes, a normalized method decision chart was drawn for different reagents . The chaSigma metrics is a standard management model for improving laboratory quality by identifying and minimizing errors. It is applicable for assessing and directly comparing the quality of analytical processes. The performance and equality are consistent with Sigma metric value  for calculating the Sigma metrics are taken from the guidelines of CLIA (et al.’s, but the unit of TEa was not specified in Nanda et al.’s report. However, given the variability of TEa provided by different guidelines, the Sigma metric vary either. A previous study has also reported that TEa targets from different sources have important impact on the interpretation and application of Sigma metrics in routine laboratory testing (Abbott reagents provide the best Sigma metric value of 2.83 for the urea assay, but this is still under the basic requirement of 3-sigma. The results of other reagents are below 1-sigma, which is an unacceptable analytical performance. Our results are comparable to the results of Singh The analytical imprecision, CV, is the measure of instability of analytical system. In daily laboratory testing, the higher CV means the assay has more variation and less precision. In this study, the CVs were calculated with the daily control data during a period of 20 consecutive days. To ensure assessments’ reliability and stability, the study is preferably conducted over a longer time period. Bias, as a main component that the sigma value is dependent on, estimates the inaccuracy of results and is usually assessed by difference in the means of results like EQA. However, different manufacturers, instruments and methods significantly influence the bias, even the Sigma metric value (There may be such a misunderstanding that high Sigma metrics level means better precision and accuracy. For example, Leadman reagents provide better Sigma metrics values due to higher calculated CV and smallest bias, while Mindray reagents show lower Sigma metrics value due to smaller CV and bigger bias. These data clearly indicate that Sigma metrics is a summary indicator combining multiple key analytical performance parameters.et al. found that the Sigma metrics values calculated at different concentrations are quite different (Burnett One major limitation of this study was that there was no corresponding calibrator available for the reagents of Mindray and Zybio, and a Randox calibrator was used instead. However, Randox calibrator was recommended by the introductions of Zybio. The replaceable calibrator was not declared in the introductions of Mindray, but the assignment of Randox calibrator can be traced back to the internationally recognized reference material or reference method which is consistent with the reagent of Mindray. In spite of this, it maybe still affects the bias and resultant Sigma metrics value for the reagent of Mindray and Zybio.etc (To conclude, currently many reagents produced in China have emerged and the qualities vary significantly from manufacturer to manufacturer. During reagent selection, it is critical to select with acceptable Sigma metrics level. When clinical laboratories plan to switch reagent systems, our results suggest that the non-verified reagent system generally has relatively lower Sigma metrics level than verified reagent system, but it needs to be noted that this result is not absolute. Besides, the Sigma metrics varies with the reagent, analyte, and concentration. Therefore, Sigma metrics analysis system is helpful for clarifying the performance of candidate non-verified reagents in clinical laboratory. This process should be performed strictly. As the Sigma metrics has been applied not only in the “established” laboratory, but also in developing countries like Ghana and India,


Gold is for the mistress—silver for the maid—Copper for the craftsman cunning at his trade!“Good!” said the Baron, sitting in his hall,“but iron—cold iron—is master of them all.”Rudyard KiplingIron is a central component of electron chains and a co-factor of many vital enzymes. Only a few bacteria are able to substitute iron with other metals, making it an essential element for virtually all life forms. In plants, iron is also required for photosynthesis and chlorophyll synthesis. The availability of iron in soils dictates the distribution of plant species in natural ecosystems and limits yield and nutritional quality of crops. Insufficient iron uptake causes retarded growth, interveinal chlorosis, and reduced fitness. Sufficient iron levels in food crops are critical to combat iron deficiency-induced anemia, one of the largest nutritional disorder worldwide. Too much iron is, however, toxic to cells. It is therefore mandatory for plants to overcome the often-restricted availability of soil iron by strategies that increase its mobility and restrict its uptake when present in excess.th edition of the ISINIP held in 2018 in Taipei, Taiwan.Despite great leaps forward in the research into plant iron nutrition over the past decades, many facets of cellular iron homeostasis still await further clarification. Moreover, attempts to increase the iron content in edible plant parts are far from having reached sufficient improvement in dietary iron intake. The International Symposium of Iron Nutrition and Interaction in Plants (ISINIP) is a biannual meeting that covers a wide range of aspects, including but not limited to iron availability in the soil, the regulation of cellular iron homeostasis, and the exploration of novel avenues for fortifying plants with iron. The collection of mini reviews, perspectives, and original papers presented in this Research Topic is in part associated with contributions to the 19Von Voithenberg et al.). New transporters have been identified also for the transport of iron through the inner mitochondrial membrane. Jain et al. show that the Arabidopsis transporters MIT1 and MIT2 are involved in iron import into mitochondria and critical for mitochondrial function.While the mechanisms underlying the uptake of iron from the soil are relatively well understood, the trafficking of iron to chloroplasts and mitochondria is less well explored. Chloroplasts are loaded with transition metals and represent the iron-richest system in plant cells. Research conducted by the group of Kathrin Philipar suggested that the ATP-binding ABC-transporter subunits ABCI10 and ABCI11 are part of a novel module of a prokaryote-type ECF/ABC transporter with similarities to components of prokaryotic multi-subunit ABC transporters putatively involved in metal ion uptake  transporter EFFLUX TRANSPORTER OF NA (ENA1) in rice, which is involved in the transport of NA to the apoplast and in the import of iron into cellular compartments such as plastids. The authors conclude that in rice NA transport by ENA is critical for cellular iron homeostasis by shielding iron ions from interacting with other molecules, causing precipitation of iron and oxidative damage to the cell.Iron is highly reactive and must be chelated throughout intra- and intercellular trafficking to avoid cellular damage. Gao et al. summarize the current knowledge regarding the complex network of bHLH proteins that cooperatively regulate iron uptake in the green lineage. The bHLH protein FIT (bHLH29) takes a central position in this regulatory network. The activity of FIT is sophisticatedly controlled by a suite of proteins, which either activate FIT or enhance its degradation. Wu and Ling review the action of proteins that have been shown to interact with FIT and post-transcriptionally fine-tune its activity.The regulation of iron uptake has to meet, but not to exceed the demand of the plant to avoid undernourishment or toxicity. Therefore, cellular iron levels are controlled by a delicate signaling network that orchestrates the demand of different plant parts in the context of rapidly and constantly changing availability of iron in the soil. A key role in the regulation of iron uptake is played by bHLH proteins. Until now, 16 bHLH transcription factors of this type were shown to be involved in the control of cellular iron homeostasis; it can be expected that this number is not yet exhaustive. In their mini review, Escherichia coli BolA proteins, may represent such common actors. In their Perspective paper, Rey et al. speculate that glutaredoxins, alone or in complex with BOLAs, may function in the maintenance of iron homeostasis in plants. More evidence for a specific function in iron signaling has accumulated for a small family of hemerythrin E3 ubiquitin ligases unique to the green lineage. HRZ (Hemerythrin RING Zinc finger) proteins in rice and their homologs BTS (BRUTUS) and BTSL (BRUTUS-LIKE) in Arabidopsis modulate the activity of specific transcription factors in a negative feedback loop, and are critical in avoiding the uptake of toxic amounts of iron. In their mini review, Rodríguez Celma et al. provide an overview on what is known about the putative function in iron sensing and signaling of this enigmatic group of regulators.While the mechanisms underlying the perception of the cellular iron status differs across the kingdoms of life, some actors involved in iron sensing appear to be commonly recruited by yeast, mammals, and plants. Glutaredoxins and partnering BOLAs, homologs of Romera et al. review the available data on this “ironic liaison,” and investigate the possibilities to use ISR-eliciting microbes as both pesticides and iron fertilizers in a more sustainable agriculture.Plant growth-promoting bacteria can trigger both induced systemic resistance (ISR) and a partial iron deficiency response, indicating overlap in the regulation of the two processes. Infection with growth-promoting bacteria improves plant growth, increases iron content, and boosts plant defenses, conferring resistance to pathogens and pests. Kaisalam et al. took a different approach toward the dissection of iron signaling pathways. The authors identified two small molecules, referred to as R3 and R6, through a chemical screen and showed that both molecules efficiently compromised the iron deficiency response of Arabidopsis, likely by targeting different signaling pathways. While R6 influences the IVc clade of bHLH proteins, R3 appears to affect a so far unexplored route toward the transcription factor FIT.Park et al. proposed that PRC2 (Polycomb Repressive Complex 2)-mediated methylation of lysine 27 of histone 3 attenuates the induction of FIT target genes, restricting iron uptake to avoid possible overload. The authors conclude that this epigenetic feature prevents maximum induction of genes involved in iron acquisition, providing an additional mechanism to fine-tune gene activity.Post-translational modifications of lysine residues of core histones such as acetylation or methylation are a key component of eukaryotic gene regulation. Carrió-Seguí et al. show that miR408 targets laccase-like multicopper oxidases with putative promiscuous activities interfering with the redox homeostasis and, ultimately, with iron signaling. The study further show that modifying miR408 levels resulted in complex phenotypes characterized by increased oxidative stress, altered lignification and, as a consequence of the latter, compromised metal transport.MicroRNAs (miRs) are a further node in the iron-signaling network. Lucena et al. addressed such interactions between iron and phosphate and found that deficiencies in one nutrient resulted in transitory induction of the expression of genes encoding proteins involved in the uptake of the other. The authors speculate that this induction is possibly triggered by phloem signals originating in the shoots that interact with hormones to balance the uptake of mineral nutrients from the soil. An antagonistic crosstalk between iron and phosphate signaling was also reported with respect to the accumulation of catecholic coumarins . The authors show that phosphate availability directly influences iron deficiency-induced accumulation of coumarins, a process that appears to reflect crosstalk between the regulators controlling the responses to iron and phosphate deficiency.The iron homeostasis network is sophisticatedly interwoven with the uptake and transport of other mineral nutrients, often in an antagonistic manner. Nicolic et al. observed an ameliorative effect of silicon on iron deficiency stress in barley, a plant with predominant Strategy II iron acquisition. Silicon significantly modulated the expression of Strategy II genes and increased the total iron content in the youngest leaves, which was associated with a lower ROS level, elevated APX and CAT activities, and increased chlorophyll levels. Moreover, silicon improved root-to-shoot iron translocation, resulting in an increased iron content in the youngest leaves.Similar to what have been previously reported for strategy I plants, Buckner et al.).Studying environmental cues such as Fe deficiency in isolation covers the processes that orchestrate the hierarchy of responses to simultaneously perceived signals and stresses. Terri Long’s group developed an image analysis pipeline to extract spatiotemporal metrics of GFP signals showed that subjecting the plants to both heat stress and iron deficiency affected the spatiotemporal function of the mitosis marker CYCB1;1 antagonistically, suggesting interplay between stress pathways when plants are exposed to multiple environmental factors that may modulate the response that is observed in experiments that singles out a specific stimulus  channel OsAKT1 affects the translocation of iron from roots to shoots, a process that is contributing to the tolerance to iron toxicity. Loss-of-function mutants of OsAKT1 exhibit altered K homeostasis, lower K+ concentrations, and hyperpolarization of the plasma membrane, leading to decreased iron loading into the xylem. A further important player in the tolerance of rice to high iron levels is the vacuolar iron reductase OsFRO1, which maintains cellular iron homeostasis by tuning the iron concentrations in the cytoplasm and in the vacuole .If in excess, iron can react with hydrogen peroxide and trigger the formation of harmful hydroxyl radicals through Fenton chemistry. The free iron concentration drastically increases with decreasing redox potential, as it is the case for instance in flooded soils. Although genetic variation in iron tolerance suggests the evolution of adaptive strategies to cope with high external iron levels, the mechanisms underlying such adaptations have not yet been fully elucidated. Aung et al. show that expression of the rice NA synthase OsNAS3 strongly increased when plants were subjected to excess iron levels; OsNAS3 knockout plants showed reduced growth and severe leaf bronzing, resulting from increased iron accumulation. The results demonstrate that NA is important for mitigating excess iron and provide a new perspective for the development of rice lines with improved tolerance to excess iron availability soils.The metal chelator NA plays multifaceted roles in metal homeostasis and appears also to be important to protect the plant from iron overload. More than 30% of the arable land are calcareous and potential iron-deficient. Low iron content of crops is the main cause of iron deficiency-induced anemia, the most common micronutrient deficiency globally. Iron-biofortification denotes the process of enhancing bioavailable iron in edible parts of staple food by agronomic approaches, biotechnology techniques, or conventional plant breeding. Biofortification is a promising approach to combat micronutrient malnutrition, which affects nearly one-third of the world’s population, particularly in resource-limited settings.Zanin et al. review the complex issue of how humic substances affect plant performance and investigate the possibility to use iron-humic complexes as an environmentally friendly source of iron fertilizer. The supposition that increasing crop production through innovative iron fertilizers is a promising alternative to traditional iron fertilization is also supported by a study by Cieschi et al. The authors describe the action of three iron-humic nanofertilizers as a natural, low cost and environmental option to improve iron uptake of soybean grown on calcareous soils by providing iron over prolonged time periods.Iron fertilizers are costly, not environmentally friendly, and often inefficient. Iron-humic complexes have long been known to promote iron nutrition in a multifaceted way by providing a readily available iron form in the soil and by directly impacting physiological and developmental programs. Dai et al. review the effects and mechanisms underlying the promotive effects on iron nutrition focusing on Leguminosae/Gramineae intercropping systems. Intercropping is particularly beneficial for calcifuge species adapted to acidic soils. Blueberry, a calcifuge, develops severe iron deficiency symptoms under alkaline conditions, drastically reducing plant growth and yield. Intercropping with the grasses Festuca rubra and Poa pratensis in combination with Fe-EDDHA applications was found to be effective in increasing fruit load and yield of blueberries while reducing the skin/flesh ratio and firmness of the berries .Intercropping of fruit trees or crops with graminaceous plants can have beneficial effects on the iron status of iron-inefficient plants and has been proven useful for low-input/resource-limited agricultural systems. Ludwig and Slamet-Loedin. The authors summarize the problems and advances of different biofortification strategies to enrich rice and wheat grain iron to combat “hidden hunger.” Rice has employed components of both strategy I and strategy II to acquire iron from the soil. Masuda et al. reported that transgenic rice with increased phytosiderophore production and Fe(III) reduction activity conferred by introducing a gene cassette comprising a regulator of the iron deficiency responses (IRO2), a barley IDS3 genome fragment to increase PS production, and a mutationally reconstructed yeast ferric reductase exhibited enhanced tolerance to iron-deficient conditions and significantly higher yield when cultivated on calcareous soil.Genetic biofortification, i.e. conventional breeding and genetic engineering, is the topic of a mini review by Eroglu et al. investigated metal localization in seeds of more than twenty species using histochemical or X-ray based techniques. The study revealed distinct seed iron storage patterns across plant lineages. The authors further report that in Rosids iron is concentrated in the innermost cell layers, the endodermis, and in the cortex.The distribution of iron in seeds in a main determinant for its bioavailability, which can be severely restricted by antinutrients such as phytate that are localized in the same cellular compartments. With the goal to investigate if iron can be stored in phytate-free compartment such as plastids, All authors wrote and approved the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The major drawback of the community-based mass drug administration (MDA) approach against schistosomiasis is that treatment is offered blindly without testing for the targeted infection. This partly contributes to the low treatment coverage. One approach to overcome this limitation is to introduce a diagnostic component in the treatment approach. This will improve drug uptake and compliance to treatment. This study is conducted to assess the feasibility and acceptability of integrating point-of-care Circulating Cathodic Antigen (POC-CCA) test to community-based directed MDA in improving treatment coverage and compliance with treatment among adults.S. mansoni infection based on the POC-CCA test conducted by CHWs, ability of CHWs to use the POC-CCA test accurately and safely and community acceptability of the POC-CCA test results from CHWs. Both quantitative and qualitative techniques have been used to collect data at study endpoint.This is a randomized control community trial in which 30 clusters were randomly assigned to either an intervention or control arm to evaluate two interventions on treatment coverage and compliance with treatment. In each cluster, 150 adult participants were enrolled. Community Health Workers (CHW) in both arms were trained on all aspects of praziquantel (PZQ) distribution and management of mild side effects. In the intervention arm, CHWs had additional training on how to use POC-CCA to diagnose intestinal schistosomiasis. In the intervention arm, participants were tested using POC-CCA test for presence of intestinal schistosomiasis and treated based on test results, while in the control arm, participants were treated with PZQ without testing. The primary outcome measure was the proportion of participants provided with PZQ between the two arms and geographical clusters. Secondary outcomes were prevalence of The study will generate evidence on the importance of integrating a diagnostic component into the community directed MDA conducted by CHWs. Findings will generate discussion on the current MDA policy and practice in Tanzania.PACTR201804003343404 (25/4/2018). Schistosoma mansoni and Schistosoma haematobium which cause intestinal and urogenital schistosomiasis respectively [S. mansoni has remained a major public health concern, especially among communities living along and on Lake Victoria islands, in the north-western region [Schistosomiasis infection is a neglected tropical disease which is highly prevalent in Sub-Saharan Africa (SSA), and is a major cause of public health burden and economic impact , 2. Withectively . Of the ectively . In Tanzectively . In Tanzn region , 6. Disan region , 8.The World Health Organization’s (WHO) strategic plan of 2012 proposed scaling up of mass drug administration (MDA) as a means of controlling schistosomiasis by 2020 . CurrentTo increase treatment coverage, community directed treatment has been implemented as an alternative approach for controlling schistosomiasis in a number of African countries to improve access to treatment for community members not included in mass drug administration program , 15. ThrPoint of care Circulating Cathodic Antigen (POC-CCA) test is a simple rapid test, easy to perform and interpret, as the results based on colour change . The tesThe pre-existing community health workers participating in MDA campaign at Kome Island offers an opportunity to improve the community-based MDA campaign by introducing a diagnostic component into the program to increase confidence of participants in the campaign and increase uptake of treatment . In addiIn this study, we hypothesize that integrating point-of-care Circulating Cathodic Antigen tests in community directed mass drug administration campaigns increases the uptake, compliance and coverage of schistosomiasis treatment among adult individuals above the 75% recommended by the WHO . The genSchistosoma mansoni infection.To determine the impact of using point-of-care the circulating cathodic antigens rapid diagnostic test by community health workers to diagnosed intestinal schistosomiasis on the uptake of praziquantel treatment among adult individuals diagnosed with Primary objective:-Schistosoma mansoni infection in adult individuals and associated factors.To assess community health worker’s ability to use point-of-care the Circulating Cathodic Antigen rapid test safely and accurately in diagnosing Schistosoma mansoni infection within the context of community directed intervention among adults.To assess community acceptability and attitude towards the use of a point-of-care diagnostic tool for Schistosoma mansoni infection using the point-of-care Circulating Cathodic Antigen rapid test.To document the experience of community health workers after participating in diagnosis of Secondary objectives:-S. mansoni infection and existence of a strong CHWs system involved in previous community-based schistosomiasis control projects. According to the population census of 2012, the Kome Island has a population of 38,062. Administratively, it has four wards and 16 villages. It is served by one health centre and two dispensaries. Each village has at least one primary school, with the exception of Isenyi village, which has three primary schools, and four villages with no primary school. The main sources of water are Lake Victoria, natural wells and streams. Schistosomiasis is highly endemic in the area and Schistosoma mansoni-related morbidities are quite high among the adult population. Recent records indicate that 42.2% of the adult population had periportal fibrosis, 68% had left liver lobe hepatomegaly and 55.2% had splenomegaly [A cluster randomized two-arm community trial was conducted, in which clusters were defined as the service population of the community health worker and it involved 30 clusters. The study was conducted at Kome Island located in Buchosa District, North-Western Tanzania Fig. . The Komnomegaly .Fig. 1ShSchistosomiasis control activities at Kome Island started in 1989 and focused mainly on research and treatment of infected individuals . From 20Community Health Workers (CHWs) are an integral part of the health care system in Tanzania , 27. ComAt Kome Island, based on previous projects, each of the then 53 sub-villages had two CHWs . However, currently there are 80 sub-villages, and the present study used existing CHWs previously trained on the community-based MDA campaign . There wThe study was designed as a cluster randomized two-arm community trial involving randomly selected sub-villages (clusters) at Kome Island. In the intervention arm, community members were tested for intestinal schistosomiasis by CHWs using the POC-CCA test and treated based on the test result Fig. . In the To assess CHWs ability to use the POC-CCA test safely and accurately in the intervention arm, a trained observer (a laboratory technician) visited the study cluster at two, four and six weeks post-training and used a checklist to assess CHWs compliance to safety procedures Table .Table 2SAt baseline, a quantitative questionnaire was used to collect data on CHW’s and community member’s (heads of households) knowledge about schistosomiasis, the previous community-based MDA strategy, acceptability to be diagnosed for intestinal schistosomiasis by CHWs and treatment with PZQ without being diagnosed for the targeted infection. At the end of the study, qualitative Focus Group Discussions (FGDs) were conducted to assess the experiences of CHWs from both arms after participating in the trial. FGDs were held with selected community members who participated in the study to assess community acceptability and attitudes towards use of POC-CCA test by CHWs.Randomization of clusters to either intervention or control arms was based on the population size of the adult individuals served by a community health worker. All CHWs serving clusters with less than 60 adult populations were excluded from the study.ralloc programme in Stata version 13. Community Health Workers was the unit of randomization and the unit of analysis (because CHW were representing each of the sub-villages they served). Thirty clusters (i.e. service population served by CHW) with ≥60 adult population were selected from all eligible clusters to participate in the study. The selected cluster were at least 7–15 km away from each other, regardless of whether they were randomized to be in the control or intervention arm. The cluster encompassed a radius around the CHW of at least 5 km and allowed a buffer zone of at least 5 km between the boundaries of different clusters. Buffer zones between the sites had no activities undertaken to prevent contamination between clusters.Randomization of the CHWs to receive the POC-CCA test plus PZQ or PZQ without POC-CCA testing was performed using the The primary outcome is the proportion of study participants provided with praziquantel drug (treatment coverage) and geographical cluster  in the intervention and control arms.Community Health Workers (CHWs) ability to use the POC-CCA rapid test accurately and safely.Community members acceptability (these are people’s perceptions and attitude on the two-treatment approach) of the POC-CCA test results offered by community health workers.Experience of Community Health Workers on the use of the POC-CCA rapid diagnostic test.S. mansoni infection based on the POC-CCA test conducted by Community Health Workers.Prevalence of Secondary outcomes are:-Individuals aged 18 years and older residing at Kome Island were enrolled after written informed consent was obtained. Participants were enrolled in the study if: (i) aged 18 years and above (ii) willing to participate and give written informed consent and (iii) living in the study area. Participants were excluded from the study if: (i) pregnant at the time of treatment (ii) history of adverse reaction to praziquantel and (iii) present with serious conditions at the time of treatment and not deemed fit to take treatment.The sample size was estimated using a simplified formula by Hayes and Bennet for cluster randomized trials , with a Community health workers attended a three-day training on schistosomiasis, covering topics that included epidemiology, the national neglected tropical diseases control program and mass drug administration approach (school-based versus community-based treatment approach), causes, signs and symptoms of intestinal schistosomiasis and standard operating procedures of the study. In addition, CHWs were trained on how to dispense praziquantel drugs using the praziquantel dose pole, based on the height of the study participants and how to record the number of tablets/pills provided to each participant Fig. . The CHWCommunity health workers in the intervention arm attended one day of additional training covering how to perform and interpret the rapid test for intestinal schistosomiasis Fig. . The topAfter the training, all community health workers were provided with praziquantel dose poles, registry booklets, writing materials and praziquantel drugs. In addition, community health workers in the intervention arm were provided with gloves, buckets, urine containers and point-of-care circulating cathodic antigens tests.Two laboratory technicians were trained on how to assess the ability of trained CHWs in the intervention arm to adhere to the safety procedures and standard operating procedures of performing the POC-CCA test. The laboratory technician acted as an observer, visited the study sites (clusters) at two, four and six weeks post-training to assess CHWs using a standardized checklist (steps of test preparation and safety measures) as shown in Table Trained CHWs in both arms recorded all demographic information of the study participants in the provided booklets. Collected demographic information included age, sex, sub-village of residence and history of taking anti-schistosomiasis treatment.S. mansoni infection using a commercially available test, the Point-of-Care Circulating Cathodic Antigen test , a rapid test for intestinal schistosomiasis [Trained CHWs in the intervention arm diagnosed study participants for somiasis , 30 based on the test results and any reported side effects. In the control arm, CHWs recorded the number of tablets/pills provided to each study participant and any side effects reported by participants.Diagnosis of intestinal schistosomiasis and provision of PZQ drugs was done at an agreed site within a cluster. Tablets were swallowed under direct observation (DOT) of CHWs. Estimation of number of PZQ tablets given to each participant was done using the praziquantel dose pole, based on the height of an individual Fig. . Study pA qualitative research approach using Focus Group Discussions (FGDs) was used to assess community acceptability of the approach. Eight community clusters out of 30 community clusters were selected (6 in each intervention arms and 2 control arms). In each of the cluster, one (1) FGDs session was conducted giving a total of 8 FGDs sessions. Each FGD consisted of 8–10 participants in each cluster. These FGDs participants included only those who participated in the study from the two arms. Focus Group Discussion participants were mobilized by CHWs from their respective cluster community and FGDs was conducted at a selected area near the cluster community. The interview guide was used as a tool to collect information on trust and confidence in CHWs, community willingness and acceptability of both approaches. For the intervention arm, additional information was collected on CHWs use of CCA rapid tests and trust of the test results from CHWs, compliance to treatment given and stigma associated with requesting urine samples from adult participants. The interviewer and the note taker were social scientists, speaking Kiswahili and the vernacular fluently. Table Focus Group Discussions with CHWs were conducted at the end of the study. A guide was used to stimulate the discussion and the discussion focused on their experience on using the CCA kits and provision of treatment after diagnosis, what were the challenges during MDA, what were the opportunities and what were the perspectives of the participants on the exercise, participants trust of the results and their recommendations to improve the MDA exercise with and without a diagnostic component. Table A trained observer visited each of the CHW in the intervention arm at one, two and six weeks post-training and used a standardized checklist Tables  and 5 toObservers were trained on how to use the check list and instructed on how to minimize observer bias. Using the checklist, the observer noted if CHWs performed each step correctly and also noted if she/he missed observing any step. The steps were divided into two parts, (i) those critical to safety and (ii) accurate performance of the test. Safety was defined as correctly wearing gloves, use of new urine container for every participant and disposal of the urine containers in approved buckets and urine samples in the toilet immediately after use. Whereas accuracy was defined as following all the necessary steps correctly to arrive at a correct diagnosis (i). Collecting urine samples in an enclosed pipette and pouring it in a test kit well (ii). Dispensing 2–3 drops of clearing buffer into the sample well (if provided by the manufacturer) (iii). Waiting for 15 min before reading test results (iv). Reading test results correctly (v). Recording the results. At each observation, the observer recorded number of POC-CCA rapid test performed since the previous visit and collected the used test kits, number of positives and number of treated participants. In addition, the observer recorded any challenges or concerns experienced by CHWs.Process monitoring approach using structured and unstructured observations methods was used to assess if activities were implemented as per study protocol. Unstructured observations mainly focused on behavior of treatment activities if participants were asked or instructed to take food/meal prior to drug intake or coming to diagnosis and treatment points after taking meals. In addition, the process monitoring focused on assessing if treatment procedures were followed, the praziquantel dose pole was used properly, recording any adverse events was done, and how CHWs managed any reported adverse effects. A trained observer and field coordinator were involved in process monitoring.The CHWs in both arms were supervised after every one weeks by field-based supervisors and observers to detect and correct any deviations from the protocol. During the visit, the supervisor and observer assessed completeness of data captured in the booklets, adherence to inclusion criteria, filling consent forms, performance of POC-CCA tests, PZQ drug administration to participants, POC-CCA test storage condition and assessment of the tests. After every two weeks during the study time, a review meeting was conducted to address any challenges. All treatment registers were reviewed and CHWs with any challenge/problem received support and re-training. The accuracy of CHWs in performing the POC-CCA test was assessed at each visit.The questionnaires which was used for the study, was developed and piloted in 70 participants from villages outside the study villages. The focus of the piloting activity was mainly to check for reliability of the questions. Field research assistants were trained on how to administer the questionnaire. For the qualitative data collection, an experienced social scientist was involved in training research assistants on the objectives of the study, how to take notes and how to conduct focus group discussions. Discussion guidelines were piloted to ensure that the meaning of the questions was clear to participants. Some questions were adapted from previous studies , 32. AllSome parts of the open checklist were coded before entered in the computer for analysis. Collected data were double entered using CSPro and the final data set were stored in MySQL database. Data were screened for consistency and errors corrected. Data analysis will be done using Stata version 15 . Descriptive statistics will be used to describe participant’s demographic information. Assessment of uptake of praziquantel treatment will be done by comparing proportion of the outcomes between clusters in the two arms. For the intervention arm, treatment coverage will be assessed based on number of individuals tested and received treatment after diagnosis as registered in the booklets. For the control arm, coverage for praziquantel treatment will be assessed from the CHWs booklets/registers.Odds ratios (OR) and 95% confidence interval (CIs) will be calculated using random effect logistic regression analysis adjusting for clustering. For assessing safety and accurate use of CCA test by CHWs, percentage of total steps, crucial steps and no-crucial steps performed correctly will be calculated for each observation cycle and a generalized estimating equation (GEE) logistic regression model for panel data will be fitted to identify factors associated with correct performance of at least 90% of the total steps. In addition, pooled mean and median percentage for reading of the 20 photographic results will be calculated for each observation cycle.Descriptive statistics will be calculated of FGD participant’s demographic variables and socio-economic characteristics. The narratives during the FGD were recorded using a digital audio recorder, transcribed verbatim in the language of the study participants (Kiswahili) and translated into English. This was followed by verification for accuracy by a second social scientist fluent in Kiswahili and English. The transcripts generated were then imported into ATLAS ti, version 8 and qualitative data analysis was done according to procedures described previously . The focThe study was approved by the joint Ethical and Review Committee of Bugando Medical Centre and Catholic University of Health and Allied Sciences (CREC/200/2017). The study received further clearance from the region, district administrative authorities and village authorities. Kiswahili translated informed consent forms were used to obtain consent from study participants. Before recruiting study participants, CHW describes the objective of the study to participants, the benefits and any involve risk for them to participate in the study. For illiterate participants, a thumb print was used to sign the consent forms after a clear description of the study objectives was given to participants. Further permission was requested from study participants and CHW to include their pictures/images during publication of the study findings. All pictures present in this methodology section of the study have received permission from all the study participants and the CHW involved in the study. The trial is number PACTR201804003343404 registered at Pan African Clinical Trial Registry.The overall aim of the present study is to document the additional role of the community health workers to use the point-of-care circulating cathodic antigen test to diagnose community members participating in mass drug administration for intestinal schistosomiasis. The study focused on the adult population who are currently not included in the on-going national MDA exercise, and who are known to have very low compliance to treatment . The intSecondarily, the study will demonstrate the ability of the CHWs to use the POC-CCA safely and accurately. During training, majority of the CHWs demonstrated ability to interpret and differentiate POC-CCA test kits with positive and negative results. However, they had challenges in identifying invalid and unused POC-CCA test, which necessitated re-training. In addition, through the use of CHWs to diagnosed intestinal schistosomiasis using the POC-CCA test(s), the study will allow precise estimate of the prevalence of intestinal schistosomiasis among adult individuals in the study setting.The inclusion of qualitative studies will help to acquire knowledge about the community perceptions and CHWs experiences after participating in the study. The focus for the community is to understand their acceptability of the diagnostic component performed by CHWs and recommendation to improve the exercise. Community acceptability is crucial for sustainability of any health intervention. Learning from CHW’s experiences, challenges and recommendation after implementing the study will help in improving the ways to integrate diagnostic component to community-based MDA run by CHWs in endemic areas.If introduction of a diagnostic component in the community-based MDA will have a positive impact on drug uptake and compliance, this will provide stakeholders with important evidence on how community-based MDA can be made innovative by integrating a diagnostic component performed by community health workers. This will have appositive impact on the risk of transmission of the disease.


The study also indicates that the use of different sets of fingerprints results in models, for which the ability to discriminate varies with respect to the set level of acceptable errors.The occurrence of mutagenicity in primary aromatic amines has been investigated using conformal prediction. The results of the investigation show that it is possible to develop mathematically proven  Primary aromatic amines (PAAs) are present in a wide variety of chemicals including drugs, cosmetics, dyes, plastics, food products, tobacco and pesticides. They are commonly used in the manufacture of pharmaceuticals; however, PAAs are a structural alert for mutagenicity. This PAA alert has been extensively reviewed in the literature and there is a clear association between this class of compounds and mutagenic activity ,5,6,7,8.Several models and software are currently available for the prediction of mutagenicity such as CAESAR , Derek Nvalid predictions on new compounds corresponding to the user-defined significance level (the percentage of acceptable predictions errors). This includes taking applicability domain into account as part of the framework .pe = expected accuracy = [(TP + FN)(TP + FP) + (TN + FP) (TN + FN)]/[(TP + FP + FN + TN)Accurate predictions of PAAs are important for the development of new products, including pharmaceuticals. Prediction methods that both minimize false positive and false negative predictions are critical since false positives may lead to further unnecessary experimental testing (to establish whether the compound is mutagenic) or force a new research direction to be considered whereas, false negatives could compromise safety of those exposed to the chemical if not detected.valid models using the conformal prediction framework for predicting mutagenicity of primary aromatic amines.It is possible to develop mathematically proven The user of the models can set the acceptable level of errors.both and empty) provides the user with information on how to use, further develop or possibly improve future models.The existence of uncertain classes of prediction (The use of different sets of fingerprints results in models where the ability to discriminate varies with respect to the set level of acceptable errors.This study using conformal prediction highlights several interesting aspects with respect to in silico model building:


The Clock Drawing Test (CDT) is a frequently employed screening tool with different scoring systems. Quantitative and semi-quantitative scoring systems, such as Sunderland’s et al. (1989), do not discriminate different error patterns. Thus, the same score can represent a number of different neuropsychological profiles. Therefore, the use of a scoring method that emphasizes qualitative aspects to determine specific error patterns is fundamental.To use a qualitative scale to analyze error patterns in the CDTs of older adults who scored 5 in a previous study.49 CDTs with score of 5 were analyzed using the qualitative scale. Linear regression and hierarchical and non-hierarchical cluster analyses were performed.The linear regression showed a significant association between the total score and all the error patterns of the qualitative scale. The hierarchical cluster yielded three groups. However, due to the heterogeneity observed among the groups, a non-hierarchical cluster analysis was performed to better understand the results. Three groups were determined with different neuropsychological profiles and patterns of errors.The qualitative scoring of the CDT is important when examining and analyzing specific neuropsychological domains in older adults, especially executive functions. The Clock Drawing Test (CDT) has been extolled as a screening tool for dementia. Features such as rapid and easy application are among the reasons for its worldwide clinical use.,Methods can be classified into quantitative, semi-quantitative and qualitative.In a previous study,,-,The new algorithm of Mende-Santos et al.Given these difficulties with scoring methods of the CDT, a historical review was performed.,,Researchers have demonstrated the advantages of the qualitative approach. Some of these benefits are: differentiation of diagnostic groups,,,Despite all the qualitative analysis advantages and growing interest in its administration,Finally, there is heterogeneity among neuropsychological profiles with cognitive decline and important aspects for diagnoses can be determined by qualitative analysis.,Therefore, the purpose of this study was to analyze error patterns in the CDTs of older adults without dementia that had a score of 5 in a specific algorithm method adapted from Sunderland et al.,The sample comprised forty-nine older adults who attended a social program in Rio de Janeiro. All subjects participated in a previous studyFrom the protocol of cognitive screening tests previously applied,Descriptive statistical analyses were performed to summarize and elucidate the sociodemographic features of the sample studied. A Kolmogorov-Smirnov test was later used to check for a normal distribution of the variables. A linear regression (stepwise) was performed to assess possible associations between the error patterns of the qualitative scale and its total score. Inter-rater reliability was assessed by comparing CDT scores of the three independent raters.A hierarchical cluster, employing the Chi-square method as a dissimilarity measure, was used to group the participants with similar error patterns and total score. A non-hierarchical cluster analysis (K-means) was then administered to confirm the results. Chi-square tests were then performed to compare differences between the cluster groups and verify the level of significance between the variables. The level of significance was set at 1%, i.e. p≤0.01.The demographic and cognitive characteristics of the sample are given in The linear regression showed a significant association between total score and all error patterns on the qualitative scale, as depicted in The investigation of inter-rater reliability of the CDT scored by the Modified Qualitative Error Analysis of RouleauDue to the heterogeneity observed through the groups yielded by the hierarchical cluster shown in The aim of this study was to analyze error patterns in CDTs and verify whether different neuropsychological profiles could be distinguished. The sample was composed of cognitively normal older adults that all scored 5 (numbers counter-clockwise or concentrated) in a previous studyFabricio et al.The initial objective was to understand why more than half of the participants in the cited study had a score of 5, which is below the cut-off point for dementia, considering that the semi-quantitative methodConsidering all subjects in the present sample had planning deficits, the discussion will address the other frequent pattern of errors. In general, conceptual deficits (misrepresentation of time), graphic difficulties (mild) and size of the clock , respectively, were the other types of errors with higher frequency. Similar results were found in the study by Fabricio et al.,,A lower performance for clock numbers and hands  occurs among healthy older adults.,,A high prevalence of graphic difficulties and size of Clock pattern of errors were also present in studies performed with healthy samples.,,,,On the other hand, stimulus-bound response and perseveration errors were less frequent in this sample. The literature confirms that both stimulus-bound response and perseveration errors are not commonly committed by older adults without dementia.Among a sample with a semi-quantitative score of 5,Different neuropsychological profiles were evident. There were similarities among the groups  represenRegarding spatial/planning deficits, group 2 had a lower frequency of this type of error. On the other hand, the two other groups could not be differentiated solely using the qualitative scale. It is important to mention that the scale only scored the final drawing. As the best methods for determining planning strategies are those that analyze the whole process of construction of the drawing,,,Comparing healthy older adults to those with cognitive decline, the frequency of all types of error increases.As the exclusion criterion of the study was based on the MMSE, this sample likely comprised older adults without dementia. However, considering the type and level of errors of Group 3 and the presence of participants with partial dependence and depressive symptoms, the study might have included individuals with MCI.These results support the initial hypothesis that the use of a qualitative scoring method that takes into account specific error patterns could be useful for specifying e type and level of cognitive decline in different subgroups. Thus, different neuropsychological profiles can be described in a sample of older adults without dementia.The neuropsychological approach in scoring systems of the CDT is an important aspect, as the quantitative and semi-quantitative scoring methods alone cannot differentiate groups.Although the scale was also useful for discriminating types of errors of executive functions from constructional abilities, the type of analysis that scores the drawing only without considering the process and the sequence used in the construction, is not the best method for determining planning strategies. This information could be a complementary aspect to allow a better qualitative description of executive functioning in elderly. Therefore, further studies with a larger sample or a study that describes the construction strategies of drawing a clock could help understand planning and organization features.


Epilepsy with Centrotemporal Spikes (ECTS) is the most common form of self-limited focal epilepsy. The pathophysiological mechanisms by which ECTS induces neuropsychological impairment in 15–30% of affected children remain unclear. The objective of this study is to review the current state of knowledge concerning the brain structural and functional changes that may be involved in cognitive dysfunctions in ECTS. Structural brain imaging suggests the presence of subtle neurodevelopmental changes over the epileptogenic zone and over distant regions in ECTS. This structural remodeling likely occurs prior to the diagnosis and evolves over time, especially in patients with cognitive impairment, suggesting that the epileptogenic processes might interfere with the dynamics of the brain development and/or the normal maturation processes. Functional brain imaging demonstrates profound disorganization accentuated by interictal epileptic spikes (IES) in the epileptogenic zone and in remote networks in ECTS. Over the epileptogenic zone, the literature demonstrates changes in term of neuronal activity and synchronization, which are effective several hundred milliseconds before the IES. In the same time window, functional changes are also observed in bilateral distant networks, notably in the frontal and temporal lobes. Effective connectivity demonstrates that the epileptogenic zone constitutes the key area at the origin of IES propagation toward distant cortical regions, including frontal areas. Altogether, structural and functional network disorganizations, in terms of: (i) power spectral values, (ii) functional and effective connectivity, are likely to participate in the cognitive impairment commonly reported in children with ECTS. These results suggest a central and causal role of network disorganizations related to IES in the neuropsychological impairment described in ECTS children. SeizureIn typical ECTS, standard EEG shows centrotemporal spikes on normal background activity . IntericTypical ECTS belongs to the spectrum of rolandic epilepsy, which encompasses typical ECTS, Atypical Benign Partial Epilepsy (ABPE), Landau-Kleffner syndrome (LKS) and epileptic encephalopathy with Continuous Spikes and Waves during slow-wave Sleep (CSWS) .ABPE is defined by atypical seizures  , 24 assoTypical ECTS and rolandic epilepsy spectrum are believed to represent a continuum of different clinical phenotypes of self-limited focal epilepsy  but someThe genetic origin of ECTS was first suggested on the basis of clinical observations  comparinNormal cognition was previously considered to be a prerequisite for the diagnosis of ECTS, which is why this form of epilepsy was initially called Benign Childhood Epilepsy with Centro-Temporal Spikes (BCECTS) . However, ~15–30% of affected children show some degree of neuropsychological impairment, including disorders of language processing –55, atteThe pathophysiological mechanisms by which ECTS induces neuropsychological impairment remain unclear. The neuropsychological impairments are not related to seizure characteristics  and are Despite the large number of studies conducted since the first description of ECTS by Nayrac and Beaussart , the patThe objective of this study is to review the functional and structural disorganizations in the epileptogenic zone, but also in remote areas, that may be involved in cognitive dysfunctions in patients with typical ECTS. Before addressing these disorganizations, we will review the current state of knowledge concerning the epileptogenic zone responsible for structural and functional interactions with distant regions.The epileptogenic zone is defined by the site of onset of epileptic seizures and their primary organization  in the thickness of the cortex . These cSome studies have reported that this structural remodeling evolves over time , 119, asFunctional correlations between structural remodeling and cognitive deficits have been recently addressed , 119. A As expected, an increase in θ, α, and ß power spectra and cortical source densities (sensor space)  has beenHowever, IES can no longer be simply considered to be due to hypersynchronization of a single homogeneous population of neurons. Using time-frequency analysis, complex sequences of desynchronization-synchronization-desynchronization have been demonstrated surrounding the IES in ECTS  Figure . These sHigher relative EEG power in the θ, α, and ß bands is still observed in the centro-temporal area regardless of the presence of IES in the EEG segments . ProfounAltogether, these results indicate profound disorganization of the epileptogenic zone, not only in the time-window around the IES, but also during the period during which IES are not detected. IES are transient biomarkers of the local complex disorganization that is consubstantial to the epileptic network in ECTS. This disorganization is effective several hundred milliseconds before the IES  and consFunctional MRI and HD-EEG studies in the interictal state, regardless of the presence of IES, have demonstrated profound changes of the spectral power in distant areas to the spike onset zone , 4B. DesFunctional connectivity measurements have been used to study functional alteration of the brain networks during the interictal state.In the human brain, inhibitory and excitatory circuits interact by integrating information at local and global levels. It has been shown that the normal brain has a small-world functional topology, which can efficiently combine functionally specialized (segregated) modules with intermodular links (integrating). This type of organization reflects an optimal balance between functional integration and segregation , 146. ThA higher connection density around the epileptogenic zone, including the motor areas, the central region and the ipsilateral temporal region, has been described in all frequency bands . FunctioThe direction of information flow between the various brain regions has also been studied in ECTS children using effective connectivity analysis. Tools such as Dynamic Causal Modeling (DCM) can be used to measure the effective connectivity between selected brain regions  . This stFunctional changes in terms of power spectral values, functional and effective connectivity and small-world properties can participate in the cognitive impairments commonly reported in children with ECTS. The effect of pathological epileptic HFO on cognitive function needs also to be considered.The epileptogenic zone induces functional changes in the frontal lobes, which play a major role in the processing and execution of higher cognitive skills and behavior . The cenIn summary, the available literature demonstrates the existence of:Complex functional disorganizations in the epileptogenic zone that are accentuated in the presence of IES on scalp EEGComplex functional changes in terms of activity and synchronization in bilateral distant networks that are accentuated in the presence of IES on scalp EEGComplex functional changes in distant areas beginning around 400 ms before the IES in the same time window as complex modifications of synchronization around IES  in the epileptogenic areaComplex functional connectivity changes including small-world network impairmentImpaired structural brain maturation in children with ECTS with no specificity for the epileptogenic area.Altogether, these results suggest a central and causal role of the functional and structural disorganizations of the neuronal network that promote the IES in the neuropsychological impairment described in children with ECTS. Two non-exclusive hypotheses can be proposed to explain the pathophysiological mechanisms of the neuropsychological impairment. First, disorganizations related to IES might directly induce transient inhibition of the networks. Second, disorganizations related to IES might induce long-lasting effects on brain functioning and maturation.This hypothesis is supported by clinical observations of transient cognitive impairment (TCI). Transient cognitive impairment (TCI) , 161 is In addition to the short-lasting effects, network disorganizations related to IES may have long-term effects on developing neural networks , 171.This abnormal brain neurodevelopment could be related to reports that certain neuropsychological deficits persist despite return to a normal EEG and clinical remission , 163, suStructural and functional disorganizations not related to IES but participating in neurocognitive impairment in ECTS should not be excluded.The complexity of the interactions between the epileptogenic network and remote functional networks has been demonstrated in ECTS. Structural brain imaging suggests the presence of subtle neurodevelopmental changes over the epileptogenic zone and over distant regions in ECTS. Functional brain imaging demonstrates profound disorganization accentuated by IES in the epileptogenic zone which are effective several hundred milliseconds before the IES. In the same time window, desynchronization and changes in neuronal activity are observed in bilateral distant networks. These functional changes are associated with the alteration of global small-world properties toward a more random network; the direction of information flow between the brain regions demonstrates that the epileptogenic zone constitutes the key area at the origin of IES propagation toward distant cortical regions.Altogether, these results support the idea that disorganizations of the neuronal network related to IES may disrupt local and distant networks with a possible impact on functional and the maturational processes. If the impact of IES on cognitive impairments remains debated, these results suggest a central and causal role of network disorganizations related to IES in the neuropsychological impairment described in ECTS children. These results suggest that therapeutic suppression of IES should reduce the risk of neuropsychological impairment in children with ECTS. However, the currently available antiepileptic drugs are not suitable and, specific treatments for disorganizations of the neuronal network related to IES need to be developed.EB-P, MM, AA, and FW conceived and designed the review, read and accepted the manuscript. EB-P, MM, FW wrote the paper.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Bisphosphonates (BPs) are a family of molecules characterized by two key properties: their ability to bind strongly to bone mineral and their inhibitory effects on mature osteoclasts and thus bone resorption. Chemically two groups of BPs are recognized, non-nitrogen-containing and nitrogen-containing BPs. Non-nitrogen-containing BPs incorporate into the energy pathways of the osteoclast, resulting in disrupted cellular energy metabolism leading to cytotoxic effects and osteoclast apoptosis. Nitrogen-containing BPs primarily inhibit cholesterol biosynthesis resulting in the disruption of intracellular signaling, and other cellular processes in the osteoclast.BPs also exert a wide range of physiologic activities beyond merely the inhibition of bone resorption. Indeed, the breadth of reported activities include inhibition of cancer cell metastases, proliferation and apoptosis in vitro. In addition, the inhibition of angiogenesis, matrix metalloproteinase activity, altered cytokine and growth factor expression, and reductions in pain have been reported. In humans, clinical BP use has transformed the treatment of both post-menopausal osteoporosis and metastatic breast and prostate cancer. However, BP use has also resulted in significant adverse events including acute-phase reactions, esophagitis, gastritis, and an association with very infrequent atypical femoral fractures (AFF) and osteonecrosis of the jaw (ONJ).Despite the well-characterized health benefits of BP use in humans, little is known regarding the effects of BPs in the horse. In the equine setting, only non-nitrogen-containing BPs are FDA-approved primarily for the treatment of navicular syndrome. The focus here is to discuss the current understanding of the strengths and weaknesses of BPs in equine veterinary medicine and highlight the future utility of these potentially highly beneficial drugs. The strBPs are Food and Drug Administration (FDA)-approved and commonly used in the US and Europe for the prevention and treatment of osteoporosis as well as to treat other bone diseases such as Paget’s disease and bone metastatic disease with remarkable efficacy in humans –13. BPs In the years since the widespread approved use of tiludronate disodium and clodronate in adult horses suffering from navicular syndrome, there have been reports of additional benefits of tiludronate use including the treatment of chronic back soreness  and loweOne of the oft stated goals of BP treatment in the horse is an increase in bone mass and strength, the result of a reduction in osteoclastic bone resorption, as observed in humans, but this parameter is largely unmeasured or ignored in equine studies . AlthougIn another interesting experimental paradigm, unilateral cast immobilization of the horse forelimb was used assess the protective effect of tiludronate on immobilization-induced bone loss . ImmobilGiven the rampant BP use in the equine industry, there are only a few reports demonstrating a positive effect of either BP approved for use in horses with navicular syndrome , 16, 27 There has been much to do in the equine popular press highlighting recent human case reports and small clinical series where it has been suggested that long term bisphosphonate therapy (> 5 years) may suppress normal bone remodeling to such an extent that endogenous bone healing is decreased . The rucn = 215) or placebo (n = 206) within 14 days of the fracture. Administration of this highly potent N-containing BP did not affect fracture healing or clinical parameters [In the HORIZON recurrent fracture clinical trial  no eviderameters . Collectrameters . Indeed,rameters . This strameters .Given the growing concerns regarding treatment length and potential BP side effects, it is time for the veterinary community to push for more research and controlled trials of the use of the BPs, as well as focused and appropriate laboratory studies in the veterinary space. In addition, the incorporation of the existing human clinical data into the setting of CE as a means to advancing understanding of the utility and limitations of BP is warranted. Furthermore, studies with several second generation BPs may be required, given the distinct pharmacology and multiple subclasses of BPs that appear to act differently in mammalian assays and human clinical trials , 52.Importantly, in view of the long half-life of BPs, it is feasible that BPs may have a significant effect on bone turnover after re-dosing, beyond the 3 monthly dose regimen currently approved in the horse. It is important to conduct additional well-designed dosing studies with appropriate bone end-points, such as imaging and serum markers of bone remodeling. Such studies are important as they may discriminate between the bone and non-bone effects of BPs and relieve concerns for adverse equine skeletal effects such as those that occur in human patients when there are significant and lasting reductions in CTX-I following BP treatment. In addition, veterinarians must consider the rationale for BP treatment. Since little evidence of changes in BMD or even bone strength changes exists following BPs in the horse, perhaps the primary utility of BP use is indeed the non-bone effect? This important distinction must be investigated.The use of BP in the horse has been complicated of late with the recent public discourse regarding the off-label use of BPs in the yearling Thoroughbred industry. While the public outcry is concerned about ‘cleaning up’ potentially abnormal radiographs in young Thoroughbreds or change in fracture risk as the young Thoroughbred reach training and racing age, this is not supported by laboratory animal research. Early preclinical rodent studies of clodronate and etidronate Fig.  convinciIn humans, BPs are currently used in the treatment of pediatric bone disorders such as osteogenesis imperfecta (OI) , where aCertainly, in the setting of OI, cyclical BPs transiently reduce pain and improve function . Doses oet. al. [With regard to the apparent analgesic effects of BPs, at least in humans, the data would suggest these are more likely to be associated with N-containing BPs although little or no mechanistic understanding exists. There is a study examining the BP analgesic effect from a meta-analysis of 8595 patients enrolled in a number of BP clinical trials . Twenty-et. al.  comparedet. al. . Althouget. al. .In the horse there is currently a dearth of information regarding the effect of single and repeated doses of clodronate and tiludronate. Well-designed and appropriately powered research by non-biased researchers with germane bone parameters as outcome measures must be completed. Only with this data can horse owners and practitioners alike make informed decisions regarding the efficacy and appropriate clinical use of these potent molecules. Certainly, clients and practitioners alike require ongoing educational efforts regarding the efficacy and appropriate clinical use of these potent molecules. Following the development of a better understanding of BP effects in the horse, appropriately designed and powered placebo-controlled studies will determine to what extent beneficial BP effects on lameness are due to the inhibition of bone resorption and ascertain the details of repeat dosing in the equine setting. Such a strategy is required to ensure safer clinical use and produce a sufficient level of evidence to ensure safety.


An 11-year-old boy was admitted to the clinic with a 1-year history of fatigue and abdominal pain. In physical examination, there were multiple small, flat brown-violet pigmentations on his buccal mucosa and lips and brown spotty pigmentations on the lip mucosa, present since his birth . His motLaboratory findings were consistent with severe iron deficiency anemia and the fecal occult blood test was positive. Gastrointestinal endoscopic examination revealed two polyps of the stomach and three polyps of the jejunum that caused bleeding were removed with forceps . Histopathologic examination revealed hamartomatous polyps. The presence of brown pigmentations and multiple gastrointestinal polyps alerted us to a possible diagnosis of Peutz-Jeghers syndrome and serine/threonine kinase 11  mutation was found positive in both the patient and his mother. It is very important to conduct a thorough physical examination and to probe the family history in cases of iron deficiency anemia that is frequently encountered in children, especially in the presence of other complaints such as abdominal pain ,2. This