NAFLD impacts patient reported outcomes (PROs). Our aim was to assess the impact of NAFLD on patients’ HRQOL.National Health and Nutrition Examination Survey (NHANES) 2001–2011 data were used to identify adult patients with NAFLD . Patients with other chronic diseases  were excluded. Subjects without any of these conditions were healthy controls. HCV RNA (+) patients were HCV-controls. All patients completed NHANES HRQOL-4 questionnaire. Linear regression determined the association between NAFLD and HRQOL components adjusting for age, gender, race, and BMI.n = 9661); 3333 NAFLD (age 51 years and BMI 34 kg/m2); 346 HCV+  and 5982 healthy controls (age 48 years and BMI 26 kg/m2). The proportion of subjects rating their health as “fair” or “poor” in descending order were HCV controls (30 %) NAFLD (20 %) and healthy controls (10 %) (p < 0.001). HRQOL-4 components scores 2–4 were lowest for HCV, followed by NAFLD and then healthy controls . After adjustment for age, gender, race, and BMI, NAFLD patients were 18–20 % more likely to report days when their physical health wasn’t good or were unable to perform daily activities as a result (p < .0001).Participants with complete data were included (NAFLD causes impairment of HRQOL. As NAFLD is becoming the most important cause of CLD, its clinical and PRO impact must be assessed. Non-alcoholic fatty liver disease (NAFLD) is an important cause of chronic liver disease worldwide –5. NAFLDNAFLD is increasingly being diagnosed in patients with nonspecific symptoms with incidental elevation of aminotransferases , 8. Besin = 49,762) were excluded if they had Alcoholic Liver Disease (ALD) , other chronic diseases such as HIV, cancer, end-stage kidney disease and/or had missing data on key variables (demographic and HRQOL),  conducted between 2001 and 2011] were used. The Center for Disease Control and Prevention (CDC) collected the data by an interviewer-administered questionnaire conducted in participants’ homes by trained interviewers. Clinical data were obtained through the use of specially designed and equipped mobile examination centers. Participants (Age (years), gender, race/ethnicity , education , annual household income  were self-reported. Anthropometric measurements  were obtained by trained staff during the medical examination.Chronic Hepatitis C diagnosis was based on a positive Hepatitis C antibody  or HCV RNA as detected by polymerase chain reaction; (2) NAFLD was determined by the Fatty Liver Index (FLI). A FLI score of 60 or more in the absence of HCV, HBV and significant alcohol consumption was NAFLD; and (3) Participants without any chronic liver diseases and a FLI score ≤ 30 were considered as Healthy Controls.The following liver diseases were identified and included in the study: (1) Four components of a participants’ health-related quality of life (HRQOL-4) over a previous 30-day period were assessed during the scheduled visit. The four component questions were: Component 1 (C1): Rate your health status as poor, fair, good, very good, or excellent. ; (b) C2 Thinking about your physical health, which includes physical illness and injury, for how many days during the past 30 days was your physical health not good; (c) C3 Thinking about your mental health, which includes stress, depression, and problems with emotions, for how many days during the past 30 days was your mental health not good; and (d) C4 About how many days did poor physical or mental health keep him/her from doing your usual activities, such as self-care, work, or recreation.For analyses, participants were divided into two groups: fair/poor coded as “1” and good/very good/excellent were coded as “0” for component 1. For components C2–C4, we coded participants who reported no poor days (50 % of participants) as “0” and coded the others who reported any number of days >0 as “1”.P < .0001). Spearman rank-order correlations were estimated to examine an association between the participants’ ranking their health condition (C1) and demographic variables stratified by liver disease. SAS V9.3 was used for analyses . To account for the complex survey design from NHANES, SAS survey command with sampling weight, stratification and clustering variables were used except for Spearman’s rank-order correlation analyses.Simple logistic regression models with linearized variance estimation and weighting were used to estimate the association between self-reported quality of life questionnaire  and liver disease while adjusting for age, gender, race, BMI, smoking, education, diabetes, and heart disease. We did not adjust household income in the final model due to a high correlation with education . NAFLD patients had significantly higher body mass index (BMI) as compared to the Controls  and HCV patients . Of the NAFLD cohort, 57.8 % were male and 72.4 % were White. Furthermore, metabolic syndrome components such as hyperlipidemia, hypertension, and insulin resistance were more common in the NAFLD group, as well as history of heart disease .A total of 9661 participants with complete data were enrolled in the study (Table p < .0001). The overall HRQOL score was lowest for HCV and then NAFLD patients.Component 1 of HRQOL was concerned with overall HRQOL assessment Table , Fig. 2.p < .001). On the other hand, HCV patients were least likely to report no days of having physical health problems (52.4 %).Component 2 was mostly involved with number of days with physical health issues. In fact, the control group was more likely to report no days of having physical health problems compared to the NAFLD group .Component 3 was concerned with number of days with mental health issues. In this context, NAFLD patients reported similar mental health issues to the controls . On the other hand, HCV patients were less likely to report no days of having mental health problem, compared to controls and NAFLD patients . HCV patients had the worst score with only 64.9 % in this group reporting zero effected day (p < .0001).Component 4 was concerned with number of days that the physical and mental health issues would prevent patients from their activities. Again, the control group had significantly fewer number of days that physical or mental health kept the participants away from doing their usual activities  during last 30-days compared to the NAFLD group  and NAFLD  remained significant Table .Table 3A such as HRQOL are surrogates for a patient’s experience . AssessiAlthough not exactly clear, the reported poor physical health in NAFLD patients may be related to fatigue. Fatigue has been shown in previous studies to be a significant problem for NAFLD patients , 14–16. It is also important to note that impairment of HRQOL in NAFLD was less pronounced than those with HCV. In fact, the mental health aspect of PROs was more profoundly affected in HCV than NAFLD. This is not a surprise given the strong association of HRQOL with depression in patients with HCV –20.In conclusion, NAFLD is associated with impairment of patients’ HRQOL. Since previous studies reporting association of NAFLD with PRO impairment (12–13) were reported from the tertiary care centers, their results could have been associated with referral bias. This current study using large population database from NHANES provides additional data documenting impairment of HRQOL in patients with NAFLD. This integrated approach to understanding the clinical and PRO burden of NAFLD can provide a more comprehensive approach to treatment and management of patients with NAFLD.This study was approved by the IRB, the approval number is NHANES IRB: 12.1074.In this study, NHANES data was used, which is available online at NHANES website.


Phellinus fungi are ubiquitously present in the environment and play an important role in shaping forest ecology. Several species of Phellinus are notorious pathogens that can affect a broad variety of tree species in forest, plantation, orchard and urban habitats; however, current detection methods are overly complex and lack the sensitivity required to identify these pathogens at the species level in a timely fashion for effective infestation control. Here, we describe eight oligonucleotide microarray platforms for the simultaneous and specific detection of 17 important Phellinus species, using probes generated from the internal transcribed spacer regions unique to each species. The sensitivity, robustness and efficiency of this Phellinus microarray system was subsequently confirmed against template DNA from two key Phellinus species, as well as field samples collected from tree roots, trunks and surrounding soil. This system can provide early, specific and convenient detection of Phellinus species for forestry, arboriculture and quarantine inspection, and could potentially help to mitigate the environmental and economic impact of Phellinus‐related diseases.Polyporoid  Phellinus genus sensu lato currently comprises 154 species  assays  regions, areas of non‐coding DNA located between the small subunit and large subunit ribosomal RNA (rRNA)‐coding genes, which are removed after transcription of the rRNA cistron  and P. weirii. Tests with template DNA sequences and field samples confirmed the sensitivity and specificity of our microarray system, which was also shown to reliably detect infections in trees before any visually identifiable symptoms of disease were observed. There is strong demand now in forestry, arboriculture and phytosanitation for pathogen detection systems that can be easily mastered and readily deployed to yield early, accurate and specific results in a very short space of time. Our Phellinus microarray system fulfils most of these practical requirements, and could potentially play a part in reducing the environmental and economic impact of Phellinus infestation worldwide.Here, we report the development of a robust high‐throughput oligonucleotide microarray system capable of simultaneously screening for multiple Phellinus species, using universal primers previously described , before disease symptoms were visually identifiable, and the detection results were subsequently confirmed through conventional morphological analyses and pathogen isolation methods. We further used our microarray system to assess tree seedlings from a local plant nursery for the presence of Phellinus infestations. Our results showed that multiple seedlings of five species  sampled were all free of Phellinus infection .We further prepared field samples from trees in Taiwan with suspected or confirmed Phellinus genus s.l. can induce decay in the roots, trunk and branches of almost all woody plants, and represent a serious threat to forest ecosystems and commercial arboriculture. Among the Phellinus species, P. weirii is considered to be one of the most destructive pests in the economically important Douglas fir forests of the Pacific Northwest in North America  of this shorter probe, the hybridization temperature of our microarrays was adjusted to 48°C; this did not compromise the specificity of longer probes.Previously, microarrays have seen limited application due to the difficulty of identifying unique sequences that can be used for inter‐species differentiation  DNA amplicon was also added to the hybridization buffer. Together, these two controls can provide verification of the amplification and hybridization process within our developed assay.Multiplex PCR‐based methods have previously been used to identify rol Fig. A. The amP. weirii or P. noxius at 1 pg of starting DNA , the Taiwan Forestry Research Institute  and the USDA Forest Product Laboratory  (Table Cultures of A) Table . All strg for 2 min, after which 0.6 volumes of isopropanol was used to precipitate nucleic acids. The precipitated DNA was washed with 500 μl of wash buffer  and re‐suspended in 20 μl of distilled deionized water containing 0.1 μl of RNase A . Concentration of DNA was determined through spectrophotometry .Genomic DNA was isolated from fresh fungal cultures, field samples or herbarium specimens, using a modified version of the CTAB method that was previously described , 1 U Prime Taq DNA polymerase  and 5 μl (∼ 1 ng) of template DNA, using the GeneAmp PCR 2400 System . Polymerase chain reaction conditions were as follows: 94°C for 4 min and 35 cycles at 95°C for 30 s, 50°C for 30 s and 72°C for 1 min, with a final extension at 72°C for 7 min. Products of PCR were subjected to electrophoresis in 1.0 % (wt/vol) agarose gel and visualized by UV illumination after ethidium bromide staining.Universal fungal primers ITS1/4 , which were added to the amplification reaction. Digoxigenin labelling was performed with either 5'‐DIG‐tagged ITS 1/4 primers or DIG‐11‐dUTP . Polymerase chain reaction conditions for labelling were the same as described above, and the labelled amplicons were used as targets in subsequent microarray hybridization reactions.Phellinus species, using the AlignX function in Vector NTI , to identify polymorphic regions specific to each species. Probes were designed according to the following criteria: (i) probe length of 25–55 bp; (ii) GC% of ∼ 40%; (iii) and Tm of 55–65°C. The Gibbs free energy (ΔG) of probes was calculated, and the presence of dimers and hairpin loops was assessed in order to minimize the formation of secondary structures. Seven additional thymine bases were added to the 3'‐end of each probe to increase sensitivity , and diluted to a final concentration of 20 μM and 10 μM for spotting on positively charged PVC (polyvinylchloride) chips  or positively charged nylon membranes  respectively. Oligonucleotide probes were spotted onto PVC chips using automatic spotter, and fixed with UV Crosslinker . Alternatively, probes were spotted onto nylon membranes using EZspot arrayer , air‐dried and exposed to UV for probe immobilization.et al., isionWorks LS Analysis Software v6.5.2 (UVP).Hybridization of amplicons to probes on microarrays was performed according to a previously described protocol (Hsiao P. weirii and P. noxius template genomic DNA was quantified using a NanoDrop ND‐1000 Spectrophotometer (NanoDrop Technologies), diluted to starting DNA concentrations of 1 ng, 100 pg, 10 pg, 1 pg, 100 fg and 10 fg, and subjected to PCR amplification with biotin‐labelled or DIG‐labelled ITS1/4 primers. The PCR products were then subjected to electrophoresis on a 1.0% (w/v) agarose gel, as well as hybridization with PVC chip or nylon membrane microarrays, according to the procedures described above.To assess array sensitivity, Phellinus species in a single assay, complex samples containing template genomic DNA from 2 (P. melleoporus and P. noxius), 3  or 4  Phellinus species were prepared, labelled with biotin‐tagged primers during amplification and hybridized to a PVC chip array. In addition, field samples collected from roots, stems, soil and herbariums throughout Taiwan were similarly prepared and hybridized to PVC chip arrays. The field samples were also subjected to conventional detection involving cultivation on PDA or semi‐selective media  C. camphora; (C) G. robusta; (D) F. microcarpa; and (E) P. campanulata are depicted here.Fig. S3. Microarray analysis results of five species of tree seedlings from a local plant nursery. Probes were arranged on arrays as indicated in Fig. F. formosana; (B) C. camphora; (C) K. elegans; (D) M. champaca; and (E) A. confusa seedlings were found to be free of Phellinus infestation through microarray analysis.Click here for additional data file.


However, little is known about the social brain phenotype of 16p11.2 CNV and how this phenotype is related to the social impairments associated with CNVs at this locus. The aim of this preliminary study was to use molecular subtyping to establish the social brain phenotype of individuals with 16p11.2 CNV and how these patterns relate to typical development and ASD.Copy number variations (CNV) within the recurrent ~600 kb chromosomal locus of n = 12), deletion carriers (n = 12), individuals with idiopathic ASD (n = 8), and neurotypical controls (n = 16). Participants watched videos containing social and nonsocial motion during electroencephalogram (EEG) acquisition.We evaluated the social brain phenotype as expressed by mu attenuation in 48 children and adults characterized as duplication carriers . Both 16p11.2 CNVs.These results indicate atypical but unique patterns of mu attenuation for deletion and duplication carriers, highlighting the need to continue characterizing the social brain phenotype associated with The online version of this article (doi:10.1186/s11689-015-9118-5) contains supplementary material, which is available to authorized users. Third, the ASD group did not differentiate between conditions, t  = 1.43, p = 0.15.The primary aim of this paper was to address group differences across conditions. Figure t = 2.80, p = .0051, but not the computer-generated environment, t  = 0.45, p = 0.65. The opposite was true for the deletion carriers, in which the nonsocial context differences  were primarily driven by differences within the computer-generated environment, t  = 6.52, p < 0.0001, but not the live action environment, t  = 0.38, p = 0.70. Duplication carriers and the ASD group did not exhibit context discrimination differences between environments.The group × environment interaction was qualified by the three-way interaction between group, context, and environment. This interaction indicated that the effect of more social mu attenuation than nonsocial for the typical group was primarily driven by differences within the live action environment, Results for model 2 are reported in Table t  = 0.36, p = 0.72. This suggests that model 1 indicated that differences in mu attenuation for certain environments in the typical and deletion carrier groups may resolve over the course of the experiment by epoch 30, such that there is no longer a difference in mu attenuation by environment.Main effects of condition and group estimated at epoch 30 were similar to the overall effects of model 1 with one exception. Unlike model 1, which predicted that more mu attenuation for the social live action condition, model 2 predicted no difference between live action and computer-generated environments, A main effect of time indicated that for subsequent epochs, the mu attenuation intercept  was expected to decrease or lessen. However, as described by the significant group × time interaction, the rate of mu attenuation lessening differed by group. The first column in Fig. p = .071, see Table Although the time × group × context interaction was marginally significant  = 2.01–4.22, ps < 0.05] until epoch 60, t  = 1.73, p = 0.083. However, the duplication carriers did not begin discriminating contexts until epoch 30 and continued to exhibit more mu attenuation for nonsocial conditions until the end of the video, ts  = 2.09–2.92, ps < 0.05. In contrast to the carrier groups, the typical group exhibited greater mu attenuation for social conditions at epoch 25, ts  = 2.0–2.6, p < 0.05. Unlike all other groups, the ASD group did not exhibit mu attenuation context discrimination at any epoch, indicating equivalent levels of mu attenuation for social and nonsocial conditions.To fulfill our objective of characterizing dynamic changes in mu attenuation related to social perception, we conducted a series of pairwise comparisons between social and nonsocial context for each group every 5 epochs from epoch 1 to epoch 60. Figure 16p11.2 carriers between (1) children and adults and (2) between 16p11.2 carriers with and without an ASD diagnosis. While these comparisons may be subjected to type I and II errors due to small sample size, these analyses were meant to qualitatively support model 1 and model 2 predictions. The patterns of mu attenuation for these comparisons are depicted in Additional file To better understand how these patterns may be related to heterogeneity within these unique samples, we conducted a series of post hoc ANOVAs to determine if patterns of mu attenuation are consistent for n = 9, F  = 16.28, p < .001] and adult duplication carriers  exhibited the overall group pattern exhibited with greater mu attenuation for nonsocial  conditions. There was no difference between context for adult deletion carriers  or child duplication carriers . However, visual inspection of the dynamic mu attenuation patterns across time .First, child deletion carriers [n = 3) nor duplication carriers (n = 3) with an ASD diagnosis exhibited context discrimination, Fs < .31, ps > .58, similar to the pattern of mu attenuation for the ASD  group. Visual inspection of the dynamic mu attenuation patterns across time , mid-experiment increase for nonsocial (16p11.2 duplication), and no motion discrimination (ASD). The duplication carriers and the typical group exhibited proportionally equivalent mu attenuation during the initial portion of social and nonsocial videos. Subsequently, after 25–30 epochs, the modulation of mu attenuation diverged for these two groups, such that the duplication carriers followed an atypical trajectory with greater mu attenuation for nonsocial conditions. It may be that while social stimuli are initially similarly salient for duplication carriers and typical individuals, the maintenance of that salience at the neurophysiological level differs, resulting in a rapid decrease in neurophysiological processes devoted to social information processing.Second, the single-trial analysis indicated that the relative patterns of mu attenuation between social and nonsocial motion vary differently for each group over time. Specifically, these patterns indicated three unique atypical patterns of mu attenuation: overall increase for nonsocial . While both CNV groups exhibit greater mu attenuation for nonsocial relative to social motion, this overall pattern of discrimination occurs during the later exposure trials for the duplication carriers. This finding reflects different temporal processing trajectories, which may be driven by unique underlying neural systems for integrating social information. One possible explanation may be that the CNV groups differ in level of motivation to engage with social stimuli. Considering the rapid rate of mu attenuation reduction during the observation of social motion for the deletion group , more aligned with the individuals with idiopathic ASD. In other words, the functional social brain phenotype associated with ASD supersedes the unique 16p11.2 deletion and 16p11.2 duplication phenotype. Each unique atypical pattern may correspond to different aspects of social cognitive impairments. For instance, individuals with ASD do not exhibit social and nonsocial motion discrimination, supporting theories of complex information dysfunction [16p11.2 CNVs and ASD, it is likely that concomitant features of social cognition  more closely resemble ASD impairments [However, within this preliminary study, individuals with function , 52. In airments –56.16p11.2 CNV and neurodevelopmental disorders, a developmental model is critical. Prior work indicates a diagnosis-independent emergence of mu attenuation across development [Given the relationship between elopment , but botelopment , 58. In 16p11.2 CNV patterns are fairly consistent across development, despite the large age range. Considering the rarity of this group of individuals, the results are informative as a preliminary investigation of the 16p11.2 CNV social brain phenotype. As genetic testing becomes more commonplace, the percentage of known individuals with 16p11.2 CNVs will increase. While investigating this CNV and other rare variants is a long-term process, these results justify continuing to use neuroimaging in order to elucidate the underlying pathogenic mechanisms involved with unique molecular subtypes.The three primary strengths of this study include the use of careful molecular subtyping as a “genetics-first” approach , neurode16p11.2 carriers exhibit atypical social perception, similar to, yet distinct from, ASD. Future research should continue to address clinical and behavioral heterogeneity by reducing variance through large, carefully matched populations. The process by which these patterns emerge in relation to developmental age may be of particular interest, as well as how mu attenuation is impacted by clinical interventions promoting social development. Additional considerations for individual differences would benefit from the inclusion of robust behavioral measures of social cognition previously linked to spectral power in the mu rhythm, such as imitative ability [In conclusion, our findings strongly support the notion that  ability .


Escherichia coli infections) should have renal imaging after a first infection’. We surveyed all microbiology laboratories in England with Clinical Pathology Accreditation to determine standard operating procedures (SOPs) for urgent microscopy, culture and reporting of children's urine and to ascertain whether the SOPs facilitate compliance with NICE guidance. We undertook a computer search in six microbiology laboratories in south-west England to determine urine submissions and urine reports in children under 3 years. Seventy-three per cent of laboratories (110/150) participated. Enterobacteriaceae that were not E. coli were reported only as coliforms (rather than non-E. coli coliforms) by 61 % (67/110) of laboratories. Eighty-eight per cent of laboratories (97/110) provided urgent microscopy for hospital and 54 % for general practice (GP) paediatric urines; 61 % of laboratories  cultured 1 μl volume of urine, which equates to one colony if the bacterial load is 106 c.f.u. l− 1. Only 22 % (24/110) of laboratories reported non-E. coli coliforms and provided urgent microscopy for both hospital and GP childhood urines; only three laboratories also cultured a 5 μl volume of urine. Only one of six laboratories in our submission audit had a significant increase in urine submissions and urines reported from children less than 3 years old between the predicted pre-2007 level in the absence of guidance and the 2008 level following publication of the NICE guidance. Less than a quarter of laboratories were providing a service that would allow clinicians to fully comply with the first line recommendations in the 2007 NICE UTI in children guidance. Laboratory urine submission report figures suggest that the guidance has not led to an increase in diagnosis of UTI in children under 3 years old.The National Institute of Care Excellence (NICE) 2007 guidance CG54, on urinary tract infection (UTI) in children, states that clinicians should use urgent microscopy and culture as the preferred method for diagnosing UTI in the hospital setting for severe illness in children under 3 years old and from the GP setting in children under 3 years old with intermediate risk of severe illness. NICE also recommends that all ‘infants and children with atypical UTI (including non- Urine testing remains important to obtain a definitive diagnosis in the management of urinary tract infection (UTI), and culture and susceptibility are needed to guide antibiotic treatment. This is just as important in UTI in children as in adults. The National Institute of Health and Care Excellence (NICE) 2007 guidance CG54, on ‘Urinary tract infection (UTI) in children: diagnosis, treatment and long-term management’, states that clinicians should refer all infants under 3 months old with suspected UTI to paediatric specialist care, and a urine sample should be sent from all children with suspected UTI for urgent microscopy and culture  NICE, 2  with a single organism should be considered a possible UTI, and >107 c.f.u. l− 1 (10 000 c.f.u. ml− 1) with a predominant organism in a mixed growth of two organisms should be considered a probable UTI, in the presence of symptoms and white blood cells. One of the other recommendations of NICE that has implications for laboratory reporting is ‘Infants and children with atypical UTI [this includes infection with non-Escherichia coli bacteria  standards for microbiology investigation state ‘Generally, a pure growth of between 10bacteria   audit the extent of the availability of urgent urine microscopy for children in hospital and in primary care, 2) audit what methods are used to culture children's urine, 3) audit how children's urine culture results are routinely reported and 4) determine, in a subset of laboratories, whether NICE guidance had led to a change in numbers of urine specimens from children under three years being submitted and/or reported. This audit scope allowed us to determine whether laboratories facilitate compliance with NICE recommendations. The audit formed part of a Healthcare Quality Improvement Partnership (HQIP) multi-centre audit (HQIP NCA 075)  that focused on three key clinical themes of the childhood UTI NICE guidance; improving the rate of diagnosis, careful clinical evaluation to identify ‘high risk’ children and selective use of follow-up investigations.E. coli bacteria.The audit questionnaire (Appendix 1) was developed by clinical microbiologists and a consultant paediatrician and piloted several times with three laboratories for ease of completion and feasibility of data entry. The 14 questions were phrased to enable us to determine whether the microscopy, culture and antibiotic susceptibility service laboratories provided met the recommendations within the NICE UTI guidance  and the To determine urine submission rates, laboratories in south-west England were asked to provide the number of all urine specimens submitted to them between 2003 and 2011 from children under 3 years of age  and the number of positive urines in this age group in whom antibiotic susceptibility results were reported. Laboratories were asked to report any changes in laboratory methodology or reporting protocols.E. coli UTI to be identified and then investigated further by clinicians. We asked that a biomedical scientist or consultant medical microbiologist who was responsible for childhood urine culture and susceptibility reports complete the questionnaire. An email reminder was sent in June 2011. As the initial response rate was under 50 %, a personalised letter was sent to all microbiology consultants in August 2011. In October the remaining 35 non-responding laboratories were phoned to confirm that they processed childhood urine specimens and therefore should be included in the non-responders.The audit questionnaire was emailed with an explanatory letter to the managers of all 164 microbiology laboratories in England with Clinical Pathology Accreditation (CPA) in May 2011. The letter and questionnaire both stated that the aim of the HQIP audit was to determine whether laboratories identify and report organisms, in line with the 2007 NICE guidance, allowing children with non-The number of responses and proportion of different responses to each question were tabulated. Data were cleaned by one of the research team, and where laboratories had ticked more than one box for an answer, comments given by participants were used to determine which answer was recorded. The 95 % confidence intervals (CI) were obtained by using the binomial distribution, with the number of responses as parameters thereto.P values for significance of change in trend and change in level were obtained. All analyses were performed in stata version 13.0  of the outcome between what it would have been in 2008 without guidance and its prediction in 2008 as a result of the guidance, and change in the trend from 2008 onwards, respectively. Since there were only three observations for laboratory 4 in the post-guidance period, the only covariate in the model was a variable equal to the year less 2008, so that year numbering began from 1. Statistical significance was determined by means of the likelihood ratio test. For laboratory 4, the significance of the trend was determined, whereas for all the other laboratories the ion 13.0 .As this was an audit it did not require ethical approval but it was undertaken in line with information governance and Caldicott principles.The questionnaire was sent to 164 laboratories; 14 of these did not process urines, five declined to take part due to lack of time, and we contacted 30 of the 35 non-responding laboratories, all of which confirmed that they did routinely perform urine cultures from children. Thus, 73 % (110/150) of laboratories undertaking childhood urine culture returned completed questionnaires. Non-responding laboratories were not atypical; they were from across England serving both rural and urban communities and included both private and NHS laboratories. Seventy-five per cent of laboratories (83/110) returned data on the number of urines submitted from children under 16 years in 2010 and population served. There was a wide range of urine submissions from children to laboratories from 438 to 36 400; this equated to a range of 1 to 161/1000 total population. Six laboratories in the South West returned detailed data on laboratory submissions.The majority of the laboratories  undertook urgent microscopy for hospitalized children; 18 % only during office hours, 25 % during office and extended hours and 57 % during all hours. Only 54 % of laboratories (59) also processed urgent specimens from children in general practice.6 c.f.u. l− 1 (>1000 c.f.u. ml− 1) with a single species may be diagnostic of UTI, or >107 c.f.u. l− 1 (10 000 c.f.u. ml− 1) predominant organism in a mixed growth of two isolates should be considered a possible UTI, in the presence of symptoms and significant numbers of white blood cells . Sixty-one per cent of laboratories  reported that they cultured 1 μl of urine from children, 10 % 2–3 μl, 5 % 5 μl, 10 % 10 μl, 5 % 20–100 μl, 7 % used a filter paper method and 2 % used an automated method. Fifty-nine per cent of laboratories  reported 107 c.f.u. l− 1 (10 000 c.f.u. ml− 1) as significant, 19 % (21/110) reported 106 c.f.u. l− 1 (>1000 c.f.u. ml− 1) as significant and 15 % (16/110) reported 108 c.f.u. l− 1 (105 c.f.u. ml− 1) as significant. Eighteen per cent of laboratories (20/110) specifically commented that they would only report the lower colony counts if in pure growth or accompanied by significant white cells.PHE operating standards state that 106 c.f.u. l− 1 (1000 c.f.u. ml− 1), 35 % (39/110) reported any growth or 100 c.f.u. ml− 1 (105 c.f.u. l− 1) and 20 % (22/110) 107 c.f.u. l− 1 (10 000 c.f.u. ml− 1). Only one laboratory reported 108 c.f.u. l− 1 (105 c.f.u. ml− 1).Laboratories reported much lower counts from suprapubic aspirates; 42 %  reported 108 c.f.u. l− 1 (105 c.f.u. ml− 1) if there was a single predominant organism over 107 c.f.u. l− 1 (10 000 c.f.u. ml− 1); another 7 % (8/110) reported these if the urine was a repeat sample, if there were significant numbers of white blood cells or if clinically indicated.Seventy-one per cent of laboratories  reported a mixed growth of 10E. coli from other non-E. coli coliforms (E. coli from other lactose-fermenting coliforms (E. coli was reported by its species name by 83 % (91/110) of laboratories; 17 % laboratories (19/110) only reported it as a coliform. Enterobacteriaceae that were not E. coli  were reported only as coliforms (rather than non-E. coli coliforms) by 61 % (67/109) of laboratories (seven of these reported Proteus spp. when present); 37 % (40/109) reported to genus or species level dependent on the organism and 1.8 % (2/109) reported these as coliforms other than E. coli. Eighty-five per cent of laboratories (17/20) who used CLED routinely reported Enterobacteriaceae that were not E. coli as coliforms, compared with 50 % of laboratories (43/86) who used chromogenic agar for primary culture.For the routine identification of uropathogens, 81 % of laboratories (89/110) used chromogenic agar, which allows the easy differentiation of E. coli coliform, most reported to species level  or genus level (8.7 %); only 9 % (10/110) of laboratories reported these as coliforms. When a non-E. coli was not an Enterobacteriaceae, all laboratories reported these to genus or species level. Two (1.8 %) laboratories routinely added a standardized comment to the urine report if the organism isolated was a non-E. coli indicating that the child may need further investigation; one other laboratory (0.9 %) linked all reports to local UTI guidance.When reporting a multi-resistant non-Six laboratories in the South West returned urine submission data; laboratories 2, 3 and 4 were only able to collect data from 2006, 2007 and 2009, respectively.P value = 0.005; 48.5 % increase in positive reports, 95 % CI 27.6–72.3 %, P = 0.004). Laboratory 1 had no increase in submissions, but had a significant increase in positives reported; this increase occurred after 2010 when the laboratory indicated that they had a change in reporting policy and started reporting antibiotic susceptibility for organisms within a mixed growth if there was a predominant organism greater than 107 c.f.u. l− 1 (104 c.f.u. ml− 1). Laboratory 3 had no significant increase in submission or an increasing trend, but the level of positives increased significantly after the guidance was published ; however, there was no change in testing or reporting policy. Laboratories 2 and 5 had no significant increases; furthermore, laboratory 5 had a significant decreasing trend in submissions .Only laboratory 6 showed a significant increase in submission of urine specimens from children under 3 years and a significant increase in urines reported with antibiotic susceptibilities between the pre-2007 level in the absence of guidance and the 2008 level following guidance ; there was no change in testing or reporting policy.Laboratory 4, with 3 years of data from 2009 to 2011, demonstrated a significant increase in specimen submission ; however, the number of positives reported with antibiotic susceptibility results decreased non-significantly by 6.7 % (8 c.f.u. l− 1 (105 c.f.u. ml− 1); 61 % (67/109) are not fully identifying most non-E. coli coliforms and, therefore, are not giving enough information on reports for clinicians to be able to refer children in whom an Enterobacteriaceae other than E. coli has been isolated. Only 22 % (24/110) of laboratories comply with both the recommendations for urgent microscopy for all children and clearly differentiating non-E. coli coliforms; only 2.7 % (3/110) of laboratories also cultured a 5 μl volume of urine allowing them to diagnose an infection with 106 c.f.u. l− 1 (>1000 c.f.u. ml− 1) with any confidence. Our audit of laboratory submissions showed that for five of the six laboratories the guidance has not significantly increased urine submissions or diagnosis of UTIs in children less than three years.Forty-six per cent of laboratories are not undertaking urgent microscopy in general practice paediatric urines and 61 % are unable to reliably diagnose a UTI with a colony count of under 10E. coli received appropriate imaging tests. Furthermore, we made no attempt to identify the quality of the sample received and the quality of the request information provided which would have also required a more in-depth audit; these two factors may have a significant effect on the reliability and interpretation of results. Such in-depth audits would be very difficult to undertake on a large number of laboratories. The urine submission audit was only undertaken in the South West and due to computer changes, only three laboratories were able to provide submission data for more than one year pre-guidance, making interpretation more difficult. This reflects ongoing problems of using routinely generated data to monitor the effect of guidance on laboratory practice and the difficulties of determining whether NICE guidance has increased the numbers of childhood UTIs diagnosed.The questionnaire survey had a very good return and, therefore, probably reflects current laboratory practice across English laboratories. The participants reported standard operating procedures; however, this cannot replace a prospective audit of clinical cases and their outcomes, including whether children with atypical UTIs caused by non-NICE guidance indicates that urgent microscopy is the preferred diagnostic method in children under 3 years, as the diagnostic performance of urine dipstick nitrite and leukocyte esterase tests, in particular the likelihood ratios, is poor (NICE 2007). In all children under 3 months and those under 3 years with risk of serious illness, referral to the hospital setting is advised. However, NICE indicates that some infants or children with intermediate risk of serious disease may be managed in primary care where urgent microscopy is preferable. Although they suggest that urine dipstick can be used in children with less serious illness, elsewhere in the guidance they state ‘that a combination of a positive leucocyte esterase with positive nitrite has the highest LR+ and is the most useful dipstick test for ruling in UTI. However, a negative result for either leucocyte esterase or nitrite has the highest LR −  and will be most useful in excluding UTI. It is important to note that in children younger than 2 years the dipsticks are less reliable in both scenarios’. . Thus, ideally we recommend that one would have a system where there is widespread access to urgent microscopy by Primary Care, given the unreliability of dipsticks in the younger children.Improved access to urgent microscopy by Primary Care may be a challenge with the ongoing reorganization of laboratory services moving to larger, more centralised laboratories well away from the point of care. Any service contracts with microbiology laboratories need to ensure that microscopy provision is available in some way at the point of care. The current relative lack of access to urgent microscopy services may delay appropriate treatment and, due to the unreliability of urine dipticks in children under 3 years, may result in general practitioners having to refer these children in whom they suspect a UTI to hospital; this may be neither cost effective nor appropriate for the child and family. Alternatively, the general practitioner may treat the patient based on symptoms and/or dipstick alone, which is less than ideal.8 c.f.u. l− 1 (105 c.f.u. ml− 1) (8 c.f.u. l− 1 (105 c.f.u. ml− 1) is generally indicative of UTI; a pure or predominant growth of an organism at over 107 c.f.u. l− 1 (104 c.f.u. ml− 1) may be considered significant if there is pyuria and the child is symptomatic . Both sets of guidance indicate that colony counts of 106 c.f.u. l− 1 (>1000 c.f.u. ml− 1) of a single species may be diagnostic of UTI in voided urine (mid-stream or clean-catch urine rather than a nappy urine) in the presence of symptoms. The PHE standard methods state that if a 1 μl or 2 μl inoculum or filter paper method is used, the method is only sensitive for screening down to 107 c.f.u. l− 1 (104 c.f.u. ml− 1) ; only 20 % of the laboratories surveyed who culture 5 μl or more would be able to diagnose an infection giving rise to 106 c.f.u. l− 1 (>1000 c.f.u. ml− 1) with any confidence. The recent Diagnosis of Urinary Tract Infection in Young Children (DUTY) study (6 c.f.u. l− 1 (>1000 c.f.u. ml− 1) and 107 c.f.u. l− 1 (104 c.f.u. ml− 1) were also considered to have UTI, although there were more in this group with another confirmed cause for their illness. If children have urine with lower counts the laboratory should probably request a repeat specimen, as the probability of UTI is increased by the isolation of the same organism from two specimens (6 c.f.u. l− 1 (>1000 c.f.u. ml− 1) as significant from these specimens. The PHE standards do not give specific recommendations for significance of counts from suprapubic aspirates but do state that ‘(SPA) is seen as the “gold standard” but is usually reserved for clarification of equivocal results from voided urine in infants and small children’ .The NICE guidance defines UTI by a combination of clinical features and the presence of bacteria in the urine. It indicates that typically a UTI is caused by a single organism, which is present in a high concentration, usually greater than 10. ml− 1) , but thaE. coli coliform or any other type of bacteria, there is an increased risk of serious underlying pathology. NICE guidance recommends that infants, children and young people (under 16 years) with atypical urinary tract infection (which includes infection with non-E. coli organisms) should have ultrasound of the urinary tract during the acute infection. It is therefore important that laboratory test reports differentiate between E. coli and non-E. coli organisms to identify whether further investigations are needed’ . Although 83 % of laboratories reported E. coli itself by name, we found the laboratories in our audit had an inconsistent approach to reporting of non-E. coli organisms in urines. Although non-Enterobacteriaceae were often reported to genus or species level, non-E. coli coliforms were frequently not identified and were reported as coliforms by 61 % of laboratories. This was a key finding of our study. Laboratories who did not use chromogenic agar were more likely not to report non-E. coli coliforms just as coliforms (75 %), although 50 % of laboratories who used chromogenic agar similarly reported these as coliforms. As children with non-E. coli coliforms are more likely to have unrecognized renal abnormalities these may go unrecognized if clinicians do not realize they should perform the renal imaging recommended for these patients by the NICE 2007 guidance , 73 % of children with recurrent UTI and 77 % with severe reflux  had a 3.4-fold relative risk of significant urinary tract abnormalities and thus obstructive uropathy . In line with childhood UTI NICE guidance and the quality standards , it would also be best practice for laboratories to indicate with a comment in the report to clinicians the need to consider renal imaging in children with non-E. coli UTI. In our audit only two laboratories added any such routine comment to reports of non-E. coli isolates.The 2013 NICE Urinary tract infection in infants, children and young people quality standard states that: ‘If a urinary tract infection is caused by a non-7 c.f.u. l− 1 (104 c.f.u. ml− 1). In the other case it is possible to speculate that urines for culture were submitted from children with a higher pre-test likelihood of UTI.Previous work has shown a wide variation in urine specimen submission to laboratories . One posThere are two major obstacles in the diagnosis of UTI in this age group: UTI symptoms in this age group overlap with symptoms of much more common viral infections and it is an onerous task to obtain urine samples from children with no bladder control. The DUTY study, which has enrolled 7000 children and has detailed urine results for over 5000, may be able to inform a more robust algorithm than currently in the NICE guidance to help clinicians identify a set of clinical criteria that if present increase the likelihood of UTI, thereby focusing efforts in obtaining urine samples to this group of children (E. coli coliform UTI need to be identified (NICE 2007) and this is now a NICE standard  the national microbiology standard method 41  should be changed to include this recommendation. Laboratories should review their laboratory Standard Microbiology Investigations and reporting for UTI and ensure it is in line with both NICE guidelines and the national standards. Laboratories could encourage referral of children with non-E. coli UTI for imaging by adding a routine comment to reports of non-E. coli isolates. NICE guidance needs more rigorous implementation through all relevant professional societies.Laboratory microscopy should be made available to support a reliable and rapid diagnosis of UTI for young children in both the primary and secondary care setting. Since NICE considers that children with non-


The group with a change in proteinuria ≤0.30 g/g (n = 1261) had lower cumulative probabilities of renal death (p < 0.001). In a linear regression model, a higher baseline proteinuria and a greater increase in proteinuria were associated with faster annual GFR decline. Cox’s analysis showed that every 1 unit increase in natural log and every 0.1 g/g increase in the change in proteinuria resulted in 67%  and 1%  greater risk of renal death respectively after adjusting for the effects of the other covariates. Our study provided a patient-based evidence to support proteinuria as a therapeutic target in advanced CKD.Current evidence of proteinuria reduction as a surrogate target in advanced chronic kidney disease (CKD) is incomplete due to lack of patient-pooled database. We retrospectively studied a multicenter cohort of 1891 patients who were enrolled in the nationwide multidisciplinary pre-end stage renal disease care program with a baseline glomerular filtration rate (GFR) <45 mL/min/1.73 m The rapidly increasing incidence and prevalence of chronic kidney disease (CKD) have posed serious problems for global public health1Since 2002, the publication of a clinical practice guideline on CKD48Proteinuria is an independent risk factor for progressive kidney function decline as well as all-cause mortality101112141516171820222326272 per year. The patients with diabetic nephropathy had higher baseline UPCR compared to non-diabetic patients (2.92 ± 2.57 vs. 1.05 ± 1.48 g/g). By using generalized additive models (GAM), patients were stratified into a high proteinuria group (baseline UPCR >1.04 g/g) and a low proteinuria group (baseline UPCR ≤1.04 g/g) (The overview of cohort formation was shown in .04 g/g) . High prA multiple linear regression analysis was applied for the independent determinants of annual GFR change . PolycysTo investigate the difference in clinical characteristics of patients with CKD with different changes in UPCR, we divided all patients into two groups by ΔUPCR of 0.30 g/g after GAM analysis . There wIn total, 367 participants (19.6%) suffered from renal death in a mean follow-up time of 32.0 ± 12.3 months. A Nelson-Aalen analysis revealed that the cumulative probabilities regarding renal death, mortality, and dialysis were lower in the low proteinuria group than those in the high proteinuria group . In anot2, and albumin levels of 3.2–4.4 g/dL also contributed to increments in odds ratio of renal death. Only a few participants had phosphate levels >7.2 mg/dL (n = 7), BMI >36.4 kg/m2 (n = 40), or albumin levels <3.2 g/dL (n = 32).A Cox’s proportional hazards ratio analysis was used to determine the independent risk factors for renal death . Every 1In this study of an advanced CKD population, we demonstrated that high baseline proteinuria (UPCR >1.04 g/g) was associated with rapid GFR decline and also predicted renal death. Additionally, less increase of proteinuria (ΔUPCR ≤0.30 g/g) was significantly associated with slower renal function decline and also predicted a lower probability of dialysis and mortality.The Modification of Diet in Renal Disease (MDRD) study showed that high proteinuria >1.0 g/day was a predictor of renal progression292 showed a significant correlation between the extent of proteinuria reduction and GFR decline192134Proteinuria reduction and residual proteinuria are predictors of renal disease progression and ESRD18233334262836Comparing to baseline proteinuria, the effects of changes in proteinuria were smaller on both GFR decline and renal death. Participants with higher baseline proteinuria reflected more severe degree of nephron dysfunction and damage3519≤0.3 g/g group, which might inferred the effectiveness of multidiscipline care in controlling proteinuria. Besides, one study showed that still a substantial number of participants with diabetes and nephropathy treated with losartan had no reduction in proteinuria262836In contrast to previous literature4244The strength of our study is that our participants received standard multidisciplinary care based on the NKF KDOQI guidelines instead of specific drug therapy, as proposed by previous literatureThere were several limitations in our study. First, due to retrospective study design, we were unable to adjust specific interventions and pharmacotherapy other than RAS blockade or to clarify the characteristics that lead to different degrees of proteinuria reduction under same treatment strategy. Second, our study population comprised exclusively of Taiwanese with advanced CKD who were referred to multi-discipline care. For examples, patients with more rapid progression who are unable to collect subsequent proteinuria were excluded. Selection bias might exist. Third, the MDRD equation was developed based on younger Caucasian subjects (aged 50.6 ± 12.7 years)465146In summary, this multicenter retrospective study including participants with advanced CKD under multidisciplinary care in nephrology clinics showed that high proteinuria was associated with rapid GFR decline and also predicted renal death. The cut-off level of UPCR 1.04 g/g could be used in clinical practice to classify a high-risk group. Additionally, proteinuria reduction was significantly associated with slower renal function decline and also predicted risk of dialysis. To aim at improving renal outcome, the clinician can adjust the treatment policy to reduce proteinuria as much as possible. Our study supported the concept that proteinuria is a therapeutic target in patients with advanced CKD.−1.154 × Age −0.203 × . We selected patients aged above 18 years with a baseline GFR <45 mL/min/per 1.73 m2 and at least two measurements of serum creatinine. We further excluded patients with less than two measurements of urine protein excretion . Patients with the interval of separate proteinuria measurements less than 9 months were also excluded. The severity of CKD was then staged based on the NKF KDOQI clinical practice guidelinesThis is a retrospective cohort study conducted in six collaborative hospitals in Taiwan from 2008 to 2011. Patients who joined the nationwide pre-ESRD care program were enrolled. The GFR was estimated by using the MDRD study equation2; or serum creatinine level ≥6 mg/dL or GFR ≤15 mL/min/per 1.73 m2 with the presence of one or more uremic symptoms that threaten life or impair quality of life8The pre-ESRD care program in Taiwan included standardized interventions and multidisciplinary care that followed the NKF KDOQI guidelines4Demographics collected through medical records and registry data in each hospital included age; sex; employment status; education; smoking habits; drinking and betel nut use; original cause of renal failure; BMI; MAP; biochemical measurements including hemoglobulin, serum albumin, calcium, phosphate, and UPCR; comorbidity history; and antihypertensive drugs. Diabetic nephropathy was defined as UPCR ≥0.5 g/g in patients with diabetes mellitus without any other cause of renal failure.2 per year. The change in covariates  was defined as the value in follow-up minus baseline value of each parameter. A repeated-measures analysis was not used to assess changes during follow-up because the times of follow-up measurements varied across the patients.Two primary outcomes were selected from our analysis. First, renal death was defined as the composite of initiation of dialysis and mortality . Second, annual GFR change was estimated as the slope of a linear model including two GFR measurements, determined in mL/min/per 1.73 mp value ≤ 0.05 was considered statistically significant. The distributional properties of continuous variables were expressed as mean , categorical variables were presented by frequency and percentage, and the cumulative hazard rates of survival outcomes were estimated by the Nelson-Aalen method.A statistical analysis was performed using the IBM SPSS Statistics 21 software  and the R 3.0.2 software . In statistical testing, a two-sided GAMs56t test, Wilcoxon rank-sum test, chi-square test, and Fisher’s exact test  were used to examine the differences in the distributions of continuous variables and categorical variables within groups stratified by UPCR as well as ΔUPCR. Next, a multivariate analysis was conducted by fitting the linear regression model and Cox’s proportional hazards model to estimate the effects of risk factors, prognostic factors, or predictors on annual GFR change, occurrence of renal death, and time to renal death, respectively.In univariate analysis, the two-sample How to cite this article: Chen, C.-H. et al. Proteinuria as a Therapeutic Target in Advanced Chronic Kidney Disease: a Retrospective Multicenter Cohort Study. Sci. Rep.6, 26539; doi: 10.1038/srep26539 (2016).


Changes in coastal upwelling strength have been widely studied since 1990 when Bakunproposed that global warming can induce the intensification of upwelling in coastalareas. Whether present wind trends support this hypothesis remains controversial, asresults of previous studies seem to depend on the study area, the length of the timeseries, the season, and even the database used. In this study, temporal and spatialtrends in the coastal upwelling regime worldwide were investigated during upwellingseasons from 1982 to 2010 using a single wind database  with high spatial resolution (0.3°). Of the major upwellingsystems, increasing trends were only observed in the coastal areas of Benguela,Peru, Canary, and northern California. A tendency for an increase inupwelling-favourable winds was also identified along several less studied regions,such as the western Australian and southern Caribbean coasts. Wind-driven coastal upwelling results from the action of winds along a coast thatgenerate an Ekman drift directed offshore. This causes pumping of cool and nutrient-richwater towards the sea surface along a narrow region close to the coast, which enhancesprimary production. Thus, coastal upwelling systems are among the most productive marineregions in the world’s oceans.Due to the ecological and economic importance of these regions, changes in upwellingstrength and timing have attracted considerable scientific interest in recent decades.In several studies, researchers have primarily focussed on analysing trends in windstrength due to climate variability and the resultant changes in the coastal upwellingprocess. In 1990, Bakunet al.Studies focused on different upwelling regions have been conducted to investigate theBakun hypothesis through an analysis of available wind data. However, these studiesreported contradictory results, which indicate that wind estimates from differentdatabases can differ in trends and variabilityThe aim of this study was to identify the temporal and spatial trends in coastalupwelling regimes worldwide using wind stress data from the National Centers forEnvironmental Prediction (NCEP) Climate Forecast System Reanalysis (CFSR)http://rda.ucar.edu/pub/cfsr.html developed by the National Oceanicand Atmospheric Administration (NOAA). Data were retrieved from the NOAA NationalOperational Model Archive and Distribution System, which is maintained by the NOAANational Climatic Data Center. Detailed information about the CFSR database can befound in Saha et al.Wind data were acquired from the NCEP CFSR database at The process used to identify upwelling areas and to calculate trends is summarizedbelow:Wind data were initially averaged at a daily scale to smooth the effect ofhigh frequency events such as breezes. In addition, upwelling eventstypically last 3–14 daysabs means absolute value. Alongshore wind stress has beenpreviously used to estimate variations in upwelling intensity16Alongshore wind stress –1, whichcorresponds to the transition from a gentle to a moderate breeze on theBeaufort scale. García-Reyes et al.–1 following Cury andRoy–1. This condition must befulfilled for at least three consecutive months. Second, the area isconsidered to be an upwelling region only if at least 10 consecutive points(about three degrees) fulfil the previous condition. This second conditiondiscards the appearance of small local areas.Trends were calculated using only the months under strong upwellingconditions. Those months were selected from the climatology data consideringTrends were calculated at each pixel as the slope of the linear regressionof the alongshore monthly wind stress anomalies versus time. Monthlyanomalies were calculated by subtracting from the alongshore wind stress ofa certain month . This database facilitates the analysis of wind behaviour at asmall scale, which is a key factor when considering coastal mesoscale effects asupwelling. Recent studies have compared this database and different wind productswith wind measured by several buoys along the Iberian Peninsula coast1112In this study, 3025 coastal points worldwide were analysed to select the majorupwelling regions and evaluate upwelling trends from 1982 to 2010. Under our termsof selection, ten upwelling systems were evaluated: Benguela, Canary, the southernCaribbean Sea, Chile, Peru, California (north and south), West Australia, Java,North Kenya, and Somalia–Oman. These systems were grouped together intoseveral macroscopic zones, namely the eastern and western coasts of the Atlantic,Pacific, and Indian Oceans. No upwelling systems along the western Pacific Oceanwere assessed. Alongshore wind stress was calculated over more than 600 points alongthis coast, and no region fulfilled the conditions required to be considered acoastal upwelling area.Two upwelling systems were analysed along this coast from south to north:Benguela  and Canary(18.89–32.63°N) . Alongt−3 Nm–2 dec–1in the northern area to −3 Nm–2 dec–1in the southern one.At the southern coast of the Benguela system, alongshore wind stress had positivevalues (upwelling-favourable conditions) throughout the year, but the highestvalues occurred during the austral spring and summer months for the area extending from 20 to 30°S, indicating upwellingreinforcement from 1958 to 2007. Narayan et al.−3 Nm–2 dec–1)in the COADS and National Center for Environmental Prediction/National Centerfor Atmospheric Research  meridional wind stress across26–36°S from 1960 to 2001 using annual data. On theother hand, annual wind stress data from European Center for Medium rangeWeather Forecasting (ECMWF) Re-Analysis  used in the samestudy9et al.et al.Previous studies reported similar results. For example, Patti −3 Nm–2 dec–1)and 27–29°N (−3 Nm–2 dec–1).Published results regarding trends in upwelling along the Canary coast arecontroversial. Trends in upwelling can be highly dependent on the length of thetime series, the selected area, and the season evaluated in the analysis.Results similar to those found in the present study were reported by Cropperet al.et al.et al.et al.−3 Nm–2 dec–1)using the same database across 24–32°N consideringannual wind stress over the last four decades. Narayan et al.−3 Nm–2 dec–1)in the ERA-40 Reanalysis meridional wind stress for the same period. Incontrast, when they used the NCEP/NCAR reanalysis these authors−3 Nm–2 dec–1)over the last four decades (1960–2001) in the same region(24–32 °N), indicating a reduction in coastal upwelling.More recently, Barton et al.et al.et al.et al.2Along the Canary coast, positive values of alongshore wind stress for almost all points. In the western region (71.25–76.56°W) a non-significant positive trend was observed in most of thearea, with maximum values of around 410−3 Nm–2 dec–1.This region was previously studied mainly in terms of upwelling occurrence usingwind and sea surface temperature data21222324Only one region along the coast of the western Atlantic Ocean fits the conditionsrequired to be considered an coastal upwelling area: the southern Caribbean Sea(62.19–76.56°W) . In thes period. The easegion 71.–76.56°WThree upwelling systems were evaluated along this coast from south to north:Chile (37.62–28.88 °S), Peru (16.39–10.15°S), and California  .−3 and –810−3 Nm–2 dec–1.Non-significant positive trends were detected at midlatitudes. Previous studiesalong this coast have reported different results. For example, Garreaud andFalveyet al.et al.et al.Along the coast of Chile, positive values of alongshore wind stress were observedthroughout the year, with the highest values during the austral spring andsummer months (e months. Signifi−3 Nm–2 dec–1).A non-significant negative trend with small values was observed at –1) at15°S, which is a prominent upwelling region along the Peru coast.Gutierrez et al.et al.et al.et al.−3 Nm–2 dec–1)was much higher than that found in the present study. Narayan et al.−3 Nm–2 dec–1)and ERA-40 Reanalysis (−3 Nm–2 dec–1)revealed a significant increasing trend in upwelling, in agreement with theresults reported by Patti et al.1415et al.−3 Nm–2 dec–1)at the Peruvian upwelling region (6–16°S) usingmeridional wind stress from the NCEP/NCAR reanalysis. Pardo et al.Along the Peru coast, alongshore winds had positive values throughout the year,with maxima from May to October only at the three southernmost points and at the northernmost one. Along thenorthern California coast, alongshore wind stress showed a similar behaviour,although the highest values were mainly observed in summer (−3 Nm–2 dec–1.Positive trends were observed north of 38.5 °N with a significancelevel higher than 90% for the northernmost region (4-610−3 Nm–2 dec–1).Along the southern California coast, the highest values of alongshore wind stresswere observed during the spring and summer months using annual wind stress data from the same dataset from 1960 to 2001 for thesame region. Although the southern area evaluated in the present study(33.88–36.06°N) is included in the region studied byMendelssohn and Schwinget al.et al.et al.−3 Nm–2 dec–1)from 34 to 40°N over the last four decades (1960–2001).This decrease is in good agreement with the results of the present study,although the trend value was much higher in our case (−3 Nm–2 dec–1).Different results were also reported in several studies that covered a widerregion. For example, Garcia-Reyes and LargierControversial results in relation to the long-term variability in coastalupwelling were also found in the California upwelling system. In terms of windspeed, Mendelssohn and Schwinget al.et al.et al.Ekman transport data also have been evaluated in different studies conductedalong the California Upwelling Ecosystem. Rykaczewski and Checkleywww.esrl.noaa.gov) can be an important source of variabilityalong the Pacific upwelling systems, its influence on the estimated trends ofConsidering that ENSO  and Java (105.62–116.87°E) .−3 Nm–2 dec–1south of 25.5°S. Non-significant negative trends were found at thenorthernmost latitudes (21.4–22.9°S). As far as we know,no studies regarding upwelling trends in terms of wind have been conducted alongthe western coast of Australia. Previous studies have shown the absence ofpersistent upwelling off this coast despite a prevailing summer wind systemfavouring upwelling. This absence of upwelling has been attributed to thepresence of the Leewin Current (LC), a warm poleward flow transportingnutrient-poor waters from the tropics3739414243444546Along the western Australian coast, maximum values of alongshore wind stress wereobserved from October to March, which corresponds to the austral spring andsummer months  and Somalia-Oman(1.72–22.01°N) .−3 Nm–2 dec–1.The coast of northern Kenya is characterized by the occurrence of an irregularupwelling linked to the northeast monsoon, which normally develops from Novemberto March. Several researchers have related the occurrence of this upwelling tohigher productivity in the area, although the fact that upwelling events are notregular has attracted relatively little scientific interest about upwellingtrends in terms of wind. This region has been studied mainly in terms of changesin chemical and biological oceanographic parameters related to the occurrence ofthe northeast and southeast monsoons, which lead to important differencesbetween the Kenya and Somalia coasts in terms of upwelling545556575859Along the northern Kenya coast, positive values of alongshore wind stress wereobserved from November to April  trend was found east (west) of71.25°W.Our analysis covered the last three decades (1982–2010), which is therecent period of strongest global warming, thus it allowed detailed analysis ofthe influence of warming processes on upwelling trends. In another study, Limaand WetheyAmong the upwelling systems analysed in the present study, Lima and WetheyThese results delve into a possible discussion about whether the Bakun hypothesisis being fulfilled or not taking into account the present study and thosementioned in Conceived and designed the experiments: R.V., I.A., F.S., M.dC. and M.G-G. Performedthe experiments: R.V. Analyzed the data: R.V., I.A., F.S., M.dC. and M.G-G.Contributed reagents/materials/analysis tools: R.V., F.S. and M.G-G. Wrote thepaper: R.V., I.A., M.dC. and M.G-G. All authors reviewed the manuscript.How to cite this article: Varela, R. et al. Has upwelling strengthenedalong worldwide coasts over 1982-2010? Sci. Rep.5, 10016; doi: 10.1038/srep10016 (2015).


The OC concentrations on all three particle size fractions were significantly related to the OC concentration of bulk soil. However, OC concentration increased slowly toward an apparent maximum on silt and clay, but this maximum was far greater than the maximum predicted by established C sequestration models. In addition, significant increases in OC associated with sand occurred when the bulk soil OC concentration exceeded 30 g C kg−1, but this increase occurred when the OC concentration on silt + clay was still far below the predicted storage capacity for silt and clay fractions. Since the OC concentrations in all fractions of Brookston clay loam soil continued to increase with increasing C (bulk soil OC content) input, we concluded that the concept of OC storage capacity requires further investigation.We test the common assumption that organic carbon (OC) storage occurs on sand-sized soil particles only after the OC storage capacity on silt- and clay-sized particles is saturated. Soil samples from a Brookston clay loam in Southwestern Ontario were analysed for the OC concentrations in bulk soil, and on the clay (<2 μm), silt (2–53 μm) and sand (53–2000 μm) particle size fractions. The OC concentrations in bulk soil ranged from 4.7 to 70.8 g C kg We further hypothesized that relationships among OC storage in bulk soil and on the sand, silt and clay particle size fractions could be profitably examined by considering a single soil type with a large range of OC concentrations resulting from a range of long-term soil and crop management practices. Accordingly, soil samples were collected from several long-term field trials on a Brookston clay loam, and relationships among OC storage in bulk soil and on the clay (<2 μm), silt (53–2 μm) and sand (>53 μm) particle size fractions were studied.−1 soil with an average of 20.8 g C kg−1. The concentration of OC associated with particle size fractions ranged from 0.4 to 66.9 g C kg−1 for sand, 1.9 to 64.4 g C kg−1 for silt, and 6.2 to 84.0 g C kg−1 for clay, with the averaged values of 6.4, 21.3, and 29.9 g C kg−1, respectively. The concentration of OC associated with particle size fractions on a bulk soil basis ranged from 0.1 to 28.3 g C kg−1 bulk soil for sand, 0.6 to 20.7 g C kg−1 bulk soil for silt, and 2.9 to 28.9 g C kg−1 bulk soil for clay, with the averaged values of 1.8, 7.1 and 11.8 g C kg−1 bulk soil, respectively.The OC concentrations in bulk soil and in the sand, silt and clay sizes are presented in 2 = 0.73, 0.96 and 0.95 for sand, silt and clay, respectively  varied greatly from 0.051 kg sand g−1 C for sand, to 0.014 kg clay g−1 C for clay, to 0.006 kg silt g−1 C for silt. Despite the fact that the clay rate constant was a factor of 2.3 greater than the silt rate constant, the predicted storage capacity for the clay fraction , over 70% of OC was associated with clay, less than 5% with sand, and the remainder with silt. Increasing bulk soil OC resulted in deceases in the proportion of OC on clay, and compensating increases in the proportions of OC on sand and silt fractions  Brookston clay loam under grain production, the proportion of OC in clay fraction dropped to 50–60%, while that in silt and sand fractions increased to 30–35% and 15%, respectively. With further increases in bulk soil OC up to 60 g C kg−1 soil, the proportion of OC associated with clay dropped to below 40%, while that in silt and sand fractions increased to about 30%  of Brookston clay loam play a key role in OC retention. In addition, the maximum soil OC associated with soil particles <53 μm diameter was much greater than the “protective capacity” or “saturation level” predicted by a recognized carbon storage model for Canadian soils (40 plus g C in clay + silt fraction kg−1 soil)When bulk soil OC was <20 g C kget al.et al.et al.et al.et al.−1 for silt + clay , of the Carter t + clay . Howevernagement . It theret al.et al.231325et al.−2 as a potential maximal OC associated with fine particles based on data from marine sediments and upland soils. Soils with OC loading less than ~1 mg C m−2 are more likely to represent agricultural soils or deep soil horizons with low C inputs24−2 of holding capacity may not be an accurate estimate of maximal OC sequestration. Our results support the view that only a limited portion of fine mineral surfaces contribute to OC sequestration and more work is needed to elucidate the impact of amount of carbon input on OC saturation levels.Clay mineral surface area has been used in place of clay concentration to estimate the sequestration potential of soils. However, using sieved (<2 mm) topsoil and labelled organic carbon, Vogel −1 soil, which are substantially below the saturation levels of 77 g C kg−1 for OC predicted by the model  is typical for the near-surface of cash-cropped Brookston clay loam in this study, and appears to be “steady state equilibrium” for the current management systems. In contrast to other studies, however, we found that not only sand OC, but also silt and clay OC, continued to increase with increasing bulk soil OC.Using the “protective capacity” concept, HassinkIn summary, the ability of clay and silt to “store” OC appears to exist for Brookston clay loam soil, but the apparent OC storage capacity of the clay and silt fraction seems much greater than the capacity predicted by existing OS sequestration models. Although the OC on the sand fraction of Brookston clay loam showed a sharp increase after the OC on the clay fraction reached a certain level, the OC on clay and silt continued to increase at a reduced rate. These results support our hypothesis that literature estimates of OC storage capacity on clay and silt can be substantially underestimated. It is also appears that soil carbon sequestration is determined by more than the “protective capacity” of the silt plus clay particle sizes.−1, and average plow layer texture of 265 g kg−1 sand, 327 g kg−1 silt, and 408 g kg−1 clay. This is why most Brookston clay loam soil in these studies and in southwestern Ontario are systematically tile drained. The Brookston clay loam soils comprise about 80% of the agricultural land in Southwestern, Ontario, Canada. Soil samples were collected from three long-term field experiments, including: i) an organic amendment study which received one-time applications  of household food waste compost , yard waste compost (75 Mg ha−1), and pig manure plus wheat straw compost (75 Mg ha−1)  at the field site  is a poorly drained lacustrine soil with an average plow layer OC concentration of 20.8 g C kg−1 soil suspension. The concentrations of OC in bulk soil, and in the sand, clay, and silt + clay fractions were determined using a CN 2000 analyzer . Because the soil is free of carbonates, the total C concentration is equivalent to the soil OC concentration. The OC in the silt fraction was calculated by weight proportion usingSoil samples were collected 10 years after initiation of the organic amendment study, 50 years after initiation of the fertilization and crop rotation study, and 24 years after initiation of the tillage study. A total of 221 soil cores (3.3 cm diameter) were obtained using a tractor-mounted Concord soil sampler, fitted with polycarbonate tube inserts. The soil cores were cut into six segments  for the organic amendment and fertilization-rotation studies, and into five segments  for the tillage study. Replicate samples from the same depth were pooled within each experiment and visible crop residues removed. The pooled samples were passed through a 2 mm sieve and air-dried, and then separated into the clay (<2 μm), clay + silt (<53 μm), and sand (53–2000 μm) particle size fractions using sonicationMsi+cl is weight percentage (%) of the silt + clay fraction, Mcl is weight percentage of the clay fraction, and Csi, Ccl and Csi+cl are the organic carbon concentrations (g C kg−1 soil) in the silt, clay, and silt + clay fractions, respectively.where Relationships among OC concentrations on different soil particle sizes and bulk soil were characterized using SigmaPlot V12.0  to (best) curve-fit various single-pool growth models to OC data. Since the OC in sand size particles grew exponentially with no apparent maximum, y (g kg−1 soil) and a (g kg−1 soil) are, respectively, the current and antecedent concentrations of OC on sand-size particles, b (kg soil g−1) is a constant describing the rate of accumulation/depletion of OC, and x (g kg−1 soil) is the concentration of OC in bulk soil. For silt and clay, an exponential growth to maximum model produced the best fits:where y (g kg−1 soil) and y0 (g kg−1 soil) are, respectively, the current and antecedent OC concentrations on silt or clay, (y0 + a) (g kg−1 soil) is the storage capacity or saturation level for OC, b (kg soil g−1) is a constant describing the rate of accumulation/depletion of OC, and x (g kg−1 soil) is the concentration of OC in bulk soil. Relationships among the proportional changes of OC in particle size fractions as related to OC content in bulk soil were also evaluated using best empirical curves .


Several prostate cancers carry homologous recombination repair mutations that respond to olaparib. Because of the mechanism, the efficacy of platinum‐based therapy can be used to predict the efficacy of poly(adenosine diphosphate‐ribose) polymerase inhibitors such as olaparib.BRCA2 mutation in the germline and case 2 had a BRCA2 mutation in a somatic chromosome only. Both patients responded well to olaparib.We experienced two neuroendocrine prostate cancer patients who achieved a response duration of more than 1 year with platinum‐based therapy. Case 1 had a Cisplatin and olaparib may overlap in response due to their medicinal action. It may be useful to consider genetic testing in some CRPC patients who have responded to cisplatin. ARandrogen receptorCABcombined androgen blockadeCRPCcastration‐resistant prostate cancerGSGleason scoreNEPCneuroendocrine prostate cancerNSEneuron‐specific enolaseOSoverall survivalPARPpoly(adenosine diphosphate‐ribose) polymerasePCprostate cancerPSAprostate‐specific antigenMany castration‐resistant prostate cancer patients who convert to neuroendocrine prostate cancer cases have a poor prognosis. However, genetic testing should be considered in neuroendocrine prostate cancer patients who might benefit from platinum‐based therapy.BRCA mutation was identified after a successful platinum regimen and the subsequent response to olaparib was confirmed.Prostate cancer (PC) can be effectively treated it with androgen deprivation therapy, but some cases have been reported to develop castration‐resistant prostate cancer (CRPC). Some of these patients may undergo neuroendocrine differentiation that is not mediated by androgen receptor (AR) signaling, resulting in neuroendocrine prostate cancer (NEPC). NEPC is successfully treated with a platinum‐based regimen similar to small cell lung cancer. We experienced two cases in which a BRCA2 mutation, so we administered olaparib. Three months after, the back pain improved, and the lymph node metastasis had shrunk and was determined to be partial response (PR)  of 4 + 5 = 9, prostate‐specific antigen (PSA) level of 40.8 ng/ml with metastasis of thoracic vertebra 3 years ago. Combined androgen blockade (CAB) therapy and proton radiation therapy (78.0 Gy/39 fr) to the prostate were performed. The PSA level decreased to <0.01 ng/ml, and the CAB was stopped after 2.5 years. Five months after stopping the CAB, he presented back pain with PSA <0.01 ng/ml. A computed tomography (CT) scan detected multiple liver, bone, and lymph node metastases with a high neuron‐specific enolase (NSE) level of 171 ng/ml Fig. . A biopsBRCA mutations. A multi‐gene cancer panel test (FoundationOne®CDx) was performed using bladder metastatic tissue collected at the time of TUR. The results showed that the patient was positive for a somatic BRCA2 mutation. The lymph node shrank from 21.0 mm to 7.5 mm over the course of 3 months after olaparib administration  performed. A pathological examination revealed positive findings for chromogranin A and an elevated NSE level, so we diagnosed NEPC. Carboplatin and etoposide combination therapy shrank the tumor. Thereafter, 17 courses were administered over 24 months, and the disease was controlled. Chemotherapy was stopped due to neuropathy and fatigue. Subsequently, the patient showed progression of pelvic lymph node metastasis Fig. . Geneticion Fig. . PSA decNEPC is an aggressive variant characterized by low or absent AR expression and gain of the neuroendocrine phenotype, but it is not responsive to therapies targeting AR signaling. De novo NEPC accounts for <2% of all prostate cancers, but treatment‐induced NEPC occurs in 10–17% of patients with CRPC as a result of transdifferentiation.Platinum‐based chemotherapy is usually used to treat NEPC. Several studies have shown platinum‐based chemotherapy results in response rates of 8.9–41%, progression‐free survival of 5.8 months, and overall survival of 9.6–12 months.BRCA1/2 have more aggressive disease than those with proficient homologous recombination repair.BRCA1, BRCA2, or ATM.BRCA1, BRCA2, or both mutations. The risk of disease progression or death was 70% lower with olaparib than with a placebo.BRCA1 and BRCA2 mutations; thus, platinum drugs are effective. We speculated that BRCA1/2 mutations might be associated with a high frequency in NEPC cases that responded to platinum therapy. However, we considered the following two points to be important. First, according to the previous reports, the frequency of HRR mutation including BRCA 1/2 mutations in treatment‐related (induced) NEPC patients is less than 20%.CRPC patients with deleterious alterations in genes involved in homologous recombination repair such as Yoshiyuki Miyazawa: Conceptualization; data curation; formal analysis; investigation; resources; validation; writing – original draft. Takanori Shimizu: Validation. Yositaka Sekine: Supervision; validation. Seiji Arai: Supervision. Akira Ohtsu: Validation. Yuji Fujizuka: Supervision. Masashi Nomura: Supervision. Hidekazu Koike: Validation. Hiroshi Matsui: Supervision. Kazuhiro Suzuki: Supervision; writing – review and editing.Kazuhiro Suzuki has potential financial conflicts of interest as below, Consultancies: Astellas Pharma, Sanofi, Janssen, Bayer, Takeda Pharmaceutical, Daiichi‐Sankyo, AstraZeneca, Grants received: Astellas Pharma, Ono Pharmaceutical, Takeda Pharmaceutical, Daiichi‐Sankyo. All other authors have no conflict of interest exists.Not applicable.We obtained informed consent from all patients.Not applicable.


Patients with bipolar disorder (BPD) are at high risk of cardiovascular diseases (CVDs) that are attributed to endothelial dysfunction. Circulating endothelial progenitor cells (cEPCs) are proposed as indicators of endothelial dysfunction. This study examined the relationship of cEPC numbers with BPD diagnosis and its clinical symptoms in patients with BPD.We recruited 48 patients with BPD and 50 healthy controls (HCs). All the patients had scores of <13 on the Young Mania Rating Scale (YMRS). In addition to the YMRS and Clinical Global Impression for BPD (CGI‐BP), bipolar patients were assessed using relevant measurements for their depression, anxiety, general psychopathology, cognitive dysfunction and deficit, somatic symptoms, quality of life, and level of disability. cEPC counts were measured using flow cytometry.p [corrp] = .00018) and positive scores in the positive and negative syndrome scale . The number of mature cEPCs was inversely correlated with YMRS (corrp = .014), CGI‐BP (corrp = .00022), and PANSS‐P (corrp = .0049) scores. In multivariate stepwise analysis, numbers of both types of cEPCs were associated with CGI‐BP.The numbers of immature and mature cEPCs in the bipolar patients did not significantly differ from those in the HCs. After correction for multiple comparisons and controlling for body mass index and smokers, the number of immature cEPCs was observed to be inversely correlated with CGI‐BP (corrected cEPC levels, an indicator of endothelial dysfunction and risk of CVDs, may be associated manic and positive symptom severities in patients with BPD. Evidence suggests that patients with BPD are more susceptible to CVDs, including ischemic heart disease, congestive heart failure, and cerebrovascular disorders  derived from the bone marrow  discovered that the cEPC numbers of patients with BPD (some of whom were treated with lithium) did not differ significantly from those of controls. However, that study did not investigate the association of cEPC numbers with the clinical symptoms of BPD . Board‐certificated senior psychiatrists interviewed all the participants using the Mini International Neuropsychiatric Interview (MINI), clinical observation, and medical charts to diagnose psychiatric disorders. The diagnosis of BPD was made based on the criteria for BPD specified in the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition, Text Revision (DSM‐IV‐TR). The exclusion criteria were any DSM‐IV‐TR diagnosis , unpredictable physical illness, pregnancy, and breastfeeding. Age‐ and gender‐matched participants without any psychiatric disorder were included in the healthy control (HC) group. The participants’ ages ranged from 20 to 65 years. This study was approved by the Institutional Review Board of Taipei Veterans General Hospital and was conducted in accordance with the Declaration of Helsinki. Written informed consent was obtained from the participants prior to their study involvement.2.2This study assessed the severity of manic symptoms by using the YMRS and Clinical Global Impressions Bipolar (CGI‐BP) scale  and Montgomery–Åsberg Depression Rating Scale (MADRS). Both scales are commonly applied in psychiatric research and clinical practice for assessing depression level. However, the HAMD‐17 does not evaluate oversleeping, overeating, and concentration difficulties, and the MADRS lacks assessment of psychomotor changes, reduced interest in daily activities, or guilt  to assess the level of general BPD psychopathology  was employed to assess the participants' objective neurocognitive function. The DSST measures the psychomotor speed of the performance in tasks requiring visual perception, spatial decision making, and motor skills. The DSST is a valid and sensitive assessment of various cognitive domains, including executive function, attention, psychomotor speed, and working memory Jaeger, . In addiThe participants also completed two other self‐administered questionnaires: the Depressive and Somatic Symptom Scale (DSSS) and the 12‐item Short‐Form Health Survey (SF‐12). The advantage of the DSSS is that it simultaneously assesses somatic and depressive symptoms, overcoming the deficiencies of other depression scales that assess few somatic elements  to assess patients' functional impairment in daily life. The SDS is a three‐item self‐reported tool for evaluating functional disability in work, school, social, and family life contexts . The intraindividual variability of immature (CD34+KDR+CD133+) and mature (CD34+KDR+) cEPCs over time was evaluated based on two measurements conducted 1 year apart in 21 patients, and the resulting intraclass correlation coefficients were 0.69, 0.75, and 0.78, respectively. Cell counts are expressed as cEPCs per 105 mononuclear cells , aspartate aminotransferase (AST), creatinine (CREAT), fasting blood glucose (FBS), triglyceride (TG), cholesterol (CHOL), high‐density lipoprotein cholesterol (HDL), and uric acid (UA) levels, were determined using standard laboratory procedures. For the measurement of cEPC counts, a 1.0‐ml sample of peripheral blood was obtained from each participant. The cEPC count was measured through flow cytometry by researchers blinded to the patients’ clinical information. The blood samples were subsequently incubated with allophycocyanin (APC)‐conjugated monoclonal antibodies against human KDR , phycoerythrin (PE)‐conjugated monoclonal antibodies against human CD133 , and fluorescein isothiocyanate (FITC)‐conjugated monoclonal antibodies against human CD34  in the dark for 30 min. Each analysis was based on 150,000 acquired events and was as reliable as 500,000 events (intraclass correlation coefficient > 0.95). When viability markers were included, the viability of cEPCs reached 96.3%. The interindividual variability of two samples obtained from 10 patients was strongly correlated . Model 1 controlled for the effect of body mass index (BMI) and smokers. Model 2 additionally controlled for systolic blood pressure (SBP), diastolic blood pressure (DBP), FBS, and HDL in addition to controlling for BMI and smokers. Model 3 additionally controlled for the use of antipsychotic drugs and mood stabilizers in addition to controlling for BMI and smokers. Models 2 and 3 are the parallel sensitivity analyses. Multivariate stepwise linear regression was used to identify the most predictive clinical symptom associated with cEPC counts with adjustment for potential confounding factors . The criterion for entry an independent variable was set as if a p‐value less than .05 and for removal as greater than .1. The statistical significance level for all univariate analyses was set at a corrected p‐value of <.05 after multiple comparison adjustment with the Benjamini–Hochberg procedure (https://tools.carbocation.com/FDR), and the threshold for statistical significance in the multivariate stepwise linear regression was set as p <.05.Statistical analyses were performed using SPSS . The distribution of categorical variables across groups was compared using the chi‐square test (and the Fisher exact test if necessary). The group differences in continuous variables were determined using the independent 33.1N = 2), heart disease (N = 1), diabetes mellitus , and hyperlipidemia (N = 2). The total YMRS score of all patients with BPD was less than 13. The mean  scores of YMRS, HAMD‐17, and MADRS in the BPD patients were 2.4 , 10.9 , and 15.1 , respectively. Table U test, all p <.05). No significant difference was observed in mature and immature cEPC counts between the patients with BPD and the HCs .In this study, 48 patients with BPD  and 50 HCs were recruited. The BPD patients were treated with the following psychotropic drugs: antipsychotic drugs , mood stabilizers , and antidepressants . Some of the patients had also received diagnoses of hypertension (3.2b) between the cEPC counts and clinical measurements related to BPD in three analytic models with Kendall tau‐b partial correlations. Both immature and mature cEPC counts were correlated inversely with YMRS in Model 1 , Model 2 , and also in Model 3 . The counts of both types of cEPCs were also correlated inversely with CGI‐BP in Model 1 , Model 2 , and Model 3 . The score of PANSS‐P were found to correlate negatively with both types of cEPCs in Model 1 , Model 2 , and Model 3 . Despite the correlation noted between immature cEPCs and YMRS scores, inverse correlations of the mature cEPCs count with the YMRS scores, immature and mature cEPC counts with the CGI‐BP scores, and immature and mature cEPC counts with the PANSS‐P scores in the three partial correlation models remained statistically significant after correction for multiple comparisons .Table 3.3p = .044), YMRS score (p = .030), CGI‐BP score (p = .002), and PANSS‐Total score  were significantly associated with the immature cEPC count. Among these results, the association between CGI‐BP scores and the immature cEPC count remained statistically significant after correction for multiple testing (corrected p = .04). The mature cEPC count was significantly associated with being a smoker (p = .013) as well as YMRS (p = .021), CGI‐BP (p = .002), PANSS‐T (p = .016), and PANSS‐G (p = .044) scores in the univariate analysis. Among these results, the association between the mature cEPC count and CGI‐BP score remained statistically significant after correction for multiple testing (corrected p = .044). In the multivariate stepwise linear analysis, the CGI‐BP score was significantly associated with both immature and mature cEPC numbers .In this study, linear regression was used to determine the relationship between the cEPC count and clinical information with simultaneous adjustment for potential confounding factors . In the analyses, cEPC counts were the dependent variables, and scores of the clinical measures were the independent variables. Table 4In this study, we explored the role of cEPCs in BPD and BPD‐related clinical presentations in patients with BPD. Although no significant difference in immature and mature cEPC counts was noted between the patients with BPD and the HCs, we discovered that the count of both types of cEPCs exhibited significant inverse correlations with YMRS, CGI‐BP, and PANSS‐P scores Table . The greEvidence has suggested that variations in the cEPC count are related to affective symptoms. Such a finding mainly originates from the effects of depressive symptom severity and depressive disorders on the cEPC count , C‐reactive protein (CRP), and interleukin 6 (IL‐6). CRP, an acute phase marker of systemic inflammation, exerts a direct inhibitory effect on cEPC differentiation, migration, proliferation, adhesion, and angiogenic activity , BPD diagnosis, and levels of clinical symptom severity. Third, adolescent patients, older patients, or patients undergoing any acute mood‐related episode were not enrolled in this study. The findings of inverse correlations of manic and positive symptom severities with cEPC counts were obtained from the data of patients whose total YMRS scores were less than 13 (Tohen et al., In conclusion, the present study reported low numbers of cEPC correlated with severe manic and positive symptom severity in patients with BPD. The inverse correlation of cEPC counts and the severity of manic and positive symptoms were independent of several cardiovascular factors. Although the medians of both types of cEPCs did not significantly differ between the patients with BPD and the HCs, our results suggest that the cEPC count may be a treatment, rather than a diagnostic, marker of endothelial dysfunction in patients with BPD who are at risk of CVDs (Gill et al., All authors have no conflict of interest and no financial support relevant to this article to disclose.Ya‐Mei Bai and Kai‐Lin Huang conceptualized and supervised the project. Ying‐Jay Liou performed data analyses and drafted the manuscript. Mu‐Hong Chen, Ju‐Wei Hsu, Kai‐Lin Huang, and Ya‐Mei Bai provided the samples and clinical data for the study. Po‐Hsun Huang designed, performed and supervised the laboratory experiments. All authors have made substantial contributions to the work, reviewed, and revised the manuscript.


Mycobacterium avium complex (MAC) lung disease. He was admitted because of fever. Pneumothorax and pleural effusion were present in the right lung, and a new consolidation appeared in the right upper lobe. Based on positive culture of the pleural effusion for CAM‐resistant M. avium and findings on chest computed tomography, he was diagnosed with pleurisy due to M. avium, with rupture of the subpleural lung parenchymal lesion into the pleural space. Additional treatment with streptomycin (SM) improved the patient's high‐grade fever. SM might be effective for pleurisy caused by CAM‐resistant MAC lung disease.A 72‐year‐old man received rifampicin, ethambutol and sitafloxacin to treat clarithromycin (CAM)‐resistant  Mycobacterium avium complex lung disease, which was treated effectively with streptomycin.Here, we describe a case of pleurisy with pneumothorax due to clarithromycin‐resistant  Mycobacterium avium complex (MAC) lung disease has increased.The prevalence of clarithromycin (CAM)‐resistant M. avium was confirmed 4 years ago, so CAM was withdrawn and aminoglycoside antibiotics were proposed; however, the patient refused therapy due to difficulty in attending the frequent hospital appointments required for intramuscular injection of SM. Subsequently, the radiological findings worsened and positive cultures continued  level were 163 cm, 48 kg, 38.4°C and 96% on room air, respectively. The white blood cell count was 6670/ml, and C‐reactive protein (CRP) was 20.1 mg/dl The test for HIV was negative. A chest x‐ray on admission revealed pneumothorax and pleural effusion in the right lung. A chest drainage tube was inserted into the right thoracic cavity to treat the pneumothorax. At that time, pleural effusion was collected. Biochemical analysis of the effusion revealed the following: protein, 5.5 g/dl ; lactate dehydrogenase (LDH), 2021 IU/L ; adenosine deaminase, 113.2 U/L; and glucose, 45 mg/dl. Sulbactam/ampicillin (ABPC/SBT) was added soon after admission as the bacteriological results of the effusion were unavailable at that point in time. Despite the treatment, the fever remained unchanged. Then, an effusion smear was negative for acid‐fast bacilli, and cultured isolates were identified as CAM‐resistant M. avium . A sputum smear was positive for acid‐fast bacilli, and cultured isolates were identified as CAM‐resistant M. avium. No other microorganisms were cultured from sputum and pleural fluid. We diagnosed pleurisy due to M. avium on the basis of ineffectiveness of ABPC/SBT and positive culture of the pleural effusion for M. avium. On Day 7, pneumothorax improved and consolidation in the right upper lobe was revealed on chest computed tomography , ethambutol (EB) and CAM from 15 years to 5 years before this visit. Because the radiological findings worsened and positive cultures continued, sitafloxacin (STFX) was added to the regimen 5 years before this visit. CAM‐resistant d Figure . On admiy Figure . We diagy Figure .Despite the combination therapy including fluoroquinolone, aminoglycoside and surgical resection, the outcomes of CAM‐resistant MAC lung disease were poor.A previous study has shown that pleural concentrations increase gradually to a maximum value about 6 h after intravenous administration of amikacin.None declared.The authors declare that appropriate written informed consent was obtained for the publication of this manuscript and accompanying images.