Summary Background Dupilumab [a monoclonal antibody blocking the shared receptor subunit for interleukin (IL)‐4 and IL‐13] is approved for patients aged ≥ 12 years with inadequately controlled, moderate‐to‐severe atopic dermatitis (AD). Dupilumab trials of up to 52 weeks demonstrated efficacy and a favourable safety profile in patients with moderate‐to‐severe AD inadequately controlled with topical medications. Objectives To further characterize the safety of dupilumab by evaluating clinical laboratory findings from three randomized, double‐blinded, placebo‐controlled phase III trials (LIBERTY AD SOLO 1 & 2 and LIBERTY AD CHRONOS). Methods Patients were randomized 1 : 1 : 1 (SOLO 1 & 2) or 3 : 1 : 3 (CHRONOS) for 16 and 52 weeks, respectively, to dupilumab weekly, every 2 weeks or placebo. CHRONOS patients received a standardized concomitant topical corticosteroid regimen. Laboratory outcomes were summarized descriptively in 1376 patients from SOLO 1 & 2 and 740 from CHRONOS. Results Treatment groups had similar results in baseline laboratory parameters. Platelets and neutrophils showed mild decreases from baseline in dupilumab vs. placebo groups. Some dupilumab‐treated patients had small transient increases in eosinophils. Grade 3 eosinophilia was reported in < 1% of dupilumab‐treated and placebo‐treated patients; no adverse events were associated with eosinophilia. Lactate dehydrogenase levels decreased from baseline during dupilumab treatment in all trials. No clinically meaningful changes were observed between treatment groups in other haematology, chemistry or urinalysis parameters. Conclusions There were no clinically important changes in routine laboratory parameters that could be attributed to dupilumab. This study supports the use of dupilumab as a systemic treatment for moderate‐to‐severe AD that does not require laboratory monitoring. What's already known about this topic? Long‐term treatment of atopic dermatitis (AD) with conventional immunosuppressive agents is limited by the risk of significant side‐effects and a need for repeated tests to monitor haematological and/or organ (e.g. liver, kidney) toxicities. Dupilumab [a monoclonal antibody blocking the shared receptor subunit for interleukin (IL)‐4 and IL‐13] is approved for the treatment of patients with inadequately controlled, moderate‐to‐severe AD. In 16‐week and 52‐week studies, dupilumab demonstrated a positive risk/benefit profile in moderate‐to‐severe AD. What does this study add? This study is the first comprehensive analysis of dupilumab laboratory safety data of the 16‐week SOLO 1 & 2 (pooled N = 1376) and 52‐week CHRONOS (N = 740) trials, demonstrating an absence of clinically important changes in haematology, serum chemistry and urinalysis parameters in patients with moderate‐to‐severe AD treated with dupilumab. Our data support the use of dupilumab as a systemic treatment for the long‐term management of moderate‐to‐severe AD without routine laboratory monitoring in clinical practice.

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SummaryPectate lyase genes have been documented as excellent candidates for improvement of fruit firmness. However, implementation of pectate lyase in regulating fruit postharvest deterioration has not been fully explored. In this report, 22 individual pectate lyase genes in tomato were identified, and one pectate lyase gene SlPL (Solyc03g111690) showed dominant expression during fruit maturation. RNA interference of SlPL resulted in enhanced fruit firmness and changes in pericarp cells. More importantly, the SlPL‐RNAi fruit demonstrated greater antirotting and pathogen‐resisting ability. Compared to wild‐type, SlPL‐RNAi fruit had higher levels of cellulose and hemicellulose, whereas the level of water‐soluble pectin was lower. Consistent with this, the activities of peroxidase, superoxide dismutase and catalase were higher in SlPL‐RNAi fruit, and the malondialdehyde concentration was lower. RNA‐Seq results showed large amounts of differentially expressed genes involved in hormone signalling, cell wall modification, oxidative stress and pathogen resistance. Collectively, these data demonstrate that pectate lyase plays an important role in both fruit softening and pathogen resistance. This may advance knowledge of postharvest fruit preservation in tomato and other fleshy fruit.

SummaryIn Arabidopsis, the miR171‐GRAS module has been clarified as key player in meristem maintenance. However, the knowledge about its role in fruit crops like tomato (Solanum lycopersicum) remains scarce. We previously identified tomato SlGRAS24 as a target gene of Sly‐miR171. To study the role of this probable transcription factor, we generated transgenic tomato plants underexpressing SlGRAS24, overexpressing SlGRAS24, overexpressing Sly‐miR171 and expressing β‐glucuronidase (GUS) under the SlGRAS24 promoter (proSlGRAS24‐GUS). Plants overexpressing SlGRAS24 (SlGRAS24‐OE) had pleiotropic phenotypes associated with multiple agronomical traits including plant height, flowering time, leaf architecture, lateral branch number, root length, fruit set and development. Many GA/auxin‐related genes were down‐regulated and altered responsiveness to exogenous IAA/NAA or GA 3 application was observed in SlGRAS24‐OE seedlings. Moreover, compromised fruit set and development in SlGRAS24‐OE was also observed. These newly identified phenotypes for SlGRAS24 homologs in tomato were later proved to be caused by impaired pollen sacs and fewer viable pollen grains. At anthesis, the comparative transcriptome results showed altered expression of genes involved in pollen development and hormone signalling. Taken together, our data demonstrate that SlGRAS24 participates in a series of developmental processes through modulating gibberellin and auxin signalling, which sheds new light on the involvement of hormone crosstalk in tomato development.

Fluorescent Ca2+ indicators have been essential for the analysis of Ca2+ signaling events within the cells of the CNS. However, a systematic evaluation of their potential adverse effects is lacking. Here we show that chemical Ca2+ indicators, but not a genetically encoded Ca2+ indicator, potently suppress the activity of Na+- and K+-dependent adenosine triphosphatase (Na,K-ATPase), independently from their Ca2+ chelating activity. Loading of commonly utilized Ca2+ indicators, including Fluo-4 acetoxymethyl (AM), Rhod-2 AM, Fura-2 AM and the Ca2+ chelator BAPTA AM, in cultures of neurons, astrocytes, cardiomyocytes and kidney proximal tubule epithelial cells suppressed Na,KATPase activity by 30–80%. Ca2+ indicators also suppressed agonist-induced activation of Na,K-ATPase, altered metabolic status and produced a dose-dependent loss of cell viability. In vivo, loading of Ca2+ indicators significantly altered brain extracellular concentrations of K+ and ATP. A critical review of the previous observations based on chemical Ca2+ indicators is therefore warranted.

We undertook a bivariate meta-analysis to assess the overall accuracy of respiratory specimen PCR assays for diagnosing Pneumocystis pneumonia. The summary sensitivity and specificity were 0.99 (95% confidence interval, 0.96 to 1.00) and 0.90 (0.87 to 0.93). Subgroup analyses showed that quantitative PCR analysis and the major surface glycoprotein gene target had the highest specificity value (0.93). Respiratory specimen PCR results are sufficient to confirm or exclude the disease for at-risk patients suspected of having Pneumocystis pneumonia.Pneumocystis pneumonia (PCP) remains a frequent opportunistic infection among immunocompromised patients, especially AIDS patients (5,14,15,24). The gold standard for diagnosing PCP is the detection of cysts and/or trophic forms by staining (5, 15). Many studies have assessed the diagnostic yield of PCR techniques for diagnosing PCP. However, the true potential role of PCR assays remains controversial. Thus, we pooled prospective cohort studies with consecutive patients and undertook a bivariate meta-analysis to assess the diagnostic accuracy of PCR on the basis of sensitivity (SEN), specificity (SPE), and positive and negative likelihood ratios (PLR and NLR, respectively) (19).We searched the MEDLINE and EMBASE databases for the English-language literature published up to May 2011. Full-text publications were included if (i) they used PCR on respiratory samples, such as bronchoalveolar lavage fluid (BALF), induced sputum (IS), or oropharyngeal wash (OW), for immunocompromised patients with pulmonary diseases or requiring bronchoscopy for suspected PCP and (ii) the prospective cohort studies were performed with consecutive patients. Studies with fewer than 10 patients with PCP were excluded. We explored potential heterogeneity by subgroup analyses (16). All analyses were performed using Stata, version 10 (Stata Corp., College Station, TX), with the Midas program (6).Thirteen reports, including 20 eligible studies, met our inclusion criteria (1-3, 7-12, 18, 21, 25, 26) (Table 1). Twelve articles reported patients with false-positive results, and follow-up was complete for most of these patients. When probable PCP (clinical and radiographic findings consistent with the diagnosis and consequent recovery with anti-PCP treatment) was included, 31.8% (65/204) of the patients had PCP. We could not extract the exact data for AIDS and non-AIDS patients from two articles (7,8), which led to discrepancies between the whole and subset populations. We found significant heterogeneity for all test performances.The test performances for the whole and subset groups are shown in Table 2. For the whole population, the area under the summary receiver operating characteristic curve and 95% confidence intervals were 0.98 (0.96 to 0.99), indicating that the PCR assay has an excellent diagnostic value for PCP. For the whole population, the percentage of heterogeneity likely due to a threshold effect was 38%, indicating a moderate influence of a diagnostic threshold effect on the performance of ...

Molecules of the diffusible signal factor (DSF)-family are a class of quorum sensing (QS) signals used by the phytopathogens Xanthomonas. Studies during the last decade have outlined how Xanthomonas cells enter the QS phase. However, information on the mechanism underlying its exit from the QS phase is limited. RpfB has recently been reported as a fatty acyl-CoA ligase (FCL) that activates a wide range of fatty acids to their CoA esters in vitro. Here, we establish an improved quantification assay for DSF-family signals using liquid chromatography-mass spectrometry in X. campestris pv. campestris (Xcc). We first demonstrated that RpfB represents a naturally occurring DSF-family signal turnover system. RpfB effectively turns over DSF-family signals DSF and BDSF in vivo. RpfB FCL enzymatic activity is required for DSF and BDSF turnover. Deletion of rpfB slightly increased Xcc virulence in the Chinese radish and overexpression of rpfB significantly decreased virulence. We further showed that the expression of rpfB is growth phase-dependent, and its expression is significantly enhanced when Xcc cells enter the stationary phase. DSF regulates rpfB expression in a concentration-dependent manner. rpfB expression is also negatively regulated by the DSF signalling components RpfC, RpfG and Clp. The global transcription factor Clp directly binds to the AATGC-tgctgc-GCATC motif in the promoter region of rpfB to repress its expression. Finally, RpfB-dependent signal turnover system was detected in a wide range of bacterial species, suggesting that it is a conserved mechanism.

Background Atopic dermatitis is a chronic inflammatory condition with substantial burden and limited treatment options for adolescents with moderate-to-severe disease. Significantly more patients treated with dupilumab vs. placebo achieved Investigator's Global Assessment 0/1 at week 16. Objective The objective of this study was to assess the impact of dupilumab treatment vs. placebo on the achievement of clinically meaningful improvements in atopic dermatitis signs, symptoms and quality of life. Methods R668-AD-1526 LIBERTY AD ADOL was a randomized, double-blinded, parallel-group, phase III clinical trial. Two hundred and fifty-one adolescents with moderate-to-severe atopic dermatitis received dupilumab 300 mg every 4 weeks (q4w; n = 84), dupilumab 200 or 300 mg every 2 weeks (q2w; n = 82), or placebo (n = 85). A post-hoc subgroup analysis was performed on 214 patients with Investigator's Global Assessment > 1 at week 16. Measures of atopic dermatitis signs, symptoms, and quality of life were assessed. Clinically meaningful improvement in one or more of three domains of signs, symptoms, and quality of life was defined as an improvement of ≥ 50% in Eczema Area and Severity Index, ≥ 3 points in Peak Pruritus Numerical Rating Scale, or ≥ 6 points in the Children's Dermatology Life Quality Index from baseline. Results Of patients receiving dupilumab q2w, 80.5% [66/82] experienced clinically meaningful improvements in atopic dermatitis signs, symptoms, or quality of life at week 16 (vs. placebo, 20/85 [23.5%], difference 57.0% [95% confidence interval 44.5-69.4]; q4w vs. placebo, 53/84 [63.1%], difference 39.6% [95% confidence interval 25.9-53.3]; both p < 0.0001). Results were similar in adolescents with Investigator's Global Assessment > 1 at week 16 (q2w, 46/62 [74.2%] vs. placebo, 18/83 [21.7%], difference 52.5% [95% confidence interval 38.5-66.6]; q4w, 38/69 [55.1%] vs. placebo, difference 33.4% [95% confidence interval 18.7-48.1]; both p < 0.0001). Conclusions Dupilumab provided clinically meaningful improvements in signs, symptoms, and quality of life in adolescents with moderate-to-severe atopic dermatitis among patients with Investigator's Global Assessment > 1 at week 16. Treatment responses should be interpreted in the context of such clinically relevant patient-reported outcome measures. Trial Registration ClinicalTrials.gov; NCT03054428.