Photobacterium  were isolated from bleached coral Madracis decactis (scleractinian) in the remote St Peter & St Archipelago (SPSPA), Mid-Atlantic Ridge, Brazil. Healthy M. decactis specimens were also surveyed, but no strains were related to them. The novel isolates formed a distinct lineage based on the 16S rRNA, recA, and rpoA gene sequences analysis. Their closest phylogenetic neighbours were Photobacterium rosenbergii, P. gaetbulicola, and P. lutimaris, sharing 96.6 to 95.8% 16S rRNA gene sequence similarity. The novel species can be differentiated from the closest neighbours by several phenotypic and chemotaxonomic markers. It grows at pH 11, produces tryptophane deaminase, presents the fatty acid C18:0, but lacks C16:0 iso. The whole cell protein profile, based in MALDI-TOF MS, distinguished the strains of the novel species among each other and from the closest neighbors. In addition, we are releasing the whole genome sequence of the type strain. The name Photobacterium sanctipauli sp. nov. is proposed for this taxon. The G + C content of the type strain A-394T (= LMG27910T = CAIM1892T) is 48.2 mol%.Five novel strains of  Photobacterium comprises 26 formally described species   were identified as P. phosphoreum, P. damselae and P. mandapamensis   and coram corals . P. jeann Brazil , whereasustralia , as welln Brazil . Photobapamensis . Coral mpamensis .Madracis decactis in the Brazilian St Peter & St Paul Archipelago (SPSPA) analyzed 403 isolates , and RNA polymerase alpha subunit (rpoA) were obtained as described previously (®). Through this method, the DNA was simultaneously fragmented and tagged with sequencing adapters. The library size distribution was accessed using the 2100 Bioanalyzer and the High Sensitivity DNA Kit (Agilent®). The accurate quantification of the library was accomplished using the 7500 Real Time PCR (Applied Biosystems®) and the KAPA Library Quantification Kit (Kapabiosystems®). Paired-end (2 ×250 bp) sequencing was performed on a MiSeq (Illumina®) using the MiSeq reagent kit v2 (500 cycles). R1 and R2 reads were quality filtered (Q > 20) and 3’ end trimmed with Prinseq v0.20.4 . The gene sequence data obtained in this study are available through the open access website TAXVIBRIO (http://www.taxvibrio.lncc.br/). The GenBank accession numbers for the 16S rRNA, recA, and rpoA genes and genome sequences are listed in ®). Spectra were generated with mMass software v5.5.0  as described previously  medium at ambient temperature (∼27 °C) after 24–48 h incubation . Gene seeviously . Primerseviously . Sequenceviously . Trees weviously . The robeviously . For geneviously  was used v0.20.4 . Ray v. rameters . Generalrameters . In siliGGDC 2.0 . This onGGDC 2.0  to generGGDC 2.0 . DDH preGGDC 2.0  with logGGDC 2.0 . StrainsGGDC 2.0  towards e v5.5.0 . Type steviously  and by BPhotobacterium  confirmed that the isolates formed a distinct lineage related to P. rosenbergii and P. gaetbulicola  with their closest neighbours, respectively. These levels of similarity are below the cut-offs determined to define a species of the family Vibrionaceae  ranged from 99.8% to 100% based on recA. Their rpoA sequences were identical. Trees based on partial sequences of the housekeeping genes recA (855 bp) and rpoA (969 bp) also confirmed their phylogenetic position in the genus Photobacterium and revealed they constituted a separate branch, clearly indicating that they belong to a new Photobacterium species (T genome are supplied in In silico DDH (%) values between A-394T and P. angustum S14, P. damselae subsp. damselae CIP 102761, P. halotolerans DSM18316, P. leiognathi lrivu.4.1 and P. profundum 3TCK were 21.5 (±2.34), 22.7 (±2.37), 20.3 (±2.31), 21.6 (±2.35) and 20.6 (±2.31) respectively. AAI between A-394T and P. leiognathi lrivu.4.1 CIP 102761 was 75%.16S rRNA gene sequence analysis revealed that the five isolates formed a tight monophyletic branch affiliated to the genus acterium . The fivacterium . P. rosetrarenae  and P. m marinum . The phybulicola . The novionaceae . The sim species –S2. Gene18:0, and absence of C16:0 iso (P. rosenbergii (LMG 22223T), P. gaetbulicola (LMG 27839T) and P. lutimaris (LMG 25278T)  . MLSA wa 25278T) . Based oPhotobacterium sanctipauli .β-glucosaminidase, β-galactosidase and arginine dihydrolase; but negative for lipase (C14), valine arylamidase, cystine arylamidase, trypsin, α-chemotrypsin, acid phosphatase, α-galactosidase, β-glucuronidase, α-glucosidase, β-glucosidase, α-mannosidase, α-fucosidase, lysine decarboxylase, ornithine decarboxylase, H2S production, urease activity, acetoin production (Voges–Proskauer) and gelatinase. Variable reactions were obtained for esterase (C4) (−), esterase lipase (C8) (−), tryptophane deaminase (w) and indole production (−) . Reduces nitrate to nitrite but not to N2. Positive for fermentation/oxidation of glucose and mannitol but negative for inositol, sorbitol, rhamnose, saccharose, amygdalin and arabinose. Melibiose (+) gave variable reactions. D-Salicin, α-D-glucose, D-mannose, D-galactose are used as sole energy sources. Does not utilize dextrin, D-raffinose, glycerol, N-acetyl-D-galactosamine, D-glucose-6-PO4, D-aspartic acid, D-serine, gelatin, glycyl-L-proline, L-alanine, L-arginine, L-aspartic acid, L-glutamic acid, L-pyroglutamic acid, L-serine, pectin, L-galactonic acid lactone, mucic acid, quinic acid, D-saccharic acid, p-hydroxy-phenylacetic acid, methyl pyruvate, D-lactic acid methyl ester, citric acid, D-malic acid, bromo-succinic acid, γ-amino-butyric acid, α-hydroxy-butyric acid, β-hydroxy-D,L-butyric acid, propionic acid, acetic acid and formic acid. The following reactions are variable within the species: citrate (−), D-maltose (−), D-trehalose (−), D-cellobiose (w), gentiobiose (−), sucrose (−), D-turanose (−), stachyose (−), α-D-lactose (−), D-melibiose (−), β-methyl-D-glucoside (−), N-acetyl-D-glucosamine (−), N-acetyl-β-mannosamine (−), N-acetyl neuraminic acid (−), D-fructose (−), 3-methyl glucose (w), D-fucose (w), L-fucose (w), L-rhamnose (−), inosine (−), D-sorbitol (−), D-mannitol (−), D-arabitol (−), myo-inositol (−), D-glucose-6-PO4 (−), L-histidine (w), D-galacturonic acid (−), D-gluconic acid (−), D-glucuronic acid (−), glucuronamide (w), L-lactic acid (−), α-keto-glutaric acid (w), L-malic acid (−), tween 40 (−) and acetoacetic acid (w). Does not assimilate any of the substrates included in the API 20 NE system. The most abundant cellular fatty acids are summed feature 3 , C16:0 (21.4%), C18:1ω7c (11.6%), C14:0 (5.2%), C12:0 and summed feature 2 , C12:03–OH(2.5%), C17:0 (1.6%), Iso-C17:0 (1.5%), Iso-C15:0 and C17:1ω8c (1.1%), and in minor amounts C13:0, C17:1ω6c, C18:0 and Unknown 12.484 (0.3–0.5%). The G + C content of the type strain (A-394T) is 48.2 mol%. The type strain is A-394T (= LMG 27910T = CAIM 1892T). It was isolated from the tissues of bleached Madracis decactis (Scleractinia) in St Peter & St Paul Archipelago, Brazil. SPSPASt Peter & St Paul ArchipelagoMLSAmultilocus sequence analysisAAIaverage amino acid identityDDHDNA-DNA hybridizationGGDCGenome-To-Genome Distance CalculatorFAMEfatty acid methyl ester analysesMALDI-TOFmatrix-assisted laser desorption/ionization time-of-flightColonies are small, beige, irregular shaped, with smooth and translucent edge and 1–2 mm in diameter after 24 h at 28 °C on MA under aerobic conditions. On TCBS colonies are green, round with a smooth border and 2–3 mm in diameter. Cells are small bacilli measuring 2–3 µm in diameter, Gram-negative, motile, facultative anaerobic, oxidase and catalase-positive. Grows well between 20 and 30 °C but not at 4 and 45 °C. No growth occurs in the absence of NaCl, but grows well under NaCl concentrations of 1%–8% (w/v). Grows at pH 6-11. Positive for alkaline phosphatase, leucine arylamidase, naphtol-AS-BI-phosphohydrolase, N-acetyl-10.7717/peerj.427/supp-1Table S1Upper GenBank accession numbers for the 16S rRNA gene, recA and rpoA housekeeping genes and genome sequences of Photobacterium sanctipauli sp. nov.; and for recA and rpoA of P. gaetbulicola LMG 27839T (data from this study). Lower Accession numbers for the Photobacterium strains’ genomes used for GGD calculation (data publicly available at GenBank).Click here for additional data file.10.7717/peerj.427/supp-2Table S2Photobacterium sanctipauli sp. nov. and source information.Strains of Click here for additional data file.10.7717/peerj.427/supp-3Table S3T (= LMG 27910T = CAIM1892T) genome determined in RAST environment.Statistics and general features of A-394Click here for additional data file.10.7717/peerj.427/supp-4Table S41, P. sanctipauli (range profile of five strains); 2, P. rosenbergii LMG 22223T . The evolutionary distances were computed using the Kimura 2-parameter method. Phylogenetic analyses were conducted in MEGA5. Bootstrap values (>50%) shown are based on 1,000 repetitions. Vibrio maritimus R-40493T was used as outgroup. Bar, 2% estimated sequence divergence.Neighbour-joining phylogenetic tree showing the position of Click here for additional data file.10.7717/peerj.427/supp-7Figure S2P. sanctipauli sp. nov based on rpoA gene sequences (969 bp). The evolutionary distances were computed using the Kimura 2-parameter method. Phylogenetic analyses were conducted in MEGA5. Bootstrap values (>50%) shown are based on 1,000 repetitions. Vibrio maritimus R-40493T was used as outgroup. Bar, 1% estimated sequence divergence.Neighbour-joining phylogenetic tree showing the position of Click here for additional data file.10.7717/peerj.427/supp-8Figure S3A−T394, A-373, A-379, A-397 and A-398) showing they are not clonal. The closely related type strains of P. rosenbergii (LMG 22223T), P. gaetbulicola (LMG 27839T) and P. lutimaris (LMG 25278T) were included in the analysis. The dendrogram was constructed using Pearson correlation coefficient and UPGMA.Comparison of the MALDI-TOF MS fingerprint profiles of the 5 novel strains (Click here for additional data file.


To evaluate CT-morphological features of lung cancers detected in a CT lung cancer screening trial, and to determine the correlation between CT-morphological features and the histopathological diagnosis of screen-detected cancers.197 solid lung cancers (192 participants) detected in all four screening rounds of the Dutch-Belgian randomized lung cancer screening trial (NELSON) were included. CT-morphological features included nodule shape, margin, location, volume, and volume-doubling time (VDT). Based on histopathology, cancers were divided into four groups: adenocarcinoma (N=114), squamous cell carcinoma (N=37), large cell carcinoma (N=28) and neuro-endocrine cancers (N=18). Data were analyzed using ANOVA, Chi-square and Fischer’s exact test.Mean participant age was 61.3 years , and 160/192 (83.3%) were male. In all four histopathologic groups, the majority of cancers had a spherical nodule shape (70.6-95.8%). Margins of malignant nodules were most often lobulated (39.3-45.9%) and spiculated (22.2-35.7%), without statistically significant difference between histopathological groups. Most cancers (63.5%) were located in the upper lobes, adenocarcinomas significantly more often  than other types of cancers (p=0.004). Adenocarcinomas had a higher mean VDT than large cell carcinomas . VDTs of other histopathological subgroups did not differ significantly.Only VDT and location in the upper versus lower lobes are associated with histopathological diagnosis of screen-detected lung cancers. No discrimination can be made between different histopathologic cancer types based on CT-morphological features alone.


Combination antiretroviral therapy (cART) may affect vitamin D [25(OH)D], parathyroid hormone (PTH), bone mineral density (BMD) and bone turnover (BT). Reduced BMD and secondary hyperparathyroidism have been reported with tenofovir (TDF). We investigated the associations between TDF and bone markers, especially in 25(OH)D-deficient patients.2) were measured using dual-energy X-ray absorptiometry. BT was assessed by serum type 1 procollagen (P1NP) and carboxy-terminal collagen cross-links (CTX). Mann–Whitney U tests compared serum markers and BT, and t-tests compared BMD according to TDF in all and 25(OH)D-deficient patients.In a single-centre longitudinal study investigating BMD in HIV-positive men, serum 25(OH)D, calcium, phosphate, PTH and alkaline phosphatase (ALP) were measured. Lumbar spine, non-dominant total hip and non-dominant femoral neck BMD  years, 94% white ethnicity, 93% MSM, diagnosed HIV positive for median 9.6  years, median CD4 547  cells/µL, HIV RNA <40 copies/mL in 87% (96% of those on cART). 25(OH)D (nmol/L) was normal (>75), insufficient (50–75), deficient (25–50) and severely deficient (<25) in 14%, 29%, 50% and 7%, respectively. Of 381 men on cART, 77% were currently on TDF. TDF was not associated with median calcium (p=0.69) or phosphate (p=0.52), but patients had higher  median ALP [81  vs. 73  IU/L, p=0.005) compared to non-TDF cART. There was no difference in the association between vitamin D and PTH according to whether someone currently was D deficiency, hyperparathyroidism, reduced BMD or increased BT, although patients on TDF had higher but normal ALP. We found no evidence to support additional monitoring of bone markers in patients on TDF regardless of 25(OH)D status.


Age is considered an independent risk factor for short and long-term mortality of elderly ICU patients, when very elderly (≥85 years) and mid-elderly (75-84 years) populations are compared to young elderly (65-74 years) ones. However, it is not yet clear if this trend remains when very elderly are compared to mid-elderly populations.a) to compare short  and long-term (6-months and 1-year) mortality of mid-elderly to very elderly patients andb) to evaluate the influence of patients' characteristics on mortality.Single center, retrospective, observational study in an 8-bed adult general ICU (January 2011-June 2014). Patients ≥75 years were divided into two age-groups, 75-84 and ≥85 years old. Characteristics on ICU admission were recorded. Patients hospitalized ≤48 hours were excluded. ICU, hospital, 6-months and 1-year after hospital discharge mortality were calculated. Univariate analysis for categorical variables was performed using Pearson’s x2 or Fisher test and Student t-test for continuous data. Multivariate analysis of the time to ICU mortality was calculated using Cox regression model and for hospital, 6-months and 1-year mortality using logistic regression analysis. P value < 0.05 was considered significant.244 patients were included, 195 in the 75-84 and 49 in the ≥85 years group. Mortality rates for the two groups were: ICU, 74/195 (37.9%) vs. 24/49 (49%) (p = 0.08), hospital, 23/121 (19%) vs. 5/25 (20%) (p = 0.90), 6-months, 15/95 (17%) vs. 4/20 (20%) (p = 0.74) and 1-year, 22/80 (27.5%) vs. 7/16 (20%) (p = 0.19), respectively. in multivariable analysis, patients with malignancy as reason for ICU admission had increased ICU mortality risk . Patients with higher APACHE II score on ICU admission had more important 1-year mortality risk .More than one third of our ICU patients were mid- or very elderly. No difference was observed between the two age-groups considering short or long-term mortality. Malignancy as a reason for ICU admission and APACHE II score negatively influenced the ICU and 1-year mortality, respectively.


H. pylori infection. H. pylori stimulate Wnt/β-catenin pathway by activating oncogenic c-Met and epidermal growth factor receptor (EGFR), or by inhibiting tumor suppressor Runx3 and Trefoil factor 1 (TFF1). H. pylori also trigger Wnt/β-catenin pathway by recruiting macrophages. Moreover, Wnt/β-catenin pathway is found involved in H. pylori-induced gastric cancer stem cell generation. Recently, by using gastroids, researchers have further revealed that H. pylori induce gastric epithelial cell proliferation through β-catenin. These findings indicate that Wnt/β-catenin is an oncogenic pathway activated by H. pylori. Therefore, this pathway is a potential therapy target for H. pylori-related gastric cancer.A section of gastric cancers presents nuclear β-catenin accumulation correlated with  Wnt/β-catenin pathway, also called canonical Wnt pathway, is crucial to embryo development and adult tissue homeostasis , 2. AberHelicobacter pylori (H. pylori) infection is a strong risk factor for gastric cancer. The underlying mechanisms include chronic inflammation in gastric mucosa, genetic and epigenetic alterations of tumor suppressor genes, activation of oncogenic signals, and generation of gastric cancer stem cells (CSC)  [H. pylori can cause DNA double-strand breaks directly [H. pylori-related gastric carcinogenesis [H. pylori induce gastric epithelial cell epithelial-mesenchymal transition (EMT), and generate potential cancer stem cells [H. pylori also stimulate some oncogenic pathways. Activation of epidermal growth factor receptor (EGFR) can resist H. pylori-induced gastric epithelial cell apoptosis [H. pylori-induced gastric carcinogenesis.) Figure . Chronic (BMDCs) , 10. H. directly , and caudirectly , 13 or bdirectly . Hypermeogenesis . H. pyloem cells , 17. H. poptosis , 19. Morβ-catenin exon 3 that encodes serine-threonine phosphorylation sites for the GSK3β [β-catenin mutation [APC mutation [APC methylation [Investigations on gastric cancer specimens showed that about 20%∼30% gastric cancers presented nuclear β-catenin accumulation . Some muhe GSK3β . These mmutation . In colomutation . Unlike mutation , 24 nor hylation  seemed tH. pylori to nuclear β-catenin accumulation. Nuclear β-catenin mainly localized in epithelial cells within the proliferative zone in antral glands, and appeared more frequently in H. pylori cytotoxin-associated gene A (CagA)-positive specimens, compared with either CagA-negative or uninfected patients [H. pylori could induce nuclear β-catenin accumulation in vivo and in vitro [H. pylori promoted gastric epithelial cell proliferation through β-catenin by using gastroids, three-dimensional organ-like structures [Several studies from a same group demonstrated the attribution of patients . CagA-poin vitro -28. Receructures .H. pylori induced phosphorylation of c-Met and gastric epithelial cell proliferation [H. pylori-induced gastric epithelial cell proliferation and atrophic gastritis [Aberrant activation of c-Met receptor occurred commonly in gastric cancers . Infectiferation , 32. Upoferation , 32. CD4astritis . C-Met-Pastritis . On the astritis . Interesastritis , 36, indH. pylori-related gastric cancer [H. pylori secretory protein HP0175 [H. pylori induced EGFR phosphorylation, and then activated PI3K/Akt pathway [H. pylori infection  was expressed in normal gastric mucosa , but free mucosa , 56. In nfection . TFF1 innfection . On the -catenin   [It has been well accepted that tumor-associated macrophages (TAMs) promote cancer development.  (MCP-1) , 59 or S (MCP-1)  in gastr (MCP-1) , 7  . The sup (MCP-1) . Macroph (MCP-1) . These oH. pylori. MiR-101 and miR-320 were inhibited by H. pylori through CagA [H. pylori [H. pylori infection with Wnt/β-catenin activation in gastric cancer (Table MicroRNAs (miRs) are small noncoding RNAs that can up- or downregulate the expression of oncogenes and tumor suppressors. Some miRs, such as miR-101, mir-124a, miR-203, miR-210 and miR-320, were downregulated by ugh CagA , 65. Hypugh CagA -68. The ugh CagA -73, indi. pylori -76. Thes. pylori -79. The H. pylori may activate Wnt/β-catenin pathway by affecting Wnt ligands, receptors or antagonists. H. pylori and TNF-α could induce Wnt10a and Wnt10b expression in gastric cancer cells [H. pylori infection could also activate Wnt co-receptor LRP6, and result in nuclear β-catenin accumulation [SFRP4 and SFRP5 methylation was found to be positively correlated with H. pylori infection [H. pylori on these molecules are still unclear.Some evidence indicates that er cells , 81. H. mulation . Secretemulation . Actuallnfection . In addinfection , Wnt7a [nfection , Wnt7b [nfection  and Wnt nfection , were alH. pylori colonized stomach gland epithelium, and promoted stem cell-related gene expression and Lgr5(+) stem cell proliferation [H. pylori also induced gastric epithelial cell EMT to generate gastric cancer stem cells, and this process was also via CagA [H. pylori-induced EMT and stem cell generation remain largely unknown. Wnt/β-catenin pathway was important for gastrointestinal progenitor cell proliferation and differentiation [H. pylori-induced gastric stem cell proliferation [H. pylori induce gastric stem cell generation and proliferation at least partly via Wnt/β-catenin pathway.Gastric stem cells are implicated in gastric cancer initiation and progression. Via CagA, feration . H. pylovia CagA . The molntiation , 91. Actntiation , 93. Recntiation . Moreoveferation . These fH. pylori in gastric carcinogenesis. H. pylori can upregulate Wnt/β-catenin activator c-Met and EGFR, and downregulate Wnt/β-catenin suppressor TFF1 and RUNX3. H. pylori can also activate Wnt/β-catenin pathway by recruiting tumor-associated macrophages. Importantly, via Wnt/β-catenin pathway, H. pylori induced gastric stem cell generation and expansion, promoting gastric cancer initiation and progression.Increasing evidence demonstrates Wnt/β-catenin as a crucial pathway stimulated by H. pylori infection, c-Met, EGFR, TFF1, Runx3, or else? Can dysregulations of these molecules synergize in gastric cancer development? Which virulent factor of H. pylori plays a dominant role in Wnt/β-catenin activation, CagA, VacA or else? Given the complexities of H. pylori strains and host factors, more work should be done to find the answers. In addition, the effects of H. pylori on Wnt ligands, receptors and antagonists, and the roles of miRs in Wnt/β-catenin activation in gastric cancer, need to be further investigated.However, there are still some questions need to be answered. Which signal molecule plays a dominant role in Wnt/β-catenin activation under H. pylori eradication can reduce the risk of gastric cancer, but it can not completely prevent H. pylori-related gastric carcinogenesis. One of the reasons is that the activation of oncogenic pathway, such as Wnt/β-catenin, has happened before H. pylori eradication.Recently, a series of therapies antagonizing Wnt/β-catenin pathway have entered clinical trials. As Wnt/β-catenin pathway is essential for tissue homeostasis, it remains elusive about their clinical efficacy and safety . H. pylo


Childhood-onset Systemic lupus erythematosus (cLES) is an autoimmune disease with a wide spectrum of clinical manifestations, characterized by periods of activity and remission. Hematological and immunological abnormalities are commonly found. Laboratory evaluation, including the cytokine profile, assists in the diagnosis, determination of disease activity and may predict future damage caused by the disease.To determine the serum levels of Th1 (IL-12), Th2 (IL-6 and IL-10) and Th17 (IL-17) cytokines in cSLE and healthy controls. To evaluate their association whit disease activity, laboratory and treatment features and Mood and anxiety disorders.We included 63 consecutive cSLE patients [median age 18 years (range 11-25)] and 59 healthy controls [median age 20 years (8-33)]. Controls were age and sex-matched to cSLE patients. SLE patients were further assessed for clinical and laboratory SLE manifestations, disease activity [SLE Disease Activity Index (SLEDAI)], damage [Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index (SDI)] and current drug exposures. Total dose of corticosteroids and other immunosuppressant medications used since the onset of disease were calculated by data obtained by careful review of the medical charts.Mood and anxiety disorders were determined through Becks Depression (BDI) and anxiety inventory (BAI). Th1 (IL-12), Th2 (IL-6 and IL-10) and Th17 (IL-17) cytokines levels were measured by ELISA and compared by non-parametric tests.Serum IL-6 (p=0.001), IL-10 (p=0.006), IL-12 (p=0.027) and IL-17 (p=0.0001) levels were increased in cSLE patients when compared to healthy controls. IL-6 levels were significantly increased in patients whit active disease (p=0.008). IL-6 (p=0.032) and IL-12 (p=0.028) levels were significantly increased in patients with active nephritis. We observed that IL-17 was associated with migraine (p=0.045), IL-6 with thrombocytopenia (p=0.022) and IL-12 with the presence of anxiety (p=0.048). No association between cytokine levels and SDI scores or medication was observed.Cytokines play a central role in cSLE, IL-6 is associated with SLEDAI and may be a biomarker of disease activity, Th1 and Th2 responses may play a role in lupus nephritis and Th1 and Th17 may play a role in neuropsychiatric symptoms in cSLE. Longitudinal studies are necessary to confirm their ability to predict SLE related manifestations.K. Peliçari: None declared, M. Postal: None declared, R. Barbosa: None declared, N. Sinicato: None declared, F. Peres: None declared, R. Marini: None declared, S. Appenzeller Grant / Research Support from: FAPESP  CNPQ


Bacillus decisifrondis E5HC-32T (DSM 11725T) is a Gram-positive, subterminal spherical spore-forming, strictly aerobic bacterium. Here, we report the 5,613,728-bp genome sequence of B. decisifrondis E5HC-32T, which is the first genome information of this species and will provide useful information for genomic taxonomy and phylogenomics of Bacillus-like bacteria. T was isolated from soil in Australia and was named as Bacillus decisifrondis sp. nov. .The type strain E5HC-32sp. nov. . Its phyd pH 8.4 . NotablyB. decisifrondis E5HC-32T (DSM 11725T) was performed via the Illumina Hiseq 2500 system. Two DNA libraries with insert sizes of 500 bp were constructed and sequenced. After filtering of the 0.67-Gb raw data, 0.66-Gb of clean data was obtained, providing approximately 100-fold coverage. The reads were assembled via SOAPdenovo software version 2  utilizing GeneMark, Glimmer, and tRNAscan-SE tools (The annotation of the genome was performed using the NCBI Prokaryotic Genomes Automatic Annotation Pipeline (PGAAP) (SE tools . A totalSE tools .LIYL00000000. The version described in this paper is version LIYL00000000.1.This whole-genome shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession no.


Bacillus sp. SM1 strain MCC2138), which was isolated from marine coastal waters and has the ability to degum raw silk fabric as well as Ramie fiber. The genome comprises 1.76 Mb with a GC content of 34.5%.Reported here is the draft genome sequence of an amylase-, protease-, DNase-, oxidase-, gelatinase-, and catalase-producing, Gram-positive diplobacillus ( It could accumulate Pb, Cr, Ni, Cu, and Co  with a 2.0× coverage generated on an Illumina platform, totaling 17,69,494 bp, with a largest contig length of 5,01,909 bp and T server  revealedThe important genes contained in the different contigs are as follows: ABC transporter ATP binding protein; redox-sensitive transcriptional regulator (AT-rich DNA binding protein); heat shock protein 60 family cochaperone GroES/GroEL; ATP pyrophosphatase subunit (EC 6.3.5.2); phosphate regulon transcriptional regulatory protein PhoB (SphR) (accession no. LXJX01000001.1); oligopeptide ABC transporter, periplasmic oligopeptide-binding protein OppA/B/C/D/F (TC3.A.1.5.1); hydrolase, tRNA; glycerate kinase (EC2.7.1.31); arginase (EC3.5.3.1); diadenylate cyclase spyDAC; bacterial checkpoint controller DisA with nucleotide-binding domain; phosphoglucosamine mutase ; programmed cell death toxin YdcE and YdcD; outer membrane lipoprotein receptor; Holo (acyl carrier protein) synthase (EC2.7.8.7); UV endonuclease UvdE family; DEAD box ATP-dependent RNA helicase CshA (EC3.6.4.13); t-RNA (accession no. NZ_LXJX01000007.1); ABC transporter permease protein YvcS; ABC transporter ATP-binding protein YvcR; beta-lactamase class A; secreted and spore coat–associated protein 1; chitinase (EC 3.2.1.14); cell-division protein FtsW; microbial collagenase (EC 3.4.24.3); transcriptional regulator, TetR family (accession no. NZ_LXJX01000017.1); cysteine synthase (EC 2.5.1.47); chaperonin (heat shock protein 33); tRNA (Ile) lysidine synthetase (EC 6.3.4.19); AbrB family transcriptional regulator (SpoVT); transcription repair coupling factor; Fin, required for the switch from sigma F to sigma G during sporulation; peptidyl-tRNA hydrolase (EC 3.1.1.29) (accession no. NZ_LXJX01000019.1).LXJX00000000. The version described in this paper is the first version, LXJX01000000.This whole-genome shotgun project has been deposited at DDBJ/ENA/GenBank under the accession number


AbstractDulichiella pattaniensis is new to science, and Melitalati latiflagella Ren & Andres, 2012 has not been previously reported from Thai Waters. Dulichiella pattaniensis is characterized by male gnathopod 2 distolateral crown with 4 spines; pleonite/urosomite formula 7-7-7-5-6-2; pereopod 5-7 dactylus with 2 accessory spines. This combination of characters has not been recorded previously in the Dulichiella. The characters of the specimens are described and illustrated. All specimens are deposited in the Princess Maha Chakri Sirindhorn Natural History Museum, Prince of Songkla University, Thailand.Two species of melitid amphipod were collected from the Gulf of Thailand.  Rotomelita longipropoda Wongkamhaeng et al., 2013 was described. In this study, we describe a new melitid species Dulichiella pattaniensis sp. n., and our observations of Melita latiflagella Ren & Andress, 2012, which has not been previously reported in Thai Waters. Figures and descriptions of both species are provided.Melitid amphipods most commonly occur in coastal and freshwater areas. Thailand has a variety of aqueous habitats including coral reefs, seagrass beds, and mangrove forests, but only one melitid amphipod, , antennaA; , gnathopodG; , headHD; , lower lipLL; , mandibleMD; , maxillaMX; , maxillipedMP; , pereopodP; , pleopodPl; , telsonT; , uropodU; , urosomeUR; , upper lipUL; , rightr; , leftl; , male♂; , female♀. Specimens of different species were deposited into the Prince of Songkla University Zoological Collection (PSUZC).Amphipods were collected from some settlement plates in an artificial reef in Ban Pak Bang Ta Wa, Pattani Bay and from sediment of Lower Songkhla Lake . The sitStout, 1912http://species-id.net/wiki/DulichiellaPageBreakmales with bunches of long slender setae. Pereopod 7 basis in female fully expanded. Pleonites dorsally serrate. Uropod 3 inner ramus scale-like; outer ramus 4 to 5× longer than wide, 2-articulate. Telson deeply cleft, lobes tapering distally to an acute point. Head antDulichiella spinosa Stout, 1912 (type by monotypy).Dulichiella appendiculata Say, 1818; Dulichiella australis Haswell, 1879; Dulichiella celestun Paz-Rios & Ardisson, 2014; Dulichiella cotesi Giles, 1890; Dulichiella cuvettensis Appadoo & Myers, 2005; Dulichiella fresnellii ; Dulichiella guinea Lowry & Springthorpe, 2007; Dulichiella lecroyae Lowry & Springthorpe, 2007; Dulichiella oahu Lowry & Springthorpe, 2007; Dulichiella pacifica Lowry & Springthorpe, 2005; Dulichiella pattaniensis sp. n.; Dulichiella spinosa Stout, 1912 (type species); Dulichiella takedai Tomikawa & Komatsu; Dulichiella terminos Lowry & Springthorpe, 2007; Dulichiella tomioka Lowry & Springthorpe, 2007; Dulichiella tulear Lowry & Springthorpe, 2007.PageBreakhttp://zoobank.org/212F51D8-E32A-4601-9E04-88292FBEB989http://species-id.net/wiki/Dulichiella_pattaniensis6°51'55"N, 101°10'7"E), artificial reef , 1 September 2010, Puttapreecha, R., PSUZC-CR-0192. Allotypes, ♀ collected with holotype; PSUZC-CR-0193; Paratype, collected with holotype ).Holotype. ♂, THAILAND, Lower Gulf of Thailand, Pattani Bay , flagellum can be 25-articulate . Antenna 2, Antenna 2 setiferous, peduncular article 2 cone gland not reaching end of article 3; article 5 subequal to 4, flagellum 9-articulate.Based on male holotype. Body length 6.3 mm (from tip of rostrum to apex of telson). Upper lip, (labrum) distally rounded. Lower lip, inner lobes well developed, pubescent. Mandible, both similar, left incisor 3 dentates, right incisor 4 dentate; left and right lacinia mobilis armed with 3 and 4 dentates respectively; palp slender with marginal setae, article 1 smooth, article 3 slightly longer than article 2. Maxilla 1, inner plate narrow with 2 apical plumose setae; outer plate with 8 apical serrate robust setae; palp 2-articulate, article 1 with 3 distal setae, article 2 with 4 apical robust setae and 4 apical setae. Maxilla 2, inner plate with mediofacial row of 29 setae and 14 apical plumose setae; outer plate broader than inner plate, distally setose. Maxilliped, inner plate borad, with 6 plumose marginal setae; outer plate margin with 11 conate robust setae, terminal with 4 plumose setae; palp 4-articulate, article 2-3 with marginal setae, article 4 tapering with fine marginal setae.Pereon.PageBreakPageBreakPageBreakPageBreakPageBreakPageBreakPageBreakPageBreakPageBreakPageBreakGnathopod 1 subchelate, smaller than gnathopod 2; coxa anterodistal corner not produced, posteroventral corner notch present, anterior margin straight; length ratio of articles from basis to dactylus 10: 3:4:7:5: 3; basis slender; merus–propodus setose; palm slightly convex, defined by posterodistal corner, without posterodistal robust setae. Gnathopod 2 sexually dimorphic; left and right gnathopods unequal in size; coxa posteroventral corner notch present; (larger) length ratio of article from basis to dactylus 8:1:3:1:10:11; propodus distolateral corner crown with 4 rounded spines, palm straight, posterodistal corner produced, upturned, fit with dactylus; dactylus apically blunt;  subchelate; length ratio of article from basis to dactylus 9:3:4:6:6:4; merus with sharp posteroventral spine; carpus subequal to propodus; palm straight, without posteroventral spine. Pereopod 3–4 alike. Pereopod 5 basis posterior margin straight, posteroventral corner rounded; carpus and propodus sparsely setose; dactylus unguis anterior margin with accessory spines. Pereopod 6–7 alike, basis, merus, carpus, propodus with long marginal setae. Pereopod 6 basis posterior margin straight, minutely castelloserrate; dactylus unguis anterior margin with accessory spines. Pereopod 7 basis posterior margin straight, with posterior margin minutely castelloserrate, posteroventral corner; dactylus unguis anterior margin with accessory spines.Pleon. Pleonite/urosomite dorsal spine formula (7-7-7-5-6-2). Pleonites 1–3 with dorsal setae. Epimera 1–3 posteroventral margin without spines above posteroventral corner. Epimeron 3 posterior margin smooth, posteroventral corner with strongly produced acute. Urosomite 1 with spine at midline, no medial gape. Urosomite 2 with dorsal setae. Urosomite 3 with dorsal setae, with 2 dorsal spines. Uropod 3 inner ramus scale-like, much shorter than outer ramus; outer ramus much longer (more than 2× length) than peduncle, 2-articulate. Telson with dorsal robust setae.Female. . Length, 7.4 mm. Gnathopod 1 coxa 4 anterodistal corner not produced, posteroventral corner notch present, anterior margin excavated. Gnathopod 2 equal, coxa subrectangular, palm crenulated, oblique with setae on margins. Pereopod 7 basis expanded, posterior margin slightly convex.This species is named after the type locality.Dulichiella pattaniensis sp. n., with pleonite/urosome formular of 7-7-7-5-6-2 has only Dulichiella cotesi, Dulichiella oahu, Dulichiella pacifica and Dulichiella tulear that share this characters. This new species can be distinguished from Dulichiella cotesi, Dulichiella oahu and Dulichiella tulear by having male gnathopod 2 (large) with 4 spines on distolateral crown while those three species have 3 spines. Dulichiella pattaniensis differs from Dulichiella pacifica in the following: the male large gnathopod 2 has a fourth spine on its distolateral crown that is not well developed vs. its well-developed fourth set of spines. The male gnathopod 1 has carpus longer than its propodus vs. a carpus subequal to its propodus. Pereopods 3-4 have a dactyli with 2 accessory spines vs. 3-4 dactyli with 1 accessory spine.Dulichiella appendiculata, Dulichiella cuvettensis, Dulichiella celestun, Dulichiella fresnellii, Dulichiella guinea, Dulichiella lecroyae, Dulichiella pacifica and Dulichiella takedai share this distinct character. Dulichiella pattaniensis can be distinguished from amphipods by having pleonite/urosome formular of 7-7-7-5-6-2 while Dulichiella appendiculata, Dulichiella cuvettensis, Dulichiella fresnellii, Dulichiella lecroye and Dulichiella pacifica pleonite/urosome formula 7-7-7-5-4-2, Dulichiella guinea 9-9-7-5-4-2 and Dulichiella celestun 9-9-9-5-6-2. Moreover, Dulichiella pattaniensis differs from Dulichiella cuvettensis, Dulichiella guinea and Dulichiella lecroyae by having 2 accessory spines on pereopods 3-4 dactyli vs. 1 accessory spine. A summary of these distinguishing characters are given in The new species also has four spines on the distolateral crown of the male gnathopod 2. Only 7 species, Ren & Andress, 2012http://species-id.net/wiki/Melita_latiflagella09°18'39.5"N, 99°46'46.4"E), 1 Feb 2012, Wongkamhaeng, K. PSUZC-CR-0191. .Lower Gulf of Thailand, Songkhla Lake .


Hyperoxia is frequently encountered in the intensive care unit (ICU) and during surgical procedures such as coronary artery bypass surgery (CABG). Higher oxygen concentrations intuitively provide a salutary oxygen reserve, but hyperoxia can induce adverse effects such as systemic vasoconstriction, reduction of cardiac output, increased microcirculatory heterogeneity and increased reactive oxygen species production. Previous studies in patients undergoing CABG surgery suggest reduced myocardial damage when avoiding extreme perioperative hyperoxia (>400 mmHg). Here, we compare moderate hyperoxia to near-physiological values.2s from moderate hyperoxia to near-physiological values decreases myocardial damage and organ injury.To investigate whether an oxygenation strategy towards lowering perioperative PaO2 target according to common practice (200-220 mmHg during cardiopulmonary bypass (CPB) and 130-150 mmHg in the ICU) versus a lower PaO2 target . The primary outcome was myocardial damage (CK-MB and Troponin-T), which was determined before surgery, at ICU admission and 2, 6 and 12 hours thereafter.In this single-center, open-label randomized-controlled trial, 50 patients scheduled for elective isolated CABG surgery were allocated to either a PaO2 during CPB was 220 mmHg, IQR (211-233) vs. 157 , respectively. In the ICU, weighted PaO2 was 107 (86-141) vs. 90 . Median maximum values of CK-MB were 25.8µg/L, IQR (20.3-32.6) vs. 24.9  and of Troponin-T 0.35 µg/L, IQR (0.30-0.46) vs. 0.42. Areas under the curve (AUC) of CK-MB  and 0.30µg/L/h (0.25-0.44) vs. 0.39  for Troponin-T. Cardiac Index, Systemic Vascular Resistance Index, and serum lactate levels (Lactatemax median 2 mmol/L IQR(1.4-2.6) vs. 2.2) were similar between groups throughout the ICU period.Baseline and surgery characteristics were not different between groups. The mean age of patients was 66 years (SD 8) vs. 68(6), respectively. Mean duration of CPB was 105 minutes (SD 24) vs. 108(28). Weighted PaOIn the present RCT, an oxygenation strategy towards near-physiological arterial oxygen tensions did not reduce myocardial damage in comparison to moderate hyperoxia. On the other hand, conservative oxygen administration did not lead to increased lactate levels.This investigation was supported by grants from ESICM NEXT Start-Up 2014 and ZonMW


To seek for possible differences in tryptase values during acute anaphylaxis induced by non-steroid anti-inflammatory drugs (NSAID) related to the suspected mechanism (selective vs non-selective reactions).Retrospective review of patients attending the Emergency Department (2009-2013) for a NSAID induced anaphylaxis and with a tryptase determination. Patients were split into 3 groups according to the allergy work-up ResultsSelective reactions (S) were defined by positive skin results and/or negative challenges with other NSAID; non-selective (NS) reactions were defined by the existence of symptoms with several NSAID or after positive challenge results. Those cases who did not comply with the previous conditions were non-conclusive (NC). We analysed: demographics, severity of the episode (Chi Square test), and rate of tryptase higher than 11.4 mcg/l (Chi Square test), median tryptase values and timing of sample obtention .86 patients were included: S group: 36 patients, NS group: 20 patients, and NC: 30 patients. Median ages were 48, 45 and 54 years, respectively. Severity of the episodes: S group: Mild: 22.2%, moderate: 30.6% and severe: 47.2%; NS group: Mild: 30%, moderate: 35% and severe: 35%; NC group: Mild: 16.7%, moderate: 26.7% and severe anaphylaxis: 56.7% (p=0.658). The most frequent eliciting drugs were metamizole , acetic derivatives  and propionic acid derivatives . The median of tryptase determinations were: S group: 13.4(2.75-98.6), NS: 12.35(3.85-53.6) and NC: 13.4mcg/l (2.82-57.1) (p=0.856). Median duration of symptoms: S group: 1.5 (0.33-12), NS: 1.75 (0.5-4); NC: 1.5 hours (0.5-6h) (p=0.582).Although there have been reports of differences in tryptase releases during anaphylaxis related to different allergens, we have found no differences among the different patterns of NSAID reactions that may help us to avoid future challenges for diagnostic confirmation.


Rictor to impair mTORC2 activity in mouse mammary epithelia, which decreased Akt phosphorylation, ductal length, secondary branching, cell motility, and cell survival. These effects were recapitulated with a pharmacological dual inhibitor of mTORC1/mTORC2, but not upon genetic disruption of mTORC1 function via Raptor deletion. Surprisingly, Akt re-activation was not sufficient to rescue cell survival or invasion, and modestly increased branching of mTORC2-impaired mammary epithelial cells (MECs) in culture and in vivo. However, another mTORC2 substrate, protein kinase C (PKC)-alpha, fully rescued mTORC2-impaired MEC branching, invasion, and survival, as well as branching morphogenesis in vivo. PKC-alpha-mediated signaling through the small GTPase Rac1 was necessary for mTORC2-dependent mammary epithelial development during puberty, revealing a novel role for Rictor/mTORC2 in MEC survival and motility during branching morphogenesis through a PKC-alpha/Rac1-dependent mechanism.Akt phosphorylation is a major driver of cell survival, motility, and proliferation in development and disease, causing increased interest in upstream regulators of Akt like mTOR complex 2 (mTORC2). We used genetic disruption of  The protein kinase mTOR is frequently activated in breast cancers, where it enhances cancer cell growth, survival, and metastastic spread to distant organs. Thus, mTOR is an attractive, clinically relevant molecular target for drugs designed to treat metastatic breast cancers. However, mTOR exists in two distinct complexes, mTORC1 and mTORC2, and the relative roles of each complex have not been elucidated. Moreover, as pathways that regulate normal tissue growth and development are often highjacked to promote cancer, understanding mTOR function in normal mammary epithelial development will likely provide insight into its role in tumor progression. In this study, we assessed the role of mTORC1 and mTORC2 complexes in normal mammary epithelial cell branching, survival, and invasion. Interestingly, while mTORC1 was not required for branching, survival and invasion of mammary epithelial cells, mTORC2 was necessary for these processes in both mouse and human models. Furthermore, we found that mTORC2 exerts its effects primarily through downstream activation of a PKC-alpha-Rac1 signaling axis rather than the more well-studied Akt signaling pathway. Our studies identify a novel role for the mTORC2 complex in mammary morphogenesis, including cell survival and motility, which are relevant to breast cancer progression. Post-natal mammary epithelial morphogenesis is a complex process during which an extensively branched ductal network develops from a rudimentary epithelial bud . Branchimammalian target of rapamycin (mTOR) regulates cellular metabolism, protein and lipid synthesis, cell survival, and cytoskeletal organization, processes that are required for proper mammary morphogenesis. mTOR regulates these processes through phosphorylation of its target substrates, including translation initiation factor 4E (eIF4E)-binding protein 1 (4E-BP1), p70S6 kinase (S6K), Akt, SGK1, and protein kinase C-alpha  . For some experiments, cells were maintained for 24 hour in Starvation Media [Growth Media without serum or EGF] prior to stimulation and/or analysis. PKC-alpha inhibitor GO6976  and adenoviral particles  were purchased. Cells were incubated with adenoviral particles (5 X 108 particle forming units/ml) for 3–5 hours at 37°C and cells were allowed to recover for 48 hours prior to experimental analysis.MCF10A and MCF10A Rictor2HPO4, 3.5 millimolar NaH2PO4, 7.7 millimolar NaN3, 0.1% bovine serum albumin, 0.2% Triton-X 100, 0.05% Tween-20] and stained with rabbit anti-pan-cytokeratin  and AF621-goat anti-rabbit (1:100), counterstained with TO-PRO-3 Iodide (Invitrogen), and imaged using the Vanderbilt Cell Imaging Shared Resource Zeiss LSM 510 confocal microscope and LSM Image Browser software. For E-cadherin staining, the primary antibody used was anti-E-cadherin (BD Transduction Laboratories) and visualized with anti-mouse Alexa 594 secondary antibody . Confocal images of 3D structures were visualized using an LSM 510 META inverted confocal microscope with a 20X/0.75 plan apochromat objective.Matrigel-emdedded organoids cultured on coverslips were fixed 8 minutes in 1:1 methanol:acetone at -20°C, permeabilized in 0.5% Triton-X 100/PBS for 10 min, blocked , sonicated 10 seconds, and cleared by centrifugation at 4°C, 13,000 x g for 5 min. Protein concentration was determined using BCA (Pierce). Proteins were separated by SDS-PAGE, transferred to nitrocellulose membranes, blocked in 3% gelatin in TBS-T [Tris-buffered saline, 0.1% Tween-20), incubated in primary antibody overnight and in HRP-conjugated anti-rabbit or anti-mouse for 1 hour, and developed using ECL substrate (Pierce). Antibodies used: alpha-actin ; AKT and S473 P-Akt ; S6 and P-S6 ; Rictor ; Raptor ; Rab11 ; PKC-alpha and T638/641 P-PKC-alpha ; Rac . GST-Pak-PBD effector pulldown assays were performed using reagents from Millipore as per manufacturer’s protocol.Cells and tissues were homogenized in ice-cold lysis buffer [50 millimolar Tris pH 7.4, 100 millimolar NaF, 120 millimolar NaCl, 0.5% NP-40, 100 micromolar NaIn situ TUNEL analysis was performed on paraffin-embedded sections using the ApopTag kit . IHC on paraffin-embedded sections was performed as described previously [Mammary glands were whole-mounted on slides, cleared of adipose, and stained with hematoxylin as described previously . Sectioneviously  using: KMethanol-fixed PMECs were probed 1 hour with GST-PBD (Millipore) diluted 1:50 in PBS. GST (lacking PBD) was used as a negative control. Samples were washed then probed with AF488-conjugated anti-GST (1:100), stained with DAPI or AF621-phalloidin, and mounted.5) were added to upper chambers of Matrigel-coated transwells in starvation medium and incubated 5 hours to score migration in response to 10% serum-containing medium in the lower chamber. Filters were swabbed and stained with 0.1% crystal violet, [MECs  has been accredited by AAALAC International since 1967 (File #000020). The AAALAC Council on Accreditation's most recent review of VU's program was done in 2011 and resulted in "Continued Full Accreditation.” Isofluorane was used for anesthesia, as well as euthanasia. For human euthanasia, cervical dislocation was used following isofluorane overdose.S1 FigA. Hematoxylin stained whole mount preparations from 6 week old RictorWT mice and RictorMGKO mice. High magnification panels show irregular ductal tracts and increased staining density in RictorMGKO samples, indicative of multiple cell layers. Data are a representation of 11 independent animals/genotype. B. Hematoxylin stained whole mount preparations from 10 week old wild-type RictorWT mice and RictorMGKO mice show sustained length defects in the ductal tracts of mammary glands lacking Rictor. C. IHC for Ki67 or TUNEL in TEBs from 6 week old wild-type RictorWT mice and RictorMGKO mice. Average percent Ki67 and TUNEL+ nuclei (± S.D.) was determined. D. Confocal analysis of primary organoids (PMOs) stained for E-cadherin (red) revealed multiple cell layers in acinar structures and poor lumen formation in Rictor-deficient (RictorFL/FL organoids infected with Ad.Cre) PMOs relative to control PMOs (RictorFL/FL organoids infected with Ad.LacZ), consistent with the phenotype in RictorMGKO epithelium in vivo.(TIF)Click here for additional data file.S2 FigRictorFL/FL mice and infected with Ad.GFP or Ad.Cre. A. Fluorescent imaging of organoids 24 hours post-infection. B. Confocal analysis of WT organoids infected with Ad.GFP. C. Organoids were fixed 10 days post-infection and subjected to immunofluorescent staining using a pan-cytokeratin antibody (red) to confirm epithelial identity and ToPro-3 nuclear marker (blue). While Ad.LacZ-infected RictorFL/FL organoids formed hollow lumens surrounded by an organized epithelial layer, Ad.Cre-infected RictorFL/FL organoids remained rounded and disorganized. Data are a representation of 3 independent organoid isolates. D. MCF10A parental or MCF10A-RictorZFN cells were assessed for invasion through Matrigel-coated transwell filters. The cells remaining in the upper chamber after 24 hours were fixed and stained with either DAPI (blue) or Annexin V-FITC (green). The average percent viable cells (± S.D.) left in the upper chamber was quantitated using Image J, Student’s T-test.Primary organoids (PMOs) were isolated from (TIF)Click here for additional data file.S3 FigA-C. PMECs and organoids from RictorFL/FL mice were coinfected with Ad.Cre and either Ad.LacZ or Ad.AktDD. A. Western analysis of PMEC lysates. B. Organoids photographed after 10 days in Matrigel culture. Average number of branches/organoid ± S.D. (right panel) is shown. N = 7 independent organoid isolates, analyzed in triplicate. Midline values indicate average, whiskers indicate S.D., Student’s T-test. C. Colony size of organoids (± S.D.) measured in pixel area. N = 7 independent organoid isolates, analyzed in triplicate, Student’s T-test.(TIF)Click here for additional data file.S4 FigA. Immunofluorescent detection of P-PKC-alpha in mammary gland sections from 6 week old virgin RictorWT and RictorMGKO mice. B.In situ detection of GTP-only control (no PBD) via IF detection of GST-PBD  on mammary gland sections from 6-week old mice. C.WT PMECs cultured ± Rac inhibitor or RictorFL/FL PMECs infected with Ad.LacZ or Ad.Cre were assessed for GTP-bound Rac via IF detection of GST-PBD . D.RictorFL/FL PMECs infected with Ad.Cre or Ad.LacZ were probed with GST-PBD (green) and counterstained with phalloidin (red). Representative images are shown. E.RictorFL/FL PMECs infected with Ad.Cre or Ad.LacZ (±Ad.caRac1) were analyzed for TUNEL. F. Whole mount analysis of endogenous mammary glands harvested from WT recipient mice.(TIF)Click here for additional data file.S5 FigWT PMECs infected with Ad.LacZ or Ad.caRac1 were grown to confluence (3–5 days after infection) then wounded using a P200 pipette tip in the presence of DMSO or rapamycin. Monolayers were imaged after 24 hours and total wounded area (arbitrary units) remaining was measured at 24 hours after wounding. Values shown are the average ± S.D. N = 6 per time point.(TIF)Click here for additional data file.S6 FigA. Western analysis of whole mammary gland lysates from 10 week old mice. B. IF for CK8, CK14 and ZO-1 in mammary gland sections. C. IHC for Ki67 or TUNEL in TEBs from 6 week old wild-type RaptorWT mice and RaptorMGKO mice. Average percent Ki67+ and TUNEL+ nuclei (± S.D.) was determined, Student’s T-test.(TIF)Click here for additional data file.


Cichorium intybus L. (chicory) in acute liver injury. Pathological observation, reactive oxygen species (ROS) detection and measurements of biochemical indexes on mouse models proved hepatic protective effect of Cichorium intybus L. Identification of active compounds in Cichorium intybus L. was executed through several methods including ultra performance liquid chromatography/time of flight mass spectrometry (UPLC-TOF-MS). Similarity ensemble approach (SEA) docking, molecular modeling, molecular docking, and molecular dynamics (MD) simulation were applied in this study to explore possible mechanisms of the hepato-protective potential of Cichorium intybus L. We then analyzed the chemical composition of Cichorium intybus L., and found their key targets. Furthermore, in vitro cytological examination and western blot were used for validating the efficacy of the selected compounds. In silico analysis and western blot together demonstrated that selected compound 10 in Cichorium intybus L. targeted Akt-1 in hepatocytes. Besides, compound 13 targeted both caspase-1 and Akt-1. These small compounds may ameliorate liver injury by acting on their targets, which are related to apoptosis or autophagy. The conclusions above may shed light on the complex molecular mechanisms of Cichorium intybus L. acting on hepatocytes and ameliorating liver injury.This study aimed at investigating the possible mechanisms of hepatic protective activity of  Cichorium intybus L.  is a perennial herb from the Asteraceae family with both alimentary use and medicinal use. It is 40–100 cm in height, usually with bright blue flowers, rarely white or pink. It has been reported that fresh chicory typically contains 68% inulin, 14% sucrose, 5% cellulose, 6% protein, 4% ash, and 3% other compounds, while dried chicory contains approximately 98% inulin and 2% other compounds + ; 1H-NMR  δ: 7.07 , 6.72 , 3.66 , 3.52 ; 13C-NMR  δ: 173.1 (C-8), 156.2 (C-4), 129.9 , 124.5 (C-1), 114.9 , 50.9 (C-1ʹ), 39.5 (C-7).Compound 13: colorless acicular crystal (MeOH); HR-ESI-TOF-MS 153.0549 [M + H]+ ; 1H-NMR δ: 7.00 , 6.67 , 3.46 , 9.27 ; 13C-NMR  δ: 176.1 (C-8), 157.4 (C-4), 131.3 , 126.8 (C-1), 116.2 , 41.1 (C-7).Compound http://sea.bkslab.org/) [http://www.geneontology.org) and KEGG Orthology (KO) (http://www.genome.jp/kegg/ko.html) [Target proteins of the small molecule compounds were obtained from the similarity ensemble approach (SEA) (ab.org/) . An assiko.html) ,23,24. Fhttp://www.pdb.org/pdb/home/home.do). Molecular docking was performed using the UCSF DOCK6.5 program , which utilized DOCK algorithm to address rigid body docking by superimposing the ligand on to a negative image of the binding pocket [We selected 2 effective small molecule compounds and 2 proteins as receptors. Before the docking, we downloaded initial three-dimensional geometric co-ordinates of the X-ray crystal structures of compounds and proteins from the Protein Data Bank (PDB)  simulations. MD simulation of the complex was carried out with the GROMACS 4.5.4   package 3, and a protease inhibitor cocktail . Following one freeze–thaw cycle, the cell lysates were centrifuged at 10,000× g at 4 °C for 20 min and boiled in sample buffer for 5 min. The proteins were then subjected to SDS–PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) and transferred to a PVDF (polyvinylidene fluoride) membrane  using a semi-dry electroblotting system. After blocking with 5% skim milk in PBS, the membranes were incubated with primary antibodies (1:1000 dilution) at 4 °C overnight. The membranes were then washed with PBS containing 0.05% Tween 20 and incubated with horseradish peroxidase (HRP)-conjugated secondary antibodies (1:5000 dilution) at room temperature for 1 h. After washing, the membranes were soaked in ECL solution  for 1 min, according to the manufacturer’s instructions, and exposed to film . Other materials used for protein analyses were purchased from Sigma-Aldrich  [The expression levels of the selected target proteins (Akt-1 and caspase-1) in liver tissues were measured by applying western blot. Liver tissues were lysed in a buffer containing 1% Triton X-100, 50 mM Tris (pH 7.5), 10 mM EDTA, 0.02% NaNMO, USA) ,31,32.t-test. p < 0.05 was considered statistically significant.All the presented data and results were confirmed in at least three independent experiments. The data are expressed as means ± S.D. Statistical comparisons were made by One-way ANOVA and Student’s Cichorium intybus L. and active compounds in Cichorium intybus L. The active compounds ameliorate liver injury by acting on Akt-1 and caspase-1, which are related to apoptosis or autophagy. In summary, our results may shed light on the complex molecular mechanisms of Cichorium intybus L. acting on hepatocytes and ameliorating liver injury.This study demonstrates the hepatic protective effect of


The oral antihyperglycemic agent metformin was associated with favorable outcome and smaller myocardial infarct size in patients with diabetes undergoing percutaneous coronary interventions (PCI) for ST-segment elevation myocardial infarction (STEMI) , 2. HoweTo determine the effect of chronic metformin treatment on cardiovascular morbidity and mortality in patients with diabetes presenting with STEMI subsequently undergoing PCI.From January 2004 until June 2013, all consecutive critically ill patients undergoing primary PCI for STEMI at the University Medical Center Groningen were included in a registry and 1-year follow-up was obtained. Our primary endpoint consisted of the composite endpoint of myocardial infarction, target vessel and target lesion revascularization, and all-cause mortality (MACE). The secondary endpoint, myocardial infarction size, was estimated using peak levels of creatine kinase (CK), the myocardial band of CK (CK-MB), troponin T, and high-sensitive troponin T (hs-troponin T). The effect of metformin on myocardial infarct size from the 2004-2010 cohort has been reported previously [In total, 4776 consecutive patients underwent primary PCI for STEMI, 719 (15%) diabetic patients were included in the final analysis and 215 (30%) patients used metformin at admission. MACE and mortality rates were 21% and 12% for patients with diabetes, 23% and 19% for metformin patients, 21% and 15% for patients on sulfonylurea, and 30% and 20% for patients on insulin, respectively. Metformin was not associated with reduced risk for MACE (adjusted hazard ratio (aHR): 1.19  0.78-1.81), P = 0.42) or survival benefit (aHR: 0.23 (CI95% 0.80-2.51), P = 0.23) compared to diabetic patients not using metformin. Insulin use was an independent predictor for MACE (aHR 1.73 (CI95% 1.13-2.65), P = 0.01) and all-cause mortality (aHR 1.81 (CI95% 1.03-3.21), P = 0.04). Baseline levels of CK, CK-MB, and hs-troponin T were comparable between both groups. Median (interquartile range) peak levels of CK, CK-MB, and hs-Troponin T were all non-significant lower in the metformin group (table Chronic metformin use in patients presenting with STEMI was associated with smaller infarct-size and not with lower MACE and mortality rates, as compared to other patients with diabetes.


To evaluate whole-body diffusion weighted magnetic resonance imaging (WB-DWI) for staging of women with cancer during pregnancy.Twenty patients diagnosed with cancer during pregnancy underwent WB-DWI additional to conventional imaging in this prospective single centre study. Reproducibility of WB-DWI between 2 readers was evaluated using Cohen’s κ statistics and accuracy was compared to conventional imaging for assessing primary tumour site, nodal metastases and visceral metastases. Histopathology after surgery or biopsy was the primary reference standard.Ten patients had breast cancer, 3 lymphoma, 2 cervical uterine cancer, 1 ovarian borderline tumour, 2 colon cancer, 1 lung cancer and 1 a conjunctival tumour. The WB-DWI readers showed very good agreement for lesion detection, κ = 0.94. With WB-DWI, reader 1 detected 38 of 41 malignant lesions, reader 2 thirty-nine lesions and conventional imaging 27. WB-DWI showed sensitivity of 95% (95% CI: 74-99) for both readers and specificity up to 99% (95% CI: 76-99) compared to 50% sensitivity (95% CI: 28-72) with 100% (95% CI: 97-100) specificity for conventional imaging. For staging distant metastases, WB-DWI sensitivities were 66.7% (95% CI: 13-98) and 100% (95% CI: 40-100) respectively for reader 1 and 2 with specificities of 94.1% (95% CI: 69-99) and 100% (95% CI: 40-100) compared to sensitivity of 33.3% (95% CI: 1.7-87) and specificity of 100% (95% CI: 77-100) for conventional imaging.WB-DWI is feasible for single-step non-invasive imaging based cancer staging during pregnancy showing additional value to conventional imaging procedures for detecting distant and nodal metastases.


Protochlamydia sp. strain HS-T3, an environmental chlamydia. This bacterium is an amoebal endosymbiont, found in Acanthamoeba isolated from a hot spring in Japan. Strain HS-T3 readily grew in mammalian cells at 37°C, a characteristic not previously reported for environmental chlamydiae.Here, we report the draft genome sequence of high-temperature-adapted  Chlamydiae is a bacterial phylum and class comprising obligate intracellular bacteria. Two distinct lineages exist, pathogenic and environmental chlamydiae, and this divergence likely occurred 0.7 to 1.4 billion years ago   , sexuallindness) , and comindness) . In conteduction . Most eneduction . Howevere, Japan .Protochlamydia HS-T3 was obtained using an Illumina HiSeq 2000 sequencer , with sequencing runs for paired-end sequences. The bacterial DNA libraries were prepared using a TruSeq DNA sample kit (Illumina). The genome was assembled into 34 contigs, ranging in size from 189 to 363,061 bp, using de novo sequence assembler software  (http://rast.nmpdr.org/) (http://www.genome.jp/kegg/) (The draft genome of MBL-EBI) . Gene prdr.org/) , and funp/kegg/) . The seqProtochlamydia sp. HS-T3 was 2,308,589 bp (2.3 Mb) with a G+C content of 38.74% and 950-fold genome coverage. The genome sequence contained 2,259 coding sequences, 40 tRNAs, and 2 ribosomal RNAs. Neighbor analysis using RAST revealed that the closest neighbor of Protochlamydia HS-T3 was Protochlamydia amoebophila UWE25 (NC_005861.1)  and type III general secretion gene clusters but lacked the genes encoding the type IV secretion system. In addition, 1,139 hypothetical proteins and 8 unknown proteins were identified.The draft genome sequence of 5 NC_00581.1 (6), 05861.1) . SimilarProtochlamydia HS-T3 has been deposited at DDBJ/EMBL/GenBank under the accession numbers BBPT01000001 through BBPT01000034 (34 entries). The version described in this paper is the first version.The draft genome sequence of


Previous studies have described improvements on lipid parameters when switching from other antiretroviral drugs to tenofovir (TDF) and impairments in lipid profile when discontinuing TDF. –3It is This was a randomized, crossover, double-blind, placebo-controlled clinical trial (NCT 01458977). Subjects with HIV-1 RNA <50 copies/mL during at least 6 months on stable DRV/r (800/100 mg QD) or LPV/r (400/100 mg BID) monotherapy, with confirmed fasting total cholesterol ≥200 or LDL-cholesterol ≥130 mg/dL and not taking lipid-lowering drugs were randomized to (A) adding TDF/FTCduring 12 weeks followed by 24 weeks without TDF/FTC, or (B) continuing without TDF/FTC for 12 weeks, adding TDF/FTC for 12 weeks and then withdrawing TDF/FTC for 12 additional weeks. Randomization was stratified by DRV/r or LPV/r use at study entry. All subjects received a specific dietary counselling. Primary endpoints were changes in median fasting total, LDL and HDL-cholesterol 12 weeks after TDF/FTC addition. Analyses were performed by ITT.46 subjects with a median age of 43 (40–48) years were enrolled in the study: 70% were male, 56% received DRV/r and 44% LPV/r. One subject withdrew the study voluntarily at week 4 and another one interrupted due to diarrhoea at week 24. Treatment with TDF/FTC decreased total, LDL and HDL-cholesterol from 235.9 to 204.9 (p<0.001), 154.7 to 127.6 (p<0.001) and 50.3 to 44.5 mg/dL (p<0.001), respectively. In comparison, total, LDL and HDL-cholesterol levels remained stable during placebo exposure. Week 12 total cholesterol (p<0.001), LDL-cholesterol (p<0.001) and HDL-cholesterol (p=0.011) levels were significantly lower in TDF/FTC versus placebo. Treatment with TDF/FTC reduced the fraction of subjects with abnormal fasting total-cholesterol (≥200 mg/dL) from 86.7% to 56.8% (p=0.001) and LDL-cholesterol (≥130 mg/dL) from 87.8% to 43.9% (p<0.001), which was not observed with placebo. There were no virological failures, and CD4 and triglyceride levels remained stable regardless of exposure.Coformulated TDF/FTC has an intrinsic lipid-lowering effect, likely attributable to TDF.


In heart failure (HF) alveolar-capillary membrane is abnormal. Surfactant-derived proteins (SPs) and plasma receptor for advanced-glycation-end-products (RAGE) have been proposed as lung damage markers.Eighty-nine chronic HF and 17 healthy subjects were evaluated by echocardiography, blood parameters, carbon monoxide lung diffusion (DLCO) and cardiopulmonary exercise test. We measured immature SP-B, mature SP-B, SP-A, SP-D and RAGE plasma levels.th–25th interquartile range) Vs. 11.1 (6.4), p<0.01; SP-A, 29.6 (20.1) Vs. 18.3 (13.5), p = 0.01; SP-D: 125 (90) Vs. 78 (58), p<0.01]. Immature SP-B, SP-A, SP-D and RAGE values were related to DLCO, peak oxygen consumption, ventilatory efficiency, and brain natriuretic peptide (BNP), whereas plasma mature SP-B was not. The DLCO Vs. immature SP-B correlation was the strongest one. At multivariate analysis, RAGE was associated to age and creatinine, SP-A to DLCO and BNP, SP-D to BNP, mature SP-B to DLCO and creatinine, and immature SP-B only but strongly to DLCO.Immature SP-B (arbitrary units), mature SP-A (ng/ml) and SP-D (ng/ml), but not mature SP-B (ng/ml) and RAGE (pg/ml) levels, were higher in HF than in controls [immature SP-B: 15.6 (13.1, 75Immature SP-B is the most reliable biological marker of alveolar-capillary membrane function in HF. Impairment of respiratory function is part of the chronic heart failure (HF) syndrome, being both lung mechanics and gas exchange altered. In all cases, RAGE and SPs have been linked to alveolar capillary membrane damage, but a comparative evaluation among RAGE and the different SPs available as markers of alveolar capillary membrane damage in HF has not been performed yet. We therefore analyzed the correlation between lung diffusion abnormalities, in terms of carbon monoxide total lung diffusion (DLCO), and RAGE and several SPs in a population of chronic stable HF patients and healthy controls, aiming to identify the ones that better correlates with gas diffusion.We studied HF patients in stable clinical conditions and healthy subjects. Patients belong to a group of individuals regularly followed up at our HF unit and were randomly recruited between February 2012 and November 2012, whereas healthy subjects were hospital staff employees or their relatives with gender and age similar to the HF patients.Study inclusion criteria for HF patients were New York Heart Association functional classes (NYHA) I to IV, echocardiographic evidence of reduced left ventricular systolic function , optimized and individually tailored drug treatment, stable clinical conditions for at least 2 months, capability/willingness to perform a maximal or nearly maximal cardiopulmonary exercise test (CPET). Patients were excluded if they had severe obstructive and/or restrictive lung disease, anemia (hemoglobin <11 g/dL), history and/or documentation of pulmonary embolism, primary valvular heart disease, pulmonary arterial hypertension, pericardial disease, exercise-induced angina, ST changes, severe arrhythmias and significant cerebrovascular, renal, hepatic and hematological disease.At enrolment, demographical and clinical data were collected. Before CPET, in both HF and healthy subjects echocardiographic evaluation, natriuretic peptide B (BNP) and blood samples measurements, standard pulmonary function tests, including DLCO, were performed.1) and lung vital capacity (VC) , and were standardized as percentages of predicted normal values. 1/VC lower than 60% was considered indicative of a severe obstructive ventilatory defect BNP test was performed on UniCel-DxI-800 Access immunoassay  with Triage-Biosite reagent . Forced expiratory volume in 1 second . 2), carbon dioxide production (VCO2) and ventilation (VE) were measured breath by breath and reported as average over 20 seconds. All tests were executed and evaluated by 2 expert readers blinded to plasma SPs and RAGE values. The anaerobic threshold (AT) was identified by a standard techniques. 2 was calculated as the slope of the linear relationship between VE and VCO2 from 1 minute after the beginning of loaded exercise to the end of the isocapnic buffering period.A CPET familiarization test was always performed. g for 10 minutes at 4°C, divided into aliquots and frozen at −80°C until assayed.Fresh blood (5 mL) was drawn into Vacutainer tubes containing citrate 0.129 mol/L as an anticoagulant. Plasma was immediately prepared by means of centrifugation at 1,500×versus the volume of the total proteins loaded and stained with MemCode. The values were also normalized versus the band volume of pooled plasma, loaded as control on each gel, and they are expressed as arbitrary units (AU). Inter-assay coefficient of variation was 12.1±2.9%.The analysis of the immature forms of SP-B, which is detectable in the 3 predominant forms. with molecular mass ranging from 17 to 42 kDa, was performed by Western blotting on plasma samples, as previously described. The quantitative analysis of the levels of the mature form (8 kDa) of SP-B was performed by an ELISA purchased from Uscn Life Science Inc. . Inter-assay and intra-assay coefficient of variation were 11.6±2.1% and 7.9±1.5%, respectively.Plasma levels of SP-A, SP-D and RAGE were determined using commercially available ELISA kits . Measurements were performed in duplicate and the results were averaged. The intra-assay and inter-assay coefficients of variation for SP-A were <5% and <10%, Limit of Detection (LOD) is 0.16 ng/ml, and cut off level is 1 ng/ml. For SP-D the intra-assay and inter-assay coefficients of variation were <3% and <4%, LOD is 0.01 ng/ml, and cut off level is 1.56 ng/ml. The intra-assay and inter-assay coefficients of variation for RAGE were <6% and <8%, respectively, minimal detectable dose is 4.12 pg/ml, and the cut off level is 78 pg/ml.th percentile–25th percentile). Categorical variables were compared with χ2 test. Unpaired t-test or non-parametric Mann-Whitney test were used when appropriate for between-group comparison. One-way analysis of variance [ANOVA] followed by Kruskal Wallis test were used to compare data for the non-normally distributed variables.Unless otherwise indicated all data are expressed as means ± SD. Data with skewed distribution are given as median and interquartile range (752 (≥15 mL/kg/min and <15 mL/kg/min), median VE vs VCO2 slope (≤30 and >30), median LVEF (≤36% and >36%) and median BNP (≤160 pg/ml and >160 pg/ml). Finally, given that it is well-reported a possible confounding role of smoke on the alveolar-capillary membrane 2 value and VE/VCO2 slope. To avoid distorted estimates of the effect, Pearson’s correlations were limited to the population with HF. A Bootstrap method, Besides between healthy subjects and HF patients, statistical analysis was also performed by subdividing HF patients according to NYHA class (I–II vs III–IV), median DLCO (≥80% vs <80%), median peak VO2) and 17 healthy subjects  . Active smokers were 30/89 and 7/17 in the HF and control group, respectively. HF etiology was cardiomyopathy due to ischemic coronary disease in 50 cases and not-ischemic in 39 cases. Among HF subjects, 44 were in NYHA class I–II and 45 in class III–IV. The LVEF in HF patients was 36±9%; the median BNP was 160 pg/mL . Treatment in patients with HF included angiotensin-converting enzyme inhibitors in 53 cases (60%), AT1 blockers in 21 cases (24%), β-blockers in 81 cases (91%), diuretics in 59 cases (66%), antialdosteronic drugs in 43 cases (48%), amiodarone in 37 cases (42%), digoxin in 4 cases (5%), antiplatelet in 59 cases (66%), and anticoagulant in 24 cases (27%).A total of 106 subjects were evaluated: 89 patients with HF  levels. Only immature SP-B and mature SP-A were higher in patients with most severe HF considering all the above reported HF severity criteria . VO2ATKG-Oxygen uptake at anaerobic threshold (mL/min/Kg). VEVCO2slope-Ventilatory efficiency. NYHA-New York Heart Association functional class. ISCHAEMIC_ETIOLOGY-. ICD-implantable cardioverter-defibrillator . CRT-cardiac resynchronization therapy . BBLOCK-Betablocker . ACEI-Ace inhibitor . AT1 BLOCK-Angiotensin converting enzyme inhibitors . ANTIALDOSTERONIC-angiotensin type 1 receptor blockers . DIURETIC-. AMIODARONE-. DIGOXIN-. ANTICOAGULANT-. ANTIPLATELET-.PAZIENT ID-Identification number. HF-Heart failure patient . SPBIMMATURE-Immature SPB (AU). SPB-Surfactant protein B (ng/mL). SPA-Surfactant protein A (ng/mL). SPD-Surfactant protein D (ng/mL). RAGE-Plasma receptor for advanced glycation end products (pg/mL). AGE-Age (years). GENDER-. IPERTENSION-. DMII-Diabetes type II . weight-(Kg). height-(cm). BMI-Body mass index (kg/m2). FUMO-smoke . LVEF-Left ventricular ejection fraction (%). Vtd-TeleDiastolic Volume (mL). Vts-TeleSystolic Volume (mL). IM-Mitral insufficiency . severityIM-Mitral insufficiency severity (1–4). GradoIT-Tricuspidalic insufficiency severity. TAPSE-Tricuspid annular plane systolic excursion (mm). PAPs-Systolic Pulmonary artery Pressure (mmHg). Hb-Hemoglobin (mg/dL). Crea-Creatinine (mg/dL). Na-Sodium (mmol/L). K-Potassium (mmol/L). Uric acid-(mg/dL). BNP-brain natriuretic petide (pg/mL). VC-Vital Capacity (L). VC_P-Vital Capacity (% of predicted). FVC-forced vital capacity (L). FVC_P-forced vital capacity (%). FEV1-forced expiratory volume in 1 second (L). FEV1_A-forced expiratory volume in 1 second (L). FEV1FVC-FEV1/FVC ratio. Dladj_ass-carbon monoxide lung diffusing capacity (mL/mm Hg/min). Dladj_perc-carbon monoxide lung diffusing capacity (% of predicted). SBPBASAL-Systolic blood pressure (mmHg). DBPBASAL-Diastolic blood pressure (mmHg). HRBASAL-Heart Rate (bpm). LOAD-Load at exercise (watt). QRPICk-Respiratory exchange ratio at peak. sbpPEAK-Systolic blood pressure at peak exercise (mmHg). DBPPEAK-Diastolic blood pressure at peak exercise (mmHg). VO(XLSX)Click here for additional data file.


E. coli (ESBLEc) increased.We conducted an annually bloodstream infection (BSI) survey into hospitals overlapping the Centre France region (2.6 million). Since 2005, the incidence of BSIs associated with ESBL-producing To improve the understanding of the pathway and the determination of the risk factors of ESBLEc-BSIs.For each BSI, were reported patient age, sex, recent hospitalization, living in nursing home, recent antibiotherapy, urinary catheterization, BSI source, death within 7days of diagnosis.BSI isolates were studied: antimicrobial susceptibility, determination of molecular mechanism associated with ESBL-production, genetic diversity of ESBLEc (MLST).E. coli BSI were identified, including 31 ESBLEc . Incidence of community acquired(CA)- and healthcare associated(HCA)-BSI were 0.47/100,000 and 0.040/1,000 PDs, respectively.During the survey , 443 For ESBLEc-CA-BSIs, male/female ratio was 1.4, median age 80, urinary BSI source in 50% of cases, recent antibiotherapy in 33 %. Most ESBLEc were resistant to fluoroquinolones (67%), SXT/TMP (67%). High genetic diversity (8 STs including 4 ST131).For ESBLEc-HCA-BSI, male/female ratio was 0.9, median age 75, urinary BSI source in 63% of cases (recent catheterization in 1/2), recent antibiotherapy in 58%. Most ESBLEC were resistant to fluoroquinolones (79%), SXT/TMP (63%). Low genetic diversity (9 STs including 7 ST131).Among BSI, ESBLEc-BSI were associated with healthcare (p=0.004), long-stay unit (p=0.018), recent antibiotherapy (p=0.002). ESBLEc were associated with resistance to fluoroquinolones, SXT/TMP and genta./tobramycine (p<0.001).Among ESBLEc-BSI, clinical determinants and BSI characteristics similar whatever the clonal group excepted for ST131 associated with long-stay unit (p=0.042).Among ST131-BSI, clinical determinants and BSI characteristics similar for ESBLEc and non ESBLEc excepted median age higher in ESBLEc (80/67).Recent antibiotherapy (and easy spread into long-stay units for ST131): likely the major risk factor for ESBLEc BSI.None declared.


KRAS wild-type and mutant. Prognostic relevance of KRAS genotype was evaluated in patients unfit for FIr-B/FOx, treated with conventional medical treatments. Consecutive MCRC patients not eligible for FIr-B/FOx regimen due to age (≥75 years) and/or comorbidities were treated with tailored conventional first-line treatments. KRAS codon 12/13 mutations were screened by direct sequencing. Activity and efficacy were evaluated and compared according to medical treatments, age (non-elderly and elderly ≥65 years), comorbidity stage , metastatic extension (liver-limited and other/multiple metastatic), and KRAS genotype, using log-rank. Selected first line treatments were medical in 37 patients (92.5%), and surgical in 3 patients (7.5%). Medical treatment regimens: triplet, 18 (45%); doublet, 15 (37.5%); mono-therapy, 4 (10%). At median follow-up of 8 months, objective response rate (ORR) was 37%, median progression-free survival (PFS) 7 months, liver metastasectomies 8% (liver-limited disease 37.5%), median overall survival (OS) 13 months. Triplet regimens failed to significantly affect clinical outcome, compared to doublet. According to KRAS genotype, ORR, PFS and OS were, respectively: wild-type 50%, 8 months, 13 months; mutant 25%, 6 months, 9 months. KRAS genotype wild-type compared to mutant significantly affected PFS, while not OS. KRAS c.35 G>A mutation (G12D) significantly affected worse PFS and OS compared to wild-type and/or other mutations. KRAS genotype, specifically the c.35 G>A KRAS mutation, may indicate poor prognosis in MCRC patients unfit for intensive medical treatments.First-line triplet chemotherapy plus bevacizumab (FIr-B/FOx) can improve efficacy of metastatic colorectal cancer (MCRC),  KRAS genotype  faces with different options and lines of treatment according to patients’ fitness , extension of metastatic disease [liver-limited (L-L) or other/multiple metastatic (O/MM)], genotype –4. Firstgenotype ,5,6. Morgenotype –9.In clinical practice, a decision-making process including functional, nutritional, and co-morbidity status is required to tailor first line medical treatment . ElderlyKRAS mutations occur in 35–45% of colorectal cancer (CRC), mostly codon 12 (80%), prevalently c.35 G>A (G12D) transversion (32.5%)  , impairi (32.5%) . In the ild-type –39; BEV activity ,37.KRAS genotype, depending on differential tumor biological aggressiveness  mutant genotype may significantly affect worse OS of MCRC patients treated with FIr-B/FOx, compared to wild-type or different other mutations . Thus, it was not a clinical trial and approval by ethics committee and institutional review board was not necessary, because patients were treated with conventional treatments without any additional medical intervention out of the best common clinical practice. Patients had histological confirmed diagnosis of MCRC, age ≥18 years, PS ≤2. Criteria to define patients unfit, or not eligible for intensive regimens were: age ≥75 years; uncontrolled severe diseases; cardiovascular disease ; thromboembolic disease, coagulopathy, preexisting bleeding diatheses; proteinuria >1 g/24 h. Patients were classified according to Cumulative Illness Rating Scale (CIRS)  for administration e (CIRS) . Treatme2/die, over 12 h (from 10:00 pm to 10:00 am), days 1–2, 8–9, 15–16, 22–23; CPT-11 , 120–160 mg/m2, days 1 and 15; l-OXP , 70–80 mg/m2, days 8 and 22; cycles every 4 weeks. Other triplet, doublet and mono-regimens were administered according to previously reported schedules , associated to weekly alternating CPT-11 or L-OXP : TFI/5-Fchedules ,42. TargKRAS codon 12 and 13 mutations by direct sequencing. KRAS exon 2 sequence was performed from PCR-amplified tumor DNA using the Big Dye V3.1 Terminator kit, electrophoresis in ABI PRISM 3130xl Genetic Analyzer, and analysis using the GeneMapper Analysis software version 4.0 .Genetic analyses were performed on paraffin-embedded tissue blocks from primary tumor and/or metastatic sites, as previously reported . GenotypKRAS genotype on clinical outcomes were evaluated. Patients were classified according to: metastatic extension, L-L and O/MM , 23 (59%) were wild-type and 16 (41%) mutant. Clinical features of the 37 patients who underwent first line medical treatments were , specifically c.35 G>A (G12D), 7 (19.4%), c.35 G>T (G12V), 6 (16.6%); codon 13, 2 (5.5%), c.37 G>T (G13V), 1 (2.7%) and c.38 G>A (G13D), 1 (2.7%).Forty patients unfit for intensive regimens, among 72 consecutive MCRC (56%), were treated with : medicalnts were : male/fents were : non-eldMedical treatments were tailored according to age and CIRS stage. Triplet regimens were administered in 18 patients (49%): non-elderly 6, young-elderly 4, old-elderly 8; CIRS primary 2, intermediate 10, secondary 6. Doublet regimens were administered in 15 patients (40%): non-elderly 3, young-elderly 3, old-elderly 9; CIRS primary 1, intermediate 4, secondary 10. Mono-regimens were administered in 4 patients (11%): young-elderly 1, old-elderly 3; CIRS primary 1, intermediate 1, secondary 2.KRAS wild-type patient (33%), with primary rectal tumor and a single L-L metastasis. Twelve patients (32%) received, at least, a second line treatment.Among the 37 patients who underwent medical treatments, 10 were not evaluable for activity: 7 (19%) did not receive at least 2 cycles of treatment; 3 were on-treatment. The intent-to-treat analysis of 27 patients showed ORR 37%  . We obseAmong 7 evaluable L-L patients, ORR was 71%; 3 performed liver metastasectomies (43%) and 1 cCR (14%); median PFS 11 months (3–13+ months); median OS 12 months (3–13+ months). Among 20 evaluable O/MM patients, ORR was 25%; median PFS 6 months (1–12 months); median OS 13 months (1+−23+ months). Clinical outcome (PFS and OS) in L-L compared to O/MM patients was not significantly different .Among 16 evaluable patients treated with triplet regimens , ORR wasMoreover, PFS and OS were not significantly different in non-elderly and young-elderly compared to old-elderly patients , and in primary and intermediate CIRS stage compared to secondary stage patients .KRAS wild-type patients evaluable for activity, ORR was 50%  . We observed 3 partial responses (25%); 5 stable diseases (42%); 4 progressive diseases (33%). Disease control rate was 67% . No liver metastasectomies were performed. Median PFS was 6 months (1–11 months), 12 events occurred (80%). Median OS was 8 months (3–18 months), 10 events occurred. KRAS wild-type compared with mutant patients showed significantly different PFS (p=0.043), but not OS   (Among 14 CI ± 27) . We obset not OS . KRAS c.ctively) , and to ctively) . No diffpatients . PFS andctively) .Patients unfit for first line FIr-B/FOx intensive regimen, due to age (≥75 years) and/or comorbidities, were prevalent (56%), mostly elderly (76%), particularly old-elderly patients (54%), prevalently PS 1–2 (59%), CIRS stage intermediate/secondary (89%), O/MM disease (79%). Most unfit MCRC patients were treated with triplet or doublet regimens , and some (19%) did not reach the first evaluation of activity at 2–3 months.Retrospective evaluations of doublets consisting of CPT-11 or OXP, associated to 5-FU or capecitabine in elderly patients eligible for clinical trials gained ORR 18–59.4%, PFS 4.9–10.0 months and OS 8.5–20.7 months ,31,46. BYoung-elderly patients eligible for FIr-B/FOx intensive regimen, prevalently characterised by PS 0 (89%) and intermediate CIRS stage (93%), reported ORR 79%, PFS 11 months, OS 21 months, equivalent to overall patients . A complKRAS wild-type and mutant (KRAS geno-type was reported as significantly affecting PFS and OS in patients treated with XelOx/BEV (KRAS c.35 G>A (G12D) mutant genotype may significantly affect worse OS, compared to wild-type or other mutations , PFS and OS were not significantly different in d mutant ,6,8,37 ad mutant . RecentlelOx/BEV . We receutations . PresentA mutant ,40, and KRAS genotype may indicate different PFS, and c.35 G>A KRAS mutant a significantly worse PFS and OS, compared to wild-type and other mutations. Present findings warrant prospective trials comparing clinical outcome in unfit patients, according to KRAS genotype.In conclusion, in MCRC patients unfit for first line intensive FIr-B/FOx regimen, tailored doublet and triplet medical treatments showed similar activity and efficacy, also according to age and comorbidities.


Klebsiella quasipneumoniae subsp. similipneumoniae, KP_Z4175. This strain, isolated as part of a hospital infection-control screening program, is resistant to multiple β-lactam antibiotics, aminoglycosides, and trimethoprim-sulfamethoxazole.We report here the draft genome sequence of a multidrug-resistant clinical isolate of  Klebsiella quasipneumoniae, formerly Klebsiella pneumoniae phylogroup KpII, was recently taxonomically reclassified as a new sister species of K. pneumoniae with two subspecies, K. quasipneumoniae subsp. quasipneumoniae and K. quasipneumoniae subsp. similipneumoniae  using the GGDC 2.1 software  generating 2 × 301-bp paired-end reads. A total of 15,256,306 reads were produced comprising 1,540,886,906 bases after adapter sequence trimming. K. quasipneumoniae subsp. similipneumoniae strain KP_Z4175, antibiotic resistance genes were identified using ResFinder version 2.1 -IIc), two fluoroquinolone resistance genes (aac(6′)Ib-cr and QnrB4), one macrolide-lincosamide-streptogramin B resistance gene (ere(A)), two phenicol resistance genes (cmlA1 and floR), one rifampin resistance gene (ARR-2), two sulfonamide resistance genes (sul1 and sul2), one tetracycline resistance gene (tet(D)), and one trimethoprim resistance gene (dfrA14). All identified resistance genes had nucleotide identities of 98.35 to 100% over 85 to 100% of the reference gene lengths. Broth microdilution testing using CLSI breakpoints for Enterobacteriaceae indicated that KP_Z4175 is resistant to gentamicin (MIC >64), tobramycin (=32), cefazolin (>64), ceftriaxone (>64), aztreonam (>64), and trimethoprim-sulfamethoxazole (>64). The isolate had intermediate resistance to ampicillin-sulbactam (=16) and pipericillin/tazobactam (=32), and was sensitive to ertapenem (≤0.03), imipenem (=1), meropenem (≤0.03), amikacin (=0.5), cefepime (=8), and ciprofloxacin (=1).To examine the antibiotic resistance profile of sion 2.1 . In addieumoniae , β-lactaLVCD00000000. The version described in this paper is version LVCD01000000.This whole-genome shotgun project has been deposited at DDBJ/ENA/GenBank under the accession number


Dysregulation of the nucleo-cytoplasmic transport of proteins plays an important role in carcinogenesis. The nuclear export of proteins depends on the activity of transport proteins, exportins. Exportins belong to the karyopherin β superfamily. Exportin-1 (XPO1), also known as chromosomal region maintenance 1 (CRM1), mediates transport of around 220 proteins. In this review, we summarized the development of a new class of antitumor drugs, collectively known as selective inhibitors of nuclear export (SINE). KPT-330 (selinexor) as an oral agent is showing activities in early clinical trials in both solid tumors and hematological malignancies.The online version of this article (doi:10.1186/s13045-014-0078-0) contains supplementary material, which is available to authorized users. The nucleo-cytoplasmic transport of proteins plays an important role in maintaining normal cellular functions. The nuclear export of proteins depends on the activity of transport proteins, exportins. Exportin-1 (XPO1), also known as chromosomal region maintenance 1 (CRM1), mediates transport of around 220 proteins . XPO1 isXPO1 binds to the cargo proteins through a leucine rich nuclear export signal (NES) and transports the proteins through a membrane pore complex via a Ran-GTP gradient -15]  . The cliin vitro with most potency. However, its use in vivo is limited by poor pharmacokinetics . KPkinetics -[25]. Th50) of 150 nmol/L and inhibited pancreatic cancer cell lines while sparing normal human pancreatic ductal epithelial cells. The in vivo effects were noted using KPT-330 (selinexor) in subcutaneous and orthoptic pancreatic cancer models in mice. Oral administration of KPT-330 led to significant tumor growth inhibition when compared with control or gemcitabine treatment . EG. EG37]. l status . The antl status . In celll status . KPT-330l status . Moreovenografts .in vitro in RCC cell lines . Tr. Tr52]. hibitors . They cohibitors ,54]. Ho. Hoet alsistance ,63],. SI. SIin vi KPT-185 . FLT3 ge KPT-185 . SINE do KPT-185 ,74]. Ko. Koin viradation . Nutlin-radation . The add,. SI. SI79]. able IC50. In vivoet al. demonstrated successful use of KPT-185 in vitro and KPT-330 in vivo in Philadelphia chromosome positive ALL (Ph + ALL) and chronic myeloid leukemia blast crisis (CML-BC). Combination with imatinib led to synergistic effects . Wa. Wa81].  effects . SINE tr effects .et al. recently. They showed that KPT-330 treatment given to heavily pretreated, refractory/relapsed AML patients had no DLT. Out of the 32 evaluable patients, 4 (12%) showed complete response (CR) with hematological recovery, 1 (3%) showed marrow CR (mCR), mCR without hematological recovery was seen in 1 (3%) patient. Partial response (PR) was seen in 2 patients (6%). Eleven patients (34%) showed progression while 12 (37%) experienced stable disease after 30 days . Pr. Pr91]. L) cells . KPT-185L) cells ,93]. KP. KPet alL) cells . In a 37L) cells .in vitro and in vivo. Su. Suin vid in vivo. C-myc ad in vivo. A studyd in vivo. BRD4, ad in vivo,99]. BR. BRin vid in vivo. JQ1, a d in vivo. This reet al. conducted an in vitro study of KPT-185 in MCL and showed that SINE increases MCL cell apoptosis primarily by increasing nuclear p53 levels -[1-[1100]-. KPT dru3 levels . They vepression . London  effects . Gutierrpatients . These dMore and more targeted small molecule inhibitors are entering clinical application quickly ,84],,-113]. S. S113]. DL and KP designed the study. All authors have contributed to data preparation, drafting and revising the manuscripts. All authors have read and approved the final manuscript.


Blood biomarkers are increasingly used to diagnose, guide therapy in, and risk-stratify community-acquired pneumonia (CAP) patients in emergency departments (EDs). How pre-analytic factors affect these markers' initial levels in this population is unknown.In this secondary analysis of consecutive ED patients with CAP from a large multicentre antibiotic stewardship trial , we usedP = 0.02) in patients with liver insufficiency; ProADM, + 13.2%  in patients with age above median; and CRP, -12.8%  with steroid pretreatment.Of 925 CAP patients , 25.5% had antibiotic pretreatment, 2.4%, corticosteroid pretreatment, 22.3% chronic renal failure, and 2.4% chronic liver insufficiency. Differences associated with pre-analytic factors averaged 6.1 ± 4.6%; the three largest statistically significant changes (95% Cl) were: PCT, +14.2% (+2.1 to +26.4%, The influence of pre-analytic factors on the examined biomarkers is marginal and not clinically relevant. Our observations reinforce the concept ofusing biomarkers in algorithms with widely- separated cutoff values and overruling criteria considering the entire clinical picture, without adjustment for pre-analytic factors.


The importance of maternal nutrition to offspring health and risk of disease is well established. Emerging evidence suggests paternal diet may affect offspring health as well.In the current study we sought to determine whether modulating pre-conception paternal B vitamin intake alters intestinal tumor formation in offspring. Additionally, we sought to identify potential mechanisms for the observed weight differential among offspring by profiling hepatic gene expression and lipid content.1638N mice  were fed diets containing replete , mildly deficient (DEF), or supplemental (SUPP) quantities of vitamins B2, B6, B12, and folate for 8 weeks before mating with control-fed wild type females. Wild type offspring were euthanized at weaning and hepatic gene expression profiled. Apc1638N offspring were fed a replete diet and euthanized at 28 weeks of age to assess tumor burden.Male Apc1638N offspring of different paternal diet groups. Although in female Apc1638N offspring there were no differences in tumor incidence or multiplicity, a stepwise increase in tumor volume with increasing paternal B vitamin intake was observed. Interestingly, female offspring of SUPP and DEF fathers had a significantly lower body weight than those of CTRL fed fathers. Moreover, hepatic trigylcerides and cholesterol were elevated 3-fold in adult female offspring of SUPP fathers. Weanling offspring of the same fathers displayed altered expression of several key lipid-metabolism genes. Hundreds of differentially methylated regions were identified in the paternal sperm in response to DEF and SUPP diets. Aside from a few genes including Igf2, there was a striking lack of overlap between these genes differentially methylated in sperm and differentially expressed in offspring.No differences in intestinal tumor incidence or burden were found between male ApcIn this animal model, modulation of paternal B vitamin intake prior to mating alters offspring weight gain, lipid metabolism and tumor growth in a sex-specific fashion. These results highlight the need to better define how paternal nutrition affects the health of offspring. B, C, D) Photomicrographs of Oil Red-O stained liver sections from DEF (22.7%), CTRL (22.3%) and SUPP (37.6%) offspring respectively. Images are from mice with the highest staining area in each group. Images taken at 100x and scale bar is 200 μm. Paternal diets: DEF, B vitamin deficient; CTRL, B vitamin replete; SUPP, B vitamin supplemented.(DOCX)Click here for additional data file.S3 FigA) CpGs 1–14 are located at -359, -357, -347, -337, -334, -320, -314, -304, -296, -288, -280, -274, -267 and -264 bp from the start of Acaca1 exon 1 respectively. B) CpGs 1–9 are located at +36, +59, +50, +27, +14, +10, -3, -14, -40 and -44 bp from the start of Elovl6 exon 1 respectively. C) CpGs 1–19 are located -122, -113, -96, - 89, -77, -73, -64, -61, -52, -50, -48, -45, -41, -39, -33, -31, -28, -26 and -23 bp from start of Gpam exon 1 respectively.D) CpGs 1–8 are located -3775, -3766, -3754, -3706, -3702, -3699, -3685, -3678 and -3665 bp from start of H19 exon 1 respectively. Unless noted, for each CpG site, 2-Way ANOVA p (group) >0.05 and p (sex) >0.05. ‘a’ denotes p <0.05 for group effect and ‘b’ denotes p v0.05 for sex effect. Unless noted, repeated measures ANOVA for group  effect yielded a p>0.05. Paternal diets: DEF, B vitamin deficient; CTRL, B vitamin replete; SUPP, B vitamin supplemented; M male; F female. n = 4 males and 8 females per group.DNA methylation was measured in target genes using bisulfite pyrosequencing. (DOCX)Click here for additional data file.S1 TableB denotes biotin modification of primer at 5’ end of FWD or 3’ end of RVS primer.(DOCX)Click here for additional data file.S2 TableValues are mean ± SEM. n = 10, 13 and 9 respectively.(DOCX)Click here for additional data file.S3 TableDMR ID is an arbitrary identifier for each differentially methylated region; CTRL, control diet; DEF, B vitamin deficient diet; m1, mean log2(636/532) for CTRL sperm; m2, mean log2(636/532) for DEF sperm; chr:position identifies the location of the DMR within the chromosome using mouse genome build mm9 coordinates; diff, mean log2(635/532) difference across the DMR; qval, q-value—an adjusted p-value indicating the false discovery rate (FDR) calculated by the Wilcoxon rank sum-test. For genes with a qval < 0.1, less than 10% of genes are expected to be false positives; Gene symbol identifies the gene annotated to a DMR and its proximity in bases to the left (-) or right (+) of the DMR up to a distance of 1000 kb where all genome coordinates and gene symbols are derived from mouse genome build mm9. n = 8/gp.(XLSX)Click here for additional data file.S4 TableDMR ID is an arbitrary identifier for each differentially methylated region; CTRL, control diet; SUPP, B vitamin supplemented diet; m1, mean log2(636/532) for CTRL sperm; m2, mean log2(636/532) for SUPP sperm; chr:position identifies the location of the DMR within the chromosome using mouse genome build mm9 coordinates; diff, mean log2(635/532) difference across the DMR; qval, q-value—an adjusted p-value indicating the false discovery rate (FDR) calculated by the Wilcoxon rank sum-test. For genes with a qval < 0.1, less than 10% of genes are expected to be false positives; Gene symbol identifies the gene annotated to a DMR and its proximity in bases to the left (-) or right (+) of the DMR up to a distance of 1000 kb where all genome coordinates and gene symbols are derived from mouse genome build mm9. n = 8/gp.(XLSX)Click here for additional data file.S5 Table* Values are mean ± SEM. Note that number weaned offspring is less than sired offspring due to maternal cannibalism that is common amongst first-time mothers. DEF, B vitamin deficient; CTRL, B vitamin replete; SUPP, B vitamin supplemented.(DOCX)Click here for additional data file.S6 TableValues are mean ± SEM. Paternal diets, DEF, B vitamin deficient; CTRL, B vitamin replete; SUPP, B vitamin supplemented. Sample size is in parentheses.(DOCX)Click here for additional data file.S7 TableSignificant differences according to Cuffdiff analysis combining male and female offspring (q<0.05). Expression values are Fragments Per Kilobase Of Exon Per Million Fragments Mapped (FPKM). CTRL, control; DEF, deficient . n = 10/gp.(XLSX)Click here for additional data file.S8 TableSignificant differences according to Cuffdiff analysis combining male and female offspring (q<0.05). Expression values are Fragments Per Kilobase Of Exon Per Million Fragments Mapped (FPKM). CTRL, control; SUPP, supplemented . n = 10/gp.(XLSX)Click here for additional data file.S9 TableCategories identified by Ingenuity Pathway Analysis®. DEF, B vitamin deficient; CTRL, B vitamin replete; SUPP, B vitamin supplemented . P values represent the range of p values of the sub-categories of each category.(DOCX)Click here for additional data file.S10 Tablen = 5/gp.(XLSX)Click here for additional data file.S11 Table(DOCX)Click here for additional data file.