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Introduction Prevalence and 1-year impact of right ventricular systolic dysfunction (RVSD) early after lung transplantation (LTx), in a population without pulmonary hypertension (PHT), is not well documented. In this retrospective study, we hypothesized that, as after heart Tx, occurrence of early RVSD after LTx in those patients (pts) would be associated with impaired 1-year survival.¹ Methods After ICU admission of LTx pts, we routinely perform a comprehensive transoesophageal echocardiographic (TOE) to check pulmonary veins’ patency. After exclusion of pts transplanted for PHT and of pts under ECMO, we retrospectively reviewed RV systolic data acquired during this TOE. At the time of TOE, all pts were mechanically ventilated. RVSD was defined based on visual assessment (RV dilation and/or RV hypokinesia) and RV fractional area change (FAC) Results From January 2004 to March 2018, 166 LTx pts underwent comprehensive TOE in ICU. Indications for LTx were COPD in 117 pts (70.5%), fibrosis in 25 (15.1%), cystic fibrosis in 7 (4.2%), bronchiectasis in 7 pts (4.2%) and other indications in 10 (6.0%). RV visual assessment was reported as abnormal in 56/166 pts (33.7%). RV FAC was reported in 112 pts and was Discussion Based on this retrospective TOE and single-center experience, prevalence of early RVSD after LTx, in a population without PHT, ranges from 34 to 39%, depending on the used criteria (visual assessment or RV FAC). However, occurrence of early RVSD after LTx was not associated with impaired 1-year survival.

An imbalance between acidogenic and methanogenic organisms during anaerobic digestion can result in increased accumulation of volatile fatty acids, decreased reactor pH, and inhibition of methane-producing Archaea. Most commonly the result of organic input overload or poor inoculum selection, these microbiological and biochemical changes severely hamper reactor performance, and there are a few tools available to facilitate reactor recovery. A small, stable consortium capable of catabolizing acetate and producing methane was propagated in vitro and evaluated as a potential bioaugmentation tool for stimulating methanogenesis in acidified reactors. Replicate laboratory-scale batch digesters were seeded with a combination of bioethanol stillage waste and a dairy manure inoculum previously observed to result in high volatile fatty acid accumulation and reactor failure. Experimental reactors were then amended with the acetoclastic consortium, and control reactors were amended with sterile culture media. Within 7 days, bioaugmented reactors had significantly reduced acetate accumulation and the proportion of methane in the biogas increased from 0.2 ± 0 to 74.4 ± 9.9 % while control reactors showed no significant reduction in acetate accumulation or increase in methane production. Organisms from the consortium were enumerated using specific quantitative PCR assays to evaluate their growth in the experimental reactors. While the abundance of hydrogenotrophic microorganisms remained stable during the recovery period, an acetoclastic methanogen phylogenetically similar to Methanosarcina sp. increased more than 100-fold and is hypothesized to be the primary contributor to reactor recovery. Genomic sequencing of this organism revealed genes related to the production of methane from acetate, hydrogen, and methanol. Electronic supplementary material The online version of this article (doi:10.1007/s00253-015-7058-3) contains supplementary material, which is available to authorized users.

Chicory (Cichorium intybus) is a perennial plant (Asteraceae) that grows wild in pasture fields in Saudi Arabia. Chicory plants displaying symptoms typically induced by phytoplasmas, such as bushy phenotype and stunt, were observed in the Mulayda region, Qassim governorate, Saudi Arabia. In this study we examined samples taken from three symptomatic chicory plants and confirmed the presence of phytoplasma DNA. Analysis of the 16S rRNA-encoding sequences showed that the plants were infected with a phytoplasma from the pigeon pea witches’-broom group (16SrIX). Sequencing of the 16S rRNA-encoding gene and the partial cpn60 sequence, computer-simulated RFLP analysis, and phylogenetic analysis of both markers revealed that the phytoplasma identified was representative of a new 16SrIX-J and cpn60 UT IX-IJ subgroup. The present study identified chicory plants as a novel host for phytoplasma strains within the pigeon pea witches’-broom phytoplasma group, and expanded the known diversity of this group.

International audience; Facioscapulohumeral dystrophy (FSHD) is characterized by the contraction of the D4Z4 array located in the sub-telomeric region of the chromosome 4, leading to the aberrant expression of the DUX4 transcription factor and the mis-regulation of hundreds of genes. Several therapeutic strategies have been proposed among which the possibility to target the polyadenylation signal to silence the causative gene of the disease. Indeed, defects in mRNA polyadenylation leads to an alteration of the transcription termination, a disruption of mRNA transport from the nucleus to the cytoplasm decreasing the mRNA stability and translation efficiency. This review discusses the polyadenylation mechanisms, why alternative polyadenylation impacts gene expression, and how targeting polyadenylation signal may be a potential therapeutic approach for FSHD.

Muscular dystrophies are characterized by weakness and wasting of skeletal muscle tissues. Several drugs targeting the myostatin pathway have been used in clinical trials to increase muscle mass and function but most showed limited efficacy. Here we show that the expression of components of the myostatin signaling pathway is downregulated in muscle wasting or atrophying diseases, with a decrease of myostatin and activin receptor, and an increase of the myostatin antagonist, follistatin. We also provide in vivo evidence in the congenital myotubular myopathy mouse model (knock-out for the myotubularin coding gene Mtm1) that a down-regulated myostatin pathway can be reactivated by correcting the underlying gene defect. Our data may explain the poor clinical efficacy of anti-myostatin approaches in several of the clinical studies and the apparent contradictory results in mice regarding the efficacy of anti-myostatin approaches and may inform patient selection and stratification for future trials., Drugs targeting myostatin reverse muscle wasting in animal models, but have limited efficacy in patients. The authors show that the myostatin pathway is downregulated in patients, possibly explaining the poor outcome of anti-myostatin approaches, and that it can be reactivated by correcting disease-causing mutations in mice.

The vaginal microbiome plays an important role in maternal and neonatal health. Imbalances in this microbiota (dysbiosis) during pregnancy are associated with negative reproductive outcomes, such as pregnancy loss and preterm birth, but the underlying mechanisms remain poorly understood. Consequently a comprehensive understanding of the baseline microbiome in healthy pregnancy is needed. We characterized the vaginal microbiomes of healthy pregnant women at 11–16 weeks of gestational age (n = 182) and compared them to those of non-pregnant women (n = 310). Profiles were created by pyrosequencing of the cpn60 universal target region. Microbiome profiles of pregnant women clustered into six Community State Types: I, II, III, IVC, IVD and V. Overall microbiome profiles could not be distinguished based on pregnancy status. However, the vaginal microbiomes of women with healthy ongoing pregnancies had lower richness and diversity, lower prevalence of Mycoplasma and Ureaplasma and higher bacterial load when compared to non-pregnant women. Lactobacillus abundance was also greater in the microbiomes of pregnant women with Lactobacillus-dominated CSTs in comparison with non-pregnant women. This study provides further information regarding characteristics of the vaginal microbiome of low-risk pregnant women, providing a baseline for forthcoming studies investigating the diagnostic potential of the microbiome for prediction of adverse pregnancy outcomes.

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The use of oligonucleotide-coupled fluorescent microspheres is a rapid, sequencing-independent, and reliable way to diagnose bacterial diseases. Previously described applications of oligonucleotide-coupled fluorescent microspheres for the detection and identification of bacteria in human clinical samples have been successfully adapted to detect and differentiate "Ca. Phytoplasma" species using as a target the chaperonin 60-encoding gene. In this chapter, we describe in detail the design and validation of oligonucleotide capture probes, and their application in the assay aiming to differentiate phytoplasma strains infecting Brassica napus and Camelina sativa plants grown in the same geographic location at the same time.

The pretreatment of plant biomass negatively impacts the economics of many bioenergy and bioproduct processes due to the thermochemical requirements for deconstruction of lignocelluluose. An effective strategy to reduce these severity requirements is to pretreat the biomass with white-rot fungi, such as Trametes versicolor, which have the innate ability to deconstruct lignocellulose with a suite of specialized enzymes. In the present study, the effects of 12 weeks of pretreatment with a wild-type strain (52J) and a cellobiose dehydrogenase-deficient strain (m4D) of T. versicolor on hardwood and Miscanthus were explored. Both strains of T. versicolor led to significant decreases of insoluble lignin and significant increases of soluble lignin after acid hydrolysis, which suggests improved lignin extractability. The glucose yields after saccharification using an enzyme cocktail containing chitinase were similar or significantly higher with 52J-treated biomass compared to untreated hardwood and Miscanthus, respectively. The fungal treated biomass, regardless of the strain used, also showed significant increases in energy content and compressive strength of pellets. Overall, the use of T. versicolor as a pretreatment agent for hardwood and Miscanthus could be an environmentally friendly strategy for conversion technologies that require delignification and saccharification, and/or processes that require densification and transport.

Geographically diverse samples from strawberry exhibiting symptoms of Strawberry Green Petal (SbGP), periwinkle plants with virescence, and blackberry, blueberry, and raspberry plants displaying yellowing and inedible fruits, were assayed for the presence of phytoplasma DNA. PCR targeting the 16S rRNA-encoding gene and chaperonin-60 (cpn60) showed that the plants were infected with phytoplasma subgroup16SrXIII-(A/I)I (SbGP/MPV). To examine the geographic distribution of this pathogen in Mexico, we designed an array of cpn60-targeted molecular diagnostic assays for SbGP/MPV phytoplasma. A fluorescent microsphere hybridization assay was designed that was capable of detecting SbGP/MPV phytoplasma in infected plant tissues, successfully differentiating it from other known phytoplasma cpn60 UT sequences, while identifying a double infection with SbGP/MPV and aster yellows (16SrI) phytoplasma. Two quantitative assays, quantitative real-time PCR (qRT-PCR) and droplet digital PCR (ddPCR), gave similar results in infected samples. Finally, a loop-mediated isothermal amplification (LAMP) assay provided rapid detection of SbGP/MPV phytoplasma DNA. Application of these assays revealed that SbGP/MPV phytoplasma is widely distributed in Central Mexico, with positive samples identified from eleven localities within three states separated by hundreds of kilometres. These results also provide tools for determining the presence and geographic distribution of this pathogen in plant and insect samples in other localities.

Biochemical methane potential (BMP) assays were conducted on byproducts from dry-grind wheat-based ethanol plants amended with feedlot manure at two input ratios. Whole stillage (WST), thin stillage (TST) and wet cake (WCK) were tested alone and with 1:1 and 2:1 ratios (VS basis) of byproduct:feedlot manure in bench-scale batch reactors. The addition of manure increased both the rate and consistency of methane production in triplicate reactors. In addition, digesters co-digesting thin stillage and cattle manure at 1:1 and 2:1 stillage:manure produced 125% and 119% expected methane based on the biomethane potential of each substrate digested individually. Bacterial community analysis using universal target amplification and pyrosequencing indicated there was a numerically dominant core of 42 bacteria that was universally present in the reactors regardless of input material. A smaller-scale analysis of the archaeal community showed that both hydrogenotrophic and acetoclastic methanogens were present in significant quantities.

While transfer of a protein encoded by a single nucleus to nearby nuclei in multinucleated cells has been known for almost 25 years, the biological consequences for gain-of-function diseases have not been considered. Here, we have investigated nuclear protein spreading and its potential consequences in two of the three most prevalent neuromuscular diseases. By performing co-cultures between diseased or control human myoblasts and murine C2C12 myoblasts, we demonstrate that in facioscapulohumeral dystrophy, although the transcription of the toxic protein DUX4 occurs in only a limited number of nuclei, the resulting protein diffuses into nearby nuclei within the myotubes, thus spreading aberrant gene expression. In myotonic dystrophy type 1, we observed that in human-mouse heterokaryons, the expression of a mutated DMPK from human nuclei titrates splicing factors produced by neighboring nuclei, inducing the mis-splicing of several pre-mRNAs in murine nuclei. In both cases, the spreading of the pathological phenotypes from one nucleus to another is observed, highlighting an additional mechanism that contributes to the dissemination and worsening of the muscle pathogenesis. These results indicate that nuclear protein spreading may be an important component of pathophysiology of gain of function muscular diseases which should be taken into consideration in the design of new therapeutic approaches.

Introduction/Objectives In Duchenne muscular dystrophy (DMD), myofibres die with a necrotic morphology, however the cell death mechanisms are largely unknown. Inflammation plays a significant part in muscle degeneration. Necroptosis is a newly identified programmed form of necrosis, which is driven by receptor interacting protein kinase 3 (RIPK3). It plays a major role in the inflammation-induced injuries in several tissues. We are currently investigating the involvement of necroptosis in DMD pathogenesis. Materials/Patients and methods In vitro, muscle cells were challenged with inflammatory stimuli, with or without various protein inhibitors. Cell toxicity was monitored by the quantification of ATP content (reflecting cell survival) and protease realease (reflecting membrane permeability) In vivo, dystrophic skeletal muscles from mdx mice (a mouse model for DMD) were analyzed. Mdx mice were crossed with RIPK3 KO mice. Results In vitro, we found that TNFα can trigger necroptosis in C2C12 cell line, suggesting that muscle cells can undergo necroptosis upon inflammatory challenge. In vivo, we found evidence of necroptosis in human and mouse dystrophin-deficient muscles. By depleting RIPK3 in mdx mice, we significantly decreased myonecrosis. Conclusions Our data suggest that the necroptosis machinery is involved in the cell death affecting myofibres in DMD pathogenesis.

Introduction Right ventricular (RV) diastolic dysfunction is known to be associated with increased mortality in heart failure or cardiac surgical patients (pts). However, after lung transplantation (LTx), the impact of early RV diastolic dysfunction on 1-year survival, in a population without pulmonary hypertension, is not well documented. Methods We routinely perform a comprehensive transoesophageal echocardiography (TOE) after ICU admission of LTx patients (pts) to check the pulmonary veins’ patency. After exclusion of pts transplanted for pulmonary hypertension and of pts under ECMO, we retrospectively reviewed the diastolic RV parameters acquired during this TOE. At that time, all pts were mechanically ventilated. RV diastolic function was explored by tricuspid peak early and late velocities (E and A waves) (pulsed-wave Doppler (PW) through the tricuspid valve), early diastolic peak velocity (Ea) of the lateral tricuspid annulus (PW-TDI), hepatic venous flow (HVF) pattern (PW Doppler), and characterized as normal or abnormal according to the algorithm validated by Shi et al. One-year survival status was recorded. Continuous variables are presented as median [interquartile range] and analysed with Mann-Whitney U test. Dichotomous variables are presented as numbers (%) and analysed with Chi-Squared test. Results From May 2008 to March 2018, RV diastolic data were available in 69 pts. Indications for LTx were COPD in 51 pts (73.9%), fibrosis in 12 (17.4%), bronchiectasis in 1 pt (1.5%) and other indications in 5 (7.2%). RV diastolic function was categorized as abnormal in 47pts. One-year survival rate was 72.3% (34/47) in the group with early RV diastolic dysfunction and 95.4% (1/22) in the group with normal RV diastolic function (P = 0.027). Discussion Based on these retrospective TOE data in this single-center experience, occurrence of early RV diastolic dysfunction after LTx seems to be associated with impaired 1-year survival. Further prospective data are needed to confirm these preliminary observations.

In order to address the hypothesis that seeds from ecologically and geographically diverse plants harbor characteristic epiphytic microbiota, we characterized the bacterial and fungal microbiota associated with Triticum and Brassica seed surfaces. The total microbial complement was determined by amplification and sequencing of a fragment of chaperonin 60 (cpn60). Specific microorganisms were quantified by qPCR. Bacteria and fungi corresponding to operational taxonomic units (OTU) that were identified in the sequencing study were isolated and their interactions examined. A total of 5477 OTU were observed from seed washes. Neither total epiphytic bacterial load nor community richness/evenness was significantly different between the seed types; 578 OTU were shared among all samples at a variety of abundances. Hierarchical clustering revealed that 203 were significantly different in abundance on Triticum seeds compared with Brassica. Microorganisms isolated from seeds showed 99-100% identity between the cpn60 sequences of the isolates and the OTU sequences from this shared microbiome. Bacterial strains identified as Pantoea agglomerans had antagonistic properties toward one of the fungal isolates (Alternaria sp.), providing a possible explanation for their reciprocal abundances on both Triticum and Brassica seeds. cpn60 enabled the simultaneous profiling of bacterial and fungal microbiota and revealed a core seed-associated microbiota shared between diverse plant genera.

A time-course analysis was conducted of thermophilic anaerobic digestion of dairy manure and wheat distillery thin stillage. Sequencing of chaperonin targets provided a phylogenetic survey of both bacteria and archaea in the digestate, along with an appraisal of the diversity of the reactor microbiome. A total of 1129 bacterial operational taxonomic units (OTU) were detected in the reactors, with OTU related to Clostridium becoming numerically dominant by day 7, and Acetivibrio-related OTU by day 35. Archaeal communities were less diverse, with 19 OTU detected representing both acetoclastic and hydrogenotrophic methanogens. Regardless of input material, the same organisms came to dominate the reactors, reflecting strong selective pressures present in the digesters. Principal coordinate analysis of the microbial communities showed that the bacterial communities clustered based on factors other than input material. Bacterial and archaeal OTU were identified with significant correlations to performance parameters, suggesting important roles in the methane production pathway.

The very nature of lignocellulosic biomass presents resistance and recalcitrance to biological and chemical degradation during enzymatic hydrolysis and the subsequent fermentation process. This leads to a very low conversion rate, which makes the process economically unfeasible. In this study, alkaline (NaOH) pretreatment was applied on barley straw, using radio-frequency-based dielectric heating to enhance accessibility and digestibility during the subsequent stage of enzymatic hydrolysis. Three levels of temperature (70, 80, and 90 °C), five levels of biomass:NaOH solution ratio (1:4, 1:5, 1:6, 1:7, and 1:8), 1 h equilibration time, screen size of 1.6 mm, 1% w/v NaOH concentration, and 20 min residence time were used for the pretreatment. The effect of the alkaline pretreatment was evaluated through chemical composition analysis of the pretreated and non-treated biomass samples. The use of NaOH solution and the biomass:NaOH solution ratio played a vital role in the breakdown of the lignified matrix. The ratio 1:6 at the four temperatures studied was determined to be the optimal treatment conditions. Radio frequency-assisted alkaline pretreatment resulted in lower acid insoluble lignin and higher total acid soluble lignin moieties. Based on the obtained data, we predict that this pretreatment will decrease the required amount and cost of enzymes by up to 64% compared to using non-treated biomass.

Naturally occurring nitrogen-fixing symbionts from root nodules of caragana (Caragana arborescens) growing in central Saskatchewan were isolated following surface sterilization of caragana root nodules and squashing and spreading of the contents on yeast extract – mannitol medium. The symbiotic nature of the strains was confirmed following inoculation onto surface-sterilized C. arborescens seed in a gnotobiotic Leonard jar system. The Rhizobium isolates from C. arborescens root nodules were intermediate in generation time (g) (mean g of 5 isolates was 6.41 h) compared with the fast growers, Rhizobium leguminosarum NRG457 (g: 4.44 h), Rhizobium tropici 899 (g: 3.19 h), and Sinorhizobium meliloti BALSAC (g: 3.45 h), but they were faster than the slow-growing Bradyrhizobium japonicum USDA 110 (g: 13.86 h) and similar to Mesorhizobium amorphae (g: 7.76 h). Nitrogen derived from fixation by measuring changes in δ15N natural abundance in plant tissue confirmed the effectiveness of the strains; approximately 80% N2 from fixation. Strain identification was carried out by determining the sequences of 3 genes: 16S rRNA-encoding genes, cpn60, and recA. This analysis determined that the symbiotic partner of Canadian C. arborescens belongs to the genus Mesorhizobium and seems more related to M. loti than to previously described caragana symbionts like M. caraganae. This is the first report of Mesorhizobium sp. nodulating C. arborescens in western Canada.

Pretreatment of biomass is required in order to efficiently access the energy potential of the substrate. Many pretreatment strategies have been described, which are typically thermochemical or the...