First case report of multivisceral transplant from a deceased cardiac death donor.

The current shortage of pediatric multivisceral donors accounts for the long time and mortality on the waiting list of pediatric patients. The use of donors after cardiac death, especially after the outbreak of normothermic regional perfusion, has increased in recent years for all solid organs except the intestine, mainly because of its higher susceptibility to ischemia-reperfusion injury. We present the first literature case of multivisceral donors after cardiac death transplantation in a 13-month-old recipient from a 2.5-month-old donor. Once exitus was certified, an extracorporeal membrane oxygenation circuit was established, cannulating the aorta and infrarenal vena cava, while the supra-aortic branches were clamped. The abdominal organs completely recovered from ischemia through normothermic regional perfusion (extracorporeal membrane oxygenation initially and beating heart later). After perfusion with the preservation solution, the multivisceral graft was uneventfully implanted. Two months later, the patient was discharged without any complications. This case demonstrates the possibility of reducing the time spent on the waiting list for these patients.

Molecular Identification and Acid Stress Response of an Acidithiobacillus thiooxidans Strain Isolated from Rio Tinto (Spain).

Acidithiobacillus thiooxidans is of paramount importance in the development of biomining technologies. Being widely recognized as an extreme acidophile, extensive research has been dedicated to understanding its significant role in the extraction of several ores in recent years. However, there still exist significant molecular uncertainties surrounding this species. This study focuses on developing a taxonomic assignment method based on the sequencing of the 16S-5S rRNA cluster, along with a qPCR-based technology enabling precise growth determination. Additionally, an approach to understanding its response to acid stress is explored through RT-PCR and MALDI-TOF analysis. Our findings indicate that when subjected to pH levels below 1, the cell inhibits central (carbon fixation and metabolism) and energy (sulfur metabolism) metabolism, as well as chaperone synthesis, suggesting a potential cellular collapse. Nevertheless, the secretion of ammonia is enhanced to raise the environmental pH, while fatty acid synthesis is upregulated to reinforce the cell membrane.

Biodegradation of Pine Processionary Caterpillar Silk Is Mediated by Elastase- and Subtilisin-like Proteases.

Pine processionary caterpillar nests are made from raw silk. Fibroin protein is the main component of silk which, in the case of pine processionary caterpillar, has some unusual properties such as a higher resistance to chemical hydrolysis. Isolation of microorganisms naturally present in silk nests led to identification of Bacillus licheniformis and Pseudomonas aeruginosa strains that in a defined minimal medium were able to carry out extensive silk biodegradation. A LasB elastase-like protein from P. aeruginosa was shown to be involved in silk biodegradation. A recombinant form of this protein expressed in Escherichia coli and purified by affinity chromatography was able to efficiently degrade silk in an in vitro assay. However, silk biodegradation by B. licheniformis strain was mediated by a SubC subtilisin-like protease. Homologous expression of a subtilisin Carlsberg encoding gene (subC) allowed faster degradation compared to the biodegradation kinetics of a wildtype B. licheniformis strain. This work led to the identification of new enzymes involved in biodegradation of silk materials, a finding which could lead to possible applications for controlling this pest and perhaps have importance from sanitary and biotechnological points of view.

Therapeutic Neuroprotection by an Engineered Neurotrophin that Selectively Activates Tropomyosin Receptor Kinase (Trk) Family Neurotrophin Receptors but Not the p75 Neurotrophin Receptor.

The neurotrophin growth factors bind and activate two types of cell surface receptors: the tropomyosin receptor kinase (Trk) family and p75. TrkA, TrkB, and TrkC are bound preferentially by nerve growth factor, brain-derived neurotrophic factor, and neurotrophin 3 (NT3), respectively, to activate neuroprotective signals. The p75 receptors are activated by all neurotrophins, and paradoxically in neurodegenerative disease p75 is upregulated and mediates neurotoxic signals. To test neuroprotection strategies, we engineered NT3 to broadly activate Trk receptors (mutant D) or to reduce p75 binding (mutant RK). We also combined these features in a molecule that activates TrkA, TrkB, and TrkC but has reduced p75 binding (mutant DRK). In neurodegenerative disease mouse models in vivo, the DRK protein is a superior therapeutic agent compared with mutant D, mutant RK, and wild-type neurotrophins and protects a broader range of stressed neurons. This work rationalizes a therapeutic strategy based on the biology of each type of receptor, avoiding activation of p75 toxicity while broadly activating neuroprotection in stressed neuronal populations expressing different Trk receptors. SIGNIFICANCE STATEMENT: The neurotrophins nerve growth factor, brain-derived neurotrophic factor, and neurotrophin 3 each can activate a tropomyosin receptor kinase (Trk) A, TrkB, or TrkC receptor, respectively, and all can activate a p75 receptor. Trks and p75 mediate opposite signals. We report the engineering of a protein that activates all Trks, combined with low p75 binding, as an effective therapeutic agent in vivo.

Delayed introduction of sirolimus in paediatric intestinal transplant recipients: indications and long-term benefits.

To review our experience using sirolimus in a single centre paediatric intestinal transplantation cohort. Intestinal transplant patients with more than 3 months follow-up were divided into two groups according to their immunosuppression regimen: tacrolimus, (TAC group, n = 45 grafts) or sirolimus (SRL group, n = 38 grafts), which included those partially or completely converted from tacrolimus to sirolimus. The indications to switch were tacrolimus side effects and immunological complications. Survival and complications were retrospectively analysed comparing both groups. SRL was introduced 9 months (0 months-16.9 years) after transplant. The main cause for conversion was worsening renal function (45%), followed by haemolytic anaemia (21%) and graft-versus-host-disease (16%). Both groups showed a similar overall patient/graft survival (P = 0.76/0.08) and occurrence of rejection (24%/17%, P = 0.36). Immunological complications did not recur after conversion. Renal function significantly improved in most SRL patients. After a median follow-up of 65.17 months, 28/46 survivors were on SRL, 26 with monotherapy, with good graft function. Over one-third of our patients eventually required SRL conversion that allowed to improve their kidney function and immunological events, without entailing additional complications or survival impairment. Further trials are warranted to clarify the potential improvement of the standard tacrolimus maintenance by sirolimus conversion or addition.

Antagonism of proNGF or its receptor p75NTR reverses remodelling and improves bladder function in a mouse model of diabetic voiding dysfunction.

AIMS/HYPOTHESIS: Although 80% of diabetic patients will suffer from voiding difficulties and urinary symptoms, defined as diabetic voiding dysfunction (DVD), therapeutic targets and treatment options are limited. We hypothesise that the blockade of the pro-nerve growth factor (NGF)/p75 neurotrophin receptor (p75NTR) axis by an anti-proNGF monoclonal antibody or by a small molecule p75NTR antagonist (THX-B) can restore bladder remodelling (represented by bladder weight) in an animal model of DVD. Secondary outcomes of the study include improvements in bladder compliance, contractility and morphology, as well as in voiding behaviour, proNGF/NGF balance and TNF-α expression. METHODS: In a streptozotocin-induced mouse model of diabetes, diabetic mice received either a blocking anti-proNGF monoclonal antibody or a p75NTR antagonist small molecule as weekly systemic injections for 4 weeks. Animals were tested at baseline (at 2 weeks of diabetes induction), and after 2 and 4 weeks of treatment. Outcomes measured were voiding function with voiding spot assays and cystometry. Bladders were assessed by histological, contractility and protein expression assays. RESULTS: Diabetic mice showed features of DVD as early as 2 weeks after diabetes diagnosis (baseline) presented by hypertrophy, reduced contractility and abnormal cystometric parameters. Following treatment initiation, a twofold increase (p < 0.05) in untreated diabetic mouse bladder weight and thickness compared with non-diabetic controls was observed, and this change was reversed by p75NTR antagonism (37% reduction in bladder weight compared with untreated diabetic mice [95% CI 14%, 60%]) after 4 weeks of treatment. However, blocking proNGF did not help to reverse bladder hypertrophy. While diabetic mice had significantly worse cystometric parameters and contractile responses than non-diabetic controls, proNGF antagonism normalised bladder compliance (0.007 [Q1-Q3; 0.006-0.009] vs 0.015 [Q1-Q3; 0.014-0.029] ml/cmH2O in untreated diabetic mice, representing 62% reduction [95% CI 8%, 110%], p < 0.05) and contractility to KCl, carbachol and electrical field stimulation (p < 0.05 compared with the diabetic group) after 2 weeks of treatment. These effects were not observed after 4 weeks of treatment with proNGF antagonist. p75NTR antagonism did not show important improvements in cystometric parameters after 2 weeks of treatment. Slightly improved bladder compliance (0.01 [Q1-Q3; 0.009-0.012] vs 0.013 [Q1-Q3; 0.011-0.016] ml/cmH2O for untreated diabetic mice) was seen in the p75NTR antagonist-treated group after 4 weeks of treatment with significantly stabilised contractile responses to KCl, carbachol and electric field stimulation (p < 0.05 for each) compared with diabetic mice. Bladder dysfunction observed in diabetic mice was associated with a significant increase in bladder proNGF/NGF ratio (3.1 [±1.2] vs 0.26 [±0.04] ng/pg in control group, p < 0.05 at week 2 of treatment) and TNF-α (p < 0.05). The proNGF/NGF ratio was partially reduced (about 60% reduction) with both treatments (1.03 [±0.6] ng/pg for proNGF antibody-treated group and 1.4 [±0.76] ng/pg for p75NTR blocker-treated group after 2 weeks of treatment), concomitant with a significant decrease in the bladder levels of TNF-α (p < 0.05), despite persistent hyperglycaemia. CONCLUSIONS/INTERPRETATION: Our findings indicate that blockade of proNGF and the p75NTR receptor in diabetes can impede the development and progression of DVD. The reported improvements in morphological and functional features in our DVD model validates the proNGF/p75NTR axis as a potential therapeutic target in this pathology. Graphical abstract.

Advances in the control of phytopathogenic fungi that infect crops through their root system.

Productivity and economic sustainability of many herbaceous and woody crops are seriously threatened by numerous phytopathogenic fungi. While symptoms associated with phytopathogenic fungal infections of aerial parts (leaves, stems and fruits) are easily observable and therefore recognizable, allowing rapid or preventive action to control this type of infection, the effects produced by soil-borne fungi that infect plants through their root system are more difficult to detect. The fact that these fungi initiate infection and damage underground implies that the first symptoms are not as easily noticeable, and therefore both crop yield and plant survival are frequently severely compromised by the time the infection is found. In this paper we will review and discuss recent insights into plant-microbiota interactions in the root system crucial to understanding the beginning of the infectious process. We will also review different methods for diminishing and controlling the infection rate by phytopathogenic fungi penetrating through the root system including both the traditional use of biocontrol agents such as antifungal compounds as well as some new strategies that could be used because of their effective application, such as nanoparticles, virus-based nanopesticides, or inoculation of plant material with selected endophytes. We will also review the possibility of modeling and influencing the composition of the microbial population in the rhizosphere environment as a strategy for nudging the plant-microbiome interactions toward enhanced beneficial outcomes for the plant, such as controlling the infectious process.

Intestinal failure in adults and children.

Intestinal failure (IF) is rare, but it represents one of the most complex medical-surgical management pathologies, both in adults and children. The first-line treatment is parenteral nutrition (PN). However, new alternatives in the field of intestinal rehabilitation have opened up in recent decades, with the rise of multidisciplinary teams and the development of new hormone therapies as the first non-symptomatic approach to IF.

Diurnal variations in intraocular pressure and central corneal thickness and the correlation between these factors in dogs.

OBJECTIVE: To study the diurnal variation in intraocular pressure (IOP) and central corneal thickness (CCT) in healthy Beagles by rebound tonometry and ultrasonic pachymetry, respectively, in addition to determining whether a correlation exists between these two variables. ANIMALS STUDIED: Twenty eyes from 10 healthy Beagle dogs were included in the study. PROCEDURES: The IOP and CCT were measured by rebound tonometry and ultrasonic pachymetry, respectively, at 2-h intervals over an 8-hour period between 10:00 and 18:00. RESULTS: The mean values (± SD) of IOP obtained were 11.45 ± 2.96 at 10:00, 10.00 ± 1.89 at 12:00, 8.25 ± 1.62 at 14:00, 7.05 ± 1.05 at 16:00, and 6.55 ± 1.36 at 18:00. The mean values (± SD) of CCT obtained were 554.95 ± 72.41 at 10:00, 549.20 ± 69.10 at 12:00, 566.15 ± 80.56 at 14:00, 545.45 ± 70.19 at 16:00, and 538.30 ± 73.33 at 18:00. The IOP and CCT of dogs were found to decrease progressively from the first to the last measurement. There were statistically significant differences between the IOP (P = 0.000) and CCT values (P = 0.032) measured at different times of the day. There was no effect or interaction between gender and eye with the dependent variables. The IOP and CCT were found to be positively correlated (r = 0.213, P = 0.034). The regression equation demonstrated that for every 100 µm increase in CCT, there was an elevation in IOP by 0.8 mmHg. CONCLUSIONS: The CCT and IOP values were lower in the afternoon/evening than in the morning, and these were positively correlated. Both findings are important for the diagnostic interpretation of IOP values in dogs.

p75NTR and Its Ligand ProNGF Activate Paracrine Mechanisms Etiological to the Vascular, Inflammatory, and Neurodegenerative Pathologies of Diabetic Retinopathy.

UNLABELLED: In many diseases, expression and ligand-dependent activity of the p75(NTR) receptor can promote pericyte and vascular dysfunction, inflammation, glial activation, and neurodegeneration. Diabetic retinopathy (DR) is characterized by all of these pathological events. However, the mechanisms by which p75(NTR) may be implicated at each stage of DR pathology remain poorly understood. Using a streptozotocin mouse model of diabetic retinopathy, we report that p75(NTR) is upregulated very early in glia and in pericytes to mediate ligand-dependent induction of inflammatory cytokines, disruption of the neuro-glia-vascular unit, promotion of blood-retina barrier breakdown, edema, and neuronal death. In a mouse model of oxygen-induced retinopathy, mimicking proliferative DR, p75(NTR)-dependent inflammation leads to ischemia and pathological angiogenesis through Semaphorin 3A. The acute use of antagonists of p75(NTR) or antagonists of the ligand proNGF suppresses each distinct phase of pathology, ameliorate disease, and prevent disease progression. Thus, our study documents novel disease mechanisms and validates druggable targets for diabetic retinopathy. SIGNIFICANCE STATEMENT: Diabetic retinopathy (DR) affects an estimated 250 million people and has no effective treatment. Stages of progression comprise pericyte/vascular dysfunction, inflammation, glial activation, and neurodegeneration. The pathophysiology of each stage remains unclear. We postulated that the activity of p75NTR may be implicated. We show that p75NTR in glia and in pericytes mediate ligand-dependent induction of inflammatory cytokines, disruption of the neuro-glia-vascular unit, promotion of blood-retina barrier breakdown, edema, and neuronal death. p75NTR-promoted inflammation leads to ischemia and angiogenesis through Semaphorin 3A. Antagonists of p75NTR or antagonists of proNGF suppress each distinct phase of pathology, ameliorate disease, and prevent disease progression. Our study documents novel mechanisms in a pervasive disease and validates druggable targets for treatment.

Use of Endophytic and Rhizosphere Actinobacteria from Grapevine Plants To Reduce Nursery Fungal Graft Infections That Lead to Young Grapevine Decline.

Endophytic and rhizosphere actinobacteria isolated from the root system of 1-year-old grafted Vitis vinifera plants were evaluated for their activities against fungi that cause grapevine trunk diseases. A total of 58 endophytic and 94 rhizosphere isolates were tested. Based on an in vitro bioassay, 15.5% of the endophytic isolates and 30.8% of the rhizosphere isolates exhibited antifungal activity against the fungal pathogen Diplodia seriata, whereas 13.8% of the endophytic isolates and 16.0% of the rhizosphere isolates showed antifungal activity against Dactylonectria macrodidyma (formerly Ilyonectria macrodidyma). The strains which showed the greatest in vitro efficacy against both pathogens were further analyzed for their ability to inhibit the growth of Phaeomoniella chlamydospora and Phaeoacremonium minimum (formerly Phaeoacremonium aleophilum). Based on their antifungal activity, three rhizosphere isolates and three endophytic isolates were applied on grafts in an open-root field nursery in a 3-year trial. The field trial led to the identification of one endophytic strain, Streptomyces sp. VV/E1, and two rhizosphere isolates, Streptomyces sp. VV/R1 and Streptomyces sp. VV/R4, which significantly reduced the infection rates produced by the fungal pathogens Dactylonectria sp., Ilyonectria sp., P. chlamydospora, and P. minimum, all of which cause young grapevine decline. The VV/R1 and VV/R4 isolates also significantly reduced the mortality level of grafted plants in the nursery. This study shows that certain actinobacteria could represent a promising new tool for controlling fungal trunk pathogens that infect grapevine plants through the root system in nurseries.IMPORTANCE Grapevine trunk diseases are a major threat to the wine and grape industry worldwide. They cause a significant reduction in yields as well as in grape quality, and they can even cause plant death. Trunk diseases are caused by fungal pathogens that enter through pruning wounds and/or the root system. Although different strategies have recently been developed to protect pruning wounds using antifungal compounds (natural or synthetic) or biocontrol agents, no tools are yet available for controlling soil pathogens that infect plants through their root system. This study shows that different actinobacterial isolates, when applied to grafts in a nursery, can significantly reduce the infection rate caused by fungal pathogens that enter through the root system. This is a new, promising, and green alternative for preventing the decline of young grapevines in nurseries and vineyards.

Diurnal variations of central corneal thickness and intraocular pressure in dogs from 8:00 am to 8:00 pm.

Diurnal variations in central corneal thickness (CCT) and intraocular pressure (IOP) and their relationships were studied in healthy dogs. Central corneal thickness was measured by ultrasonic pachymetry and IOP by applanation tonometry in 16 beagle dogs. Measurements were taken every 90 min over 12 h (08:00 am to 08:00 pm). The mean CCT and IOP values obtained during the sampling period were 545.6 ± 21.7 µm (range: 471 to 595 µm) and 15 ± 2.2 mmHg (range: 10 to 19 mmHg), respectively. The CCT and IOP showed statistically significant decreases at 6:30 pm and 5:00 pm, respectively (P < 0.001). Central corneal thickness and IOP values were lower in the afternoon/evening than in the morning and were positively correlated. Both findings are important for the diagnostic interpretation of IOP values in dogs.

Variations diurnes de l'épaisseur de la cornée centrale et de la pression intraoculaire chez les chiens, de 8 heures à 20 heures. Les variations quotidiennes de l'épaisseur de la cornée centrale (ECC) par pachymétrie ultrasonore, ainsi que la pression intraoculaire (PIO) obtenue par tonométrie à aplanissement, ont été étudiées chez seize chiens beagle en bonne santé. La relation entre ces deux paramètres a aussi été évaluée. Les mesures ont été effectuées toutes les 90 minutes durant douze heures (08:00 am à 08:00 pm). Les valeurs moyennes de ECC et PIO obtenues durant la période de l'expérience ont été respectivement de 545,6 ± 21,7 µm (valeurs extrêmes de 471 à 595 µm) et de 15 ± 2,2 mmHg (valeurs extrêmes de 10 à 19 mmHg). La mesure de l'épaisseur cornéenne ainsi que la pression intraoculaire ont montré statistiquement une baisse significative après 6:30 pm et 05:00 pm (P < 0,001) respectivement. Les valeurs de l'ECC et de la PIO étaient plus basses l'après-midi que le matin, avec une corrélation positive entre les deux.(Traduit par les auteurs).

Uncommon acute neurologic presentation of canine distemper in 4 adult dogs.

Four uncommon cases of canine distemper (CD) were diagnosed in vaccinated adult dogs. All dogs had acute onset of neurologic signs, including seizures, abnormal mentation, ataxia, and proprioceptive deficits. Polymerase chain reaction for CD virus was positive on cerebrospinal fluid in 2 cases. Due to rapid deterioration the dogs were euthanized and CD was confirmed by postmortem examination.

Rare présentation neurologique aiguë de la maladie de Carré chez 4 chiens adultes. Quatre cas peu communs de maladie de Carré chez des chiens adultes vaccinés. Tous les cas ont présenté un début aigu ou suraigu des signes neurologiques, comportant principalement des crises épileptiques, altération de l'état mental, ataxie, et déficits proprioceptifs. Dans deux cas, la PCR a été positive à la maladie de Carré dans le liquide céphalorachidien. En raison de la progression rapide des signes, les chiens ont été euthanasiés et la maladie de Carré confirmée par la nécropsie.(Traduit par Ana Roman).

GD2 and GD3 gangliosides as diagnostic biomarkers for all stages and subtypes of epithelial ovarian cancer.

Background: Ovarian cancer (OC) is the deadliest gynecological cancer, often diagnosed at advanced stages. A fast and accurate diagnostic method for early-stage OC is needed. The tumor marker gangliosides, GD2 and GD3, exhibit properties that make them ideal potential diagnostic biomarkers, but they have never before been quantified in OC. We investigated the diagnostic utility of GD2 and GD3 for diagnosis of all subtypes and stages of OC. Methods: This retrospective study evaluated GD2 and GD3 expression in biobanked tissue and serum samples from patients with invasive epithelial OC, healthy donors, non-malignant gynecological conditions, and other cancers. GD2 and GD3 levels were evaluated in tissue samples by immunohistochemistry (n=299) and in two cohorts of serum samples by quantitative ELISA. A discovery cohort (n=379) showed feasibility of GD2 and GD3 quantitative ELISA for diagnosing OC, and a subsequent model cohort (n=200) was used to train and cross-validate a diagnostic model. Results: GD2 and GD3 were expressed in tissues of all OC subtypes and FIGO stages but not in surrounding healthy tissue or other controls. In serum, GD2 and GD3 were elevated in patients with OC. A diagnostic model that included serum levels of GD2+GD3+age was superior to the standard of care (CA125, p<0.001) in diagnosing OC and early-stage (I/II) OC. Conclusion: GD2 and GD3 expression was associated with high rates of selectivity and specificity for OC. A diagnostic model combining GD2 and GD3 quantification in serum had diagnostic power for all subtypes and all stages of OC, including early stage. Further research exploring the utility of GD2 and GD3 for diagnosis of OC is warranted.

The Grapevine Microbiome to the Rescue: Implications for the Biocontrol of Trunk Diseases.

Grapevine trunk diseases (GTDs) are one of the most devastating pathologies that threaten the survival and profitability of vineyards around the world. Progressive banning of chemical pesticides and their withdrawal from the market has increased interest in the development of effective biocontrol agents (BCAs) for GTD treatment. In recent years, considerable progress has been made regarding the characterization of the grapevine microbiome, including the aerial part microbiome (flowers, berries and leaves), the wood microbiome, the root environment and vineyard soil microbiomes. In this work, we review these advances especially in relation to the etiology and the understanding of the composition of microbial populations in plants affected by GTDs. We also discuss how the grapevine microbiome is becoming a source for the isolation and characterization of new, more promising BCAs that, in the near future, could become effective tools for controlling these pathologies.

Chronic and acute models of retinal neurodegeneration TrkA activity are neuroprotective whereas p75NTR activity is neurotoxic through a paracrine mechanism.

In normal adult retinas, NGF receptor TrkA is expressed in retinal ganglion cells (RGC), whereas glia express p75(NTR). During retinal injury, endogenous NGF, TrkA, and p75(NTR) are up-regulated. Paradoxically, neither endogenous NGF nor exogenous administration of wild type NGF can protect degenerating RGCs, even when administered at high frequency. Here we elucidate the relative contribution of NGF and each of its receptors to RGC degeneration in vivo. During retinal degeneration due to glaucoma or optic nerve transection, treatment with a mutant NGF that only activates TrkA, or with a biological response modifier that prevents endogenous NGF and pro-NGF from binding to p75(NTR) affords significant neuroprotection. Treatment of normal eyes with an NGF mutant-selective p75(NTR) agonist causes progressive RGC death, and in injured eyes it accelerates RGC death. The mechanism of p75(NTR) action during retinal degeneration due to glaucoma is paracrine, by increasing production of neurotoxic proteins TNF-α and α(2)-macroglobulin. Antagonists of p75(NTR) inhibit TNF-α and α(2)-macroglobulin up-regulation during disease, and afford neuroprotection. These data reveal a balance of neuroprotective and neurotoxic mechanisms in normal and diseased retinas, and validate each neurotrophin receptor as a pharmacological target for neuroprotection.

Using Rhizosphere Phosphate Solubilizing Bacteria to Improve Barley (Hordeum vulgare) Plant Productivity.

On average less than 1% of the total phosphorous present in soils is available to plants, making phosphorous one of the most limiting macronutrients for crop productivity worldwide. The aim of this work was to isolate and select phosphate solubilizing bacteria (PSB) from the barley rhizosphere, which has other growth promoting traits and can increase crop productivity. A total of 104 different bacterial isolates were extracted from the barley plant rhizosphere. In this case, 64 strains were able to solubilize phosphate in agar plates. The 24 strains exhibiting the highest solubilizing index belonged to 16 different species, of which 7 isolates were discarded since they were identified as putative phytopathogens. The remaining nine strains were tested for their ability to solubilize phosphate in liquid medium and in pot trials performed in a greenhouse. Several of the isolated strains (Advenella mimigardefordensis, Bacillus cereus, Bacillus megaterium and Burkholderia fungorum) were able to significantly improve levels of assimilated phosphate, dry weight of ears and total starch accumulated on ears compared to non-inoculated plants. Since these strains were able to increase the growth and productivity of barley crops, they could be potentially used as microbial inoculants (biofertilizers).

Inhibiting the MNK1/2-eIF4E axis impairs melanoma phenotype switching and potentiates antitumor immune responses.

Melanomas commonly undergo a phenotype switch, from a proliferative to an invasive state. Such tumor cell plasticity contributes to immunotherapy resistance; however, the mechanisms are not completely understood and thus are therapeutically unexploited. Using melanoma mouse models, we demonstrated that blocking the MNK1/2-eIF4E axis inhibited melanoma phenotype switching and sensitized melanoma to anti-PD-1 immunotherapy. We showed that phospho-eIF4E-deficient murine melanomas expressed high levels of melanocytic antigens, with similar results verified in patient melanomas. Mechanistically, we identified phospho-eIF4E-mediated translational control of NGFR, a critical effector of phenotype switching. Genetic ablation of phospho-eIF4E reprogrammed the immunosuppressive microenvironment, exemplified by lowered production of inflammatory factors, decreased PD-L1 expression on dendritic cells and myeloid-derived suppressor cells, and increased CD8+ T cell infiltrates. Finally, dual blockade of the MNK1/2-eIF4E axis and the PD-1/PD-L1 immune checkpoint demonstrated efficacy in multiple melanoma models regardless of their genomic classification. An increase in the presence of intratumoral stem-like TCF1+PD-1+CD8+ T cells, a characteristic essential for durable antitumor immunity, was detected in mice given a MNK1/2 inhibitor and anti-PD-1 therapy. Using MNK1/2 inhibitors to repress phospho-eIF4E thus offers a strategy to inhibit melanoma plasticity and improve response to anti-PD-1 immunotherapy.

Alternative Splicing of a Receptor Intracellular Domain Yields Different Ectodomain Conformations, Enabling Isoform-Selective Functional Ligands.

Events at a receptor ectodomain affect the intracellular domain conformation, activating signal transduction (out-to-in conformational effects). We investigated the reverse direction (in-to-out) where the intracellular domain may impact on ectodomain conformation. The primary sequences of naturally occurring TrkC receptor isoforms (TrkC-FL and TrkC.T1) only differ at the intracellular domain. However, owing to their differential association with Protein Disulfide Isomerase the isoforms have different disulfide bonding and conformations at the ectodomain. Conformations were exploited to develop artificial ligands, mAbs, and small molecules, with isoform-specific binding and biased activation. Consistent, the physiological ligands NT-3 and PTP-sigma bind both isoforms, but NT-3 activates all signaling pathways, whereas PTP-sigma activates biased signals. Our data support an "in-to-out" model controlling receptor ectodomain conformation, a strategy that enables heterogeneity in receptors, ligands, and bioactivity. These concepts may be extended to the many wild-type or oncogenic receptors with known isoforms.

New mutations associated with Hirschsprung disease.

INTRODUCTION: Hirschsprung Disease is caused by an impairment in cell migration from the neural crest to the gastrointestinal tract, resulting in an absence of neurons in the myenteric plexus. Many mutations in several genes have been associated to Hirschsprung disease; most of them affecting the RET proto-oncogen pathway. The purpose of this study is the description of novel and known mutations in genes associated to Hirschsprung disease and their prognostic implications. MATERIAL AND METHODS: Retrospective analysis of patients with Hirschsprung disease and positive genetic studies evaluated from 1970 to 2013. RESULTS: We found 21 positive genetic studies in the global series, 17 of them involving the RET proto-oncogene: Two of the mutations are novel and they have not been reported in the medical literature. CONCLUSIONS: The RET protooncogene is the main gene associated with Hirschsprung disease. There are still multiple unknown mutations related to the pathogenesis of the disease. The study of this gene must be part of the work-up of all patients with Hirschsprung disease, as well as their first degree relatives if the mutation is associated with MEN2A and MEN2B syndromes.