Heterogeneous oxygen availability affects the titer and topology but not the fidelity of plasmid DNA produced by Escherichia coli

Jaen KE, Sigala J-C, Olivares-Hernandez R, Niehaus K, Lara AR. Heterogeneous oxygen availability affects the titer and topology but not the fidelity of plasmid DNA produced by Escherichia coli. BMC BIOTECHNOLOGY. 2017;17(1): 60.Background: Dissolved oxygen tension (DOT) is hardly constant and homogenously distributed in a bioreactor, which can have a negative impact in the metabolism and product synthesis. However, the effects of DOT on plasmid DNA (pDNA) production and quality have not been thoroughly investigated. In the present study, the effects of aerobic (DOT >= 30% air sat.), microaerobic (constant DOT = 3% air sat.) and oscillatory DOT (from 0 to 100% air sat.) conditions on pDNA production, quality and host performance were characterized. Results: Microaerobic conditions had little effect on pDNA production, supercoiled fraction and sequence fidelity. By contrast, oscillatory DOT caused a 22% decrease in pDNA production compared with aerobic cultures. Although in aerobic cultures the pDNA supercoiled fraction was 98%, it decreased to 80% under heterogeneous DOT conditions. The different oxygen availabilities had no effect on the fidelity of the produced pDNA. The estimated metabolic fluxes indicated substantial differences at the level of the pentose phosphate pathway and TCA cycle under different conditions. Cyclic changes in fermentative pathway fluxes, as well as fast shifts in the fluxes through cytochromes, were also estimated. Model-based genetic modifications that can potentially improve the process performance are suggested. Conclusions: DOT heterogeneities strongly affected cell performance, pDNA production and topology. This should be considered when operating or scaling-up a bioreactor with deficient mixing. Constant microaerobic conditions affected the bacterial metabolism but not the amount or quality of pDNA. Therefore, pDNA production in microaerobic cultures may be an alternative for bioreactor operation at higher oxygen transfer rates

An Autophagy-Related Kinase Is Essential for the Symbiotic Relationship between Phaseolus vulgaris and Both Rhizobia and Arbuscular Mycorrhizal Fungi

Eukaryotes contain three types of lipid kinases that belong to the phosphatidylinositol 3-kinase (PI3K) family. In plants and Saccharomyces cerevisiae, only PI3K class III family members have been identified. These enzymes regulate the innate immune response, intracellular trafficking, autophagy, and senescence. Here, we report that RNAi-mediated downregulation of common bean (Phaseolus vulgaris) PI3K severely impaired symbiosis in composite P. vulgaris plants with endosymbionts such as Rhizobium tropici and Rhizophagus irregularis. Downregulation of Pv-PI3K was associated with a marked decrease in root hair growth and curling. Additionally, infection thread growth, root-nodule number, and symbiosome formation in root nodule cells were severely affected. Interestingly, root colonization by AM fungi and the formation of arbuscules were also abolished in PI3K loss-of-function plants. Furthermore, the transcript accumulation of genes encoding proteins known to interact with PI3K to form protein complexes involved in autophagy was drastically reduced in these transgenic roots. RNAi-mediated downregulation of one of these genes, Beclin1/Atg6, resulted in a similar phenotype as observed for transgenic roots in which Pv-PI3K had been downregulated. Our findings show that an autophagy-related process is crucial for the mutualistic interactions of P. vulgaris with beneficial microorganismsInstituto de Fisiología y Recursos Genéticos VegetalesFil: Estrada-Navarrete, Georgina. Universidad Nacional Autónoma de México. Instituto de Biotecnología. Departamento de Biología Molecular de Plantas; MéxicoFil: Cruz-Mireles, Neftaly. Universidad Nacional Autónoma de México. Instituto de Biotecnología. Departamento de Biología Molecular de Plantas; MéxicoFil: Lascano, Hernán Ramiro. Consejo Nacional de Investigaciones Científicas y Técnicas. Unidad de Estudios Agropecuarios (UDEA); ArgentinaFil: Lascano, Hernán Ramiro. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Fisiología y Recursos Genéticos Vegetales. ArgentinaFil: Alvarado-Affantranger, Xóchitl. Universidad Nacional Autónoma de México. Instituto de Biotecnología. Laboratorio Nacional de Microscopía Avanzada; MéxicoFil: Hernández-Barrera, Alejandra. Universidad Nacional Autónoma de México. Instituto de Biotecnología. Departamento de Biología Molecular de Plantas; MéxicoFil: Barraza, Aarón. Universidad Nacional Autónoma de México. Instituto de Biotecnología. Departamento de Biología Molecular de Plantas; MéxicoFil: Olivares, Juan E. Universidad Nacional Autónoma de México. Instituto de Biotecnología. Departamento de Biología Molecular de Plantas; MéxicoFil: Arthikala, Manoj-Kumar. Universidad Nacional Autónoma de México. Escuela Nacional de Estudios Superiores-Unidad León; MéxicoFil: Cárdenas, Luis. Universidad Nacional Autónoma de México. Instituto de Biotecnología. Departamento de Biología Molecular de Plantas; MéxicoFil: Quinto, Carmen. Universidad Nacional Autónoma de México. Instituto de Biotecnología. Departamento de Biología Molecular de Plantas; MéxicoFil: Sanchez, Federico. Universidad Nacional Autónoma de México. Instituto de Biotecnología. Departamento de Biología Molecular de Plantas; Méxic

Mejoras en el diseño agronómico de la instalación de riego dirigidas al aumento de la productividad técnica y económica del agua en limonero temprano

El objetivo del trabajo fue incrementar la productividad en limonero temprano mediante mejoras en el diseño agronómico de la red de riego. El ensayo se llevó a cabo en una parcela experimental en Torre Pacheco (Murcia), en árboles adultos de limonero ‘Fino 49’ injertados sobre Citrus macrophylla Wester. Se han evaluado dos diseños del sistema de riego: diseño convencional (2L), con dos tuberías portagoteros (6 goteros árbol-1); y un diseño con mayor superficie mojada (3L), con tres tuberías (9 goteros árbol-1). Partiendo de los resultados agronómicos, se ha realizado un análisis económico comparativo entre dos diseños del sistema de riego. El diseño 3L fue el más productivo técnica y económicamente. El Producto Bruto Económico (PBE) fue un 14% superior en el diseño 3L respecto al 2L; el Producto Bruto Técnico (PBT) lo fue en un 6,2%. Este resultado se debe a dos motivos: por un lado y en mayor medida, al incremento en la proporción de limón de primer corte del diseño 3L y, por otro lado, a la disminución de limón derivado a la industria. El Margen Bruto (MB) sigue la misma pauta que el Producto Bruto (PB), ya que los costes diferenciales (CD) son de poca envergadura. El precio ponderado del kg de limón medio (PBE/PBT) es de 0,367 € kg-1 y 0,341 € kg-1, en 3L y 2L, respectivamente, y supone un ingreso de 2.631 € extras por hectárea y año a favor del sistema 3L. Por último, resaltar que el diseño 3L destaca principalmente por ser es más productivo económicamente, en relación al agua aplicada (€ m-3)

Distributed predictive secondary control with soft constraints for optimal dispatch in hybrid AC/DC microgrids

Hybrid AC/DC microgrids (H-MGs) are a prominent solution for integrating distributed generation and modern AC and DC loads. However, controlling these systems is challenging as multiple electrical variables need to be controlled and coordinated. To provide flexibility to the control system, these variables can be regulated to specific values or within secure bands. This paper proposes a set of distributed model predictive control schemes for the secondary control level to control certain variables to specific values and other variables within secure pre-defined bands into H-MGs. Specifically, optimal dispatch of active and reactive power is achieved while frequency and voltages are regulated within secure bands in H-MGs. Dynamic models of AC generators, DC generators and interlinking converters along with their novel multi-objective cost functions are developed in constrained distributed predictive optimisation problems to simultaneously achieve the aforementioned objectives via information sharing. Extensive simulation work validates the performance of this proposal

Truncating FLNC Mutations Are Associated With High-Risk Dilated and Arrhythmogenic Cardiomyopathies

BACKGROUND: Filamin C (encoded by the FLNC gene) is essential for sarcomere attachment to the plasmatic membrane. FLNC mutations have been associated with myofibrillar myopathies, and cardiac involvement has been reported in some carriers. Accordingly, since 2012, the authors have included FLNC in the genetic screening of patients with inherited cardiomyopathies and sudden death. OBJECTIVES: The aim of this study was to demonstrate the association between truncating mutations in FLNC and the development of high-risk dilated and arrhythmogenic cardiomyopathies. METHODS: FLNC was studied using next-generation sequencing in 2,877 patients with inherited cardiovascular diseases. A characteristic phenotype was identified in probands with truncating mutations in FLNC. Clinical and genetic evaluation of 28 affected families was performed. Localization of filamin C in cardiac tissue was analyzed in patients with truncating FLNC mutations using immunohistochemistry. RESULTS: Twenty-three truncating mutations were identified in 28 probands previously diagnosed with dilated, arrhythmogenic, or restrictive cardiomyopathies. Truncating FLNC mutations were absent in patients with other phenotypes, including 1,078 patients with hypertrophic cardiomyopathy. Fifty-four mutation carriers were identified among 121 screened relatives. The phenotype consisted of left ventricular dilation (68%), systolic dysfunction (46%), and myocardial fibrosis (67%); inferolateral negative T waves and low QRS voltages on electrocardiography (33%); ventricular arrhythmias (82%); and frequent sudden cardiac death (40 cases in 21 of 28 families). Clinical skeletal myopathy was not observed. Penetrance was >97% in carriers older than 40 years. Truncating mutations in FLNC cosegregated with this phenotype with a dominant inheritance pattern (combined logarithm of the odds score: 9.5). Immunohistochemical staining of myocardial tissue showed no abnormal filamin C aggregates in patients with truncating FLNC mutations. CONCLUSIONS: Truncating mutations in FLNC caused an overlapping phenotype of dilated and left-dominant arrhythmogenic cardiomyopathies complicated by frequent premature sudden death. Prompt implantation of a cardiac defibrillator should be considered in affected patients harboring truncating mutations in FLNC.Instituto de Salud Carlos III [PI11/0699, PI14/0967, PI14/01477, RD012/0042/0029, RD012/0042/0049, RD012/0042/0066, RD12/0042/0069]; Spanish Ministry of Economy and Competitiveness [SAF2015-71863-REDT]; Plan Nacional de I+D+I; Plan Estatalde I+D+I, European Regional Development Fund; Health in Code SLS

Establishment and Characterization of Immortalized Neuronal Cell Lines Derived From the Spinal Cord of Normal and Trisomy 16 Fetal Mice, an Animal Model of Down Syndrome

We report the establishment of continuously growing cell lines from spinal cords of normal and trisomy 16 fetal mice. We show that both cell lines, named M4b (derived from a normal animal) and MTh (trisomic) possess neurological markers by immunohistochemistry (neuron specific enolase, synaptophysin, microtubule associated protein-2 [MAP-2], and choline acetyltransferase) and lack glial traits (glial fibrillary acidic protein and S10

Aortic disease in Marfan syndrome is caused by overactivation of sGC-PRKG signaling by NO

AbstractThoracic aortic aneurysm, as occurs in Marfan syndrome, is generally asymptomatic until dissection or rupture, requiring surgical intervention as the only available treatment. Here, we show that nitric oxide (NO) signaling dysregulates actin cytoskeleton dynamics in Marfan Syndrome smooth muscle cells and that NO-donors induce Marfan-like aortopathy in wild-type mice, indicating that a marked increase in NO suffices to induce aortopathy. Levels of nitrated proteins are higher in plasma from Marfan patients and mice and in aortic tissue from Marfan mice than in control samples, indicating elevated circulating and tissue NO. Soluble guanylate cyclase and cGMP-dependent protein kinase are both activated in Marfan patients and mice and in wild-type mice treated with NO-donors, as shown by increased plasma cGMP and pVASP-S239 staining in aortic tissue. Marfan aortopathy in mice is reverted by pharmacological inhibition of soluble guanylate cyclase and cGMP-dependent protein kinase and lentiviral-mediated Prkg1 silencing. These findings identify potential biomarkers for monitoring Marfan Syndrome in patients and urge evaluation of cGMP-dependent protein kinase and soluble guanylate cyclase as therapeutic targets.</jats:p

Aplicación de la oxifertirrigación para optimizar los recursos hídricos en cítricos, basado en la aplicación de peróxido de hidrógeno en el agua de riego

En este trabajo se estudia la respuesta fisiológica y agronómica de la aplicación de la oxifertirrigación química, basada en la aplicación de peróxido de hidrógeno como fuente de oxígeno a nivel radicular en cítricos. El ensayo tuvo lugar durante dos campañas (2018-2019 y 2019-2020) en árboles adultos de mandarino híbrido ‘Ortanique’ ubicados en una parcela experimental del IMIDA en Torre Pacheco (Murcia). Se establecieron dos tratamientos, un tratamiento ‘Control’ (0 ppm de H2O2) y otro identificado como ‘OXI’ (50-100 ppm de H2O2 durante todo el ciclo de cultivo). Ambos tratamientos recibieron la misma cantidad de agua y de fertilizante. El H2O2 se aplicó de forma continua con una bomba dosificadora a la red de riego. Los resultados más destacados mostraron que el estado hídrico de los árboles del tratamiento ‘OXI’ fue muy similar al control. Respecto a los parámetros de intercambio gaseoso, la aplicación de H2O2 estimuló una mayor apertura estomática en el mes de septiembre de ambas campañas. Sin embargo, las ligeras alteraciones fisiológicas no han supuesto cambios sustanciales en la biometría de la planta. En la segunda campaña, la aplicación de H2O2 en el riego favoreció la acumulación de N, K y Fe en hoja, lo que permitiría reducir la dosis de fertilizante. La respuesta productiva y la eficiencia en el uso del agua no presentaron una clara mejora a la aplicación de H2O2 en el agua de riego. Los mayores niveles de N en el tratamiento ‘OXI’ afectaron negativamente a la calidad del fruto, reduciendo el porcentaje de zumo y aumentando el porcentaje de corteza. En cambio, la aplicación de H2O2 disminuyó el índice de madurez de la fruta, lo que resulta interesante de cara a retrasar la recolección en variedades tardías

Riego de un cultivo de citricos con agua marina desalinizada. resultados preliminares en suelo y planta

La escasez de agua y la creciente presión sobre los recursos hídricos en las regiones semiáridas ha extendido la utilización para el riego de recursos hídricos no convencionales, como el agua marina desalinizada (AMD). Debido a su composición en Cl-, Na+ y B3+, el riego con AMD podría causar problemas agronómicos y afectar al suelo y a los cultivos a medio y largo plazo. En este estudio, se regó una parcela de mandarinos durante 20 meses con (i) agua proporcionada por la Comunidad de Regantes del Campo de Cartagena (CR), (ii) agua marina desalinizada (AMD) y (iii) mezcla de agua 50% CR y 50% AMD (AM). Se evaluó el efecto sobre la dinámica y acumulación de los iones tóxicos Cl-, Na+ y B3+ en el suelo y en la planta. La [B3+] del agua AMD fue superior a la de CR, acumulándose en el suelo, con una concentración un 25% superior a la encontrada con CR al final del ensayo. La [B3+] en la capa superficial del suelo se correlacionó con la [B3+] en el agua y con la [B3+] en la hoja. Aunque tras 20 meses los árboles regados con AMD tuvieron una [B3+] foliar un 25% superior a la de árboles regados con CR, no presentaron síntomas de toxicidad. Las [Cl-] y [Na+] del agua fueron similares en los tres tipos de agua, superando los umbrales a partir de los cuales pueden producir toxicidad en cítricos. Las concentraciones de Cl- y Na+ en hoja permanecieron por debajo del umbral de toxicidad establecido para cítricos. Los resultados obtenidos son preliminares ya que este estudio debería extenderse durante un periodo más largo para obtener datos más concluyentes acerca de los efectos a largo plazo de la utilización de AMD tanto en el suelo como en la planta

Accuracy of clinical pallor in the diagnosis of anaemia in children: a meta-analysis

BACKGROUND: Anaemia is highly prevalent in children of developing countries. It is associated with impaired physical growth and mental development. Palmar pallor is recommended at primary level for diagnosing it, on the basis of few studies. The objective of the study was to systematically assess the accuracy of clinical signs in the diagnosis of anaemia in children. METHODS: A systematic review on the accuracy of clinical signs of anaemia in children. We performed an Internet search in various databases and an additional reference tracking. Studies had to be on performance of clinical signs in the diagnosis of anaemia, using haemoglobin as the gold standard. We calculated pooled diagnostic likelihood ratios (LR's) and odds ratios (DOR's) for each clinical sign at different haemoglobin thresholds. RESULTS: Eleven articles met the inclusion criteria. Most studies were performed in Africa, in children underfive. Chi-square test for proportions and Cochran Q for DOR's and for LR's showed heterogeneity. Type of observer and haemoglobin technique influenced the results. Pooling was done using the random effects model. Pooled DOR at haemoglobin <11 g/dL was 4.3 (95% CI 2.6–7.2) for palmar pallor, 3.7 (2.3–5.9) for conjunctival pallor, and 3.4 (1.8–6.3) for nailbed pallor. DOR's and LR's were slightly better for nailbed pallor at all other haemoglobin thresholds. The accuracy did not vary substantially after excluding outliers. CONCLUSION: This meta-analysis did not document a highly accurate clinical sign of anaemia. In view of poor performance of clinical signs, universal iron supplementation may be an adequate control strategy in high prevalence areas. Further well-designed studies are needed in settings other than Africa. They should assess inter-observer variation, performance of combined clinical signs, phenotypic differences, and different degrees of anaemia