Fourier transform mid infrared spectroscopy applications for monitoring the structural plasticity of plant cell walls

[EN] Fourier transform mid-infrared (FT-MIR) spectroscopy has been extensively used as a potent, fast and non-destructive procedure for analyzing cell wall architectures, with the capacity to provide abundant information about their polymers, functional groups, and in muro entanglement. In conjunction with multivariate analyses, this method has proved to be a valuable tool for tracking alterations in cell walls. The present review examines recent progress in the use of FT-MIR spectroscopy to monitor cell wall changes occurring in muro as a result of various factors, such as growth and development processes, genetic modifications, exposition or habituation to cellulose biosynthesis inhibitors and responses to other abiotic or biotic stresses, as well as its biotechnological applicationsSIThe authors thank Antonio Encina, Penélope García-Angulo, and María de Castro for their helpful scientific discussion, and to Denise Phelps for the English revision of the manuscrip

Production of Encecalin in Cell Cultures and Hairy Roots of Helianthella quinquenervis (Hook.) A. Gray

[EN] Plant cell and organ cultures of Helianthella quinquenervis, a medicinal plant whose roots are used by the Tarahumara Indians of Chihuahua, Mexico, to relieve several ailments, were established to identify and quantify some chromenes with biological activity, such as encecalin, and to evaluate their potential for biotechnological production. Gas chromatography–mass spectrometry (GC-MS) analysis corroborated the presence of quantifiable amounts of encecalin in H. quinquenervis cell cultures (callus and cell suspensions). In addition, hairy roots were obtained through three transformation protocols (prick, 45-s sonication and co-culture), using wild type Agrobacterium rhizogenes A4. After three months, cocultivation achieved the highest percentage of transformation (66%), and a comparable production (FW) of encecalin (110 g/g) than the sonication assay (120 g/g), both giving far higher yields than the prick assay (19 g/g). Stable integration of rolC and aux1 genes in the transformed roots was confirmed by polymerase chain reaction (PCR). Hairy roots from cocultivation (six months-old) accumulated as much as 1086 g/g (FW) of encecalin, over three times higher than the cell suspension cultures. The production of encecalin varied with growth kinetics, being higher at the stationary phase. This is the first report of encecalin production in hairy roots of H. quinquenervis, demonstrating the potential for a future biotechnological production of chromenes.SIThe initial phase of the conservation project was supported by the International Cooperative Biodiversity Group (NIH UO1 TW00316), CONACYT (U47512-Z) and PAPIIT-UNAM (IN205907-3). This work was partially supported by the Ministerio Español de Economía y Competitividad (AGL2011-30545-C02-02) and by Universidad de León through the financial support to Research Groups, by a fellowship from CONACYT, Mexico, to J.M.H.-A., and by the Unitat de Fisiologia Vegetal, Facultat de Farmàcia, Universitat de Barcelona, Spain

Aplicaciones del cultivo in vitro en la caracterización del efecto del ancimidol en paredes celulares de maíz (Zea mays L.) y en la producción de encecalina en "Helianthella quinquenervis" (Hook.) a. Gray = Applications of in vitro cultures in the characterization of the effects of ancymidol on maize (Zea mays L.) cell walls and in the production of encecalin by Helianthella quinquenervis (Hook.) A. Gray

213 p.El cultivo de células y tejidos de plantas comprende a un conjunto de técnicas que, en condiciones asépticas, permiten la inducción de una variedad de respuestas al controlar las condiciones físicas y químicas de los cultivos. Es una herramienta importante tanto en los estudios básicos como en los aplicados ya que tiene importantes aplicaciones comerciales. En la investigación básica, se ha avanzado en el conocimiento de la bioquímica y fisiología de la pared celular, así como en los mecanismos de la síntesis y degradación de la celulosa, y su relación con otras moléculas en la pared celular. Para profundizar en el conocimiento del metabolismo de la pared celular, se ha hecho uso de inhibidores de la biosíntesis de celulosa (CBIs por sus siglas en inglés), estos son un grupo de moléculas de diversa naturaleza química cuyo sitio de acción es desconocido en la mayoría de los casos; sin embargo, algunos de ellos se usan como herbicidas al bloquear la formación de la pared celular. Recientemente, se ha propuesto al ancimidol (un inhibidor de la enzima ent-kaurenasa en la síntesis de giberelinas) como un probable CBI. Esta propuesta está basada principalmente en la promoción de malformaciones celulares por parte del ancimidol. Con el fin de determinar el papel del ancimidol como inhibidor de la biosíntesis de celulosa, se ha llevado a cabo un estudio completo del efecto del ancimidol solo y en presencia de giberelina tanto a nivel celular (determinando cómo afecta a los parámetros de crecimiento, viabilidad y morfología del cultivo), como a nivel subcelular (analizando los cambios que produce en la composición y estructura de la pared celular) en células en suspensión de maíz, a través de un enfoque bioquímico y molecular por medio de la cuantificación de celulosa, análisis FTIR, incorporación de [14C]Glc y [3H]Ara en las fracciones de pared celular, así como la regulación en la expresión de algunos genes ZmCesA y ZmKO. Otra de las líneas del cultivo in vitro es la obtención de metabolitos secundarios de plantas (MSP) de interés para las industrias agroquímica, farmacéutica y alimentaria. Una de sus ventajas es que a través del cultivo in vitro los recursos pueden ser utilizados de manera sostenible ya que la producción no depende de las condiciones ambientales. En este caso, abordamos la producción de MSP de Helianthella quinquenervis (Hook) A. Gray una planta herbácea perteneciente a la familia Asteraceae, que es conocida por los indios rarámuri (tarahumara) de Chihuahua, México, como «rarésoa» o «corsonero», donde las raíces secas y en polvo se usan como preparados terapéuticos para combatir o aliviar la tos, dolores musculares, enfermedades gastrointestinales y úlceras. El extracto crudo de las raíces de H. quinquenervis ha demostrado actividad antimicrobiana, así como actividad antiprotozoaria. El análisis fitoquímico de las raíces y las partes aéreas de H. quinquenervis ha permitido el aislamiento e identificación de diversos cromenos (benzopiranos), prenil acetofenonas y benzofuranos (euparina). Algunos de estos cromenos, como la encecalina y la desmetilencecalina, poseen una citotoxicidad marginal contra varias líneas celulares tumorales humanas y un efecto inhibidor del crecimiento vegetal. Las fuentes de estos compuestos siguen siendo las plantas silvestres, como en la mayoría de las plantas medicinales, por lo que su explotación pone en riesgo sus poblaciones. La producción de MSP de plantas (MSP) de interés a escala comercial por parte de H. quinquenervis es difícil de alcanzar, porque está sujeta a condiciones ambientales fluctuantes y porque ocurre en concentraciones muy bajas. En este trabajo de doctorado, describimos el uso del cultivo in vitro de plantas en la investigación básica (con cultivos de maíz expuestos al ancimidol para estudiar su efecto en el metabolismo de la pared celular) y en la investigación aplicada, con el desarrollo de diferentes estrategias para la producción de encecalina y otros MSP relacionados con potencial interés comercial por medio del cultivo de células y órganos de Hellianthella quinquenervis (callos, células en suspensión y raíces pilosas o hairy roots). Esta es la primera vez que se han descrito tanto la generación de raíces transformadas por Agrobacterium rhizogenes (A4) en Helianthella quinquenervis, como la cuantificación de la producción de encecalina por CG-MS y MM en cultivos in vitr

Production of Encecalin in Cell Cultures and Hairy Roots of Helianthella quinquenervis (Hook.) A. Gray

Plant cell and organ cultures of Helianthella quinquenervis, a medicinal plant whose roots are used by the Tarahumara Indians of Chihuahua, Mexico, to relieve several ailments, were established to identify and quantify some chromenes with biological activity, such as encecalin, and to evaluate their potential for biotechnological production. Gas chromatography–mass spectrometry (GC-MS) analysis corroborated the presence of quantifiable amounts of encecalin in H. quinquenervis cell cultures (callus and cell suspensions). In addition, hairy roots were obtained through three transformation protocols (prick, 45-s sonication and co-culture), using wild type Agrobacterium rhizogenes A4. After three months, cocultivation achieved the highest percentage of transformation (66%), and a comparable production (FW) of encecalin (110 μg/g) than the sonication assay (120 μg/g), both giving far higher yields than the prick assay (19 μg/g). Stable integration of rolC and aux1 genes in the transformed roots was confirmed by polymerase chain reaction (PCR). Hairy roots from cocultivation (six months-old) accumulated as much as 1086 μg/g (FW) of encecalin, over three times higher than the cell suspension cultures. The production of encecalin varied with growth kinetics, being higher at the stationary phase. This is the first report of encecalin production in hairy roots of H. quinquenervis, demonstrating the potential for a future biotechnological production of chromenes

Frequency and management of maternal infection in health facilities in 52 countries (GLOSS): a 1-week inception cohort study

Background Maternal infections are an important cause of maternal mortality and severe maternal morbidity. We report the main findings of the WHO Global Maternal Sepsis Study, which aimed to assess the frequency of maternal infections in health facilities, according to maternal characteristics and outcomes, and coverage of core practices for early identification and management. Methods We did a facility-based, prospective, 1-week inception cohort study in 713 health facilities providing obstetric, midwifery, or abortion care, or where women could be admitted because of complications of pregnancy, childbirth, post-partum, or post-abortion, in 52 low-income and middle-income countries (LMICs) and high-income countries (HICs). We obtained data from hospital records for all pregnant or recently pregnant women hospitalised with suspected or confirmed infection. We calculated ratios of infection and infection-related severe maternal outcomes (ie, death or near-miss) per 1000 livebirths and the proportion of intrahospital fatalities across country income groups, as well as the distribution of demographic, obstetric, clinical characteristics and outcomes, and coverage of a set of core practices for identification and management across infection severity groups. Findings Between Nov 28, 2017, and Dec 4, 2017, of 2965 women assessed for eligibility, 2850 pregnant or recently pregnant women with suspected or confirmed infection were included. 70·4 (95% CI 67·7–73·1) hospitalised women per 1000 livebirths had a maternal infection, and 10·9 (9·8–12·0) women per 1000 livebirths presented with infection-related (underlying or contributing cause) severe maternal outcomes. Highest ratios were observed in LMICs and the lowest in HICs. The proportion of intrahospital fatalities was 6·8% among women with severe maternal outcomes, with the highest proportion in low-income countries. Infection-related maternal deaths represented more than half of the intrahospital deaths. Around two-thirds (63·9%, n=1821) of the women had a complete set of vital signs recorded, or received antimicrobials the day of suspicion or diagnosis of the infection (70·2%, n=1875), without marked differences across severity groups. Interpretation The frequency of maternal infections requiring management in health facilities is high. Our results suggest that contribution of direct (obstetric) and indirect (non-obstetric) infections to overall maternal deaths is greater than previously thought. Improvement of early identification is urgently needed, as well as prompt management of women with infections in health facilities by implementing effective evidence-based practices