Nutritional Performance of Five Citrus Rootstocks under Different Fe Levels.

Iron is an essential micronutrient for citrus, playing an important role in photosynthesis and yield. The aim of this paper was to evaluate the tolerance to Fe deficiency of five citrus rootstocks: sour orange (S), Carrizo citrange (C), Citrus macrophylla (M), Troyer citrange (T), and Volkamer lemon (V). Plants were grown for 5 weeks in nutrient solution that contained the following Fe concentrations (in µM): 0, 5, 10, 15, and 20. At the end of the experiment, biomass (dry weight-DW), leaf area, total leaf chlorophyll (CHL), and the activity of root chelate reductase (FCR) were recorded. Additionally, the mineral composition of roots (R) and shoots (S) was evaluated. Principal component analysis was used to study the relationships between all parameters and, subsequently, the relations between rootstocks. In the first component, N-S, P-S, Ca-S, Cu-S, Zn-S, Mn-S, Zn-R, and Mn-R concentrations were related to leaf CHL and FCR. Increases in leaf CHL, Mg-R, and DW (shoots and roots) were inversely related to Cu-R, which was shown in the second component. The values obtained were consistent for V10, C15, and C20, but in contrast for S0 and S5. In conclusion, micronutrient homeostasis in roots and shoots of all rootstocks were affected by Fe stress conditions. The Fe/Cu ratio was significantly related to CHL, which may be used to assist rootstock performance.

Effects of foliar application of organic acids on strawberry plants.

The large economic costs and environmental impacts of iron-chelate treatments has led to the search for alternative methods and compounds to control iron (Fe) deficiency chlorosis. Strawberry plants (Fragaria x ananassa) were grown in Hoagland's nutrient solution in a greenhouse with two levels of Fe: 0 and 10 µM Fe(III)-EDDHA. After 20 days, plants growing without Fe showed typical symptoms of Fe deficiency chlorosis in young leaves. Then, the adaxial and abaxial sides of one mature or one young leaf in each plant were brushed with 10 mM malic (MA), citric (CA) or succinic (SA) acids. Eight applications were done over a two-week period. At the end of the experiment, the newly emerged (therefore untreated), young and mature leaves were sampled for nutritional and metabolomic analysis, to assess the effectiveness of treatments. Leaf regreening was monitored using a SPAD-502 apparatus, and the activity of the ferric chelate-reductase activity (FCR) was measured using root tips. Iron deficiency negatively affected biomass and leaf chlorophyll but did not increase FCR activity. Application of succinic acid alleviated the decrease in chlorophyll observed in other treatments, and the overall nutritional balance in the plant was also changed. The concentrations of two quinic acid derivatives increased under Fe deficiency and decreased in plants treated with succinic acid, and thus they are proposed as Fe stress markers. Data suggest that foliage treatments with carboxylates may be, in some cases, environmentally friendly alternatives to Fe(III)-chelates. The importance of Fe mobilization pathways in the formulation of new fertilizers is also discussed.

Silencing of the FRO1 gene and its effects on iron partition in Nicotiana benthamiana.

To evaluate the dynamic role of the ferric-chelate reductase enzyme (FCR) and to identify possible pathways of regulation of its activity in different plant organs an investigation was conducted by virus-induced gene silencing (VIGS) using tobacco rattle virus (TRV) to silence the ferric reductase oxidase gene (FRO1) that encodes the FCR enzyme. Half of Nicotiana benthamiana plants received the VIGS vector and the rest remained as control. Four treatments were imposed: two levels of Fe in the nutrient solution (0 or 2.5 µM of Fe), each one with silenced or non-silenced (VIGS-0; VIGS-2.5) plants. Plants grown without iron (0; VIGS-0) developed typical symptoms of iron deficiency in the youngest leaves. To prove that FRO1 silencing had occurred, resupply of Fe (R) was done by adding 2.5 µM of Fe to the nutrient solution in a subset of chlorotic plants (0-R; VIGS-R). Twelve days after resupply, 0-R plants had recovered from Fe deficiency while plants containing the VIGS vector (VIGS-R) remained chlorotic and both FRO1 gene expression and FCR activity were considerably reduced, consequently preventing Fe uptake. With the VIGS technique we were able to silence the FRO1 gene in N. benthamiana and point out its importance in chlorophyll synthesis and Fe partition.

Cirugía tiroidea en programas de cooperación en África subsahariana / Thyroid surgery at a volunteer program in Sub-Saharan Africa

INTRODUCCIÓN: Nuestro objetivo es describir la experiencia adquirida en un programa de cooperación dedicado a la cirugía tiroidea en Camerún. Su interés radica en la imposibilidad para seguir las guías internacionales de tratamiento en áreas de recursos imitados debido a la falta de estudios preoperatorios y a la dificultad para conseguir medicación hormonal sustitutiva de por vida. MÉTODOS: Estudio observacional de una cohorte de 16 pacientes operados de enfermedad tiroidea en Dschang (Camerún) en junio de 2015. La técnica quirúrgica empleada fue la hemitiroidectomía con istmectomía en la enfermedad unilateral y la tiroidectomía subtotal bilateral en la enfermedad bilateral. RESULTADOS: Todos los pacientes eran de raza negra, 15 mujeres y un hombre, con edad media de 41 años. Se realizaron 5tiroidectomías subtotales, 9hemitiroidectomías y 2istmectomías. Cuatro pacientes presentaban componente intratorácico. Fue necesaria la sección de músculos pretiroideos en un caso. Se visualizaron las glándulas paratiroides y los nervios laríngeos recurrentes en el 86 y el 84% de los casos, respectivamente. Se dejaron drenajes en 7 casos y la mediana de duración de la intervención fue de 75 min. Como complicaciones cabe destacar un hematoma cervical que precisó reintervención y 2infecciones de herida quirúrgica. No hubo hipocalcemias clínicas ni lesiones recurrenciales apreciables. La estancia media fue de 2,3 días. A largo plazo, los pacientes con tiroidectomías bilaterales presentaban niveles elevados de TSH. CONCLUSIONES: La cirugía tiroidea en países subdesarrollados, adaptando los protocolos y técnicas que utilizamos en nuestro medio (evitando la tiroidectomía total), tiene una tasa de complicaciones asumible. No deben realizarse tiroidectomías bilaterales salvo que se disponga de estudios funcionales y se asegure previamente la disponibilidad de la hormona tiroidea

INTRODUCTION: The aim of this study is to demonstrate our experience at a volunteer surgical program in Cameroon, which is of special interest given to the inability to adopt international treatment guidelines for thyroid surgery in areas of limited resources due to the lack of preoperative testing and to the difficulty to obtain sustitutive hormonal treatment. METHODS: This is a prospective observational study that includes 16 cases of thyroid surgery in Dschang (Cameroon) during June 2015. The patients were previously selected by a local medical team. All patients were black, 15 women and one man, with a mean age of 41 years. The surgical technique used for the removal of unilateral disease was hemithyroidectomy with isthmectomy and bilateral subtotal thyroidectomy for bilateral disease. RESULTS: Five subtotal thyroidectomies, 9hemithyroidectomies and 2isthmectomies were performed. Prethyroid muscles were divided only in one case. We visualized 86% of the parathyroid glands and 84% of the recurrent laryngeal nerves. The main complications observed were one symptomatic cervical haematoma that required reoperation and 2surgical wound infections. There were no clinical episodes of hypocalemia or recurrent nerve lesion. The mean length of stay was 2.3 days. At follow-up, all bilateral thyroidectomies developed high TSH levels. CONCLUSIONS: Thyroid surgery is safe in developing countries adopting protocols and techniques we use in our environment (avoiding total thyroidectomy). Bilateral thyroidectomies should not be performed unless functional studies are available in the follow-up and a thyroid hormone supplement stock guaranteed whenever necessary

Thyroid surgery at a volunteer program in Sub-Saharan Africa. / Cirugía tiroidea en programas de cooperación en África subsahariana.

INTRODUCTION: The aim of this study is to demonstrate our experience at a volunteer surgical program in Cameroon, which is of special interest given to the inability to adopt international treatment guidelines for thyroid surgery in areas of limited resources due to the lack of preoperative testing and to the difficulty to obtain sustitutive hormonal treatment. METHODS: This is a prospective observational study that includes 16 cases of thyroid surgery in Dschang (Cameroon) during June 2015. The patients were previously selected by a local medical team. All patients were black, 15 women and one man, with a mean age of 41 years. The surgical technique used for the removal of unilateral disease was hemithyroidectomy with isthmectomy and bilateral subtotal thyroidectomy for bilateral disease. RESULTS: Five subtotal thyroidectomies, 9hemithyroidectomies and 2isthmectomies were performed. Prethyroid muscles were divided only in one case. We visualized 86% of the parathyroid glands and 84% of the recurrent laryngeal nerves. The main complications observed were one symptomatic cervical haematoma that required reoperation and 2surgical wound infections. There were no clinical episodes of hypocalemia or recurrent nerve lesion. The mean length of stay was 2.3 days. At follow-up, all bilateral thyroidectomies developed high TSH levels. CONCLUSIONS: Thyroid surgery is safe in developing countries adopting protocols and techniques we use in our environment (avoiding total thyroidectomy). Bilateral thyroidectomies should not be performed unless functional studies are available in the follow-up and a thyroid hormone supplement stock guaranteed whenever necessary.

The memory of iron stress in strawberry plants.

To provide information towards optimization of strategies to treat Fe deficiency, experiments were conducted to study the responses of Fe-deficient plants to the resupply of Fe. Strawberry (Fragaria × ananassa Duch.) was used as model plant. Bare-root transplants of strawberry (cv. 'Diamante') were grown for 42 days in Hoagland's nutrient solutions without Fe (Fe0) and containing 10 µM of Fe as Fe-EDDHA (control, Fe10). For plants under Fe0 the total chlorophyll concentration of young leaves decreased progressively on time, showing the typical symptoms of iron chlorosis. After 35 days the Fe concentration was 6% of that observed for plants growing under Fe10. Half of plants growing under Fe0 were then Fe-resupplied by adding 10 µM of Fe to the Fe0 nutrient solution (FeR). Full Chlorophyll recovery of young leaves took place within 12 days. Root ferric chelate-reductase activity (FCR) and succinic and citric acid concentrations increased in FeR plants. Fe partition revealed that FeR plants expressively accumulated this nutrient in the crown and flowers. This observation can be due to a passive deactivation mechanism of the FCR activity, associated with continuous synthesis of succinic and citric acids at root level, and consequent greater uptake of Fe.

Changes in the concentration of organic acids in roots and leaves of carob-tree under Fe deficiency.

Several fruit trees are able to cope with iron (Fe) deficiency when grown in calcareous soils in the Mediterranean region, although information regarding well adapted slow-growing species is scarce, and the mechanisms activated by these species are not described in the literature. A crucial issue related to tolerance is the need to transport Fe over relatively long distances inside the plant. To evaluate the possible role of organic acids in the movement of Fe in tolerant plants, we studied the concentration of low molecular weight organic acids in several organs of 1-year old carob plants grown for 55 days in nutrient solutions without Fe (0µM Fe) or with 1µM Fe and 10µM Fe. Roots, stems and leaves were harvested, and the biomass, Fe and organic acid contents quantified. Total leaf chlorophyll (Chl) was evaluated in young leaves over the experimental period and the activity of root ferric chelate-reductase (FC-R; EC 1.16.1.17) was determined after 35 days, when deficiency symptoms appeared. Iron chlorosis was observed only at the end of the experiment in plants grown in the absence of Fe, and these plants had a smaller DW of leaves and also significant greater activity of root FC-R. Iron deficiency (Fe0 and Fe1 treatments) induced significant changes in the concentrations of succinic, malic, citric and fumaric acids, which increased in roots, or in basal, middle and apical leaves. There were significant correlations between most organic acids (with the exceptions of 2-oxoglutaric and tartaric acids) and leaf Chl. Analysis of each type of leaf showed that more succinic and malic acids were present in young chlorotic leaves while the reverse was true for quinic acid. These changes in organic acids followed a root-to-foliage pathway that was similar in all leaf types and particularly evident in young chlorotic leaves. We hypothesised that it was associated with Fe transport from roots to aboveground tissues, as there were significant differences in Fe contents between treatments with and without Fe.

Development and recovery of iron deficiency by iron resupply to roots or leaves of strawberry plants.

Bare-root transplants of strawberry (Fragaria ananassa Duch. cv. 'Selva') were transferred to nutrient solutions with or without iron (Fe). After six weeks of growth, plants grown in solution lacking Fe were chlorotic and showed morphological changes in roots typical of Fe deficiency. Subsequently, four treatments were applied for nine days: plants grown in continued absence of Fe (Fe0); plants grown in continued presence of 10 µM Fe (Fe10); foliar application of ferrous sulphate every two days to chlorotic plants (Fe-leaves); and growth of chlorotic plants in solution with ferrous sulphate (Fe-solution). After six days, the chlorophyll (Chl) content in leaves of Fe-solution plants was similar to that in Fe10 plants. Under the Fe-leaves treatment, a slight regreening of new leaves was observed only by the end of the experiment. After nine days, ferric chelate reductase (FC-R) activity was unchanged in Fe10 but increased in Fe0 plants. The FC-R activity of Fe-solution plants was similar to the initial value for chlorotic plants, whereas it was reduced drastically under the Fe-leaves treatment. The Fe concentration in leaves of Fe0 and Fe10 was similar, whereas the Fe-solution and Fe-leaves treatments enhanced leaf Fe concentration. In contrast to the Fe-solution treatment, foliar application of Fe did not increase the Fe concentration in roots. Under our experimental conditions, FC-R activity in strawberry appeared to be deactivated rapidly by pulses of Fe applied by foliar sprays. Deactivation was slower if Fe was applied directly to roots, which suggested that the plants had greater opportunity to take Fe.

Control of volume-sensitive chloride channel inactivation by the coupled action of intracellular chloride and extracellular protons.

The volume-sensitive chloride current (I(ClVol)) exhibit a time-dependent decay presumably due to channel inactivation. In this work, we studied the effects of chloride ions (Cl(-)) and H(+) ions on I(ClVol) decay recorded in HEK-293 and HL-60 cells using the whole-cell patch clamp technique. Under control conditions ([Cl(-)](e) = [Cl(-)](i) = 140 mM and pH(i) = pH(e) = 7.3), I(ClVol) in HEK cells shows a large decay at positive voltages but in HL-60 cells I(ClVol) remained constant independently of time. In HEK-293 cells, simultaneously raising the [Cl(-)](e) and [Cl(-)](i) from 25 to 140 mM (with pH(e) = pH(i) = 7.3) increased the fraction of inactivated channels (FIC). This effect was reproduced by elevating [Cl(-)](i) while keeping the [Cl(-)](e) constant. Furthermore, a decrease in pH(e) from 7.3 to 5.5 accelerated current decay and increased FIC when [Cl(-)] was 140 mM but not 25 mM. In HL-60 cells, a slight I(ClVol) decay was seen when the pH(e) was reduced from 7.3 to 5.5. Our data show that inactivation of I(ClVol) can be controlled by changing either the Cl(-) or H(+) concentration or both. Based on our results and previously published data, we have built a model that explains VRAC inactivation. In the model the H(+) binding site is located outside the electrical field near the extracellular entry whilst the Cl(-) binding site is intracellular. The model depicts inactivation as a pore constriction that happens by simultaneous binding of H(+) and Cl(-) ions to the channel followed by a voltage-dependent conformational change that ultimately causes inactivation.

The MAP kinase TVK1 regulates conidiation, hydrophobicity and the expression of genes encoding cell wall proteins in the fungus Trichoderma virens.

Mitogen-activated protein (MAP) kinases modulate morphological and genetic processes, which determine cell fate. The tvk1 gene encodes a MAP kinase of Trichoderma virens and its deletion promotes an unusual conidiation phenotype in submerged culture. Here, it is reported that the morphology, physiology and expression of genes encoding cell wall proteins from Trichoderma are significantly affected by Tvk1. Morphological changes were evident in the cell walls of aerial conidia produced by a MAPK null mutant when compared to those produced by the wild-type. Unexpectedly, conidia produced in submerged culture by the Deltatvk1 strain were highly hydrophobic, whereas in aerial conidia hydrophobicity was severely reduced. In addition, the Deltatvk1 strain was unable to break the liquid-air interface when the fungus grew in rich medium; however, when it grew in minimal medium the fungus produced large filaments which were much more efficient at breaking the interface than the wild-type. Through cDNA subtractive hybridization between the wild-type and Deltatvk1 grown in submerged culture, five genes encoding hydrophobin-like proteins and two additional genes encoding cell wall proteins were identified. Four hydrophobin-encoding genes (Tv-hfb1, Tv-srh1, tv-cfth1 and Tv-qid3) and a gene encoding a clock-controlled-like protein (Tv-ccg14/TvSm1) were upregulated in Deltatvk1, whereas genes encoding a cell wall protein (tv-qid74) and an additional hydrophobin (tv-hfb3) were absent in the mutant strain. Clear differences in gene expression were shown during conidiation and emergence from the liquid-air interface, suggesting different functions of the corresponding proteins in these two phenomena. The results support a model in which Tvk1 regulates morphology and genes encoding cell wall proteins during development of Trichoderma.

[Intrathoracic gossypiboma interpreted as bronchogenic carcinoma. Another false positive with positron emission tomography]. / Textiloma intratorácico interpretado como carcinoma broncogénico. Otro falso positivo de la tomografía por emisión de positrones.

Gossypibomas from inflammatory reactions to textile foreign bodies are a rare postoperative complication and are easily confused with neoplastic processes because of their diversity of symptoms and radiographic signs. Positron emission tomography (PET) is seldom used to diagnose gossypibomas and PET findings can result in false positives for a diagnosis of neoplastic disease. We describe the case of a 56-year-old man in whom PET findings showed an intrathoracic mass suggesting a tumor. The final diagnosis was gossypiboma, identified 23 years after pneumothorax surgery.

Textiloma intratorácico interpretado como carcinoma broncogénico. Otro falso positivo de la tomografía por emisión de positrones / Intrathoracic gossypiboma interpreted as bronchogenic carcinoma: another false positive with positron emission tomography

Los textilomas, reacciones inflamatorias contra cuerpos extraños de tipo textil, representan una complicación posquirúrgica poco frecuente. La diversidad de síntomas y signos radiológicos con que se manifiestan contribuye a que su diagnóstico se confunda con un proceso de tipo neoplásico. Los hallazgos descritos con la tomografía por emisión de positrones (PET), técnica poco habitual en el manejo diagnóstico de esta entidad, pueden condicionar falsos positivos neoplásicos. Describimos el caso de un varón de 56 años, que 23 años después de una cirugía de neumotórax presentó una masa intratorácica indicativa de neoplasia en la PET, cuyo diagnóstico final fue de textiloma

Gossypibomas from inflammatory reactions to textile foreign bodies are a rare postoperative complication and are easily confused with neoplastic processes because of their diversity of symptoms and radiographic signs. Positron emission tomography (PET) is seldom used to diagnose gossypibomas and PET findings can result in false positives for a diagnosis of neoplastic disease. We describe the case of a 56-year-old man in whom PET findings showed an intrathoracic mass suggesting a tumor. The final diagnosis was gossypiboma, identified 23 years after pneumothorax surgery

Cross talk between a fungal blue-light perception system and the cyclic AMP signaling pathway.

Blue light regulates many physiological and developmental processes in fungi. In Trichoderma atroviride the complex formed by the BLR-1 and BLR-2 proteins appears to play an essential role as a sensor and transcriptional regulator in photoconidiation. Here we demonstrate that the BLR proteins are necessary for carbon deprivation induced conidiation, even in the absence of light, pointing to the existence of an unprecedented cross talk between light and carbon sensing. Further, in contrast to what has been found in all other fungal systems, clear BLR-independent blue-light responses, including the activation of protein kinase A (PKA) and the regulation of gene expression, were found. Expression of an antisense version of the pkr-1 gene, encoding the regulatory subunit of PKA, resulted in a nonsporulating phenotype, whereas overexpression of the gene produced colonies that conidiate even in the dark. In addition, overexpression of pkr-1 blocked the induction of early light response genes. Thus, our data demonstrate that PKA plays an important role in the regulation of light responses in Trichoderma. Together, these observations suggest that the BLR complex plays a general role in sensing environmental cues that trigger conidiation and that such a role can be separated from its function as a transcription factor.

Three decades of fungal transformation: novel technologies.

Fungi are lower eukaryotes that play important roles in many human activities, including biotechnological processes, phytopathology, and biomedical research. In addition, they are excellent models for molecular and genetic studies. An important key in the advancement of genetics and molecular biology of a given organism is the development of genetic transformation systems. This technology makes possible the analysis and manipulation of the genome of the organism of interest. Thirty years from the first report of transformation of a fungus, transformation of many other fungi has been achieved. However, the development of gene tagging systems generally applicable to a wide range of filamentous fungi has remained elusive. A widely used gene tagging strategy for filamentous fungi is restriction enzyme mediated integration (REMI). In recent years numerous reports have been published describing the effective application of REMI. However, REMI shows certain disadvantages for some fungi. Recently a very promising alternative strategy has been reported based on the use of the soil bacterium Agrobacterium tumefaciens. Using this system a well-defined DNA segment (T-DNA) is transferred, which integrates by illegitimate recombination and is 100-1000 times more efficient than conventional methods. The T-DNA can be used as an efficient tool to generate recombinant strains where DNA is integrated as a single copy, allowing the generation of collections of gene-tagged mutants of the fungus of interest.