Transcriptomic analysis of the interaction geminivirus-tomato

Geminiviridae family is one of the main families of plant pathogenic viruses with large relevance as they cause great losses worldwide in commercial crops and crops destined to food production. Geminiviruses present a little single-stranded DNA genome and a capsid composed of two twin icosahedral parts. Tomato Yellow Leaf Curl Virus (TYLCV) belongs to the Begomovirus genus and is transmitted by the whitefly Bemisia tabaci. With only 6 viral proteins, this geminivirus must create a proper environment for viral replication, transcription and propagation. Behind the apparent simplicity of geminiviruses lies a complex network of molecular interactions with their host and even their natural vector, which induces a wide variety of transcriptional, post-transcriptional and chromatinic changes in both the plant and the geminivirus. In order to study these changes and decipher the effects of the transmission vector on the infection, we carried out a global approximation of the TYLCV-tomato interaction to generate integrated single-base resolution maps by NGS (next-generation sequencing) of the transcriptome, smallRNAome and methylome of the pathogen and the host. Tomato plants (Moneymaker) were infected with TYLCV under controlled conditions of light and temperature using Agrobacterium tumefaciens or its natural vector. Apical tissue from these plants was collected at different time points (2, 7, 14 and 21 days after inoculation), and three biological replicas were generated for each treatment and time. Total RNA and DNA was extracted and analysed by RNA-Seq, smallRNA-Seq and Bisulfite-Seq. The transcriptome of the tomato-TYLCV interaction will be presented and discussed.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Eye drop Self-medication: Comparative Questionnaire-based study of two Latin American cities.

A broad spectrum of ocular symptoms are treated by self-medication with commercial eye-drops.  This behavior threatens individuals' visual health. In Latin America, evidence is poor. Objective:  To  detect,  characterize  and  compare  patterns  of  ophthalmic  self-medication  between  Córdoba (Argentina) and Barranquilla (Colombia).Design:  Analytic, cross-sectional and comparative population-based study. Setting: Two private tertiary care ophthalmology centers from Córdoba, Argentina, and Barranquilla, Colombia.Participants:  Patients 18 years of age or older who consulted for the first time in this two institutions duringAugust-November 2009, were included. A number of 570 patients were enrrolled.Methods:  Data collected through a semi-structured questionnaire. Main outcome measure: To determine thefrequency of self-medication with eyedrops on a specific population of two cities in Latin America.Results:  Comparable  rates  of  ocular  self-medication  were  found  (25.6%  and  25.7%  for  Cordoba  and Barranquilla, respectively). The percentage of men and women who self-medicated was not significantly different between both samples. The major source of eye drops recommendation in the Argentineans patients was the pharmacist (31%); while the social source was predominant in Colombian individuals (53%). In Cordoba, the most frequently used product was a non-steroidal anti-inflammatory drop in combination with a vasoconstrictive agent (32%); while in Barranquilla, antibiotic eye drops were preferred (33%). Self-medication was higher between the ages of 31 and 50 years old in Argentinean citizens (28%) and between 18 to 31 years old in the Colombiancommunity (39%). This habit was found mostly in patients who completed university studies in Cordoba (33%); in Barranquilla, individuals with lower educational level practice more this behavior (36%).Conclusion:  In both populations, patients commonly treat ocular conditions by self-medicating. Currently, anincreasing number of eye drops are obtainable without prescription and a high percentage of self-medicated patients in both samples ignore the possible side effects of the used medication.Fil: Marquez, Gabriel. Fundación VER; ArgentinaFil: Hildegard Piñeros-Heilbron. Fundación Oftalmológica del Caribe; ColombiaFil: Sanchez, Victoria M.. Fundación VER; ArgentinaFil: Torres, Victor Eduardo Roque. Fundación VER; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones y Estudios sobre Cultura y Sociedad. Universidad Nacional de Córdoba. Centro de Investigaciones y Estudios sobre Cultura y Sociedad; ArgentinaFil: Gramajo, Ana L.. Fundación VER; ArgentinaFil: Juarez, Claudio P.. Fundación VER; Argentina. Fundación Oftalmológica del Caribe; ColombiaFil: Peña, Fernando. Fundación Oftalmológica del Caribe; ColombiaFil: Luna, José D.. Fundación VER; Argentin

Study of the functional domains of the PTGS suppressor V2 from geminivirus Beet curly top virus (BCTV)

Geminiviruses constitute a group of plant viruses that infect vegetable crops all over the world. Among the Geminiviridae family, the genera Mastrevirus, Begomovirus and Curtovirus are the most abundant. Suppression of gene silencing is a key mechanism for viral infection in plants. In begomovirus, V2 is a strong posttranscriptional gene silencing suppressor. We recently showed that V2 from curtovirus Beet curly top virus (BCTV) is a PTGS suppressor by impairing the RDR6/SGS3 pathway, as V2 from begomovirus. In order to identify the domains involved in the suppression activity and viral pathogenicity, we performed an alignment of several begomovirus and curtovirus V2 proteins. A protein kinase C (PKC) phosphorylation motif essential for suppression activity in begomovirus (P1) was found in all analysed sequences. We also found similar hydrophobic profiles, with two hydrophobic domains (H1 and H2) followed by a long hydrophilic domain. Then we generated BCTV V2 mutant proteins and performed transient assays in Nicotiana benthamiana plants to test their suppression activity. We also expressed them from a Potato virus X-derived vector to check the symptoms produced. Additionally, their subcellular localization was determined. Finally, we produced BCTV viruses mutated in the different domains and N. benthamiana plants were infected, analysing virus levels and symptoms produced. The results showed that P1, H1 and H2 are involved in the suppression activity and viral pathogenicity.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Geminivirus replication protein decreases PCNA sumoylation at two acceptor sites

Geminiviruses are plant viruses with circular, single-stranded DNA (ssDNA) genomes that infect a broad range of plants causing substantial crop diseases worldwide. They replicate in nuclei of infected cells by using host DNA replication machinery and an essential protein encoded in their genome designated Rep (replication-associated protein). This multifunctional protein induces the accumulation of the host factors involved in replication and it is capable of interacting with a lot of plant proteins including PCNA (Proliferating Cell Nuclear Antigen), a processivity factor that coordinates a wide range of processes involved in maintenance, duplication and transmission of the genome, and the sumoylation enzyme that conjugates SUMO to target proteins (SUMO-conjugating enzyme- SCE). PCNA modification by SUMO, and also ubiquitin, has long been known to be of key importance for determining how DNA damage is processed by the replisome and for maintenance of overall genome integrity. In yeast, PCNA sumoylation has been associated to DNA repair involving homologous recombination (HR). Previously, we reported that Rep ectopic expression does not result in broad changes in the sumoylation pattern of plant cells, but it modifies the sumoylation state of selected host proteins. In this work, we show, using a reconstituted sumoylation system in Escherichia coli, that tomato PCNA is sumoylated at two residues, K254 and K164, and that co-expression of the Rep protein suppresses PCNA sumoylation at these lysines. Finally, we confirm that PCNA is sumoylated and that Rep also interferes with PCNA sumoylation in planta.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Delayed Corneal Epithelial Healing after Intravitreal Bevacizumab: A Clinical and Experimental Study

Purpose: To report corneal epithelial defects (CEDs) and delayed epithelial healing after intravitreal bevacizumab (IVB) injection and to describe delayed corneal epithelial healing with topical administration of bevacizumab in an experimental rabbit model. Methods: A retrospective chart review was performed on 850 eyes of 850 patients with neovascular eye disease and diabetic macular edema who had received 1.25 to 2.5 mg IVB. In the experimental arm of the study, photorefractive keratectomy was used to create a 3 mm CED in the right eyes of 18 New Zealand rabbits which were then randomized to three equal groups. All rabbits received topical antibiotics, additionally those in group A received topical bevacizumab and animals in group B were treated with topical corticosteroids. The rate of epithelial healing was assessed at different time points using slitlamp photography. Results: In the clinical study, seven eyes of seven subjects developed CEDs the day after IVB injection. All of these eyes had preexisting corneal edema. The healing period ranged from 3 to 38 days (average 11 days) despite appropriate medical management. In the experimental study, topical bevacizumab and corticosteroids both significantly hindered corneal epithelial healing at 12 and 24 hours. Conclusion: Bevacizumab was demonstrated to cause CEDs in clinical settings. Moreover, corneal epithelial healing was delayed by topical application of bevacizumab, in the experimental model. These short-term results suggest that corneal edema may be considered as a risk factor for epithelial defects after IVB

Relation between TNF-alpha and exercise induced bronchoconstriction

Introduction: Exercise-induced bronchoconstriction (EIB) describes an acute airway narrowing that occurs as a result of exercise. EIB is associated with a hyperosmolar environment that lead to airway smooth muscle contraction. The proinflammatory cytokine, TNF-alpha, has a prominent role in the development of airway hiperresponsiveness (AHR). A single bout of heavy exercise can trigger EIB. The aim of this research was to compare the response of TNF-alpha over time with the presence or absence of EIB in marathon runners. Methods: Thirty-eight male runners, who did not have any chronic or acute airway disease, performed maximal cardiopulmonary exercise testing on a treadmill coupled to the gas analyzer (FitMate ™, Cosmed, Italy). Was used a ramp protocol with fixed slope of 1% and an increase of load (1km/h) every minute. Spirometry was performed before (pre), immediately after, five, ten, fifteen and twenty minutes after exercise stress testing. Were considered positive for BIE those athletes that presented a decrease in expiratory volume in one second greater than 10% in any of the spirometric tests. Blood samples were obtained the day before the marathon, immediately and after 72 hours after the race. Cytokines were determined in serum by ELISA. Was used analysis of variance (ANOVA) with two factors, group and time of blood collection, considering repeated measures on the time factor. In the case of statistical significance for interaction group and blood was concluded that the behavior over time was significantly different between EIB negative (-) and EIB positive (+), in this case applied the method of Tukey\u27s multiple comparisons to determine the moment that occurred these differences (p\u3c0.05). Results: We observed a statistically significant difference (p=0.048) in the behavior of the concentration of TNF-alpha between groups EIB- and EIB+. The measure pre of the EIB+ have an higher concentration of TNF-alpha compared to those obtained immediately after the marathon for EIB+ and the results obtained at the three moments for EIB-. (Figure 1). Conclusion: Our results suggests that an inflammatory status in serum of the marathon runners presenting BIE, before the race may be associated with bronchoconstriction triggering

Efficient phosphor-free, white light emission from ordered arrays of GaN/InGaN nanocolumnar LEDs grown by Selective Area MBE

We fabricate and characterize novel LEDs based on InGaN/GaN nanocolumns grown on patterned substrates, leading to the periodically ordered growth of emitters directly producing white ligh

Control of the morphology on selective area growth of GaN nanocolumns by rf-plasma-assisted

Selective area growth (SAG) of GaN nanocolumns (NCs), making use of patterned or masked (nanoholes) substrates, yields a periodic, homogeneous distribution of nanostructures, that makes their processing much easier compared with self-assembled ones. In addition, the control on the diameter and density of NCs avoids dispersion in the electrooptical characteristics of the heterostructures based on this type of material (embedded InGaN/GaN quantum disks for example). Selective area growth using a mask with nanohole arrays has been demonstrated by rf-plasma-assisted MBE [1, 2]

Whole genome, transcriptome, smallRNAome and methylome profiling during tomato-geminivirus interaction

Contribución a congresoTomato Yellow Leaf Curl Virus (TYLCV) belongs to the Begomovirus genus and istransmitted by the whitefly Bemisia tabaci. With only seven viral proteins, TYLCV must create a proper environment for viral replication, transcription, and propagation. Behind the apparent simplicity of geminiviruses lies a complex network of molecular interactions with their host and their natural vector, which induces a wide variety of transcriptional, post-transcriptional and chromatin changes in the host. To understand this virus-host interaction at a genetic and epigenetic level, we carried out a global approach to generate the transcriptome, smallRNAome and methylome of the TYLCV-tomato interaction. Total RNA and DNA was extracted from tomato infected plants (three biological replicates) and analysed at 2, 7, 14 and 21-day postinfection (dpi). Analysis of the changes in host transcription during the infection and its correlation with changes in sRNA profiles (microRNA and phasiRNA) and DNA methylation patterns will be presented and discussed.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech