Surgical resection as a curative intervention for solitary renal cell carcinoma metastasis to the nasal cavity.

Persistent unilateral nasal obstruction with recurrent epistaxis in an adult should raise suspicion of malignancy. Renal cell carcinoma accounts for 90% of all renal malignancies but rarely manifests as a nasal mass. We describe a case of clear cell renal cell carcinoma metastasizing to the nasal cavity.

Effect of heterologous expression of FT gene from Medicago truncatula in growth and flowering behavior of olive plants.

Olive (Olea europaea L. subsp. europaea) is one of the most important crops of the Mediterranean Basin and temperate areas worldwide. Obtaining new olive varieties adapted to climatic changing conditions and to modern agricultural practices, as well as other traits such as biotic and abiotic stress resistance and increased oil quality, is currently required; however, the long juvenile phase, as in most woody plants, is the bottleneck in olive breeding programs. Overexpression of genes encoding the 'florigen' Flowering Locus T (FT), can cause the loss of the juvenile phase in many perennials including olives. In this investigation, further characterization of three transgenic olive lines containing an FT encoding gene from Medicago truncatula, MtFTa1, under the 35S CaMV promoter, was carried out. While all three lines flowered under in vitro conditions, one of the lines stopped flowering after acclimatisation. In soil, all three lines exhibited a modified plant architecture; e.g., a continuous branching behaviour and a dwarfing growth habit. Gene expression and hormone content in shoot tips, containing the meristems from which this phenotype emerged, were examined. Higher levels of OeTFL1, a gene encoding the flowering repressor TERMINAL FLOWER 1, correlated with lack of flowering. The branching phenotype correlated with higher content of salicylic acid, indole-3-acetic acid and isopentenyl adenosine, and lower content of abscisic acid. The results obtained confirm that heterologous expression of MtFTa1 in olive induced continuous flowering independently of environmental factors, but also modified plant architecture. These phenotypical changes could be related to the altered hormonal content in transgenic plants.

Suppressing the rhamnogalacturonan lyase gene FaRGLyase1 preserves RGI pectin degradation and enhances strawberry fruit firmness.

Plant rhamnogalacturonan lyases (RGLyases) cleave the backbone of rhamnogalacturonan I (RGI), the "hairy" pectin and polymer of the disaccharide rhamnose (Rha)-galacturonic acid (GalA) with arabinan, galactan or arabinogalactan side chains. It has been suggested that RGLyases could participate in remodeling cell walls during fruit softening, but clear evidence has not been reported. To investigate the role of RGLyases in strawberry softening, a genome-wide analysis of RGLyase genes in the genus Fragaria was performed. Seventeen genes encoding RGLyases with functional domains were identified in Fragaria × ananassa. FaRGLyase1 was the most expressed in the ripe receptacle of cv. Chandler. Transgenic strawberry plants expressing an RNAi sequence of FaRGLyase1 were obtained. Three transgenic lines yielded ripe fruits firmer than controls without other fruit quality parameters being significantly affected. The highest increase in firmness achieved was close to 32%. Cell walls were isolated from ripe fruits of two selected lines. The amount of water-soluble and chelated pectins was higher in transgenic lines than in the control. A carbohydrate microarray study showed a higher abundance of RGI epitopes in pectin fractions and in the cellulose-enriched fraction obtained from transgenic lines. Sixty-seven genes were differentially expressed in transgenic ripe fruits when compared with controls. These genes were involved in various physiological processes, including cell wall remodeling, ion homeostasis, lipid metabolism, protein degradation, stress response, and defense. The transcriptomic changes observed in FaRGLyase1 plants suggest that senescence was delayed in transgenic fruits.

Embodied Conversational Agents Providing Motivational Interviewing to Improve Health-Related Behaviors: Scoping Review.

BACKGROUND: Embodied conversational agents (ECAs) are advanced human-like interfaces that engage users in natural face-to-face conversations and interactions. These traits position ECAs as innovative tools for delivering interventions for promoting health-related behavior adoption. This includes motivational interviewing (MI), a therapeutic approach that combines brief interventions with motivational techniques to encourage the adoption of healthier behaviors. OBJECTIVE: This study aims to identify the health issues addressed by ECAs delivering MI interventions, explore the key characteristics of these ECAs (eg, appearance, dialogue mechanism, emotional model), analyze the implementation of MI principles and techniques within ECAs, and examine the evaluation methods and primary outcomes of studies that use ECAs providing MI interventions. METHODS: We conducted a scoping review following the PRISMA-ScR (Preferred Reporting Items for Systematic Reviews and Meta-Analyses Extension for Scoping Reviews) methodology. Our systematic search covered the PubMed, Scopus, IEEE Xplore, ACM Digital, and PsycINFO databases for papers published between January 2008 and December 2022. We included papers describing ECAs developed for delivering MI interventions targeting health-related behaviors and excluded articles that did not describe ECAs with human appearances and without the necessary evaluation or MI explanation. In a multistage process, 3 independent reviewers performed screening and data extraction, and the collected data were synthesized using a narrative approach. RESULTS: The initial search identified 404 articles, of which 3.5% (n=14) were included in the review. ECAs primarily focused on reducing alcohol use (n=5, 36%), took on female representations (n=9, 64%), and gave limited consideration to user ethnicity (n=9, 64%). Most of them used rules-driven dialogue mechanisms (n=13, 93%), include emotional behavior to convey empathy (n=8, 57%) but without an automatic recognition of user emotions (n=12, 86%). Regarding MI implementation, of 14 studies, 3 (21%) covered all MI principles, 4 (29%) included all processes, and none covered all techniques. Most studies (8/14, 57%) conducted acceptability, usability, and user experience assessments, whereas a smaller proportion (4/14, 29%) used randomized controlled trials to evaluate behavior changes. Overall, the studies reported positive results regarding acceptability, usability, and user experience and showed promising outcomes in changes in attitudes, beliefs, motivation, and behavior. CONCLUSIONS: This study revealed significant advancements in the use of ECAs for delivering MI interventions aimed at promoting healthier behaviors over the past 15 years. However, this review emphasizes the need for a more in-depth exploration of ECA characteristics. In addition, there is a need for the enhanced integration of MI principles, processes, and techniques into ECAs. Although acceptability and usability have received considerable attention, there is a compelling argument for placing a stronger emphasis on assessing changes in attitudes, beliefs, motivation, and behavior. Consequently, inclusion of more randomized controlled trials is essential for comprehensive intervention evaluations.

Cholesterol goals, statin use and residual cardiovascular risk estimated by SMART score: Study of a Nicaraguan population.

Introduction: Adverse cardiovascular events that arise in patients with established cardiovascular disease have prompted researchers to seek variables that can help estimate residual cardiovascular risk and aid in its reduction. In Latin-America, there is limited data assessing this type of risk. Objective: Estimate residual cardiovascular risk in ambulatory patients diagnosed with Chronic Coronary Syndrome (CCS) using the SMART-Score scale seen at five clinics in Nicaragua; determine the prevalence of patients that achieve a serum LDL level of <55 mg/dL; and describe the use of statins in these patients. Methods: A total of 145 participants previously diagnosed with CCS seen regularly in ambulatory visits were enrolled. A survey was completed, including epidemiological variables that allowed the calculation of a SMART score. Data analysis was conducted using SPSS version 21.0. Results: A 46.2% of participants were male, the average age was 68.7 years (11.4 SD), 91% had hypertension, 80.7% had a BMI ≥25. Under the SMART Score risk classification per Dorresteijn et al. the following risk distribution was found: 2.8% low, 31% moderate, 20% high, 13.1% very high and 33.1% extremely high. Per the risk classification of Kaasenbrood et al., 2.8% were in the 0-9% group, 31% in the 10-19%, 20% in 20-29% and 46.2% were in the ≥30% group. A 64.8% did not meet LDL goals. Conclusion: There is an inadequate control of cLDL levels in patients with CCS, and the appropriate available therapeutic resources aren't being utilized. It is important to achieve a proper control of lipid levels in order to improve cardiovascular outcomes, despite currently being far from these goals.

National survey study of nirmatrelvir-ritonavir prescribing processes and access barriers during the severe acute respiratory coronavirus virus 2 (SARS-CoV-2) omicron surge of the coronavirus disease 2019 (COVID-19) pandemic.

In this national survey, we found that individual patient assessments by pharmacists were more common at facilities using centralized prescribing for nirmetralvir-ritonavir (Paxlovid) than decentralized prescribing. Provider discomfort was initially less with centralized prescribing, but later, there was no difference in provider discomfort based on prescribing mechanism.

CRISPR/Cas9 editing of the polygalacturonase FaPG1 gene improves strawberry fruit firmness.

Firmness is one of the most important fruit quality traits in strawberries. The postharvest shelf life of this soft fruit is highly limited by the loss of firmness, where cell wall disassembly plays an important role. Previous studies demonstrated that the polygalacturonase FaPG1 has a key role in remodelling pectins during strawberry softening. In this study, FaPG1 knockout strawberry plants have been generated using the CRISPR/Cas9 system delivered via Agrobacterium tumefaciens. Ten independent lines, cv. "Chandler", were obtained, and all of them were successfully edited as determined by PCR amplification and T7 endonuclease assay. The targeted mutagenesis insertion and deletion rates were analyzed using targeted deep sequencing. The percentage of edited sequences varied from 47% up to almost 100%, being higher than 95% for seven of the selected lines. Phenotypic analyses showed that 7 out of the eight lines analyzed produced fruits significantly firmer than the control, ranging from 33 to 70% increase in firmness. There was a positive relationship between the degree of FaPG1 editing and the rise in fruit firmness. Minor changes were observed in other fruit quality traits, such as colour, soluble solids, titratable acidity or anthocyanin content. Edited fruits showed a reduced softening rate during postharvest, displayed a reduced transpirational water loss, and were less damaged by Botrytis cinerea inoculation. The analysis of four potential off-target sites revealed no mutation events. In conclusion, editing the FaPG1 gene using the CRISPR/Cas9 system is an efficient method for improving strawberry fruit firmness and shelf life.

FaMYB123 interacts with FabHLH3 to regulate the late steps of anthocyanin and flavonol biosynthesis during ripening.

In this work, we identified and functionally characterized the strawberry (Fragaria × ananassa) R2R3 MYB transcription factor FaMYB123. As in most genes associated with organoleptic properties of ripe fruit, FaMYB123 expression is ripening-related, receptacle-specific, and antagonistically regulated by ABA and auxin. Knockdown of FaMYB123 expression by RNAi in ripe strawberry fruit receptacles downregulated the expression of enzymes involved in the late steps of anthocyanin/flavonoid biosynthesis. Transgenic fruits showed a parallel decrease in the contents of total anthocyanin and flavonoid, especially malonyl derivatives of pelargonidin and cyanidins. The decrease was concomitant with accumulation of proanthocyanin, propelargonidins, and other condensed tannins associated mainly with green receptacles. Potential coregulation between FaMYB123 and FaMYB10, which may act on different sets of genes for the enzymes involved in anthocyanin production, was explored. FaMYB123 and FabHLH3 were found to interact and to be involved in the transcriptional activation of FaMT1, a gene responsible for the malonylation of anthocyanin components during ripening. Taken together, these results demonstrate that FaMYB123 regulates the late steps of the flavonoid pathway in a specific manner. In this study, a new function for an R2R3 MYB transcription factor, regulating the expression of a gene that encodes a malonyltransferase, has been elucidated.

Length of preoperative hospital stay is the dominating risk factor for surgical site infection in neurosurgery: A cohort data-driven analysis.

Background: The number of days of preoperative hospital stay (PHS) is a modifiable variable that has shown contradictory surgical site infection (SSI) risk factor results in neurosurgery. We sought to pinpoint the day of PHS length related with a marked increase of risk of SSI. Methods: From a tertiary teaching hospital, January 2015-December 2017, prospectively collected nonpercutaneous neurosurgery procedures with standard antibiotic prophylaxis and 1-year follow-up were evaluated. SSI risk factors were assessed through multiple logistic regression models with different thresholds of PHS. Results: A total of 1012 procedures were included in the study. Incidence of SSI was 4.4%. The median PHS was higher in those with SSI than in those without (1 day, interquartile range [IQR]: 7 vs. 0 days, IQR: 1, respectively, P = 0.002). By the amount of six days of PHS, this exposure risk past the threshold of significance for impact on wound infection (OR 2.8; CI 1.23-6.39, P = 0.014). Operative time past 4 h (OR 2.11; CI 1.12-3.98; P = 0.021), and in some models, previous surgery at same admission were also identified by multivariate analysis as increasing postoperative SSI risk. Conclusion: The gradual increase of the SSI OR associated with longer PHS days was the highest risk factor of SSI in our cohort of patients. Studies directed to reduce this complication should consider the PHS.

Reconstrucción cervical con conexión vaginal secundaria a agenesia cervical. Reporte de caso / Cervical reconstruction with vaginal conexion secundary to cervical agenesia. Case report

Resumen ANTECEDENTES: La agenesia cervical congénita es una anomalía poco común del sistema de Müller, que generalmente se reconoce durante la menarquia. Hasta hace poco, el tratamiento de primera elección era la histerectomía, debido a que no se consideraba una enfermedad compatible con la fertilidad. Hoy en día, gracias a las técnicas de reproducción asistida, se han desarrollado procedimientos quirúrgicos menos agresivos que sí preservan la fertilidad de las pacientes. OBJETIVO: Revisar un caso de agenesia cervical a partir de su tratamiento quirúrgico y seguimiento de la reconstrucción cervical con la conexión entre la vagina y el útero, revisar la bibliografía y cómo establecer el pronóstico a largo plazo. CASO CLÍNICO: Paciente de 12 años llevada a consulta ante la ausencia de menstruación. El procedimiento diagnóstico transcurrió sin que se encontraran alteraciones; sin embargo, en la laparoscopia diagnóstica operatoria efectuada a los 14 años, indicada por dolor pélvico cíclico, se advirtieron la ausencia del cuello del útero, un hematometra y endometriosis grado IV. En el segundo tiempo quirúrgico, practicado en conjunto con el cirujano pediatra, se practicó, por laparotomía, la comunicación entre el útero y la vagina. En la actualidad, la paciente tiene 19 años y sus menstruaciones son cíclicas, no dolorosas. En el último ultrasonido se visualizó la conexión entre el útero y la vagina. CONCLUSIONES: Hoy en día, la anastomosis útero vaginal es un procedimiento capaz de mantener la función del útero y permitir el embarazo, con los cuidados adecuados.

Abstract BACKGROUND: Congenital cervical agenesis is a rare anomaly of the Müllerian system, usually recognized during menarche. Until recently, the treatment of first choice was hysterectomy, because it was not considered a disease compatible with fertility. Today, thanks to assisted reproductive techniques, less aggressive surgical procedures have been developed that do preserve the fertility of patients. OBJECTIVE: To review a case of cervical agenesis from its surgical treatment and follow-up cervical reconstruction with the connection between the vagina and the uterus, review the literature and how to establish the long-term prognosis. CLINICAL CASE: A 12 years old female patient was brought for consultation due to absence of menstruation. The diagnostic procedure was carried out without finding any alterations; however, in the operative diagnostic laparoscopy performed at the age of 14, indicated by cyclic pelvic pain, the absence of the cervix, a hematometra and endometriosis grade IV were noticed. In the second surgery, performed in conjunction with the pediatric surgeon, the communication between the uterus and the vagina was performed by laparotomy. At present, the patient is 19 years old and her menses are cyclical, not painful. In the last ultrasound the connection between the uterus and the vagina was visualized. CONCLUSIONS: Today, utero-vaginal anastomosis is a procedure capable of maintaining the function of the uterus and allowing pregnancy, with proper care.

Modification of 13-hydroperoxide lyase expression in olive affects plant growth and results in altered volatile profile.

The C6 aldehydes, alcohols, and the corresponding esters are the most important compounds of virgin olive oil aroma. These C6 volatile compounds are synthesized via the 13-hydroperoxide lyase (13-HPL) branch of the lipoxygenase pathway. In this investigation, a functional analysis of the olive (Olea europaea L.) 13-HPL gene by its overexpression and silencing in olive transgenic lines was carried out. With this aim, sense and RNAi constructs of the olive 13-HPL gene were generated and used for the transformation of embryogenic olive cultures. Leaves from overexpressing lines showed a slight increase in 13-HPL gene expression, whereas RNAi lines exhibited a strong decrease in their transcript levels. Quantification of 13-HPL activity in two overexpressing and two RNAi lines showed a positive correlation with levels of transcripts. Interestingly, RNAi lines showed a high decrease in the content of C6 volatiles linked to a strong increase of C5 volatile compounds, altering the volatile profile in the leaves. In addition, the silencing of the 13-HPL gene severely affected plant growth and development. This investigation demonstrates the role of the 13-HPL gene in the biogenesis of olive volatile compounds and constitutes a functional genomics study in olive related to virgin olive oil quality.

Olive (Olea europaea L.) Genetic Transformation: Current Status and Future Prospects.

Olive (Olea europaea L.) is the most characteristic and important oil crop of the Mediterranean region. Traditional olive cultivation is based on few tens cultivars of ancient origin. To improve this crop, novel selections with higher tolerance to biotic and abiotic stress, adaptable to high-density planting systems and resilient to climate change are needed; however, breeding programs are hindered by the long juvenile period of this species and few improved genotypes have been released so far. Genetic transformation could be of great value, in the near future, to develop new varieties or rootstocks in a shorter time; in addition, it has currently become an essential tool for functional genomic studies. The recalcitrance of olive tissues to their in vitro manipulation has been the main bottleneck in the development of genetic transformation procedures in this species; however, some important traits such as fungal resistance, flowering or lipid composition have successfully been manipulated through the genetic transformation of somatic embryos of juvenile or adult origin, providing a proof of the potential role that this technology could have in olive improvement. However, the optimization of these protocols for explants of adult origin is a prerequisite to obtain useful materials for the olive industry. In this review, initially, factors affecting plant regeneration via somatic embryogenesis are discussed. Subsequently, the different transformation approaches explored in olive are reviewed. Finally, transgenic experiments with genes of interest undertaken to manipulate selected traits are discussed.

Elucidating the role of polygalacturonase genes in strawberry fruit softening.

To disentangle the role of polygalacturonase (PG) genes in strawberry softening, the two PG genes most expressed in ripe receptacles, FaPG1 and FaPG2, were down-regulated. Transgenic ripe fruits were firmer than those of the wild type when PG genes were silenced individually. Simultaneous silencing of both PG genes by transgene stacking did not result in an additional increase in firmness. Cell walls from ripe fruits were characterized by a carbohydrate microarray. Higher signals of homogalacturonan and rhamnogalacturonan I pectin epitopes in polysaccharide fractions tightly bound to the cell wall were observed in the transgenic genotypes, suggesting a lower pectin solubilization. At the transcriptomic level, the suppression of FaPG1 or FaPG2 alone induced few transcriptomic changes in the ripe receptacle, but the amount of differentially expressed genes increased notably when both genes were silenced. Many genes encoding cell wall-modifying enzymes were down-regulated. The expression of a putative high affinity potassium transporter was induced in all transgenic genotypes, indicating that cell wall weakening and loss of cell turgor could be linked. These results suggest that, besides the disassembly of pectins tightly linked to the cell wall, PGs could play other roles in strawberry softening, such as the release of oligogalacturonides exerting a positive feedback in softening.

Exploring the Use of Fruit Callus Culture as a Model System to Study Color Development and Cell Wall Remodeling during Strawberry Fruit Ripening.

Cell cultures derived from strawberry fruit at different developmental stages have been obtained to evaluate their potential use to study different aspects of strawberry ripening. Callus from leaf and cortical tissue of unripe-green, white, and mature-red strawberry fruits were induced in a medium supplemented with 11.3 µM 2,4-dichlorophenoxyacetic acid (2,4-D) under darkness. The transfer of the established callus from darkness to light induced the production of anthocyanin. The replacement of 2,4-D by abscisic acid (ABA) noticeably increased anthocyanin accumulation in green-fruit callus. Cell walls were isolated from the different fruit cell lines and from fruit receptacles at equivalent developmental stages and sequentially fractionated to obtain fractions enriched in soluble pectins, ester bound pectins, xyloglucans (XG), and matrix glycans tightly associated with cellulose microfibrils. These fractions were analyzed by cell wall carbohydrate microarrays. In fruit receptacle samples, pectins were abundant in all fractions, including those enriched in matrix glycans. The amount of pectin increased from green to white stage, and later these carbohydrates were solubilized in red fruit. Apparently, XG content was similar in white and red fruit, but the proportion of galactosylated XG increased in red fruit. Cell wall fractions from callus cultures were enriched in extensin and displayed a minor amount of pectins. Stronger signals of extensin Abs were detected in sodium carbonate fraction, suggesting that these proteins could be linked to pectins. Overall, the results obtained suggest that fruit cell lines could be used to analyze hormonal regulation of color development in strawberry but that the cell wall remodeling process associated with fruit softening might be masked by the high presence of extensin in callus cultures.

Heterologous Expression of the AtNPR1 Gene in Olive and Its Effects on Fungal Tolerance.

The NPR1 gene encodes a key component of systemic acquired resistance (SAR) signaling mediated by salicylic acid (SA). Overexpression of NPR1 confers resistance to biotrophic and hemibiotrophic fungi in several plant species. The NPR1 gene has also been shown to be involved in the crosstalk between SAR signaling and the jasmonic acid-ethylene (JA/Et) pathway, which is involved in the response to necrotrophic fungi. The aim of this research was to generate transgenic olive plants expressing the NPR1 gene from Arabidopsis thaliana to evaluate their differential response to the hemibiotrophic fungus Verticillium dahliae and the necrotroph Rosellinia necatrix. Three transgenic lines expressing the AtNPR1 gene under the control of the constitutive promoter CaMV35S were obtained using an embryogenic line derived from a seed of cv. Picual. After maturation and germination of the transgenic somatic embryos, the plants were micropropagated and acclimated to ex vitro conditions. The level of AtNPR1 expression in the transgenic materials varied greatly among the different lines and was higher in the NPR1-780 line. The expression of AtNPR1 did not alter the growth of transgenic plants either in vitro or in the greenhouse. Different levels of transgene expression also did not affect basal endochitinase activity in the leaves, which was similar to that of control plants. Response to the hemibiotrophic pathogen V. dahliae varied with pathotype. All plants died by 50 days after inoculation with defoliating (D) pathotype V-138, but the response to non-defoliating (ND) strains differed by race: following inoculation with the V-1242 strain (ND, race 2), symptoms appeared after 44-55 days, with line NPR1-780 showing the lowest disease severity index. This line also showed good performance when inoculated with the V-1558 strain (ND, race 1), although the differences from the control were not statistically significant. In response to the necrotroph R. necatrix, all the transgenic lines showed a slight delay in disease development, with mean area under the disease progress curve (AUDPC) values 7-15% lower than that of the control.

Plant Regeneration via Somatic Embryogenesis in Mature Wild Olive Genotypes Resistant to the Defoliating Pathotype of Verticillium dahliae.

Regeneration capacity, via somatic embryogenesis, of four wild olive genotypes differing in their response to defoliating Verticillium dahliae (resistant genotypes StopVert, OutVert, Ac-18 and the susceptible one, Ac-15) has been evaluated. To induce somatic embryogenesis, methodologies previously used in wild or cultivated olive were used. Results revealed the importance of genotype, explant type, and hormonal balance in the induction process. Use of apical buds obtained from micropropagated shoots following a methodology used in cultivated olive (4 days induction in liquid 1/2 MS medium supplemented with 30 µM TDZ-0.54 µM NAA, followed by 8 weeks in basal 1/2 MS medium) was adequate to obtain somatic embryos in two genotypes, StopVert and Ac-18, with a 5.0 and 2.5% induction rates, respectively; however, no embryogenic response was observed in the other two genotypes. Embryogenic cultures were transferred to basal ECO medium supplemented with 0.5 µM 2iP, 0.44 µM BA, and 0.25 µM indole-3-butyric acid (IBA) for further proliferation. Somatic embryos from StopVert were maturated and germinated achieving a 35.4% conversion rate. An analysis of genetic stability on StopVert, using Simple Sequence Repeats (SSRs) and Random Amplified Polymorphic DNA (RAPDs) markers, was carried out in embryogenic callus, plants regenerated from this callus and two controls, micropropagated shoots used as explant source, and the original mother plant. Polymorphism was only observed in the banding pattern generated by RAPDs in 1 of the 10 callus samples evaluated, resulting in a variation rate of 0.07%. This is the first time in which plants have been regenerated via somatic embryogenesis in wild olive.

An atypical HLH transcriptional regulator plays a novel and important role in strawberry ripened receptacle.

BACKGROUND: In soft fruits, the differential expression of many genes during development and ripening is responsible for changing their organoleptic properties. In strawberry fruit, although some genes involved in the metabolic regulation of the ripening process have been functionally characterized, some of the most studied genes correspond to transcription factors. High throughput transcriptomics analyses performed in strawberry red receptacle (Fragaria x ananassa) allowed us to identify a ripening-related gene that codes an atypical HLH (FaPRE1) with high sequence homology with the PACLOBUTRAZOL RESISTANCE (PRE) genes. PRE genes are atypical bHLH proteins characterized by the lack of a DNA-binding domain and whose function has been linked to the regulation of cell elongation processes. RESULTS: FaPRE1 sequence analysis indicates that this gene belongs to the subfamily of atypical bHLHs that also includes ILI-1 from rice, SlPRE2 from tomato and AtPRE1 from Arabidopsis, which are involved in transcriptional regulatory processes as repressors, through the blockage by heterodimerization of bHLH transcription factors. FaPRE1 presented a transcriptional model characteristic of a ripening-related gene with receptacle-specific expression, being repressed by auxins and activated by abscisic acid (ABA). However, its expression was not affected by gibberellic acid (GA3). On the other hand, the transitory silencing of FaPRE1 transcription by agroinfiltration in receptacle produced the down-regulation of a group of genes related to the ripening process while inducing the transcription of genes involved in receptacle growth and development. CONCLUSIONS: In summary, this work presents for the first time experimental data that support an important novel function for the atypical HLH FaPRE1 during the strawberry fruit ripening. We hypothesize that FaPRE1 modulates antagonistically the transcription of genes related to both receptacle growth and ripening. Thus, FaPRE1 would repress the expression of receptacle growth promoting genes in the ripened receptacle, while it would activate the expression of those genes related to the receptacle ripening process.

The Strawberry FaWRKY1 Transcription Factor Negatively Regulates Resistance to Colletotrichum acutatum in Fruit Upon Infection.

Strawberry (Fragaria ×ananassa) is a major food crop worldwide, due to the flavor, aroma and health benefits of the fruit, but its productivity and quality are seriously limited by a large variety of phytopathogens, including Colletotrichum spp. So far, key factors regulating strawberry immune response remain unknown. The FaWRKY1 gene has been previously proposed as an important element mediating defense responses in strawberry to Colletotrichum acutatum. To get further insight into the functional role that FaWRKY1 plays in the defense mechanism, Agrobacterium-mediated transient transformation was used both to silence and overexpress the FaWRKY1 gene in strawberry fruits (Fragaria ×ananassa cv. Primoris), which were later analyzed upon C. acutatum inoculation. Susceptibility tests were performed after pathogen infection comparing the severity of disease between the two agroinfiltrated opposite halves of the same fruit, one half bearing a construct either for FaWRKY1 overexpression or RNAi-mediated silencing and the other half bearing the empty vector, as control. The severity of tissue damage was monitored and found to be visibly reduced at five days after pathogen inoculation in the fruit half where FaWRKY1 was transiently silenced compared to that of the opposite control half and statistical analysis corroborated a significant reduction in disease susceptibility. Contrarily, a similar level of susceptibility was found when FaWRKY1 overexpression and control fruit samples, was compared. These results unravel a negative regulatory role of FaWRKY1 in resistance to the phytopathogenic fungus C. acutatum in strawberry fruit and contrast with the previous role described for this gene in Arabidopsis as positive regulator of resistance against the bacteria Pseudomonas syringae. Based on previous results, a tentative working model for WRKY75 like genes after pathogen infection is proposed and the expression pattern of potential downstream FaWRKY1 target genes was also analyzed in strawberry fruit upon C. acutatum infection. Our results highlight that FaWRKY1 might display different function according to species, plant tissue and/or type of pathogen and underline the intricate FaWRKY1 responsive defense regulatory mechanism taking place in strawberry against this important crop pathogen.