NAC61 regulates late- and post-ripening osmotic, oxidative, and biotic stress responses in grapevine

During late- and post-ripening stages, grape berry undergoes profound biochemical and physiological changes whose molecular control is poorly understood. Here, we report the role of NAC61, a grapevine NAC transcription factor, in regulating different processes involved in berry ripening progression. NAC61 is highly expressed during post-harvest berry dehydration and its expression pattern is closely related to sugar concentration. The ectopic expression of NAC61 in Nicotiana benthamiana leaves resulted in low stomatal conductance, high leaf temperature, tissue collapse and a higher relative water content. Transcriptome analysis of grapevine leaves transiently overexpressing NAC61 and DNA affinity purification and sequencing analyses allowed us to narrow down a list of NAC61-regulated genes. Direct regulation of the stilbene synthase regulator MYB14, the osmotic stress-related gene DHN1b, the Botrytis cinerea susceptibility gene WRKY52, and NAC61 itself was validated. We also demonstrate that NAC61 interacts with NAC60, a proposed master regulator of grapevine organ maturation, in the activation of MYB14 and NAC61 expression. Overall, our findings establish NAC61 as a key player in a regulatory network that governs stilbenoid metabolism and osmotic, oxidative, and biotic stress responses that are the hallmark of late- and post-ripening grape stages.NAC61 regulates stilbene biosynthesis and abiotic/biotic stress responses that hallmark late- and post-ripening developmental stages in grapes. NAC61 self-activates and synergistically interacts with the master ripening regulator NAC60

Acute Lower Limb Ischaemia as a Presenting Sign of Atrial Myxoma: Case Report and Scoping Review of the Literature

Objective: Cardiac myxomas (CMs) are the most common primary cardiac tumour in adults. They are a rare cause of peripheral embolisation and may present as acute lower limb ischaemia (ALI). A scoping review was undertaken and a case of ALI due to CM embolisation is presented in this paper. Methods: MEDLINE, Scopus, and Embase were systematically searched for studies reporting data on ALI as a presentation of CM embolisation. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses Extension for Scoping Reviews (PRISMA-ScR) was followed. Results: A healthy 26 year old female presented to the emergency department with bilateral ALI. Urgent bilateral aorto-iliac embolectomy and distal embolectomy of the left femoropopliteal axis were performed. The retrieved embolic material exhibited a yellowish appearance and jelly like consistency, and histological analysis provided a diagnosis of a myxomatous embolus. Transoesophageal echocardiography confirmed the left atrial origin of a myxomatous tumour, but the residual mass was considered too small for further excision. At a two year clinical follow up, the patient was alive and well without recurrence. Between 1989 and 2023, 59 patients with ALI due to CM embolisation were identified in the literature. An in hospital mortality rate of 12.1% (n = 7) was reported, while the in hospital complication and re-intervention rates were 34.5% (n = 20) and 27.6% (n = 16), respectively. No post-discharge deaths, complications, or re-interventions were reported; fasciotomies were the most reported (n = 10). Post-discharge follow up was reported in 22 (37.3%) patients. Mean follow up was 18.0 ± 18.8 months (range 1–120), and 86.4% of patients (n = 19) were alive and well at last follow up. Conclusion: This review and the associated case report underline that CM embolisation should be considered in healthy young patients presenting with cryptogenic ALI. Early transoesophageal echocardiography and histological analysis of the retrieved embolus are recommended to minimise misdiagnosis in these populations

The transcription factor VviNAC60 regulates senescence- and ripening-related processes in grapevine

: Grapevine (Vitis vinifera L.) is one of the most widely cultivated fruit crops because the winemaking industry has huge economic relevance worldwide. Uncovering the molecular mechanisms controlling the developmental progression of plant organs will prove essential for maintaining high-quality grapes, expressly in the context of climate change, which impairs the ripening process. Through a deep inspection of transcriptomic data, we identified VviNAC60, a member of the NAC transcription factor family, as a putative regulator of grapevine organ maturation. We explored VviNAC60 binding landscapes through DNA affinity purification followed by sequencing and compared bound genes with transcriptomics datasets from grapevine plants stably and transiently overexpressing VviNAC60 to define a set of high-confidence targets. Among these, we identified key molecular markers associated with organ senescence and fruit ripening. Physiological, metabolic, and promoter activation analyses showed that VviNAC60 induces chlorophyll degradation and anthocyanin accumulation through the up-regulation of STAY-GREEN PROTEIN 1 (VviSGR1) and VviMYBA1, respectively, with the latter being up-regulated through a VviNAC60-VviNAC03 regulatory complex. Despite sharing a closer phylogenetic relationship with senescence-related homologues to the NAC transcription factor AtNAP, VviNAC60 complemented the non-ripening(nor) mutant phenotype in tomato (Solanum lycopersicum), suggesting a dual role as an orchestrator of both ripening- and senescence-related processes. Our data support VviNAC60 as a regulator of processes initiated in the grapevine vegetative- to mature-phase organ transition and therefore as a potential target for enhancing the environmental resilience of grapevine by fine-tuning the duration of the vegetative phase

BIOCHEMICAL AND FUNCTIONAL CHARACTERIZATION OF THE BACTERIAL PsaR1 SENSOR IN THE PSEUDOMONAS SYRINGAE PV. ACTINIDIAE-KIWIFRUIT INTERACTION

Kiwifruit bacterial canker, caused by Pseudomonas syringae pv. actinidiae (Psa), is responsible for important economic losses in all major areas of kiwifruit production worldwide, including Italy. As for many other bacterial diseases, current plant defense strategies against Psa are mainly based on the use of copper-containing products, which raise eco-toxicological problems. Revision and restriction processes regarding the use of high amounts of copper in agriculture impose an urgent study of new solutions, efficient and eco-compatible, avoiding at the same time the occurrence of new resistance to active molecules. Innovative strategies are based for instance on the application of targeted treatments for \u201cweakening\u201d the pathogen, i.e. to reduce its virulence within its host. However, this requires improving our knowledge regarding molecular mechanisms controlling bacterial virulence induction. A key regulator of bacterial virulence is the so-called \u2018quorum-sensing\u2019 (QS), that links bacterial density to gene expression. This mechanism allows bacteria to communicate within the bacterial community and with their environment, via small diffusible molecules. The prototypical QS system of Gram-negative bacteria consists of a LuxI-type synthase that produces the signal molecules acyl homoserine lactones (AHLs) and a cognate LuxR-type receptor/regulator that senses signal specific threshold concentration. An interesting subgroup of LuxR receptors lacks a genetically linked LuxI and has been termed \u201csolos\u201d. These \u201csolos\u201d are assumed to sense AHLs from neighboring bacteria, bacterial molecules other than AHLs or still unknown plant-produced compounds in the case of phytopathogenic bacteria. Interestingly, Psa does not produce AHLs but possesses three LuxR solos, which likely contribute to Psa virulence. As a first candidate for a targeted inhibition strategy against Psa, we are currently investigating the biochemical properties of the sensor PsaR1. To that purpose, several tentative have been made to obtain the soluble recombinant sensor in a heterologous system. Once achieved, we demonstrated that it does not bind AHLs, thus excluding the possibility to sense AHLs from neighboring bacteria, and we are currently setting a chemical screening, based on thermal shift assay, to identify the class(es) of molecules able to bind to the sensor. On the other hand, we aim to identify the pathway(s) regulated by PsaR1 during Psa interaction with kiwifruit, during different phases of the infection. Thus, a microarray analysis is being performed to compare the transcriptomic profiles of wild-type and psaR1 knockout Psa strains at both exponential and stationary growth phase, in conditions mimicking the interaction with the host plant, i.e. minimal medium supplemented with kiwifruit extract

Exploring the participation of VviNAC factors in the transcriptional regulatory network which governs grapevine maturation processes

Grapevine (Vitis vinifera L.) is one of the most important fruit crops as it is widely cultivated, and the winemaking industry has a huge worldwide economic relevance. The global warming has affected viticulture altering the maturation processes; in particular, the anticipation of the onset of the berries ripening (veraison) has changed the physiological characteristics of grapes and, consequently, has negatively influenced the wine quality. In this contest, uncovering the molecular mechanisms of the ripening could provide the key in maintaining high quality grapes and wine. For all these reasons, the identification and characterization of master regulators controlling the transition from vegetative-to-mature growth are the challenging but fundamental tasks of this research project. At first, to provide insight into the transcriptional programs controlling the development of grapevine, a global gene expression atlas was generated (Fasoli et al., 2012). Combining this dataset with a berry-specific one (Massonnet et al., 2017), an integrated network analysis was performed (Palumbo et al., 2014) and a new category of genes (switch genes), which are significantly up regulated during the developmental shift and inversely correlated with many genes suppressed during the mature growth phase, was identified. Moreover, many transcription factors are present among them, strongly indicating that they could represent master regulators of the developmental phase transition; between them, the plant-specific NAC (NAM/ATAF/CUC) transcription factors represent an interesting family due to their key role in plant development processes and stress responses (Jensen et al., 2014). Fourteen VviNACs genes were selected for functional characterization as key candidates of the major transcriptome reprogramming during grapevine development: VviNAC01, VviNAC03 (D’Incà, 2017), VviNAC08, VviNAC11 (D’Incà, 2017), VviNAC13 (D’Incà, 2017), VviNAC15, VviNAC17, VviNAC18, VviNAC26, VviNAC33 (D’Incà, 2017), VviNAC38, VviNAC39, VviNAC60 (D’Incà, 2017) and VviNAC61. Different approaches have been carried out to investigate the function of VviNACs gene: the transient over expressions, DAP-seq (DNA Affinity Purification and sequencing, Bartlett et al., 2017) and, only for VviNAC60, the ChIP-seq (Chromatin Immunoprecipitation sequencing, Kaufmann et al., 2010). All of them are powerful techniques used to identify possible targets of the selected transcription factor regulations. VviNAC01 showed its important role in the ethylene pathways; VviNAC03 did not reveal a well-defined identity but seems to be plant growth related; VviNAC08 seems to have a possible role in the gibberellin-related and circadian mechanisms; VviNAC11 seems to be related to the control of the auxin pathways and the chlorophylls degradation; VviNAC13 revealed a probable action in the lignin and phenylpropanoid metabolic processes; VviNAC15, regulating many other TFs, highlighted its role in the regulation mechanisms orchestration; VviNAC17 appeared to be a regulator of the jasmonic acid-induced gene expression; VviNAC18 analyses reported its role in the chlorophyll degradation; VviNAC26 presented the regulation of many genes related to the sugars biosynthesis and the anthocyanin synthesis; VviNAC33 terminates the photosynthetic activity and organ vegetative growth; no data are available for VviNAC38; VviNAC39 resulted to up regulate the transport of sugar and lipids and the ubiquitin-conjugating; VviNAC60 revealed a lot of hormones related up regulated direct target genes and many transcription factors, highlighting again the important and major role of this transcription factor in the grapevine maturation processes; VviNAC61 revealed a predominant role in the regulation of the aromatic compounds biosynthesis. Concerning VviNAC60, ChIP-seq data were also obtained and one interesting gene, the SRG1- SENESCENCE-RELATED GENE 1 OXIDOREDUCTASE (VIT_10s0003g02400), was found. In order to define a regulation and co-regulation network between VviNACs, some candidate targets genes taken from the different DAP-seq datasets (VviNAC01, VviNAC05, VviNAC08, VviNAC34, VviNAC37 and VviNAC61) were tested by Dual Luciferase Reporter Assays to see by which of the selected TFs were actually regulated. The obtained results showed that VviNAC01 directly repressed VviNAC05 expression, whereas activated the VviNAC08 one; moreover, VviNAC01 was validated as repressor of its own transcription. VviNAC03 resulted a repressor of VviNAC05. VviNAC11 directly up regulated VviNAC34, VviNAC37 and VviNAC61. VviNAC13 resulted to regulate VviNAC34 and VviNAC37 expression. VviNAC15 activated VviNAC34. VviNAC17 acted as a repressor of VviNAC05, VviNAC08 and VviNAC61 expression. VviNAC18 resulted a direct activator of VviNAC05. VviNAC26 positively regulated the expression of VviNAC05 and directly down regulated the VviNAC08, VviNAC34 and VviNAC61 expression. VviNAC33 was found as a direct activator of VviNAC05, VviNAC08, VviNAC34, VviNAC37 and VviNAC61 expression. VviNAC60 showed to induced VviNAC05, VviNAC34 and VviNAC61 expression. This PhD thesis lights up the possible roles of some VviNACs in the grapevine development and presents a preliminary regulatory network between this family members; further analysis must be conducted to completely elucidate this complex regulation system

Gait Alteration in Individual with Limb Loss: The Role of Inertial Sensors

: Amputation has a big impact on the functioning of patients, with negative effects on locomotion and dexterity. In this context, inertial measurement units represent a useful tool in clinical practice for motion analysis, and in the development of personalized aids to improve a patient's function. To date, there is still a gap of knowledge in the scientific literature on the application of inertial sensors in amputee patients. Thus, the aim of this narrative review was to collect the current knowledge on this topic and stimulate the publication of further research. Pubmed, Embase, Scopus, and Cochrane Library publications were screened until November 2022 to identify eligible studies. Out of 444 results, we selected 26 articles focused on movement analysis, risk of falls, energy expenditure, and the development of sensor-integrated prostheses. The results showed that the use of inertial sensors has the potential to improve the quality of life of patients with prostheses, increasing patient safety through the detection of gait alteration; enhancing the socio-occupational reintegration through the development of highly technologic and personalized prosthesis; and by monitoring the patients during daily life to plan a tailored rehabilitation program

Immersive Virtual Reality in Post-Stroke Rehabilitation: A Systematic Review

In recent years, next to conventional rehabilitation’s techniques, new technologies have been applied in stroke rehabilitation. In this context, fully immersive virtual reality (FIVR) has showed interesting results thanks to the level of immersion of the subject in the illusional world, with the feeling of being a real part of the virtual environment. This study aims to investigate the efficacy of FIVR in stroke rehabilitation. PubMed, Web of Science and Scopus were screened up to November 2022 to identify eligible randomized controlled trials (RCTs). Out of 4623, we included 12 RCTs involving post-acute and chronic stroke survivors, with a total of 350 patients (234 men and 115 women; mean age 58.36 years). High heterogeneity of the outcomes considered, the results showed that FIVR provides additional benefits, in comparison with standard rehabilitation. In particular, results showed an improvement in upper limb dexterity, gait performance and dynamic balance, influencing patient independence. Therefore, FIVR represents an adaptable, multi-faceted rehabilitation tool that can be considered in post-stroke rehabilitation, improving the compliance of the patients to the treatment and increasing the level of functioning and quality of life of stroke survivors

The geek and the chemist: Antioxidant capacity measurements by DPPH assay in beverages using open source tools, consumer electronics and 3D printing

Microcontrollers and single-board computers are widespread tools for innovative educational labs, for prototyping and for accomplish everyday tasks by expert users. Moreover, these modules are opening new exciting possibilities in the area of biological and chemical assays. In this study a Raspberry Pi computer assembled with 3D printed parts and inexpensive opto-electronic components were employed to analyse the antioxidant capacity of several bottled tea performing diphenylpicryl-hydrazyl (DPPH) tests. A dedicated python software allowed the execution of the analysis controlling the device through a small LCD touch screen or remotely through secure connections with other devices. The rRaspberry Pi-based measurements were compared with a research-grade spectrophotometer showing excellent correlation (R2 = 0.9996) and no significant differences (p < 0.05) in the range of measured values. We strongly believe that this approach could support diagnostics progress in resource-poor countries and open new opportunities in research and education