Characterization of Transcriptome Dynamics during Early Fruit Development in Olive (Olea europaea L.).

In the olive (Olea europaea L.), an economically leading oil crop worldwide, fruit size and yield are determined by the early stages of fruit development. However, few detailed analyses of this stage of fruit development are available. This study offers an extensive characterization of the various processes involved in early olive fruit growth (cell division, cell cycle regulation, and cell expansion). For this, cytological, hormonal, and transcriptional changes characterizing the phases of early fruit development were analyzed in olive fruit of the cv. 'Picual'. First, the surface area and mitotic activity (by flow cytometry) of fruit cells were investigated during early olive fruit development, from 0 to 42 days post-anthesis (DPA). The results demonstrate that the cell division phase extends up to 21 DPA, during which the maximal proportion of 4C cells in olive fruits was reached at 14 DPA, indicating that intensive cell division was activated in olive fruits at that time. Subsequently, fruit cell expansion lasted as long as 3 weeks more before endocarp lignification. Finally, the molecular mechanisms controlling the early fruit development were investigated by analyzing the transcriptome of olive flowers at anthesis (fruit set) as well as olive fruits at 14 DPA (cell division phase) and at 28 DPA (cell expansion phase). Sequential induction of the cell cycle regulating genes is associated with the upregulation of genes involved in cell wall remodeling and ion fluxes, and with a shift in plant hormone metabolism and signaling genes during early olive fruit development. This occurs together with transcriptional activity of subtilisin-like protease proteins together with transcription factors potentially involved in early fruit growth signaling. This gene expression profile, together with hormonal regulators, offers new insights for understanding the processes that regulate cell division and expansion, and ultimately fruit yield and olive size.

Disentangling the link between leaf photosynthesis and turgor in fruit growth.

Despite the importance of understanding plant growth, the mechanisms underlying how plant and fruit growth declines during drought remain poorly understood. Specifically, it remains unresolved whether carbon or water factors are responsible for limiting growth as drought progresses. We examine questions regarding the relative importance of water and carbon to fruit growth depending on the water deficit level and the fruit growth stage by measuring fruit diameter, leaf photosynthesis, and a proxy of cell turgor in olive (Olea europaea). Flow cytometry was also applied to determine the fruit cell division stage. We found that photosynthesis and turgor were related to fruit growth; specifically, the relative importance of photosynthesis was higher during periods of more intense cell division, while turgor had higher relative importance in periods where cell division comes close to ceasing and fruit growth is dependent mainly on cell expansion. This pattern was found regardless of the water deficit level, although turgor and growth ceased at more similar values of leaf water potential than photosynthesis. Cell division occurred even when fruit growth seemed to stop under water deficit conditions, which likely helped fruits to grow disproportionately when trees were hydrated again, compensating for periods with low turgor. As a result, the final fruit size was not severely penalized. We conclude that carbon and water processes are able to explain fruit growth, with importance placed on the combination of cell division and expansion. However, the major limitation to growth is turgor, which adds evidence to the sink limitation hypothesis.

The Effects of Microaggressions on Depression in Young Adults of Color: Investigating the Impact of Traumatic Event Exposures and Trauma Reactions.

Microaggressions are a common way that individuals experience racism in the United States. The current study examined the extent to which microaggressions contribute to mental health difficulties, namely trauma reactions and depression, after controlling for other traumatic event exposures. We sought to address gaps in the literature by quantitatively assessing the associations among microaggressions, posttraumatic stress symptoms, and depression symptoms. Participants were 140 young adults of color (68.8% female) who were recruited online. Linear regression analyses evidenced that microaggressions were uniquely associated with depression symptoms, B = 0.27, after controlling for lifetime traumatic event exposures, with this association partially mediated by trauma reactions, B = 0.49. These results suggest that microaggressions are a clinically relevant factor in understanding mental health problems reported by Black, Indigenous, and People of Color in the United States and warrant analysis, assessment, and intervention through a trauma lens.

Cell Wall Composition and Ultrastructural Immunolocalization of Pectin and Arabinogalactan Protein during Olea europaea L. Fruit Abscission.

Cell wall modification is integral to many plant developmental processes where cells need to separate, such as abscission. However, changes in cell wall composition during natural fruit abscission are poorly understood. In olive (Olea europaea L.), some cultivars such as 'Picual' undergo massive natural fruit abscission after fruit ripening. This study investigates the differences in cell wall polysaccharide composition and the localization of pectins and arabinogalactan protein (AGP) in the abscission zone (AZ) during cell separation to understand fruit abscission control in 'Picual' olive. To this end, immunogold labeling employing a suite of monoclonal antibodies to cell wall components (JIM13, LM5, LM6, LM19 and LM20) was investigated in olive fruit AZ. Cell wall polysaccharide extraction revealed that the AZ cell separation is related to the de-esterification and degradation of pectic polysaccharides. Moreover, ultrastructural localization showed that both esterified and unesterified homogalacturonans (HGs) localize mainly in the AZ cell walls, including the middle lamella and tricellular junction zones. Our results indicate that unesterified HGs are likely to contribute to cell separation in the olive fruit AZ. Similarly, immunogold labeling demonstrated a decrease in both galactose-rich and arabinose-rich pectins in AZ cell walls during ripe fruit abscission. In addition, AGPs were localized in the cell wall, plasma membrane and cytoplasm of AZ cells with lower levels of AGPs during ripe fruit abscission. This detailed temporal profile of the cell wall polysaccharide composition, and the pectins and AGP immunolocalization in the olive fruit AZ, offers new insights into cell wall remodeling during ripe fruit abscission.

Spatio-temporal immunolocalization of extensin protein and hemicellulose polysaccharides during olive fruit abscission.

MAIN CONCLUSION: Immunocytochemical and molecular analyses reveal that the disassembly of the cell wall may be mediated by changes in the level and subcellular location of extensin protein and hemicelluloses during olive-fruit abscission. Although cell-wall modification is believed to underlie the changes in organ abscission, information concerning the changes in cell-wall proteins and hemicellulose polysaccharides is still limited. The aim of this work was to analyze the spatio-temporal patterns of the distribution of different extensin proteins and hemicelluloses in the abscission zone (AZ) during natural ripe-fruit abscission in olive (Olea europaea L.). In this study, we employed immunogold labeling in the ripe-fruit AZ during olive AZ cell separation, using an expanded set of monoclonal antibodies that recognize different types of hemicelluloses (LM11, LM15, and LM21), callose (anti-(1,3)-ß-D-glucan) and extensin (JIM19) epitopes, and transmission electron microscopy imaging. Our data demonstrate that AZ cell separation was accompanied by a loss of the JIM19 extensin epitopes and a reduction in the detection of the LM15 xyloglucan epitopes in AZ cell walls, whereas AZ cells were found to be enriched with respect to the xylan and callose levels of the cell wall during olive ripe-fruit abscission. By contrast, AZ cell-wall polysaccharide remodeling did not involve mannans. Moreover, in ripe-fruit AZ, quantitative RT-PCR analysis revealed that OeEXT1, OeEXT2, OeXTH9, and OeXTH13 genes were downregulated during abscission, whereas the expression of OeXTH1, OeXTH5, and OeXTH14 genes increased during abscission. Taken together, the results indicate that AZ cell-wall dynamics during olive ripe-fruit abscission involves extensin protein and hemicellulose modifications, as well as related expressed genes. This is the first study available demonstrating temporal degradation of extensin protein and hemicelluloses in the AZ at the subcellular level.

Transcriptome and Hormone Analyses Revealed Insights into Hormonal and Vesicle Trafficking Regulation among Olea europaea Fruit Tissues in Late Development.

Fruit ripening and abscission are the results of the cell wall modification concerning different components of the signaling network. However, molecular-genetic information on the cross-talk between ripe fruit and their abscission zone (AZ) remains limited. In this study, we investigated transcriptional and hormonal changes in olive (Olea europaea L. cv Picual) pericarp and AZ tissues of fruit at the last stage of ripening, when fruit abscission occurs, to establish distinct tissue-specific expression patterns related to cell-wall modification, plant-hormone, and vesicle trafficking in combination with data on hormonal content. In this case, transcriptome profiling reveals that gene encoding members of the α-galactosidase and ß-hexosaminidase families associated with up-regulation of RabB, RabD, and RabH classes of Rab-GTPases were exclusively transcribed in ripe fruit enriched in ABA, whereas genes of the arabinogalactan protein, laccase, lyase, endo-ß-mannanase, ramnose synthase, and xyloglucan endotransglucosylase/hydrolase families associated with up-regulation of RabC, RabE, and RabG classes of Rab-GTPases were exclusively transcribed in AZ-enriched mainly in JA, which provide the first insights into the functional divergences among these protein families. The enrichment of these protein families in different tissues in combination with data on transcript abundance offer a tenable set of key genes of the regulatory network between olive fruit tissues in late development.

Exogenous Salicylic Acid Improves Phenolic Content and Antioxidant Activity in Table Grapes.

Grapes contain high contents of phenolics, which are known to possess health promoting properties. Exogenous application of phytoregulators, mainly methyl jasmonate and abscisic acid, to grapevines to enhance phenolic content has been reported (Portu et al. Sci Hortic 240: 378-386, 2018; Ranjbaran et al. J Faculty Agric Kyushu Univ 56: 263-267, 2011). However, these phytohormones possess some drawbacks that can be overcome by using other phytoregulators as an alternative. In this work the effect of an additional phytohormone, salicylic acid, to grapevines on the phenolics and antioxidant activity of grapes was investigated. To our knowledge, salicylic acid has been earlier applied to grapevines to affect grape ripening and quality (Lóay. Egyptian J Basic Applied Sci 4: 227-230, 2017). However, this is the first time it is applied to increase the total phenolic content. As a result of our study, total phenol content and the free radical scavenging activity increased with 100 mg l-1 of salicylic acid. In particular, the total phenol content increased from 768.3 to 1843.5 mg 100 g-1 and the IC50 values decreased from 45.2 to 13.2 mg ml-1. Also the contents of individual phenolics mostly increased significantly with 100 mg l-1 of salicylic acid, except anthocyanins. Higher concentrations of salicylic acid (ie, 500 mg l-1vs 100 mg l-1) did not result in higher contents of phenolics. Therefore, 100 mg l-1 was selected as the best salicylic acid concentration to be used in the treatment. The application of exogenous salicylic acid to grapevines is an interesting agronomic practice to obtain table grapes with improved health-promoting properties.

Regulation of sterol content and biosynthetic gene expression during flower opening and early fruit development in olive.

Phytosterols are lipophilic membrane components essential not only for diverse cellular functions but also are biosynthetic precursors of the plant hormone, brassinosteroid (BR). However, the interaction between phytosterol and BR during early fleshy-fruit growth remains largely uncharacterized. In olive, phytosterols are important lipids because they affect oil quality, but phytosterol composition during flowering and early fruit development has not been explored. Here, we first investigated the temporal changes in phytosterol composition, and biosynthetic gene expression that occurred during olive flower opening and early fruit growth. Next, we analyzed the interrelationship between phytosterol and BR, whose levels we manipulated through the application of exogenous BRs (24-epibrassinolide, EBR) or a BR biosynthesis inhibitor (brassinazole, Brz). In this report, the profiling of phytosterol measurement revealed that ß-sitosterol is the most abundant in olive reproductive organs. Our data demonstrate that both OeCYP51 and OeSMT2 genes are upregulated during floral anthesis in good agreement with the rise in cholesterol and ß-sitosterol contents in olive flower. By contrast, the OeCYP51 and OeSMT2 genes displayed different expression patterns during early olive-fruit development. Furthermore, our data show that exogenous EBR enhanced the early olive-fruit growth, as well as the OeSMT2 transcript and ß-sitosterol levels, but decreased the OeCYP51 transcript, squalene, campesterol and cholesterol levels, whereas the Brz treatment exerted the opposite effect. Overall, our findings indicate an up-regulation of ß-sitosterol biosynthesis by BR at the transcriptional level during early olive-fruit growth, providing a valuable tool to unravel the physiological function of SMT2 in future studies.

How cognitive performance changes according to the ankle-brachial index score in an elderly cohort? Results from the Toledo Study of Healthy Ageing.

In the ageing process, the vascular system undergoes morphological and functional changes that may condition brain functioning; for this reason, the aims of this study were to assess the effect of vascular function indirectly measured by ankle-brachial index (ABI) on both cognitive performance at baseline and change in cognitive performance at end of follow-up. We developed a prospective, population-based, cohort study with 1147 participants aged > 65 years obtained from the Toledo Study for Healthy Ageing who had cognitive assessment and measured ABI in the first wave (2006-2009) were selected for the cross-sectional analysis. Those participants who also performed the cognitive assessment in the second wave (2011-2013) were selected for the prospective analysis. Cognitive impairment diagnosis and symptoms and/or history of cardio/neurovascular disease were used as exclusion criteria. Multivariate segmented regression model was used to assess the associations between ABI and cognitive performance in both the cross-sectional and prospective analyses. As ABI score decreased from 1.4, the cross-sectional analysis showed a higher decrease in cognitive performance and the prospective analysis showed a higher degree of worsening in cognitive performance. Our findings suggest that the ABI, a widespread measure of vascular health in primary care, may be a useful tool for predicting cognitive performance and its evolution.

Carotid-femoral pulse wave velocity score, an estimator of cognitive performance in the elderly: results from the Toledo Study for Healthy Aging.

Ageing-related changes in the vascular wall influence the function of different organs; for this reason, we assessed how arterial stiffness measured by carotid-femoral pulse wave velocity (cf-PWV) modulates: the basal cognitive performance and the change in cognitive performance over the follow-up time. We developed a prospective, population-based cohort study with 1581 participants aged > 65 years were obtained from the Toledo Study for Healthy Aging. Participants from the second wave (2011-2013) were selected for the cross-sectional analysis. Those who also performed the cognitive assessment in the third wave (2015-2017) were selected for the prospective analysis. Arterial stiffness was evaluated by cf-PWV. Multivariate segmented regression models were used to evaluate the association between cf-PWV scores and basal neuropsychological evaluation scores and change of neuropsychological evaluation scores along follow-up. Cross-sectional analysis showed that as cf-PWV grew within the cf-PWV (5- < 10) category an improvement was observed in 7-min test, free short-term memory, and hole peg test. Furthermore, in the cf-PWV (> 13-18) category a decrease was observed in total short-term memory, free long-term memory, and total long-term memory. Prospective analysis showed a progressive worsening of cognitive function as cf-PWV increases within the cf-PWV (> 13-18) category in 7-min test, object denomination, immediate and short-term memory, and hole peg test, while in the cf-PWV (5- < 10) category, there was observed a decrease in Cumulative Executive Dysfunction Index score and short-term memory. In conclusion, a higher cf-PWV score is associated with worse cognitive performance, and with a worse evolution, reinforcing the need to plan interventions to delay arterial stiffness and its consequences.

Variations in Fruit Ploidy Level and Cell Size between Small- and Large-Fruited Olive Cultivars during Fruit Ontogeny.

Olive (Olea europaea L.) is one of the major oil fruit tree crops worldwide. However, the mechanisms underlying olive fruit growth remain poorly understood. Here, we examine questions regarding the interaction of endoreduplication, cell division, and cell expansion with olive fruit growth in relation to the final fruit size by measuring fruit diameter, pericarp thickness, cell area, and ploidy level during fruit ontogeny in three olive cultivars with different fruit sizes. The results demonstrate that differences in the fruit size are related to the maximum growth rate between olive cultivars during early fruit growth, about 50 days post-anthesis (DPA). Differences in fruit weight between olive cultivars were found from 35 DPA, while the distinctive fruit shape became detectable from 21 DPA, even though the increase in pericarp thickness became detectable from 7 DPA in the three cultivars. During early fruit growth, intense mitotic activity appeared during the first 21 DPA in the fruit, whereas the highest cell expansion rates occurred from 28 to 42 DPA during this phase, suggesting that olive fruit cell number is determined from 28 DPA in the three cultivars. Moreover, olive fruit of the large-fruited cultivars was enlarged due to relatively higher cell division and expansion rates compared with the small-fruited cultivar. The ploidy level of olive fruit pericarp between early and late growth was different, but similar among olive cultivars, revealing that ploidy levels are not associated with cell size, in terms of different 8C levels during olive fruit growth. In the three olive cultivars, the maximum endoreduplication level (8C) occurred just before strong cell expansion during early fruit growth in fruit pericarp, whereas the cell expansion during late fruit growth occurred without preceding endoreduplication. We conclude that the basis for fruit size differences between olive cultivars is determined mainly by different cell division and expansion rates during the early fruit growth phase. These data provide new findings on the contribution of fruit ploidy and cell size to fruit size in olive and ultimately on the control of olive fruit development.

Comparative transcriptional profiling analysis of olive ripe-fruit pericarp and abscission zone tissues shows expression differences and distinct patterns of transcriptional regulation.

BACKGROUND: In fleshy fruit, abscission of fully ripe fruit is a process intimately linked to the ripening process. In many fruit-tree species, such as olive (Olea europaea L. cv. Picual), there is a coupling of the full ripening and the activation of the abscission-zone (AZ). Although fully ripe fruit have marked physiological differences with respect to their AZs, dissimilarities in gene expression have not been thoroughly investigated. The present study examines the transcriptome of olive fruit and their AZ tissues at the last stage of ripening, monitored using mRNA-Seq. RESULTS: Roche-454 massive parallel pyrosequencing enabled us to generate 397,457 high-quality EST sequences, among which 199,075 were from ripe-fruit pericarp and 198,382 from AZ tissues. We assembled these sequences into 19,062 contigs, grouped as 17,048 isotigs. Using the read amounts for each annotated isotig (from a total of 15,671), we identified 7,756 transcripts. A comparative analysis of the transcription profiles conducted in ripe-fruit pericarp and AZ evidenced that 4,391 genes were differentially expressed genes (DEGs) in fruit and AZ. Functional categorization of the DEGs revealed that AZ tissue has an apparently higher response to external stimuli than does that of ripe fruit, revealing a higher expression of auxin-signaling genes, as well as lignin catabolic and biosynthetic pathway, aromatic amino acid biosynthetic pathway, isoprenoid biosynthetic pathway, protein amino acid dephosphorylation, amino acid transport, and photosynthesis. By contrast, fruit-enriched transcripts are involved in ATP synthesis coupled proton transport, glycolysis, and cell-wall organization. Furthermore, over 150 transcripts encoding putative transcription-factors (TFs) were identified (37 fruit TFs and 113 AZ TFs), of which we randomly selected eight genes and we confirmed their expression patterns using quantitative RT-PCR. CONCLUSION: We generated a set of EST sequences from olive fruit at full ripening, and DEGs between two different olive tissues, ripe fruit and their AZ, were also identified. Regarding the cross-talk between fruit and AZ, using qRT-PCR, we confirmed a set of TF genes that were differentially expressed, revealing profiles of expression that have not previously been reported, this offering a promising beginning for studies on the different transcription regulation in such tissues.

Transcriptome analysis of mature fruit abscission control in olive.

Mature fruit abscission (MFA) is a genetically controlled process, through poorly characterized mechanisms in fleshy fruit that include extensive transcriptional changes. While global transcriptome analyses have been used to investigate immature fruit abscission in fleshy fruit, no global gene expression changes specific to MFA have been described. Here we use pyrosequencing to characterize the transcriptomes of the olive abscission zone (AZ) during cell separation in order to understand MFA control at the stage of AZ activation. Analysis of gene expression from these AZs reveals that membrane microdomains involving sterols/sphingolipids and remorins together with signaling proteins are potentially involved in MFA. This is accompanied by gene activity related to sphingolipid turnover, suggesting potentially the involvement of long-chain base metabolism in regulating MFA. Activation of vesicle trafficking involving small GTPases is probably required for cell wall modifications during abscission. Analysis of transcription factors indicates that most members of the MYB and bZIP families are abundantly represented in the fruit AZ, and is consistent with a model by which most of the key transcription factors during abscission may regulate downstream processes mostly related to ABA. The data provide the first thorough analysis available for a comprehensive picture of the array of cellular responses controlled by gene expression that lead to MFA in fleshy fruit.

Inflorescence Emergence and Flowering Response of Olive Cultivars Grown in Olive Reference Collection of Portugal (ORCP).

In olive trees, fluctuations in the onset of phenological stages have been reported due to weather conditions. The present study analyses the reproductive phenology of 17 olive cultivars grown in Elvas (Portugal) in 3 consecutive years (2012-2014). Through 2017-2022, the phenological observations continued with four cultivars. The phenological observations followed the BBCH scale. Over the course of the observations, the bud burst (stage 51) occurred gradually later; a few cultivars did not follow this trend in 2013. The flower cluster totally expanded phase (stage 55) was achieved gradually earlier, and the period between stages 51-55 was shortened, especially in 2014. Date of bud burst showed a negative correlation with minimum temperature (Tmin) of November-December, and, in 'Arbequina' and 'Cobrançosa', the interval stage 51-55 showed a negative correlation with both the Tmin of February and the Tmax of April, whereas in 'Galega Vulgar' and 'Picual' there was instead a positive correlation with the Tmin of March. These two seemed to be more responsive to early warm weather, whereas 'Arbequina' and 'Cobrançosa' were less sensitive. This investigation revealed that olive cultivars behaved differently under the same environmental conditions and, in some genotypes, the ecodormancy release may be linked to endogenous factors in a stronger way.

Development and validation of EDU-COV-NEE-NEAE questionnaire to measure educational response, ICT usage and impact of COVID-19 pandemic on students with special educational needs.

The COVID-19 pandemic forced changes in the educational response, which involved the intensive use of Information and Communication Technology (ICT) skills, among other interventions. The circumstances required both families and teachers to make an extra effort to adapt to a reality that was expected to last for a long time. These changes had a special impact on students with Specific Needs of Educational Support (SNES) and Special Educational Needs (SEN). This study aimed to design and validate a questionnaire to assess the impact of the online training model used during the pandemic on teachers of students with SNES and SEN. We used qualitative, exploratory, and confirmatory factor analyses. The final 16-item scale was composed of three factors. Factor1, was labeled as "Educational Response.", factor2 was labeled "ICT Usage and Online Teaching," and Factor 3 was labeled "Impact." The questionnaire demonstrated good validity and reliability. This short questionnaire is recommended for assessing or monitoring the impact of changes in educational interventions involving ICT competencies due to COVID-19. However, they can also be easily adapted to similar circumstances or educational settings.

Hormonal Content and Gene Expression during Olive Fruit Growth and Ripening.

The cultivated olive (Olea europaea L. subsp. europaea var. europaea) is one of the most valuable fruit trees worldwide. However, the hormonal mechanisms underlying the fruit growth and ripening in olives remain largely uncharacterized. In this study, we investigated the physiological and hormonal changes, by ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS), as well as the expression patterns of hormone-related genes, using quantitative real-time PCR (qRT-PCR) analysis, during fruit growth and ripening in two olive cultivars, 'Arbequina' and 'Picual', with contrasting fruit size and shape as well as fruit ripening duration. Hormonal profiling revealed that olive fruit growth involves a lowering of auxin (IAA), cytokinin (CKs), and jasmonic acid (JA) levels as well as a rise in salicylic acid (SA) levels from the endocarp lignification to the onset of fruit ripening in both cultivars. During olive fruit ripening, both abscisic acid (ABA) and anthocyanin levels rose, while JA levels fell, and SA levels showed no significant changes in either cultivar. By contrast, differential accumulation patterns of gibberellins (GAs) were found between the two cultivars during olive fruit growth and ripening. GA1 was not detected at either stage of fruit development in 'Arbequina', revealing a specific association between the GA1 and 'Picual', the cultivar with large sized, elongated, and fast-ripening fruit. Moreover, ABA may play a central role in regulating olive fruit ripening through transcriptional regulation of key ABA metabolism genes, whereas the IAA, CK, and GA levels and/or responsiveness differ between olive cultivars during olive fruit ripening. Taken together, the results indicate that the relative absence or presence of endogenous GA1 is associated with differences in fruit morphology and size as well as in the ripening duration in olives. Such detailed knowledge may be of help to design new strategies for effective manipulation of olive fruit size as well as ripening duration.

Regulation of polyamine metabolism and biosynthetic gene expression during olive mature-fruit abscission.

Exogenous ethylene and some inhibitors of polyamine biosynthesis can induce mature-fruit abscission in olive, which could be associated with decreased nitric oxide production as a signaling molecule. Whether H2O2 also plays a signaling role in mature-fruit abscission is unknown. The possible involvement of H2O2 and polyamine in ethylene-induced mature-fruit abscission was examined in the abscission zone and adjacent cells of two olive cultivars. Endogenous H2O2 showed an increase in the abscission zone during mature-fruit abscission, suggesting that accumulated H2O2 may participate in abscission signaling. On the other hand, we followed the expression of two genes involved in the polyamine biosynthesis pathway during mature-fruit abscission and in response to ethylene or inhibitors of ethylene and polyamine. OeSAMDC1 and OeSPDS1 were expressed differentially within and between the abscission zones of the two cultivars. OeSAMDC1 showed slightly lower expression in association with mature-fruit abscission. Furthermore, our data show that exogenous ethylene or inhibitors of polyamine encourage the free putrescine pool and decrease the soluble-conjugated spermidine, spermine, homospermidine, and cadaverine in the olive abscission zone, while ethylene inhibition by CoCl2 increases these soluble conjugates, but does not affect free putrescine. Although the impact of these treatments on polyamine metabolism depends on the cultivar, the results confirm that the mature-fruit abscission may be accompanied by an inhibition of S-adenosyl methionine decarboxylase activity, and the promotion of putrescine synthesis in olive abscission zone, suggesting that endogenous putrescine may play a complementary role to ethylene in the normal course of mature-fruit abscission.

New drugs against multidrug-resistant Gram-negative bacteria: a systematic review of patents.

Background: Antimicrobial resistance has been a threat to human health ever since the accelerated consumption of antibiotics began. Materials & methods: The present systematic review was carried out using a free and specialized online database - Espacenet - and a survey for patents of antimicrobial agents from 2010 to 2021, selecting 33 recent patents that claimed compounds with antimicrobial activity against resistant strains of Gram-negative bacteria. Results: Some different and new approaches to the development of the patented antibacterial agents were identified, such as antimicrobial peptides, nanomaterials and natural extracts. Conclusion: Some alternatives to modern antibiotics with diminished effectiveness due to antimicrobial resistance were spotted. Nevertheless, many challenges remain to establish a robust and sustainable antibacterial R&D pipeline.

Polyamine-induced modulation of genes involved in ethylene biosynthesis and signalling pathways and nitric oxide production during olive mature fruit abscission.

After fruit ripening, many fruit-tree species undergo massive natural fruit abscission. Olive (Olea europaea L.) is a stone-fruit with cultivars such as Picual (PIC) and Arbequina (ARB) which differ in mature fruit abscission potential. Ethylene (ET) is associated with abscission, but its role during mature fruit abscission remains largely uncharacterized. The present study investigates the possible roles of ET and polyamine (PA) during mature fruit abscission by modulating genes involved in the ET signalling and biosynthesis pathways in the abscission zone (AZ) of both cultivars. Five ET-related genes (OeACS2, OeACO2, OeCTR1, OeERS1, and OeEIL2) were isolated in the AZ and adjacent cells (AZ-AC), and their expression in various olive organs and during mature fruit abscission, in relation to interactions between ET and PA and the expression induction of these genes, was determined. OeACS2, OeACO2, and OeEIL2 were found to be the only genes that were up-regulated in association with mature fruit abscission. Using the inhibition of ET and PA biosynthesis, it is demonstrated that OeACS2 and OeEIL2 expression are under the negative control of PA while ET induces their expression in AZ-AC. Furthermore, mature fruit abscission depressed nitric oxide (NO) production present mainly in the epidermal cells and xylem of the AZ. Also, NO production was differentially responsive to ET, PA, and different inhibitors. Taken together, the results indicate that PA-dependent ET signalling and biosynthesis pathways participate, at least partially, during mature fruit abscission, and that endogenous NO and 1-aminocyclopropane-1-carboxylic acid maintain an inverse correlation, suggesting an antagonistic action of NO and ET in abscission signalling.

Tissue-specific expression of olive S-adenosyl methionine decarboxylase and spermidine synthase genes and polyamine metabolism during flower opening and early fruit development.

Polyamines (PAs) are required for cell growth and cell division in eukaryotic and prokaryotic organisms. The present study is aimed at understanding the developmental regulation of PA biosynthesis and catabolism during flower opening and early fruit development in relation to fruit size and shape. Two full-length cDNA clones coding for S-adenosyl methionine decarboxylase (SAMDC) and spermidine synthase (SPDS) homologs, key steps in the PA biosynthesis pathway, in the stone-fruit of olive (Olea europaea L.) were identified and the spatial and temporal organization of these genes were described. In olive flowers, OeSAMDC gene transcripts were highly expressed in ovary wall, placenta and ovules, while OeSPDS transcript was confined to the ovules of ovary at anthesis stage. A correlation was detected between the SAMDC enzyme activity/accumulation transcript and spermidine (Spd) and spermine (Spm) levels during flower opening, implying that the synthesis of decarboxylated SAM might be a rate-limiting step in Spd and Spm biosynthesis. OeSAMDC and OeSPDS transcripts were co-expressed in fruit mesocarp and exocarp at all developmental stages analyzed as well as in nucellus, integuments and inner epidermis tissues of fertilized ovules. In contrast, the OeSAMDC and OeSPDS genes had different expression patterns during early fruit development. The results provide novel data about localization of PA biosynthesis gene transcripts, indicating that transcript levels of PA biosynthesis genes are all highly regulated in a developmental and tissue-specific manner. The differences between the two olive cultivars in the fruit size in relation to the differences in the accumulation patterns of PAs are discussed.