Climate Change Dependence in Ex Situ Conservation of Wild Medicinal Plants in Crete, Greece.

Over 80% of the global population addresses their primary healthcare needs using traditional medicine based on medicinal plants. Consequently, there's a rising demand for these plants for both household and industrial use at local, regional, national, and international levels. However, wild harvesting has negatively impacted natural ecosystems. Cultivating medicinal species has been proposed as a conservation strategy to alleviate this pressure. Yet, in this age of global climate change concerns, smallholder farmers' views on the benefits of such cultivation clash with the uncertainties of climate change impacts, amplifying their anxieties. In this context, the climate change dependence of ex situ cultivation of ten wild medicinal taxa with significant ethnopharmacological interest in Crete, Greece, were studied, projecting their potential habitat suitability under various future climate scenarios. The results demonstrated species-specific effects. Based on the potential cultivation area gains and losses, these effects can be categorized into three groups. We also outlined the spatial patterns of these gains and losses, offering valuable insights for regional management strategies benefiting individual practitioners.

Mining the ZINC database of natural products for specific, testosterone-like, OXER1 antagonists.

OXER1, the receptor for the oxidized arachidonic acid metabolite 5-oxo-ETE has been reported to play a significant role in inflammatory responses, being responsible for leucocyte chemotactic responses. Recently, we have identified OXER1 (GPR170) as a membrane receptor for androgens in prostate and breast cancer cells. Testosterone action via OXER1 induces specific Ca2+ release from intracellular organelles, modifies polymerized actin distribution induces apoptosis and decreases cancer cell migration. These actions are antagonized by 5-oxo-ETE. In addition, 5-oxo-ETE through a Gαi protein decreases cAMP, an action antagonized by testosterone. In this work, we mined the ZINC15 database, using QSAR, for natural compounds able to signal through Gαi and Gßγ simultaneously, mimicking testosterone actions, as well as for specific Gßγ interactors, inhibiting 5-oxo-ETE tumor promoting actions. We were able to identify four druggable Gαßγ and seven Gßγ specific OXER1 interactors. We further confirmed by bio-informatic methods their binding to the 5-oxo-ETE/testosterone binding groove of the receptor, their ADME properties and their possible interaction with other receptor and/or enzyme targets. Two compounds, ZINC04017374 (Naphthofluorescein) and ZINC08589130 (Puertogaline A) were purchased, tested in vitro and confirmed their OXER1 Gßγ and Gαßγ activity, respectively. The methodology followed is useful for a better understanding of the mechanism by which OXER1 mediates its actions, it has the potential to provide structural insights, in order to design small molecular specific interactors and ultimately design new anti-inflammatory and anti-cancer agents. Finally, the methodology may also be useful for identifying specific agonists/antagonists of other GPCRs.

A Mixture of Essential Oils from Three Cretan Aromatic Plants Inhibits SARS-CoV-2 Proliferation: A Proof-of-Concept Intervention Study in Ambulatory Patients.

INTRODUCTION: The need for effective therapeutic regimens for non-critically ill patients during the COVID-19 pandemic remained largely unmet. Previous work has shown that a combination of three aromatic plants' essential oils (CAPeo) (Thymbra capitata (L.) Cav., Origanum dictamnus L., Salvia fruticose Mill.) has remarkable in vitro antiviral activity. Given its properties, it was urgent to explore its potential in treating mild COVID-19 patients in primary care settings. METHODS: A total of 69 adult patients were included in a clinical proof-of-concept (PoC) intervention study. Family physicians implemented the observational study in two arms (intervention group and control group) during three study periods (IG2020, n=13, IG2021/22, n=25, and CG2021/22, n=31). The SARS-CoV-2 infection was confirmed by real-time PCR. The CAPeo mixture was administered daily for 14 days per os in the intervention group, while the control group received usual care. RESULTS: The PoC study found that the number and frequency of general symptoms, including general fatigue, weakness, fever, and myalgia, decreased following CAPeo administration. By Day 7, the average presence (number) of symptoms decreased in comparison with Day 1 in IG (4.7 to 1.4) as well as in CG (4.0 to 3.1), representing a significant decrease in the cumulative presence in IC (-3.3 vs. -0.9, p < 0.001; η2 = 0.20) on Day 7 and on Day 14 (-4.2 vs. -2.9, p = 0.027; η2 = 0.08). DISCUSSION/CONCLUSIONS: Our findings suggest that CAPeo possesses potent antiviral activity against SARS-CoV-2 in addition tο its effect against influenza A and B and human rhinovirus HRV14 strains. The early and effective impact on alleviating key symptoms of COVID-19 may suggest this mixture can act as a complementary natural agent for patients with mild COVID-19.

New Antagonists of the Membrane Androgen Receptor OXER1 from the ZINC Natural Product Database.

OXER1 (oxoeicosanoid receptor 1) was deorphanized in 1993 and found to be the specific receptor for the arachidonic acid metabolite 5-oxo-ETE. Recently, we have reported that androgen binds to this receptor also, being a membrane androgen receptor, triggering a number of its membrane-mediated actions (cell migration, apoptosis, cell proliferation, Ca2+ movements). In addition, our previous work suggested that a number of natural monomeric and oligomeric polyphenols interact with OXER1, acting similar to testosterone. Here, we interrogated the natural product chemical space and identified nine polyphenolic molecules with interesting in silico pharmacological activities as putative OXER1 antagonists. The molecule with the best pharmacokinetic-pharmacodynamic properties (ZINC15959779) was purchased and tested on OXER1, in prostate cancer cell cultures. It showed that it has actions similar to those of testosterone in inhibiting cAMP, while it had no action in intracellular Ca2+ mobilization or actin cytoskeleton rearrangement/migration. These results are discussed under the prism of structure-activity relationships and in silico models of the OXER1 binding groove. We suggest that these compounds, together with the previously reported (poly)phenolic compounds, can be lead structures for the exploration of the anti-inflammatory and antiproliferative effects of OXER1 antagonists.

From wild harvest towards precision agriculture: Use of Ecological Niche Modelling to direct potential cultivation of wild medicinal plants in Crete.

Understanding the distribution of wild medicinal plants and areas that are suitable for cultivation of these plants is important for both conservation and agriculture. Here, we study ten taxa with known ethnopharmacological uses, which have been used extensively in traditional medicine and as culinary supplements. We aim to (1) predict and map the potential habitat suitability for these taxa across the study area, (2) investigate spatial patterns that could have management implications, such as niche similarities among the taxa and suitability "hotspots" with the use of novel indices, and (3) develop a platform where parts of this information can be accessed and utilized by all interested groups, from the policy-maker level to the individual practitioner level. Ecological Niche Models developed for each study taxon, based on topographic, bioclimatic, soil, and land use variables had high predictive power and were used as the basis for suitability visualization. A series of informative indices were also calculated and mapped, revealing spatial patterns not readily observable from the single-taxon predictions, and providing valuable information to managers. Finally, a web-based, easy-to-use application was also created, where the predicted suitability scores for the study area can be made accessible to anyone interested. The application can provide information both in a visual form (i.e. maps of predicted suitability) and in a numerical form (i.e. estimated suitability scores for all taxa in a given geographical location). This study provides the scientific tools to make a step towards cultivating a group of economically important wild medicinal plants in Crete, as well as the tools to disseminate this information to decision makers and practitioners, and eventually integrate the research findings in local agricultural practices.

Exogenous induction of thermogenesis in Arum concinnatum by salicylic acid.

Arum concinnatum Schott is a highly thermogenic species, with the temperature of the appendix exceeding ~10.9°C above the ambient temperature during thermogenesis, whereas the rates of respiration of the male florets in intact inflorescences peak at 0.92µmol s-1 g-1, which is the highest rate so far measured among the plants. Here, we attempt the ex situ exogenous induction of thermogenesis in whole inflorescences and in separate appendices of the spadix, and explore the thermogenic patterns under controlled laboratory conditions of light and temperature. Mature but unopened inflorescences and appendices showed thermogenic responses when treated with salicylic acid (SA), but not when treated with distilled water (control). With regard to light conditions, the responses revealed only one significant difference for inflorescences, which concerns the higher maximum temperature in the continuous light treatment compared with continuous dark. Along the ambient temperature gradient, at the lowest temperature edge individuals remained stable close to ambient temperature and to control. These findings suggest that, in general, ex situ exogenous induction of thermogenesis can be achieved in whole inflorescences and in separate appendices of spadix of A. concinnatum using SA. This study also indicates that SA acts independently of light conditions, while exogenous induction of thermogenesis takes place within an ambient temperature range.

Post-market outcome of an extract of traditional Cretan herbs on upper respiratory tract infections: a pragmatic, prospective observational study.

BACKGROUND: The beneficial effects of traditional herbs of Crete, Greece for the treatment of upper respiratory tract infections have been reported in observational and laboratory studies. Following a published, double blind, randomized, placebo controlled trial, this study aimed to assess the effectiveness of an extract of three Cretan herbs on the treatment of upper respiratory tract infections, upon its market release. METHODS: An observational study was conducted in Heraklion, Crete, Greece. Participants were patients presenting at selected pharmacies with symptoms of upper respiratory tract infection, choosing to receive the extract for their treatment. Patients' symptoms (local, general, total) where recorded at three time points within 1 week, using a questionnaire developed based on the Wisconsin Upper Respiratory System Survey. For each patient, symptoms were scored on a 0-7 Likert scale and three indexes were calculated: the score of local symptoms, the score of general symptoms and the total score of symptoms. Effectiveness was assessed by examining the reduction in these indexes over the 1-week observation period. RESULTS: Mean score of general symptoms was 19.1 (SE: 0.9) in day 1, dropping to 8.6 (SE: 0.6) and 3.1 (SE: 0.4) in days 4 and 7 respectively. Mean score of local symptoms declined from 7.9 (SE: 0.5) in day 1 to 2.3 (SE: 0.3) in day 4 and to 0.5 (SE: 0.1) in day 7. Total score of symptoms reached 27.0 (SE: 1.2) in day 1, decreasing to 10.9 (SE: 0.8) in day 4 and to 3.5 (SE: 0.5) in day 7. The percentage of participants reporting fever was 82.1% at baseline, 8.0% in day 4 and 2.0% in day 7 (p < 0.0001 for paired differences). Multiple regression models indicated that supplementary medication intake did not seem to affect symptoms' severity or the day patients reported that their symptoms ceased completely. CONCLUSIONS: This pragmatic study added evidence about the potential therapeutic effects of an extract of Cretan herbs on the amelioration of upper respiratory tract infection symptoms.

Environmental (in)dependence of a hybrid zone: Insights from molecular markers and ecological niche modeling in a hybrid zone of Origanum (Lamiaceae) on the island of Crete.

The role of environment and the relative significance of endogenous versus exogenous selection in shaping hybrid zones have been crucial issues in the studies of hybridization. Recent advances in ecological niche modeling (ENM) offer new methodological tools, especially in combination with the genotyping of individuals in the hybrid zone. Here, we study the hybrid zone between the widely known spices Origanum onites and Origanum vulgare ssp. hirtum in Crete. We analyze the genetic structure of both parental taxa and their hybrid Origanum × intercendens using AFLP markers on 15 sympatric and 12 allopatric populations and employ ecological niche modeling and niche similarity tests to study their niche patterns. We complement these analyses with seed viability measurements. Our study revealed that the hybridizing taxa O. onites and O. vulgare ssp. hirtum and the resulting genotypic classes showed geographical and environmental niche similarities based on the predictions of ENMs and the subsequent similarity tests. The occurrence of the hybrid zone is not directly dependent on environmental factors which favor the fitness of the hybrid compared to the parental taxa, but rather on aspects such as historical factors and management practices, which may contribute to the localization and maintenance of the contact zone between parental species. Our results suggest that if a minimum required niche differentiation between genotypic classes is not achieved, environmental dependence might not have a prominent role on the outcome of the hybridization.

Chemical and genetic characterization of Phlomis species and wild hybrids in Crete.

The genus Phlomis is represented in the island of Crete (Greece, Eastern Mediterranean) by three species Phlomis cretica C. Presl., Phlomis fruticosa L., the island endemic Phlomis lanata Willd. and three hybrids Phlomis x cytherea Rech.f. (P. cretica x P. fruticosa), Phlomis x commixta Rech.f. (P. cretica x P. lanata) and Phlomis x sieberi Vierh. (P. fruticosa x P. lanata). This work describes (a) the profile of hybrids and parental species concerning their volatile compounds, (b) the suitability of ribosomal nuclear (ITS region), chloroplast (trnH-psbA), and AFLP markers to identify hybrids and (c) their competence to characterize the different chemotypes of both hybrids and their parental species. The cluster analysis and PCA constructed from chemical data (volatile oils) suggest that there are three groups of taxa. Group IA includes P. cretica and P. fruticosa, group IB includes P. x cytherea, whereas group II consists of P. x commixta, P. x sieberi and P. lanata. Volatile compounds detected only in the hybrids P. x sieberi and P. x commixta correspond to the 3% of the total compounds, value that is much higher in P. x cytherea (21%). Neighbor-joining, statistical parsimony analysis and the observations drawn from ribotypes spectrum of ITS markers divided Phlomis species in two groups, P. lanata and the complex P. cretica/P. fruticosa. In contrast to the ITS region, the plastid DNA marker follows a geographically related pattern. Neighbor-Net, PCA and Bayesian assignment analysis performed for AFLP markers separated the genotypes into three groups corresponding to populations of P. cretica, P. fruticosa, and P. lanata, respectively, while populations of P. x commixta, P. x cytherea, and P. x sieberi presented admixed ancestry. Most of the P. x cytherea samples were identified as F1 hybrids by Bayesian assignment test, while those of P. x commixta and P. x sieberi were identified as F2 hybrids. Overall, high chemical differentiation is revealed in one of the three hybrids, which is likely related with niche variation. Moreover, molecular markers show potential to identify Phlomis taxa.