IN-VITRO REGENERATION OF CALENDULA MARITIMA GUSS. (ASTERACEAE), A THREATENED PLANT ENDEMIC TO WESTERN SICILY

Calendula maritima is a critically endangered endemic plant of Western Sicily. Besides habitat destruction, the hybridization with the contiguous congener species C. fulgida is a major threat to its conservation. For this reason, seed-based propagation and seed storage are not appropriate for conservation purposes. In the present paper we describe a rapid and prolific in vitro plant regeneration method by direct organogenesis from leaves of C. maritima. Leaf explants were cultured on solid Murashige and Skoog (MS) medium in the presence of several plant growth regulator combinations. The best shoot multiplication rate (2.5 shoots/explant) was obtained on the medium containing 4.4 µM 6-benzylaminopurine in combination with 10 µM ß-naphthoxyacetic acid. Regenerated shoots were successfully rooted on solid MS medium supplemented with several auxins and the best result was obtained with 1.0 µM indole-3-acetic acid (35% of plantlets rooted). Plantlets were thereafter established in the greenhouse (survival frequency 75%) and no phenotypic variations were observed between regenerants and the mother plant

Increased Zygote-Derived Plantlet Formation through In Vitro Rescue of Immature Embryos of Highly Apomictic Opuntia ficus-indica (Cactaceae)

O. ficus-indica (prickly pear cactus) is an important forage and food source in arid and semiarid ecosystems and is the most important cactus species in cultivation globally. The high degree of apomixis in the species is a hindrance in plant breeding programs where genetic segregation is sought for the selection of superior genotypes. To understand if in ovulo embryo rescue could increase the proportion of zygotic seedlings, we compared the mature seed-derived seedlings with those regenerated from in vitro embryo rescue at 20, 25, 30, 35, and 40 post-anthesis days (PADs) in four Italian cultivars. The seedlings were classified as apomictic or zygotic based on molecular marker analysis using inter-sequence single repeat (ISSR) primers. Multiple embryos were recovered from all the cultured immature ovules, and plantlets were regenerated and acclimatized to the field post hardening, with success rates ranging from 62% (‘Senza spine’) to 83% (‘Gialla’). The level of polyembryony differed among cultivars and recovery dates, with the highest being ‘Rossa’, producing 4.8 embryos/ovule at 35 PADs, and ‘Gialla’, the lowest, with 2.7 at 40 PADs. The maximum number of embryos observed within a single ovule was 14 in ‘Trunzara bianca’. ISSR analysis revealed that ovule culture at 35 PADs produced the highest percentage of zygotic seedlings in all the cultivars, from 51% (‘Rossa’) to 98% (‘Gialla’), with a high genotype effect as well. Mature seeds produced much fewer seedlings per seed, ranging from 1.2 in ‘Trunzara bianca’ to 2.0 in ‘Rossa’ and a lower percentage of zygotic seedlings (from 14% in ‘Rossa’ to 63% in ‘Gialla’). Our research opens a pathway to increase the availability of zygotic seedlings in O. ficus-indica breeding programs through in ovulo embryo culture

Long-Term Field Evaluation of Conventional vs. Micropropagated Plants of Chrysanthemum cinerariifolium

Pyrethrum is a perennial herbaceous plant endemic to the eastern coast of the Adriatic Sea, and introduced in large areas of nearly all continents, where it is cultivated for the industrial extraction of pyrethrins. Pyrethrins are a group of six closely related monoterpene esters, widely used as natural insecticides. The world production of natural pyrethrins is lower than the market demand, and a wider introduction of this crop within the Mediterranean agrosystems could be an appealing opportunity for farmers and manufacturers. The availability of adequate amounts of selected plant material to bring into cultivation is, however, one of the major issues. Therefore, the in vitro propagation of elite pyrethrum genotypes could be a suitable alternative to conventional propagation methods. In this paper, we present the results of a 9-year field comparison between pyrethrum plants coming from an in vitro propagation protocol and plants obtained by cutting from the same mother plants. Furthermore, since plantlets derived from in vitro regeneration may experience ploidy changes, we evaluated the stability of the ploidy level of pyrethrum micropropagated plants by flow cytometry (FCM) analysis. FCM screening revealed no differences among the morphotypes and between them and the mother plant. Likewise, the field evaluation of plants gave no significant differences between flower yields in both groups. Hence, micropropagation was confirmed as an easy, efficient and reproducible method to obtain large quantities of selected pyrethrum genotypes

Efficacy of Cannabidiol for Delta-9-Tetrahydrocannabinol-Induced Psychotic Symptoms, Schizophrenia, and Cannabis Use Disorders: A Narrative Review

Although cannabis’ major psychoactive component, ∆-9-tetrahydrocannabinol (THC), has been linked to both earlier onset and poorer outcomes of psychotic disorders, Cannabidiol (CBD) seems to have different pharmacological mechanisms and potential therapeutic properties. However, no clinical study has investigated CBD for the treatment of co-occurring psychotic and cannabis use disorders so far, even though its utility seems grounded in a plausible biological basis. The aim of this work is thus to provide an overview of available clinical studies evaluating the efficacy of CBD for psychotic symptoms induced by THC, schizophrenia, and cannabis use disorders. After searching for relevant studies in PubMed, Cochrane Library, and ClinicalTrials.gov, we included 10 clinical studies. Available evidence suggests that CBD may attenuate both psychotic-like symptoms induced by THC in healthy volunteers and positive symptoms in individuals with schizophrenia. In addition, preliminary data on the efficacy of CBD for cannabis use disorders show mixed findings. Evidence from ongoing clinical studies will provide insight into the possible role of CBD for treating psychotic and cannabis use disorders

Pre-Discharge Predictors of 1-Year Rehospitalization in Adolescents and Young Adults with Severe Mental Disorders: A Retrospective Cohort Study

BACKGROUND AND OBJECTIVES: Readmissions of youths hospitalized for a severe mental disorder are common events and bear a remarkable human, social, and economic burden. The current study aimed at evaluating predictors of 1-year rehospitalization in a sample of adolescents and young adults with severe mental disorders. MATERIALS AND METHODS: Data for ≤25-year-old inpatients with a severe mental disorder and consecutively admitted between 1 January 2016 and 30 June 2019 were collected. Subjects were retrospectively assessed over a follow-up period of one year after the index discharge to track readmissions—i.e., the primary outcome variable. Standard descriptive statistics were used. The association between variables and 1-year rehospitalization was estimated using the univariate Cox proportional hazards regression model. We then carried out a multivariable Cox regression model, also estimating the covariate-adjusted survivor function. Hazard ratios (HRs) with related 95% confidence intervals (95% CIs) were provided. RESULTS: The final sample included 125 individuals. The multivariable Cox regression model estimated that co-occurring substance use disorders (HR = 2.14; 95% CI: 1.08 to 4.26; p = 0.029) and being admitted for a suicide attempt (HR = 2.49; 95% CI: 1.13 to 5.49; p = 0.024) were both significant predictors of 1-year rehospitalization. CONCLUSIONS: Our study showed that comorbid substance use disorders and being admitted for a suicide attempt were predictors of early readmission in youths with severe mental disorders. Although their generalizability is limited, our findings could contribute to improve the quality of young patients’ mental health care by identifying vulnerable subjects who may benefit from tailored interventions to prevent rehospitalizations

Overcoming sexual sterility in conservation of endangered species: the prominent role of biotechnology in the multiplication of Zelkova sicula (Ulmaceae), a relict tree at the brink of extinction

Biotechnology provides valuable tools to support conservation of plant species, especially in case of threatened taxa or when dealing with seed unavailability, low viability or sterility. However, plant cell culture methods have often to face problems associated with tissue recalcitrance to in vitro systems. Recalcitrance can be related to a variety of triggering factors, involving many efforts and manipulations within one or more of the micropropagation stages before obtaining successful results. An in vitro propagation protocol was developed for Zelkova sicula, a very rare and endangered relict tree, endemic to Sicily (Southern Italy). The species revealed extremely recalcitrant to in vitro culture approaches, but after many trials throughout a number of years an effective micropropagation protocol was completed. The rooting rate was about 84% of the treated explants, 8% of which were successfully acclimatized outdoor and reintroduced in the wild within a comprehensive conservation project. The technique allowed to overcome the problems of sexual sterility of this species, hence contributing concretely to contrast the problems connected with its conservation. However, additional efforts need to be carried out in order to refine the acclimatization step and further improve the whole process effectiveness

Off-label long acting injectable antipsychotics in real-world clinical practice: a cross-sectional analysis of prescriptive patterns from the STAR Network DEPOT study

Introduction Information on the off-label use of Long-Acting Injectable (LAI) antipsychotics in the real world is lacking. In this study, we aimed to identify the sociodemographic and clinical features of patients treated with on- vs off-label LAIs and predictors of off-label First- or Second-Generation Antipsychotic (FGA vs. SGA) LAI choice in everyday clinical practice. Method In a naturalistic national cohort of 449 patients who initiated LAI treatment in the STAR Network Depot Study, two groups were identified based on off- or on-label prescriptions. A multivariate logistic regression analysis was used to test several clinically relevant variables and identify those associated with the choice of FGA vs SGA prescription in the off-label group. Results SGA LAIs were more commonly prescribed in everyday practice, without significant differences in their on- and off-label use. Approximately 1 in 4 patients received an off-label prescription. In the off-label group, the most frequent diagnoses were bipolar disorder (67.5%) or any personality disorder (23.7%). FGA vs SGA LAI choice was significantly associated with BPRS thought disorder (OR = 1.22, CI95% 1.04 to 1.43, p = 0.015) and hostility/suspiciousness (OR = 0.83, CI95% 0.71 to 0.97, p = 0.017) dimensions. The likelihood of receiving an SGA LAI grew steadily with the increase of the BPRS thought disturbance score. Conversely, a preference towards prescribing an FGA was observed with higher scores at the BPRS hostility/suspiciousness subscale. Conclusion Our study is the first to identify predictors of FGA vs SGA choice in patients treated with off-label LAI antipsychotics. Demographic characteristics, i.e. age, sex, and substance/alcohol use co-morbidities did not appear to influence the choice towards FGAs or SGAs. Despite a lack of evidence, clinicians tend to favour FGA over SGA LAIs in bipolar or personality disorder patients with relevant hostility. Further research is needed to evaluate treatment adherence and clinical effectiveness of these prescriptive patterns

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. For example, a key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process versus those that measure fl ux through the autophagy pathway (i.e., the complete process including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defi ned as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost higher eukaryotes and some protists such as Dictyostelium ) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the fi eld understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation it is imperative to delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so not all Atg proteins can be used as a specific marker for an autophagic process. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field

Potential use of new diphenylurea derivativesin micropropagation of Capparis spinosa L.

A protocol for in vitro multiplication of caper (Capparis spinosa L. subsp. rupestris) from nodal segments collected from mature plants was developed. For shoot multiplication, one auxin (indol-3-butyric acid, IBA) and cytokinins of two different classes were used: the N6-substituted adenine derivatives 6-benzylamino purine (BAP), and the two synthetic phenylurea derivatives Nphenyl- N0-benzothiazol-6-ylurea (PBU) and N-phenyl-N0- (1,2,3-thidiazol-5-yl) urea (thidiazuron, TDZ). Maximum shoot production was achieved from explants cultured with the adeninic cytokinin BAP (4 lM) and the auxin IBA (0.5 lM). New shoots longer than 1 cm were used for rooting. To induce root formation, three auxins [indole-3- butyric acid (IBA), 1-naphthaleneacetic acid (NAA) and 3-Indoleacetic acid (IAA)] and two synthetic phenylurea derivatives [N,N-bis-(2,3-methylenedioxyphenyl)urea (2,3-MDPU) and N,N-bis-(3,4-methylenedioxyphenyl)urea (3,4-MDPU)] were used. All rooting compounds tested stimulated the formation of roots. However, the best result in terms of a high percentage of rooted shoots having a well-developed root system with many lateral roots was achieved with the synthetic phenylurea 2,3-MDPU (1 lM) with 93.7% of well rooted plantlets. About 80% of rooted plantlets were successfully acclimatized and transferred to the greenhouse

Influence of Culture Conditions on In Vitro Asymbiotic Germination of Anacamptis longicornu and Ophrys panormitana (Orchidaceae)

This study is the first approach to in vitro asymbiotic germination of two species of Sicilian threatened terrestrial orchids, Anacamptis longicornu and Ophrys panormitana. Seeds were collected in the wild and cultured in two different media—Orchimax medium (OM) and Murashige and Skoog (MS)—and exposed to different photoperiods and temperatures to evaluate the best conditions for the specific stages of development. The germination of A. longicornu was very high on OM (95.5%) and lower on MS medium (21.4%), whereas O. panormitana germinated only on OM medium, with significantly lower percentages (12.0%), compared with A. longicornu. This difference is caused by variation in quality and quantity of nutrients used, primarily by nitrogen source. The results show that temperature and photoperiod widely affect seed germination and development. Although further investigations on asymbiotic and symbiotic germination are needed for the improvement of conservation of Mediterranean terrestrial orchids, our results contribute to the conservation of this group of plants