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BACKGROUND AND AIMS: Current laboratory methods for opioid detection involve an initial screening with immunoassays which offers efficient but non-specific results and a subsequent liquid chromatography-tandem mass spectrometry (LC-MS/MS) confirmation which offers accurate results but requires extensive sample preparation and turnaround time. Direct Analysis in Real Time (DART) tandem mass spectrometry is evaluated as an alternative approach for accurate opioid detection with efficient sample preparation and turnaround time. MATERIALS AND METHODS: DART-MS/MS was optimized by testing the method with varying temperatures, operation modes, extraction methods, hydrolysis times, and vortex times. The method was evaluated for 12 opioids by testing the analytical measurement range, percent carryover, precision studies, stability, and method-to-method comparison with LC-MS/MS. RESULTS: DART-MS/MS shows high sensitivity and specificity for the detection of 6-acetylmorphine, codeine, hydromorphone, oxymorphone, hydrocodone, naloxone, buprenorphine, norfentanyl, and fentanyl in urine samples. However, its performance was suboptimal for norbuprenorphine, morphine and oxycodone. CONCLUSION: In this proof-of-concept study, DART-MS/MS is evaluated for its rapid quantitative definitive testing of opioids drugs in urine. Further research is needed to expand its application to other areas of drug testing.

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Carpaine, a major alkaloid present in Carica papaya leaves, has been shown to increase platelet counts in patients suffering from thrombocytopenia. Numerous commercial papaya leaf products are available, but few provide proper bioactive ingredient information. We present herein a technique for rapid screening of carpaine in these products using DART-MS. The results indicate that carpaine was detected in various forms (powder, solution) of papaya leaves. Its presence was confirmed by examining the mass pattern when conducted on a standard solution at both low and high voltages (+10 V and +90 V), using MS1 and MS2 data obtained from LC-QTOF-MS/MS. The protonated molecule was identified at m/z = 479.38, with a fragment ion at m/z = 240.20. LOD for identifying carpaine in powder and solution matrices were 5.0 × 10-5 %w/w and 0.05 µg/mL, respectively. The proposed method has been successfully validated with the AOAC International standards and can be used to identify carpaine in papaya leaf products.

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To comply with a more circular and environmentally friendly European common agricultural policy, while also valorising sunflower by-products, an ultrasound assisted extraction (UAE) was tested to optimise ethanol-wash solutes (EWS). Furthermore, the capabilities of DART-HRMS as a rapid and cost-effective tool for determining the biochemical changes after valorisation of these defatted sunflower EWS were investigated. Three batches of EWS were doubly processed into optimised EWS (OEWS) samples, which were analysed via DART-HRMS. Then, the metabolic profiles were submitted to a univariate analysis followed by a partial least square discriminant analysis (PLS-DA) allowing the identification of the 15 most informative ions. The assessment of the metabolomic fingerprinting characterising EWS and OEWS resulted in an accurate and well-defined spatial clusterization based on the retrieved pool of informative ions. The outcomes highlighted a significantly higher relative abundance of phenolipid hydroxycinnamoyl-glyceric acid and a lower incidence of free fatty acids and diglycerides due to the ultrasound treatment. These resulting biochemical changes might turn OEWS into a natural antioxidant supplement useful for controlling lipid oxidation and to prolong the shelf-life of foods and feeds. A standardised processing leading to a selective concentration of the desirable bioactive compounds is also advisable.

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BACKGROUND: When developing new antimalarial drugs, considering their potential use during pregnancy as preventive or curative therapy is crucial. This prevents the parasite from affecting embryonic development and reduces maternal and fetal death risks. Consequently, understanding the exposure and safety of antimalarial drugs during pregnancy is crucial, with well-designed animal studies playing a key role in this assessment. METHODS: As part of the drug development program for cabamiquine, a series of developmental and reproductive toxicity studies were conducted in rats and rabbits. Additionally, the zebrafish embryo model was used to further improve embryo exposure, minimize confounding factors related to maternal toxicity, and assess developmental risks of cabamiquine. RESULTS: In these studies, although maternal toxicity was observed, there were no cabamiquine-related adverse effects on fertility, embryonic, or fetal development at maternal exposures representing significant multiples (up to five and 10 times higher in rabbit and rats, respectively) than the exposure at the anticipated efficacious human dose. Similarly, no adverse effects were observed on ZF embryonic development, even though cabamiquine concentrations in the embryos were 10-fold higher than nominal concentrations. CONCLUSIONS: The results obtained in a full set of reproductive toxicity studies did not provide evidence of detrimental effects on the conceptuses and progeny at maternally nontoxic doses and exposures, still representing a multiple of the anticipated systemic exposures in women of childbearing potential (WOCBP). Cabamiquine can therefore be considered a suitable therapeutic option for WOCBP and pregnant women living in malaria-endemic regions by significantly reducing maternal and infant malaria death rates.

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Objective.In diffusing alpha-emitters radiation therapy ('Alpha DaRT'), the diffusion-leakage (DL) model is used to determine the spatial distributions of the emitters and the corresponding alpha dose, critical for a successful treatment. This work first introduces a finite volume (FV) approach to develop numerical schemes to simulate the DL model in one, two and three dimensions then presents how variations over realistic ranges of the DL model parameters related to desorption, diffusion and leakage processes affect the alpha dose distribution and the position of the clinically significant alpha particle10Gy isodose. This work also presents the effects of three modeling approximations: two source geometry approximations (solid cylinder instead of hollow, pixelized cross section instead of circular), and one dosimetric approximation (single-source dose superposition instead of multiple-sources direct dose calculation).Approach.The introduced FV approach was used to obtain spatial distributions of the emitters, from which the corresponding alpha dose distributions were calculated under the assumption of a local deposition of the alpha particles' energies. Variation ranges of the DL model parameters were based on previously published data. For each modeling approximation studied, the error and relative error on the alpha dose distribution were calculated and the displacement of the10Gy isodose was evaluated.Main results.Over realistic ranges, the desorption probabilities, diffusion lengths, and leakage probabilities affect the position of the alpha particle10Gy isodose by∼0.1mm,∼1.5mm and∼0.5mm, respectively. The three modeling approximations studied have a negligible effect on the alpha particle10Gy isodose position, with displacements⩽0.01mm.Significance.This work quantitatively evaluates the relative importance of different parameters and approximations in Alpha-DaRT alpha dose calculations based on their impact not only on the dose variation at a given distance from the source but also on the displacement of clinically significant isodoses.

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This study explores the relationship between government and citizen in the digital age and proposes a specific path to realize the value co-creation of government and citizen in the digital age. Quantitative analysis of 40 policy texts on "digital government construction" of 30 provincial and municipal governments in China before November 2022 was conducted using NVivo12. The case of Minsheng Cloud revealed that provincial and municipal government departments have not paid sufficient attention to citizen participation. Furthermore, citizens' participation ability and enthusiasm are not high. The study found that in the process of strengthening the construction of digital government, "government optimisation", "citizen participation", "governance guarantee", "data governance" and "government digitization" are the key elements. We discuss the feasibility, necessity and coupling of the value co-creation theory to establish the benign interaction between government and citizens and realize co-creation. We find that the "DART" model can be co-created based on value, and that the continuous Improving the four aspects of "dialogue", "access", "transparency" and "risk assessment" will undoubtedly help realize the value co-creation of government and citizens in the digital era, and form a governance pattern of co-construction and sharing. It is therefore of great significance to promote the government's digital construction and digital governance capability to accelerate the construction of digital China and promote the modernisation of the national governance system and governance capability.

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One of the biggest challenges for in vivo gene therapy are vectors mediating highly selective gene transfer into a defined population of therapy-relevant cells. Here we present DARPin-targeted AAVs (DART-AAVs) displaying DARPins specific for human and murine CD8. Insertion of DARPins into the GH2/GH3 loop of the capsid protein 1 (VP1) of AAV2 and AAV6 resulted in high selectivity for CD8-positive T cells with unimpaired gene delivery activity. Remarkably, the capsid core structure was unaltered with protruding DARPins detectable. In complex primary cell mixtures, including donor blood or systemic injections into mice, the CD8-targeted AAVs were by far superior to unmodified AAV2 and AAV6 in terms of selectivity, target cell viability, and gene transfer rates. In vivo, up to 80% of activated CD8+ T cells were hit upon a single vector injection into conditioned humanized or immunocompetent mice. While gene transfer rates decreased significantly under non-activated conditions, genomic modification selectively in CD8+ T cells was still detectable upon Cre delivery into indicator mice. In both mouse models, selectivity for CD8+ T cells was close to absolute with exceptional detargeting from liver. The CD8-AAVs described here expand strategies for immunological research and in vivo gene therapy options.

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The mass spectral database of tree species built by US Fish and Wildlife Service has thousands of entries and has been a valuable resource to combat illegal logging and international trade. The database was and continues to be constructed using a particular ambient ionization time-of-flight mass spectrometry (TOF-MS) platform in the agency branch in Ashland, OR, with which queries of unknown wood samples are investigated exclusively. Laboratories that operate different MS instruments also have an interest in using the database if they can produce valid matches to known samples compatible with the database. Four species were selected for inter-laboratory comparison using Orbitrap MS instruments and the equivalent TOF-MS platform with direct analysis in real time ionization of institution-sourced wood samples. Identities of the known samples were confirmed by examination of their microscopic wood anatomy. Orbitrap analysis was able to identify each species as confidently as the TOF instruments, often with less variation in spectra but not necessarily greater mass accuracy or better-matched signal abundance to the control database. The Orbitrap program also had to be doubled to two scanned mass ranges appended for greater peak intensity, before spectra could be correctly matched to the database, but the program was successful.

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With the advent of soft ionization techniques such as electrospray (ESI) and matrix-assisted laser desorption/ionization (MALDI) to produce intact gas-phase ions from nonvolatile macromolecules, mass spectrometry has become an essential technique in the field of polymeric materials. However, (co)polymers of very high molecular weight or with reticulated architectures still escape ESI or MALDI, mainly due to solubility issues. Strategies developed to tackle such an analytical challenge all rely on sample degradation to produce low-mass species amenable to existing ionization methods. Yet, chain degradation needs to be partial and controlled to generate sufficiently large species that still contain topological or architectural information. The present article reviews the different analytical degradation strategies implemented to perform mass spectrometry of these challenging synthetic polymers, covering thermal degradation approaches in sources developed in the 2000s, off-line sample pre-treatments for controlled chemical degradation of polymeric substrates, and most recent achievements employing reactive ionization modes to perform chemolysis on-line with MS.

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The initial favorable efficacy and safety profile for Alpha DaRT have been demonstrated (NCT04377360); however, the longer-term safety and durability of the treatment are unknown. This pooled analysis of four prospective trials evaluated the long-term safety and efficacy of Alpha DaRT for the treatment of head and neck or skin tumors. A total of 81 lesions in 71 patients were treated across six international institutions, with a median follow-up of 14.1 months (range: 2-51 months). Alpha DaRT sources were delivered via a percutaneous interstitial technique and placed to irradiate the tumor volume with the margin. The sources were removed two to three weeks following implantation. A complete response was observed in 89% of treated lesions (n = 72) and a partial response in 10% (n = 8). The two-year actuarial local recurrence-free survival was 77% [95% CI 63-87]. Variables, including recurrent versus non-recurrent lesions, baseline tumor size, or histology, did not impact long-term outcomes. Twenty-seven percent of patients developed related acute grade 2 or higher toxicities, which resolved with conservative measures. No grade 2 or higher late toxicities were observed. These data support the favorable safety profile of Alpha DaRT, which is currently being explored in a pivotal US trial.

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Salinity tolerance was studied in chickpea accessions from a germplasm collection and in cultivars from Kazakhstan. After NaCl treatment, significant differences were found between genotypes, which could be arranged into three groups. Those that performed poorest were found in group 1, comprising five ICC accessions with the lowest chlorophyll content, the highest leaf necrosis (LN), Na+ accumulation, malondialdehyde (MDA) content, and a low glutathione ratio GSH/GSSG. Two cultivars, Privo-1 and Tassay, representing group 2, were moderate in these traits, while the best performance was for group 3, containing two other cultivars, Krasnokutsky-123 and Looch, which were found to have mostly green plants and an exact opposite pattern of traits. Marker-trait association (MTA) between 6K DArT markers and four traits (LN, Na+, MDA, and GSH/GSSG) revealed the presence of four possible candidate genes in the chickpea genome that may be associated with the three groups. One gene, ATP-binding cassette, CaABCC6, was selected, and three haplotypes, A, D1, and D2, were identified in plants from the three groups. Two of the most salt-tolerant cultivars from group 3 were found to have haplotype D2 with a novel identified SNP. RT-qPCR analysis confirmed that this gene was strongly expressed after NaCl treatment in the parental- and breeding-line plants of haplotype D2. Mass spectrometry of seed proteins showed a higher accumulation of glutathione reductase and S-transferase, but not peroxidase, in the D2 haplotype. In conclusion, the CaABCC6 gene was hypothesized to be associated with a better response to oxidative stress via glutathione metabolism, while other candidate genes are likely involved in the control of chlorophyll content and Na+ accumulation.

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During its temporary tenure, the placenta has extensive and specialized functions that are critical for pre- and post-natal development. The consequences of chemical exposure in utero can have profound effects on the structure and function of pregnancy-associated tissues and the life-long health of the birthing person and their offspring. However, the toxicological importance and critical functions of the placenta to embryonic and fetal development and maturation have been understudied. This narrative will review early placental development in humans and highlight some in vitro models currently in use that are or can be applied to better understand placental processes underlying developmental toxicity due to in utero environmental exposures.

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Background: Internet hospitals, online health communities, and other digital health APPs have brought many changes to people's lives. However, digital health resources are experiencing low continuance intention due to many factors, including information security, service quality, and personal characteristics of users. Methods: We used cross-sectional surveys and structural equation modeling analysis to explore factors influencing user willingness to continue using digital health resources. Results: Information quality (ß = 0.31, p < 0.05), service quality (ß = 0.19, p < 0.05), platform reputation (ß = 0.34, p < 0.05), and emotional support (ß = 0.23, p < 0.05) have significant positive effects on user value co-creation behavior. Additionally, user trust and perceived usefulness could mediate the association between user value co-creation behavior and continuance intention, with mediation effects of 0.143 and 0.125, respectively. User involvement can positively moderate the association between user value co-creation behavior and user trust (ß = 0.151, t = 2.480, p < 0.001). Also, user involvement can positively moderate the association between value co-creation behavior and perceived usefulness (ß = 0.103, t = 3.377, p < 0.001). Conclusion: The keys to solving the problem of low continuance intention are improving the quality and service level of digital health resources, and promoting users' value co-creation behavior. Meanwhile, enterprises should build a good reputation, create a positive communication atmosphere in the community, and enhance user participation and sense of belonging.

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Direct-to-Mass Spectrometry and ambient ionization techniques can be used for biochemical fingerprinting in a fast way. Data processing is typically accomplished with vendor-provided software tools. Here, a novel, open-source functionality, entitled Tidy-Direct-to-MS, was developed for data processing of direct-to-MS data sets. It allows for fast and user-friendly processing using different modules for optional sample position detection and separation, mass-to-charge ratio drift detection and correction, consensus spectra calculation, and bracketing across sample positions as well as feature abundance calculation. The tool also provides functionality for the automated comparison of different sets of parameters, thereby assisting the user in the complex task of finding an optimal combination to maximize the total number of detected features while also checking for the detection of user-provided reference features. In addition, Tidy-Direct-to-MS has the capability for data quality review and subsequent data analysis, thereby simplifying the workflow of untargeted ambient MS-based metabolomics studies. Tidy-Direct-to-MS is implemented in the Python programming language as part of the TidyMS library and can thus be easily extended. Capabilities of Tidy-Direct-to-MS are showcased in a data set acquired in a marine metabolomics study reported in MetaboLights (MTBLS1198) using a transmission mode Direct Analysis in Real Time-Mass Spectrometry (TM-DART-MS)-based method.

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Breeding resilient cultivars with increased tolerance to environmental stress and enhanced resistance to pests and diseases demands pre-breeding efforts that include understanding genetic diversity. This study aimed to evaluate the genetic diversity and population structure of 265 pea accessions. The diversity arrays technology (DArT) genotyping method was employed to identify single-nucleotide polymorphisms (SNPs) and silico markers. After stringent filtering, 6966 SNP and 8,454 silico markers were selected for diversity analysis. Genetic diversity was estimated by grouping accessions based on plant material type, geographic origin, growth habit, and seed color. Generally, diversity estimations obtained using SNPs were similar to those estimated using silico markers. The polymorphism information content (PIC) of the SNP markers ranged from 0.0 to 0.5, with a quarter of them displaying PIC values exceeding 0.4, making them highly informative. Analysis based on plant material type revealed narrow observed heterozygosity (Ho = 0.02-0.03) and expected heterozygosity (He = 0.26-0.31), with landrace accessions exhibiting the highest diversity. Geographic origin-based diversity analysis revealed Ho = 0.02-0.03 and He = 0.22 to 0.30, with European accessions showing the greatest diversity. Moreover, private alleles unique to landrace (4) and European (22) accessions were also identified, which merit further investigation for their potential association with desirable traits. The analysis of molecular variance revealed a highly significant genetic differentiation among accession groups classified by seed color, growth habit, plant material types, and geographic origin (p < 0.01). Principal coordinate analysis and neighbor-joining cluster analysis revealed weak clustering of accessions at different grouping levels. This study underscores the significance of genetic diversity in pea collections, offering valuable insights for targeted breeding and conservation efforts. By leveraging genomic data and exploring untapped genetic resources, pea breeding programs can be fortified to ensure sustainable plant protein production and address future challenges in agriculture.

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OBJECTIVES: We describe an artwork observation training program proposed to the first-year psychiatry residents in the school of medicine of Montpellier-Nîmes (Université de Montpellier). This course aimed at improving the ability to observe, tolerate ambiguity in healthcare, work as a team, communicate, and develop empathy. We also evaluated the impact of this program on the enhancement of emotional and communication skills in a pilot study. MATERIALS AND METHODS: The artwork observation program based on Visual Thinking Strategies consisted of three sessions of ninety minutes involving two groups of eight medical students. First-year volunteer psychiatry residents were recruited. Before the sessions participants completed the Groningen Reflection ability Scale to evaluate reflection skills. Then, all the participants examined paintings and drawings of the Atger collection, were encouraged to observe carefully, and to engage in active group discussion by means of a structured questioning. Following the session, participants responded to a short survey to assess potential improvement of socio-emotional and communication skills. RESULTS: Fifteen psychiatry residents participated in the pilot study, eight were males. The median age was twenty-five years. The artwork observation program was well accepted by the participants. All the respondents reported improvement of emotional and communication skills, except regarding three empathy items, including the ability to feel the suffering of others which was increased only for 62% of respondents. DISCUSSION: Our program relying on Visual Thinking Strategies may improve the ability to observe carefully, to perceive the mental states of others, and to tolerate uncertainty. Training of such skills may encourage the adoption of self-reflective attitudes. CONCLUSIONS: Such an artistic innovative program should be extended to undergraduate students in medical curriculum.

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Chickpea (Cicer arietinum L.) is a very important food legume and needs improved drought tolerance for higher seed production in dry environments. The aim of this study was to determine diversity and genetic polymorphism in zinc finger knuckle genes with CCHC domains and their functional analysis for practical improvement of chickpea breeding. Two CaZF-CCHC genes, Ca04468 and Ca07571, were identified as potentially important candidates associated with plant responses to drought and dehydration. To study these genes, various methods were used including Sanger sequencing, DArT (Diversity array technology) and molecular markers for plant genotyping, gene expression analysis using RT-qPCR, and associations with seed-related traits in chickpea plants grown in field trials. These genes were studied for genetic polymorphism among a set of chickpea accessions, and one SNP was selected for further study from four identified SNPs between the promoter regions of each of the two genes. Molecular markers were developed for the SNP and verified using the ASQ and CAPS methods. Genotyping of parents and selected breeding lines from two hybrid populations, and SNP positions on chromosomes with haplotype identification, were confirmed using DArT microarray analysis. Differential expression profiles were identified in the parents and the hybrid populations under gradual drought and rapid dehydration. The SNP-based genotypes were differentially associated with seed weight per plant but not with 100 seed weight. The two developed and verified SNP molecular markers for both genes, Ca04468 and Ca07571, respectively, could be used for marker-assisted selection in novel chickpea cultivars with improved tolerance to drought and dehydration.

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PURPOSE: Diffusing alpha-emitters radiation therapy (DaRT) is a brachytherapy technique using α-particles to treat solid tumours. The high linear energy transfer (LET) and short range of α-particles make them good candidates for the targeted treatment of cancer. Treatment planning of DaRT requires a good understanding of the dose from α-particles and the other particles released in the 224Ra decay chain. METHODS: The Geant4 Monte Carlo toolkit has been used to simulate a DaRT seed to better understand the dose contribution from all particles and simulate the DNA damage due to this treatment. RESULTS: Close to the seed α-particles deliver the majority of dose, however at radial distances greater than 4 mm, the contribution of ß-particles is greater. The RBE has been estimated as a function of number of double strand breaks (DSBs) and complex DSBs. A maximum seed spacing of 5.5 mm and 6.5 mm was found to deliver at least 20 Gy RBE weighted dose between the seeds for RBEDSB and RBEcDSB respectively. CONCLUSIONS: The DNA damage changes with radial distance from the seed and has been found to become less complex with distance, which is potentially easier for the cell to repair. Close to the seed α-particles contribute the majority of dose, however the contribution from other particles cannot be neglected and may influence the choice of seed spacing.

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In this chapter, we will first consider the overall goal of nonclinical safety testing during drug development and have a brief overview of its regulatory background. We will then discuss some basic requirements of safety/toxicity testing before concentrating on the safety testing of RNA vaccines and developing a sample RNA vaccine safety testing program.

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BACKGROUND: Together with other elevated areas, the Mountains of Central Asia are significantly threatened by ongoing climate change. The presence of refuges during the glaciations makes the region extremely rich in species, especially endemic ones. However, the limited potential for colonisation of other habitats makes rocky-related species with 'island-like' distribution, particularly vulnerable to climate change. To understand the processes underlying species response to climate warming, we assessed differences in ecological niches and phylogenetic relationship of two geographically disjunctive alpine species belonging to the genus Sergia. The taxa are considered Tertiary relicts, endemic to the Tian Shan and Pamir-Alai Mountains. To illustrate range dynamics and differences in occupied niches of Sergia species, we used Ecological Niche Modelling of current and future distribution. Whereas, to reconstruct the phylogenetic relationship within and between Sergia and other related Campanulaceae species from the region we used molecular data (ITS, cpDNA, DArTseq-derived SNPs). RESULTS: The results reveal that the genus Sergia is a polyphyletic group, and its representatives differ geographically, ecologically and genetically. Both S. regelii and S. sewerzowii constitute a common clade with Asyneuma group, however, S. sewerzowii is more closely related to Campanula alberti (a species that has never previously been considered closely related to the genus Asyneuma or Sergia) than to S. regelii. Sergia sewerzowii is adapted to lower elevations with higher temperatures, while S. regelii prefers higher elevations with lower temperatures. The future distribution models demonstrate a dramatic loss of S. regelii range with a shift to suitable habitats in higher elevations, while the potential range of S. sewerzowii increases and shifts to the north. CONCLUSIONS: This study shows that S. regelii and S. sewerzowii have a long and independent evolution history. Sergia regelii and S. sewerzowii significantly differ in realised niches. These differences are mirrored in the response of the studied endemics to future climate warming. As suitable habitats shrink, rapid changes in distribution can lead to species' range loss, which is also directly related to declines in genetic variability. The outcomes of this paper will help to more precisely assess the impact of climate changes on rocky-related plant species found in this world's biodiversity hotspot.

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