Search Results (47)

Porlieria chilensis Johnst. (guayacán), an endemic Chilean species native to the sclerophyllous forests, is experiencing a significant population decline. Typically growing as a shrub or small tree on sunny slopes and rocky soils, its populations have been severely impacted by overexploitation and habitat degradation, leading to its classification as Vulnerable (VU). Mature, well-developed individuals have become increasingly rare, now mostly replaced by shrubby regrowth. This study presents the first reported in vitro propagation method for P. chilensis, aimed at establishing a protocol for its mass propagation. A 92.0% survival rate was achieved for nodal segments cultured on Murashige and Skoog (MS) medium after surface disinfection with 1.5% sodium hypochlorite for 15 min. Shoot elongation was successfully stimulated by supplementing the medium with 8.88 μM of 6-benzylaminopurine (BAP), resulting in an average shoot length of 6.9 cm. For ex vitro rooting, plants were transferred to 200 mL containers filled with a 2:1 (v/v) peat/perlite substrate. High rooting rates were obtained (87.5% and 93.1%) when treated with 1968 μM and 1476 μM of indole-3-butyric acid (IBA), respectively. Moreover, 89.8% of the plants survived acclimatization under controlled conditions. These results highlight the strong potential of in vitro propagation as an effective strategy for the conservation and restoration of P. chilensis populations. Full article

Synthetic seed technology is an innovative in vitro technique that provides improved storage capabilities for vegetative propagules. Its success mostly depends on the encapsulation matrix’s composition and the encapsulation procedure. The present study focuses on optimizing an encapsulation protocol for short-term storage and germplasm exchange using micro-cuttings of Salix tetrasperma. Among the different synthetic seed types evaluated, double-layered synthetic seeds (DLSs) exhibited the highest re-growth (93.6%) on MS medium supplemented with meta-Topolin (mT) (5.0 µM) and α-naphthalene acetic acid (NAA) (0.5 µM) after 8 weeks of culture. Viability assessment of non-embryogenic synthetic seeds during low-temperature storage (4 °C) demonstrated the enhanced resilience of double-layered synthetic seeds (DLSs) compared to their single-layered (SLS) counterparts. Following acclimatization in Soilrite^®-filled cups, 82% of the plantlets were successfully established in a greenhouse after four weeks. The increased activity and concentration of antioxidants in DLS-derived plantlets suggest the potential role of the extra layer of alginate in mitigating the effects of low-temperature stress during storage. SCoT molecular analysis confirmed the genetic integrity of the synthetic seed-derived plants. Full article

The objective of this study was an assessment of the anti-biofilm properties of fluoride non-thermal atmospheric plasma (FNTAP) generated using argon and hydrocarbon fluoride gas 1,1,1,2-tetrafluoroethane (TFE). These properties were evaluated by measuring the destruction and recovery of in vitro dual-species biofilms of Streptococcus mutans and Streptococcus sanguinis exposed to FNTAP at 5 or 10 standard cubic centimeters per minute (sccm) or argon non-thermal atmospheric plasma (ArNTAP) for 1 or 2 min, using resazurin-based reagent viability assays, colony forming units (CFU), culture media pH and live/dead staining. Both ArNTAP and FNTAP resulted in significant immediate reductions in bacterial load as compared to the control. Although ArNTAP did not significantly reduce biofilm regrowth, FNTAP treatment showed a bacterial load reduction of more than 5 log units of biofilm regrowth. FNTAP treatments significantly reduced the acidification of the culture medium after recovery incubation, indicating reduced living bacteria, with a pH of 6.92 ± 0.02 and 6.90 ± 0.03, respectively, for the 5 sccm and 10 sccm FNTAP treatments, as compared to a pH of 5.83 ± 0.26 for the ArNTAP treatment, and a significantly acidic pH of 4.76 ± 0.04 for the no-treatment groups. Our results suggest that FNTAP has exceptional anti-biofilm effects, and future directions of our research include the assessment of potential applications of FNTAP in clinical settings. Full article

Background/Objectives: Non-replicating persisters (NRPs) of Mycobacterium abscessus are a bacterial subpopulation that can survive in the host under unfavorable conditions, such as hypoxia or nutrient starvation. The eradication of these bacteria is difficult, which is one reason for the long treatment duration and treatment failure. The drug discovery process should therefore contain methods to screen activity against NRPs. Methods: A hypoxic environment is used to generate NRPs of M. abscessus that are termed low-oxygen persisters (LOPs). For this, an oxidation process is used to transition a replicating culture of M. abscessus distributed in microtiter plates within a sealable box into LOPs. Colony counting, automated object counting, bactericidal activity determination of known agents, and confocal laser scanning microscopy are used to study the obtained culture. Results: The obtained culture shows typical attributes of non-replicating cells, such as significantly reduced replication, the reversibility of the LOP state under aerobic conditions, delayed regrowth on solid medium, altered morphological patterns on a single-cell level, and phenotypical resistance against a variety of clinically relevant antimycobacterial compounds. The study reveals metronidazole and niclosamide as bactericidal against M. abscessus LOPs. These compounds can be used as LOP verification compounds within the described model. Conclusions: Our model is easily implemented and quickly identifies compounds that are inactive under hypoxic conditions. It can therefore accelerate the identification of clinically effective antimycobacterial drug substances, and can be a helpful tool during the drug development process. Full article

Assessing vegetation changes in alpine arid and fragile ecosystems is imperative for informed ecological restoration initiatives and adaptive ecosystem management. Previous studies primarily employed the Normalized Difference Vegetation Index (NDVI) to reveal vegetation dynamics, ignoring the spatial heterogeneity alterations caused by bare soil. In this study, we used a comprehensive analysis of NDVI and its spatial heterogeneity to examine the vegetation changes across the Three-River Headwaters Region (TRHR) over the past two decades. A random forest model was used to elucidate the underlying causes of these changes. We found that between 2000 and 2022, 9.4% of the regions exhibited significant changes in both NDVI and its spatial heterogeneity. These regions were categorized into six distinct types of vegetation change: improving conditions (62.1%), regrowing conditions (11.0%), slight degradation (16.2%), medium degradation (8.4%), severe degradation (2.0%), and desertification (0.3%). In comparison with steppe regions, meadows showed a greater proportion of improved conditions and medium degradation, whereas steppes had more instances of regrowth and slight degradation. Climate variables are the dominant factors that caused vegetation changes, with contributions to NDVI and spatial heterogeneity reaching 68.9% and 73.2%, respectively. Temperature is the primary driver of vegetation dynamics across the different types of change, with a more pronounced impact in meadows. In severely degraded steppe and meadow regions, grazing intensity emerged as the predominant driver of NDVI change, with an importance value exceeding 0.50. Notably, as degradation progressed from slight to severe, the significance of this factor correspondingly increased. Our findings can provide effective information for guiding the implementation of ecological restoration projects and the sustainable management of alpine arid ecosystems. Full article

Cryopreservation, storing biological material in liquid nitrogen (LN, −196 °C), offers a valuable option for the long-term conservation of non-orthodox seeds and vegetatively propagated species in the sector of agrobiodiversity and wild flora. Although the large-scale cryobanking of germplasm collections has been increasing worldwide, the wide application of cryopreservation protocols in wild flora is hampered by difficulties in vitro propagation and a lack of universal cryopreservation protocols, among others. This study established a systematic approach to developing an in vitro culture and droplet-vitrification cryopreservation procedure for shoot tips of Scrophularia kakudensis. The standard procedure includes a two-step preculture with 10% sucrose for 31 h and with 17.5% sucrose for 16 h, osmoprotection with loading solution C4-35% (17.5% glycerol + 17.5% sucrose, w/v) for 30 min, cryoprotection with A3-80% (33.3% glycerol + 13.3% dimethyl sulfoxide + 13.3% ethylene glycol + 20.1% sucrose, w/v) at 0 °C for 60 min, and cooling and rewarming using aluminum foil strips. After unloading, a three-step regrowth procedure starting with an ammonium-free medium with growth regulators was essential for developing normal plantlets from cryopreserved shoot tips. Liquid overlay on the gelled medium two weeks after inoculation resulted in vigorous growth during subcultures. Moreover, liquid overlay increased LN regeneration by up to 80%, i.e., 23% higher than no liquid overlay. Full article

Savannas play a key role in estimating emissions. Climate change has impacted the Cerrado savanna carbon balance. We used the burned area product and long-term field inventories on post-fire vegetation regrowth to estimate the impact of the fire on greenhouse gas emissions and net carbon dioxide (CO₂) emissions in the Cerrado savanna between 1985 and 2020. We estimated the immediate emissions from fires, CO₂ emissions by plant mortality, and CO₂ removal from vegetation regrowth. The burned area was 29,433 km²; savanna fires emitted approximately 2,227,964 Gg of CO₂, 85,057 Gg of CO, 3010 Gg of CH₄, 5,103 Gg of NO_x, and 275 Gg of N₂O. We simulated vegetation regrowth according to three fire regime scenarios: extreme (high fire frequency and short fire interval), intermediate (medium fire frequency and medium fire interval), and moderate (low fire frequency and long fire interval). Under the extreme and intermediate scenarios, the vegetation biomass decreased by 2.0 and 0.4% (ton/ha-year), while the biomass increased by 2.1% under a moderate scenario. We converted this biomass into CO₂ and showed that the vegetation regrowth removed 63.5% of the total CO₂ emitted (2,355,426 Gg), indicating that the Cerrado savanna has been a source of CO₂ to the atmosphere. Full article

The modification of fire regimes and their impact on vegetation recovery, soil properties, and fuel structure are current key research areas that attempt to identify the thresholds of vegetation’s susceptibility to wildfires. This study aimed to evaluate the vulnerability of Mediterranean pine forests (Pinus halepensis Mill. and Pinus pinaster Aiton) to wildfires, analyzing two major forest fires that occurred in Yeste (Spain) in 1994 and 2017, affecting over 14,000 and 3200 hectares, respectively. Four recovery regions were identified based on fire severity—calculated using the delta Normalized Burn Ratio (dNBR) index—and recurrence: areas with high severity in 2017 but not in 1994 (UB94-HS17), areas with high severity in 1994 but not in 2017 (HS94-UB17), areas with high severity in both fires (HS94-HS17), and areas unaffected by either fire (UB94-UB17). The analysis focused on examining the recovery patterns of three spectral indices—the Normalized Difference Vegetation Index (NDVI), Normalized Moisture Index (NDMI), and Normalized Burn Ratio (NBR)—using the Google Earth Engine platform from 1990 to 2023. Additionally, the Relative Recovery Indicator (RRI), the Ratio of Eighty Percent (R80P), and the Year-on-Year average (YrYr) metrics were computed to assess the spectral recovery rates by region. These three spectral indices showed similar dynamic responses to fire. However, the Mann–Kendall and unit root statistical tests revealed that the NDVI and NDMI exhibited distinct trends, particularly in areas with recurrence (HS94-HS17). The NDVI outperformed the NBR and NDMI in distinguishing variations among regions. These results suggest accelerated vegetation spectral regrowth in the short term. The Vegetation Recovery Capacity After Fire (VRAF) index showed values from low to moderate, while the Vulnerability to Fire (V2FIRE) index exhibited values from medium to high across all recovery regions. These findings enhance our understanding of how vegetation recovers from fire and how vulnerable it is to fire. Full article

Over 30 years of plant vitrification, droplet vitrification (DV) of in vitro propagules and slow freezing of dormant buds are typical methods of large-scale cryobanking worldwide. One-step sucrose preculture and Plant Vitrification Solution 2 (PVS2) cryoprotection in solution-based vitrification often face unacceptably low regeneration, and the results are on a case-by-case basis depending on the plant species, like a blind test. The absence of a universal protocol applicable across all plant diversity is considered one of the limiting factors. For wild flora, limits of source material available and difficulties in in vitro propagation make it worse to re-optimize the protocol steps for new species. Since cryoprotectant toxicity is the most crucial barrier to the vitrification of organized explants, selecting alternative plant vitrification solutions (PVS) based on the cytotoxicity of cryoprotectants is vital. This review proposes the concept of donor plant vigor (DPV), which refers to the donor plant properties that determine the potential to regenerate normal plantlets under various cryopreservation procedures. DV is a multi-stage procedure with many factors from stage (1) material preparation to (2) pre-liquid nitrogen (pre-LN) (preculture, osmoprotection, cryoprotection), (3) LN (cooling), (4) warming conditions (rewarming, unloading), and (5) regrowth. Since the cytotoxicity of PVS is a primary limiting factor in DV approaches, DPV is crucial for coping with the toxicity of PVS. The DPV is innate and can be maximized with appropriate material preparations, i.e., vigorously growing in subcultures aided by a liquid overlay on top of the gelled medium, selecting proper explants, optimizing the two-step preculture conditions, and media supplements. Developing the DV protocol starts with testing the material with a tentative standard protocol, which includes a two-step preculture (10% sucrose for 31 h and 17.5% sucrose for 16 h), osmoprotection with C4-35%, cryoprotection with A3-80% (60 min at 0 °C), cooling, and rewarming using aluminum foil strips. Using a three-step regrowth initially with ammonium-free regrowth medium, regrowth of shoot tips in one plate following the successive stages of the tentative standard protocol for shoot tips, i.e., fresh, PC, OP, CP (LNC), and LN, is a valuable tool to characterize the sensitivity of the material and to standardize the procedure by tuning the cryoprotection and cytotoxicity of cryoprotectants. A-series PVS (A3-90%, A3-80%, A3-70%) and B-series PVS (PVS3, B5-85%) can be tested based on the DPV. These alternative PVSs have been applied in over 30 pieces of literature with an 8.5~67.3% increase in LN regeneration compared to PVS2 and Plant Vitrification Solution 3 (PVS3) treatments. Using this approach as an alternative to blind condition screening would be influential in broadening the cryopreservation of diverse wild species and problem materials. Full article

Synthetic seed technology and cold storage methods provide genetic uniformity, paving the way for pest- and disease-free plants that are easy to handle. The aim of this study was to develop protocols for the cold storage of nonencapsulated and alginate-capsulated explants of Cannabis sativa L. Axillary shoot tips derived from in vitro-grown plants that were used as explants and stored for up to 9 months at 4 °C in the dark. Somatic seeds were produced in 3% sodium alginate and Murashige and Skoog (MS) medium salt and stored for up to 3 months. After 6 months of cold storage, the highest regrowth of 45% was recorded for the nonencapsulated explants. The recovery of somatic seeds was 90% under the same storage condition after 3 months. Well-developed, regenerated plants from encapsulated explants successfully acclimatized. Full article

Cryopreservation in liquid nitrogen (LN, −196 °C) is a unique option for the long-term conservation of threatened plant species with non-orthodox or limitedly available seeds. In previous studies, a systematic approach was used to develop a droplet-vitrification (DV) cryopreservation protocol for Postemon yatabeanus shoot tips that includes preculture with 10% sucrose, osmoprotection with C4-35%, cryoprotection with A3-80% vitrification solution, and a three-step regrowth starting with the ammonium-free medium. The tricarboxylic acid (TCA) cycle is a crucial component of plant cell metabolism as it is involved in redox state regulation and energy provision. We hypothesized that organic acids (OAs) associated with the TCA and its side reactions indirectly indicate metabolism intensity and oxidative stress development in shoot tips under the cryopreservation procedure. In this study, the contents of 14 OAs were analyzed using gas chromatography–tandem mass spectrometry (GC-MS/MS) in P. yatabeanus shoot tips in a series of treatments including individual steps of the DV procedure, additional stress imposed by non-optimum protocol conditions (no preculture, no osmoprotection, various vitrification solution composition, using vials instead of aluminum foils, etc.) and regrowth on different media with or without ammonium or growth regulators. The possible relation of OA content with the total cryoprotectant (CPA) concentration and shoot tips regeneration percentage was also explored. Regeneration of cryopreserved shoot tips reduced in descending order as follows: standard protocol condition (91%) > non-optimum vitrification solution (ca. 68%) > non-optimum preculture (60–62%) > regrowth medium (40–64%) > no osmoprotection, cryopreservation in vials (28–30%). Five OAs (glycolic, malic, citric, malonic, and lactic) were the most abundant in the fresh (control) shoot tips. The dynamic pattern of OAs during the DV procedure highly correlated (r = 0.951) with the total CPA concentration employed: it gradually increased through the preculture, osmoprotection, and cryoprotection, peaked at cooling/rewarming (6.38-fold above control level), and returned to the fresh control level after 5 days of regrowth (0.89-fold). The contents of four OAs (2-hydroxybutyric, 3-hydroxypropionic, lactic, and glycolic) showed the most significant (10-209-fold) increase at the cooling/rewarming step. Lactic and glycolic acids were the major OAs at cooling/rewarming, accounting for 81% of the total OAs content. The OAs were categorized into three groups based on their dynamics during the cryopreservation protocol, and these groups were differently affected by protocol step modifications. However, there was no straightforward relationship between the dynamics of OAs and shoot tip regeneration. The results suggest that active modulation of OAs metabolism may help shoot tips to cope with osmotic stress and the chemical cytotoxicity\ of CPAs. Further intensive studies are needed to investigate the effect of cryopreservation on cell primarily metabolism and identify oxidative stress-related biomarkers in plant materials. Full article

Since the 1970s, certain areas within the Three-Rivers Headwater Region (TRHR) of China have faced severe land degradation due to the combined effects of climate change and human activities, leading to restricted ecological service functions and hindering the achievement of sustainable development goals (SDGs). Land degradation in the TRHR has received widespread attention. However, the current research mainly focuses on single-dimensional degradation and lacks a comprehensive evaluation of patterns and structures, as well as above-ground and underground assessments. To address this gap, this study employed the SDG indicator 15.3.1 framework, comprehensively considering fragmentation and habitat quality index based on land cover changes, grassland degradation index, and soil water erosion index. These indexes represent the three land degradation pathways of landscape degradation, vegetation degradation, and soil erosion. This study assessed land degradation patterns in the TRHR from 2000 to 2020. Results show that approximately 44.67% of the TRHR experienced land degradation during this period, mainly in meadow-dominated regions. Additionally, 5.64% of the regions experienced the superimposition of two or more land degradation pathways, with the frequent coexistence of soil erosion and grassland degradation, accounting for 4.1% of the affected areas. Landscape degradation affected approximately 2.39% of the regions, characterized by increased grassland fragmentation or habitat quality degradation. In terms of grassland degradation, 22.26% of the regions showed medium degradation, while 7.21% and 5.63% experienced moderate and severe degradation, respectively. Moreover, approximately 13.36% of the region faced a worsening situation of soil erosion. Approximately 55.34% of the study area underwent land improvement, with significant enhancements mainly concentrated in the western and eastern regions. The regrowth of grassland in the western region and the enhancement and homogenization of grassland productivity in the eastern region played pivotal roles in promoting land improvement. This study provides critical insights into the land degradation pattern in the TRHR over the past 20 years, offering valuable references for formulating and implementing measures to protect and construct the ecological security barrier of the plateau. Full article

The objective of this study is to assess the suitability of vitrification cryo-plate (V cryo-plate) and dehydration cryo-plate (D cryo-plate) methods for the long-term conservation of eight autochthonous Prunus domestica L. genotypes originating from the Balkan Peninsula region. In vitro shoot tips were briefly pre-cultured for 1 day at 23 °C in the dark on a medium containing 0.3 M sucrose and then embedded in calcium alginate gel within the wells of the aluminum cryo-plates. In the V cryo-plate protocol, dehydration was carried out at room temperature using the following vitrification solutions: original plant vitrification solution 2 (PVS2) and 90% PVS2 solution (for 20 and 40 min) and plant vitrification solution 3 (PVS3) (for 60 and 80 min). In the D cryo-plate protocol, desiccation was performed for 2, 2.5, or 3 h over silica gel at 23 °C. The effect of different treatments was evaluated by monitoring the regrowth of both non-frozen and cryo-preserved explants. After cryo-preservation, five genotypes achieved regrowth rates over 40% in at least one of the applied protocols, while two genotypes showed regrowth rates of around 10%. A significant improvement in regrowth success for all genotypes using both cryo-plate methods was achieved by pre-culturing shoot tips for 7 days on a medium containing 0.5 M sucrose in complete darkness at 4 °C. Shoots regenerated from cryo-preserved explants were further monitored in vitro. By the third subculture, they had not only regained but had even exceeded the multiplication capacity (index of multiplication, length of axial, and lateral shoots) of shoots regenerated from dissection controls. Following multiplication, the cryo-preserved shoots were successfully rooted and rooting ability was assessed by monitoring the percentage of rooting, number and length of roots, and height of rooted plantlets. Full article

Biofuel is encouraged because of its low impact on climate change. A new framework was developed to accurately assess the climate change impacts (CCI) of biofuel by integrating the atmospheric carbon cycle model and vegetation carbon dynamic models. Forests with different growth rates (fast, medium, slow) and three collection intensities (71%, 52%, 32%) of logging residues were presumed to test the performance of this framework. The CCI of biofuel was analyzed under two functional units: 1 GJ of biofuels and 1 ha of forests to supply biofuels. According to this study, increasing the forest growth rate could decrease the CCI in both functional units. Increasing the collection intensity could decrease the CCI of 1 GJ of biofuel but increase the CCI of 1 ha of forest land (unless the impacts were negative in fast-growth forests with high and medium collection intensities). Producing bioethanol resulted in a lower CCI (−3.1–67.7 kg CO₂ eq/GJ) compared to bio-diesel (29.3–94.7 kg CO₂ eq/GJ). Hence, collecting all available logging residues (without inhibiting forest regrowth) to produce low CCI biofuels such as bioethanol was found to be the optimal option for achieving high mitigation effects. Full article

Cryopreservation is considered to be one of the most effective methods for long-term storage of plant genetic resources, particularly for ornamental species. However, there is a very little research on cryopreservation of lilacs. In this study, for the first time the cryopreservation protocol (a variation of a pregrowth-dehydration method) was successfully applied to two cultivars of Syringa vulgaris: ‘Aucubaefolia’ and ‘Polina Osipenko’. Explants of both cultivars were able to withstand the different steps of the protocol, and high survival and regrowth percentages were obtained after exposure to liquid nitrogen (67–100% and 63–88%, respectively). The current study is mainly focused on the preculture conditions of the applied method. Based on our results, we propose the use of paclobutrazol (PBZ) with the combination of 6-benzylaminopurine (BAP) and thidiazuron (TDZ) in the preculture medium for increasing explant tolerance to subsequent dehydration and freezing. During post-LN recovery, the explants appeared morphologically normal, and after 12–16 weeks after thawing, they were propagated and cultured as normal plantlets. Therefore, the reported method is effective for long-term storage of lilac meristems and could be used to create a cryobank of achievements in lilac breeding. Full article