Strawberry ( Fragaria × ananassa ) is an economically important fruit crop worldwide. Although substantial progress has been achieved through conventional breeding, improvement efforts are constrained by long breeding cycles, complex octoploid genetics, extensive heterozygosity, and strong genotype × environment interactions, all of which limit the speed and precision of trait development. Moreover, introgression of desirable traits through conventional hybridization often leads to linkage drag and unpredictable phenotypic outcomes, particularly for disease resistance and fruit quality traits. In contrast, genetic transformation and genome editing enable precise modification of target genes without disrupting elite genetic backgrounds, providing a powerful approach for the timely and targeted introduction of desirable traits. However, the successful application of these technologies relies on robust and reproducible in vitro regeneration platforms, which are essential for recovering edited plants, overcoming genotype-dependent barriers, and accelerating precision breeding in strawberries. In this study, an efficient in vitro regeneration system was developed to support genome-editing applications in strawberries. Optimal regeneration conditions were established for the cultivar Festival using in vitro leaf explants by evaluating thidiazuron (TDZ) and 6-benzylaminopurine (BAP), applied alone or in combination with low concentrations of naphthaleneacetic acid or 2,4-dichlorophenoxyacetic acid, under both light and dark culture conditions. Dark incubation markedly enhanced leaf regeneration efficiency, significantly increasing both regeneration frequency and the number of regenerated shoots compared with light conditions. The genotype dependency of regeneration was further assessed by validating the selected conditions across four additional cultivars (Albion, Chandler, Monterey, and Rocco). Notably, this study represents the first report of successful in vitro regeneration of the strawberry cultivars Monterey and Rocco. Finally, all regenerated shoots were successfully rooted and acclimatized using an optimized regeneration, rooting, and acclimatization protocol.

To investigate the role of hydrogel composition and nanozymes incorporation on the inflammatory response of encapsulated murine microglia. Sodium alginate (SA) hydrogels containing i) hyaluronic acid (HA) of low-100 kDa or high-1 MDa molecular weight and ii) Mn3O4 nanoparticles (Mn3O4-NPs) were prepared and characterized. The inflammatory response of encapsulated microglia was assessed. HA enhanced cell-biomaterial interactions and reduced cell aggregation. Under normal culture conditions, HA induced a pro-inflammatory response, particularly with high-MW HA, with a 7-fold increase in IL-6 expression and a 2.2-fold increase in TNF-α secretion. Under prolonged LPS stimulation, HA promoted an anti-inflammatory shift, with a 2-fold decrease in IL-1β expression and restored IL-4 expression to basal levels. Mn3O4‑NP-loaded hydrogels showed strong H2O2 scavenging capability, protecting cells from oxidative stress. This effect may be partially counterbalanced by •OH generation at high Mn3O4‑NP concentrations. Under short-term LPS stimulation, Mn3O4 promoted a slight pro-inflammatory response, with a 1.3-fold and 1.2-fold increase in IL-1β and TNF-α expression. However, prolonged LPS exposure reflected an anti-inflammatory scenario, with increased IL-4 and a 2.4-fold increase in Arg-1 expression. SA hydrogels are promising carriers for antioxidant nanozymes and microglia, modulating cell response under prolonged inflammation while protecting cells from oxidative stress.

Patient-derived malignant effusion organoids guide tailored therapy and dissect third-generation EGFR-tyrosine kinase inhibitor resistance mechanisms in lung cancer.

Brief guide to Drosophila maintenance and husbandry.

To overcome the depletion of wild resources and limited seed propagation of Valeriana fauriei, an adventitious root suspension culture system was established and optimized, and methyl jasmonate (MeJA)-elicited metabolic responses were then evaluated using gas chromatography-mass spectrometry (GC-MS)-based untargeted metabolomics. Culture conditions were optimized using a single-factor medium screening experiment combined with a uniform design for four culture-condition factors. The results showed that the optimized culture system increased the 28 d proliferation coefficient of Valeriana fauriei adventitious roots to 27.07. A total of 181 significant differential metabolites were identified and classified into four clusters according to their changing trends. The first cluster increased and then decreased, reaching the maximum value at 80 mg·L−1 MeJA. The second cluster decreased and then increased, reaching the minimum value at 80 mg·L−1 MeJA. The third cluster increased, then decreased, and then increased again, reaching the maximum value at 40 mg·L−1 MeJA and the minimum value at 160 mg·L−1 MeJA. The fourth cluster increased continuously with increasing MeJA concentration. Subsequently, KEGG pathway enrichment analysis was performed for the metabolite sets of the four clusters. Combined analysis indicated that 80 mg·L−1 MeJA was the treatment concentration that most strongly affected the metabolic biosynthesis of V. fauriei adventitious roots. Under this treatment, pathways related to membrane transport, amino acid metabolism, translation, nucleotide metabolism, and the biosynthesis of other secondary metabolites were the most significantly enriched.

Biofloc treatment is an eco-friendly and affordable technology which has been adopted over the last three decades to improve aquaculture sustainability. In this study, fingerlings were randomly distributed into different aquariums and fed diets with varying protein levels (15, 20, 25 and 30%). This study investigated the effect of different crude protein (CP) levels on growth, immunity and meat quality of Nile Tilapia (Oreochromis niloticus) under zero exchange heterotrophic culture conditions. Results showed that fish fed 25 and 30% protein diets had the highest growth rate, feed efficiency and protein efficiency ratio. Weigh gain (WG) and specific growth rate (SGR) significantly increase in fish fed 25 and 30 % CP levels (p<0.05). Proximate analysis showed that the fish fed 25 and 30% CP had the highest protein content, lowest lipid content and improved fatty acid profile. Additionally, fish fed 30% protein had the lowest mortality rate and highest survival rate. Immune parameters (WBCs, plasma parameters, lysozyme and antioxidants enzymes activity) were significantly enhanced in fish fed 25 and 30% CP (p<0.05). The best growth performance was obtained when the fish fed on the 25 and 30% CP in diet and can be used as sustainable production of fish under zero exchange heterotrophic culture system.

Portunid crabs, including the genus Scylla and genus Portunus, are highly commercial due to their tasty flavour and high meat yield. However, until now, crab demand is still heavily dependent on wild-caught crabs, and the domestication of crabs is necessary for the production of viable seed in hatcheries and the grow-out of portunid crabs. To enhance portunid crabs farming, reproductive technologies should be improved, and this information still needs to be compiled and reviewed. As such, the purpose of this paper is to review reproductive technologies for sustainable portunid crab’s aquaculture. This manuscript provides a narrative review of reproduction technology, with an emphasis on the portunid crab from two genera, Scylla and Portunus, covering literature from 2000 up to 2025, which focuses on practical hatchery applications. Three methods, including hormonal injections, dietary feeding, and manipulation of culture conditions, can be implemented in hatcheries to obtain as many spawned crabs as possible. Pregnenolone, 5-HT, GnRH, methyl farnesoate, prostaglandin, thyroxine and estrogen have been proven to support ovarian maturation in portunid crabs. Formulated diet rich in lipids, adding astaxanthin or using natural diet from fish, blood cockle and squid has been shown to stimulate gonad development and increase spawning rates. Manipulating temperature and photoperiod can also lead to improvements in the egg incubation period and hatching success, with higher water temperatures and longer light periods yielding better results. Through this review, knowledge on reproductive technologies has been gathered and could be further implemented during the farming of portunid crabs, making hatchery-based reproduction more reliable.

Unraveling the potential and challenges of photosynthetic microalgae for oxygenating engineered tissues.

This study explored the biotechnological production of lactobionic acid through the oxidation of whey, a by-product generated during milk and dairy processing, using the bacterium Gluconobacter frateurii. The bacterial strain used in the experiments was isolated from rotten apples. Identification was carried out by PCR amplification of the 16S rRNA gene, followed by nucleotide sequencing. Comparison of the obtained sequence with those in the NCBI GenBank database confirmed, with 99% similarity, that the isolated microorganism was Gluconobacter frateurii. The identified strain was cultivated in liquid whey-based media to assess its capacity for lactobionic acid production. Based on bacterial growth and cell count analysis, whey derived from industrial acid-treated curd was determined to be the most suitable substrate. The bacterium Gluconobacter frateurii was grown in pretreated whey medium, and lactose concentrations were monitored at 24, 36, 48, 60, 72, 84, 98, 115, and 154 hours after the onset of fermentation. The lowest lactose concentration, 1.18 g/L, was observed after 115 hours, indicating that a fermentation period of approximately four days is sufficient under liquid culture conditions. Following the completion of fermentation, the formation of lactobionic acid was confirmed using thin-layer chromatography (TLC). The product was then isolated through recrystallization, achieving a lactose conversion yield of 74%. The purified lactobionic acid was further analyzed to determine its physicochemical properties and biological activity. This research demonstrates that whey, commonly regarded as a waste product of dairy manufacturing, can be effectively utilized through biotechnological methods to produce lactobionic acid, a high-value compound with important applications in the pharmaceutical and cosmetic industries.

Alpha-linolenic acid (ALA) is an essential omega-3 fatty acid and a vital component in food applications. In this study, we investigated a range of physicochemical culture conditions—including pH, temperature, and carbon source—to evaluate biomass and ALA accumulation in Chlorella sp. L166 and its mutant, C-12. The study aimed to identify favorable culture conditions and evaluate the feasibility of using diluted bean curd (tofu) wastewater as a low-cost medium. Under mixotrophic cultivation, ALA content was determined via GC-MS, and the removal efficiencies of total nitrogen (TN), total phosphorus (TP), and chemical oxygen demand (COD) were simultaneously monitored. The results showed that L166 achieved its highest ALA accumulation at pH 6.0 and 23 °C with maltose. C-12 exhibited appropriate ALA accumulation at pH 7.0 and 23 °C with maltose and reached its maximum biomass at pH 8.0 and 25 °C with glucose. After 8 days of cultivation in threefold-diluted tofu wastewater, C-12’s ALA content reached 6.1 mg/g, significantly higher than that observed in BG11 medium. Meanwhile, both strains removed 81.2–83.2% of TN, 35.7–36.0% of TP, and 42.6–43.5% of COD. This study provides preliminary data on the effects of culture conditions on microalgal ALA production, highlighting the potential for future practical applications of C-12.

Testicular steroidogenesis is fundamental to male reproductive health, but its disruption by environmental and emerging chemicals remains insufficiently characterized due to limitations in existing test systems. Traditional animal models pose ethical and logistical challenges, while the validated H295R assay-based on a female adrenal carcinoma cell line-fails to reflect male gonadal steroidogenesis. This review uses a semi-systematic approach to evaluate over 1500 studies employing in vitro models, including primary Leydig cells, Leydig cell lines, stem cell-derived Leydig-like cells, and advanced 3D testicular systems. We assess species origin, developmental relevance, culture conditions, and the extent to which these models replicate key steroidogenic pathways. Most models rely on rodent-derived, cancerous cell lines cultured in two-dimensional monolayers, with limited representation of human and immature Leydig cells. A targeted full-text analysis examined the effects of 23 reference chemicals on testosterone, progesterone, androstenedione, and estrogen levels across the H295R assay and eight testicular in vitro models. Forskolin, genistein, prochloraz, and ketoconazole showed consistent effects and may serve as promising reference compounds. However, data for most chemicals in testicular models are scarce or inconsistent-particularly for androstenedione and progesterone-underscoring the need for improved model standardization. We propose future directions to enhance predictive power, including the development of hormone-responsive, species- and stage-specific models cultured under hormone-controlled conditions. Such advances are essential to improve chemical safety assessment and facilitate regulatory acceptance of alternative test methods.

Generation of human hematopoietic stem and progenitor cells (HSPCs) from pluripotent stem cells (PSCs) holds significant promise in disease modeling, drug screening, and the development of cell and gene therapies for various hematologic and nonhematologic disorders. However, efficient and consistent derivation of functional HSPCs remains a major challenge hindering their practical use in biomedical research and applications. One of the contributing factors is the use of human serum albumin (HSA), which has been widely regarded as essential for supporting ex vivo maintenance, expansion, and differentiation of human stem cells and their derivatives. The lot-to-lot variation of this critical reagent can contribute to inconsistent laboratory results and high manufacturing costs. In this study, we address the manufacturing and regulatory challenges associated with using HSA in HSPC differentiation and propose an approach using a caprolactam-based polymer as an alternative. Our differentiation method demonstrates robust HSPC generation across various embryonic and induced PSC lines and culture conditions, highlighting its adaptability and reliability. Importantly, PSC-derived HSPCs exhibit functional versatility in differentiation capacity into myeloid and lymphoid lineages, as validated through colony-forming assays as well as directed red blood cell and NK cell differentiations. These findings suggest that HSA is dispensable in HSPC differentiation and replacing it with synthetic polymers has the potential to mitigate lot-to-lot reagent variation, improve HSPC production consistency, lower manufacturing costs, and expedite clinical applications.

Therapeutic applications require large amounts of extracellular vesicles (EVs) that cannot be obtained by standard laboratory protocols. Since culturing parameters and isolation methods can significantly affect the molecular composition and therapeutic efficacy of EVs, the development of a new scale-up protocol should be followed by the molecular fingerprinting and validation of therapeutic potential in vivo. We developed a new scale-up protocol based on microcarrier culture (3D) of immortalized human dental pulp stem cells in a spinning bioreactor and subsequent isolation of EVs by 2-step tangential flow filtration (TFF) and size exclusion chromatography (SEC). A new scale-up protocol increased EV yields by 463-fold. When compared with ultracentrifugation (UC), isolation using TFF/SEC substantially reduced the complexity of proteomic cargo, whereas culture conditions (2D vs. 3D) affected miRNA, but not mRNA and proteomic content of the EVs. We next compared the therapeutic efficacy of both EV products in 6-hydroxydopamine rat model of Parkinson's disease (PD). The same amounts of EVs derived from standard 2D cultures by UC and a new large-scale protocol were intranasally administered to PD rats, where they similarly improved gait and cognitive functions, preserved nigrostriatal tyrosine hydroxylase density and suppressed neuroinflammation. Notably, both EV preparations were enriched in proteins and miRNAs associated with anti-oxidative and anti-inflammatory responses. Our protocol allows large-scale production of EVs that are therapeutically effective in the pre-clinical model of PD.

Fungi represent a prolific source of structurally diverse secondary metabolites, yet the extent to which culture conditions reshape the metabolic profile and functional bioactivity remains incompletely understood. In this exploratory study, ten fungal strains belonging to genera Penicillium and Aspergillus were cultivated in Yeast Extract Sucrose (YES) and Czapek Yeast Autolysate (CYA) media and analysed using untargeted LC-HRMS metabolomics. The objective of this study was to evaluate how culture medium influences metabolic profiles and to investigate medium-dependent metabolic variation and its relation to cytotoxic, antibacterial, and antifungal activities. Global metabolic profiling revealed moderate but statistically significant medium-associated metabolite variation, with discriminant metabolites predominantly enriched under CYA conditions. Putative structural annotation suggested patterns consistent with differential regulation of isoprenoid-derived sterols, terpenoids, alkaloid-like metabolites, and aromatic polyketides. While antimicrobial activities displayed a heterogeneous, strain-dependent pattern with limited correlation to individual metabolites, cytotoxic activity co-varied with metabolite composition in OPLS regression modelling. Sterols and terpenoid-related features emerged as major contributors to cytotoxicity. Given the absence of biological replication and the limited sample size inherent to this pilot study, all findings should be considered hypothesis-generating and interpreted within an exploratory framework. These results suggest that nutrient composition influences biosynthetic pathway activation while functional outcomes remain strongly dependent on strain-specific metabolic capacity. This work provides a systematic framework and targeted hypothesis for future investigations into condition-dependent fungal chemical diversity in natural product discovery.

Adipose-derived mesenchymal stem cells (ADSCs) have emerged as an important cell source in regenerative medicine because of their accessibility, abundance, multilineage differentiation potential, and paracrine activity. However, ADSCs are not biologically uniform, and their properties are strongly influenced by donor-related factors, anatomical origin of adipose tissue, and technical procedures used for cell isolation, expansion, and characterization. This review summarizes current advances in defining the source-dependent characteristics of ADSCs, with particular emphasis on donor age, metabolic status, adipose depot specificity, isolation methods, culture conditions, and source-related molecular and functional heterogeneity. Evidence from transcriptomic, epigenetic, immunophenotypic, and secretome studies indicates that ADSCs from different sources may differ substantially in proliferation and differentiation capacity, immunomodulatory activity, and therapeutic performance. Major challenges remain in translating these findings into clinical practice, including donor variability, inconsistent manufacturing workflows, lack of standardized potency assays, and insufficient integration of source-stratified strategies into product development. Emerging directions such as single-cell and multi-omics profiling, cell-free secretome-based therapeutics, and source-aware manufacturing frameworks may improve precision and reproducibility in ADSC-based therapies. A clearer understanding of ADSC source dependency will be essential for optimizing donor selection, improving product consistency, and advancing the safe and effective clinical translation of regenerative medicine applications.

Optimization of biotechnological processes is traditionally limited by time-consuming trial-and-error approaches and the complexity of simultaneously optimizing multiple, often conflicting objectives. This applies particularly to plant tissue culture medium design, which therefore serves as the application case in this study. Recent advances in machine learning and evolutionary algorithms offer powerful alternatives, yet 80% of published studies rely on licensed software, and systematic data-driven optimization frameworks remain scarce. This creates significant barriers to adoption in both academic and commercial plant biotechnology. We introduce ADAM (Advanced Design and AI-Driven Modeling for Plant Tissue Culture Media Optimization), an open-access, web-based platform that transforms protocol development into a data-driven computational process. ADAM implements a complete ML-EA workflow through five integrated modules: 1. Design of Experiments (five different concepts) for systematic parameter exploration, 2. Data Preparation with automated quality control, and 3. Model Building using nine machine learning algorithms with automated selection. The platform enables Optimization (4.) through four advanced evolutionary algorithms (genetic algorithm, particle swarm optimization, NSGA-II, SMS-EMOA) for single- and multi-objective problems, with Evaluation (5.) tools to compare original versus optimized solutions. Validation across two plant tissue culture applications showed that ADAM's models matched or exceeded the predictive performance of manually optimized approaches in the original studies. The platform successfully identified multiple optimal culture conditions balancing conflicting objectives, providing experimentally testable predictions that reduce the trial-and-error cycle. Deployed as a browser-based application requiring neither specialized hardware nor software licenses, ADAM democratizes advanced AI optimization for plant biotechnology, eliminating traditional barriers to entry while maintaining the rigor and flexibility required for scientific research.

Bu araştırma, 3 adet yerli ve 60 adet yabancı orijinli Achillea millefolium L. genotiplerinin Isparta ekolojik koşullarında verim ve kalite özelliklerini belirlemek amacıyla 2023 yılında yürütülmüştür. Çalışmada çiçeklenme süresi 10–41 gün, bitki boyu 21.8–72.0 cm, çiçek çapı 4.3–15.5 cm ve çiçekçik sayısı 26.0–419.0 adet/kapitulum arasındadır. Taze çiçek verimi 246.9–2829.7 kg/da, kuru çiçek verimi ise 82.6–1017.7 kg/da aralığında ölçülmüş ve PI 439888, PI 661151 ve PI 439892 genotipleri yüksek verim potansiyeliyle dikkat çekmiştir. Uçucu yağ oranları %0.05-0.50 arasında değişmiştir. Taze herba ve uçucu yağ verimi ile uçucu yağ oranı ve rengine göre seçilen 10 genotip ve yerli genotiplerde; 1,8-sineol (%3.1–22.4), borneol (%2.5–18.3), kamfen (%1.8–12.7), α-pinen (%4.2–28.9), β-pinen (%3.7–19.5), sabinen (%1.2–9.8), mirsen (%0.9–7.4), β-karyofilen (%2.1–10.6) ve karyofilen oksit (%1.5–8.3) ana bileşenler olarak belirlenmiştir. Sonuçlar, A. millefolium genotiplerinin kültür koşullarında başarıyla yetiştirilebildiğini ve doğal popülasyonlardan kültüre alınan materyalin sürdürülebilir ve ekonomik bir üretim potansiyeline sahip olduğunu göstermektedir. Bu çalışma, civanperçeminin endüstriyel kullanımı, kalite standardizasyonu ve gelecekte yürütülecek adaptasyon ve ıslah çalışmalarına bilimsel temel oluşturmaktadır.

Spermatogenesis relies on the highly specialized interaction between germ cells and a supportive somatic niche, yet replicating this environment in vitro remains a major challenge. Primary human Sertoli cells (hSCs), key architects of this niche, often lose their phenotype under conventional culture conditions, limiting the establishment of a stable blood-testis barrier (BTB)-associated phenotype and their ability to support germ cells. Here, we present a structurally organized, human ECM-derived connective tissue equivalent (CTE) designed to support long-term maintenance and organization of hSCs in vitro. The CTE, generated from fibroblast-secreted matrix enriched in laminin, fibronectin, and collagen IV, reproduces key biochemical and physical features of a supportive microenvironment. hSCs introduced into the CTE as single cells or pre-formed spheroids were evaluated for survival, structural organization, phenotypic stability, and ECM remodeling. Both configurations supported progressive expression and organization of BTB-associated proteins (ZO-1, OCLDN) together with upregulation of Sertoli cell-associated markers, including SOX9 and ABP, with the spheroid-based model showing improved structural cohesion, integration within the construct, and more evident junctional organization over time. Overall, this bioactive human-derived platform supports long-term maintenance of hSC phenotype and barrier-associated features in vitro, providing a promising basis for future human testis models and co-culture studies.

Influence of culture conditions and dietary regimes on survival, population dynamics and naupliar production of the copepod Paramphiascella fulvofasciata (Rosenfield & Coull, 1974)

Actinomycetes associated with plants are those that live on the surface, inside, or outside the plants, and have a reciprocal relationship with the plants. These actinomycetes have recently been found to possess many beneficial activities. The Northwest region of Vietnam is home to various valuable medicinal plants, of which the health-healing compounds, the microorganisms, particularly actinomycetes, are a promising research subject. In this study, we screened 136 endophytic and surface-associated actinomycetous strains isolated from 17 medicinal plants collected in the Northwest region of Vietnam for their antimicrobial activities against Escherichia coli, Bacillus subtilis, Candida albicans, Aspergillus fumigatus, and Aspergillus flavus. The results showed that 5 strains were antagonistic to Escherichia coli, 22 strains were antagonistic to Bacillus subtilis, 3 strains were antagonistic to all the three test fungi, 6 strains were antagonistic to more than one test microorganism, and 2 strains were antagonistic to all test microorganisms. Three actinomycetous strains (AL12.3, AT4.1, and AS10.10) with outstanding activities were identified as Streptomyces spp. The optimal conditions for their growth were determined to be as follows: ISP4 medium with potato extract, temperature at 30 °C, culture duration of 6-7 days for strain AL12.3, and 3-4 days for strains AT4.1 and AS10.10. A preliminary experiment in this study showed that when cultured on a medium supplemented with the aqueous extract of its cognate plant, strain AT4.1 displayed an enhanced antibacterial activity against E. coli and B. subtilis, compared to culture conditions without the extract.