Applied Biosciences

Flow cytometry is widely applied to infer the ploidy and genome size (GS) of plant nuclei. The conventional approach of sample preparation, reliant on fresh plant material to release intact nuclei, often results in poor yields of nuclei in conditions when a plant material cannot be kept fresh due to logistical constraints. Previous attempts to use frozen plant material were mainly limited to ploidy analysis and relied on chopping methods, which restrict the material input and often result in poor nuclei yield, especially in frozen samples, due to incomplete disruption. Here, we present a modified protocol for GS estimation using frozen plant material that facilitates larger volumes of tissue to be processed while improving debris removal. Nuclei isolated from this protocol can also be used for DNA or RNA extraction. Genome size estimates from frozen material are similar to those from fresh material, with a reduction in error range, although not always significant (p > 0.05). In certain species, frozen samples can yield substantially more nuclei than fresh material. With the addition of specific debris compensation algorithms, coefficient of variation (CV%) can be maintained below 5%. This method has special value in estimating the GS of samples collected from remote locations and frozen for use in plant genome sequencing. Freezing preserves high-quality DNA and RNA, enabling the same sample to be used for both flow cytometry and genome sequencing. Full article

An aqueous leaf extract of Egeria densa was used to green-synthesize iron (II) and iron (III) oxide nanoparticles from ferrous sulphate and ferric chloride, respectively. The successful green synthesis of the nanoparticles was confirmed through UV–visible spectroscopy, and the colour of the mixtures changed from light-yellow to green-black and reddish-brown for FeO–NPs and Fe₂O₃–NPs, respectively. The morphological characteristics of the nanoparticles were determined using an X-ray diffractometer (XRD), a Fourier transform infrared spectrophotometer (FTIR), a transmission electron microscope (TEM), and energy-dispersive X-ray spectroscopy (EDX). The UV–Vis spectrum of the FeO–NPs showed a sharp peak at 290 nm due to the surface plasmon resonance, while that of the Fe₂O₃–NPs showed a sharp peak at 300 nm. TEM analysis revealed that the FeO–NPs were oval to hexagonal in shape and were clustered together with an average size of 18.49 nm, while the Fe₂O₃-NPs were also oval to hexagonal in shape, but some were irregularly shaped, and they clustered together with an average size of 27.96 nm. EDX analysis showed the presence of elemental iron and oxygen in both types of nanoparticles, indicating that these nanoparticles were essentially present in oxide form. The XRD patterns of both the FeO–NPs and Fe₂O₃–NPs depicted that the nanoparticles produced were crystalline in nature and exhibited the rhombohedral crystal structure of hematite. The FT-IR spectra revealed that phenolic compounds were present on the surface of the nanoparticles and were responsible for reducing the iron salts into FeO–NPs and Fe₂O₃–NPs. Conclusively, this work demonstrated for the first time the ability of Elodea aqueous extract to synthesize iron-based nanoparticles from both iron (II) and iron (III) salts, highlighting its versatility as a green reducing and stabilizing agent. The dual-path synthesis approach provides new insights into the influence of the precursor oxidation state on nanoparticle formation, thereby expanding our understanding of plant-mediated nanoparticle production and offering a sustainable route for the fabrication of diverse iron oxide nanostructures. Furthermore, it provides a simple, cost-effective, and environmentally friendly method for the synthesis of the FeO–NPs and Fe₂O₃–NPs using Egeria densa. Full article

This review examines the history of consumption, life cycle, and culture conditions of seven edible mucilaginous terrestrial cyanobacterial strains—Nostoc flagelliforme, Nostoc commune, Nostoc sphaeroides, Nostoc sphaericum, Nostoc verrucosum, Aphanothece sacrum, and Nostochopsis lobatus—as resilient and sustainable food sources in the face of climate change. Traditionally consumed across various cultures and known for their resilience in extreme environments, these cyanobacteria offer high nutritional value, including proteins, vitamins, and essential fatty acids, making them promising candidates for addressing food security. Their ability to fix nitrogen reduces reliance on synthetic fertilizers, enhancing agricultural applications by improving soil fertility and minimizing dependence on fossil fuel-derived chemicals. Unlike conventional crops, these cyanobacteria require minimal resources and do not compete for arable land, positioning them as ideal candidates for low-impact food production. Despite these advantages, the review highlights the need for scalable and cost-effective cultivation methods to fully realize their potential in supporting a resilient global food supply. Additionally, it underscores the importance of ensuring their safety for consumption, particularly regarding toxin content. Full article

Carnitine palmitoyltransferase 1 (CPT1), which catalyzes the rate-limiting step of fatty acid oxidation, has been implicated in therapeutic approaches to several human diseases characterized by aberrant lipid metabolism. The isoform-specific quantification of CPT1 activity is essential in the characterization of small molecule inhibitors of CPT1, but several existing means to quantify enzymatic activity, including the use of radioisotope-labeled carnitine, are not amenable to scalable, high throughput screening. Here, we demonstrate that mitochondrial extracts from Expi293 cells transfected with a CPT1a plasmid are a reliable and robust source of catalytically active human CPT1. Moreover, with a source of catalytically active enzyme in hand, we modified a previously reported colorimetric method of coenzyme A (CoA) easily scalable to a 96-well format for the screening of CPT1a inhibitors. This assay platform was validated by two previously reported inhibitors of CPT1a: R-etomoxir and perhexiline. To further demonstrate the applicability of this method in small molecule screening, we prepared and screened a library of 87 known small molecule APIs, validating the inhibitory effect of chlorpromazine on CPT1. Full article

Prostate cancer (PCa) is one of the most common malignancies in men, where early and accurate detection is crucial. While PSA testing has been the diagnostic standard, its limited specificity leads to unnecessary biopsies and missed significant cancers. Urinary biomarkers such as PCA3 and TMPRSS2-ERG and multi-marker assays (MyProstateScore, SelectMDx, and ExoDx) offer a promising alternative. This narrative review examines their diagnostic performance and clinical utility with the aim of understanding whether they can be integrated with the established tests and exams already in use. A literature search of PubMed, Scopus, and Medline identified some relevant recent studies (2010–2025). The findings show that PCA3 and TMPRSS2-ERG improve specificity over PSA, while multi-marker tests enhance risk stratification and reduce unnecessary procedures. MPS integrates urinary biomarkers with PSA, achieving over 95% sensitivity and negative predictive value for clinically significant cancers. SelectMDx demonstrates ~90% negative predictive value, and ExoDx assesses urinary exosomes to predict aggressive disease. Despite their advantages, challenges persist, including variability in performance, cost, and accessibility. Urinary biomarkers represent a major step toward more precise, less invasive diagnostics, with future research needed to optimize clinical integration and cost-effectiveness. Full article

Experimental research demands the selection of appropriate models to align with study objectives and conditions. Traditional experimental models, such as in vivo animal studies and in vitro systems like organoids, present nutraceutical and pharmaceutical research limitations such as high cost, ethical concerns, long lifespan, and difficult genetic manipulation. Caenorhabditis elegans has proved to be a valuable model as a result of its genetic and physiological similarities to higher organisms, fully sequenced genome, short life cycle, and transparency. These features enable high-throughput screening, molecular pathway analysis, and lifespan and healthspan assays. C. elegans has significantly advanced the discovery of bioactive molecules with therapeutic potential, shedding light on aging, neurodegeneration, metabolic disorders, and immune responses. Its utility in pharmacokinetics and validation of nutraceuticals underscores its role in longevity and metabolic health research. Additionally, its conserved stress response, apoptosis, and pathogen recognition pathways facilitate the study of pharmacological interventions for inflammation, oxidative stress, and infections. This study evaluates the applicability of C. elegans as a model for in vivo screening, analyses its role in drug efficacy testing, and discusses relevant advancements, associated difficulties, and what to expect of C. elegans in research. Full article

Common beans are a vital source of protein, vitamins, and minerals. Increasing common beans productivity is crucial for improving food security and farmers’ incomes globally. This study evaluated the growth and yield responses of common beans to integrated organic and inorganic fertilizers under field conditions at the Faculty of Agriculture, Kabul University. The trial was repeated over two consecutive growing seasons in 2020 and 2021, using a randomized complete block design with 18 treatments and three replications. The fertilizers used included urea (N) (0, 60, and 90 kg/ha), diammonium phosphate (D) (0, 50, and 100 kg/ha), and farmyard manure (O) (0 and 5000 kg/ha). The results show that integrated fertilizers, particularly O5000N60D50, O5000N60D100, O5000N90D50, and O5000N90D100, significantly increased growth and yield parameters. In 2020, the grain yield increased significantly (p < 0.05) by 75.6, 76.7, and 68.4% with the O5000N60D50, O5000N60D100, and O5000N90D100 treatments, respectively. In 2021, O5000N60D50, O5000N60D100, and O5000N90D50 showed significant yield increases of 94.7, 89.6, and 97.9%, respectively. The grain yield strongly correlated with the SPAD value (r = 0.84), number of pods per plant (r = 0.71), and number of seeds per pod (r = 0.66) in 2020, and it more strongly correlated with the SPAD value (r = 0.91), number of pods per plant (r = 0.77), and number of seeds per pod (r = 0.76) in 2021. A principal component analysis highlighted the effectiveness of organic–inorganic fertilizer combinations, particularly O5000N60D50, in enhancing productivity while potentially reducing inorganic fertilizer application. This study demonstrates that integrating organic and inorganic fertilizers enhances sustainable crop productivity and reduces negative environmental impacts, particularly in regions facing nutrient depletion and drought conditions. Full article

The ECM is composed of a considerable number of biochemically and structurally diverse constituents. ECM is a highly dynamic system that constantly receives and sends biological, chemical and mechanical signals. Several studies suggest that mechanical signals derived from the extracellular microenvironment regulate skin regeneration and wound healing. Tests measuring collagen contraction showed a significant difference in contraction activation in samples treated with the 2% colostrum derivative mixture compared to the control. The analysis of the supernatant showed an inhibition of metalloproteinase-2 expression and an increase in collagen secretion by fibroblasts in treatment samples. Our hypothesis is that the molecules extracted and purified from bovine colostrum can restore the ECM environment qualitative and quantitative characteristics, thus permitting, through a mechanical action, the restoration of the wound due to the transduction of the signal activated by the integrins. Full article

Melanogenesis is regulated by melanogenic enzymes such as tyrosinase (TYR), TRP-1, and TRP-2, whose expression is controlled by the microphthalmia-associated transcription factor (MITF). Various signaling pathways, including cAMP/PKA, MAPK/ERK, Wnt/β-catenin, and PI3K/Akt, are involved in this process and have been a focal point of research for treating pigmentation disorders. However, developing effective therapies for conditions like vitiligo remains a significant challenge. In this study, the effects of umckalin on melanogenesis and its molecular mechanisms were investigated using B16F10 cells, a mouse melanoma cell line widely used as a model for melanin production studies. B16F10 cells produce melanin via melanosomes and express key melanogenic enzymes such as TYR, TRP-1, and TRP-2, making them a reliable model system. Our findings demonstrate that umckalin promotes melanogenesis in a concentration-dependent manner by upregulating TRP-1 expression and activating the MITF signaling pathway. Additionally, umckalin modulated key signaling pathways, including GSK3β/β-catenin and MAPK, to enhance melanogenesis. In conclusion, umckalin enhances melanogenic enzyme activity by activating critical signaling pathways, thereby promoting melanin synthesis. These findings suggest that umckalin could be a promising candidate for developing therapeutic agents for pigmentation disorders such as vitiligo. Further studies are required to explore its mechanisms and clinical applications in greater detail. Full article

The role of oxidants and antioxidants in inflammatory bowel disease (IBD) has been actively explored since the early 1980s, starting with the role of the respiratory burst of neutrophils and ischemia in bowel pathology. Since that time, the enzymatic components contributing to the pool of reactive oxygen species, including superoxide, H₂O₂, and lipid hydroperoxides, and the counteracting antioxidants—catalase, glutathione peroxidases (Gpx), peroxiredoxins (PRDX), superoxide dismutases, and others—have been fleshed out. My perspective on IBD is from the role of the balance or imbalance of enzymatic oxidant sources and enzymatic antioxidants in the inflammatory process. I will present evidence on the involvement of oxidant and antioxidant processes in IBD based, as much as possible, on my experiences with Gpxs. This evidence will be discussed in terms of both the immune system and local bowel oxidant and antioxidant systems. As Gpxs are generally selenium-dependent, possible deficiencies in selenium uptake in active IBD and the impact on Gpx expression will be explored. The more recently introduced ferroptosis, an iron-dependent lipid peroxidation-based pathological process, will be reviewed for its possible involvement in IBD. Full article

Accurate primer and probe design is essential for molecular applications, including PCR, qPCR, and molecular multiparameter assays like microarrays. The novel software tool AssayBLAST addresses this need by simulating interactions between oligonucleotides and target sequences. AssayBLAST handles large sets of primer and probe sequences simultaneously and supports comprehensive assay designs by allowing users to identify off-target binding, calculate melting temperatures, and ensure strand specificity, a critical but often overlooked aspect. AssayBLAST performs two optimized BLAST-based searches for each primer or probe sequence, checking the forward and reverse strands for off-target interactions and strand-specific binding accuracy. The results are compiled into a mapping table containing binding sites, mismatches, and strand orientation, allowing users to validate large sets of oligonucleotides across predefined custom databases for a complete and optimal theoretical assay design. AssayBLAST was evaluated against experimental Staphylococcus aureus microarray data, achieving 97.5% accuracy in predicting probe–target hybridization outcomes. This high accuracy demonstrates the method’s effectiveness in reliably using BLAST hits and mismatch counts to predict microarray results. AssayBLAST provides a reliable, scalable solution for in silico primer and probe validation, effectively supporting large-scale assay designs and optimizations. Its accurate prediction of hybridization outcomes demonstrates its utility in enhancing the efficiency and reliability of molecular assays. Full article

This review summarizes recent findings on the diverse roles of bacterial sialidases in microbial biology. Bacterial sialidases, also known as neuraminidases, are exog α-lycosidases that cleave terminal sialic acid residues from a number of complex compounds designated as sialoglycoconjugates (glycoproteins, glycolipids and oligosaccharides). Metabolically, they are involved in sialic acid catabolism, providing energy, carbon and nitrogen sources. Catabolic degradation of sialic acids is a physiological feature that can be considered an important virulence factor in pathogenic microorganisms. Sialidases play a pivotal role in host–pathogen interactions and promotion of bacterial colonization. The activity of these enzymes enables bacterial adhesion, biofilm formation, tissue invasion, and also provides immune evasion by exposing cryptic receptors and modifying immune components. Many different perspectives are being developed for the potential application of sialidases. In the field of medicine, they are being explored as appropriate targets for antimicrobials, vaccines, diagnostic preparations and in tumor immunotherapy. In the field of enzymatic synthesis, they are used for the regioselective production of oligosaccharide analogs, enzymatic separation of isoenzymes and as a tool for structural analysis of sialylated glycans, among other applications. Full article

Hematologic malignancies, including leukemia, lymphoma, and multiple myeloma, pose significant therapeutic challenges due to their heterogeneity and high relapse rates. Nanotechnology has emerged as a promising avenue for precision drug delivery in these malignancies, allowing for enhanced drug concentration at tumor sites and reducing systemic toxicity. Recent developments in nanocarriers—such as liposomes, polymeric nanoparticles, and inorganic nanoparticles—have enabled targeted approaches, utilizing molecular markers specific to malignant cells to increase therapeutic efficacy while minimizing adverse effects. Evidence from preclinical and clinical studies underscores the potential of nanotechnology to improve patient outcomes by facilitating controlled release, improved bioavailability, and reduced toxicity. However, translating these advancements into clinical practice requires further research to validate their safety and efficacy. This review provides a comprehensive analysis of the latest innovations in nanotechnology for targeted drug delivery in hematologic malignancies, addressing current achievements and future directions for integrating these approaches into Clinical Hemato-Oncology. Full article

The nutritional value of Inga paterno seeds remains largely unexplored. Given the global protein deficiency, underutilized legumes like I. paterno could serve as alternative protein sources. This study evaluated the effect of sprouting on the composition, protein digestibility (PD) as soluble protein (SP), amino acid profile, free amino acids by UHPLC, and nutritional indicators of I. paterno seed flour. Seeds were sprouted for 0, 2, 4, 6, 8, or 10 days, then dried, milled, and analyzed. The seeds reached 100% sprouting after six days. Sprouting led to a 54.36% decrease in protein content but a 109% increase in the lipid fraction by day six. PD doubled after 8–10 days of sprouting. Additionally, total amino acid content significantly increased and the chemical score of majority essential amino acids tripled. After in vitro digestion, sprouted flour released higher amounts of free amino acids, particularly aspartic acid (from 9.10 ± 0.18 to 19.65 ± 0.97 mg/L), histidine (from 33.48 ± 0.61 to 46.29 ± 2.34 mg/L), alanine (from 16.32 ± 0.40 to 23.74 ± 0.07 mg/L), and lysine (from no detected to 7.12 ± 0.36 m/L). These findings suggest that sprouted I. paterno seeds could be a valuable, digestible protein source with enhanced nutritional quality, making them a promising ingredient for the food industry. Full article

Lung cancer is by far the leading cause of cancer death among both men and women, making up almost 25% of all cancer deaths Each year, more people die of lung cancer than colon, breast, and prostate cancer combined. The early detection of lung cancer is critical for improving patient outcomes, and automation through advanced image analysis techniques can significantly assist radiologists. This paper presents the development and evaluation of a computer-aided diagnostic system for lung cancer screening, focusing on pulmonary nodule segmentation in low-dose CT images, by employing HighRes3DNet. HighRes3DNet is a specialized 3D convolutional neural network (CNN) architecture based on ResNet principles which uses residual connections to efficiently learn complex spatial features from 3D volumetric data. To address the challenges of processing large CT volumes, an efficient patch-based extraction pipeline was developed. This method dynamically extracts 3D patches during training with a probabilistic approach, prioritizing patches likely to contain nodules while maintaining diversity. Data augmentation techniques, including random flips, affine transformations, elastic deformations, and swaps, were applied in the 3D space to enhance the robustness of the training process and mitigate overfitting. Using a public low-dose CT dataset, this approach achieved a Dice coefficient of 82.65% on the testing set for 3D nodule segmentation, demonstrating precise and reliable predictions. The findings highlight the potential of this system to enhance efficiency and accuracy in lung cancer screening, providing a valuable tool to support radiologists in clinical decision-making. Full article

The genus Quercus (Fagaceae) is one of the most widely distributed and species-diverse trees in the Northern Hemisphere. The present study addresses the investigation of the phyto-chemical profile by ten assays, the antioxidant activity scavenging of DPPH radicals, total phenolic content, total flavonoids, and antimicrobial activity against three pathogenic bacteria with the foliage of two species of red oak (Quercus sartorii and Quercus rysophylla). Both species of oak showed a high phenolic content in the aqueous extract (22,342.10 ± 3076.5 mg GAE/kg of plant and 17,747.14 ± 1139.9 mg GAE/kg of plant, respectively). In the flavonoid content, Q. sartorii showed a higher amount in the ethanolic extract (24,587.42 ± 996.3 mg QE/kg of plant), while for Q. rysophylla, it was methanolic extract (19,875.66 ± 2754.01 QE/kg of plant). In the DPPH radical scavenging activity, Q. sartorii showed the highest percentage of inhibition in the methanolic extract (81.14 ± 1.7%), while in Q. rysophylla, it was the ethanolic extract (82.60 ± 2.7%). In the antimicrobial tests, inhibition halos were obtained in the strains Acinetobacter baumannii and Staphylococcus aureus of both species. All this gives a guideline to comprehensively elucidate the metabolites present in these two species for further study and application in the dispute against pathogenic bacteria or in diseases related to the imbalance of reactive oxygen species (ROS). Full article

Cannabis sativa is used for multiple purposes, notably for its medicinal properties. It produces various secondary metabolites, including cannabinoids, terpenes, and flavonoids, which have therapeutic value and typically produce high amounts in female plants. The growth of the global cannabis market has led to intensive breeding efforts to develop elite cultivars with enhanced secondary metabolite profiles. As a dioecious and anemophilous plant, it produces staminate and pistillate inflorescences on separate plants and relies on wind for pollination, rendering traditional propagation methods challenging owing to high genetic recombination in progeny. Consequently, asexual propagation (micropropagation) is commonly employed to maintain female clones entirely. Micropropagation/direct organogenesis is a tissue culture technique that produces numerous disease-free clone plants in vitro more rapidly than traditional rooted cuttings. Factors such as sterilization, hormonal balance, explant type, nutrient additives, carbon source, pH, and environment influence the success of cultivar-specific micropropagation. In this review, we discussed how these factors affect cannabis micropropagation based on recent findings, emphasizing the importance of optimizing cultivar-specific protocols for long-term germplasm conservation and efficient breeding based on a mechanistic background. Full article

Accurate segmentation of Regions of Interest (ROI) in lung Computed Tomography (CT) is crucial for early lung cancer diagnosis and treatment planning. However, the variability in size, shape, and location of lung lesions, along with the complexity of 3D spatial relationships, poses significant challenges. In this work, we propose SALM (Segment Anything in Lung Model), a deep learning model for 2D and 3D ROI segmentation. SALM leverages Vision Transformers, proposing an adaptation of positional encoding functions to effectively capture spatial relationships in both 2D slices and 3D volumes using a single, unified model. Evaluation on the LUNA16 dataset demonstrated strong performance in both modalities. In 2D segmentation, SALM achieved a Dice score of 93% on 124,662 slices. For 3D segmentation using 174 3D images from the same dataset, SALM attained a Dice score of 81.88%. We also tested SALM on an external database (PleThora) on a subset of 255 pulmonary CT from diseased patients, where it achieved a Dice score of 78.82%. These results highlight SALM’s ability to accurately segment lung ROI in both 2D and 3D, demonstrating its potential to improve the accuracy and efficiency of computer-aided diagnosis for lung cancer. Full article

Fusobacterium necrophorum subspecies necrophorum, a resident of the rumen, is the causative agent of bovine liver abscesses. Currently, tylosin, a macrolide, is used in the feed to reduce liver abscesses. Because macrolides are medically important antibiotics, their use in food animal production is of public health concern. There is significant interest in finding antimicrobial alternatives. Bacteriophages that lyse subsp. necrophorum have the potential to replace tylosin. Our objective was to isolate bacteriophages lytic to subsp. necrophorum. Pooled ruminal fluid from slaughtered cattle and pooled sewage samples were collected and incubated overnight with lysine, and subsp. necrophorum strains and filtrates were spotted on F. necrophorum lawns. Phage plaques were harvested and purified. Bacteriophage isolation frequencies were compared between sample types, sampling dates, and necrophorum strains. Overall relative frequency of isolated bacteriophages lytic to subsp. necrophorum was 17.1%. The frequency of bacteriophage isolation ranged from 0 to 25.4% for ruminal fluid, and from 13.7 to 32.0% for sewage. Isolation frequency was significantly higher in sewage than in ruminal fluid samples (p < 0.01). Isolation rates varied significantly between necrophorum strains. Sewage was a rich source of bacteriophages lytic to necrophorum, which have the potential to be used to prevent liver abscesses. Full article