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Santalum album is an economically important plant in the craft, spices and medicine industries. The main chemical constituents found in sandalwood essential oils are sesquiterpenes. 3-Hydroxy-3-methylglutaryl monoacyl-coenzyme A reductase (HMGR) is one of the rate-limiting enzymes required for the synthesis of sandal sesquiterpenes, but there are no studies on the HMGR gene in S. album. In this study, the full-length ORFs of the upper rate-limiting enzyme genes SaHMGR1 and SaHMGR2, which lie upstream of the MVA metabolic pathway of sandal sesquiterpenes, were cloned for the first time. Bioinformatics and phylogenetic analyses were conducted. The results showed that SaHMGR1 and SaHMGR2 had typical domains of HMGR class I enzymes in the HMGR superfamily, including four catalytic sites, six NADPH-binding sites, five substrate binding regions, four inhibitor binding sites, and several dimer interface regions. A phylogenetic analysis showed that SaHMGR1 and SaHMGR2 were highly conserved relative to corresponding genes in other plants. An analysis of subcellular localization showed that these SaHMGR genes were located in the endoplasmic reticulum. SaHMGR1 and SaHMGR2 were detected by real-time PCR in roots, sapwood, heartwood, young leaves, mature leaves and twigs. Highest expression was in roots. SaHMGR1 expression was higher in mature leaves than in heartwood while SaHMGR2 expression was lower in mature leaves than in heartwood. Expression in Escherichia coli strain DH5α with plasmid pET-32a (+) was also used to verify the functionality of both HMGR proteins, which catalyzed the formation of MVA from HMG-CoA. In E. coli, the enzymatic activity of SaHMGR1 was higher than that of SaHMGR2. These findings provide a basis for further studies on the function of SaHMGR genes and the regulation of sesquiterpene biosynthesis in S. album., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Β-Gals are a subgroup of the glycoside hydrolase (GH) family of enzymes, which possess the Glyco_hydro_35 (GH35) domain. Although studies have been conducted on the β-Gal gene family in numerous plant species, no such research has been conducted on beans. The purpose of this study was to determine the gene expression levels of β-Gal genes in the leaf tissue of P. vulgaris under salt and drought stress using quantitative real-time polymerase chain reaction (qRT-PCR) and to perform a comprehensive analysis of β-Gal gene family members using bioinformatics tools. In the bean genome, 25 Pvul-βGAL proteins with amino acid numbers ranging from 291 to 1119, molecular weights from 32.94 to 126.56 kDa, and isoelectric points from 5.46 to 9.08 were identified. Both segmental and tandem duplication have occurred in β-Gal genes in the bean genome, and Pvul-BGAL genes have been subject to negative selection in the evolutionary process. For a deeper comprehension of the evolutionary proximity of Pvul-BGAL genes, a phylogenetic tree and synteny map were drawn together with Arabidopsis thaliana and Glycine max β-Gal genes. The expression profiles of β-Gal genes in different tissues of the bean were determined in silico. In addition, the expression profiles of β-Gal genes in the leaves of bean plants subjected to drought and salt stress were analyzed, and the role of β-Gal genes in salt and drought stress was estimated. In this study, the role of β-Gal gene family in abiotic stress response and the characterization of β-Gal genes in beans were determined for the first time and will provide a basis for future functional genomics studies., Competing Interests: Declarations. Conflict of interest: The authors have no conflicts of interest to declare. Ethics approval and consent to participate: This research study complies with research publishing ethics. In addition, the manuscript meets all applicable standards with regard to the ethics of experimentation and research, and there is no duplicate publication, fraud, plagiarism, or concerns about animal or human experimentation., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Potato (Solanum tuberosum L.), an important horticultural crop is a member of the family Solanaceae and is mainly grown for consumption at global level. Starch, the principal component of tubers, is one of the significant elements for food and non-food-based applications. The genes associated with biosynthesis of starch have been investigated extensively over the last few decades. However, a complete regulation pathway of constituent of amylose and amylopectin are still not deeply explored. The current in-silico study of genes related to amylose and amylopectin synthesis and their genomic organization in potato is still lacking. In the current study, the nucleotide and amino acid arrangement in genome and twenty-two genes linked to starch biosynthesis pathway in potato were analysed. The genomic structure analysis was also performed to find out the structural pattern and phylogenetic relationship of genes. The genome mining and structure analysis identified ten specific motifs and phylogenetic analysis of starch biosynthesis genes divided them into three different clades on the basis of their functioning and phylogeny. Quantitative real-time PCR (qRT-PCR) of amylose biosynthesis pathway genes in three contrast genotypes revealed the down-gene expression that leads to identify potential cultivar for functional genomic approaches. These potential lines may help to achieve higher content of resistant starch., Competing Interests: Declarations. Competing interests: The authors declare that there are no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The Human T-cell lymphotropic virus type 1 (HTLV-1), an oncogenic virus belonging to the Deltaretrovirus genus, expresses various proteins, including Tax and HBZ, which can affect many cellular pathways. In this study, we have investigated the role of the circadian rhythm signaling pathway, a key regulator of human health, in the pathogenesis of HTLV-1 using Boolean Network analysis and laboratory methods. After an extensive search of the circadian rhythm pathway, we analyzed the relationships between the genes of this pathway using the R programming language and the BoolNet package. Subsequently, we examined the impact of viral proteins on the cellular clock rhythm genes. Finally, we identified three genes, PER2, CRY1, and DEC1, as the main checkpoints from the attractors obtained. These three genes and two viral genes, Tax and HBZ, were quantitatively assessed on two groups of individuals, including ten asymptomatic carriers infected with HTLV-1 and ten healthy individuals using the qRT-PCR method. Our results showed that the expression level of PER2 and DEC1 genes was significantly higher in the asymptomatic carriers compared to the healthy control group. Also, we recorded positive correlations between PER2 and DEC1, CRY1 and DEC1, and negative correlations between HBZ and CRY1 and DEC1. In this study, we suggested that in asymptomatic carriers, the virus might try to induce a chronic infection by escaping from the immune system due to an alteration in circadian rhythm pathways. We also detected three promising genes in this pathway that could have therapeutic or diagnostic value in these individuals. However, this possibility requires further research in different periods, different groups (e.g., ATLL and HAM/TSP), and examining a more significant number of circadian rhythm genes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025. Published by Elsevier B.V.)

Papaya (Carica papaya L.) is a commercially important fruit crop cultivated worldwide due to its nutritional and medicinal values. Papaya ringspot disease (PRSD), caused by the papaya ringspot virus (PRSV), poses a significant threat to papaya cultivation, resulting in substantial yield losses. In this study, two independent field experiments were conducted at Bagalkote located in the Northern Dry Zone of Karnataka state of India. The first experiment aimed to identify the optimal planting month for papaya to effectively manage PRSV disease. The second experiment was conducted to determine the susceptible papaya growth stage for PRSV infection. The results revealed that early planting of papaya in June or late planting in March were identified as the most optimal planting times across the majority of growth stages, as they exhibited the lowest disease incidence along with superior growth and yield characteristics compared to other planting months. In contrast, planting during the winter season (September to January) resulted in high disease severity due to an increased aphid population. Conversely, planting during periods of low aphid activity (spring season) delayed disease onset until the monsoon. By the time the aphid population increased, the plants had already progressed beyond the flowering and fruit-bearing stages. In the second experiment, the severity and frequency of foliar symptoms on PRSV-inoculated papaya plants were significantly higher in those inoculated at the early growth stage compared to those inoculated at a later growth stage. This indicates that the early growth stage (up to 180 days after transplanting) is a critical period for PRSV infection, necessitating the implementation of effective disease management measures during this time to minimize disease spread and enhance growth and yield. Furthermore, plants inoculated at the early stage exhibited a higher viral titer, more severe symptoms, and a higher percent transmission rate compared to those inoculated at a later stage. These findings were supported by qRT-PCR analysis, which demonstrated a highly significant and positive correlation between early inoculation and disease severity., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Osteoarthritis (OA) is a widespread chronic bone and joint disease for which there is currently no effective preventive or therapeutic treatment. Accumulating evidence indicates that circular RNAs (circRNAs), a class of noncoding RNAs, play critical roles in OA. Therefore, in this study, we aimed to reveal an unexplored circTBCK and elucidate its mechanism of action in the pathological process of OA. The different expression of circTBCK was obtained both in vitro and in vivo. In the in vivo model, mice were induced via destabilization of the medial meniscus (DMM) surgery, while in vitro model, mouse cells like primary chondrocytes of newborn mice and ATDC5 cell line were treated with IL-1β treatment (10 ng/mL for 24 h). The level of circTBCK was examined by quantitative real-time polymerase chain reaction (qRT-PCR). After circTBCK was overexpressed or knocked down, IL-1β treatment was performed, and then, chondrocyte viability was detected via a Cell Counting Kit-8 (CCK-8) assay at 0, 24, 48, or 72 h. To assess type II collagen (Collagen II) expression, immunofluorescence (IF) analysis was used. The levels of mRNAs and proteins related to proliferation, the extracellular matrix (ECM) and autophagy were determined by qRT-PCR and Western blotting. Compared with OA treatment, primary chondrocytes with treatment of both circTBCK overexpression and IL-1βincreased the expression of anabolic factors-Collagen II and SRY-box transcription factor 9 (SOX9), proliferation-related molecules-Ki-67 and proliferating cell nuclear antigen (PCNA), and autophagy-related molecules-Microtubule-associated protein 1 light chain 3 (LC3), B-cell lymphoma 1 (Bcl1), and autophagy-related 5 (Atg5) and decreased Sequestosome 1 (SQSTM1 or P62). In contrast, knockdown of circTBCK aggravated the chondrocyte degeneration induced by IL-1β. Overall, our findings suggest that circTBCK, an unexplored circRNA, could regulate autophagy, proliferation, and the extracellular matrix (ECM) to mitigate the development of OA, suggesting a possible target for OA prevention and therapy., Competing Interests: Declarations. Conflict of interest: The authors declare that they have no competing interests. Ethical approval: All animals were kept in a pathogen-free environment and fed adlib. The procedures for care and use of animals were approved by the Ethical Committee of Animal Experiments and Experiments Animal Welfare Capital Medical University (Ethics Batch No. AEEI-2023-280) and were performed in accordance with the committee’s guidelines., (© 2025. The Author(s).)

Introduction: This study aimed to assess the expression changes of BTG1, PGI, and PGII in tissues and serum of patients with gastric cancer, atrophic gastritis, and healthy individuals., Methods: QRT-PCR was used to measure BTG1, PGI, and PGII expression in 30 cancers, 30 atrophic gastritis, and 30 healthy tissue samples. Serum levels of PGI and PGII were measured using ELISA. Statistical tests included the Mann-Whitney U and independent T-test. Covariates like tumour stage and H. pylori status were considered., Results: BTG1 expression was significantly lower in cancer and gastritis tissues. Serum PGI and PGII levels were significantly reduced in cancer patients ( P  ≤ 0.001)., Discussion: The PGI/PGII ratio in serum emerged as a strong non-invasive biomarker for distinguishing cancer from healthy individuals. While BTG1 provides insights into gastric carcinogenesis, its clinical utility is limited due to the need for tissue samples. The serum-based PGI/PGII ratio shows greater promise as a non-invasive screening tool for GC.

Background: The transition from vegetative to floral development is a critical process regulated by a variety of complex internal and external factors. This study aims to characterize the Flowering Locus D (FLD) gene in relation to the floral induction pathway and examine its expression profiles under different photoperiodic conditions in Indian bean (Lablab purpureus (L.) Sweet)., Methods: The LprFLD gene was PCR-amplified and sequenced using gene-specific primers for both photosensitive (GNIB-22, GP-189) and photoinsensitive (GNIB-21) varieties of Indian bean. Exon prediction, phylogenetic analysis, and in silico domain prediction were performed, followed by gene expression analysis using qRT-PCR for contrasting phenotypes., Results: The complete LprFLD gene sequence was characterized, containing five exons and showing significant similarity to GmFLD and VuFLD. Phylogenetic analysis of FLD homologs from 27 taxa revealed the closest evolutionary relationships among members of the Phaseoleae tribe, including Lablab purpureus, Vignaumbellata, Vigna angularis, Vignaradiata, Vigna unguiculata, Phaseolus vulgaris, Glycine max, Glycinesoja, and Cajanus cajan. In silico domain prediction identified two conserved domains, SWIRM and amino oxidase, in the FLD homologs of Indian bean, soybean, cowpea, Arabidopsis and rice. Gene expression analysis showed that the relative expression of LprFLD in trifoliate leaves corresponded with flowering in both photosensitive and photoinsensitive genotypes under long-day conditions. Overall, the expression level of LprFLD was lower in trifoliate leaves and shoot tips., Conclusion: This is the first report on the characterization of the LprFLD locus in Indian bean. Phylogenetic analysis confirmed its close evolutionary relationship with other Phaseoleae species. qRT-PCR results suggest that the elevated expression of LprFLD in trifoliate leaves coincides with flowering time in both photosensitive and photoinsensitive genotypes under long-day conditions, indicating that other loci may also regulate floral transition under varying photoperiods in Indian bean., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s), under exclusive licence to Springer Nature B.V.)

There is growing evidence linking neutrophil extracellular traps (NETs) to tumor genesis, growth, distant metastasis, and tumor-related thrombosis. However, the roles of NETs-related genes (NETRGs) on LGG prognosis remain unclear. The purpose of this study was to integrate multiple machine learning techniques and experiment validation to develop a reliable NETs-based signature that opens up novel approaches for assessing the prognosis and treatment response of LGG patients. Consensus clustering, k-means clustering and Nonnegative Matrix Factorization was used for the TCGA-LGG dataset and identified two NETs-related subgroups. The prognostic hallmark and nomogram for LGG were developed, which consist of five differentially expressed NETRGs (FPR1, PTAFR, SLC11A1, ICAM1, LTF) based on nine analytic approaches. The ROC curves and calibration curves of our NETRGs signature and nomogram exhibited strong and robust prognosis prediction abilities in both the TCGA-LGG training set and CGGA-325, CGGA-693 validation sets. The prognosis for LGG individuals in the low-risk category was better. The TISCH was used to examine the five NETRGs at the single-cell level. Common immunological checkpoints were expressed at greater levels in high-risk individuals. LGG individuals in the low-risk category posses a higher likelihood of being sensitive to Carmustine and Vincristine, as indicated by the drug sensitivity analysis. The qRT-PCR experiment and immunohistochemistry images confirmed that the expression of FPR1, PTAFR, SLC11A1 and ICAM1 are higher in low-grade oligodendroglioma. The NETRGs signature and nomogram can accurately and conveniently predict the LGG patients' prognosis, which can facilitate individualized treatment and the improvement of prognosis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025. Published by Elsevier Ltd.)

The increasing temperature is a major threat to plant growth and development. It severely alters various biochemical and physiological processes and ultimately affects the overall crop yield. The membrane-based remorin protein-encoding genes (REM) were previously reported as significantly involved in the regulation of various biotic and abiotic stressors. However, these REM genes were not studied in Cajanus Cajan, the sixth most important legume crop due to its rich protein source and traditional medicinal plant. In this study, 17 REM gene orthologs were identified in C. cajan against A. thaliana REM genes and verified through the presence of conserved REM-related domains. Phylogenetic analysis revealed that REM genes were divided into six different groups. All the REM genes were unevenly distributed on 11 chromosomes of C. cajan. 3D protein structures and intron-exon organization indicate conserved evolutionary pattern within C. cajan. Various core, hormone-responsive, and stress-responsive cis-regulatory elements were found in promoter regions of REM genes, including TATA-box, CAAT-box, MYB, and G-box. The total estimation of antioxidant enzyme activity revealed the increase in POD and SOD activity, potentially due to a defense mechanism in response to high temperature. RNA sequencing data processing reveals higher expression of CcREM genes in leaf and flower, including CcREM1.1 and CcREM1.2. Furthermore, the differential change in expression was observed in response to high-temperature stress. Among these genes, one upregulated gene (CcREM1.3) and two downregulated genes (CcREM6.1 and CcREM6.5) are potential candidate targets for heat stress response, followed by qRT-PCR validation. Our findings suggest that CcREM1-like and CcREM6-like genes hold significant potential for future climate-smart heat-tolerant breeding of C. cajan., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

The aim of this study was to deeply explore the pathogenesis of obesity type 2 diabetes mellitus (O-T2DM) and search for potential biomarkers through high-throughput RNA sequencing technology. The study included 15 patients with O-T2DM and 15 healthy controls, and peripheral blood samples were collected for transcriptome analysis. The results showed that compared with the control group, there were 442 circRNAs and 2756 mRNAs with significant differential expression in the O-T2DM group. Through weighted gene co-expression network analysis (WGCNA) and pathway enrichment analysis, it was found that the differentially expressed mRNAs were mainly enriched in signaling pathways such as T cell receptor, cell senescence, cytotoxicity mediated by NK cells, IL-17, lipids and atherosclerosis, and the oxidative phosphorylation pathway was activated, and apoptosis was inhibited. Based on the ceRNA theory, a regulatory network was constructed, and key circRNAs such as hsa_circ_0060614 were screened out, which may regulate the expression of the MT2A gene by adsorbing hsa-mir-4668-3p, and the expression levels of the three were significantly increased in O-T2DM patients. This study provides a new perspective for the research on the molecular mechanism of O-T2DM and an important theoretical basis for the development of personalized treatment and precision medicine for it., Competing Interests: Declarations. Ethics approval and consent to participate: The study was approved by the Ethics Committee of Beijing University of Traditional Chinese Medicine (BUCM), with the ethical batch number of (2017BZHYLL0105). All procedures were performed in accordance with the Declaration of Helsinki, and all participants received written informed consent. Consent for publication: Both written and verbal consent was obtained from the subjects of the study. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Introduction: The basic helix-loop-helix (bHLH) proteins are a large family of transcription factors that are essential to physiology, metabolism, and development. However, the available information is limited about the bHLH gene family in Chimonobambusa utilis , which is widely cultivated in China because of its high-quality and economic value. C. utilis cultivars exhibit five natural color variations in their shoot sheaths, but the molecular mechanism behind this color diversity remains unclear., Methods: De novo assembly was employed to obtain gene sequences. To identify pathways related to color formation, GO enrichment analysis was performed on the 44,255 functionally annotated unigenes., Results: The transcriptomic analysis of C. utilis yielded a total of 195,977 transcripts and 75,137 unigenes after removing redundancy. The enrichment results revealed that four pathways were most strongly associated with color formation. Phylogenetic, conserved motif, and protein-protein interaction analyses, along with qRT-PCR validation, confirmed CubHLH17 's role in red sheath color., Discussion: This research not only deepens insights into the functional roles of CubHLH genes but also lays the foundation for genetic improvement of bamboo species. We suggest that these findings will contribute to both scientific research and commercial bamboo cultivation through gene editing technology in the future., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2025 Tong, Zeng, Guo, Li, Li, Chen and Liu.)

Background: Monkeypox virus (MPXV) has emerged as a significant global health concern with outbreaks worldwide. While MPXV is primarily known for its dermatological and systemic manifestations, it can also cause central nervous system (CNS) complications. This systematic review describes the demographic, clinical, diagnostic, and therapeutic characteristics of MPXV-associated CNS neuroinflammatory disorders., Methods: We systematically reviewed the literature to identify cases of MPXV-associated CNS neuroinflammatory disorders. Data on demographics, systemic and neurological manifestations, diagnostic methods, treatment strategies, and outcomes were extracted and analyzed., Results: Eighteen cases of MPXV-associated neuroinflammatory disorders were identified. The mean age of patients was 27.8 years (range: 28 days to 43 years), with a male predominance (66.7%). Diagnosis included The most common diagnoses were acute disseminated encephalomyelitis in nine cases (50.0%), encephalitis/meningoencephalitis in seven cases (38.9%, isolated transverse myelitis in one case (5.6%), and transverse myelitis with encephalitis in one case (5.6%). The latency between the onset of systemic symptoms and neurological involvement averaged 6.2 days. MPXV detection was confirmed in 13 of 18 (72.2%) cases, primarily using quantitative real-time polymerase chain reaction from various biological specimens. Among the 12 cases with documented treatment, the most commonly administered therapies were tecovirimat (58.3%) and intravenous methyl-prednisolone (66.7%). Outcomes were reported in 17 cases, with complete recovery in 29.4%, partial recovery in 41.2%, and death in 29.4% of patients., Conclusions: MPXV-associated neuroinflammatory disorders of the CNS are rare but cause significant complications. The findings underscore the need for clinical vigilance, advanced diagnostic approaches, and targeted therapeutic strategies. Further research is essential to elucidate mechanisms underlying MPXV neurovirulence and develop effective treatments for these life-threatening conditions., Competing Interests: Declarations. Ethics approval and consent to participate: As this study is a systematic review of published literature, no ethics approval or patient consent was required. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Background: The Philadelphia chromosome (Ph) represented a finding of chronic myeloid leukemia (CML) in most cases which formed from t (9; 22) (q34; q11) resulting in the Breakpoint cluster region-Abelson tyrosine-protein kinase1 (BCR-ABL1) fusion gene. Assuming CCE's inaccuracies in diagnosing CML and FISH's limitations with low BCR-ABL1 percentages, a Predicted-FISH (Pred-FISH) was developed. This model predicts treatment response during follow-up by integrating qRT-PCR results, White Blood Cell (WBC) counts, and Cytogenetic Response data., Methods: Quantitative Real-Time Polymerase Chain Reaction Analysis (qRT-PCR), fluorescence in situ hybridization (FISH), and Conventional Cytogenetic Examination (CCE or Karyotyping) have been used in the detection and follow-up of CML patients. The study included 110 individuals, divided into three groups: 31.82% (35 individuals) were newly diagnosed CML patients, another 22.73% (25 individuals) were healthy control samples, and the remaining 45.45% (50 individuals) were previously diagnosed CML patients., Results: Include BCR-ABL1 fusion gene levels detected by qRT-PCR, Ph chromosome presence t (9; 22) (q34; q11) observed by CCE, and WBC counts. The FISH test, used to confirm disease in new patients before treatment, was compared to CCE results due to its insensitivity in certain conditions. Data from CCE, FISH, qRT-PCR, and WBC for newly diagnosed patients provided a standard for evaluating the Predicted-FISH., Conclusion: The FISH technique excels in disease detection with over 98% accuracy and high sensitivity. QRT-PCR is effective for monitoring CML and BCR-ABL1 gene levels, indicating MMR and DMR. CCE, while useful for posttreatment monitoring, is less accurate in measuring treatment response over time., (© 2025 The Author(s). Journal of Clinical Laboratory Analysis published by Wiley Periodicals LLC.)

Background: One of the primary challenges that the expanding population faces is water scarcity. Thus, a global imperative has been established to safeguard extant water resources and optimize their utility through sustainable practices and efficient management. In the present investigation, Azolla pinnata, a pteridophyte (fern), was employed to phytoremediate Cr (VI) from chromium-polluted water. The potential of this treated water for agricultural purposes was verified through the use of Vicia faba plants., Results: In vitro, A. pinnata effectively remediates Cr (VI) from an array of liquid concentrations (0.05 to 90 ppm) in a ratio of 25:1 {volume (mL): fresh weight of Azolla (g)} after 2 days incubation period at room temperature. At low concentrations (0.1 ppm), the phytoremediation capacity peaked at 70%, falling to 19.53% at a high concentration (90 ppm). Upon continuous irrigation with Cr-polluted water (0.05 to 50 ppm), the in vivo pot experiment on Vicia faba plants revealed high Cr accumulation in the roots reached 52.5 mg Kg -1 dry weight (Dwt) at the 50 ppm Cr treatment. Nevertheless, a reduced Cr content of 19.5 mg Kg -1 Dwt was observed when the plants were irrigated with 50 ppm Cr-polluted water that had been treated with Azolla. At 50 ppm of Cr, Azolla's treatment significantly increased shoot length, fresh weight, and Chl a content to 25.25 cm, 3.4 g, and 6.5 mg g -1 Dwt, respectively, up from 10.25, 1.8, and 4.7 in untreated plants. The chromosomal aberrations were significantly induced in the dividing cells of all Cr treatments, with the highest value of 4.8% at 50 ppm. This value was reduced to 2.88% at the same concentration when treated with Azolla. At a concentration of 10 ppm Cr, the mitotic index was significantly improved to 6.99% when combined with Azolla, as opposed to 3.63% when the same concentration was used without Azolla. The DNA degradation assay showed partial DNA degradation at 50 ppm Cr, which the Azolla treatment eliminated. Furthermore, the gene expression levels of both the PM H + -ATPase and the calcium-dependent protein kinase CDPK5 were upregulated in response to Cr, despite the fact that the expression level was altered in a dose- and concentration-dependent manner by Azolla treatment., Conclusion: Azolla exhibits substantial potential for reducing the detrimental effects of chromium stress including oxidative stress on plants. It modulates stress-related gene expression, protects DNA integrity, enhances cell mitosis, and reduces chromosomal damage. These results indicate that Azolla has the potential to be a valuable asset in phytoremediation strategies for chromium-contaminated environments, and that it may enhance plant survival and growth under Cr stress conditions., Key Message: Azolla pinnata can be effectively utilized as an environmentally-friendly method to remediate chromium-contaminated water for agricultural usage., Competing Interests: Declarations. Experimental research and field studies on plants: “All relevant institutional, national and international guidelines and legislation were compiled or adhered to the production of this study”. Ethics approval and consent to participate: This article does not contain any studies with human participants or animals performed by any of the authors. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Candida auris causes prolonged colonization and bloodstream infections in hospitalized patients. Different C. auris clades vary in their geographic origin, disease spectrum, and antifungal resistance, but biological basis underlying such variations needs further examination. Therefore, we investigated susceptible and multidrug-resistant C. auris isolates, obtained early in the New York outbreak, to benchmark their pathogenic potential in caterpillar worms of greater wax moth Galleria mellonella . Healthy G. mellonella worms responded to C. auris in a strain-specific, dose-responsive pattern. Three drug-resistant, clade I C. auris 17-1, 18-1, and 18-2 isolates caused high mortality, while a control, drug-susceptible, clade II C. auris 16-1 caused lower mortality ( P < 0.001) with graded inocula (1 × 10 4 to 1 × 10 7 CFU). Virulence correlated with fungal growth in hemolymph as C. auris 17-1, 18-1 and 18-2 reached high fungal cell load in the infected larvae, while C. auris 16-1 multiplied less proficiently. The degree of melanization was higher in worms infected with more pathogenic C. auris . Hemocoel histopathology showed more fungal elements and about 50 granulomas per profile for C. auris 17-1, 18-1 and 18-2 compared to 5 to 10 granulomas for C. auris 16-1. With more pathogenic C. auris isolates, the expression of gallerimycin, ceropin, and galiomycin increased significantly ( P < 0.05). Thus, the induction of G. mellonella immune effector peptides, histopathological responses, and melanization are proportionate to C. auris pathogenic potential. However, G. mellonella from different vendors showed unpredictable quality upon delivery that impacted feasibility and reproducibility of planned studies. Our results indicate utility of G. mellonella experimental model for screening of C. auris pathogenesis depending upon quality of worms commercially available in the USA.IMPORTANCEThe New York metropolitan area continues to suffer from the largest, ongoing outbreak of drug-resistant Candida auris . It is necessary to gather more information to determine if C. auris isolates from this outbreak vary in their ability to cause more severe disease in affected patients. Therefore, we studied the experimental model of greater wax moth Galleria mellonella . We found that susceptible and drug-resistant NY C. auris isolates caused measurable disease in G. mellonella . There was a good correlation between pathogenic potential of C. auris and melanin in the infected worms, fungal elements in worm hemolymph, uptick in the genes involved in insect immunity, and histopathological changes. The worms sourced from various commercial vendors were of variable quality. We confirm that G. mellonella remains a facile model to study C. auris experimental disease.

Cervical cancer remains a significant health burden worldwide, emphasizing the need for early detection and intervention. DNA replication is perturbed in cancer cells, and the minichromosome maintenance protein 10 plays an important role in origin firing. By analyzing the MCM10 mRNA expression in healthy controls, precancerous lesions, and cervical cancer using qRT-PCR, we can infer if it can be considered a biomarker. We collected cervical smear samples from patients and performed MCM10 expression analysis to set up thresholds for risk stratification. We also investigated the HPV status among the patient samples with precancerous lesions and cervical cancer and found 70 % of them to be positive. Our results demonstrated a significant upregulation of MCM10 mRNA expression in tumor samples (n = 40, 7.83 ± 1.2) and precancerous lesions (n = 54, 5.69 ± 1.4) compared to normal (n = 50, 4.27 ± 0.80) with a R 2 value of 0.59, confirming its role in the progression and development of cervical cancer. In conclusion, this study emphasizes the potential role of MCM10 as a biomarker. Our study would improve early detection rates, and we propose MCM10-based community screening for risk stratification, prevention, and prognosis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

TCP transcription factors are a unique class of transcription factors that play important roles in alleviating abiotic stresses such as drought and salt. In this study, the whole-genome data of three cultivated varieties, namely, "SFS", "Sang" and "OT3098v2", were utilized to identify and analyze the members of the TCP gene family in oats, and their responses to two abiotic stresses, drought and salt, were also investigated. Results showed that there are 83, 65, and 30 non-redundant TCP genes in the three oats, with the highest number of TCP genes specific to the "SFS", reaching 22 genes. The oat TCP genes can be classified into three subfamilies: PCF, CIN, and CYC/TB1. Most AsTCP genes have important motifs, Motif 1 and Motif 2, which are part of the bHLH domain. Additionally, various cis-acting elements related to hormone response, abiotic stress, light response, and growth and development were found in the promoters of AsTCP genes. The main amplification mechanism of the oat TCP gene family is fragment duplication. Two tandem duplications, AsTCP058 / AsTCP059 and AsTCP023 / AsTCP025 , are stably present in the three oats. The highest number of AsTCP collinear relationships exist in the "SFS" with 89 pairs. After drought and salt stress treatments, significant differences in gene expression were observed among different oat cultivars and treatment periods. Genes that showed significant expression changes under both treatments ( AsTCP021 , AsTCP033 , AsTCP044 , AsTCP053 , and AsTCP058 ) may play important roles in oat's response to abiotic stresses. Notably, AsTCP053 gene was significantly upregulated at 24 h of stress treatment and showed a more sensitive response to salt stress. This study provides insights into the functional characterization of the oat TCP gene family and its molecular mechanisms underlying stress tolerance., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2025 Nie, Zhao, Guo, Zhang, Han and Liu.)

This study was carried out to quantify changes in the activities of antioxidative enzymes such as catalase (CAT), peroxidase (POX) and superoxide dismutase (SOD) in two resistant onion cultivars 'Saba - HS' and 'Saba' and two susceptible cultivars 'Golden Eye' and 'Savannah Sweet' following an inoculation with Fusarium oxysporum f. sp. cepae (FOC) at seedling stage. Further, we assessed the expression of transcription factors (TFs), R1, PR5 and RGA29-genes that are involved in conferring resistance post-inoculation using qRT-PCR. The results revealed that the least disease severity occurred with the resistant 'Saba - HS' (4.7%) and 'Saba' (6.7%) cultivars, whereas the highest disease severity occurred with the susceptible 'Golden Eye' (89.6%) and 'Savannah Sweet' (88.9%) cultivars, respectively. Both resistant genotypes 'Saba' and 'Saba-HS' showed a significant increase in CAT, POX, and SOD activities at a transcriptional level. CAT activity was upregulated 4.81-fold in 'Saba' and 4.22-fold in 'Saba-HS' followed by POX where 'Saba' showed an increase by 3.53-fold and 'Saba-HS' by 2.35-fold, and SOD 'Saba' 17.46-fold and 'Saba-HS' 22.95-fold, relatively. Marker genes, RGA29, R1 and PR5, were also upregulated in the resistant-genotypes by 3.83, 4.78 and 5.01-fold, in comparison to their-controls, respectively. Similar trends were recorded for the biomass growth parameters (BGPs)., Competing Interests: Declaration of competing interest The authors state that, this research was conducted in the absence of any commercial and or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Feline infectious peritonitis virus (FIPV) remains as one of the leading causes of morbidity and mortality in young cats from shelters and catteries worldwide. Since little is known about the molecular characteristics of currently circulating FIPV strains in Long Island, New York, samples from two shelter cats submitted to the Pathology Diagnostic Services of the Long Island University College of Veterinary Medicine, with gross and microscopic lesions consistent with those of FIP were processed for virus isolation, molecular characterization and full-length genome decoding. The younger shelter cat, a 1-year-old male (A15) was found dead without previous signs of illness. Postmortem examination revealed gross and microscopic lesions characterized by vasculitis, necrosis, hemorrhage, and pyogranulomatous inflammation confined to the colon and associated lymph nodes. The second cat, a 7-year-old spayed female (A37) had an identical clinical history and similar but widespread lesions, including fibrinous peritoneal effusion, cecal, colonic, renal, and hepatic involvement. The gross and microscopic diagnosis of FIP in these cats was confirmed by immunohistochemistry (IHC) demonstration of feline coronavirus antigen using mouse anti-FIPV3-70 monoclonal antibody. Virus isolation from saved frozen kidney and colon tissue was performed through several subsequent blind passages in MDCK and Vero cell lines. Confirmation of the FIPV isolation was done through qRT-PCR, IFA, western blot using N protein antibodies, and NGS of the full-length genome sequencing. The full-length genome sequences of the virus isolate from the two cats were decoded using next-generation sequencing (NGS) and deposited in the GenBank as accession numbers PQ192636 and PQ202302. The genome size of these isolates was (29355 and 29321) nucleotides (nt) in length, respectively. While their genome organization was consistent with other FIPV genomes as follows (5'UTR-ORF1ab-S-3abc-M-E-7b-3'UTR-3'), marked differential mutations were observed in the ORF1a/b, S, 3Abc, and 7b protein genes of the two FIPV isolates. One notable deletion of 34 nucleotides was observed in the 7b genes of one of these isolates but was absent in the other. We confirmed the potential recombination events during the evolution of those two FIPV field isolates with the potential parent virus as FECoV-US isolated in 1970 and the potential minor parent as the Canine coronavirus. Our results provide a comprehensive molecular analysis of two novel FIPV isolates causing fatal disease in shelter cats from Long Island. Diagnostic surveillance with molecular characterization and sequencing analysis of circulating FIPV strains within animal shelters may help early detect unique emerging clinical and pathological manifestations of the disease and develop more targeted prophylactic and therapeutic approaches to control it.

Key Message: Melatonin increases Pb tolerance in P. ovata seedlings via the regulation of growth and stress-related phytohormones, ROS scavenging and genes responsible for melatonin synthesis, metal chelation, and stress defense. Lead (Pb) is a highly toxic heavy metal that accumulates in plants through soil and air contamination and impairs its plant growth and development. Because of its pharmaceutical importance, improvements in Plantago ovata yield against abiotic stresses are necessary. Melatonin (MEL) is a stress-alleviating biostimulator and our results showed a reduction in Pb induced phytotoxicity by enhancing plant growth attributes and balancing protective osmolytes. Pb-induced reactive oxygen species accumulation, including superoxide and peroxide free radicals and their mitigation through enzymatic antioxidants, was demonstrated in presence of MEL. Cell viability and Pb bioaccumulation were determined to understand the extent of cellular damage. Moreover, MEL increased secondary metabolite (flavonoids and anthocyanins) contents by 2-3-fold at the lowest Pb concentrations. Similar increases in the relative expression of genes (PoPAL and PoPPO), which are responsible for the production of non-enzymatic antioxidants, were observed. Notably, the upregulation of the PoCOMT gene up to 4-fold indicates increased melatonin production, as manifested in the phytomelatonin level. MEL supplementation also increased the auxin (IAA) level by 3-fold in the 100 µM Pb treatment group, while the abscisic acid (ABA) level decreased (1.4-fold) and the expression of PoMYB (a stress-related transcription factor) increased (up to 2.66-fold). Additionally, we found extreme downregulation (up to 18-fold) in the relative expression of PoMT 2 (a metal binding thiol compound) with melatonin treatment, which is otherwise upregulated (by 6-fold) during Pb stress. In the current study, these effects collectively revealed that MEL contribute to enhanced plant growth and Pb stress tolerance., Competing Interests: Declarations. Conflict of interest: The authors declare that they have no competing interests., (© 2025. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: Energy metabolism and pyroptosis are integral to the pathogenesis of diabetic nephropathy (DN). However, the precise roles of energy metabolism and pyroptosis in DN development remain unclear. This study aims to elucidate the roles of energy metabolism- and pyroptosis-related differentially expressed genes (EMAPRDEGs) in DN development., Methods: EMAPRDEGs were identified by querying the GeneCards and Gene Expression Omnibus (GEO) databases. Subsequent analyses included Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment, Gene Set Enrichment Analysis (GSEA), and Protein-Protein Interaction (PPI) network analysis. Additionally, mRNA-miRNA, mRNA-drug, and mRNA-transcription factor (TF) interaction networks were constructed. Differential expression and receiver operating characteristic (ROC) curve analyses were performed to evaluate the diagnostic potential of EMAPRDEGs. Immune cell infiltration in DN was assessed using the ssGSEA algorithm, and the expression levels of EMAPRDEGs in DN tissues were validated by quantitative real-time PCR (qRT-PCR)., Results: Thirteen EMAPRDEGs were identified, with GO and KEGG analyses indicating their involvement in energy metabolism pathways. GSEA revealed significant enrichment of these genes in biological pathways associated with diabetic nephropathy. PPI network analysis highlighted the central role of these genes within the relevant pathways. Predictive modeling demonstrated interactions between EMAPRDEGs, 69 miRNAs, and 117 TFs. Immune infiltration analysis showed substantial alterations in immune cell populations, with ADH1B and PC showing a significant correlation with natural killer cells and memory B cells. ROC curve analysis confirmed the diagnostic potential of EMAPRDEGs for diabetic nephropathy. qRT-PCR validated the expression patterns of CASP1 , IL-18 , PDK4 , and FBP1 , which were consistent with the bioinformatics predictions., Conclusion: Bioinformatics analysis identified 13 candidate EMAPRDEGs, among which CASP1 , IL-18 , PDK4 , and FBP1 emerge as potential biomarkers for diabetic nephropathy., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2025 Published by Elsevier Ltd.)

Background: Oxidative damage has been implicated in multiple neurodegenerative diseases, including epilepsy. Selenium, in the form of selenoproteins is an integral part of the human antioxidant defense system. Though a relationship between the altered selenium levels and epilepsy has been reported, limited evidence is available about the expression pattern of selenoproteins in epileptic patients., Objective: This study aimed to determine the serum selenium levels in idiopathic epileptic and healthy individuals. Expression profiling of selenoproteins (GPx1, TRxR1 and SEPW1) both at mRNA and protein levels was also evaluated., Methods: Serum selenium levels of 30 patients with idiopathic generalized epilepsy and their age and gender matched 30 healthy controls were measured. Protein levels of Serum Glutathione Peroxidase 1 (GPx1), Thioredoxin Reductase 1 (TRxR1) and Selenoprotein W (SEPW1) were estimated using ELISA. mRNA expression of GPx1, TRxR1 and SEPW1 were determined using qRT-PCR., Results: The mean values for serum selenium levels in cases and controls were 37.6 ± 2.0 µmol/ml and 38.9 ± 2.7 µmol/ml, respectively. Selenium levels in cases were significantly lower as compared to controls (p = 0.031). No statistically significant differences were observed between the serum levels of selenoproteins GPx1, TRxR1 and SEPW1 in epileptic patients and the healthy group. GPx1 and TRxR1 expression was found to be down regulated (0.34 and 0.13 folds respectively) whereas SEPW 1 was found to be 0.04 folds up regulated in epileptic patients compared to the healthy subjects., Conclusion: Selenium deficiency observed in epileptic patients suggests the association between serum selenium levels and epilepsy. This study provides the information about the selenium status in Pakistani population and helps in understanding the role of selenium in the prevention of epilepsy., Competing Interests: Declarations. Ethics approval and consent to participate: “This study was performed in line with the principles of the Declaration of Helsinki. Approval was granted by the Institutional review board, Dow University of Health Sciences, Karachi (IRB-1438/duhs/Approval/2020 dated 30th January-2020).” Written informed consent was obtained from all individual participants included in the study.” The design and ethical parameters of the study were approved by Institutional Review Board of DUHS, Karachi (Ref: IRB-1143/DUHS/Approval/2018/). Informed consent was attained by all study participants above 18 years of age. Under-aged patients such as children or individuals who were under the age of 18 were included with signed consent from parents and guardians. Anonymity of the data was safeguarded to preserve the privacy of the records. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Metabolic reprogramming, vital for cancer cells to adapt to the altered microenvironment, remains a topic requiring further investigation for different tumor types. Our study aims to elucidate shared metabolic reprogramming across breast (BRC), colorectal (CRC), and lung (LUC) cancers. Leveraging gene expression data from the Gene Expression Omnibus and various bioinformatics tools like MSigDB, WebGestalt, String, and Cytoscape, we identified key/hub metabolism-related genes (MRGs) and their interactions. The functional characteristics including survival parameters and expression of the key MRGs were analyzed and validated through Gene Expression Profiling Interactive Analysis 2 and qRT-PCR. In addition, we employed machine learning algorithms such as k-nearest neighbours (KNN), support vector regressor (SVR), and extreme gradient boosting (XGBoost) to assess MRGs' effectiveness in predicting overall patient survival. Among 11,384 DEGs analyzed, 540 overlapped across BRC, CRC, and LUC, with 46 MRGs and 20 key/hub MRGs involved in all studied cancer types. Of these, 11 key MRGs were prognostically significant. The qRT-PCR validation of key MRGs in specific cancer cell lines confirmed their expression profiles, with some showing cell-type-specific patterns. SVR exhibited remarkable accuracy in predicting overall survival, emphasizing its clinical utility. Our integrated approach combining bioinformatics analyses and experimental validations underscores the potential of MRGs as biomarkers for metabolic therapies, with machine learning models enhancing predictive capabilities for patient outcomes., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Carbapenem-resistant Klebsiella pneumoniae poses a severe risk to global public health, necessitating the immediate development of novel therapeutic strategies. The current study aimed to investigate the effectiveness of the green algae Arthrospira maxima (commercially known as Spirulina) both in vitro and in vivo against carbapenem-resistant K. pneumoniae. In this study, thirty carbapenem-resistant K. pneumoniae isolates were collected, identified, and then screened for their susceptibility to several antibiotics and carbapenemase production genes using PCR. Both bla KPC and bla OXA-48 genes were the most predominant detected carbapenemase genes in the tested isolates. The phytochemical profiling of A. maxima algal extract was conducted using LC-MS/MS in a positive mode technique. The minimum inhibitory concentrations (MIC) of the algal extract ranged from 500 to 1000 µg/mL. The algal extract also resulted in decreasing the membrane integrity and distortion in the bacterial cells as revealed by scanning electron microscope. The bioactive compounds that were responsible for the antibacterial action were fatty acids, including PUFAs, polysaccharides, glycosides, peptides, flavonoids, phycocyanin, minerals, essential amino acids, and vitamins. Moreover, A. maxima algal extract revealed an antibiofilm activity by crystal violet assay and qRT-PCR. A murine pneumonia model was employed for the in vivo assessment of the antibacterial action of the algal extract. A. maxima showed a promising antibacterial action which was comparable to the action of colistin (standard drug). This was manifested by improving the pulmonary architecture, decreasing the inflammatory cell infiltration, and fibrosis after staining with hematoxylin and eosin and Masson's trichrome stain. Using immunohistochemical investigations, the percentage of the immunoreactive cells significantly decreased after using monoclonal antibodies of the tumor necrosis factor-alpha and interleukin six. So, A. maxima may be considered a new candidate for the development of new antibacterial medications., Competing Interests: Declarations. Ethics approval and consent to participate: The study was conducted according to the guidelines approved by the Research Ethics Committee of the Faculty of Pharmacy, Tanta University. The approval code is TP/RE/4/24 p-002. Consent to participate is not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Severe COVID-19 and MIS-C are rare but serious outcomes associated with SARS-CoV-2 infection. The onset of MIS-C often involves the gastrointestinal system, suggesting a potential connection with gut microbiota. This study aims to compare the gut microbiota of children with severe COVID-19 and those with MIS-C using various biomolecular approaches. Gut microbiota composition and specific microbial modulations were analyzed using fecal samples collected at hospital admission. The study included hospitalized patients (mean age 6 ± 5 years) diagnosed with severe COVID-19 (37 patients) or MIS-C (37 patients). Microbial differences were assessed using both NGS and qRT-PCR methodologies. In 75% of cases, pharmacological treatments included antibiotics and corticosteroids, which influenced the microbiota composition. Early age was found to have the most significant impact on microbiota diversity. Significant differences in alpha and beta diversity were observed between COVID-19 and MIS-C patients, particularly concerning low-abundance species. Levels of Bacteroides spp., Bifidobacterium spp., and Akkermansia muciniphila were comparable between groups, while an increased activity of Bifidobacterium spp. was noted in children with positive fecal samples ( p = 0.019). An in-depth evaluation of lesser-known gut species may be key to reducing the risk of severe outcomes and developing microbiota-based biomarkers for the early diagnosis of MIS-C.

Introduction: In this study, we sought to determine the sub-chronic toxicological effects of AEC on zebrafish embryos., Methods: We utilized fish early life stage (FELS) and fish embryo toxicity (FET) tests, vascular, neurological, and renal transgenic zebrafish lines, and gene expression anal-ysis of the zebrafish tissue., Results: In the FET tests, AEC caused several abnormalities in the larvae, with the LC50 at 24 hpf being 4.076 ± 0.221 mg/L and 3.296 ± 0.127 mg/L at 96 hpf. In the FELS test, AEC was shown to be lethal following 16 days of exposure at 0.5 mg/L, 1 mg/L and 2 mg/L. Some of the transgenic zebrafish lines exhibited slight changes in fluorescent signaling pat-terns after exposure to AEC at 1 mg/L and 2 mg/L. Notable results of the gene expression analysis revealed: gpx4b and got2 were downregulated in the liver; HIF1a was downregulated at 0.25 mg/L and 0.5 mg/L concentrations, NOTCH1a and fli-1 genes were downregulated at all concentrations, and A2b was upregulated in the vasculature; a1T, ngn1, elavl3, syn2a, mbp, gap43 were down-regulated in the nervous system; and wt1b was downregulated in the kidney., Discuccion: Altogether, the results of our study indicate the potential for AEC to cause harm to organisms., Competing Interests: Declaration of conflicting interestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Visceral leishmaniasis, a life-threatening vector-borne illness that disproportionately affects children and elderly immunocompromised people, is a primary tropical neglected disease. No apoptotic partner proteins have yet been reported in Leishmania donovani, while their identification could contribute to knowledge on parasite cell death and the establishment of alternative therapeutics. We searched for mammalian Bcl-2 family protein orthologs and found one anti-apoptotic and two pro-apoptotic orthologs in L. donovani. A pro-death aquaporin protein, due to its characteristic BH3 domain known to interact with pro-apoptotic proteins in mammalian Bcl-2 family proteins, was also included in this study. Molecular docking and molecular dynamics simulations were conducted to assess protein-protein interactions between the identified apoptotic proteins and mimic mammalian intrinsic apoptotic pathways. The results showed that both pro-apoptotic proteins interacted with the hydrophobic pocket of the anti-apoptotic ortholog, forming a stable complex. This interaction may represent a critical event in an apoptotic pathway in L. donovani. To further characterise it, we used CRISPR-Cas9 approaches to target the identified proteins. Pure knocked population mutants, and episomal over-expressing mutant cells were exposed to apoptotic stimuli. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and quantitative expression profiling suggested that these proteins are involved in the parasite's apoptosis and could play a role in its survival., (© K. Kumar et al., published by EDP Sciences, 2025.)

The aging immune system undergoes significant changes, leading to a state known as immunosenescence. Understanding the molecular mechanisms underlying immunosenescence is crucial for developing targeted interventions to enhance immune functions in older individuals. This bio-protocol review focuses on the application of quantitative reverse transcription-polymerase chain reaction (qRT-PCR) for the mRNA quantification of cytokine-inducible SH2-containing protein (CISH), an immune regulator overexpressed in T-cell responses from older adults. We outline a comprehensive protocol for the quantitative assessment of CISH mRNA expression, providing a valuable tool for researchers investigating immunosenescence., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Developing melon genotypes with resistance to Fusarium oxysporum f. sp. Melonis-(FOM) race1.2 is a major goal in any breeding program. In this study, we identified the role of 11 gene markers that contribute to polygenic resistance during the FOM1.2-melon interaction. qRT-PCR analysis elucidated upregulation of candidate marker genes AMT, DXPR, Fom-2, GLUC, GalS, GRF3, MLO, PRK, RuBlsCo, TLP and WRKY in resistant 'Shante-F1' and 'Khatouni', and susceptible 'Shante-T' and 'Shahabadi' at 7, 14 and 21 days post-inoculation (dpi). We also studied changes in defence-related enzyme activity: chitinase (CHI), β-1,3-glucanase (GLU) and peroxidase (POX) in melon roots. AMT, GLUC and DXPR transcripts were upregulatied in leaf and root tissues of the resistant 'Shante-F1' and 'Shahabadi'. Transcript levels for GalS and GRF3 increased 6.77- and 6.83-fold in roots of 'Shante-F1' at 7 dpi, whereas in PRK, TLP and WRKY theye increased by 7.84-, 5.15- and 12.26-fold at 14 dpi, respectively. However, transcript levels increased by 5.18-fold for Fom-2 and 8.46-fold for MLO at 21 dpi. Also, RBC transcript level peaked at 14 dpi with 4.9-fold increase in leaves of resistant genotypes, whereas AMT increased 2.94-fold at 21 dpi, and GLUC and DXPR increased 7.11- and 2.91-fold at 14 dpi in 'Shante-F', respectively. Defence-related-enzyme activity was also upregulated three-fold in resistant varieties. The dynamic shifts in the melon transcriptome induced by FOM1.2 emphasize that resistance mechanisms are predominantly regulated through signalling pathways involving CHI, GLU, and POX defence response. Surprisingly, the AMT gene, basically resistant to downy mildew, Pseudoperonospora cubensis; GLUC, MLO and PRK resistant to powdery mildew (Sphaerotheca fusca); TLP and WRKY resistant to Phytophthora blight (Phytophthora capsici); and GRF3 and RBC resistant to root knot nematodes (Meloidogyne spp.) were upregulated in resistant genotypes, indicating a dual role of these genes in resistance to more than one disease at a time., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

FOXO transcription factors belong to the forkhead protein family and are distinguished by their unique forkhead (FKH) DNA-binding domain. In the realm of mammals, four FOXO paralogs are recognized: FOXO1, FOXO3, FOXO4, and FOXO6. These paralogs are evolutionary counterparts of the daf-16 gene discovered in the nematode C. elegans. A key feature shared by these paralogs is a consensus binding site known as the DAF-16 family protein-binding site (DBE: 5'-TTGTTTAC-3'). The functional outcome of FOXO transcription factors primarily hinges on their affinity for these specific binding sites within the promoters of their target genes. Nevertheless, it is worth noting that many of these target genes exhibit tissue-specific expression patterns. Consequently, there is not a single FOXO target gene whose expression can reliably serve as a universal indicator of FOXO activity across all cell types and tissues or in response to all stimuli. In light of these considerations, we present a collection of target genes that, when collectively assessed, can accurately gauge FOXO activation. In this chapter, we outline a specific protocol for utilizing quantitative reverse transcription polymerase chain reaction (qRT-PCR) to measure the expression levels of these genes., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Bioactive glass (BAG) has emerged as an effective bone graft substitute due to its diverse qualities of biocompatibility, bioactivity, osteoblast adhesion and enhanced revascularization. However, inferior osteogenic capacity of BAG compared to autologous bone grafts continues to limiting it's wide-spread clinical applications towards repairing of bone fractures and healing. In this study, we have fabricated BAG composites with 0.5 to 2 wt% bismuth ferrite (BF, a multiferroic material) with an aim to generate in-situ electrical charges pertinent to early-stage bone regeneration thus mimicking natural bone, which is a piezoelectric material. The fabricated BAG composites were characterised in terms of microstructures, phase analysis, remanent polarization, wettability and subsequently evaluated for in vitro cell proliferation and osteogenesis with and without magnetic field exposure (200 mT, 30 min./day). Pre-osteoblast cells from mice (MC3T3-E1) seeded on these composites exhibited excellent cell growth without any cytotoxicity, which is further supported by FITC/DAPI staining and a live/dead assay. The results of Alizarin Red S assay and increased levels of Alkaline Phosphatase (ALP) activity, at 21 days of culture, suggest that the BAG-BF composites promote in vitro osteogenic differentiation of pre-osteoblast cells. The enhanced osteogenesis of BAG-BF composites was also confirmed through qRT-PCR analysis, which showed rapid upregulation of osteoblastogenic specific genes namely RunX-2, Collagen-1, Bone Sialo Protein, and ALP after 21 days. Additionally, the osteogenic differentiation was assessed by the Western Blot technique, which revealed significantly higher band intensity of osteogenic markers in BAG-1.5 BF and BAG-2 BF composites than pure BAG. These findings clearly demonstrate that in-situ electrical stimulation and osteoconductive capacity of BF reinforced BAG composites have positive impact on osteoblast cell development, bone formation, and healing., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: Neural tube defects (NTDs) are defined as an incomplete closure of the neural tube (NT), with a prevalence of 1.2 per 1000 live births around the world. Methylation of the maternally imprinted gene Insulin-like growth factor 2 (IGF2) is one of the epigenetic mechanisms that contribute significantly to the development of NTDs. Its important functions are the fetal growth and metabolism, as well as cell division and differentiation., Objectives: In a couple of studies, individual changes in IGF2 with single nucleotide variants (SNVs) have been found to be linked with NTDs as well as with other diseases. However, there is more need of studying so that we can better understand precisely this mechanism at the molecular level. In this study we tried to investigate the relationship between genetic variants of IGF2 and NT closure by screening all SNVs of IGF2 using both in silico methods and clinical experiments on human samples., Methods: We enrolled 98 consecutive mothers carrying fetuses with NTDs as cases and 76 age-matched mothers with healthy babies as controls. Following an in silico analysis of the IGF2 gene, we designed three primers targeting a total of 180 exonic, 579 intronic, and 46 variants in the 3' untranslated region (3' UTR) for polymerase chain reaction (PCR) and subsequent sequencing. Statistical analyses were performed to assess associations between these genetic variants and NTDs. Additionally, quantitative real-time PCR (qRT-PCR) was conducted to analyze the mRNA expression levels of the target gene., Results: This is the first study to enlist 11 non-synonymous variants [rs750845881(D93N), rs1290256492(P87H), rs755066389(P87T), rs150610908(G65D), rs762200142(R64C), rs868067982(G49C), rs778465733(R48L), rs1018841144(R48C), rs1057518115(C33R), rs112276039(C22R), rs1240151267(L16W)] in the regulation of IGF2 gene functions through in silico analysis. Further clinical validation in maternal samples showed a significant association between rs3213225 (C > T) (OR-2.076; CI-1.123-3.840; p-0.0209) and rs734351 (C > T) (OR-2.148; CI-1.167-3.956; p-0.0148) with increased disease risk. RNA expression study validated and supported all of these SNVs, indicating a strong correlation with NTD., Conclusions: Our research strongly suggests that genetic variations in IGF2 may help explain the risk association with NTDs and could be valuable for diagnostic and therapeutic applications., (© 2025 Wiley Periodicals LLC.)

Introduction: Side by side with tooth decay, periodontitis remains one of the most common oral diseases and is increasingly recognized as a serious public health concern worldwide., Objectives: The present study aims at comparing the levels of 5 specific miRNAs (miR-29b-3p, miR-34a-5p, miR-155-5p, miR-181a-5p, and miR-192-5p) in patients with periodontal disease and healthy controls., Methods: The pathogenic mechanism is related to the activation of immune response and significant alteration of coding and noncoding genes, including miRNA. The study includes 50 subjects (17 with periodontal disease and 33 healthy controls) with a mean age of 45.3 y. In both periodontitis patients and healthy controls, a panel of 5 miRNAs (miR-29b-3p, miR-34a-5p, miR-155-5p, miR-181a-5p, and miR-192-5p) is examined by determining their expression levels with quantitative reverse transcription polymerase chain reaction., Results: The periodontitis patients express high levels of all the investigated miRNAs. Receiver operating characteristic curve analysis shows an area under the curve (AUC) of 0.69 to 0.74 for individual transcripts with the highest AUC value observed for miR-192, followed by miR-181a., Conclusions: The study indicates that the 5-miRNA panel can be used as biomarker for periodontitis. In this way, all implantology procedures and treatment options for patients diagnosed with periodontitis can be improved for better long-term results, predictability, and follow-up frequency., Knowledge Transfer Statement: The discovery of a miRNA panel as a potential biomarker for periodontitis offers major opportunities for practical application. Our study can improve diagnostic accuracy; researchers can develop new theories on molecular mechanisms and biomarker discovery., Competing Interests: Declaration of Conflicting InterestsThe authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

The N6-methyladenosine (m 6 A) methyltransferase METTL3 has been demonstrated to function in mediating m 6 A modification, but its role in ischemic stroke (IS) has not been fully elucidated. This study aimed to explore the downstream mechanism of METTL3-mediated m 6 A modification in IS. GSE16561 and GSE22255 were downloaded from the Gene Expression Omnibus database for analysis of differentially expressed genes (DEGs), and it was found that METTL3 mRNA was downregulated in IS. Then quantitative real-time polymerase chain reaction was used to verify the downregulation of METTL3 mRNA in the peripheral blood of IS patients and the cortexes of transient middle cerebral artery occlusion mice. By combining DEGs with the m 6 A-downregulated genes in GSE142386 which performed methylated RNA immunoprecipitation sequencing (MeRIP-seq) on METTL3-deficient and control endothelial cells, a total of 131 genes were identified as the METTL3-mediated m 6 A-modified genes in IS. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that the genes were mainly involved in cytokine-cytokine receptor interaction, MAPK signaling pathway and NF-kappa B signaling pathway. CTSS and SBK1 were further screened as the key METTL3-mediated m 6 A-modified genes by random forest model and PCR validation. The ROC curve analysis showed that the combination with CTSS and SBK1 was of good diagnostic value for IS, with the AUC of 0.810, sensitivity of 0.780, and specificity of 0.773. Overall, we found that METTL3-mediated m 6 A modification may influence the occurrence and development of IS by participating in inflammation-related biological processes, and two key m 6 A-modified genes mediated by METTL3 (CTSS and SBK1) can be used as diagnostic biomarkers for IS., Competing Interests: Declarations. Conflict of interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Ethical Approval: This study was approved by the ethical committee of Guangxi Medical University and the Ethics Committee of Guangxi University of Chinese Medicine, and performed in accordance with the Declaration of Helsinki. Written informed consent was obtained from each participant after the full description of the study. All animal experiments were in compliance with the National Institutes of Health Guide for Care and Use of Laboratory Animals and were approved by the Animal Ethics Committee of Guangxi Medical University. Consent to Participate: Informed consent was obtained from all individual participants included in the study., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

In our laboratory, we study thrombopoiesis and hemostasis using zebrafish as a model organism to unravel the mechanisms of differentiation and development of thrombocytes. We have shown in our earlier work that thrombocytes are functional equivalents of platelets and have transcriptional machinery similar to megakaryocytes. We recently found evidence that hox genes play a role in their development. We used piggyback gene knockdown and thrombocyte quantification assays to understand the influence of these ancient developmental regulators on thrombopoiesis. In this chapter, we describe methods used to discover these hox genes., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Blackgram is an important short-duration grain legume, but its yield is highly affected by various stresses. Among biotic stresses, yellow mosaic disease (YMD) is known as a devastating disease that leads to 100% yield loss under severe conditions. The cultivated lines possess resistance, but exploring more diverse sources of resistance may be useful for pyramiding to improve the durability of said resistance. Some wild Vigna species have potentially demonstrated a high level of resistance. R-genes, including gene families of leucine-rich repeats (LRRs) and leucine-rich repeat receptor-like kinases (LRR-RLKs), are known for modulating the resistance in plants against various biotic stresses. The first comprehensive analysis of the LRR and LRR-RLK gene families in mungbean is reported in the present study. A total of forty-six candidate genes were identified and grouped into eight clades. Protein motif analysis showed that the "Pkinase domain" and "LRR domains" were conserved in most of the R-proteins. The expression of candidate genes viz. VrNBS&#95;TNLRR-8 , VrLRR&#95;RLK-20 , VrLRR&#95;RLK-17 , and VrLRR&#95;RLK-19 demonstrated significantly up-regulated expression upon YMD infection in control and salicylic acid-primed (SA-primed) plants. The analysis provides insight into the diversity and robust candidate genes for functional studies modulating YMD resistance altered by salicylic acid.

The NAC gene family plays a crucial role in plant growth, development, and responses to biotic and abiotic stresses. Paspalum Vaginatum , a warm-season turfgrass with exceptional salt tolerance, can be irrigated with seawater. However, the NAC gene family in seashore paspalum remains poorly understood. In this study, genome-wide screening and identification were conducted based on the NAC ( NAM ) domain hidden Markov model in seashore paspalum, resulting in the identification of 168 PvNAC genes. A phylogenetic tree was constructed, and the genes were classified into 18 groups according to their topological structure. The physicochemical properties of the PvNAC gene family proteins, their conserved motifs and structural domains, cis-acting elements, intraspecific collinearity analysis, GO annotation analysis, and protein-protein interaction networks were analyzed. The results indicated that the majority of PvNAC proteins are hydrophilic and predominantly localized in the nucleus. The promoter regions of PvNACs are primarily enriched with light-responsive elements, ABRE motifs, MYB motifs, and others. Intraspecific collinearity analysis suggests that PvNACs may have experienced a large-scale gene duplication event. GO annotation indicated that PvNAC genes were essential for transcriptional regulation, organ development, and responses to environmental stimuli. Furthermore, the protein interaction network predicted that PvNAC73 interacts with proteins such as BZIP8 and DREB2A to form a major regulatory hub. The transcriptomic analysis investigates the expression patterns of NAC genes in both leaves and roots under varying durations of salt stress. The expression levels of 8 PvNACs in roots and leaves under salt stress were examined and increased to varying degrees under salt stress. The qRT-PCR results demonstrated that the expression levels of the selected genes were consistent with the FPKM value trends observed in the RNA-seq data. This study established a theoretical basis for understanding the molecular functions and regulatory mechanisms of the NAC gene family in seashore paspalum under salt stress.

Background: As a master immune system regulator, transforming growth factor β1 (TGF-β1) is closely linked to the complicated pathophysiology and development of systemic sclerosis (SSc), a multisystem fibrotic disease., Objective: We aim to evaluate the transcriptional levels of TGF-β1 mRNA in PBMCs, assess the TGF-β1 serum levels of SSc patients, and compare them with those of healthy subjects., Methods: PBMCs were isolated from whole blood of 50 SSc patients and in 30 healthy controls. After total RNA was extracted from isolated PBMCs, complementary DNA (cDNA) synthesis was performed. Afterward, the expression of TGF-β1 mRNA was assessed using quantitative real-time PCR using the SYBR Green, GAPDH, and TGF-β1 specific primers. The serum levels of TGF-β1 were determined using a commercially available ELISA kit., Results: There was a significant upregulation of TGF-β1 relative expression (p < 0.0001), when SSc patients were compared to the control group. The diffuse subgroup was more common in patients with elevated TGF-β1 mRNA expression (p < 0.0001). However, an insignificant difference was observed between the disease subsets of SSc. Serum TGF- β1 levels were upregulated in SSc patients (78.35 ± 23.16) compared to healthy subjects (61.06 ± 15.90), and were considerably higher in SSc patients with ILD (p < 0.01) and positive anti-topo-Isomerase antibody (p < 0.0001)., Conclusion: In patients with SSc, elevated levels of TGF-β1 in serum and their correlation with clinical symptoms imply that this cytokine may serve as a marker for fibrotic and vascular involvement in SSc., Competing Interests: Declarations. Ethical approval: The Institutional Ethics Committee (IEC-SKIMS) approved the study vide protocol number #RP77/2019. Consent to participate: After giving their informed consent and having the right to withdraw at any moment, participants willingly decided to participate in this study. Consent for publication: The patients who participated in this study gave written informed consent for participation and publication of this study. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Plant viruses need host factors to complete their life cycle. The eukaryotic translation initiation factor 4E (eIF4E) and its isoforms are the host factors essential for infection by plant viruses in the genus Potyvirus. The present study describes the characterization of eIF4E gene in banana cultivar Nendran, and its interaction with the viral genome-linked protein (VPg) of banana bract mosaic virus (BBrMV). The in-silico analysis of 853 bp coding mRNA region of eIF4E revealed the presence of five exons. The deduced amino acid sequence had 99.58% identity with the predicted protein sequence of the eIF4E-1 in Musa acuminata subspecies malaccensis. Expression profiling of eIF4E gene in banana cultivar Nendran using qRT-PCR revealed up to 16.9-fold up-regulation of the gene in response to BBrMV infection, indicating its role in disease development. The three-dimensional models of the eIF4E and VPg proteins were generated to deduce the precise interaction between the two proteins. Using the online server ZDOCK, ten interaction complexes were obtained. In nine out of the ten complexes, the maximum number of residues of the eIF4E interacted with the Tyrosine-63rd residue of VPg. The corresponding nucleotides were identified as the potential targets for editing the host eIF4E gene to impart potyvirus resistance., Competing Interests: Declarations. Competing Interests: The authors declare that they have no conflict of interest in the publication., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Ovarian cancer (OC) is often associated with an unfavorable prognosis. Given the crucial involvement of lysosomes in tumor advancement, lysosome-related genes (LRGs) hold promise as potential therapeutic targets., Methods: To identify differentially expressed lysosome-related genes (DE-LRGs), we performed a matching analysis between differentially expressed genes (DEGs) in OC and the pool of LRGs. Genes with prognostic significance were analyzed using multiple regression analyses to construct a prognostic risk signature. The model's efficacy was validated through survival analysis in various cohorts. We further explored the model's correlation with clinical attributes, tumor microenvironment (TME), mutational patterns, and drug sensitivity. The quantitative real-time polymerase chain reaction (qRT-PCR) validated gene expression in OC cells., Results: A 10-gene prognostic risk signature was established. Survival analysis confirmed its predictive accuracy across cohorts. The signature served as an independent prognostic element for OC. The high-risk and low-risk groups demonstrated notable disparities in terms of immune infiltration patterns, mutational characteristics, and sensitivity to therapeutic agents. The qRT-PCR results corroborated and validated the findings obtained from the bioinformatic analyses., Conclusions: We devised a 10-LRG prognostic model linked to TME, offering insights for tailored OC treatments., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Sepsis is one of the leading causes of death among seriously ill patients worldwide, affecting more than 30 million people annually and accounting for 1-2% of hospitalizations. By analyzing gene expression omnibus (GEO) data set, our team explored the relationship between m6A methylation gene and poor prognosis of sepsis. The purpose of this present study is to examine new detection markers for patients with poor prognosis, provide theoretical basis for timely intervention and improve the survival rate of patients. First, GSE54514 transcriptome data were extracted from the GEO database 31 patients with sepsis related death and 72 sepsis survivors. Key genes were screened from differentially expressed genes (DEGs), least absolute shrinkage and selection operator (LSAAO) and random forest (RF). And then, METTL3, WTAP and RBM15 were further verified by quantitative reverse transcription PCR (qRT-PCR). The constructed nomogram model showed high accuracy in predicting death. These three genes are mainly involved in chemokine signaling pathway, differentiation of monocytes and T cells, and phagocytosis of immune cells. The analysis showed that a high m6A score subtype is linked to lower immunosuppression and higher survival rates in clinical samples, suggesting better immune responses and outcomes for these patients. Finally, the protective effect of METTL3 in sepsis was demonstrated in mouse sepsis model applied with METTL3 inhibitor, by conducting cell flow cytometry analysis, enzyme-linked immunosorbent assay (ELISA) and hematoxylin-eosin (HE) staining. In conclusion, these findings provide potential biomarkers and targets for early precision diagnosis and treatment., Competing Interests: Declarations. Institutional review board statement: This study was approved by the Animal Care and Use Committee of Anhui Medical University (Ethics Committee Approval Number: LLSC20200404). Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Quantitative real-time PCR (qRT-PCR) is an essential tool for analyzing and selecting stable reference genes. In order to screen for suitable reference genes under high-temperature stress conditions in Arabidopsis , this study measured the relative expression levels of 17 candidate reference genes using qRT-PCR. Among these, four are traditional reference genes, while the remaining thirteen are candidate reference genes with no previous reports on their use as reference genes. The expression stability of the candidate reference gene expression was analyzed and evaluated using five methods: ΔCt, geNorm, NormFinder, BestKeeper, and RefFinder. The results indicated that the LHCB4.1 and LHCB5 genes displayed the highest stability in expression under high-temperature stress conditions. To verify the stability of the reference genes, we treated Arabidopsis with high-temperature stress, used the selected LHCB4.1 and LHCB5 as references, and analyzed the expression of the heat-responsive gene HSFA2 using qRT-PCR. The results showed that when LHCB4.1 and LHCB5 were used individually or in combination as reference genes, the relative expression of HSFA2 significantly increased and remained consistent under high-temperature treatment. This indicates that both LHCB4.1 and LHCB5 are suitable reference genes for qRT-PCR analysis in Arabidopsis exposed to high-temperature stress. The novel reference genes identified in this study will serve as a reliable reference standard for gene expression studies in Arabidopsis under high-temperature stress, thereby enhancing the accuracy and comparability of experimental data.

Isolation of high-quality RNA from abaca is very challenging due to the presence of polyphenols, polysaccharides, and its high fiber content. In this study, we compared six extraction methods across three tissue types and different developmental stages ( in-vitro -grown young versus field-grown mature tissue). The Invitrogen PureLink RNA kit proved to be the most efficient in extracting RNA from young abaca tissues (leaves, pseudostem, and corm). The quality of RNA extracted from young tissues was further assessed by RNA-seq applications, with raw sequencing reads mapping back to the M. textilis reference genome at rates of 86.0%-90.4%. The SDS-TRIzol-method modified with an added on-column DNAse I treatment was used to extract RNA from mature tissues (leaves, midrib, and pseudostem). RNA isolated from five M. textilis cultivars and across three mature tissue types showed RNA yield per 100 mg of fresh weight ranges from 0.57 to 10.94 µg and RNA integrity number (RIN) scores of more than 7.0 for all tissue types. Our improved SDS-Trizol method for RNA extraction described here is simple and yields good quality RNAs from mature abaca tissues while the PureLink RNA kit is suitable for extracting RNA from young abaca samples amenable to RT-qPCR and next-generation sequencing studies.

Anoectochilus roxburghii (Wall.) Lind. has significant medicinal and economic value, and the social demand for this species is increasing annually. Laser light sources have different luminescent mechanisms compared with ordinary light sources and are also important factors regulating the synthesis of functional metabolites in A. roxburghii. However, the regulatory mechanism through which A. roxburghii responds to blue laser light has not been investigated. Previous studies have shown that blue-laser treatment results in more red leaves than blue- or white-light treatment. Here, the differences in the effects of laser treatment on A. roxburghii were analyzed by transcriptome sequencing. Gene Ontology analysis revealed that the membranes, calcium ion binding, brassinosteroid-mediated signaling pathway and response to salicylic acid play important roles in the response of A. roxburghii to blue laser light. Kyoto Encyclopedia of Genes and Genomes analysis revealed the involvement of multiple pathways in the response to blue-laser treatment, and among these, beta biosynthesis, flavone and flavonol biosynthesis, thiamine metabolism, limonene and pinene biosynthesis, and peroxisomes play core roles. Cytoscape interaction analysis of the differentially expressed miRNA targets indicated that novel&#95;miR&#95;66, novel&#95;miR&#95;78 and novel&#95;miR&#95;212 were most likely involved in the effect of blue laser light on A. roxburghii. Metabolic content measurements showed that blue laser light increased the beet red pigment, thiamine, total flavonoid and limonene contents, and qPCR analysis confirmed that novel&#95;miR&#95;21, novel&#95;miR&#95;66, novel&#95;miR&#95;188 and novel&#95;miR&#95;194 might participate in the blue-laser signaling network through their target genes and thereby regulate the functional metabolite accumulation in A. roxburghii. This study provides a scientific basis for high-yield A. roxburghii production., (© The Author(s) 2024. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Plant plasma membrane (PM) H + -ATPase functions as a proton-motive force by exporting cellular protons to establish a transmembrane chemical gradient of H + ions and an accompanying electrical gradient. These gradients are crucial for plant growth and development and for plant responses to abiotic and biotic stresses. In this study, a comprehensive identification of the PM H + -ATPase gene family was conducted across four cotton species. Specifically, 14 genes were identified in the diploid species Gossypium arboreum and Gossypium raimondii , whereas 39 and 43 genes were identified in the tetraploid species Gossypium hirsutum and Gossypium barbadense , respectively. The characteristics of this gene family were subsequently compared and analyzed using bioinformatics. Chromosomal localization and collinearity analyses elucidated the distribution characteristics of this gene family within the cotton genomes. Gene structure and phylogenetic analyses demonstrated the conservation of this family across cotton species, whereas the examination of cis -acting elements in gene promoters highlighted their involvement in environmental stress and hormone response categories. An expression profile analysis revealed eight genes whose expression was upregulated under salt stress conditions, and quantitative real-time PCR results suggested that the cotton PM H + -ATPase genes may play crucial roles in conferring resistance to salt stress. These findings establish a robust foundation for subsequent investigations into the functions of cotton PM H + -ATPase genes and may offer valuable insights for selecting genes for resistance breeding programs.

This paper outlines a practical method for validating quantitative-qualitative techniques used to detect genetic material through qRT-PCR, specifically focusing on SARS-CoV-2 testing and adhering to ISO/IEC 17025:2018 accreditation standards. Despite the prevalence of quantitative-qualitative screening in genetic testing, comprehensive validation guidelines remain a notable gap in the field. Such guidelines could be applied to other molecular testing areas that rely on these techniques, particularly those involving sample handling, automated extraction, and amplification processes, which can significantly impact results. This work describes the statistical approaches behind qRT-PCR protocols, followed by a technical characterization profile of the validation process. Modifications to the gold standard method allowed us to establish a technical limit of detection (LOD) of 5,09 copies/reaction at a 95 % confidence interval., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors. Published by Elsevier Ltd.)

Background and Purpose: Chlorogenic acid (CA) is a natural chemical that promises antiaging activity against photoaging skin damage. This research examined CA activities in mitigating skin photoaging., Experimental Approach: UV-exposed human skin fibroblast cells were subjected to CA at 6.25, 12.5, and 25 μg/mL. The protein levels of cell secretion, such as cyclooxygenase (COX)-2, nitric oxide (NO), and interleukin (IL)-6 were measured using ELISA and colorimetry methods. Meanwhile, the mRNA expressions of glutathione peroxidase (GPX)-1, tissue inhibitor metalloproteinase (TIMP)-1, matrix metalloproteinase (MMP)-1, caspase (CASP)-3, CASP-8, and fibroblast growth factor (FGF)-2 were quantified using the qRT-PCR method., Findings/results: CA treatment reduced inflammatory and aging biomarkers. CA at 6.25 μg/mL lowered NO, COX-2, and IL-6 levels to 89.44 μmol/L, 8.10 ng/mL, and 62.75 pg/mL, respectively. CA at 25 μg/mL resulted in the most significant down-regulation of MMP-1, CASP-3, and CASP-8 genes' expression (3.27, 1.25, and 3.59, respectively). Furthermore, treatment with CA at 25 µg/mL demonstrated the most notable activity in up-regulating antioxidant markers, specifically GPX-1, and extracellular matrix (ECM) integrity markers, including TIMP-1 and FGF-2 genes' expression., Conclusion and Implications: CA imposes its anti-aging activity by decreasing inflammatory and aging biomarkers, and increasing cellular antioxidant and ECM integrity., Competing Interests: The authors stated that there were no conflict of interest in this study., (Copyright: © 2024 Research in Pharmaceutical Sciences.)

Potato ( Solanum tuberosum ) is the fourth largest staple food crop globally. However, potato cultivation is frequently challenged by various diseases during planting, significantly impacting both crop quality and yield. Pathogenic microorganisms must first breach the plant's cell wall to successfully infect potato plants. Cellulose, a polysaccharide carbohydrate, constitutes a significant component of plant cell walls. Within these walls, cellulose synthase ( CesA ) plays a pivotal role in cellulose synthesis. Despite its importance, studies on StCesAs (the CesA genes in potato) have been limited. In this study, eight CesA genes were identified and designated as StCesA1-8 , building upon the previous nomenclature ( StCesA1-4 ). Based on their phylogenetic relationship with Arabidopsis thaliana, these genes were categorized into four clusters (CesA I to CesA IV). The genomic distribution of StCesAs spans seven chromosomes. Gene structure analysis revealed that StCesAs consist of 12 to 14 exons. Notably, the putative promoter regions harbor numerous biologically functional cis -acting regulatory elements, suggesting diverse roles for StCesAs in potato growth and development. RNA-seq data further demonstrated distinct expression patterns of StCesAs across different tissues. Additionally, quantitative real-time PCR (QRT-PCR) results indicated significant up-regulation of StCesA5 expression under biotic stresses, implicating its potential involvement in potato disease resistance., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Gong, Ma, Dusengemungu and Feng.)

Key Message: Fusarium oxysporum disrupts redox homeostasis in Vigna mungo, likely by interfering with salicylic acid signaling, which can be ameliorated by boosting PAL and its related pathways via salicylic acid pretreatment. Fusarium oxysporum, a widespread soil-borne fungus, significantly threatens global crops. This study centers on elucidating the infection strategies employed by F. oxysporum against a new and underexplored host Vigna mungo, a leguminous crop of high agronomic value, and the defense mechanisms that can be activated against the infection, aiming to uncover how these responses can be leveraged to develop potential countermeasures. Building on prior work demonstrating the in vitro antifungal efficacy of phytohormones, including salicylic acid (SA), this study further investigates SA pretreatment at 100 µM, which previously reduced reactive oxygen species (ROS) and improved germination under Fusarium stress. Through a comprehensive analysis of V. mungo plants pretreated with SA and subjected to F. oxysporum infection, we observed that fungal exposure reduced growth, chlorophyll content, and levels of proteins, phenolics and flavonoids, while increasing stress markers and antioxidant activity. SA pretreatment mitigated these effects by boosting antioxidant molecules and activating the phenylalanine ammonia-lyase (PAL) pathway, thereby enhancing endogenous SA and ROS scavenging. Furthermore, qRT-PCR analysis confirmed SA-mediated upregulation of antioxidant (catalase and peroxidase), fungal stress response genes ((pathogenesis-related gene 4 (PR4) and defensin (DEF)) and SA synthesis and regulator genes (PAL and WRKY70) involved in plant systemic resistance, while LC-MS data revealed an altered metabolic profile with increased phytoalexins and antioxidants synthesis. Overall, SA pretreatment confers resistance against F. oxysporum in V. mungo by modulating endogenous SA and metabolic profile to activate key defense pathways and redox homeostasis, highlighting its potential in plant defense strategies and reinforcing our proposed model of SA action., Competing Interests: Declarations. Conflict of interest: The authors declare no relevant financial or nonfinancial conflicts of interest. Ethical approval: Ethical approval was not necessary for this study. Consent to publish: All the authors have provided consent for publication., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)