BioTech

15 pages, 1726 KiB

Open AccessArticle

Enhancing In Vitro Regeneration in Three Sweet Potato Genotypes: Interplay Between Disinfectant, Explant Age, and Genotype

by El Hadj Hussein Tapily, Kan Modeste Kouassi, Marius Konan Kouassi, John Steven S. Seka, Fidèle Tiendrébéogo and Justin S. Pita

BioTech 2025, 14(3), 63; https://doi.org/10.3390/biotech14030063 - 19 Aug 2025

Viewed by 569

Abstract

Regenerating sweet potato from field-derived plant material requires careful management of several critical factors, including the effectiveness of the disinfectant, the age of the explant, and the genotype used. In this context, establishing a reliable aseptic protocol is essential for successful in vitro [...] Read more.

Regenerating sweet potato from field-derived plant material requires careful management of several critical factors, including the effectiveness of the disinfectant, the age of the explant, and the genotype used. In this context, establishing a reliable aseptic protocol is essential for successful in vitro culture. This study aimed to assess the effects of two disinfectants (sodium hypochlorite and mercuric chloride), three sweet potato genotypes (Nakabo, Boyapleu, and Irene), and three explant ages (2, 3, and 4 weeks) on clean culture establishment and regeneration efficiency from nodal explants. The findings revealed that regeneration success is significantly influenced by the type and concentration of disinfectant, explant age, and genotype. Treatment with 10% sodium hypochlorite markedly reduced contamination, achieving clean culture and regeneration rates of 75.72 ± 3.36% and 86.83 ± 3.02%, respectively, regardless of explant age. In contrast, higher concentrations of mercuric chloride induced necrosis in the explants. The highest clean culture rate (93.82 ± 1.16%) was observed in 3-week-old explants, which also showed a regeneration rate of 54.93 ± 3.19%. Furthermore, the Boyapleu and Irene genotypes demonstrated good suitability for in vitro culture, whereas the Nakabo genotype performed poorly under the tested conditions. Full article

(This article belongs to the Section Industry, Agriculture and Food Biotechnology)

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16 pages, 2083 KiB

Open AccessArticle

Effects of Glycerol and Phenolics on Myceliophthora heterothallica Endoxylanase Expressed in K. phaffii

by Jéssica de Araujo Zanoni, Izabela Karolina Costa Zilli, Guilherme de Paula Pretto, Flavio Augusto Vicente Seixas, Marcela Marques de Freitas Lima, Eliana Gertrudes de Macedo Lemos, Eleni Gomes, Gabriel Zazeri and Gustavo Orlando Bonilla-Rodriguez

BioTech 2025, 14(3), 62; https://doi.org/10.3390/biotech14030062 - 18 Aug 2025

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Abstract

Industrial applications of xylanases in high-temperature settings are limited by enzyme instability. This study evaluated glycerol and phenolic compounds as modulators of the catalytic and structural properties of a recombinant Myceliophthora heterothallica endoxylanase (rMhXyn) expressed in Komagataella phaffii. Glycerol (20% v/ [...] Read more.

Industrial applications of xylanases in high-temperature settings are limited by enzyme instability. This study evaluated glycerol and phenolic compounds as modulators of the catalytic and structural properties of a recombinant Myceliophthora heterothallica endoxylanase (rMhXyn) expressed in Komagataella phaffii. Glycerol (20% v/v) significantly improved thermostability (5-fold increase in half-life at 55 °C), decreased the activation energy for catalysis, and enhanced structural rigidity as evidenced by molecular dynamics simulations (reduced RMSD and Rg). In contrast, phenolic acids provided only short-term stabilization at moderate temperatures and did not confer structural benefits. Enzyme kinetics revealed that glycerol enhanced catalytic turnover (↑V_max), while phenolic compounds modified both K′ and cooperativity (Hill coefficient). Thermodynamic analysis supported glycerol’s stabilizing effect, with increased ∆H_(D) and a positive shift in ∆S_(D). These results suggest glycerol as a superior stabilizer for rMhXyn in high-temperature bioprocesses such as lignocellulosic biomass hydrolysis. These findings highlight the potential of targeted additives to improve enzyme performance for biotechnological applications. Full article

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18 pages, 1363 KiB

Open AccessArticle

Rosemary Extract: Phytochemical Composition and Potential for Eliminating Polymicrobial Biofilm of Candida albicans and Multidrug-Resistant Bacteria

by Tuana Mendonça Faria Cintra, Raquel Teles de Menezes, Lara Steffany de Carvalho, Leticia de Miguel Nazario, Leandro Wang Hantao, Maria Cristina Marcucci, Luciane Dias de Oliveira and Vanessa Marques Meccatti-Domiciano

BioTech 2025, 14(3), 61; https://doi.org/10.3390/biotech14030061 - 13 Aug 2025

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Abstract

Herbal medicines can be promising for the treatment of infections caused by multidrug-resistant microorganisms. This study aimed to evaluate Rosmarinus officinalis (Rosemary) hydroalcoholic extract (RHE) regarding its phytochemical composition and potential for eliminating polymicrobial biofilm of Candida albicans with multidrug-resistant bacteria (Acinetobacter [...] Read more.

Herbal medicines can be promising for the treatment of infections caused by multidrug-resistant microorganisms. This study aimed to evaluate Rosmarinus officinalis (Rosemary) hydroalcoholic extract (RHE) regarding its phytochemical composition and potential for eliminating polymicrobial biofilm of Candida albicans with multidrug-resistant bacteria (Acinetobacter baumannii, Klebsiella pneumoniae, and Pseudomonas aeruginosa). The extraction and quantification of the extract (flavonoids and phenols) were performed, and its antioxidant activity (DPPH) and the presence of bio-active compounds were investigated using high-performance liquid chromatography with Diode Array Detection (HPLC-DAD) and Gas Chromatography–Mass Spectrometry (GC-MS). The minimum inhibitory concentration (MIC) and minimum microbicidal concentration (MMC) were determined, and the extract’s action on polymicrobial biofilms was evaluated using the MTT assay. Data were analyzed using one-way ANOVA and Tukey’s tests, as well as Kruskal–Wallis and Dunn’s tests, with a significance level of 5%. RHE showed compatible amounts of flavonoids and phenols, with an EC50 of 19.53 µg/mL. Through HPLC-DAD and GC-MS, biomolecules such as rosmarinic acid and α-Pinene were identified. The extract exhibited microbicidal activity and antibiofilm action, with reduction percentages of up to 69.6% (p < 0.05), showing superior performance compared to 0.12% chlorhexidine against C. albicans + A. baumannii. In conclusion, RHE may be a promising therapeutic agent against multidrug-resistant pathogens. Full article

(This article belongs to the Topic From Plant to Pharmacology: Understanding the Metabolism of Natural Products)

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20 pages, 979 KiB

Open AccessArticle

Influence of Nutrient Medium Composition on the Redistribution of Valuable Metabolites in the Freshwater Green Alga Tetradesmus obliquus (Chlorophyta) Under Photoautotrophic Growth Conditions

by Elvira E. Ziganshina and Ayrat M. Ziganshin

BioTech 2025, 14(3), 60; https://doi.org/10.3390/biotech14030060 - 11 Aug 2025

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Abstract

The study of microalgae has led to significant progress in recent decades. The current microalgal biomass yield is unsatisfactory, except for certain species that are cultivated for the nutraceutical and pharmaceutical industries. In this study, the growth efficiency and biochemical composition of Tetradesmus [...] Read more.

The study of microalgae has led to significant progress in recent decades. The current microalgal biomass yield is unsatisfactory, except for certain species that are cultivated for the nutraceutical and pharmaceutical industries. In this study, the growth efficiency and biochemical composition of Tetradesmus obliquus at high levels of nutrients were characterized. Increasing the NH₄⁺-N content in the medium to 164 mg L⁻¹ allowed the algae to steadily accumulate biomass (6.14 ± 0.28 g L⁻¹) with a moderate content of starch. Optimizing the levels of N, P, and S allowed the biomass productivity to increase from the average 0.45 to 0.88 g L⁻¹ day⁻¹. A further increase of NH₄⁺-N to 410 mg L⁻¹ and other nutrients’ concentration allowed the algae to accumulate biomass (7.50 ± 0.28 g L⁻¹), enriched with protein and pigments. The algae cultivated with the high load of nutrients reached 100%, 84%, and 96% removal of N, P, and S, respectively. Adding the NaHCO₃ to the photobioreactor for pH adjustment (instead of NaOH) did not significantly improve the growth parameters or affect the composition of the algal cells. In general, our study will improve the comprehensive understanding of culture-based approaches to study the perspective use of the alga T. obliquus. Full article

(This article belongs to the Special Issue A Sustainable Approach to Macroalgae and Microalgae: From Cultivation to Compound Recovery and Biotechnological Applications)

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12 pages, 505 KiB

Open AccessArticle

Effect of Ultrasonic Pretreatment on the Extraction Process of Essential Oils from Grapefruit (Citrus paradisi) By-Products

by Francisco Cadena-Cadena, Joe Luis Arias-Moscoso, Leandris Argentel-Martínez, Jony R. Torres Velazquez, Dulce Alondra Cuevas-Acuña, Nydia Estrellita Buitimea Cantua and Bartolo Concha-Frías

BioTech 2025, 14(3), 59; https://doi.org/10.3390/biotech14030059 - 7 Aug 2025

Viewed by 331

Abstract

This study evaluated the effect of ultrasonic pulse-assisted extraction on the yield and antioxidant activity of essential oils from grapefruit (Citrus paradisi) by-products using hydrodistillation and Soxhlet solvent extraction (hexane, acetone, ethanol). Ultrasound was applied at 40% amplitude for 20 min [...] Read more.

This study evaluated the effect of ultrasonic pulse-assisted extraction on the yield and antioxidant activity of essential oils from grapefruit (Citrus paradisi) by-products using hydrodistillation and Soxhlet solvent extraction (hexane, acetone, ethanol). Ultrasound was applied at 40% amplitude for 20 min before extraction. Results showed that ultrasound significantly increased extraction yield with hexane (from 2.6 ± 0.58% to 7.6 ± 1.5%) and acetone (from 8.6 ± 0.96% to 12 ± 1.4%), while ultrasound-assisted hydrodistillation nearly doubled the yield (from 0.7 ± 0.03% to 1.5 ± 0.49%). In contrast, ultrasound decreased yield with ethanol by 3%. Antioxidant activity measured by TEAC assay was highest in acetone extracts without ultrasound (13,366.5 ± 7.66 mmol TE/g) and ethanol extracts (12,606.8 ± 0.51 mmol TE/g). However, ultrasound combined with ethanol increased DPPH scavenging activity from 1073.5 ± 1.07 µg/mL to 4933.3 ± 0.71 µg/mL and maintained high flavonoid content (9.41 ± 0.15 mg/mL) and phenolics (5.33 ± 0.09 mg/mL). Ultrasound-assisted hydrodistillation also enhanced antioxidant capacity, with DPPH values rising from 51.82 ± 5.56 µg/mL to 2413.03 ± 3.17 µg/mL. These findings demonstrate that ultrasound effectively enhances essential oil extraction and antioxidant activity depending on the solvent used, underscoring the potential of this clean technology for valorizing citrus by-products. Full article

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21 pages, 763 KiB

Open AccessReview

Pathway Analysis Interpretation in the Multi-Omic Era

by William G. Ryan V., Smita Sahay, John Vergis, Corey Weistuch, Jarek Meller and Robert E. McCullumsmith

BioTech 2025, 14(3), 58; https://doi.org/10.3390/biotech14030058 - 29 Jul 2025

Viewed by 421

Abstract

In bioinformatics, pathway analyses are used to interpret biological data by mapping measured molecules with known pathways to discover their functional processes and relationships. Pathway analysis has become an essential tool for interpreting large-scale omics data, translating complex gene sets into actionable experimental [...] Read more.

In bioinformatics, pathway analyses are used to interpret biological data by mapping measured molecules with known pathways to discover their functional processes and relationships. Pathway analysis has become an essential tool for interpreting large-scale omics data, translating complex gene sets into actionable experimental insights. However, issues inherent to pathway databases and misinterpretations of pathway relevance often result in “pathway fails,” where findings, though statistically significant, lack biological applicability. For example, the Tumor Necrosis Factor (TNF) pathway was originally annotated based on its association with observed tumor necrosis, while it is multifunctional across diverse physiological processes in the body. This review broadly evaluates pathway analysis interpretation, including embedding-based, semantic similarity-based, and network-based approaches to clarify their ideal use-case scenarios. Each method for interpretation is assessed for its strengths, such as high-quality visualizations and ease of use, as well as its limitations, including data redundancy and database compatibility challenges. Despite advancements in the field, the principle of “garbage in, garbage out” (GIGO) shows that input quality and method choice are critical for reliable and biologically meaningful results. Methodological standardization, scalability improvements, and integration with diverse data sources remain areas for further development. By providing critical guidance with contextual examples such as TNF, we aim to help researchers align their objectives with the appropriate method. Advancing pathway analysis interpretation will further enhance the utility of pathway analysis, ultimately propelling progress in systems biology and personalized medicine. Full article

(This article belongs to the Topic Computational Intelligence and Bioinformatics (CIB))

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23 pages, 2175 KiB

Open AccessArticle

Fetal Health Diagnosis Based on Adaptive Dynamic Weighting with Main-Auxiliary Correction Network

by Haiyan Wang, Yanxing Yin, Liu Wang, Yifan Wang, Xiaotong Liu and Lijuan Shi

BioTech 2025, 14(3), 57; https://doi.org/10.3390/biotech14030057 - 28 Jul 2025

Viewed by 362

Abstract

Maternal and child health during pregnancy is an important issue in global public health, and the classification accuracy of fetal cardiotocography (CTG), as a key tool for monitoring fetal health during pregnancy, is directly related to the effectiveness of early diagnosis and intervention. [...] Read more.

Maternal and child health during pregnancy is an important issue in global public health, and the classification accuracy of fetal cardiotocography (CTG), as a key tool for monitoring fetal health during pregnancy, is directly related to the effectiveness of early diagnosis and intervention. Due to the serious category imbalance problem of CTG data, traditional models find it challenging to take into account a small number of categories of samples, increasing the risk of leakage and misdiagnosis. To solve this problem, this paper proposes a two-step innovation: firstly, we design a method of adaptive adjustment of misclassification loss function weights (MAAL), which dynamically identifies and increases the focus on misclassified samples based on misclassification rates. Secondly, a primary and secondary correction network model (MAC-NET) is constructed to carry out secondary correction for the misclassified samples of the primary model. Experimental results show that the method proposed in this paper achieves 99.39% accuracy on the UCI publicly available fetal health dataset, and also obtains excellent performance on other domain imbalance datasets. This demonstrates that the model is not only effective in alleviating the problem of category imbalance, but also has very high clinical utility. Full article

(This article belongs to the Section Computational Biology)

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16 pages, 982 KiB

Open AccessArticle

Impact of Cattle Breed in scRNA-Seq Reference on Muscle Fiber Type Deconvolution from Bulk RNA-Seq: A Comparison of Software Tools

by Raphael P. Moreira, Marcelo R. Vicari, Henrique A. Mulim, Theresa M. Casey, Jacquelyn Boerman, Xing Fu and Hinayah R. Oliveira

BioTech 2025, 14(3), 56; https://doi.org/10.3390/biotech14030056 - 25 Jul 2025

Viewed by 386

Abstract

While bulk RNA sequencing provides a comprehensive view of transcriptomes, it lacks cell type specificity. Single-cell RNA sequencing (scRNA-seq) overcomes this limitation by providing detailed insights at the individual cell level, though it involves higher costs. Deconvolution methods can estimate cell type proportions [...] Read more.

While bulk RNA sequencing provides a comprehensive view of transcriptomes, it lacks cell type specificity. Single-cell RNA sequencing (scRNA-seq) overcomes this limitation by providing detailed insights at the individual cell level, though it involves higher costs. Deconvolution methods can estimate cell type proportions in bulk RNA-seq data, but their results may vary based on the scRNA-seq reference data and software used. This study investigates the estimation of muscle fiber type proportions through deconvolution analysis of Longissimus dorsi muscle bulk RNA-seq data from late-gestation Holstein Friesian multiparous cows. Four software tools (i.e., CIBERSORTx, Cellanneal, DeconvR-NNLS, and DeconvR-RLM) were compared using scRNA-seq reference data from Brahman and Wagyu cattle breeds, which included proportions of types I, IIa, and IIx myofibers. Kruskal–Wallis and Dunn’s tests revealed that the breed of reference data significantly influenced the proportions of type IIa and IIx muscle fibers across different deconvolution methods. To the best of our knowledge, this is the first study to show that the cattle breed used in reference scRNA-seq data can substantially impact deconvolution outcomes, highlighting a critical consideration for accurate cell type proportion estimation in livestock genomics. These findings suggest that future deconvolution studies should carefully consider breed compatibility between reference and target datasets. Full article

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14 pages, 1901 KiB

Open AccessArticle

The Role of Microarray in Modern Sequencing: Statistical Approach Matters in a Comparison Between Microarray and RNA-Seq

by Isaac D. Raplee, Samiksha A. Borkar, Li Yin, Guglielmo M. Venturi, Jerry Shen, Kai-Fen Chang, Upasana Nepal, John W. Sleasman and Maureen M. Goodenow

BioTech 2025, 14(3), 55; https://doi.org/10.3390/biotech14030055 - 5 Jul 2025

Viewed by 510

Abstract

Gene expression analysis is crucial in understanding cellular processes, development, health, and disease. With RNA-seq outpacing microarray as the chosen platform for gene expression, is there space for array data in future profiling? This study involved 35 participants from the Adolescent Medicine Trials [...] Read more.

Gene expression analysis is crucial in understanding cellular processes, development, health, and disease. With RNA-seq outpacing microarray as the chosen platform for gene expression, is there space for array data in future profiling? This study involved 35 participants from the Adolescent Medicine Trials Network for HIV/AIDS Intervention protocol. RNA was isolated from whole blood samples and analyzed using both microarray and RNA-seq technologies. Data processing included quality control, normalization, and statistical analysis using non-parametric Mann–Whitney U tests. Differential expression analysis and pathway analysis were conducted to compare the outputs of the two platforms. The study found a high correlation in gene expression profiles between microarray and RNA-seq, with a median Pearson correlation coefficient of 0.76. RNA-seq identified 2395 differentially expressed genes (DEGs), while microarray identified 427 DEGs, with 223 DEGs shared between the two platforms. Pathway analysis revealed 205 perturbed pathways by RNA-seq and 47 by microarray, with 30 pathways shared. Both microarray and RNA-seq technologies provide highly concordant results when analyzed with consistent non-parametric statistical methods. The findings emphasize that both methods are reliable for gene expression analysis and can be used complementarily to enhance the robustness of biological insights. Full article

(This article belongs to the Section Computational Biology)

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25 pages, 4957 KiB

Open AccessArticle

Monitoring of the Single-Cell Behavior of an Escherichia coli Reporter Strain Producing L-phenylalanine in a Scale-Down Bioreactor by Automated Real-Time Flow Cytometry

by Prasika Arulrajah, Sophi Katharina Riessner, Anna-Lena Heins and Dirk Weuster-Botz

BioTech 2025, 14(3), 54; https://doi.org/10.3390/biotech14030054 - 3 Jul 2025

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Abstract

Large-scale bioprocesses often suffer from spatial heterogeneities, which impact microbial performance and often lead to phenotypic population heterogeneity. To better understand these effects at the single-cell level, this study applied, for the first time, automated real-time flow cytometry (ART-FCM) to monitor L-phenylalanine production [...] Read more.

Large-scale bioprocesses often suffer from spatial heterogeneities, which impact microbial performance and often lead to phenotypic population heterogeneity. To better understand these effects at the single-cell level, this study applied, for the first time, automated real-time flow cytometry (ART-FCM) to monitor L-phenylalanine production with an Escherichia coli triple reporter strain in a fed-batch process with glycerol as the carbon source. The strain was cultivated in both a well-mixed stirred-tank bioreactor (STR) and a scale-down two-compartment bioreactor (TCB), consisting of an STR and a coiled flow inverter (CFI) in bypass, to simulate spatial heterogeneities. ART-FCM enabled autonomous, high-frequency sampling every 20 min, allowing for real-time tracking of fluorescence signals linked to growth (rrnB-mEmerald), oxygen availability (narGHIJ-CyOFP1), and product formation (aroFBL-mCardinal2). The STR exhibited uniform reporter expression and higher biomass accumulation, while the TCB showed delayed product formation and pronounced phenotypic diversification depending on the set mean residence time in the CFI. Single-cell fluorescence distributions revealed that the shorter mean residence time in the CFI resulted in pronounced subpopulation formation, whereas longer exposure attenuated heterogeneity, indicating transcriptional adaptation. This finding highlights a critical aspect of scale-down studies: increased exposure duration to perturbations can enhance population robustness. Overall, this study demonstrates the relevance of ART-FCM, in combination with a multi-reporter strain, as a pioneering tool for capturing dynamic cellular behavior and correlating it to process performance, providing deeper insights into microbial heterogeneity under fluctuating bioprocess conditions. Full article

(This article belongs to the Section Industry, Agriculture and Food Biotechnology)

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11 pages, 2802 KiB

Open AccessCommunication

Investigation of the Cytotoxicity of Cu(II), Au(III), and Pd(II) Complexes with 2,4-Dithiouracil and 6-Propyl-2-thiouracil Derivatives

by Petya Marinova, Denica Blazheva, Aleksandar Slavchev and Petia Genova-Kalou

BioTech 2025, 14(3), 53; https://doi.org/10.3390/biotech14030053 - 1 Jul 2025

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Abstract

This study investigates the cytotoxic properties of metal complexes incorporating thio-uracil derivatives, specifically 2,4-dithiouracil and 6-propyl-2-thiouracil. The research focuses on the cytotoxic effects of Cu(II) and Pd(II) complexes with 6-propyl-2-thiouracil, as well as mixed-ligand transition metal Cu(II) and Au(III) complexes of 2,4-dithiouracil with [...] Read more.

This study investigates the cytotoxic properties of metal complexes incorporating thio-uracil derivatives, specifically 2,4-dithiouracil and 6-propyl-2-thiouracil. The research focuses on the cytotoxic effects of Cu(II) and Pd(II) complexes with 6-propyl-2-thiouracil, as well as mixed-ligand transition metal Cu(II) and Au(III) complexes of 2,4-dithiouracil with 2-thiouracil and uracil. Cytotoxic activity was assessed against human cervical carcinoma cells (HeLa) and normal kidney cells from the African green monkey. The results demonstrated that incorporating Cu(II) and Au(III) into the compound structures significantly enhanced their cytotoxic effects. Notably, all tested complexes exhibited a stronger inhibitory effect on cancer cell proliferation compared to normal cells, with the palladium(II) complex of 6-propyl-2-thiouracil showing the lowest CD₅₀ value against the tumor cell line (0.00064 mM), which were 149 times lower than that of the ligand (0.0955 mM). These findings suggest that thio-uracil-based metal complexes, particularly those containing palladium (II) and gold(III), hold significant potential for further development as anticancer agents. Full article

(This article belongs to the Section Medical Biotechnology)

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15 pages, 634 KiB

Open AccessReview

Reactive Molecules in Cigarette Smoke: Rethinking Cancer Therapy

by Vehary Sakanyan

BioTech 2025, 14(3), 52; https://doi.org/10.3390/biotech14030052 - 27 Jun 2025

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Abstract

Science has made significant progress in detecting reactive oxygen species (ROS) in tobacco smoke, which is an important step for precision cancer therapy. An important advance is also the understanding that superoxide can be produced by electrophilic molecules. The dual action of hydrogen [...] Read more.

Science has made significant progress in detecting reactive oxygen species (ROS) in tobacco smoke, which is an important step for precision cancer therapy. An important advance is also the understanding that superoxide can be produced by electrophilic molecules. The dual action of hydrogen peroxide, directly or via electrophilic molecules, in the development of oxidative stress allows for the identification of target proteins that can potentially stop unwanted signals in cancer development. However, despite advances in proteomics, reliable inhibitors to stop ROS-associated cancer progression have not yet been proposed for the treatment of tobacco cigarette smokers. This is likely due to an imperfect understanding of the diversity of molecular mechanisms of anti-ROS action. Fluorescent protein detection in living cells, called in-gel, offers a direct route to a better understanding of the rapid interaction of ROS and electrophilic compounds with targeted proteins. It seemed that the traditional paradigm of pharmaceutical innovation “one drug, one disease” did not solve the problem of tobacco smoking causing cancer. However, among the various therapeutic treatments for tobacco smokers, the best way to combat cancer today is smoking cessation, which fits into the “one-cure” paradigm. Full article

(This article belongs to the Section Medical Biotechnology)

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13 pages, 14235 KiB

Open AccessArticle

Expression and Biological Activity Analysis of Recombinant Fibronectin3 Protein in Bacillus subtilis

by Chaozheng Lu, Guangxin Xu, Yin Tian, Zhiwei Yi and Xixiang Tang

BioTech 2025, 14(3), 51; https://doi.org/10.3390/biotech14030051 - 23 Jun 2025

Viewed by 498

Abstract

Fibronectin (FN), a primary component of the extracellular matrix (ECM), features multiple structural domains closely linked to various cellular behaviors, including migration, spreading, adhesion, and proliferation. The FN3 domain, which contains the RGD sequence, is critical in tissue repair because it enables interaction [...] Read more.

Fibronectin (FN), a primary component of the extracellular matrix (ECM), features multiple structural domains closely linked to various cellular behaviors, including migration, spreading, adhesion, and proliferation. The FN3 domain, which contains the RGD sequence, is critical in tissue repair because it enables interaction with integrin receptors on the cell surface. However, the large molecular weight of wild-type FN presents challenges for its large-scale production through heterologous expression. Therefore, this study focused on cloning the FN3 functional domain of full-length FN for expression and validation. This study selected Bacillus subtilis as the expression host due to its prominent advantages, including efficient protein secretion, absence of endotoxins, and minimal codon bias. The recombinant vector pHT43-FN3 was successfully constructed through homologous recombination technology and transformed into Bacillus subtilis WB800N. The FN3 protein was successfully expressed after induction with IPTG. Following purification of the recombinant FN protein using a His-tag nickel column, SDS-PAGE analysis showed that the molecular weight of FN3 was approximately 27.3 kDa. Western blot analysis confirmed the correct expression of FN3, and the BCA protein assay kit determined a protein yield of 5.4 mg/L. CCK8 testing demonstrated the good biocompatibility of FN3. In vitro cell experiments showed that FN3 significantly promoted cell migration at a 20 μg/mL concentration and enhanced cell adhesion at 10 μg/mL. In summary, this study successfully utilized Bacillus subtilis to express the FN3 functional domain peptide from FN protein and has validated its ability to promote cell migration and adhesion. These findings not only provide a strategy for the expression of FN protein in B. subtilis, but also establish an experimental foundation for the potential application of FN3 protein in tissue repair fields such as cutaneous wound healing and cartilage regeneration. Full article

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13 pages, 2298 KiB

Open AccessArticle

In Silico Identification of Pathogenicity Effectors on Fusarium oxysporum f. sp. vanillae

by Felipe Roberto Flores-de la Rosa, Cristian Matilde-Hernández, Nelly Abigail González-Oviedo, Humberto José Estrella-Maldonado, Liliana Eunice Saucedo-Picazo and Ricardo Santillán-Mendoza

BioTech 2025, 14(3), 50; https://doi.org/10.3390/biotech14030050 - 20 Jun 2025

Viewed by 523

Abstract

Vanilla is a highly valuable spice in multiple industries worldwide. However, it faces a serious problem due to a disease known as root and stem rot, caused by the fungus Fusarium oxysporum f. sp. vanillae. Little is known about the pathogenicity mechanisms [...] Read more.

Vanilla is a highly valuable spice in multiple industries worldwide. However, it faces a serious problem due to a disease known as root and stem rot, caused by the fungus Fusarium oxysporum f. sp. vanillae. Little is known about the pathogenicity mechanisms this fungus employs to establish the disease, making it imperative to elucidate mechanisms such as the presence of pathogenicity effectors in its genome. The aim of the present study was to determine the presence of the SIX gene family in the genome of three strains of F. oxysporum associated with root rot: two pathogenic strains and one non-pathogenic endophyte strain. Additionally, the complete effectorome of these strains was predicted and compared to exclude effectors present in the endophytic strain. Our results show that only the SIX9 gene is present in the strains associated with the disease, regardless of their pathogenic nature. Furthermore, no variation was observed in the SIX9 gene among these strains, suggesting that SIX9 is not involved in pathogenicity. Instead, we identified 339 shared effectors among the three strains, including the non-pathogenic strain, strongly suggesting that these genes are not relevant for establishing root rot but may play a role in endophytic colonization. The highly virulent strain IXF41 exhibited eight exclusive pathogenicity effectors, while the moderately virulent strain IXF50 had four. Additionally, one effector was identified as shared between these two strains but absent in the endophytic strain. These effectors and their promoters were characterized, revealing the presence of several cis-regulatory elements responsive to plant hormones. Overall, our findings provide novel insights into the genomic determinants of virulence in F. oxysporum f. sp. vanillae, offering a foundation for future functional studies and the development of targeted disease management strategies. Full article

(This article belongs to the Section Computational Biology)

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