{"title":"Application of silica monoliths for improved storage stability of metabolites in human plasma.","authors":"Kazuhiro Kawamura, Eiichiro Fukusaki","doi":"10.1016/j.jbiosc.2025.02.007","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.02.007","url":null,"abstract":"<p><p>In metabolomic studies, sample collection and analysis are typically performed at separate locations, necessitating the transport and storage of samples. However, sample transport and storage conditions are often constrained by the available facilities. Specifically, metabolite levels in biological and food samples can fluctuate due to the activity of endogenous enzymes, depending on the transport and storage conditions. Therefore, in this study, we aimed to achieve metabolite stabilization during storage by sampling human plasma on silica monoliths. Silica monoliths maintained the metabolite samples in a dry state, enabling their transport and storage at high temperatures. Plasmas stored at room temperature and refrigerated were measured using gas chromatography/mass spectrometry (GC/MS), and the fluctuations in metabolites between normal storage and storage on silica monoliths were compared. As a result, fluctuations in several metabolites such as glucose and amino acids were observed during normal storage. However, these were suppressed during storage on silica monoliths. Overall, our findings highlight the efficiency of silica monoliths for sample transport and storage at high temperatures.</p>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-03-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143630501","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Carine Souza da Silva, Gustavo Miranda Pires Santos, Gabriele Rodrigues Conceição, Ian da Silva Andrade, Alana Nogueira Silva, Rodrigo Miranda Pires Santos, Paulo Fernando de Almeida, Fabio Alexandre Chinalia
{"title":"Role of low-level alternating current and impedance for enhancing microalgae biomass and lipid production.","authors":"Carine Souza da Silva, Gustavo Miranda Pires Santos, Gabriele Rodrigues Conceição, Ian da Silva Andrade, Alana Nogueira Silva, Rodrigo Miranda Pires Santos, Paulo Fernando de Almeida, Fabio Alexandre Chinalia","doi":"10.1016/j.jbiosc.2025.02.001","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.02.001","url":null,"abstract":"<p><p>Microalgae hold significant potential for producing value-added bioproducts in pharmaceutical, cosmetic, food, and biofuel industries, with a global market value estimated at US$ 11.8 billion in 2023. Innovations in culturing systems, such as electric stimulation, aim to enhance growth performance, as it can improve cellular processes, including nutrient uptake and lipid accumulation. This study investigates the effect of low alternating electrical currents (μA) on the growth and lipid production of the halotolerant microalga Dunaliella salina across varying salt concentrations (3.5 % and 8.5 % Conway medium). Applying electric stimulation at 50, 750, and 990 μA for 30 min daily over 15 days resulted in significant enhancements, particularly at 3.5 % salinity, where lipid content increased by 144 %. The findings indicate that electrical stimulation notably reduced the lag phase and increased exponential growth rates, with superior growth coefficients correlating with higher medium impedance rather than direct current levels.</p>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-03-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143604997","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Development of an enzymatic cascade for semi de novo ATP production using thermophilic enzymes.","authors":"Takuma Suzuki, Suryatin Alim Gladwin, Kentaro Miyazaki, Hiroya Tomita, Kohsuke Honda","doi":"10.1016/j.jbiosc.2025.02.005","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.02.005","url":null,"abstract":"<p><p>Industrial production of ATP has mostly relied on extraction from living cells. Although microbial and enzymatic ATP production have also been developed, the former suffers from complexity in product separation, while the latter requires expensive substrates, making their practical use difficult. To tackle these problems, we newly developed an enzymatic cascade for ATP production, which does not use expensive substrates, by assembling 16 thermophilic enzymes prepared through a heat-purification from the crude extract of recombinant Escherichia coli. This cascade consists of two modules: an ATP regeneration module based on a non-oxidative glycolysis and an ADP supply module. The ATP regeneration module can provide the energy required for phosphorylation of AMP and ADP to ATP while simultaneously supplying ribose-5-phosphate, a building block of adenosine phosphates, from inexpensive starch and inorganic phosphate. Ribose-5-phosphate is then adenylated with exogenously supplied adenine in the ADP supply module and further phosphorylated to ATP. This ATP production cascade is not accompanied by CO<sub>2</sub> emission and is expected to be a novel ATP manufacturing platform with less environmental impact. In the present study, ATP production with 100 % molar conversion yield was achieved from 1 mM adenine. However, increasing the initial adenine concentration resulted in lower yields. Enzyme characterization and docking simulations revealed that this decline was due to non-competitive inhibition of certain enzymes by ATP, which could potentially be mitigated through protein engineering.</p>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-03-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143597122","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Shuai Zhang, Bingjie Cheng, Qing Liao, Xuewu Huang, Mengjiao Mi, Ming Huang, Yue Wu, Shuyan Wu, Xiaoyuang Wang, Xiaoqing Hu
{"title":"Multi-gene metabolic engineering of Pichia pastoris to synthesize ectoine.","authors":"Shuai Zhang, Bingjie Cheng, Qing Liao, Xuewu Huang, Mengjiao Mi, Ming Huang, Yue Wu, Shuyan Wu, Xiaoyuang Wang, Xiaoqing Hu","doi":"10.1016/j.jbiosc.2025.02.006","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.02.006","url":null,"abstract":"<p><p>As a promising osmolyte, ectoine has been widely applied in cosmetics, food, and pharmaceutical industries in recent years, therefore its biomanufacturer has attracted increasing interest. Ectoine-producing isolates were previously screened from halophilic microorganisms. After ectoine synthetase was identified, genetic engineering of Escherichia coli, Corynebacterium glutamicum, and Hansenula polymorpha were employed to produce ectoine. However, Pichia pastoris, another successful host capable of high-density cell culture, had not yet been exploited as an ectoine-synthesizing host. In this study, therefore, P. pastoris was employed for the first time to produce ectoine through multi-gene metabolic engineering. Firstly, Chromohalobacter salexigens HZS/E, a halophilic isolate producing 46.96 mg/mL ectoine, was identified, while ectABC encoding ectoine synthetase was cloned. Later, ectABC was introduced into P. pastoris GS115 under the control of two different promoters. The results showed that P<sub>GAP</sub>-based HZS02 accumulated 8.03 g/L, 12.62 % higher than 7.13 g/L produced by P<sub>AOX</sub>-based HZS01. Finally, to enhance the supply of the precursor l-aspartate-β-semialdehyde, three genes (aspC, aK, and asD) were individually and collectively overexpressed. The highest ectoine yield was achieved at 10.88 g/L by GS115/pGAPZ A-ectABC-aspC-aK-asD. This study demonstrated that P. pastoris was a highly effective host for ectoine biosynthesis.</p>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-03-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143585665","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Enhancing specimen preparation for transmission electron microscopy: Trypan Blue staining and low-melting-point agar embedding for ultra-thin cell sections.","authors":"Noriyuki Ishii, Yoshito Koja, Keiichi Noguchi, Masafumi Yohda, Shin Takeda","doi":"10.1016/j.jbiosc.2025.02.003","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.02.003","url":null,"abstract":"<p><p>The observation of ultrathin sections of cells and other specimens using transmission electron microscopy (TEM) is a standard technique in many biological research laboratories. Cells are typically collected, dehydrated through an ascending series of alcohols, and embedded in resin blocks before being sectioned with an ultramicrotome. This multi-step process can be time consuming and error prone. To address these challenges, we introduced modifications to improve sample visualization while avoiding hazardous substances like osmium tetroxide and epoxy resins (e.g., Araldite), which are increasingly regulated internationally. Specifically, we stained cells with Trypan Blue and used low melting point agar, facilitating visual identification of target areas and enabling precise embedding. As a result, visual tracking of samples prior to embedding for TEM was improved, preventing the cutting of empty blocks and ensuring efficient preparation of ultrathin sections.</p>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-03-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143585684","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Antibiofilm action of phytochemicals on Enterobacteriaceae.","authors":"Anuj Rohatgi, Pratima Gupta","doi":"10.1016/j.jbiosc.2025.01.007","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.01.007","url":null,"abstract":"<p><p>The biofilm-associated infections pose a great threat to human health. The available drugs are not effective due to the formation of biofilm and limited access to underlying pathogens. The initiation of biofilm formation occurs through adhesion, facilitated by the adhesin protein MrkD1P in the fimbriae tip. This study targeted the MrkD1P protein and employed plant phenols to inhibit biofilm formation in Escherichia coli, Salmonella typhi, and Klebsiella pneumoniae, as major Enterobacteriaceae species. A homology model was constructed for the MrkD1P protein, and 44 phenolic derivatives were assessed for their interaction with this protein. Caffeic acid and 3-hydroxybenzoic acid exhibited the best binding-free energies of 29.61 kcal/mol and 24.24 kcal/mol, respectively. Using a microtiter plates-based minimum biofilm inhibitory concentration assay, it was found that doses of these compounds ranging from 2 to 256 mg/mL effectively reduced biofilm formation. The biofilm inhibition assay demonstrated over 80 % reduction of biofilms in all tested species at inhibitory doses. Further analysis through field emission gun scanning electron micrographs revealed that the compounds disintegrated fimbriae on cell surfaces. Additionally, the re-formation assay demonstrated the inability of biofilm-associated cells to re-form the biofilm on fresh surfaces due to fimbriae inhibition. This study highlights the antibiofilm capabilities of caffeic acid and 3-hydroxybenzoic acid, indicating their potential as effective treatments for illnesses caused by Enterobacteriaceae biofilms.</p>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-02-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143537184","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Norihiko Takemoto, Wenhui Hao, Hiroki Matsuzawa, Jun Sakurai, Yota Tsuge
{"title":"Controlling the carbon flux between glycolysis and the pentose phosphate pathway via targeted protein degradation in Corynebacterium glutamicum.","authors":"Norihiko Takemoto, Wenhui Hao, Hiroki Matsuzawa, Jun Sakurai, Yota Tsuge","doi":"10.1016/j.jbiosc.2025.02.002","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.02.002","url":null,"abstract":"<p><p>Controlling the carbon flux is important for efficient production of value-added chemicals using microbial cell factories. In this study, we developed a system to control the carbon flux in Corynebacterium glutamicum via targeted protein degradation. We employed an SspB-dependent protein degradation system targeting the SsrA tag and applied it to control the carbon flux. First, we selected a degradation tag efficiently recognized by the ClpXP protease in C. glutamicum using green fluorescent protein (GFP) as a model protein. Among the four tags examined in this study, a mutant SsrA tag with DAS residues from Escherichia coli resulted in specific GFP degradation only when the adaptor SspB was induced in C. glutamicum. Next, we applied this system to control the carbon flux. We selected phosphoglucoisomerase (PGI) encoded by pgi, as a target protein, to control the carbon flux between glycolysis and pentose phosphate pathway (PPP) and 1,5-diaminopentane as a model product to evaluate this control system. Compared with the parental strain, the specific growth rate of the engineered strain decreased by 36 %, whereas the yield and production rate of 1,5-diaminopentane increased by 193 % and 70 %, respectively. This is the first report on the application of a protein degradation system to control carbon flux in C. glutamicum. The system developed in this study can be widely applied for designing C. glutamicum cell factories for efficient production of varied value-added chemicals.</p>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-02-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143515818","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Mika Nishimura, Kazuki Takahashi, Masahito Hosokawa
{"title":"Recent advances in single-cell RNA sequencing of Bacteria: Techniques, challenges, and applications.","authors":"Mika Nishimura, Kazuki Takahashi, Masahito Hosokawa","doi":"10.1016/j.jbiosc.2025.01.008","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.01.008","url":null,"abstract":"<p><p>Single-cell RNA sequencing (scRNA-seq) has revolutionized our understanding of cellular heterogeneity in complex biological systems. While this technology has been widely applied to eukaryotic cells, its adaptation to bacterial systems has been challenging due to the unique characteristics of bacterial transcripts. This review surveys the recent developments in bacterial scRNA-seq techniques, highlighting the technical challenges, methodological innovations, and emerging applications in microbiology. We discuss the key differences between eukaryotic and bacterial RNA-seq approaches, focusing on the strategies to overcome limitations such as the lack of poly-A tails in bacterial mRNAs and the low RNA content in individual bacterial cells. The review covers various bacterial scRNA-seq methods, including plate-based, split-pool barcoding, and droplet-based techniques, comparing their strengths and limitations in terms of sensitivity, throughput, and applicability to different bacterial species. Furthermore, we explore the biological insights gained from these techniques, such as identifying rare cell states, characterization of antibiotic responses, and analysis of bacterial communities. Finally, we discuss future perspectives and potential applications of bacterial scRNA-seq in understanding microbial physiology, host-pathogen interactions, and complex microbial ecosystems. This comprehensive overview aims to provide researchers with a clear understanding of the current state and future directions of single-cell transcriptomics in bacteria.</p>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-02-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143472473","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Application of deep learning for evaluation of the growth rate of Daphnia magna.","authors":"Shinsuke Inagaki, Yohei Kondo, Pijar Religia, Nikko Adhitama, Yasuhiko Kato, Eiji Watanabe, Hajime Watanabe","doi":"10.1016/j.jbiosc.2025.01.006","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.01.006","url":null,"abstract":"<p><p>For the safe use of chemicals widely used in human activities, it is crucial to assess their ecological impacts when released into the environment. Daphnia, a well-established environmental indicator species, is commonly used to evaluate the biological effects of chemicals and testing methods have been established. Among various indicators, the growth rate is one of the important parameters, but it requires significant time and effort to measure. In this study, we applied deep learning-based image recognition techniques to extract images of Daphnia from live imaging and assess their size. The estimated size of Daphnia, derived from images processed through deep learning, showed a high correlation with measured values, demonstrating the capability to measure Daphnia size from the images while they are swimming. This approach enables non-invasive measurements of Daphnia size without complicated procedures, which not only streamlines ecological impact assessments but also presents a valuable technique for ecological studies, such as analyzing the size distribution of zooplankton.</p>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-02-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143425452","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Importance of dataset design in developing robust U-Net models for label-free cell morphology evaluation","authors":"Takeru Shiina , Kazue Kimura , Yuto Takemoto , Kenjiro Tanaka , Ryuji Kato","doi":"10.1016/j.jbiosc.2025.01.004","DOIUrl":"10.1016/j.jbiosc.2025.01.004","url":null,"abstract":"<div><div>Advances in regenerative medicine highlighted the need for label-free cell image analysis to replace conventional microscopic observation for non-invasive cell quality evaluation. Image-based evaluation provides an efficient, quantitative, and automated approach to cell analysis, but segmentation remains a critical and challenging step. In this study, we investigated how training dataset design influenced the robustness of U-Net models for cell segmentation, focusing on challenges posed by limited data availability in cell culture. Using 2592 image pairs from four cell types representing key morphological categories, we constructed 42 investigation patterns to evaluate the effects of dataset size, dataset content, and morphological diversity on model performance. Our results showed that robust segmentation models could be developed with approximately 10 raw images captured using a 4× objective lens, a much smaller dataset than typically assumed. The dataset content was found to be crucial: training dataset images that captured commonly observed cell patterns yielded more robust models compared to those capturing rare or irregular cell patterns, which often impaired model performance with large deviations. Additionally, including both spindle and round cell morphologies in the training datasets improved model robustness when tested across all four cell types, while datasets restricted to a single morphology type could not achieve robust models. These findings highlight the importance of curating datasets that capture representative yet diverse cell morphologies. By addressing critical questions about dataset design, this study provides actionable guidance for the effective use of deep learning-based cell segmentation models in manufacturing and research applications.</div></div>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":"139 4","pages":"Pages 329-339"},"PeriodicalIF":2.3,"publicationDate":"2025-02-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143399168","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}