Biomanufacturing harnesses engineered cells for the large-scale production of biochemicals, biopharmaceuticals, biofuels, and biomaterials, playing a vital role in mitigating global environmental crises, achieving carbon peaking and neutrality, and driving the green transformation of the economy and society. The effective design and construction of these engineered cells require precise and comprehensive computational models. Recent technological breakthroughs including high-throughput sequencing, mass spectrometry, spectroscopy, and microfluidic devices, coupled with advances in data science, artificial intelligence, and automation, have enabled the rapid acquisition of large-scale biological datasets, thereby facilitating a deeper understanding of cellular dynamics and the construction of mechanism-based models with enhanced accuracy. This review systematically summarises the mathematical frameworks employed in cellular modelling. It begins by evaluating prevalent mathematical paradigms, such as network topology analyses, stochastic processes, and kinetic equations, critically assessing their applicability across various contexts. The discussion then categorises modelling strategies for specific cellular processes, including cellular growth and division, morphogenesis, DNA replication, transcriptional regulation, metabolism, signal transduction, and quorum sensing. We also examine the recent progress in developing whole-cell models through the integration of diverse cellular processes. The review concludes by addressing key challenges such as data scarcity, unknown mechanisms, multi-dimensional data integration, and exponentially escalating computational complexity. Overall, this work consolidates the mathematical models for the precise simulation of cellular processes, thereby enhancing our understanding of the molecular mechanisms governing cellular functions and contributing to the future design and optimisation of engineered organisms.
Models, Biological ; Cell Physiological Phenomena ; Cell Engineering/methods* ; Humans

Biomanufacturing has emerged as a crucial driving force for efficient material conversion through engineered cells or cell-free systems. However, the intrinsic spatiotemporal heterogeneity, complexity, and dynamic characteristics of these processes pose significant challenges to systematic understanding, optimization, and regulation. This review summarizes essential methodologies for multi-omics data acquisition and analyses for bioprocesses and outlines modelling approaches based on multi-omics data. Furthermore, we explore practical applications of multi-omics and modelling in fine-tuning process parameters, improving fermentation control, elucidating stress response mechanisms, optimizing nutrient supplementation, and enabling real-time monitoring and adaptive adjustment. The substantial potential offered by integrating multi-omics with computational modelling for precision bioprocessing is also discussed. Finally, we identify current challenges in bioprocess optimization and propose the possible solutions, the implementation of which will significantly deepen understanding and enhance control of complex bioprocesses, ultimately driving the rapid advancement of biomanufacturing.
Fermentation ; Genomics/methods* ; Biotechnology/methods* ; Proteomics/methods* ; Models, Biological ; Metabolomics/methods* ; Bioreactors ; Multiomics

OBJECTIVE To evaluate the cost-effectiveness of the first-line treatment using the combination therapy of sugemalimab and chemotherapy （hereinafter referred to as the “combination therapy”） for advanced esophageal squamous cell carcinoma （ESCC） with high programmed death-ligand 1 （PD-L1） expression from the perspective of the Chinese healthcare system. METHODS A partitioned survival model was constructed based on data from the GEMSTONE-304 study. The model cycle was set at 3 weeks， with a study duration of 10 years and a discount rate of 5%. The primary output parameters of the model included total costs， quality-adjusted life year （QALY）， incremental costs， and incremental cost-effectiveness ratio （ICER）. Cost- utility analysis was employed to assess the economic feasibility of the combination therapy compared to chemotherapy alone. The robustness of the base case analysis results was evaluated through univariate sensitivity analysis， probabilistic sensitivity analysis， and scenario analysis. RESULTS The ICER of the combination therapy compared to chemotherapy alone was 288 430.35 yuan/QALY， significantly exceeding the willingness-to-pay （WTP） threshold of 173 354.52 yuan/QALY which was set at 1.94 times the per capita gross domestic product （GDP） in 2023. The price of sugemalimab was the primary factor influencing the ICER. When the WTP threshold was set at 1.94 times the per capita GDP （173 354.52 yuan/QALY）， the probability of the combination therapy being cost-effective compared to chemotherapy alone was 0. The combination therapy only became cost-effective compared to chemotherapy alone when the price of the drug dropped to 6 107.41 yuan per box （600 mg）. CONCLUSIONS From the perspective of the Chinese healthcare system， the combination therapy for first-line treatment of advanced ESCC with high PD-L1 expression is not cost-effective； the combination therapy is cost-effective when the price of sugemalimab decreas by 50.65%.

ObjectiveTo explore the efficacy and predictive indicators of stellate ganglion block （SGB） as an adjunctive intervention for chronic subjective tinnitus and accumulate experience for the application of SGB in the clinical treatment of tinnitus. MethodsA retrospective review was conducted on the data of chronic subjective tinnitus patients who received SGB intervention， with unsatisfactory outcomes otherwise. Pure tone audiometry （PTA）， tinnitus loudness evaluation and Pittsburgh sleep quality index （PSQI） were used. The tinnitus handicap inventory （THI） scores were compared before and after SGB intervention. Correlation analysis and linear regression equations were employed to identify the potential indicators predicting the effectiveness of SGB intervention. Statistical analysis was performed by SPSS 24.0 software. ResultsBy April 2023， a total of 107 patients with chronic subjective tinnitus had undergone SGB intervention， including 67 male and 40 female， with a mean age of （45.32±11.40） years old and an average tinnitus history of （20.32±24.64） months ［16 （12~20）］. Only 7 patients （6.54%） quitted the intervention for personal reasons， which demonstrated good compliance with the intervention. No patients experienced adverse reactions such as infection at the injection site， hematoma， nerve injury， local anesthetic intoxication and so on， which revealed good safety. After SGB intervention， THI scores decreased to below 36 points in 77 patients and decrease by 10 points or more in 12 of the remaining patients， with a total effective rate of 89%. A paired sample t-test showed a significant difference in THI scores before and after SGB intervention （t=15.575， P<0.001）， indicating good improvement. Pearson correlation analysis suggested that pre-intervention THI scores and subjective tinnitus loudness were significantly positively correlated with the improvement level of THI scores （P<0.05）. Further stepwise linear regression analysis found that "pre-intervention THI scores" had statistical significance （P<0.001）， with a regression coefficient of 0.308， predicting a 17.4% improvement level in THI scores. ConclusionsDue to its good and safe short-term effects， SGB intervention can be used as a supplementary option for chronic subjective tinnitus when other interventions are not ideal， especially for patients with higher THI scores. However， further research is needed to clarify the long-term efficacy and underlying mechanisms， in order to establish a more solid theoretical basis for SGB intervention in the treatment of subjective tinnitus.

Amino acids are the basic building blocks of protein that are very important to the nutrition and health of humans and animals, and widely used in feed, food, medicine and daily chemicals. At present, amino acids are mainly produced from renewable raw materials by microbial fermentation, forming one of the important pillar industries of biomanufacturing in China. Amino acid-producing strains are mostly developed through random mutagenesis- and metabolic engineering-enabled strain breeding combined with strain screening. One of the key limitations to further improvement of production level is the lack of efficient, rapid, and accurate strain screening methods. Therefore, the development of high-throughput screening methods for amino acid strains is very important for the mining of key functional elements and the creation and screening of hyper-producing strains. This paper reviews the design of amino acid biosensors and their applications in the high-throughput evolution and screening of functional elements and hyper-producing strains, and the dynamic regulation of metabolic pathways. The challenges of existing amino acid biosensors and strategies for biosensor optimization are discussed. Finally, the importance of developing biosensors for amino acid derivatives is prospected.
Animals ; Humans ; Amino Acids ; Biosensing Techniques ; Metabolic Engineering ; High-Throughput Screening Assays ; China

Promoter is an important genetic tool for fine-tuning of gene expression and has been widely used for metabolic engineering. Corynebacterium glutamicum is an important chassis for industrial biotechnology. However, promoter libraries that are applicable to C. glutamicum have been rarely reported, except for a few developed based on synthetic sequences containing random mutations. In this study, we constructed a promoter library based on the native promoter of odhA gene by mutating the -10 region and the bystanders. Using a red fluorescent protein (RFP) as the reporter, 57 promoter mutants were screened by fluorescence imaging technology in a high-throughput manner. These mutants spanned a strength range between 2.4-fold and 19.6-fold improvements of the wild-type promoter. The strongest mutant exhibited a 2.3-fold higher strength than the widely used strong inducible promoter P_trc. Sequencing of all 57 mutants revealed that 55 mutants share a 1-4 bases shift (4 bases shift for 68% mutants) of the conserved -10 motif "TANNNT" to the 3' end of the promoter, compared to the wild-type promoter. Conserved T or G bases at different positions were observed for strong, moderate, and weak promoter mutants. Finally, five promoter mutants with different strength were employed to fine-tune the expression of γ-glutamyl kinase (ProB) for L-proline biosynthesis. Increased promoter strength led to enhanced L-proline production and the highest L-proline titer of 6.4 g/L was obtained when a promoter mutant with a 9.8-fold higher strength compared to the wild-type promoter was used for ProB expression. The use of stronger promoter variants did not further improve L-proline production. In conclusion, a promoter library was constructed based on a native C. glutamicum promoter P_odhA. The new promoter library should be useful for systems metabolic engineering of C. glutamicum. The strategy of mutating native promoter may also guide the construction of promoter libraries for other microorganisms.
Corynebacterium glutamicum/metabolism* ; Gene Library ; Metabolic Engineering ; Promoter Regions, Genetic/genetics*

One-carbon compounds such as methanol and methane are cheap and readily available feedstocks for biomanufacturing. Oxidation of methanol to formaldehyde catalyzed by methanol dehydrogenase (MDH) is a key step of microbial one-carbon metabolism. A variety of MDHs that depend on different co-factors and possess different enzymatic properties have been discovered from native methylotrophs. Nicotinamide adenine dinucleotide (NAD)-dependent MDHs are widely used in constructing synthetic methylotrophs, whereas this type of MDH usually suffers from low methanol oxidation activity and low affinity to methanol. Consequently, methanol oxidation is considered as a rate-limiting step of methanol metabolism in synthetic methylotrophs. To accelerate methanol oxidation, thereby improving the methanol utilization efficiency of synthetic methylotrophs, massive researches have focused on discovery and engineering of MDHs. In this review, we summarize the ongoing efforts to discover, characterize, and engineer various types of MDHs as well as the applications of MDHs in synthetic methylotrophs. Directed evolution of MDH and construction of multi-enzyme complexes are described in detail. In the future prospective part, we discuss the potential strategies of growth-coupled protein evolution and rational protein design for acquisition of superior MDHs.
Alcohol Oxidoreductases/genetics* ; Carbon ; Methane ; Methanol

Aspergillus niger is a vital industrial workhouse widely used for the production of organic acids and industrial enzymes. This fungus is a crucial cell factory due to its innate tolerance to a diverse range of abiotic conditions, high production titres, robust growth during industrial scale fermentation, and status as a generally recognized as safe (GRAS) organism. Rapid development of synthetic biology and systems biology not only offer powerful approaches to unveil the molecular mechanisms of A. niger productivity, but also provide more new strategies to construct and optimize the A. niger cell factory. As a new generation of genome editing technology, the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR associated (Cas) system brings a revolutionary breakthrough in targeted genome modification for A. niger. In this review, we focus on current advances to the CRISPR/Cas genome editing toolbox, its application on gene modification and gene expression regulation in this fungal. Moreover, the future directions of CRISPR/Cas genome editing in A. niger are highlighted.
Aspergillus niger/genetics* ; CRISPR-Cas Systems/genetics* ; Clustered Regularly Interspaced Short Palindromic Repeats/genetics* ; Gene Editing ; Genome

Corynebacterium glutamicum is an important workhorse of industrial biotechnology, especially for amino acid bioindustry. This bacterium is being used to produce various amino acids at a level of over 6 million tons per year. In recent years, enabling technologies for C. glutamicum metabolic engineering have been developed and improved, which accelerated construction and optimization of microbial cell factoriers, expanding spectra of substrates and products, and facilitated basic researches on C. glutamicum. With these technologies, C. glutamicum has become one of the ideal microbial chasses. This review summarizes recent key technological developments of enabling technologies for C. glutamicum metabolic engineering and focuses on establishment and applications of CRISPR-based genome editing, gene expression regulation, adaptive laboratory evolution, and biosensor technologies.
Amino Acids ; Biotechnology ; Corynebacterium glutamicum/genetics* ; Gene Editing ; Metabolic Engineering

Amino acids are important compounds with a wide range of applications in the food, medicine and chemical industries. Corynebacterium glutamicum is a powerful workhorse commonly used in industrial amino acid production, with the scale of more than one million tons. In addition to its efficient anabolism, the effective exporters also ensure the high amino acid production by C. glutamicum. In this review, the research progress of amino acid exporter of C. glutamicum is summarized, to provide the foundation for further improving amino acid production by C. glutamicum via metabolic engineering.
Amino Acids ; Corynebacterium glutamicum/genetics* ; Metabolic Engineering