Prompt:
Classify every fixed-vocabulary term as present or absent in the evidence. Vocabulary: bioreactor, cell culture, fermentation, chromatography, purification, filtration, viability, titer, potency, sterility, viral vector, gene therapy, cell therapy, monoclonal antibody, recombinant protein, dissolved oxygen. Return JSON with exactly two alphabetized keys: {"absent":[...],"present":[...]}. Preserve vocabulary order within each list. JSON whitespace is ignored.

Evidence:
Vascular endothelial growth factor (VEGF) is widely used in regenerative medicine and therapeutic research. However, the purification of recombinant VEGF largely relies on affinity chromatography, which requires expensive chromatographic columns, specialized equipment, and multistep processing. These column-based workflows increase operational complexity and cost, particularly for large-scale production. Therefore, the development of an alternative purification strategy to conventional chromatography-based purification for VEGF is needed. In this study, we developed a chromatography-free VEGF purification strategy using an anti-VEGF-scFv–calsequestrin (CSQ) fusion protein that enables calcium-dependent affinity precipitation. The fusion protein retained strong binding affinity for VEGF ( K d = 1.1 nM) while exhibiting rapid and reversible Ca 2 ⁺-dependent polymerization. Upon CaCl₂ addition, the anti-VEGF-scFv-CSQ–VEGF complex rapidly formed aggregates, enabling efficient separation of VEGF from impurities. Using this strategy, VEGF was purified within 30 min with a purity of 94% and a yield of 93%. SEC-HPLC analysis confirmed a purity of 94.3%, and host cell protein contamination was reduced from 1.44 × 10 4 ppm to 774 ppm. The fusion protein also maintained stable purification performance over five repeated cycles, with VEGF recovery consistently maintained above 85%. These findings demonstrate that the scFv-CSQ fusion protein enables rapid separation of VEGF through calcium-dependent polymerization. This column-free mechanism reduces operational cost and technical complexity, highlighting its potential as an alternative to conventional chromatography-based purification. The online version contains supplementary material available at 10.1186/s40643-026-01063-y.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
The preparation of perillartine, a potent marketed sweetener, and N ‐tert‐butyl‐α‐phenylnitrone (PBN), a most commonly used free‐radical spin‐trap used in personal care formulation, endowed with antioxidant, neuroprotective, and anti‐aging properties, was studied by vibrating and planetary ball‐milling. The reactivity in polymeric jars, such as PTFE and in‐house made polyoxymethylene (POM), was evaluated and compared to the most commonly used stainless steel and zirconium oxide milling media. This investigation revealed an insightful correlation while also accounting for the energy involved in each transformation. Assessment of the processes by green chemistry metrics and tools ( e.g. , Chem21, DOZN 3.0 and EcoScale) showed that mechanochemical approaches offer a Safe and Sustainable by Design (SSbD) approach also complying with the IUPAC guiding principles of a responsible chemistry. The reduced solvent consumption, the simplified purification procedures, and lower associated environmental and safety hazards are strongly aligned with the green chemistry principles. Keywords: active ingredients, biomass transformation, green chemistry metrics, mechanochemistry, polymeric jars

Candidates:
A. The evidence does not state that this investigation revealed an insightful correlation while also accounting for the energy involved in each transformation.
B. The preparation of perillartine, a potent marketed sweetener, and N ‐tert‐butyl‐α‐phenylnitrone (PBN), a most commonly used free‐radical spin‐trap used in personal care formulation, endowed with antioxidant, neuroprotective, and anti‐aging properties, was not studied by vibrating and planetary ball‐milling.
C. This investigation revealed an insightful correlation while also accounting for the energy involved in each transformation.
D. The preparation of perillartine, a potent marketed sweetener, and N ‐tert‐butyl‐α‐phenylnitrone (PBN), a most commonly used free‐radical spin‐trap used in personal care formulation, endowed with antioxidant, neuroprotective, and anti‐aging properties, was studied by vibrating and planetary ball‐milling.
E. The evidence does not state that assessment of the processes by green chemistry metrics and tools ( e.g.
F. Assessment of the processes by green chemistry metrics and tools ( e.g.
G. The reactivity in polymeric jars, such as PTFE and in‐house made polyoxymethylene (POM), was evaluated and compared to the most commonly used stainless steel and zirconium oxide milling media.
H. The reactivity in polymeric jars, such as PTFE and in‐house made polyoxymethylene (POM), was not evaluated and compared to the most commonly used stainless steel and zirconium oxide milling media.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Africa is home to 15% of the global population and bears nearly a quarter of the world′s disease burden. However, it faces significant challenges in biomedical research and healthcare due to limited access to essential reagents. This review examines the critical need for localized reagent production to strengthen biomedical research and healthcare across the continent. The reagent market in Africa is characterized by varying levels of development, and the current production capacity falls short of meeting the biomedical research and healthcare demands. The reliance on imported reagents results in high costs, lengthy procurement processes, and vulnerability to supply chain disruptions. Establishing reagent production facilities across the continent can enhance self‐sufficiency, reduce costs, and improve biomedical research tailored to the unique health challenges faced by African populations. Such efforts can be supported by strategic public–private partnerships and international collaborations, leveraging the successes of the existing pharmaceutical investments across the continent and local capacity‐building initiatives. Regional collaboration, market development, technical training, infrastructure, regulatory frameworks, and sustainable funding are essential to ensure long‐term success. Developing skilled human capital and creating a unified market through initiatives like the African Continental Free Trade Area can attract investment and ensure economic sustainability of local reagent production. Together, these measures can significantly improve biomedical research output and healthcare response capabilities, preparing the continent for future health emergencies. Keywords: biomedical research, funding, local manufacturing, reagent production, sustainability

Candidates:
A. This review examines the critical need for localized reagent production to strengthen biomedical research and healthcare across the continent.
B. The reagent market in Africa is characterized by varying levels of development, and the current production capacity falls short of meeting the biomedical research and healthcare demands.
C. The reagent market in Africa is not characterized by varying levels of development, and the current production capacity falls short of meeting the biomedical research and healthcare demands.
D. However, it faces significant challenges in biomedical research and healthcare due to limited access to essential reagents.
E. Africa is home to 15% of the global population and bears nearly a quarter of the world′s disease burden.
F. The evidence does not state that this review examines the critical need for localized reagent production to strengthen biomedical research and healthcare across the continent.
G. The evidence does not state that however, it faces significant challenges in biomedical research and healthcare due to limited access to essential reagents.
H. Africa is home to 16% of the global population and bears nearly a quarter of the world′s disease burden.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["16015 L", "49.28 µg/mL", "96.9%", "95.9%", "16016 L", "48.28 µg/mL", "1000 µg/mL", "1001 µg/mL"]

Evidence:
L-ASNase has attracted attention in many biomedical and food safety applications. Therefore, this study was designed to identify a novel and promising candidate for the sustainable biosynthesis of extracellular L-ASNase from P. ostreatus AUMC 16015 grown on various agricultural substrates under solid-state fermentation (SSF). Also, the enzyme’s wide-ranging bioactivities were examined, involving its antioxidant, anti-inflammatory, and antitumor properties, while evaluating its potential applications in food processing. Optimal P. ostreatus AUMC 16015 L-ASNase production was 56.47 U/mL, which was attained under SSF conditions where the enzyme yield increased by 2.46-fold compared to pre-optimization conditions. Enzyme high purity was validated by a single distinct band at approximately 48 kDa on both SDS-PAGE and native PAGE analyses. The enzyme demonstrated high substrate specificity ( K m = 7.7 mM; V max = 167.78 U/mL). Functionally, it exhibited strong antioxidant activity (2,2-diphenyl-1-picrylhydrazyl) (DPPH) IC 50 = 48.28 µg/mL) and a robust anti-hemolytic effect (95.9% at 1000 µg/mL). L-ASNase exhibited its most potent inhibitory effect against Caco-2 cells at an IC 50 of 5.49 ± 0.03 µg/mL, followed by MCF-7, which showed a slightly higher IC 50 of 5.86 ± 0.08 µg/mL. Furthermore, L-ASNase significantly mitigated potato chips acrylamide formation, achieving a 9.6-fold decrease after 120 min of treatment. Additionally, Gas chromatography-mass spectrometry ( GC-MS) showed that the potato’s chemical profile was significantly changed by L-ASNase treatment, with the introduction of numerous bioactive substances and the elimination of some potentially dangerous components. The biochemical activity of the purified L-ASNase suggested potential biomedical and food applications. This study is a trial for cost-effective enzyme production and supports a circular bioeconomy by converting waste into useful bioproducts. Future work should focus on scaling up production and testing its effects in living organisms to unlock this enzyme’s full commercial and medical potential. Keywords: L-ASNase, P. ostreatus AUMC 16015, SSF, Biomedical and food applications


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Keywords: Greenhouse, Hydroponic system, Plant molecular farming, Nicotiana benthamiana , Sustainable, Controlled-environment agriculture (CEA)

Candidates:
A. The evidence does not state that keywords: Greenhouse, Hydroponic system, Plant molecular farming, Nicotiana benthamiana , Sustainable, Controlled-environment agriculture (CEA)
B. Keywords: Greenhouse, Hydroponic system, Plant molecular farming, Nicotiana benthamiana , Sustainable, Controlled-environment agriculture (CEA)


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Barcoded rabies viral tracing enables high-throughput mapping of brain-wide inputs to individual neurons while allowing integration with transcriptomic profiling. Here, we present a protocol to produce and apply barcoded rabies virus in mice for single-neuron input mapping. We describe steps to generate barcode plasmid libraries, produce barcoded rabies virus, and isolate target brain regions. We then detail procedures for preparing single-cell suspensions or bulk RNA, building barcode amplicon libraries from input regions, and performing initial analysis of the sequencing data. For complete details on the use and execution of this protocol, please refer to Tan et al. 1 , 2 Subject areas: Bioinformatics, Cell Biology, Cell culture, Neuroscience, Sequence analysis, Sequencing, Single Cell

Candidates:
A. Barcoded rabies viral tracing enables high-throughput mapping of brain-wide inputs to individual neurons while allowing integration with transcriptomic profiling.
B. The evidence does not state that we describe steps to generate barcode plasmid libraries, produce barcoded rabies virus, and isolate target brain regions.
C. The evidence does not state that barcoded rabies viral tracing enables high-throughput mapping of brain-wide inputs to individual neurons while allowing integration with transcriptomic profiling.
D. The evidence does not state that we then detail procedures for preparing single-cell suspensions or bulk RNA, building barcode amplicon libraries from input regions, and performing initial analysis of the sequencing data.
E. We describe steps to generate barcode plasmid libraries, produce barcoded rabies virus, and isolate target brain regions.
F. Here, we present a protocol to produce and apply barcoded rabies virus in mice for single-neuron input mapping.
G. The evidence does not state that here, we present a protocol to produce and apply barcoded rabies virus in mice for single-neuron input mapping.
H. We then detail procedures for preparing single-cell suspensions or bulk RNA, building barcode amplicon libraries from input regions, and performing initial analysis of the sequencing data.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Antimicrobial food pads play a key role in the food packaging industry by improving food safety and quality. To control the growth of bacterial pathogens in fish fillets, this study aims to develop a bioactive system using absorbent food pads composed of bacterial cellulose (BC) infused with tangerine essential oil (TEO) as an antibacterial agent. The effects of active BC-pads, gamma irradiation, and their combination on artificially inoculated bacteria (previously isolated) were studied for 6 days at 4 °C using fish fillet samples as a model food system. The results revealed that the initial population counts of Klebsiella oxytoca, Serratia ficaria, Enterobacter cloacae, and Kocuria rosea, were 5.47, 5.63, 5.14, and 5.25 log CFU/g, respectively. These counts increased during the storage period, reaching 8.80 log CFU/g as a mean for all tested strains. After six days of storage, BC, BC/TEO, BC + 1.0 kGy, and BC/TEO + 1.0 kGy pads reduced the initial load of the four bacteria in the fish fillet samples by [1.59, 1.68, 1.92, 2.35], [3.21, 3.12, 3.36, 3.72], [3.53, 3.26, 3.26, 3.37] and [4.97, 5.02, 4.95, 4.83] log CFU/g, respectively. The combination of BC and TEO with low-dose gamma radiation (1.0 kGy) enhances the decontamination effectiveness of fish fillets through synergistic effects. FTIR analysis revealed a slight shift in the absorbance peaks of some functional group interactions and the formation of new bonds between BC and TEO in the presence of gamma irradiation. The study has successfully developed sustainable, eco-friendly, functional bioactive food pads that reduce microbial growth and prevent spoilage of fish fillets during storage. The online version contains supplementary material available at 10.1186/s40643-026-01061-0. Keywords: Food pads, Bacterial cellulose, Hurdle technology, Antimicrobial, Tangerine essential oil, Gamma irradiation, Klebsiella oxytoca , Serratia ficaria , Enterobacter cloacae , Kocuria rosea

Candidates:
A. The results revealed that the initial population counts of Klebsiella oxytoca, Serratia ficaria, Enterobacter cloacae, and Kocuria rosea, were 6.47, 5.63, 5.14, and 5.25 log CFU/g, respectively.
B. To control the growth of bacterial pathogens in fish fillets, this study aims to develop a bioactive system using absorbent food pads composed of bacterial cellulose (BC) infused with tangerine essential oil (TEO) as an antibacterial agent.
C. The evidence does not state that antimicrobial food pads play a key role in the food packaging industry by improving food safety and quality.
D. The effects of active BC-pads, gamma irradiation, and their combination on artificially inoculated bacteria (previously isolated) were studied for 6 days at 4 °C using fish fillet samples as a model food system.
E. The evidence does not state that to control the growth of bacterial pathogens in fish fillets, this study aims to develop a bioactive system using absorbent food pads composed of bacterial cellulose (BC) infused with tangerine essential oil (TEO) as an antibacterial agent.
F. The effects of active BC-pads, gamma irradiation, and their combination on artificially inoculated bacteria (previously isolated) were studied for 7 days at 4 °C using fish fillet samples as a model food system.
G. The results revealed that the initial population counts of Klebsiella oxytoca, Serratia ficaria, Enterobacter cloacae, and Kocuria rosea, were 5.47, 5.63, 5.14, and 5.25 log CFU/g, respectively.
H. Antimicrobial food pads play a key role in the food packaging industry by improving food safety and quality.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
2-phenylethanol (2-PE) is a high-value aromatic alcohol. Its bioproduction is limited by pathway bottlenecks, limited by a rate-limiting decarboxylation step, product toxicity, and volatilization. We sought to build an efficient Escherichia coli ( E. coli ) platform by optimizing the segment of the Ehrlich route that converts L-phenylalanine (L-Phe) to 2-PE, together with a food-grade soybean-oil overlay for in situ product recovery (ISPR). Whole-cell assays showed that phenylpyruvate decarboxylation was the main bottleneck when the yeast decarboxylase Aro10 was used. A phylogeny-guided screen identified Lactococcus lactis KivD as a superior substitute in E. coli . After replacing Aro10 with KivD and tuning expression, leveraging endogenous glutamate dehydrogenase (GDH) for NAD(P)H recycling enabled sufficient cofactor regeneration, yielding 49.5 mM 2-PE from 50 mM L-Phe with 99.0% conversion. At the shake-flask scale, a fed-batch L-Phe feeding strategy coupled with a 2:1 soybean-oil overlay for in situ product removal reduced toxicity and volatilization losses, producing 130 mM 2-PE (15.9 g/L). Using soybean oil as an ISPR phase effectively sequestered 2-PE from the aqueous phase, minimizing volatilization and increasing the overall recovered yield to 94.2% based on L-Phe consumption. Replacing the rate-limiting decarboxylation step with KivD, balancing redox through cofactor recycling, and coupling the pathway to a mild oil-overlay recovery increased flux and enabled high-titer 2-PE in E. coli . The workflow of bottleneck substitution plus biocompatible in situ recovery can be transferred to related aromatic alcohols and provides a foundation for de novo routes and high-cell-density fermentations that target still higher titers. The online version contains supplementary material available at 10.1186/s13036-026-00667-4.

Candidates:
A. 2-phenylethanol (2-PE) is a high-value aromatic alcohol.
B. The evidence does not state that we sought to build an efficient Escherichia coli ( E.
C. Its bioproduction is limited by pathway bottlenecks, limited by a rate-limiting decarboxylation step, product toxicity, and volatilization.
D. 3-phenylethanol (2-PE) is a high-value aromatic alcohol.
E. Its bioproduction is not limited by pathway bottlenecks, limited by a rate-limiting decarboxylation step, product toxicity, and volatilization.
F. coli ) platform by optimizing the segment of the Ehrlich route that converts L-phenylalanine (L-Phe) to 3-PE, together with a food-grade soybean-oil overlay for in situ product recovery (ISPR).
G. We sought to build an efficient Escherichia coli ( E.
H. coli ) platform by optimizing the segment of the Ehrlich route that converts L-phenylalanine (L-Phe) to 2-PE, together with a food-grade soybean-oil overlay for in situ product recovery (ISPR).


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Moreno-Corona et al. identify compound-heterozygous WDR75 variants in a patient with hypogammaglobulinemia and autism spectrum disorder. These variants impair pre-ribosomal RNA processing and trigger nucleolar stress, suggesting a new ribosomopathy.

Candidates:
A. identify compound-heterozygous WDR76 variants in a patient with hypogammaglobulinemia and autism spectrum disorder.
B. These variants impair pre-ribosomal RNA processing and trigger nucleolar stress, suggesting a new ribosomopathy.
C. identify compound-heterozygous WDR75 variants in a patient with hypogammaglobulinemia and autism spectrum disorder.
D. The evidence does not state that these variants impair pre-ribosomal RNA processing and trigger nucleolar stress, suggesting a new ribosomopathy.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
The baculovirus expression vector system is an established platform for large-scale production of (glyco)proteins, subunit vaccines, virus-like particles, and recombinant adeno-associated virus (rAAV) vectors. We engineered a novel bacmid vector (BAC6) to improve genetic stability by deletion of the non-homologous repeat (hr) origin of DNA replication (ori) and to preserve product integrity and recovery through deletion of chitinase and cathepsin. Tn7-based transposition in E. coli and homologous recombination in insect cells were combined in BAC6 to drive expression from the odv-e56 and polyhedrin loci, respectively. Virus growth kinetics of BAC6 were similar to the parental bacmid, and genetic stability was investigated for at least eight serial passages at high multiplicity of infection. Next-generation sequencing was used to identify mutations, deletions, and defective interfering particle (DIP) formation, which became apparent only in later passages. With BAC6, the enrichment of DIPs originating from the non-hr ori was prevented. BAC6 versatility was demonstrated by high-yield (12 mg/L) severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike production in suspension Sf9 insect cells. Finally, rAAV production with BAC6, simultaneously employing both transgene insertion sites, resulted in yields of 5.8e10 AAV capsids/mL and 2.3e10 genome copies/mL. BAC6 provides insertion of multiple transgenes at two different loci and is non-inferior to commercial baculovirus expression vectors with regard to genetic stability and productivity. Keywords: genome editing, bacmid, baculovirus expression vector system, illumina next generation sequencing, SARS-CoV-2, adeno-associated virus, AAV

Candidates:
A. We engineered a novel bacmid vector (BAC7) to improve genetic stability by deletion of the non-homologous repeat (hr) origin of DNA replication (ori) and to preserve product integrity and recovery through deletion of chitinase and cathepsin.
B. The baculovirus expression vector system is an established platform for large-scale production of (glyco)proteins, subunit vaccines, virus-like particles, and recombinant adeno-associated virus (rAAV) vectors.
C. We engineered a novel bacmid vector (BAC6) to improve genetic stability by deletion of the non-homologous repeat (hr) origin of DNA replication (ori) and to preserve product integrity and recovery through deletion of chitinase and cathepsin.
D. coli and homologous recombination in insect cells were combined in BAC7 to drive expression from the odv-e56 and polyhedrin loci, respectively.
E. Virus growth kinetics of BAC6 were similar to the parental bacmid, and genetic stability was investigated for at least eight serial passages at high multiplicity of infection.
F. Virus growth kinetics of BAC7 were similar to the parental bacmid, and genetic stability was investigated for at least eight serial passages at high multiplicity of infection.
G. coli and homologous recombination in insect cells were combined in BAC6 to drive expression from the odv-e56 and polyhedrin loci, respectively.
H. The baculovirus expression vector system is not an established platform for large-scale production of (glyco)proteins, subunit vaccines, virus-like particles, and recombinant adeno-associated virus (rAAV) vectors.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Genetically modified cell therapies (GMCT) hold potential for the treatment of cancer, autoimmune conditions, and rare diseases. A major determinant of GMCT success is modification efficiency, yet transducing certain cells of interest, such as primary natural killer (NK) cells, remains challenging. Major barriers to widespread adoption of GMCT are cost and difficulty of scale. Therefore, improving transduction rates with accessible, good manufacturing practices (GMP) compatible reagents may help reduce this barrier and prevent manufacturing failure. We show that using a modified viscous transduction medium (VTM) made with methyl-cellulose significantly increased the transduction efficiency of primary human NK cells without affecting viability, expansion, or function. VTM outperformed commercially available transduction enhancers. Transduction with VTM significantly improved the yield of anti-CD22 chimeric antigen receptor (CAR)-NK cells. Moreover, VTM was similarly effective at improving the transduction of primary T cells and hematopoietic stem cells. Overall, VTM may provide a simple, cost-effective, and GMP-compliant solution to increase the yield of genetically modified immune cells for immunotherapy. This may improve the efficacy of immunotherapies and reduce the overall costs of production.

Candidates:
A. The evidence does not state that genetically modified cell therapies (GMCT) hold potential for the treatment of cancer, autoimmune conditions, and rare diseases.
B. Major barriers to widespread adoption of GMCT are not cost and difficulty of scale.
C. Therefore, improving transduction rates with accessible, good manufacturing practices (GMP) compatible reagents may help reduce this barrier and prevent manufacturing failure.
D. A major determinant of GMCT success is modification efficiency, yet transducing certain cells of interest, such as primary natural killer (NK) cells, remains challenging.
E. A major determinant of GMCT success is not modification efficiency, yet transducing certain cells of interest, such as primary natural killer (NK) cells, remains challenging.
F. Major barriers to widespread adoption of GMCT are cost and difficulty of scale.
G. Genetically modified cell therapies (GMCT) hold potential for the treatment of cancer, autoimmune conditions, and rare diseases.
H. The evidence does not state that therefore, improving transduction rates with accessible, good manufacturing practices (GMP) compatible reagents may help reduce this barrier and prevent manufacturing failure.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["31%", "30%", "20%", "21%"]

Evidence:
Chimeric antigen receptor (CAR) T cell therapies represent a significant advancement for treating hematological malignancies, particularly in relapsed/refractory cases. Despite their clinical success, the high cost of CAR T cell therapies remains a major barrier to broader implementation. A significant proportion of these costs stems from the dependency on viral vectors and the limited understanding of transduction mechanisms. This work evaluates the impact of physical and chemical parameters during transduction using a spinoculation process. Physical parameters, such as spinoculation duration and speed, were identified as key drivers of transduction efficiency, contributing to a 20%–30% increase in transduction. Similarly, the addition of LentiBOOST and polybrene enhanced transduction efficiency by approximately 1–2-fold compared with control conditions without these supplements. Given that both physical and chemical parameters influence transduction efficiency, a quality-by-design approach was used to systematically investigate their potential synergistic or antagonistic interactions. This systematic approach highlighted the cytotoxic impact of polybrene and demonstrated that LentiBOOST is critical to drive transduction, particularly in CD4 subsets. The optimized process led to a 2–3-fold improvement in transduction without compromising CAR T cell growth or functionality and was shown to be compatible with serum- and xeno-free medium, supporting its translational potential.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["801 nm", "31%", "216%", "30%", "800 nm", "215%"]

Evidence:
Membrane fouling remains a major challenge in water treatment, biomedical, and pharmaceutical fields, yet understanding its mechanisms at the membrane interface remains difficult. Herein, we report a 3D impedimetric microfluidic membrane-mimic (IM3) cassette that integrates a porous membrane between microfluidic channels for real-time fouling investigation via electrochemical impedance spectroscopy (EIS). System validation using fluorescein and KCl electrolytes demonstrated reproducible charge-transfer resistance ( R ct ) values and confirmed cassette robustness. Colloidal fouling studies using 800 nm polystyrene latex (PS) beads demonstrated a clear concentration-dependent behavior. High particle loading (10 5 particles/mL) caused rapid, severe impedance increases, indicating extensive pore blockage and cake layer formation. Low loading (10 1 particles/mL) showed minimal changes, suggesting negligible fouling, as confirmed by scanning electron microscopy. The distribution of relaxation times (DRT) analysis revealed a single dominant relaxation peak at 10 –2 s that grew stronger as fouling severity increased, confirming that pore blockage raised interfacial resistance through a unified charge-transfer mechanism rather than caused diffusion-limited processes. A quantitative fouling model using EIS-measured R ct changes captured temporal and concentration dependencies through an exponential growth framework. Maximum fouling extent ( F max ) increased significantly with particle concentration (30% at 10 1 to 215% at 10 5 particles/mL), demonstrating concentration-dependent pore blockage. The fouling rate constant ( k ) showed weak concentration dependence and plateaued at high loadings, indicating the limitation by available deposition sites rather than particle transport. These findings show that for PS beads, fouling is mainly governed by the extent of surface deposition (driven by concentration) rather than by the particle arrival rate (controlled by kinetics). The developed modular IM3 cassette enables customization of different membranes, channel geometries, and flow configurations. These results establish a quantitative framework linking EIS data to physically meaningful fouling phenomena, offering a versatile platform for the mechanistic investigation of dynamic membrane fouling behavior under varying particulate conditions.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
The filamentous fungus Aspergillus niger is a well-established cell factory in biotechnology. Its productivity depends on macromorphological development which remains difficult to control, partly because the relationship between seed culture and reactor-specific shear force conditions has not been systematically investigated. This study examined how high or low shear forces affect pellet development at both micro- and macromorphological levels in stirred-tank reactors (STR, high shear regime) and rocking-motion bioreactors (RMB, low shear regime). A. niger seed cultures with initially either large or small pellets were used to inoculate batch STR or RMB. Comparable cultivation conditions were applied so that fermentations differed mainly in shear force regime. Growth characteristics and pellet macromorphologies were analysed using 2D and 3D image analyses, enabling us to classify pellets according to three different classes based on their inner pellet architecture. The distribution of these classes depended on both the macromorphologies of the seed culture and the reactor type. Under high shear forces in the STR, pellets underwent breakage shortly after stirrer activation, were limited in their size to an average diameter of 500–600 µm, and formed a homogeneous population. In addition, broken pellets occurred predominantly under STR conditions. In contrast, cultivations in RMB preserved the initial pellet architecture, allowed the formation of larger pellets (median diameter ~ 800 µm) and supported pellet fusion, thus resulting in a more heterogeneous macromorphological population. Notably, glucose uptake rate correlated with the surface-to-volume ratio of the pellet populations, i.e., glucose became faster consumed under STR conditions accompanied with lower biomass yields and higher protein secretion. Citric acid production, however, was detectable in both STR and RMB only when reactors were inoculated with seed cultures characterised by a loose pellet morphology. Overall, our study demonstrates how shear regime and seed culture morphology jointly shape pellet architecture, population heterogeneity and productivity in scale-up processes. Such a comprehensive understanding of morphological developments is instrumental for optimising bioprocesses and future predictive modelling approaches. • 2D/3D analysis of defined seed cultures in different shear-induced environments • High shear restricts and homogenises pellets, while low shear maintains heterogeneity • Highest citric acid and total protein levels were found in smaller, compact pellets. The online version contains supplementary material available at 10.1007/s00253-026-13822-0.

Candidates:
A. The filamentous fungus Aspergillus niger is not a well-established cell factory in biotechnology.
B. niger seed cultures with initially either large or small pellets were used to inoculate batch STR or RMB.
C. The evidence does not state that its productivity depends on macromorphological development which remains difficult to control, partly because the relationship between seed culture and reactor-specific shear force conditions has not been systematically investigated.
D. Its productivity depends on macromorphological development which remains difficult to control, partly because the relationship between seed culture and reactor-specific shear force conditions has not been systematically investigated.
E. The evidence does not state that this study examined how high or low shear forces affect pellet development at both micro- and macromorphological levels in stirred-tank reactors (STR, high shear regime) and rocking-motion bioreactors (RMB, low shear regime).
F. The filamentous fungus Aspergillus niger is a well-established cell factory in biotechnology.
G. This study examined how high or low shear forces affect pellet development at both micro- and macromorphological levels in stirred-tank reactors (STR, high shear regime) and rocking-motion bioreactors (RMB, low shear regime).
H. niger seed cultures with initially either large or small pellets were not used to inoculate batch STR or RMB.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["61.2–97.9%", "62.2–97.9%"]

Evidence:
Inadequate sludge treatment poses a significant risk of environmental pollution. To reduce pollution, utilizing waste sludge as an energy source offers a sustainable solution to mitigate pollution. Since sludges contain abundant organics, they are expected to produce more valuable organics, such as volatile fatty acids (VFAs). Although sludge fermentation has been widely studied, direct yields of VFA production from different sludge types are still limited, particularly for sludge materials originating from Finnish wastewater treatment plants. In this study, laboratory-scale bioreactors were built to examine VFA production from digester feed sludge, digested sludge, and ammonia removal sludge, which were collected from local wastewater treatment plants. Among the tested substrates, digested feed sludge resulted in the highest VFA yield, reaching 171.6 ± 6.3 mg/g volatile solids after 11 days of incubation, which indicated its superior potential for VFA generation. In contrast, bioreactors fed with digested sludge or ammonia removal sludge showed no clear increasing trend in VFA production. The addition of sawdust led to lower overall VFA yields, approximately half of those obtained with digested feed sludge alone, and increasing the sludge proportion did not result in further yield enhancement. Acetic acid was the dominant VFA in all bioreactors, accounting for 61.2–97.9% of total VFAs. Microbial community analysis indicated the prevalence of phyla Bacillota and Pseudomonadota, with Lactobacillus being relatively abundant in bioreactors exhibiting higher VFA production. These findings suggest that sludge type plays an important role in determining VFA production performance and may support more informed selection of sludge substrates for VFA-oriented sludge valorisation. Keywords: Volatile fatty acids, Waste sludge valorisation, Microbial community, Sludge fermentation, Bioprocess sustainability, Resource recovery


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Bottom-up proteomics workflows encompass several key stages, including sample preparation, data acquisition, and data analysis. Of these, sample preparation is the initial and critical stage, as it significantly influences the depth, reproducibility, and reliability of subsequent mass spectrometry–based analyses. While several main digestion strategies exist, including in-gel, in-solution, and filter-aided methods, each presents distinct trade-offs in terms of throughput, contamination removal, and applicability to complex biological matrices. The Suspension Trapping (S-Trap) method offers a compelling alternative by efficiently capturing and digesting proteins while removing interferents like sodium dodecyl sulfate (SDS), which can compromise downstream LC–MS/MS performance. This protocol details a S-Trap workflow optimized for biofluid proteomics, specifically plasma, serum, and cerebrospinal fluid (CSF). We describe two complementary formats: a manual tube-based procedure for individual or small-batch samples and a 96-well-plate-based system enabling high-throughput processing. The protocol integrates optional high-abundance protein depletion to enhance coverage of low-abundance analytes and includes steps for reduction, alkylation, digestion, and peptide elution for low total protein content samples, such as plasma, serum, and cerebrospinal fluid. By providing a detailed protocol, this work aims to improve the consistency and accessibility of S-Trap-based sample preparation, facilitating robust and reproducible discoveries in bottom-up proteomics. Key features • Plasma/serum/cerebrospinal fluid sample preparation for bottom-up proteomics. • Lab Suspension Trapping (S-Trap)-based digestion for efficient detergent removal and high peptide recovery. • Optimized for challenging samples (e.g., CSF, plasma) with low protein concentration or high lipid content. • Includes both single-tube and high-throughput 96-well plate formats for flexible experimental design. Keywords: Mass spectrometry, LC-MS, Proteomics, Sample preparation, S-trap

Candidates:
A. Bottom-up proteomics workflows encompass several key stages, including sample preparation, data acquisition, and data analysis.
B. The Suspension Trapping (S-Trap) method offers a compelling alternative by efficiently capturing and digesting proteins while removing interferents like sodium dodecyl sulfate (SDS), which cannot compromise downstream LC–MS/MS performance.
C. While several main digestion strategies exist, including in-gel, in-solution, and filter-aided methods, each presents distinct trade-offs in terms of throughput, contamination removal, and applicability to complex biological matrices.
D. Of these, sample preparation is the initial and critical stage, as it significantly influences the depth, reproducibility, and reliability of subsequent mass spectrometry–based analyses.
E. The evidence does not state that while several main digestion strategies exist, including in-gel, in-solution, and filter-aided methods, each presents distinct trade-offs in terms of throughput, contamination removal, and applicability to complex biological matrices.
F. Of these, sample preparation is not the initial and critical stage, as it significantly influences the depth, reproducibility, and reliability of subsequent mass spectrometry–based analyses.
G. The evidence does not state that bottom-up proteomics workflows encompass several key stages, including sample preparation, data acquisition, and data analysis.
H. The Suspension Trapping (S-Trap) method offers a compelling alternative by efficiently capturing and digesting proteins while removing interferents like sodium dodecyl sulfate (SDS), which can compromise downstream LC–MS/MS performance.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Polysaccharides from Chroogomphus rutilus (CRPs) exhibit promising bioactivities; however, conventional alkaline extraction (CAE) often results in low yields and structural disruption. In this study, a support vector regression (SVR)–optimized ultrasonic-assisted alkaline extraction (UAAE) process was developed to maximize yield while preserving conformational integrity. Compared with the quadratic response surface methodology model, SVR showed superior predictive accuracy and robustness, with the testing R 2 increasing from 0.8751 to 0.9027 and RMSE reduced by 11.7%. SVR also exhibited narrower residual dispersion and improved stability in the high-yield region (>17%), confirming its suitability for nonlinear, multivariable bioprocess optimization. Under the optimized conditions—ultrasonic temperature 56.5 °C, ultrasonic time 35 min, soaking time 138 min, liquid–solid ratio 26 mL/g, NaOH concentration 0.55 mol/L, and ultrasonic power 325 W—the extraction yield reached 20.09 ± 0.10%, representing a 53.7% increase compared with CAE. High-performance gel permeation chromatography revealed two dominant molecular weight (Mw) fractions for UAAE-derived CRP at 7.54 × 10 5 and 1.32 × 10 4 Da, whereas CAE-derived products exhibited a trimodal distribution dominated by low-Mw species at 1.43 × 10 3 Da. Spectroscopic, microscopic, and thermal analyses demonstrated that UAAE more effectively preserved ordered helical conformations, and improved structural stability compared with CAE. CRPs obtained under optimized conditions also showed enhanced antioxidant activity, including ABTS• + and hydroxyl radical scavenging capacities. The integrating acoustic cavitation with alkaline treatment and SVR-based modeling provides an efficient, data-driven strategy for sustainable production of high-quality fungal polysaccharides with preserved bioactive conformations.

Candidates:
A. In this study, a support vector regression (SVR)–optimized ultrasonic-assisted alkaline extraction (UAAE) process was developed to maximize yield while preserving conformational integrity.
B. The evidence does not state that polysaccharides from Chroogomphus rutilus (CRPs) exhibit promising bioactivities; however, conventional alkaline extraction (CAE) often results in low yields and structural disruption.
C. SVR also exhibited narrower residual dispersion and improved stability in the high-yield region (>18%), confirming its suitability for nonlinear, multivariable bioprocess optimization.
D. Compared with the quadratic response surface methodology model, SVR showed superior predictive accuracy and robustness, with the testing R 3 increasing from 0.8751 to 0.9027 and RMSE reduced by 11.7%.
E. Compared with the quadratic response surface methodology model, SVR showed superior predictive accuracy and robustness, with the testing R 2 increasing from 0.8751 to 0.9027 and RMSE reduced by 11.7%.
F. In this study, a support vector regression (SVR)–optimized ultrasonic-assisted alkaline extraction (UAAE) process was not developed to maximize yield while preserving conformational integrity.
G. SVR also exhibited narrower residual dispersion and improved stability in the high-yield region (>17%), confirming its suitability for nonlinear, multivariable bioprocess optimization.
H. Polysaccharides from Chroogomphus rutilus (CRPs) exhibit promising bioactivities; however, conventional alkaline extraction (CAE) often results in low yields and structural disruption.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Optimizing transient expression systems in fish embryos is crucial for rapid gene function analysis. Here, we established an efficient system in medaka ( Oryzias latipes ) embryos by evaluating nucleic acid type and injection site. Our results revealed that injecting the elongation factor 1αA ( ef1αA ) promoter‐driven plasmid into the yolk yielded the highest expression on day 1 post‐fertilization. Using this optimized system, we investigated cytosolic 5′‐nucleotidase 1a ( nt5c1a ), which is involved in the metabolism of inosine monophosphate (IMP), an umami flavor compound. In silico analysis revealed that medaka had two nt5c1a paralogs: nt5c1aa and nt5c1ab . While nt5c1ab retains conserved substrate‐recognition sequences and exhibits significant IMP degradation activity, nt5c1aa has lost these functions. Structural analysis using AlphaFold revealed that the Nt5c1aa L305P mutation causes local conformational changes near the substrate‐binding site, potentially altering substrate orientation without disrupting the overall protein fold. Our expression system demonstrated that this single L305P substitution partially restored IMP‐degrading activity in Nt5c1aa, confirming that residue 305 is a key determinant of its functional divergence. Our findings provide a robust foundation for molecular breeding to enhance umami flavor in farmed fish. Specifically, the targeted manipulation of these nt5c1a paralogs could facilitate developing breeds with maximized IMP accumulation in muscle tissues.

Candidates:
A. Optimizing transient expression systems in fish embryos is not crucial for rapid gene function analysis.
B. Using this optimized system, we investigated cytosolic 6′‐nucleotidase 1a ( nt5c1a ), which is involved in the metabolism of inosine monophosphate (IMP), an umami flavor compound.
C. Here, we established an efficient system in medaka ( Oryzias latipes ) embryos by evaluating nucleic acid type and injection site.
D. Our results revealed that injecting the elongation factor 2αA ( ef1αA ) promoter‐driven plasmid into the yolk yielded the highest expression on day 1 post‐fertilization.
E. Optimizing transient expression systems in fish embryos is crucial for rapid gene function analysis.
F. Using this optimized system, we investigated cytosolic 5′‐nucleotidase 1a ( nt5c1a ), which is involved in the metabolism of inosine monophosphate (IMP), an umami flavor compound.
G. The evidence does not state that here, we established an efficient system in medaka ( Oryzias latipes ) embryos by evaluating nucleic acid type and injection site.
H. Our results revealed that injecting the elongation factor 1αA ( ef1αA ) promoter‐driven plasmid into the yolk yielded the highest expression on day 1 post‐fertilization.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
This data article presents an open ultrasonic pulse – echo dataset acquired from NMC – 622 cathode slurries monitored under static conditions to characterise their post – mixing structural recovery behaviour. Slurries were prepared at varying solid contents of 63, 65, 67 and 69 wt.% and transferred into a fixed test container for ultrasonic monitoring. A 5 MHz contact transducer was used to record continuous A – Scan waveform under consistent acquisition settings (raw time – domain signals paired with frequency – domain representation). From each waveform, reflection amplitude and time – of – flight (ToF) features extracted from the slurry – air interface echo provides a compact representation of evolving acoustic response over time. The dataset is intended for benchmarking of signal processing pipelines, development of physics – informed feature extraction, and machine – learning models for slurry metrology and coating readiness assessment. By Packing signals, minimal metadata, and lightweight visualisation and analysis script, the resources enable reproducible reuse across materials – processing and signal – processing communities. Keywords: Ultrasonic testing, Lithium-ion battery manufacturing, Cathode slurry, Structural recovery, Process monitoring, Production line parameterisation

Candidates:
A. A 5 MHz contact transducer was used to record continuous A – Scan waveform under consistent acquisition settings (raw time – domain signals paired with frequency – domain representation).
B. A 6 MHz contact transducer was used to record continuous A – Scan waveform under consistent acquisition settings (raw time – domain signals paired with frequency – domain representation).
C. This data article presents an open ultrasonic pulse – echo dataset acquired from NMC – 622 cathode slurries monitored under static conditions to characterise their post – mixing structural recovery behaviour.
D. The evidence does not state that from each waveform, reflection amplitude and time – of – flight (ToF) features extracted from the slurry – air interface echo provides a compact representation of evolving acoustic response over time.
E. From each waveform, reflection amplitude and time – of – flight (ToF) features extracted from the slurry – air interface echo provides a compact representation of evolving acoustic response over time.
F. Slurries were prepared at varying solid contents of 64, 65, 67 and 69 wt.% and transferred into a fixed test container for ultrasonic monitoring.
G. Slurries were prepared at varying solid contents of 63, 65, 67 and 69 wt.% and transferred into a fixed test container for ultrasonic monitoring.
H. This data article presents an open ultrasonic pulse – echo dataset acquired from NMC – 623 cathode slurries monitored under static conditions to characterise their post – mixing structural recovery behaviour.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Automated phenotyping in ophthalmology requires accurate standardization of clinical terms to facilitate interoperability and research. This study evaluates the suitability of the human phenotype ontology (HPO) for automated extraction of ophthalmic phenotypes from narrative documentation. We developed a locally operated AI pipeline combining text segmentation and negation detection based on a small language model (PHI-4) with a dense retrieval approach using an augmented multilingual HPO catalog. Synonyms were incorporated into the HPO during training on anonymized consecutive physician letters. To validate the pipeline, 175 anterior segment and fundus descriptions from randomly picked medical records were manually annotated with HPO terms as ground truth. Overall, 342 HPO terms were identified manually (on average 2.53 terms per document), with 341 retrieved by the pipeline (on average 2.52 terms per document). Performance metrics showed a median Jaccard similarity of 0.67, precision of 0.83, recall of 0.82, and F1 score of 0.80. These results demonstrate that our AI pipeline effectively extracts standardized HPO terms from free-text ophthalmic findings. Integration of this pipeline into clinical information systems may enhance data interoperability and reduce manual coding workload in ophthalmology practices in the future. The online version contains supplementary material available at 10.1038/s41598-026-51512-z.

Candidates:
A. Synonyms were not incorporated into the HPO during training on anonymized consecutive physician letters.
B. We developed a locally operated AI pipeline combining text segmentation and negation detection based on a small language model (PHI-4) with a dense retrieval approach using an augmented multilingual HPO catalog.
C. The evidence does not state that this study evaluates the suitability of the human phenotype ontology (HPO) for automated extraction of ophthalmic phenotypes from narrative documentation.
D. The evidence does not state that automated phenotyping in ophthalmology requires accurate standardization of clinical terms to facilitate interoperability and research.
E. Automated phenotyping in ophthalmology requires accurate standardization of clinical terms to facilitate interoperability and research.
F. Synonyms were incorporated into the HPO during training on anonymized consecutive physician letters.
G. We developed a locally operated AI pipeline combining text segmentation and negation detection based on a small language model (PHI-5) with a dense retrieval approach using an augmented multilingual HPO catalog.
H. This study evaluates the suitability of the human phenotype ontology (HPO) for automated extraction of ophthalmic phenotypes from narrative documentation.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Whole‐cell biocatalysis offers a sustainable alternative to traditional chemical synthesis for producing pharmaceutically relevant, often chiral, amines and amino acids. Saccharomyces cerevisiae has emerged as a privileged microbial chassis due to its robustness, ease of genetic manipulation, and GRAS status. This concise review summarizes recent advances in metabolic and genetic engineering of S. cerevisiae for amine biocatalysis, focusing on strategies to overcome bottlenecks such as enzyme gene expression, cofactor regeneration, and precursor channeling. The first section covers state‐of‐the‐art methods for engineered strain construction, including genomic editing, optimization of gene expression (copy number, promoters, terminators, codon usage), and metabolic engineering (pathway balancing, compartmentalization, cofactor supply, transport proteins, auxiliary enzymes, and enzyme targeting via signal peptides), all enhancing product yields and enabling complex amine synthesis. The central section critically discusses compound families accessible via engineered S. cerevisiae , including various amines, amino alcohols, and amino acids such as l ‐carnitine, ergothioneine, halogenated tryptamine, serotonin, psilocybin, spermidine, l ‐ornithine, and mycosporine derivatives. Bioproduction of complex alkaloids, such as tropine derivatives (hyoscyamine and scopolamine) and ergot alkaloids, is also reviewed. Finally, current challenges and future perspectives are outlined, highlighting the integration of systems and synthetic biology tools to establish S. cerevisiae as a scalable platform for industrial amine production.

Candidates:
A. Saccharomyces cerevisiae has emerged as a privileged microbial chassis due to its robustness, ease of genetic manipulation, and GRAS status.
B. Whole‐cell biocatalysis offers a sustainable alternative to traditional chemical synthesis for producing pharmaceutically relevant, often chiral, amines and amino acids.
C. The evidence does not state that this concise review summarizes recent advances in metabolic and genetic engineering of S.
D. The evidence does not state that saccharomyces cerevisiae has emerged as a privileged microbial chassis due to its robustness, ease of genetic manipulation, and GRAS status.
E. This concise review summarizes recent advances in metabolic and genetic engineering of S.
F. cerevisiae for amine biocatalysis, focusing on strategies to overcome bottlenecks such as enzyme gene expression, cofactor regeneration, and precursor channeling.
G. The evidence does not state that whole‐cell biocatalysis offers a sustainable alternative to traditional chemical synthesis for producing pharmaceutically relevant, often chiral, amines and amino acids.
H. The evidence does not state that cerevisiae for amine biocatalysis, focusing on strategies to overcome bottlenecks such as enzyme gene expression, cofactor regeneration, and precursor channeling.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
In times of a climate crisis caused by extreme emissions of green-house gases on a global scale, mitigation solutions need to be found. One solution is the system of carbon capture and utilization (CCU), where C1 gases, such as carbon monoxide (CO), carbon dioxide (CO 2 ), or methane, are either redirected from industrial off-gas streams or directly air-captured. A biotechnological process for CCU is the use of Methanothermobacter marburgensis for CO 2 fixation and production of value-added compounds. In this study, we focused on valine production, an amino acid important for human or feedstock nutrition. We demonstrated overproduction of valine from CO 2 in M. marburgensis with temperature-induced promoters. Here, we reached a 12.9-fold increase in valine production between the OFF- and ON states of the inducible promoter with a maximal specific production rate of 14.17 mg gCDW −1 h −1 of valine in closed batch experiments. In the second approach for valine production, we overexpressed acetolactate synthase genes with resistance to allosteric valine inhibition from Methanothermobacter thermautotrophicus recombinant in M. marburgensis . We identified a strong reduction in allosteric inhibition towards valine. This resulted in specific valine productivity of up to 40 mg gCDW −1 h −1 and states the highest specific productivity on an individual amino acid in methanogens. With those findings, we expanded the toolbox for genetic modification of M. marburgensis by a thermo-inducible promoter system and applied protein engineering for enhanced production of value-added compounds to M. marburgensis . This proof of concept shows the feasibility of archaeal cell factories generation via genetic engineering for industrial production of value-added compounds with thermophilic methanogens.

Candidates:
A. A biotechnological process for CCU is the use of Methanothermobacter marburgensis for CO 2 fixation and production of value-added compounds.
B. The evidence does not state that in this study, we focused on valine production, an amino acid important for human or feedstock nutrition.
C. One solution is the system of carbon capture and utilization (CCU), where C1 gases, such as carbon monoxide (CO), carbon dioxide (CO 2 ), or methane, are either redirected from industrial off-gas streams or directly air-captured.
D. The evidence does not state that in times of a climate crisis caused by extreme emissions of green-house gases on a global scale, mitigation solutions need to be found.
E. In times of a climate crisis caused by extreme emissions of green-house gases on a global scale, mitigation solutions need to be found.
F. In this study, we focused on valine production, an amino acid important for human or feedstock nutrition.
G. A biotechnological process for CCU is the use of Methanothermobacter marburgensis for CO 3 fixation and production of value-added compounds.
H. One solution is the system of carbon capture and utilization (CCU), where C2 gases, such as carbon monoxide (CO), carbon dioxide (CO 2 ), or methane, are either redirected from industrial off-gas streams or directly air-captured.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Extracellular vesicles (EVs) are nanoscale mediators of intercellular communication with diverse molecular cargoes that reflect their cell of origin. Advances in isolation, detection, and single‐particle analytics have revealed increasing molecular and functional heterogeneity, while exposing limitations in how EV identity and activity are currently defined. The field has expanded rapidly; however, translational progress is constrained by an incomplete mechanistic understanding, a lack of standardized measurements, and inconsistencies in regulatory classification. This review provides a critical synthesis of current EV research from an analytical and translational perspective, emphasizing the measurement science needed to define, compare, and benchmark EV preparations as therapeutic products. We discuss evolving regulatory frameworks and recent updates to the MISEV guidelines, highlighting the need for operational definitions grounded in source material, isolation method, and molecular markers. Updated workflow considerations are presented across EV production and characterization, with a focus on orthogonal analyses, quantification of co‐isolates, and potency assays that support reproducibility and quality control. Together, these advances and ongoing challenges underscore the need for analytical rigor to transform EV research from descriptive studies into a reproducible and quantitative measurement science.

Candidates:
A. Advances in isolation, detection, and single‐particle analytics have revealed increasing molecular and functional heterogeneity, while exposing limitations in how EV identity and activity are currently defined.
B. The field has expanded rapidly; however, translational progress is constrained by an incomplete mechanistic understanding, a lack of standardized measurements, and inconsistencies in regulatory classification.
C. This review provides a critical synthesis of current EV research from an analytical and translational perspective, emphasizing the measurement science needed to define, compare, and benchmark EV preparations as therapeutic products.
D. Extracellular vesicles (EVs) are not nanoscale mediators of intercellular communication with diverse molecular cargoes that reflect their cell of origin.
E. Advances in isolation, detection, and single‐particle analytics have revealed increasing molecular and functional heterogeneity, while exposing limitations in how EV identity and activity are not currently defined.
F. The evidence does not state that this review provides a critical synthesis of current EV research from an analytical and translational perspective, emphasizing the measurement science needed to define, compare, and benchmark EV preparations as therapeutic products.
G. Extracellular vesicles (EVs) are nanoscale mediators of intercellular communication with diverse molecular cargoes that reflect their cell of origin.
H. The field has expanded rapidly; however, translational progress is not constrained by an incomplete mechanistic understanding, a lack of standardized measurements, and inconsistencies in regulatory classification.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Given the biotechnological potential of yeast‐derived oils for oleochemical production, genes encoding lipid metabolism enzymes are key targets for metabolic engineering. Genetic engineering tools such as Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9, Transcription Activator‐Like Effector Nucleases (TALENs), Zinc‐Finger Nucleases (ZFNs), RNA interference (RNAi), and integrative plasmids can be used to modulate fatty acid biosynthesis and optimize lipid production. Among them, the CRISPR/Cas9 system, recognized for its simplicity and efficiency, has been deployed as a tool to create oleaginous yeast strains with high lipid productivity and features suitable for application in biorefineries. Species such as Cutaneotrichosporon oleaginosus , Rhodotorula toruloides , Candida spp., and Yarrowia lipolytica have already been engineered using CRISPR/Cas9 to enhance the production of fatty acids and their derivatives. However, designing and constructing an efficient CRISPR/Cas9 platform for oleaginous yeasts faces several hurdles, including low transformation efficiency, difficulties in expressing Cas9 and sgRNAs efficiently and consistently, the lack of well‐characterized promoters, limited availability of PAM sequences, and poorly understood DNA repair mechanisms. Here, we address the application of the CRISPR/Cas9 system in oleaginous yeasts, laying out the challenges to developing efficient platforms and highlighting key trends in the field. We compare and discuss alternative CRISPR‐Cas9 expression strategies to provide an overview of the current landscape and support the development of new approaches.

Candidates:
A. Given the biotechnological potential of yeast‐derived oils for oleochemical production, genes encoding lipid metabolism enzymes are key targets for metabolic engineering.
B. Among them, the CRISPR/Cas9 system, recognized for its simplicity and efficiency, has been deployed as a tool to create oleaginous yeast strains with high lipid productivity and features suitable for application in biorefineries.
C. Genetic engineering tools such as Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9, Transcription Activator‐Like Effector Nucleases (TALENs), Zinc‐Finger Nucleases (ZFNs), RNA interference (RNAi), and integrative plasmids can be used to modulate fatty acid biosynthesis and optimize lipid production.
D. Given the biotechnological potential of yeast‐derived oils for oleochemical production, genes encoding lipid metabolism enzymes are not key targets for metabolic engineering.
E. Among them, the CRISPR/Cas10 system, recognized for its simplicity and efficiency, has been deployed as a tool to create oleaginous yeast strains with high lipid productivity and features suitable for application in biorefineries.
F. Species such as Cutaneotrichosporon oleaginosus , Rhodotorula toruloides , Candida spp., and Yarrowia lipolytica have already been engineered using CRISPR/Cas9 to enhance the production of fatty acids and their derivatives.
G. Genetic engineering tools such as Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas10, Transcription Activator‐Like Effector Nucleases (TALENs), Zinc‐Finger Nucleases (ZFNs), RNA interference (RNAi), and integrative plasmids can be used to modulate fatty acid biosynthesis and optimize lipid production.
H. Species such as Cutaneotrichosporon oleaginosus , Rhodotorula toruloides , Candida spp., and Yarrowia lipolytica have already been engineered using CRISPR/Cas10 to enhance the production of fatty acids and their derivatives.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
To address agrochemical pollution and improve microbial screening efficiency, this study aimed to develop an integrated pipeline for isolating high-yielding lipopeptide-producing Bacillus strains and validating their agricultural applications. A tiered screening strategy combining hemolytic activity assay, plate confrontation, and oil-spreading assay was established to isolate strains from rhizosphere soils. Lipopeptide production was identified by MALDI-TOF MS and quantified via HPLC. Genomic analysis was performed to elucidate the genetic basis of lipopeptide synthesis. Crude lipopeptide extracts were evaluated for antimicrobial activity via plate assays, and for plant growth-promoting effects under pot and field conditions. The screening successfully identified potent Bacillus producers with multiple plant growth-promoting traits. Strain A1C-8 produced 1140.7 mg/L surfactin, Bv1 produced 450.1 mg/L iturin, and C5C-6 produced 707.8 mg/L fengycin. Crude extracts exhibited strong antimicrobial activity, with inhibition rates of 54.6% against Staphylococcus aureus and 273.3% against Rhizoctonia solani . Genomic analysis confirmed a robust genetic basis for lipopeptide synthesis. Notably, field trials demonstrated that lipopeptide extracts alone significantly enhanced crop growth: maize ear fresh weight increased by 32.46 % under 100 mg/L treatment, peanut pod number per plant rose by 42.75 % under 200 mg/L treatment, and sunflower fresh seed weight per capitulum improved by 35 % with 50 mg/L application. This study provides an integrated pipeline from strain screening to field validation, highlighting Bacillus lipopeptides as sustainable alternatives to agrochemicals for combined biocontrol and plant growth promotion.

Candidates:
A. Lipopeptide production was not identified by MALDI-TOF MS and quantified via HPLC.
B. Genomic analysis was not performed to elucidate the genetic basis of lipopeptide synthesis.
C. The evidence does not state that to address agrochemical pollution and improve microbial screening efficiency, this study aimed to develop an integrated pipeline for isolating high-yielding lipopeptide-producing Bacillus strains and validating their agricultural applications.
D. A tiered screening strategy combining hemolytic activity assay, plate confrontation, and oil-spreading assay was not established to isolate strains from rhizosphere soils.
E. To address agrochemical pollution and improve microbial screening efficiency, this study aimed to develop an integrated pipeline for isolating high-yielding lipopeptide-producing Bacillus strains and validating their agricultural applications.
F. A tiered screening strategy combining hemolytic activity assay, plate confrontation, and oil-spreading assay was established to isolate strains from rhizosphere soils.
G. Lipopeptide production was identified by MALDI-TOF MS and quantified via HPLC.
H. Genomic analysis was performed to elucidate the genetic basis of lipopeptide synthesis.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["31 °C", "1.5–1.5%", "30 °C", "0.5–1.5%", "95%", "96%"]

Evidence:
Cytokines are small glycosylated polypeptides that orchestrate immune responses and are widely produced in recombinant form for therapeutic and research purposes. This review highlights Komagataella phaffii as an efficient host for the heterologous expression of recombinant cytokines from human and other species. A systematic search of PubMed, Scopus, and Web of Science identified studies addressing upstream and downstream processes involved in cytokine expression. Molecular design strategies such as codon optimization, vector design, and signal peptide selection are discussed together with process parameters including temperature, inducer concentration, and bioreactor conditions, as well as recovery and purification of biologically active cytokines. Optimal production is often associated using multi-copy vectors driven by the AOX1 promoter, α-factor secretion signals, methanol induction (0.5–1.5% v/v), temperatures below 30 °C, and co-feeding strategies. Downstream purification commonly yields products exceeding 95% purity. Strategies such as PEGylation, albumin fusion, and antibody fusion are also described to improve cytokine stability and half-life. This review integrates and critically analyzes molecular and bioprocessing advances that establish K. phaffii as a powerful platform for recombinant cytokine production.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
High-content imaging (HCI) involves the automated acquisition and quantitative analysis of cell phenotypes from microscopy images. These studies often rely on screening, which can involve thousands of chemical or genetic perturbations that produce terabytes of microscopy data. To extract meaningful biological insights, these data must be processed into quantitative features through a technique known as image-based profiling. A major analytical bottleneck is curating the high-dimensional, single-cell data derived from various image-analysis tools. These datasets suffer from inconsistent schemas, inefficient file formats, and undocumented ontological relationships. These challenges reduce reproducibility and slow progress in downstream applications. To solve these issues, we introduce CytoTable, a software package for harmonizing single-cell image-based profiling. CytoTable enables modular, portable, and cross-language data integration through a robust, reproducible, and scalable engine that harmonizes single-cell readouts from multiple image-analysis tools, preparing for feature integration with software in the Cytomining ecosystem such as Pycytominer. Keywords: image-based profiling, open-source software, single-cell harmonization, microscopy image analysis, reproducibility, high-content imaging

Candidates:
A. High-content imaging (HCI) involves the automated acquisition and quantitative analysis of cell phenotypes from microscopy images.
B. A major analytical bottleneck is curating the high-dimensional, single-cell data derived from various image-analysis tools.
C. The evidence does not state that high-content imaging (HCI) involves the automated acquisition and quantitative analysis of cell phenotypes from microscopy images.
D. To extract meaningful biological insights, these data must be processed into quantitative features through a technique known as image-based profiling.
E. A major analytical bottleneck is not curating the high-dimensional, single-cell data derived from various image-analysis tools.
F. The evidence does not state that to extract meaningful biological insights, these data must be processed into quantitative features through a technique known as image-based profiling.
G. These studies often rely on screening, which can involve thousands of chemical or genetic perturbations that produce terabytes of microscopy data.
H. These studies often rely on screening, which cannot involve thousands of chemical or genetic perturbations that produce terabytes of microscopy data.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Large quantities of spoiled date palm fruits are generated annually and are often discarded, resulting in environmental and economic losses. Developing sustainable strategies to convert this waste into valuable products is therefore an important challenge. This study aimed to establish an integrated bioprocess that utilizes spoiled date fruits as a low-cost substrate to simultaneously produce biodiesel-grade lipids, natural pigments, and fungal chitosan using Talaromyces atroroseus . The objectives included optimizing substrate concentration and pretreatment conditions, evaluating lipid and pigment production, and recovering chitosan from residual fungal biomass. The results demonstrated that moderate substrate levels supported maximum lipid and pigment yields, while dilute acid pretreatment significantly enhanced sugar release from spoiled date fruits and fungal productivity. The extracted lipids were rich in C16–C18 fatty acids and complied with international biodiesel quality standards. The produced pigments showed good pH and thermal stability along with strong antioxidant activity. Additionally, chitosan recovered from de-oiled biomass exhibited suitable structural properties, and pigment–chitosan composites displayed enhanced antioxidant performance. Therefore, this work presents a sustainable biorefinery approach that transforms agricultural waste into multiple high-value products, supporting waste valorization, renewable energy production, and circular bioeconomy initiatives and offering clear benefits to society and industry.

Candidates:
A. This study aimed to establish an integrated bioprocess that utilizes spoiled date fruits as a low-cost substrate to simultaneously produce biodiesel-grade lipids, natural pigments, and fungal chitosan using Talaromyces atroroseus .
B. The objectives included optimizing substrate concentration and pretreatment conditions, evaluating lipid and pigment production, and recovering chitosan from residual fungal biomass.
C. The evidence does not state that this study aimed to establish an integrated bioprocess that utilizes spoiled date fruits as a low-cost substrate to simultaneously produce biodiesel-grade lipids, natural pigments, and fungal chitosan using Talaromyces atroroseus .
D. Large quantities of spoiled date palm fruits are generated annually and are often discarded, resulting in environmental and economic losses.
E. The evidence does not state that the objectives included optimizing substrate concentration and pretreatment conditions, evaluating lipid and pigment production, and recovering chitosan from residual fungal biomass.
F. Large quantities of spoiled date palm fruits are not generated annually and are often discarded, resulting in environmental and economic losses.
G. Developing sustainable strategies to convert this waste into valuable products is not therefore an important challenge.
H. Developing sustainable strategies to convert this waste into valuable products is therefore an important challenge.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Microalgae are tiny organisms that play an essential role in nature, especially in places with harsh environmental conditions. In this study, we examined a microalga called Chloroidium saccharophilum , which we found in Laguna Blanca, Chile. This is important because this species had never been reported in Chile before or in this region of South America. Its presence in such an extreme environment makes it valuable for scientific and biotechnological research. This microalga is naturally resistant to very salty water, strong sunlight, and temperature changes—conditions that would harm many other organisms. These special abilities give it potential as a natural source of useful compounds such as antioxidants. Antioxidants help protect cells from damage caused by stress and inflammation, which is relevant for health-related products and other applications. Although the microalga showed strong growth, it produced only small amounts of antioxidants under the conditions we tested. This suggests that more research is needed to discover the best environmental factors that encourage higher production of these beneficial compounds. Learning about organisms that thrive in extreme areas can support new sustainable technologies and contribute to economic and social development in remote regions.

Candidates:
A. The evidence does not state that in this study, we examined a microalga called Chloroidium saccharophilum , which we found in Laguna Blanca, Chile.
B. This is not important because this species had never been reported in Chile before or in this region of South America.
C. Microalgae are not tiny organisms that play an essential role in nature, especially in places with harsh environmental conditions.
D. Microalgae are tiny organisms that play an essential role in nature, especially in places with harsh environmental conditions.
E. This is important because this species had never been reported in Chile before or in this region of South America.
F. Its presence in such an extreme environment makes it valuable for scientific and biotechnological research.
G. The evidence does not state that its presence in such an extreme environment makes it valuable for scientific and biotechnological research.
H. In this study, we examined a microalga called Chloroidium saccharophilum , which we found in Laguna Blanca, Chile.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
The association between dietary fiber and phenolic compounds allows the latter to reach the colon, where most polysaccharides undergo fermentation. This bioprocessing weakens the matrix and promotes the release of the phenolic compounds, which then exert beneficial effects on intestinal function. Although this notion is widely accepted, supporting evidence remains scarce. In this study, we subjected grape pomace skin to in vitro digestion to obtain an indigestible fraction suitable for SHIME bioreactors. Throughout these stages, we observed a sequential increase in the release of phenolic compounds, with colonic fermentation playing an important role. Although we did not observe an increase in short-chain fatty acid (SCFA) production by the gut microbiota, we performed a repeated-challenge design on differentiated Caco-2 monolayers. With this approach, we found that the phenolic-rich ferment prevented the transepithelial electrical resistance (TEER) drop on the second challenge and modulated the transcriptomic profile assessed by RNA-seq. Our findings indicate that the Caco-2 cellular responses mentioned above were SCFA-independent and likely due to the differential impact of phenolic compound load after colonic fermentation of grape pomace skin.

Candidates:
A. Although this notion is widely accepted, supporting evidence remains scarce.
B. This bioprocessing weakens the matrix and promotes the release of the phenolic compounds, which then exert beneficial effects on intestinal function.
C. The evidence does not state that this bioprocessing weakens the matrix and promotes the release of the phenolic compounds, which then exert beneficial effects on intestinal function.
D. Although this notion is not widely accepted, supporting evidence remains scarce.
E. In this study, we subjected grape pomace skin to in vitro digestion to obtain an indigestible fraction suitable for SHIME bioreactors.
F. The evidence does not state that in this study, we subjected grape pomace skin to in vitro digestion to obtain an indigestible fraction suitable for SHIME bioreactors.
G. The association between dietary fiber and phenolic compounds allows the latter to reach the colon, where most polysaccharides undergo fermentation.
H. The evidence does not state that the association between dietary fiber and phenolic compounds allows the latter to reach the colon, where most polysaccharides undergo fermentation.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Genetically modified (GM) lactic acid bacteria (LAB) are gaining attention as tools for innovation in the food sector, health applications, and industrial processes. LAB have long been used safely due to their GRAS/QPS status, making them suitable for improving fermentation and synthesizing specific and beneficial metabolites. Advances in genomics and gene editing have significantly expanded the available tools, ranging from classical mutagenesis to site-specific recombination, homologous recombination in non-coding regions, CRISPR-based systems, and food-grade chromosomal integration. These approaches enable the insertion of desired genes and the development of engineered strains with tailored functionalities. GM-LAB are also being studied as live delivery systems for therapeutic molecules, including cytokines, hormones, antimicrobial peptides, and vaccine antigens. Engineered strains of Lactococcus lactis and Lactobacillus spp. have yielded promising outcomes in applications such as mucosal immunization, modulation of inflammatory and metabolic responses, and inhibition of pathogenic microorganisms, including multidrug-resistant bacteria. From an industrial perspective, several studies highlight their potential for cost-effective recombinant protein production and the synthesis of high-value metabolites through fermentation. However, within the European Union, their use is subject to stringent regulatory oversight, requiring comprehensive molecular and environmental risk assessments, careful evaluation of horizontal gene transfer, and a preference for markerless chromosomal integrations. Despite these constraints, GM-LAB offer significant potential to improve food quality, sustainability, and human health.

Candidates:
A. Genetically modified (GM) lactic acid bacteria (LAB) are gaining attention as tools for innovation in the food sector, health applications, and industrial processes.
B. These approaches enable the insertion of desired genes and the development of engineered strains with tailored functionalities.
C. Genetically modified (GM) lactic acid bacteria (LAB) are not gaining attention as tools for innovation in the food sector, health applications, and industrial processes.
D. The evidence does not state that advances in genomics and gene editing have significantly expanded the available tools, ranging from classical mutagenesis to site-specific recombination, homologous recombination in non-coding regions, CRISPR-based systems, and food-grade chromosomal integration.
E. Advances in genomics and gene editing have significantly expanded the available tools, ranging from classical mutagenesis to site-specific recombination, homologous recombination in non-coding regions, CRISPR-based systems, and food-grade chromosomal integration.
F. The evidence does not state that these approaches enable the insertion of desired genes and the development of engineered strains with tailored functionalities.
G. The evidence does not state that lAB have long been used safely due to their GRAS/QPS status, making them suitable for improving fermentation and synthesizing specific and beneficial metabolites.
H. LAB have long been used safely due to their GRAS/QPS status, making them suitable for improving fermentation and synthesizing specific and beneficial metabolites.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Microbial lipids have emerged as a promising sustainable alternative to plant- and petroleum-derived oils, with applications spanning biofuels, oleochemicals, nutraceuticals, and specialty materials. Significant advances in metabolic engineering and strain development have increased lipid production capacity across diverse microorganisms. Numerous reviews have summarized the biological and metabolic advances in this field, highlighting significant progress in metabolic engineering and strain development that has increased lipid production capacity across diverse microorganisms. However, translating these gains into economically viable industrial processes remains a major challenge. This review examines process engineering strategies for microbial lipid production across the full bioprocessing pipeline, from laboratory-scale strain evolution to industrial-scale operation. We discuss recent developments in adaptive laboratory evolution, systems-guided strain optimization, and robustness engineering, emphasizing their implications for process performance. Key bioprocess parameters—including substrate selection, nutrient limitation strategies, reactor design, oxygen transfer, and process control—are critically evaluated for their impact on lipid yield, productivity, and scalability. Furthermore, downstream processing considerations and techno-economic constraints are analyzed in the context of large-scale implementation. By integrating strain-level innovations with process engineering principles, this review highlights current bottlenecks, emerging solutions, and future directions for achieving efficient and scalable microbial lipid biomanufacturing.

Candidates:
A. Numerous reviews have summarized the biological and metabolic advances in this field, highlighting significant progress in metabolic engineering and strain development that has increased lipid production capacity across diverse microorganisms.
B. However, translating these gains into economically viable industrial processes remains a major challenge.
C. Significant advances in metabolic engineering and strain development have increased lipid production capacity across diverse microorganisms.
D. The evidence does not state that however, translating these gains into economically viable industrial processes remains a major challenge.
E. The evidence does not state that microbial lipids have emerged as a promising sustainable alternative to plant- and petroleum-derived oils, with applications spanning biofuels, oleochemicals, nutraceuticals, and specialty materials.
F. The evidence does not state that numerous reviews have summarized the biological and metabolic advances in this field, highlighting significant progress in metabolic engineering and strain development that has increased lipid production capacity across diverse microorganisms.
G. Microbial lipids have emerged as a promising sustainable alternative to plant- and petroleum-derived oils, with applications spanning biofuels, oleochemicals, nutraceuticals, and specialty materials.
H. The evidence does not state that significant advances in metabolic engineering and strain development have increased lipid production capacity across diverse microorganisms.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Akkermansia muciniphila , a next-generation probiotic in the human intestinal mucus layer, exhibits significant health-promoting properties. However, traditional static culture systems fail to replicate the dynamic peristaltic environment of the gastrointestinal tract, limiting understanding of its metabolic characteristics. This study employed an improved gastrointestinal bioreactor simulating intestinal peristalsis to investigate A. muciniphila growth dynamics and metabolomic profiles under dynamic conditions. Dynamic cultivation significantly enhanced bacterial growth. Biomass reached 1.32 ± 0.03 g/L in bovine heart infusion (BHI) medium and 2.03 ± 0.05 g/L in BHI supplemented with 2.5 g/L porcine mucin. These values represent increases of 45.05% and 123.08% relative to static BHI cultures, respectively. Dynamic conditions markedly elevated short-chain fatty acid production (acetic, propionic, isobutyric, isovaleric acids). Untargeted metabolomics identified 1463 metabolites with 1294 showing significant differential expression. Dynamic cultivation substantially altered amino acid biosynthesis, fatty acid, purine, and pyrimidine metabolism. These findings advance the understanding of A. muciniphila physiology and provide insights into its metabolic characteristics under simulated intestinal conditions.

Candidates:
A. However, traditional static culture systems fail to replicate the dynamic peristaltic environment of the gastrointestinal tract, limiting understanding of its metabolic characteristics.
B. muciniphila growth dynamics and metabolomic profiles under dynamic conditions.
C. Akkermansia muciniphila , a next-generation probiotic in the human intestinal mucus layer, exhibits significant health-promoting properties.
D. This study employed an improved gastrointestinal bioreactor simulating intestinal peristalsis to investigate A.
E. The evidence does not state that muciniphila growth dynamics and metabolomic profiles under dynamic conditions.
F. The evidence does not state that this study employed an improved gastrointestinal bioreactor simulating intestinal peristalsis to investigate A.
G. The evidence does not state that akkermansia muciniphila , a next-generation probiotic in the human intestinal mucus layer, exhibits significant health-promoting properties.
H. The evidence does not state that however, traditional static culture systems fail to replicate the dynamic peristaltic environment of the gastrointestinal tract, limiting understanding of its metabolic characteristics.