The Hub supports core research in vaccine manufacturing. It comprises manufacturing research themes essential to address the main limitations in vaccine development, manufacture and rapid response. To embrace the diversity of antigen and vaccine type, the Grand Challenges (GC) work is demonstrated using four common modalities (VLPs, RNA, viral vectored and conjugate vaccines) widely applicable to known outbreak pathogens. The work will not be limited to viral diseases as bacterial pathogens are a constant global threat to human and animal health.
For most bacterial pathogens there are no currently licensed vaccines, and this represents a serious gap in national security particularly against biothreat agents and multiantibiotic resistant bacteria. The Hub will address two GC themes. In order to be better prepared for a future pandemic response, GC1 will develop new tools and technologies to speed up manufacture, but also facilitate distribution, storage and administration. To meet the Hub vision as a sustainability leader in the sector, GC2 will focus on tools that support better understanding of the impact on the environment and will facilitate waste re-use and facility design for (re)manufacture.
WP 1.1 Improved Platforms
Microbial pathogens engineering
Led by Brendan Wren (LSHTM), Catherine Green (University of Oxford), Martina Micheletti (UCL)
PDRAs: Elizabeth Atkins (LSHTM), Ivana Stolfa (UCL)
The aim of this work is to develop two new recombinant vaccines against bacterial pathogens using a novel Protein-Glycan Conjugation Technology (PGCT), which will be produced as an exemplar in collaboration with UCL (microscale and scaling studies, WP1.2) and Oxford (Clinical Biomanufacturing Facility). These vaccines will protect against Francisella tularensis (biothreat, protein coupled to lipopoylysaccharides) and Streptococcus pneumoniae serotype 1 (multi-antibiotic resistance threat, protein coupled to capsule).
Completed year one deliverables:
- A Streptococcus pneumoniae serotype 1 vaccine has been constructed and the product tested and quantified.
- Ten Streptococcus pneumoniae capsular polysaccharide serotypes have been cloned, towards a standardised procedure for optimally cloning all Streptococcus pneumoniae 100 serotypes.
- A prototype glycoconjugate vaccine against Francisella tularensis has been developed.
Progress on year two deliverables:
- Expression of the Streptococcus pneumoniae serotype 1 vaccine has been improved from nanograms to micrograms in small-scale bacterial This is currently being scaled up in shake flasks for immunological tests in mice. This improvement was achieved by modifying the oligosaccharyltransferase (OST) expression plasmids to put them under arabinose induction, and by optimising the growth media.
- The Francisella tularensis prototype vaccine is now being expressed in improved coli strains as well as in a CLM24 strain. Strain named Crow has a mutation to detoxify lipid A, and strain named Raven is a derivative of Crow with a copy of the OST pglB integrated onto the bacterial chromosome. In a previous study the best glycosylation levels for a glycoconjugate vaccine against Campylobacter jejuni were seen with both a copy of pglB on the chromosome and on a plasmid, and we will be testing if the same principle applies here. These are currently with Ivana Stolfa at UCL to begin development work on using both manual and automated (Tecan) systems.
Novel platform development
Led by Nicola Stonehouse (University of Leeds)
PDRA: Emma Wroblewski (University of Leeds)
The aim of this work is to develop a platform antigen display technology, VelcroVax, where the capsid/core of hepatitis B virus is used as a scaffold in order to ‘present’ antigens. This system mirrors analogous two-component VLP vaccine delivery platforms using technologies in which a small peptide spontaneously forms a covalent (isopeptide) bond with its partner protein (e.g. Spytag/Spycatcher). These systems further demonstrate the validity of this approach to vaccine platform development, but VelcroVax is potentially more compatible with the recognition and presentation of oligomeric structures such as trimeric viral glycoproteins.
Completed year one deliverables:
Work is progressing on scaling up the production of virus-like particles (VLPs). Trials have been undertaken using a range of different chromatography approaches with the VLPs and the team is now in the process of optimising buffer conditions and evaluating yields in order to balance losses with purity.
Work is underway to explore alternative yeast strains and expression approaches. The group has compared constitutive expression with their previous approaches of AOX induction in terms of yields and is now starting to use alternative yeast promotors with their Pichia constructs. These should allow for more controllable expression of the protease and may increase VLP yield.
Progress on year two deliverables:
Alongside the work on scaling up production of virus-like particles (VLPs), the team at the University of Leeds is undertaking Pichia fermentation experiments with colleagues at UCL to produce EVD68 VLPs. This builds on previous work with polio VLPs, where scaling studies were undertaken at the Centre for Process Innovation (CPI) and funded by the original VaxHub grant.
Studies aimed at the characterisation of the VelcroVax VLPs platform in terms of the efficiency of dual antigen presentation have begun. Previous ELISA assays are being supplemented with mass spectrometry to assess the mass of decorated VLPs and therefore the degree of decoration.
Strategies to enhance immunogenicity – VLP focus
Led by Sandy Douglas and Sumi Biswas (University of Oxford)
PDRA: Catherine Cherry (University of Oxford)
The aim of this work is to optimise an existing VLP platform, produce a sufficient supply for trials and develop suitable analytics for the characterisation of the VLPs and evaluation of their stability.
Completed year one deliverables:
Two different approaches for producing recombinant antigens have been evaluated. In both cases, the potential to provide a sufficient supply of recombinant antigen for conjugation to VLP for a phase I clinical trial was established.
To support analytical development, HPLC-SEC has been used for evaluating the degree of conjugation of antigen to VLP, alongside the evaluation of antigen-VLP particle stability in formulations with different excipients.
Progress on year two deliverables:
Into year two, the two antigen production approaches will be further optimised for production at larger scale in bioreactors.
Strategies to enhance immunogenicity – Adenoviral vector focus
Led by Sarah Gilbert (University of Oxford)
RA: Reshma Kailath (University of Oxford)
This work focuses on the optimization of adenoviral vectored vaccines and will adopt two different approaches to improving immunogenicity: (i) optimised production and purification methods to increase the proportion of infectious particles, and (ii) use of lipid nanoparticles encapsulation to increase infectivity.
Completed year one deliverables:
Data on encapsulation of adenoviruses with different formulations were chosen based on a thorough literature review and expert recommendations. The ChAdOx1 were encapsulated in these liposomes. The characterisation analysis included Dynamic Light Scattering (DLS) and negative stain microscopy and yielded promising results. Among the various formulations, one was selected for transduction studies based on EM results, which demonstrated promising encapsulation with that particular formulation.
Data on in vitro infectivity of encapsulated adenovirus in different cell lines were collected. Multiple transduction studies were conducted using varying concentrations of liposomes complexed with adenoviruses. However, no increase in infectivity was observed with this formulation. To assess encapsulation efficiency, the team needed to identify a cell line resistant to ChAdOx1 infection, specifically one lacking CAR receptors. Several human and animal cell lines were tested, including A549, MDCK, CHO, CHO-K1, and BHK21.
Progress on year two deliverables:
The custom-synthesised cationic lipopeptide was received in December 2024. The lipopeptide:DNA complex is referred to as lipoplexes from herein. For the lipoplex formulations, two DNA constructs expressing the GFP protein were used: plasmid DNA (standard mammalian expression plasmid) and BAC DNA (containing the ChAdOx1 Adenovirus genome). The standard mammalian expression plasmid was used to test the effectiveness of the lipoplex and ChAdOx1 Adenovirus genome is being used as this is the focus of the research.
The initial experiment was designed to optimise the DNA encapsulation efficiency by determining the optimal N/P ratio required for effective encapsulation using the lipopeptide in combination with a helper lipid. A gel retardation assay was used for this assessment, testing the lipopeptide with varying DNA concentrations across different N/P ratios. The results indicated that at certain N/P ratios with higher DNA amounts, some DNA remained unbound. Furthermore, when the lipoplex was treated with nuclease to eliminate unbound DNA, no changes in band patterns were observed, confirming effective DNA encapsulation.
To assess cellular entry, a transduction study was performed in two distinct human cell lines: A549 (lung carcinoma cells) and 293A (human embryonic kidney cells). Multiple control samples were included, such as DNA alone (to assess passive entry), using commercial transfection reagent (lipofectamine3000) as a positive control and Empty lipopeptide to evaluate cytotoxicity. Fluorescence was visualised using EVOS microscope confirming that the lipoplex formulation effectively enhanced DNA entry through evaluation of GFP expression in both cell lines.
The next phase of the study will involve testing the formulations under various pH conditions to assess whether they maintain a positive charge at both low and high pH as different cell compartments have acidic and alkaline pH’s. This will be crucial for ensuring efficient DNA entry into cells during transfection. Once these conditions are optimised, further testing will be conducted in additional cell lines to evaluate the broader applicability of the lipoplex formulations.
Modular antigen display on nanoparticles
Led by Stefanie Frank and Michael Thomas (UCL)
PDRA: Yuqian Ou (UCL)
Studies in this WP on antigen display on protein nanoparticles are aligned with WP1.3 on ‘Enhancing mRNA stability by novel encapsulation strategies’. As WP1.3 develops, the group will be creating combinations of antigen display and antigen encapsulation with the aim to improve immunogenicity and delivery.
Completed year one deliverables:
Dr Yuqian Ou was recruited in Jan 2025. She will work across WP1.1 and WP1.3 but will initially focus on WP1.3. Previous work on modular antigen display in our group will provide the background for this WP and is currently being prepared for publication with Dr Ou’s support.
Progress on year two deliverables:
First steps will focus on the design of particles displaying molecules that aid in the targeted delivery of particles created in WP1.3.
WP 1.2 Rapid development and manufacture
Process intensification and distributed manufacture & Digitalised automated workflow
Led by Sandy Douglas and Catherine Green (University of Oxford)
RA: Jacqueline Vieira (University of Oxford)
Completed year one deliverables:
- Experiments were planned with a new Expi293F inducible cell line due to higher viral productivity as well as relatively shorter doubling time. All subsequent experiments will be performed using Expi293F inducible A higher viral titre and higher cell specific productivity were obtained from the Expi293F inducible cell line compared to the Trex HEK cells. A full media replacement was also carried out to further evaluate the two cell lines’ performance.
- Expi293F inducible cell line will be further investigated to understand any limitations in productivity.
Progress on year two deliverables:
Once the new cell line is suitably characterised, the aim will be to understand and characterise the relationship between upstream process conditions/downstream purification and product quality (e.g HCP content).
Process intensification and distributed manufacture & Digitalised automated workflow
Led by Duygu Dikicioglu and Martina Micheletti (UCL)
PDRA: Cheng Zhang (Apr-Aug 2024), Ryan Mellor (Oct 2024-present) (UCL)
Extremely rapid deployment of technologies for optimal process development is not trivial and an efficient approach necessitates consideration of the whole bioprocess. The aim of this work is to develop an automated system that mimics the whole manufacturing process and can be developed at short notice and deployed to support pandemic response.
Completed year one deliverables:
The initial PDRA, Dr Cheng Zhang, was recruited in April 2024 but subsequently left and was replaced by Dr Ryan Mellor who is in post and ensuring continuation of the project activities. Synthace and Tecan training was completed in December 2024 (Synthace) and early 2025 (Tecan).
The Synthace software setup has been completed, and the software is ready for use. Tecan platform setup and maintenance has also been completed and is available for staff to familiarise themselves with after completing the required training.
Progress on year two deliverables:
Work has begun on developing an online platform for building and sharing vaccine bioprocess workflow, the first release of which is now online for internal feedback. Following Tecan training, work will begin to implement automated experiments, initially based on the work of Samaras et al (2021), and then expanding the capabilities of the software and hardware later in 2025.
Development and optimisation of automated data capture and analysis pipeline for the validated workflow will begin in Q2 2025 with development and validation of at least one additional automated workflow for bioprocessing applications planned for Q3 2025. In Q4 2025, work is planned for the implementation of remote operation capabilities and demonstration of successful workflow execution from a remote location.
Microscale processing and robust scaling strategies
Led by Martina Micheletti (UCL)
PDRA: Ivana Stolfa (UCL)
The aim of this work is to use microscale processing, scale-down tools and knowledge of scaling fundamentals for experimental design and data collection on different vaccine technologies (from WP1.1) for validation and scaling purposes. This work package aims to advance Protein Glycan Coupling Technology (PGCT) for the production of glycoconjugate vaccines. The team will develop a flexible production platform, using as a prototype the conjugation of Francisella tularensis O-antigen to the Pseudomonas aeruginosa carrier protein exotoxin A, facilitated by the Campylobacter jejuni PglB oligosaccharyltransferase. Francisella tularensis is classified as a Class A bioterrorism agent due to its low infectious rate, high fatality rate and ease of aerosol distribution.
Currently, no vaccines are available to protect against Francisella tularensis infection.
The focus will be on optimising production at bench scale (20 mL), microscale (2 mL) using the Tecan Freedom Evo, and bioreactor scale. The previously developed manufacturing process has already been adapted for the production of a recombinant F. tularensis glycoconjugate vaccine expressed in Escherichia coli.
Completed year one deliverables:
The PDRA on this project has been recruited and is in post. A scoping literature exercise has been carried out to inform publication of an overview of scaling strategies to better define and understand gaps.
Familiarisation and knowledge transfer with other researchers has started to map out challenges to inform miniaturisation strategies.
Progress on year two deliverables:
Process optimisation is currently underway using a Design of Experiment (DOE) approach, evaluating key parameters such as optical density at induction (OD600) and growth temperature of E. coli to maximise yield.
Upon completion of this work package, efforts will shift to assessing the use of a defined medium to further enhance process yield and translating the optimised process to bioreactor scale.
Next steps are:
- Adaptation of previously developed protocol for PGCT technology to produce bacterial vaccine against Francisella Tularensis (strain, periplasmic extraction protocol, induction, alternative method to detect protein and glycan)
- Evaluation of bacterial growth conditions (media composition, temperature, induction time) manually
- Automated evaluation of bacterial growth conditions (media composition, temperature, induction time)
- Scale-up of PGCT technology using bioreactors
Product impurities and separation
Led by Daniel Bracewell (UCL)
DRA: Braulio Carrillo Sanchez (UCL)
The aim of this work is to develop processes able to remove product- related impurities that affect infectivity for adenovirus-vectored products. In addition, the team will focus on downstream processing opportunities by investigating the potential of a polishing chromatography step based on affinity chromatography using both commercial and novel ligands.
Completed year one deliverables:
Dr Braulio Carrillo Sanchez was recruited in January 2025 as a PDRA to focus on characterisation of adenovirus particle heterogeneity from affinity separations.
Identifying potential new cell-based analytics for critical quality attributes of adenovirus based on the receptor the virus uses to gain entry into cells – the team has discussed the shortcomings and advantages of current cell- based assays with University of Oxford colleagues in the VaxHub network, to determine possible routes forward with particular focus on cell types and end point analytics.
Identifying which critical quality attributes / features of adenovirus might be suitable for the development of separations to reduce product heterogeneity – ongoing interest in getting a definitive understanding of adenovirus fibre proteins and infectivity as a quality attribute. Promising structural biology insights are being obtained from work carried out by a previous doctoral student in Professor Bracewell’s team. Within the VaxHub network, the team has engaged in collaborative discussions with colleagues at the University of Oxford on how this could be investigated further.
Progress on year two deliverables:
- Integration of a novel Ad5 affinity peptide separation process at UCL Biochemical Engineering, first introduced by collaborators at North Carolina State This approach enables mild bioprocessing conditions during adenovirus-vectored vaccine production that could promote the recovery of functional viral particles.
- Further characterisation of viral particle populations separated by affinity The team will seek to expand the use of the L1-52/55k marker for immature/non-infective virions and correlate this with in vitro infectivity and recovery performance.
- Exploring the feasibility of super-resolution microscopy for viral particle visualisation and characterisation at the single particle level. This will seek to leverage the single-molecule localisation microscopy (SMLM) capabilities of the ONI Nanoimager instrument.
WP 1.3 Mucosal and thermostable formulations
Bio-based excipients and adjuvants
Led by James Winterburn (University of Manchester)
PDRA: Phavit Wongsirichot (University of Manchester, July- August 2024)
The aim of this work is to explore how fermentation processes for the production of novel materials suitable for vaccine formulation and delivery, e.g. biosurfactants, can be engineered to be robust, scalable and with consideration of both technical feasibility and economic viability, and materials produced tested within the Hub.
Completed year one deliverables:
The initial focus of this work has been an investigation of sustainable formulations and exploration of adjuvants. During Year one a rigorous scoping exercise was conducted to identify suitable candidate molecules.
Progress on year two deliverables:
- (Re)recruitment of a PDRA with required expertise
- Production of molecules of interest, as identified from literature review– to be started once PDRA is in post.
- Production of bio-based materials for others in VaxHub Sustainable – ongoing collaboration with Professor Sudaxshina Murdan (UCL) on bio-based formulation.
Enhancing mRNA stability by novel encapsulation strategies
Led by Stefanie Frank and Michael Thomas (UCL)
PDRA: Yuqian Ou (UCL)
This work package focuses on exploring novel encapsulation strategies for mRNA vaccines using self-assembling protein nanoparticles to improve stability, storage, and administration.
Completed year one deliverables:
Dr Yuqian Ou joined the team in January 2025 as PDRA working across WP1.1 and 1.3.
Encapsulation of RNA into nanoparticles was completed using an in vitro disassembly and reassembly approach based on recent studies in the group (Van de Steen et al., 2024, DOI: 10.1021/acsabm.3c01153). Particle disassembly and reassembly were optimised and reproducible loading of small reporter RNA achieved. The encapsulated reporter RNA has been demonstrated to have resistance to RNase digestion in vitro.
Progress on year two deliverables:
Next, analytical tools will be applied to characterise the stability of loaded reporter RNA molecules. Further steps will include incorporating larger mRNA molecules and evaluating their integrity and stability. Cell-uptake studies of loaded particles are also in progress.
In vitro models will be developed with subsequent evaluations focusing on particle endocytosis pathways and the efficiency of mRNA translation.
Mucosal vaccination strategies and Platform sublingual formulations
Led by Sudaxshina Murdan (UCL) and Nicola Stonehouse (University of Leeds)
PDRAs: Mohamed Yousif (UCL) and Emma Wroblewski (University of Leeds)
This work aims to progress mucosal vaccination and investigate effective mucosal adjuvants.
Completed year one deliverables:
Polio type 1 vaccine (PV1) virus-like particles (VLPs) have been investigated. VLPs produced in Leeds were tested at UCL in vitro and in vivo studies in an animal model. Immunogenicity of VLPs following sublingual and/or parenteral delivery, using several dosing regimens and vaccine adjuvants, were tested in vivo in the mouse model.
In vivo testing of new vaccine adjuvants by the sublingual route has been achieved with several metal nanoparticles tested as vaccine adjuvants.
As the VLP and metal nanoparticles used as an antigen and potential adjuvants, respectively, metal nanoparticles did not enhance antigen-specific systemic and mucosal immune responses to sublingually administered VLPs.
Progress on year two deliverables:
Potential of metal nanoparticles to dose-spare VLPs with respect to enhancing systemic and mucosal immune responses. An in vivo study to investigate any dose-sparing effects of metal nanoparticles on VLPs will be carried out in June – July 2025. Methods to stabilise purchased metal nanoparticles with respect to their particle size, shape and surface properties was developed using freeze-drying and appropriate cryo- and lyo-protectants. Metal nanoparticles of silver, gold and zinc oxide, in a range of particle sizes (10–400 nm) were successfully freeze-dried using protectants while maintaining their physico-chemical properties. Freeze- drying will enable the inclusion of the metal nanoparticles in future freeze- dried vaccines.
Efficacy of a new adjuvant to enhance systemic and mucosal immune responses and cellular immunity to sublingually and parenterally administered VLPs and standard polio vaccine (IPV). The team acquired training in the measurement of cellular immunity and an experiment will be conducted in the US in a new collaborator’s laboratory.
Work will be carried out in June – August 2025 on systemic and mucosal immune responses following sublingual immunisation with VelcroVax VLPs decorated with RSV-G protein antigens. At the same time the team will work to answer a fundamental question on whether the vaccine adjuvant effect of metal nanoparticles with respect to systemic and mucosal immune responses is similar, regardless of the nature of the VLP (e.g. polio and VelcroVax VLPs).
The next steps in this work package will be to explore the ability of sublingual immunisation to elicit cellular immune responses.
WP 2.1 Understanding bioprocess sustainability impact
LCA methodology for hotspot analysis and decision- making and Development of a comprehensive inventory for biotechnological manufacture
Led by Brenda Parker (UCL)
RA: Rita Morais (UCL); PDRA: recruitment ongoing
This work will address two key issues, while learning from recognised sustainability Project Partners. Firstly, within the context of the LCA framework the team will work on a “best practice” document to ensure that the scope and preparation of the life cycle inventory can be refined so that outputs are meaningful. This will facilitate hotspot analysis or decision- making in process design. Secondly, the team will source data and boost the range of well-defined inputs to enable more precise life cycle inventories reflective of the actual materials used in bioprocesses. The study will examine the process and environment using a cradle-to-gate approach, including an Inventory analysis, a detailed impact assessment followed by an interpretation and process review stage.
Completed year one deliverables:
Rita Morais joined the VaxHub Sustainable team as a Research Assistant for Sustainable Bioprocess Design in April 2024, focusing on researching opportunities for industrial symbiosis and the circular economy within the industry. Rita has been actively creating a biomaterials library, working on the next iteration of the water bioremediation project, and participating in meetings with companies to learn of opportunities for industrial symbiosis.
Work has been successful in meeting with industry partners to scope sustainability topics and priorities for LCA research. The team has met with members of the AstraZeneca team online, and following Sartorius’ visit to UCL Biochemical Engineering in April 2024, where the team presented their overall sustainability goals, they have begun to build a SharePoint structure to be able to capture information about industry priorities to enable the assessment of common themes, as well as to track progress.
Work on the SharePoint site is ongoing, Rita Morais has been liaising with the Hub Partners Relations Manager in regards to data capture about sustainability during partner meetings.
A related student research project is now completed to benchmark the conventional plasmid DNA manufacturing route with a view to a sustainability evaluation for the doggy-bone DNA route.
Public engagement activity on water recycling and remediation resulted in work from the RA being presented at the Water Pressure Exhibition, Hamburg – opened March 2024. An output of this public engagement activity is the Augmented Polycultures installation, featured in the Water Pressure exhibition at the Museum of Arts and Crafts in Hamburg. This installation represents a decentralised approach to bioremediation of greywater and surface run-off for future buildings, through the use of algae biofilms inoculated in 3D printed ceramic tiles. The Augmented Polycultures installation at the Museum für Gestaltung Zürich was included as part of the Water: Designing for the Future exhibition in 2024, Zurich, Switzerland. Following the installation and exhibition inauguration, the museum produced an engaging video to explain the project to a broader audience.
Investigation of waste streams as future resource and Design for (re)manufacture
Led by Brenda Parker (UCL)
RA: Rita Morais (UCL)
This work aims to answer the fundamental question of how we might plan for future facilities to be embedded within a circular economy and aims to address key aspects around the use of waste and novel facility design within the broader vaccine manufacturing industry.
Completed year one deliverables:
The aim was to construct a 1m2 prototype for water remediation and development of a testing protocol to look at N and P capture from wastewater. As a preliminary step to the prototype, a smaller-scale version was developed to evaluate the system. 3D-printed clay tiles were printed using a WASP, in which a variation of textures was tested in terms of fabrication feasibility. A draft testing protocol was established, and initial experiments are currently underway.
Progress on year two deliverables:
Building on the findings of the preliminary work in constructing a 1m2 prototype for water remediation and development of a testing protocol to look at N and P capture from wastewater, the design and conceptualization of the full-scale prototype are being completed. Discussions with the engineering workshop regarding fabrication have taken place, with production set to begin following necessary design adjustments. The next phase will focus on refining the design based on insights from previous small-scale tile tests, after which multiple design variations will be 3D-printed for the prototype.
Regarding beta testing of the biomaterials library with users and linkage of the site with robotic capabilities at UCL, discussions are ongoing on how to integrate this library within the Bio-Id fabrication workshop. The materials library has been successfully developed and is set to be tested within the Bio-Id context soon. Additionally, a system for adding new information is in development, along with comprehensive documentation of all fabrication tools used in each project to ensure seamless integration with the existing university platform.
Sustainability champions amongst the Hub membership will be identified and a workshop organised for the champions to engage with design methodology and propose potential exploratory projects linked to current waste challenges. Planning for the workshop has begun, the workshop will focus on engaging sustainability champions with design methodologies to explore innovative solutions for current waste challenges. Its structure is being developed to encourage collaboration and generate actionable project ideas.
One industry case study will be identified for the circular economy approach using bio-integrated design and subsequent material development. The team has been conducting initial research on collaboration opportunities with companies to repurpose their waste streams. Currently the team is identifying references for circular systems and potential partners to develop innovative solutions that enhance their upcycling process. Next steps involve meeting with the identified companies to explore possible collaborations and integration into their upcycling chain.
