3D imaging and analysis to unveil the impact of microparticles on the pellet morphology of filamentous fungi.

Controlling the morphology of filamentous fungi is crucial to improve the performance of fungal bioprocesses. Microparticle-enhanced cultivation (MPEC) increases productivity, most likely by changing the fungal morphology. However, due to a lack of appropriate methods, the exact impact of the added microparticles on the structural development of fungal pellets is mostly unexplored. In this study synchrotron radiation-based microcomputed tomography and three-dimensional (3D) image analysis were applied to unveil the detailed 3D incorporation of glass microparticles in nondestructed pellets of Aspergillus niger from MPEC. The developed method enabled the 3D analysis based on 375 pellets from various MPEC experiments. The total and locally resolved volume fractions of glass microparticles and hyphae were quantified for the first time. At increasing microparticle concentrations in the culture medium, pellets with lower hyphal fraction were obtained. However, the total volume of incorporated glass microparticles within the pellets did not necessarily increase. Furthermore, larger microparticles were less effective than smaller ones in reducing pellet density. However, the total volume of incorporated glass was larger for large microparticles. In addition, analysis of MPEC pellets from different times of cultivation indicated that spore agglomeration is decisive for the development of MPEC pellets. The developed 3D morphometric analysis method and the presented results will promote the general understanding and further development of MPEC for industrial application.

Breaking down barriers: comprehensive functional analysis of the Aspergillus niger chitin synthase repertoire.

BACKGROUND: Members of the fungal kingdom are heterotrophic eukaryotes encased in a chitin containing cell wall. This polymer is vital for cell wall stiffness and, ultimately, cell shape. Most fungal genomes contain numerous putative chitin synthase encoding genes. However, systematic functional analysis of the full chitin synthase catalogue in a given species is rare. This greatly limits fundamental understanding and potential applications of manipulating chitin synthesis across the fungal kingdom. RESULTS: In this study, we conducted in silico profiling and subsequently deleted all predicted chitin synthase encoding genes in the multipurpose cell factory Aspergillus niger. Phylogenetic analysis suggested nine chitin synthases evolved as three distinct groups. Transcript profiling and co-expression network construction revealed remarkably independent expression, strongly supporting specific role(s) for the respective chitin synthases. Deletion mutants confirmed all genes were dispensable for germination, yet impacted colony spore titres, chitin content at hyphal septa, and internal architecture of submerged fungal pellets. We were also able to assign specific roles to individual chitin synthases, including those impacting colony radial growth rates (ChsE, ChsF), lateral cell wall chitin content (CsmA), chemical genetic interactions with a secreted antifungal protein (CsmA, CsmB, ChsE, ChsF), resistance to therapeutics (ChsE), and those that modulated pellet diameter in liquid culture (ChsA, ChsB). From an applied perspective, we show chsF deletion increases total protein in culture supernatant over threefold compared to the control strain, indicating engineering filamentous fungal chitin content is a high priority yet underexplored strategy for strain optimization. CONCLUSION: This study has conducted extensive analysis for the full chitin synthase encoding gene repertoire of A. niger. For the first time we reveal both redundant and non-redundant functional roles of chitin synthases in this fungus. Our data shed light on the complex, multifaceted, and dynamic role of chitin in fungal growth, morphology, survival, and secretion, thus improving fundamental understanding and opening new avenues for biotechnological applications in fungi.

Selective anomer crystallization from aqueous solution: Monitoring lactose recovery under mutarotation limitation via attenuated total reflection Fourier-transform spectroscopy and theoretical rate analysis.

Lactose is typically produced via cooling crystallization either from whey or whey permeate (edible grade) or from aqueous solution (pharmaceutical grade). While in solution, lactose is present in 2 anomeric forms, α- and ß-lactose. During cooling crystallization under standard process conditions, only α-lactose crystallizes, depleting the solution of α-anomer. In practice, mutarotation kinetics are often assumed to be much faster than crystallization. However, some literature reports limitation of crystallization by mutarotation. In the present research, we investigate the influence of operating conditions on mutarotation in lactose crystallization and explore the existence of an operation regimen where mutarotation can be disregarded in the crystallization process. Therefore, we study crystallization from aqueous lactose solutions by inline monitoring of concentrations of α- and ß-lactose via attenuated total reflection Fourier-transform spectroscopy. By implementing a linear cooling profile of 9 K/h to a minimum temperature of 10°C, we measured a remarkable increase in ß/α ratio, reaching a maximum of 2.19. This ratio exceeds the equilibrium level by 36%. However, when the same cooling profile was applied to a minimum temperature of 25°C, the deviation was significantly lower, with a maximum ß/α ratio of 1.72, representing only an 8% deviation from equilibrium. We also performed a theoretical assessment of the influence of process parameters on crystallization kinetics. We conclude that mutarotation needs to be taken into consideration for efficient crystallization control if the crystal surface area and supersaturation are sufficiently high.

Mechanical, physical and thermal properties of composite materials produced with the basidiomycete Fomes fomentarius.

BACKGROUND: To achieve climate neutrality, fundamentally new concepts of circularity need to be implemented by the building sector as it contributes to 40% of anthropogenic CO2 emission. Fungal biotechnology can make a significant contribution here and help eliminate fossil dependency for building material production. Recently, we have shown that the medicinal polypore Fomes fomentarius feeds well on renewable lignocellulosic biomass and produces composite materials that could potentially replace fossil fuel-based expanded polystyrene as insulation material. RESULTS: In this study, we explored the mechanical, physical, and thermal properties of F. fomentarius-based composite materials in more detail and determined key performance parameters that are important to evaluate the usability of F. fomentarius-based composite materials in the construction sector. These parameters were determined according to European standards and included compressive strength, modulus of elasticity, thermal conductivity, water vapour permeability, and flammability of uncompressed composites as well as flexural strength, transverse tensile strength, and water absorption capacity of heat-pressed composites, among others. We could show that uncompressed composites obtained from F. fomentarius and hemp shives display a thermal conductivity of 0.044 W (m K)-1 which is in the range of natural organic fibres. A water vapour permeability of 1.72 and classification into flammability class B1 clearly surpasses fossil-based insulation materials including expanded polystyrene and polyurethane. We could furthermore show that heat-pressing can be used to reliably generate stiff and firm particleboards that have the potential to replace current wood-based particleboards that contain synthetic additives. X-ray microcomputed tomography finally visualized for the first time the growth of hyphae of F. fomentarius on and into the hemp shive substrates and generated high-resolution images of the microstructure of F. fomentarius-based composites. CONCLUSION: This study demonstrates that fungal-based composites produced with F. fomentarius partially meet or even exceed key performance parameters of currently used fossil fuel-based insulation materials and can also be used to replace particleboards.

Investigating the Role of Odorant-Polymer Interactions in the Aroma Perception of Red Wine: A Density Functional Theory-Based Approach.

The aroma of red wine results from the intricate interplay between aroma compounds (odorants) and complex polymers generated during fermentation. This study combines density functional theory (DFT), human sensory experiments, and nuclear magnetic resonance to investigate the impact of odorant-polymer interactions on wine aroma. Molecular aggregation patterns of odorants with polymer segments are identified, indicating the crucial role of intermolecular noncovalent interactions, such as hydrogen bonds and van der Waals interactions, in stabilizing odorant-polymer conformations. Certain odorants, including 3-isobutyl-2-methoxypyrazine and cis-whisky lactone, exhibit high binding affinity to specific polymer segments, such as (+)-catechin and p-coumaric acid, resulting in substantial changes in the perceived aroma. Their strong binding affinities correlate with changes in sensory experiments for binary mixtures. The results provide insights into the molecular mechanisms of odorant-polymer interactions in red wine with the potential of DFT calculations as a tool for predicting and tailoring red wine aroma.

On the reproducibility of extrusion-based bioprinting: round robin study on standardization in the field.

The outcome of three-dimensional (3D) bioprinting heavily depends, amongst others, on the interaction between the developed bioink, the printing process, and the printing equipment. However, if this interplay is ensured, bioprinting promises unmatched possibilities in the health care area. To pave the way for comparing newly developed biomaterials, clinical studies, and medical applications (i.e. printed organs, patient-specific tissues), there is a great need for standardization of manufacturing methods in order to enable technology transfers. Despite the importance of such standardization, there is currently a tremendous lack of empirical data that examines the reproducibility and robustness of production in more than one location at a time. In this work, we present data derived from a round robin test for extrusion-based 3D printing performance comprising 12 different academic laboratories throughout Germany and analyze the respective prints using automated image analysis (IA) in three independent academic groups. The fabrication of objects from polymer solutions was standardized as much as currently possible to allow studying the comparability of results from different laboratories. This study has led to the conclusion that current standardization conditions still leave room for the intervention of operators due to missing automation of the equipment. This affects significantly the reproducibility and comparability of bioprinting experiments in multiple laboratories. Nevertheless, automated IA proved to be a suitable methodology for quality assurance as three independently developed workflows achieved similar results. Moreover, the extracted data describing geometric features showed how the function of printers affects the quality of the printed object. A significant step toward standardization of the process was made as an infrastructure for distribution of material and methods, as well as for data transfer and storage was successfully established.

Influence of Flow Rate, Particle Size, and Temperature on Espresso Extraction Kinetics.

Brewing espresso coffee (EC) is considered a craft and, by some, even an art. Therefore, in this study, we systematically investigated the influence of coffee grinding, water flow rate, and temperature on the extraction kinetics of representative EC components, employing a central composite experimental design. The extraction kinetics of trigonelline, caffeine, 5-caffeoylquinic acid (5-CQA), and Total Dissolved Solids (TDS) were determined by collecting and analyzing ten consecutive fractions during the EC brewing process. From the extraction kinetics, the component masses in the cup were calculated for Ristretto, Espresso, and Espresso Lungo. The analysis of the studied parameters revealed that flow rate had the strongest effect on the component mass in the cup. The intensity of the flow rate influence was more pronounced at finer grindings and higher water temperatures. Overall, the observed influences were minor compared to changes resulting from differences in total extracted EC mass.

Synchrotron radiation-based microcomputed tomography for three-dimensional growth analysis of Aspergillus niger pellets.

Filamentous fungi produce a wide range of relevant biotechnological compounds. The close relationship between fungal morphology and productivity has led to a variety of analytical methods to quantify their macromorphology. Nevertheless, only a µ-computed tomography (µ-CT) based method allows a detailed analysis of the 3D micromorphology of fungal pellets. However, the low sample throughput of a laboratory µ-CT limits the tracking of the micromorphological evolution of a statistically representative number of submerged cultivated fungal pellets over time. To meet this challenge, we applied synchrotron radiation-based X-ray microtomography at the Deutsches Elektronen-Synchrotron [German Electron Synchrotron Research Center], resulting in 19,940 3D analyzed individual fungal pellets that were obtained from 26 sampling points during a 48 h Aspergillus niger submerged batch cultivation. For each of the pellets, we were able to determine micromorphological properties such as number and density of spores, tips, branching points, and hyphae. The computed data allowed us to monitor the growth of submerged cultivated fungal pellets in highly resolved 3D for the first time. The generated morphological database from synchrotron measurements can be used to understand, describe, and model the growth of filamentous fungal cultivations.

Development and Characterization of a 96-Well Exposure System for Safety Assessment of Nanomaterials.

In this study, a 96-well exposure system for safety assessment of nanomaterials is developed and characterized using an air-liquid interface lung epithelial model. This system is designed for sequential nebulization. Distribution studies verify the reproducible distribution over all 96 wells, with lower insert-to-insert variability compared to non-sequential application. With a first set of chemicals (TritonX), drugs (Bortezomib), and nanomaterials (silver nanoparticles and (non-)fluorescent crystalline nanocellulose), sequential exposure studies are performed with human lung epithelial cells followed by quantification of the deposited mass and of cell viability. The developed exposure system offers for the first time the possibility of exposing an air-liquid interface model in a 96-well format, resulting in high-throughput rates, combined with the feature for sequential dosing. This exposure system allows the possibility of creating dose-response curves resulting in the generation of more reliable cell-based assay data for many types of applications, such as safety analysis. In addition to chemicals and drugs, nanomaterials with spherical shapes, but also morphologically more complex nanostructures can be exposed sequentially with high efficiency. This allows new perspectives on in vivo-like and animal-free approaches for chemical and pharmaceutical safety assessment, in line with the 3R principle of replacing and reducing animal experiments.

Predicting the essential oil composition in supercritical carbon dioxide extracts from hop pellets using mathematical modeling.

Supercritical fluid extraction from hops (Humulus lupulus L.) can be used to extract essential oil for the flavoring of beer. With a special focus on the oil composition being linked to the hop aroma, the influence of pressure and temperature on the extraction kinetics of seven oil components (ß-myrcene, α-humulene, ß-caryophyllene, 2-methylbutyl isobutyrate, undecanone, linalool, and α-pinene) is analyzed and modeled in this article. Supercritical CO2 extraction from hop pellets was conducted at pressure-temperature combinations of 90/100/110 bar and 40/45/50 °C. The extract composition over time, analyzed by gas chromatography, was used for the parameterization of two existing mechanistic models: an internal-mass-transfer-control (IMTC), and a broken-and-intact-cells (BIC) model. The IMTC model was found to effectively describe most extraction kinetics and hence applied in this study. In contrast to previous studies, the IMTC model parameters were not only fitted to individual extraction curves from different experiments but also correlated to temperature and pressure as a further step towards model-based prediction. Using the parameterized model, the extract composition was predicted at 95 bar/48 °C, 105 bar/42 °C, and 105 bar/48 °C. Extraction yields were found to be higher at lower temperatures and higher pressures in general. The sensitivity towards pressure was observed to differ between components and to be particularly higher for ß-myrcene compared with α-humulene. Changes of the essential oil composition with a variation in pressure and temperature were predicted correctly by the model with a mean relative deviation from experimental data of 11.7% (min. 1.2%, max. 36.2%).

Effects and interactions of metal oxides in microparticle-enhanced cultivation of filamentous microorganisms.

Filamentous microorganisms are used as molecular factories in industrial biotechnology. In 2007, a new approach to improve productivity in submerged cultivation was introduced: microparticle-enhanced cultivation (MPEC). Since then, numerous studies have investigated the influence of microparticles on the cultivation. Most studies considered MPEC a morphology engineering approach, in which altered morphology results in increased productivity. But sometimes similar morphological changes lead to decreased productivity, suggesting that this hypothesis is not a sufficient explanation for the effects of microparticles. Effects of surface chemistry on particles were paid little attention, as particles were often considered chemically-inert and bioinert. However, metal oxide particles strongly interact with their environment. This review links morphological, physical, and chemical properties of microparticles with effects on culture broth, filamentous morphology, and molecular biology. More precisely, surface chemistry effects of metal oxide particles lead to ion leaching, adsorption of enzymes, and generation of reactive oxygen species. Therefore, microparticles interfere with gene regulation, metabolism, and activity of enzymes. To enhance the understanding of microparticle-based morphology engineering, further interactions between particles and cells are elaborated. The presented description of phenomena occurring in MPEC eases the targeted choice of microparticles, and thus, contributes to improving the productivity of microbial cultivation technology.

Inline imaging reveals evolution of the size distribution and the concentration of microbubbles in dissolved air flotation.

Dissolved air flotation (DAF) is an efficient process to remove impurities from fresh or salt water. As the removal is based on the agglomeration of impurities on the generated microbubbles, the size distribution and concentration of air bubbles are key parameters in dissolved air flotation. However, the development of microbubbles in the whole flotation process remains unexplored. In this study, we show that state-of-the-art inline microscopy enables the image acquisition of bubbles in DAF. Based on image analysis, thousands of microbubbles (10-200 µm) were analyzed within 6-12 min experiments. Consequently, bubble size distributions and bubble concentrations can be determined with moderate effort. Bubble size distributions were measured in a lab-scale DAF comprising a saturation unit, a decompression valve in/after which the bubbles are formed, and the actual flotation tank. The state of the microbubbles is not only determined at different positions within the tank but also in the supply pipe from the decompression valve to the tank. All bubble size distributions were unimodal and can be described well with Burr XII distributions. For fresh water, bubble size increased while bubble concentration decreased along the supply pipe between the decompression valve and the inlet of the flotation tank, indicating bubble coalescence. Compared to freshwater, saltwater inhibited this bubble coalescence in the pipe. Within the flotation tank, the bubble size did not change drastically for neither salt- nor freshwater. However, the bubble concentration decreased for both waters, which could be explained by dilution effects. Our results demonstrate that the developed inline method is a promising tool to study the evolution of microbubbles in flotation systems. Further, it might also be applied to investigate microbubbles in other processes such as fermentation, decomposition of organic compounds, and fouling mitigation in membranes.

Investigation of the Kinetics of Hysteresis Effects in Silica Gel.

Mathematical models can provide estimates for the shelf life of water-sensitive products like food or pharmaceuticals. This study presents a simple kinetic model using two first-order reactions for the evaporation and condensation of water. Furthermore, the model can be simplified to contain only one free parameter, the reaction rate constant k, which has been validated for silica gel at a relative humidity between 0% and 80% with experimental data. The experimental data shows the hysteresis effect of the silica gel in the region between 30% and 80% relative humidity and its dependence on the relative humidity earlier in the process. It also shows there are multiple equilibrium water contents at a relative humidity of 40%, depending on the previous relative humidity. The relative humidity barely influences the fitted reaction rate constant during adsorption. However, during the desorption process, not only the current relative humidity but also the history of relative humidity have an influence. A higher relative humidity in the previous step can slow down the desorption rate in the following step.

Key odorant melanoidin interactions in aroma staling of coffee beverages.

Fractionation of high molecular weight (HMW) coffee melanoidins of varying roasting intensity and evaluation of aroma binding activity via nuclear magnetic resonance spectroscopy revealed two fractions 5-10 kDa and > 50 kDa with high and selective binding affinity toward key coffee odorants. Quantification of monosaccharides, amino acids and phenolic acids, as well as aliphatic organic acids in hydrolysates of HMW material indicated the importance of aromatic residues in the form of hydroxycinnamic acids for the formation of non-covalent interactions with odorants. Caffeic acid concentrations were up to four times higher in fractions 5-10 kDa and > 50 kDa compared to 30-50 kDa and 10-30 kDa fractions. A minimization strategy was developed involving alkaline treatment of the most affine HMW fractions followed by reconstitution at typical coffee concentrations. These reconstituted HMW fractions exhibited up to 25% less aroma binding compared to native fractions of coffee.

From spores to fungal pellets: A new high-throughput image analysis highlights the structural development of Aspergillus niger.

Many filamentous fungi are exploited as cell factories in biotechnology. Cultivated under industrially relevant submerged conditions, filamentous fungi can adopt different macromorphologies ranging from dispersed mycelia over loose clumps to pellets. Central to the development of a pellet morphology is the agglomeration of spores after inoculation followed by spore germination and outgrowth into a pellet population, which is usually very heterogeneous. As the dynamics underlying population heterogeneity is not yet fully understood, we present here a new high-throughput image analysis pipeline based on stereomicroscopy to comprehensively assess the developmental program starting from germination up to pellet formation. To demonstrate the potential of this pipeline, we used data from 44 sampling times harvested during a 48 h submerged batch cultivation of the fungal cell factory Aspergillus niger. The analysis of up to 1700 spore agglomerates and 1500 pellets per sampling time allowed the precise tracking of the morphological development of the overall culture. The data gained were used to calculate size distributions and area fractions of spores, spore agglomerates, spore agglomerates within pellets, pellets, and dispersed mycelia. This approach eventually enables the quantification of culture heterogeneities and pellet breakage.

Interferometric Probing of Physical and Chemical Properties of Solutions: Noncontact Investigation of Liquids.

Interferometry is a highly versatile tool for probing physical and chemical phenomena. In addition to the benefit of noncontact investigations, even spatially resolved information can be obtained by choosing a suitable setup. This review presents the evolution of the various setups that have evolved since the first interferometers were developed in the mid-nineteenth century and highlights the benefits, limitations, and typical areas of application. This review focuses on interferometry based on electromagnetic waves in the near-infrared and visible range applied to liquid samples, categorizes the chemical/physical properties (e.g., pressure, temperature, composition) and phenomena (e.g., evaporation, crystal growth, diffusion, thermophoresis) that can be assessed, and presents a comprehensive literature review of specific existing applications. Finally, it discusses some fundamental open questions with respect to geometric considerations and overlapping effects.

Filamentous fungal applications in biotechnology: a combined bibliometric and patentometric assessment.

BACKGROUND: Processes and products employing filamentous fungi are increasing contributors to biotechnology. These organisms are used as cell factories for the synthesis of platform chemicals, enzymes, acids, foodstuffs and therapeutics. More recent applications include processing biomass into construction or textile materials. These exciting advances raise several interrelated questions regarding the contributions of filamentous fungi to biotechnology. For example, are advances in this discipline a major contributor compared to other organisms, e.g. plants or bacteria? From a geographical perspective, where is this work conducted? Which species are predominantly used? How do biotech companies actually use these organisms? RESULTS: To glean a snapshot of the state of the discipline, literature (bibliometry) and patent (patentometry) outputs of filamentous fungal applications and the related fields were quantitatively surveyed. How these outputs vary across fungal species, industrial application(s), geographical locations and biotechnological companies were analysed. Results identified (i) fungi as crucial drivers for publications and patents in biotechnology, (ii) enzyme and organic acid production as the main applications, (iii) Aspergillus as the most commonly used genus by biotechnologists, (iv) China, the United States, Brazil, and Europe as the leaders in filamentous fungal science, and (v) the key players in industrial biotechnology. CONCLUSIONS: This study generated a summary of the status of filamentous fungal applications in biotechnology. Both bibliometric and patentometric data have identified several key trends, breakthroughs and challenges faced by the fungal research community. The analysis suggests that the future is bright for filamentous fungal research worldwide.

Influence of Monovalent Salts on α-Glycine Crystal Growth from Aqueous Solution: Molecular Dynamics Simulations at Constant Supersaturation Conditions.

The growth of α-glycine crystals from aqueous solution is investigated at constant supersaturations by utilizing the constant chemical potential molecular dynamics method. The study considers two faces (010) and (011) that predominantly determine the α-glycine crystal morphology. The general Amber force field (GAFF) with two different charge sets derived from semi-empirical calculations using the complete neglect of differential overlap method (CNDO) and from density functional calculations using the double-numerical plus d- and p-polarization basis set (DNP) is applied to describe α-glycine. The extended simple point charge model is used to simulate water. It is observed that the GAFF/DNP set leads to a much slower integration of glycine molecules into the crystal structure than the GAFF/CNDO set. The GAFF/CNDO set, however, causes the growth even at concentrations well below the experimental solubility. For the GAFF/DNP set, the influence of potassium chloride (KCl) and sodium chloride (NaCl) on the face growth rates is investigated. The parameters recently proposed by Yagasaki et al. [J. Chem. Theory Comput. 2020, 16, 2460-2473] are used to describe salt ions, as standard GAFF parameters lead to the unexpected formation of salt clusters at a concentration lower than the experimental solubility value. According to our simulation results, both salts suppress the growth of the (011) and (010) faces. The inhibiting effect of NaCl is much stronger than that of KCl for the (011) face, while both salts have a similar inhibiting effect on the (010) face. The results are in line with the experimental observations of the impact of salt ions on the α-glycine growth rates for the (011) face reported in literature.

Understanding and controlling filamentous growth of fungal cell factories: novel tools and opportunities for targeted morphology engineering.

Filamentous fungal cell factories are efficient producers of platform chemicals, proteins, enzymes and natural products. Stirred-tank bioreactors up to a scale of several hundred m³ are commonly used for their cultivation. Fungal hyphae self-assemble into various cellular macromorphologies ranging from dispersed mycelia, loose clumps, to compact pellets. Development of these macromorphologies is so far unpredictable but strongly impacts productivities of fungal bioprocesses. Depending on the strain and the desired product, the morphological forms vary, but no strain- or product-related correlations currently exist to improve process understanding of fungal production systems. However, novel genomic, genetic, metabolic, imaging and modelling tools have recently been established that will provide fundamental new insights into filamentous fungal growth and how it is balanced with product formation. In this primer, these tools will be highlighted and their revolutionary impact on rational morphology engineering and bioprocess control will be discussed.

Universal law for diffusive mass transport through mycelial networks.

Filamentous fungal cell factories play a pivotal role in biotechnology and circular economy. Hyphal growth and macroscopic morphology are critical for product titers; however, these are difficult to control and predict. Usually pellets, which are dense networks of branched hyphae, are formed during industrial cultivations. They are nutrient- and oxygen-depleted in their core due to limited diffusive mass transport, which compromises productivity of bioprocesses. Here, we demonstrate that a generalized law for diffusive mass transport exists for filamentous fungal pellets. Diffusion computations were conducted based on three-dimensional X-ray microtomography measurements of 66 pellets originating from four industrially exploited filamentous fungi and based on 3125 Monte Carlo simulated pellets. Our data show that the diffusion hindrance factor follows a scaling law with respect to the solid hyphal fraction. This law can be harnessed to predict diffusion of nutrients, oxygen, and secreted metabolites in any filamentous pellets and will thus advance the rational design of pellet morphologies on genetic and process levels.