Sascha Schäuble

Filamentous soil bacteria of the genus Streptomyces are a major source of bioactive natural products, including antibiotics. However, their industrial cultivation is complicated by complex morphologies ranging from dispersed mycelia to dense pellets, which are closely linked to productivity. Microparticles, such as talc or aluminium oxide, are commonly added to modulate macromorphology, yet their effects across diverse Streptomyces species remain insufficiently characterized. Here, we systematically evaluated the impact of cellulose particles on growth, morphology, and metabolite production in 13 Streptomyces species ( Streptomyces spp.). Cellulose addition significantly reduced pellet size and altered metabolite profiles in a species‐dependent manner, resulting predominantly in a decrease of natural product abundance. Without cellulose addition, metabolite profiles correlated with biosynthetic potential encoded in the genomes, whereas this relationship did not hold in the presence of cellulose. With the addition of cellulose, a decrease in peptide signals present across species was observed. Cellulose‐associated mechanical stress led to species‐specific metabolomic shifts characterized by increased amino acids, lipids, and other degradation products, consistent with enhanced mechanical cellular disruption, which might have decreased proteolytic turnover during the stationary phase in some species that were more prone to mechanical stress. Although pellet size reduction is often associated with increased abundance of natural products, these findings demonstrate that the influence of solid particles on Streptomyces natural products and metabolism is species‐dependent. This work provides a foundation for future studies using dynamic systems, such as co‐cultivation with cellulolytic organisms, to improve natural product discovery.

The commensal and pathogenic lifestyles of the opportunistic fungal pathogen Candida albicans require complex signaling networks regulated by protein kinases. To investigate the role of C. albicans protein kinases at the intestinal epithelial interface, we screened a comprehensive protein kinase deletion library for the capacity of the mutants to damage intestinal epithelial cells (IEC). Mutants showing altered IEC cytotoxicity relative to the wild type were further analyzed for their growth and morphology, focusing on hyper-damaging strains to identify kinases that rather prevent host cell damage. Deletion of CRK1 caused increased IEC-specific damage, despite slower growth, reduced hyphal length, and reduced adhesion as compared to wild-type cells. While tissue invasion levels and the formation of transcellular tunnels of the crk1 Δ/Δ mutant were increased, the translocation capacity through the IEC barrier was reduced. Transcriptional and metabolic profiling suggested a role for Crk1 in metabolic adaptation to carbon and nitrogen sources, which was validated by showing that high glucose and amino acids are required for crk1 Δ/Δ to cause increased IEC damage. Deletion of CRK1 rendered C. albicans more susceptible to cell wall and membrane stressors, but caused higher resistance to a catalase-specific and histidine biosynthesis inhibitor. This phenotypic pattern of medium- and epithelial cell type-specific cytotoxicity displayed by a C. albicans protein kinase mutant suggests that Crk1 regulates processes linked to carbon and amino acid metabolism that are relevant to interactions with intestinal epithelial cells. IMPORTANCE: Microbial signal transduction pathways regulate adaptation to changing environmental conditions and facilitate the success of many microbes during interactions with their hosts. The fungal pathobiont Candida albicans exists as a harmless commensal on mucosal surfaces of most humans but can also cause superficial and invasive infections under certain circumstances. Both lifestyles require complex signaling networks, predominantly regulated by protein kinases. The C. albicans genome was predicted to encode 108 protein kinases, yet nearly 50% remain uncharacterized. We aimed to dissect the role of C. albicans protein kinases during the transition from commensal to pathogen. We showed that multiple protein kinase genes are involved in epithelial cell damage. Particularly, the protein kinase gene Crk1 was of interest because deletion of CRK1 caused increased damage to intestinal epithelial cells under distinct conditions. Our study links Crk1 with regulation of metabolic processes relevant for commensalism and pathogenicity of C. albicans .

Fungi harbor unique primary and secondary metabolic pathways that represent a hidden treasure of biochemical and natural compounds. Their metabolism is central to their ability to interact with, to adapt to and to survive in host environments, and to cause human disease and great harm. Among these is Lichtheimia corymbifera, an emerging causative agent of mucormycosis that has been classified as high priority fungal pathogen by the World Health Organization and is gaining increasing importance as a model organism for research on invasive fungal infections. We reconstructed a genome-scale L. corymbifera metabolic model and show substantial differential metabolic activity to process carbohydrate or amino acid carbon sources. We furthermore adapt the model to proteome changes depending on the presence of human peripheral blood mononuclear cells (PBMCs) and show fungal sphingolipid metabolic activity changes next to changes in cytoskeleton and tight junction associated PBMC activity over three days of cultivation. These insights underline the possibility that L. corymbifera can potentially scavenge host-derived lipids to fortify its own cell membrane. We demonstrate that in silico metabolic predictions can provide testable hypotheses and can lead to the identification of metabolic processes which are essential for the development of targeted antifungal drugs and novel solutions for balancing host challenges.

SUMMARY: One of the enduring paradoxes of sepsis is that organs fail despite little evidence of irreversible tissue injury. Emerging evidence suggests that this state reflects a regulated metabolic shutdown within host tissues, yet whether such hypometabolism contributes to pathology or promotes survival remains unclear. This phenomenon resembles torpor, a physiological state of profound hypometabolism induced by environmental stress and mediated through reversible protein phosphorylation. Septic hypometabolism is identified here as a conserved tissue-specific metabolic adaptation which is characterized by transient activation of Glycogen Synthase Kinase (GSK)3β. This activation reduced disease severity of bacterial sepsis without affecting the hosts pathogen burden, indicating that GSK3β activity promotes disease tolerance to infection. Consistent with these findings, plasma signatures associated with GSK3β inhibition correlated with worse clinical outcomes in patients with sepsis. Together, these results define septic hypometabolism as a torpor-like response and identify reversible phosphorylation as a key mechanism governing host adaptation to severe bacterial infection.

Background Whether the lung microbiome represents a stable microbial colonization or a transient ecosystem shaped by continuous microbial turnover and controlled by host immunity remains unresolved. The murine lung microbiome largely consists of species from the former Lactobacillus genus with Ligilactobacillus murinus as a dominant species, bacterial genera such as Streptococcus , Staphylococcus , Mammaliicoccus , Enterococcus and other less frequently detected bacteria. Here, we directly addressed the question of persistence and host interaction of a dominant murine lung commensal in vivo and focused on the host immune response towards lung commensal bacteria. Results We developed a transformation strategy for stable genomic integration of a green fluorescent protein (GFP)-encoding gene to track the fate of a lung bacterium. Following intranasal administration of GFP-labeled L. murinus in mice, bacteria were readily detected in the lungs at early time points but declined rapidly and became undetectable after 72 hours, as determined by quantification of viable bacteria and qPCR. Flow cytometry and fluorescence imaging revealed efficient uptake of GFP-labeled bacteria by lung phagocytes. These findings indicate that even dominant members of the murine pulmonary microbiota normally detected at low abundances are transiently present in the lungs without causing infection. We further analyzed the effects of moderate and high bacterial concentrations. While moderate bacterial loads were efficiently controlled without clinical effects, high concentrations induced severe lethargy, indicating a threshold-dependent host response. Finally, we demonstrated that pulmonary commensals such as L. murinus , Staphylococcus xylosus , and Mammaliicoccus sciuri, as well as conidia of the opportunistic lung pathogen Aspergillus fumigatus , are phagocytosed at comparable rates in macrophage assays. Conclusions Our data demonstrate that even lung-adapted bacterial species fail to establish stable colonization and are instead subject to rapid immune-mediated elimination contributing to the maintenance of a low microbial burden in the lungs. While this homeostatic balance supports health, elevated bacterial loads trigger immune activation and, at high levels, lead to health deterioration. Together, these results support a model of a highly dynamic and transient lung microbiome, maintained by continual microbial immigration rather than long-term colonization. Accounting for the lung microbiome dynamics is essential for understanding host-microbiota interactions and respiratory health.

The saprophytic mold Aspergillus fumigatus produces small (2-3 µm) airborne spores (conidia) that can reach the lung alveoli upon inhalation. There, they encounter the surfactant-rich environment of the alveolar epithelium and initiate swelling and germination. Primary alveolar macrophages are essential for the rapid clearance of conidia and maintenance of pulmonary homeostasis. However, A. fumigatus remains the leading cause of invasive pulmonary aspergillosis in immunocompromised patients, and the mechanisms governing fungal clearance versus invasion remain poorly understood. In this study, we adapted a previously established monocyte-derived alveolar-like macrophage (ALM) model to investigate early host-pathogen interactions upon A. fumigatus challenge. Given the requirement of GM-CSF for maintaining alveolar macrophage identity and function, we included GM-CSF differentiated macrophages (GM-M), as a widely used reference model. Primary alveolar macrophages (pAM), isolated from human lung biopsies were used to validate the physiological relevance of the ALM model. Combined phenotypic, functional and transcriptomic analyses demonstrated that ALMs closely resemble pAMs under both steady-state and infection conditions across multiple time points and fungal burdens. Notably, fungal dual RNA-sequencing revealed a significant upregulation of fungal virulence-associated factors during interaction with ALM, which was not observed in GM-M co-cultures. Collectively, these findings support the use of ALMs as a robust, experimentally accessible and physiologically relevant in vitro model for investigating early A. fumigatus infection, providing new insights into host-pathogen dynamics at the alveolar interface.

Natural killer (NK) cells contribute to the innate immune system and are pivotal for the defence against opportunistic pathogens, including fungi. Aspergillus fumigatus (AF), a filamentous mold, can cause invasive pulmonary aspergillosis in immunocompromised patients, e.g. in patients after allogeneic stem cell transplantation (alloSCT). In this pilot study, we challenged NK cell samples from alloSCT recipients collected 90, 120, and 180 days after transplantation and from healthy individuals with AF and characterize the proteome response differences. We identified 2259 differentially abundant proteins between the NK cell proteomes of alloSCT recipients and healthy individuals. Among these, 1118 proteins were differentially abundant at all time points and 1931 proteins specifically at day 180 post-alloSCT. Following stimulation of NK cells with AF, we found a profoundly different early proteome (day 90, n=1652 proteins), while at day 180, only 77 proteins remained significantly differentially abundant. We identified, among others, a major differentially abundant protein cluster related to IL27RA (including OAS, STAT1, and MX). Furthermore, for selected markers [granzyme A (GZMA), Neural Cell Adhesion Molecule 1 (NCAM1/CD56), perforin-1 (PRF1)], we confirmed our proteome data by flow cytometry in NK cells from an independent second patient and healthy individual cohort. In conclusion, we demonstrate the advantage of combining comprehensive proteomic profiling with targeted flow cytometry to investigate NK cell responses to AF. Our data analysis connects STAT1 with IL27RA as well as granzyme, IFNg, and NCAM1 activity, which may be exploited towards future therapeutics warranting confirmation in larger study cohorts.

Fungi, and particularly fungal pathogens, are having an increasing impact on human health and economy. At the same time, modern high-throughput technologies offer insights deep into the molecular level and thus mechanisms, giving scientists new opportunities to identify fungal biomarkers and essential components for survival and virulence. This wealth of data, however, is most often only analyzed in the context of a specific scientific question, while many more projects may benefit from a multifaceted view on e.g. fungal gene expression under various conditions. The prime challenge is the limited access to readily pre-processed data and circumventing technological biases introduced by different sequencing platforms and software tools across different projects. We here present FungiNetDB, a web platform comprising 139 fungal pathogenicity datasets and statistical analysis of more than 2000 different pairwise gene expression comparisons. FungiNetDB thus resembles a most comprehensive fungal transcriptomics resource, which can be explored without any programming knowledge, allows highly customizable filtering and cross-project comparisons and download of all offered data tables and visualizations.

Candida albicans is a leading cause of invasive candidiasis, a life-threatening infection with high mortality despite antifungal therapy. Echinocandins are first-line agents due to their fungicidal activity and low toxicity profile. Beyond direct killing, they may also modulate host immunity, e.g. through fungal cell wall remodeling. However, the systemic impact of echinocandins on human immune cells remains incompletely understood, particularly in the context of drug resistance. Using an ex vivo human whole-blood infection model, we assessed the effects of caspofungin during infection with echinocandin-susceptible (ECHS/FKSwt) and echinocandin-resistant (ECHR/FKSmut) C. albicans strains. Infection alone triggered a strong and conserved transcriptional immune program enriched for innate recognition and inflammatory pathways. Caspofungin treatment during infection with the susceptible strain selectively amplified these responses, engaging a sequential cascade of host defenses that linked pathogen recognition with innate effector functions and adaptive polarisation. In contrast, caspofungin induced only minimal transcriptional changes in resistant strain infections or in uninfected blood. These findings demonstrate that caspofungin efficacy arises not only from its fungicidal activity but also from amplification of host immunity, an effect primarily dependent on fungal susceptibility. This dual mode of action highlights a previously underappreciated role of echinocandins in shaping antifungal immunity and provides new mechanistic insight into drug-host–pathogen interactions during systemic candidiasis.

The fungus Candida albicans is a frequent colonizer of humans but also an opportunistic pathogen causing superficial to severe infections, especially in vulnerable individuals. Its broad metabolic flexibility is key for the fungal adaptation to host environments, evasion from immune attack, and virulence. Amino acid metabolism and homeostasis, in particular, are critical for fungal fitness—illustrated by a rapid metabolic shift in response to amino acid starvation to restore intracellular metabolic balance. To investigate the cellular mechanisms underlying such compensatory metabolic processes, we performed data-driven genome-scale metabolic modeling based on transcriptional metabolic profiles of amino acid-starved cells to identify condition-specific fungal metabolic fluxes and pathway activities specific to cellular response to amino acid starvation. Most prominently, we predicted altered activity of the shikimate pathway upon amino acid limitation and identified a simultaneous induction of aromatic amino acid (AAA) biosynthesis and a metabolic gene cluster required for the catabolism of hydroxybenzenes. Further phenotypic and transcriptional analyses not only verified the transcription factor Zcf25 as the central regulator of the catechol-branch of this pathway, but also condition-specific co-regulation of AAA and hydroxybenzene metabolism mediated by Zcf25 and the transcriptional regulator of amino acid metabolism Stp2. These findings propose a so far unknown metabolic link between amino acid and hydroxybenzene metabolism in C. albicans , therewith adding another layer to its metabolic plasticity. IMPORTANCE: The opportunistic human fungal pathogen Candida albicans possesses a remarkable metabolic plasticity, which is essential for both fungal commensalism and virulence and influences its physiology and behavior in multiple ways. The investigation of such processes particularly benefits from the emergence of multi-omics and in silico approaches. In this study, we combined a multi-omics approach with genome-scale metabolic modeling to investigate the fungal metabolic adaptation to amino acid utilization and starvation. Most strikingly, we found an altered activity of the shikimate pathway upon amino acid starvation, accompanied by a simultaneous induction of two metabolic gene clusters required for the metabolism of hydroxybenzenes. Further analyses revealed so far unknown potential functional and regulatory links between both metabolic pathways, which provide starting points for future research leading to a better understanding of the fungal adaptation to dynamic host conditions.