Natalia Schiefermeier

Introduction: The lung environment is defined by unique biological boundary conditions, including complex alveolar geometry, extracellular matrix composition and mechanical forces generated during respiration. These factors were shown to regulate alveolar permeability, surfactant secretion, cell contractility and apoptosis, but their role in fungal infections remains unknown. Aspergillus fumigatus is a critical fungal pathogen that causes severe pulmonary infections in immunocompromised individuals. Our study addresses a knowledge gap by investigating how boundary conditions affect A. fumigatus conidia interactions with alveolar epithelial cells.Methods: We applied micropatterned substrates to confine cells into defined shapes and densities, allowing precise control over geometric conditions and extracellular matrix composition. Using cell line stably expressing the phagolysosomal protein Lamp1-NeonGreen and multiplane fluorescent microscopy, we evaluated A. fumigatus conidia binding and internalization efficiency.Results: We observed significantly faster and more efficient A. fumigatus conidia internalization in cells confined on micropatterns compared to previously reported studies using cell monolayers. Altering cell geometry, density and extracellular matrix composition strongly affected conidia binding and localization to Lamp1⁺ phagolysosomes. Cells on X-shaped or multicellular micropatterns showed higher internalization rates, particularly at the periphery, suggesting spatial heterogeneity in pathogen uptake. Additionally, changes in extracellular matrix composition influenced the intracellular trafficking of A. fumigatus conidia.Discussion: Our findings emphasize the essential role that local mechanical and biochemical cues play in shaping the interactions between fungal pathogens and alveolar cells. Understanding how lung boundary conditions change in disease states will provide important insights into fungal infection outcomes.

Introduction: The lung environment is defined by unique biological boundary conditions, including complex alveolar geometry, extracellular matrix composition and mechanical forces generated during respiration. These factors were shown to regulate alveolar permeability, surfactant secretion, cell contractility and apoptosis, but their role in fungal infections remains unknown. Aspergillus fumigatus is a critical fungal pathogen that causes severe pulmonary infections in immunocompromised individuals. Our study addresses a knowledge gap by investigating how boundary conditions affect A. fumigatus conidia interactions with alveolar epithelial cells. Methods: We applied micropatterned substrates to confine cells into defined shapes and densities, allowing precise control over geometric conditions and extracellular matrix composition. Using cell line stably expressing the phagolysosomal protein Lamp1-NeonGreen and multiplane fluorescent microscopy, we evaluated A. fumigatus conidia binding and internalization efficiency. Results: We observed significantly faster and more efficient A. fumigatus conidia internalization in cells confined on micropatterns compared to previously reported studies using cell monolayers. Altering cell geometry, density, and extracellular matrix composition strongly affected conidia binding and localization to Lamp1 ⁺ phagolysosomes. Cells on X-shaped or multicellular micropatterns showed higher internalization rates, particularly at the periphery, suggesting spatial heterogeneity in pathogen uptake. Additionally, changes in extracellular matrix composition influenced the intracellular trafficking of A. fumigatus conidia. Discussion: Our findings emphasize the essential role that local mechanical and biochemical cues play in shaping the interactions between fungal pathogens and alveolar cells. Understanding how lung boundary conditions change in disease states will provide important insights into fungal infection outcomes.

Aspergillus fumigatus is an opportunistic fungal pathogen that causes serious lung diseases in immunocompromised patients. The lung surfactant produced by alveolar type II and Clara cells in the lungs is an important line of defense against A. fumigatus. The surfactant consists of phospholipids and surfactant proteins (SP-A, SP-B, SP-C and SP-D). The binding to SP-A and SP-D proteins leads to the agglutination and neutralization of lung pathogens as well as the modulation of immune responses. SP-B and SP-C proteins are essential for surfactant metabolism and can modulate the local immune response; however, the molecular mechanisms remain unclear. We investigated changes in the SP gene expression in human lung NCI-H441 cells infected with conidia or treated with culture filtrates obtained from A. fumigatus. To further identify fungal cell wall components that may affect the expression of SP genes, we examined the effect of different A. fumigatus mutant strains, including dihydroxynaphthalene (DHN)-melanin-deficient ΔpksP, galactomannan (GM)-deficient Δugm1 and galactosaminogalactan (GAG)-deficient Δgt4bc strains. Our results show that the tested strains alter the mRNA expression of SP, with the most prominent and consistent downregulation of the lung-specific SP-C. Our findings also suggest that secondary metabolites rather than the membrane composition of conidia/hyphae inhibit SP-C mRNA expression in NCI-H441 cells.

Background: The implementation of standardized assessments in physiotherapeutic practice strongly supports diagnostic and treatment plans. Previous studies reported insufficient usage of standardized assessments due to lack of time, lack of knowledge, lack of resources and other barriers. Physiotherapy in outpatient settings became essential for the rehabilitation of patients with post COVID-19 condition but it remains unknown to what extent assessments are implemented into the evaluation of these patients. In this study, we explored the current use and barriers regarding the implementation of physiotherapeutic assessments to evaluate patients with post COVID-19 condition. Methods: A cross-sectional online survey was carried out among 180 physiotherapists working in outpatient settings in Austria and South Tyrol. Results: The majority of physiotherapists (88%) indicated that standardized assessments are useful, though less than a fifth of participants actually implement assessments in practice. Among implementation barriers, “insufficient experience” (41.8%) and “lack of knowledge” (36.6%) were mentioned most often. Concerning specific post COVID-19 assessments, the evaluation of “physical and respiratory function”, “quality of life” and “activities of daily living” were stated to be of particular relevance. Conclusions: Our study revealed a low implementation rate and identified the main barriers regarding the non-usage of standardized assessments for post COVID-19 patients. Trial registration: The Private University for Health Sciences and Health Technology (UMIT TIROL), and the Research Committee for Scientific Ethical Questions granted approval for the survey (RCSEQ, Hall in Tirol, Austria, Number 2834).

The rehabilitation needs of COVID-19 survivors are increasingly recognized, with a focus on combating respiratory and neuromuscular dysfunctions. The aim here was to explore the perception of Austrian physiotherapists and physiotherapy students on post-COVID-19 rehabilitation care and to identify barriers for the application of sufficient rehabilitation. We analysed current knowledge and practical skills in respiratory physiotherapy, performing a cross-sectional national survey among physiotherapists working in outpatient settings and physiotherapy students in their last academic year of bachelor-level education in Austria. Out of 255 survey participants, one-third already had inquiries to treat post-COVID-19 patients, and the majority of respondents expected a further increased inflow of patients with rehabilitation needs (64.2%). Only 11.2% of respondents reported feeling sufficiently informed about post-COVID-19 rehabilitation. A total of 68.2% of students and up to 48.1% of physiotherapists favoured a COVID-19-specific adaptation already in the basic academic education, and 74.1% of survey participants indicated interest in attending specific training. Concerning respiratory physiotherapy, our data showed discrepancies between the estimation of the importance of specific examination and treatment techniques and the level of current experience. There is a clear lack of experience in implementing effective device-based respiratory therapy. Our data indicate an urgent need to develop new education and training programs with a focus on the interdisciplinary rehabilitation of patients with post-COVID-19 syndrome.

Inhaled Aspergillus fumigatus spores can be internalized by alveolar type II cells. Cell lines stably expressing fluorescently labeled components of endocytic pathway enable investigations of intracellular organization during conidia internalization and measurement of the process kinetics. The goal of this report was to evaluate the methodological appliance of cell lines for studying fungal conidia internalization. We have generated A549 cell lines stably expressing fluorescently labeled actin (LifeAct-mRuby2) and late endosomal protein (LAMP1-NeonGreen) following an evaluation of cell-pathogen interactions in live and fixed cells. Our data show that the LAMP1-NeonGreen cell line can be used to visualize conidia co-localization with LAMP1 in live and fixed cells. However, caution is necessary when using LifeAct-mRuby2-cell lines as it may affect the conidia internalization dynamics.

Monocytes are important players to combat the ubiquitously present fungus Aspergillus fumigatus. Recruitment of monocytes to sites of fungal A. fumigatus infection has been shown in vivo. Upon exposure to A. fumigatus in vitro, purified murine and human blood monocytes secrete inflammatory cytokines and fungicidal mediators. Mononuclear tissue phagocytes are phenotypically and functionally different from those circulating in the blood and their role in antifungal defenses is much less understood. In this study, we identified a population of migrating CD43⁺ monocytes in cells isolated from rat distal lungs. These cells are phenotypically different from alveolar macrophages and show distinct locomotory behavior on the surface of primary alveolar cells resembling previously described endothelial patrolling monocytes. Upon challenge, the CD43⁺ monocytes internalized A. fumigatus conidia resulting in inhibition of their germination and hyphal growth. Thus, migrating lung monocytes might play an important role in local defense against pulmonary pathogens.

Monocytes are important players to combat ubiquitously present fungus Aspergillus fumigatus . Recruitment of monocytes to sites of fungal infection was shown in vivo , and purified murine and human blood monocytes are able to induce inflammatory and fungicidal mediators as well as the host cell and the fungal transcriptional responses upon exposure to A.fumigatus . Mononuclear tissue phagocytes are phenotypically and functionally different from those circulating in the blood and their role in antifungal defences is much less understood. In this study, we identified a population of migrating CD43 + monocytes in cells isolated from rat distal lungs. These cells phenotypically different from alveolar macrophages, showed clearly distinct locomotory behaviour on the surface of primary alveolar cells resembling previously described endothelial patrolling. The CD43 + monocytes internalized live A.fumigatus conidia resulting in inhibition of conidial germination and hyphal growth. Thus, migrating lung monocytes might play an important role in local defence against pulmonary pathogens.

Dietary intake of sodium, potassium, calcium and magnesium has a strong impact on personal health. In order to understand possible correlations with regional occurrence of diseases and to develop new dietary strategies, it is necessary to evaluate food choices in defined geographic areas. The aim of this study was to analyze daily consumption and major food sources of electrolytes with an emphasis on dietary patterns. In this representative, cross-sectional study, daily foods of 463 adults were assessed with two nonconsecutive 24-h recalls. Our results show high sodium and low potassium consumption in all age groups in both men and women. Furthermore, more than half of investigated persons had low calcium, and 40% indicated low dietary magnesium intake. Only 1% of our study population reached the recommended values for all electrolytes, while 13% consumed adequate levels of three electrolytes. Moreover, 14% did not reach reference levels for any of the four minerals. A further comparison of dietary patterns and food preferences showed significant differences in major food groups including nonalcoholic drinks, fruits, vegetables, legumes, milk products, vegetable oil, bread and sweets. Our results are important for further evaluations of nutrition intake and the development of new dietary strategies.