Biology subjects

van Cappellen, W.

Publications and source records attributed to van Cappellen, W..

3 recordsLinked to original sources

bioRxiv2024

Early Fibrin Biofilm Development in Cardiovascular Infections

The single most common microbe causing cardiovascular infections is Staphylococcus aureus (S. aureus). S. aureus produces coagulase that converts fibrinogen to fibrin, which is incorporated into biofilms. This process aids in adherence to intravascular structures, defense against the host immune system, and resistance to antimicrobial treatment. Despite its significance, fibrin formation in S. aureus biofilms remains poorly understood. Therefore, this study aimed to elucidate the early development of cardiovascular biofilms. Clinically isolated coagulase-positive S. aureus and coagulase-negative Streptococcus gordonii (S. gordonii) from patients with cardiovascular infections, and a coagulase mutant S. aureus {Delta}coa, were grown in tryptic soy broth (TSB), Iscoves Modified Dulbeccos Medium (IMDM), and pooled human plasma, with or without porcine heart valves. Bacterial growth, metabolic activity, and bacterial fibrinogen utilization were measured over 24 hr at 37 {degrees}C. Time-lapse confocal microscopy was used to visualize and track biofilm development. S. aureus exhibited more growth in TSB and human plasma than S. gordonii and S. aureus {Delta}coa, but showed similar growth as S. aureus {Delta}coa in IMDM. Peak metabolic activity for all isolates was highest in TSB and lowest in human plasma. The presence of porcine valves caused strain-dependent alterations in time to peak metabolic activity. Confocal imaging revealed fibrin-based biofilm development exclusively in the coagulase-producing S. aureus strains. Between 2 and 6 hr of biofilm development, 74.9% (p=0.034) of the fibrinogen from the medium was converted to fibrin. Variations in fibrin network porosity and density were observed among different coagulase-producing S. aureus strains. Fibrin formation is mediated by S. aureus coagulase and first strands occurred within 3 hr for clinical strains after exposure to human plasma. This study stresses the importance of experimental design given the bacterial changes due to different media and substrates and provides insights into the early pathogenesis of S. aureus cardiovascular biofilms. HighlightsO_LIBacterial growth and activity are medium and substrate dependent C_LIO_LICoagulase is necessary for Staphylococcus aureus fibrin biofilm development C_LIO_LIFibrin strands begin forming in Staphylococcus aureus biofilms within 3 hours C_LI

microbiology↗

bioRxiv2021

Structured illumination microscopy with noise-controlled image reconstructions

Super-resolution structured illumination microscopy (SIM) has become a widely used method for biological imaging. Standard reconstruction algorithms, however, are prone to generate noise-specific artefacts that limit their applicability for lower signal-to-noise data. Here, we present a physically realistic noise model that explains the structured noise artefact and that is used to motivate new complementary reconstruction approaches. True Wiener-filtered SIM optimizes contrast given the available signal-to-noise ratio, flat-noise SIM fully overcomes the structured noise artefact while maintaining resolving power. Both methods eliminate ad-hoc user adjustable reconstruction parameters in favour of physical parameters, enhancing objectivity. The new reconstructions point to a trade-off between contrast and a natural noise appearance. This trade-off can be partly overcome by additional notch filtering, but at the expense of a decrease in signal-to-noise ratio. The benefits of the proposed approaches are demonstrated on focal adhesion and tubulin samples in 2D and 3D, and on nano-fabricated fluorescent test patterns.

biophysics↗

bioRxiv2020

Super-resolution imaging of RAD51 and DMC1 in DNA repair foci reveals dynamic distribution patterns in meiotic prophase

The recombinase RAD51, and its meiosis-specific paralog DMC1 localize at DNA double-strand break (DSB) repair sites in meiotic prophase nuclei. While both proteins are required during meiotic homologous recombination, their spatial organization during meiotic DSB repair is not fully understood. Using super-resolution microscopy on mouse spermatocyte nuclei, we aimed to define their relative position at DSB foci, and how these vary in time. We show that a large fraction of meiotic DSB repair foci (38%) contained a single RAD51 cluster and a single DMC1 cluster (D1R1 configuration) that were partially overlapping (average center-center distance around 70 nm). The majority of the rest of the foci had a similar combination of a major RAD51 and DMC1 cluster, but in combination with additional clusters (D2R1, D1R2, D2R2, or DxRy configuration) at an average distance of around 250 nm. As prophase progressed, less D1R1 and more D2R1 foci were observed, where the RAD51 cluster in the D2R1 foci elongated and gradually oriented towards the distant DMC1 cluster. This correlated with more frequently observed RAD51 bridges between the two DMC1 clusters. D1R2 foci frequency was more constant, and the single DMC1 cluster did not elongate, but was observed more frequently in between the two RAD51 clusters in early stages. D2R2 foci were rare (<10%) and nearest neighbour analyses also did not reveal pair formation between D1R1 foci. In the absence of the transverse filament of the synaptonemal complex (connecting the chromosomal axes of homologs), early configurations were more prominent, and RAD51 elongation occurred only transiently. This in-depth analysis of single cell landscapes of RAD51 and DMC1 accumulation patterns at DSB repair sites at super-resolution thus revealed the variability of foci composition, and defined functional consensus configurations that change over time. AUTHOR SUMMARYMeiosis is a specific type of cell division that is central to sperm and egg formation in sexual reproduction. It forms cells with a single copy of each chromosome, instead of the two copies that are normally present. In meiotic prophase, homologous chromosomes must connect to each other, to be correctly distributed between the daughter cells. This involves the formation and repair of double-strand breaks in the DNA. Here we used super-resolution microscopy to elucidate the localization patterns of two important DNA repair proteins: RAD51 and DMC1. We found that repair sites most often contain a single large cluster of both proteins, with or without one additional smaller cluster of either protein. RAD51 protein clusters displayed lengthening as meiotic prophase progressed. When chromosome pairing was disturbed, we observed changes in the dynamics of protein accumulation patterns, indicating that they actually correspond to certain repair intermediates changing in relative frequency of occurrence. These analyses of single meiotic DNA repair foci reveal the biological variability in protein accumulation patterns, and the localization of RAD51 and DMC1 relative to each other, thereby contributing to our understanding of the molecular basis of meiotic homologous recombination.

developmental biology↗