Biology subjects

Hepp, C.

Publications and source records attributed to Hepp, C..

3 recordsLinked to original sources

bioRxiv2025

High-throughput Single-Virion DNA-PAINT Reveals Structural Diversity, Cooperativity and Flexibility during Selective Packaging in Influenza

Influenza A, a negative-sense RNA virus, has a genome that consists of eight single-stranded RNA segments. Influenza co-infections can result in re-assortant viruses that contain gene segments from multiple strains, causing pandemic outbreaks with severe consequences for human health. The outcome of re-assortment is likely influenced by a selective sequence-specific genome packaging mechanism. To uncover the contributions of individual segment pairings to selective packaging, we set out to statistically analyse packaging defects and inter-segment distances in individual A/Puerto Rico/8/34 (H1N1) (PR8) virus particles. To enable such analysis, we developed a multiplexed DNA-PAINT approach capable of assessing the segment stoichiometry of more than 10,000 individual virus particles in one experiment; our approach can also spatially resolve the individual segments inside complete virus particles with a localisation precision of ~10 nm. Our results show the influenza genome can be assembled through multiple pathways in a redundant and cooperative process guided by preferentially interacting segment pairs and aided by synergistic effects that enhance genome assembly, driving it to completion. Our structural evidence indicates that the interaction strength of segment pairs affects the spatial configuration of the gene segments, which appears to be preserved in mature virions. As our method quantified the interactions of whole influenza segments instead of identifying individual sequence-based interactions, our results can serve as a template to quantify the contributions of individual sequence motifs to selective packaging.

microbiology↗

bioRxiv2025

Tunable fluorogenic DNA probes drive fast and high-resolution single-molecule fluorescence imaging

A main limitation of single-molecule fluorescence (SMF) measurements is the "high concentration barrier", describing the maximum concentration of fluorescent species tolerable for sufficient signal-to-noise ratio (SNR). To address this barrier in several SMF applications, we design fluorogenic probes based on short ssDNAs, fluorescing only upon hybridising to their complementary target sequence. We engineer the quenching efficiency and fluorescence enhancement upon duplex formation through screening several fluorophore-quencher combinations, label lengths, and sequence motifs, which we utilise as tuning screws to adapt our labels to different experimental designs. Using these fluorogenic probes, we can perform SMF experiments at concentrations of 10 {micro}M fluorescent labels; this concentration is 100-fold higher than the operational limit for standard TIRF experiments. We demonstrate the ease of implementing these probes into existing protocols by performing super-resolution imaging with DNA-PAINT, employing a fluorogenic 6 nt-long imager; through the faster acquisition of binding events, the imaging of viral genome segments could be sped up significantly to achieve extraction of 20-nm structural features with only [~]150 s of imaging. The exceptional tunability of our probe design will overcome concentration barriers in SMF experiments and unlock new possibilities in super-resolution imaging, molecular tracking, and smFRET. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=89 SRC="FIGDIR/small/634148v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@1d51a20org.highwire.dtl.DTLVardef@1a64fb7org.highwire.dtl.DTLVardef@1bbb4bforg.highwire.dtl.DTLVardef@578c63_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2020

Megafauna decline have reduced pathogen dispersal which may have increased emergent infectious diseases

The Late Quaternary extinctions of megafauna (defined as animal species >44.5 kg) reduced the dispersal of seeds and nutrients, and likely also microbes and parasites. Here we use body-mass based scaling and range maps for extinct and extant mammal species to show that these extinctions led to an almost seven-fold reduction in the movement of gut-transported microbes, such as Escherichia coli (3.3 km2/day to 0.5 km2/day). Similarly, the extinctions led to a seven-fold reduction in the mean home ranges of vector-borne pathogens (7.8 km2 to 1.1 km2). To understand the impact of this, we created an individual-based model where an order of magnitude decrease in home range increased maximum aggregated microbial mutations 4-fold after 20,000 years. We hypothesize that pathogen speciation and hence endemism increased with isolation, as global dispersal distances decreased through a mechanism similar to the theory of island biogeography. To investigate if such an effect could be found, we analysed where 145 zoonotic diseases have emerged in human populations and found quantitative estimates of reduced dispersal of ectoparasites and fecal pathogens significantly improved our ability to predict the locations of outbreaks (increasing variance explained by 8%). There are limitations to this analysis which we discuss in detail, but if further studies support these results, they broadly suggest that reduced pathogen dispersal following megafauna extinctions may have increased the emergence of zoonotic pathogens moving into human populations.

ecology↗