The [PSI+] prion phenotype in yeast manifests as a white, pink, or red color pigment. Experimental manipulations destabilize prion phenotypes, and allow colonies to exhibit [psi-] (red) sectored phenotypes within otherwise completely white colonies. Further investigation of the size and frequency of sectors that emerge as a result of experimental manipulation is capable of providing critical information on mechanisms of prion curing, but we lack a way to reliably extract this information. Images of experimental colonies exhibiting sectored phenotypes offer an abundance of data to help uncover molecular mechanisms of sectoring, yet the structure of sectored colonies is ignored in traditional biological pipelines. In this study, we present [PSI]-CIC, the first computational pipeline designed to identify and characterize features of sectored yeast colonies. To overcome the barrier of a lack of manually annotated data of colonies, we develop a neural network architecture that we train on synthetic images of colonies and apply to real images of [PSI+] , [psi-] , and sectored colonies. In hand-annotated experimental images, our pipeline correctly predicts the state of approximately 95% of colonies detected and frequency of sectors in approximately 89.5% of colonies detected. The scope of our pipeline could be extended to categorizing colonies grown under different experimental conditions, allowing for more meaningful and detailed comparisons between experiments. Our approach streamlines the analysis of sectored yeast colonies providing a rich set of quantitative metrics and provides insight into mechanisms driving the curing of prion phenotypes.
Symbiotic marine bacteria that are transmitted through the environment are susceptible to abiotic factors (salinity, temperature, physical barriers) that can influence their ability to colonize their specific hosts. Given that many symbioses are driven by host specificity, environmentally transmitted symbionts are more susceptible to extrinsic factors depending on conditions over space and time. In order to determine whether the population structure of environmentally transmitted symbionts reflects host specificity or biogeography, we analysed the genetic structure of Sepiola atlantica (Cephalopoda: Sepiolidae) and their Vibrio symbionts (V. fischeri and V. logei) in four Galician Rías (Spain). This geographical location is characterized by a jagged coastline with a deep-sea entrance into the land, ideal for testing whether such population barriers exist due to genetic isolation. We used haplotype estimates combined with nested clade analysis to determine the genetic relatedness for both S. atlantica and Vibrio bacteria. Analyses of molecular variance (AMOVA) were used to estimate variation within and between populations for both host and symbiont genetic data. Our analyses reveal a low percentage of variation among and between host populations, suggesting that these populations are panmictic. In contrast, Vibrio symbiont populations show certain degree of genetic structure, demonstrating that the hydrology of the rías is driving bacterial distribution (and not host specificity). Thus, for environmentally transmitted symbioses such as the sepiolid squid-Vibrio association, abiotic factors can be a major selective force for determining population structure for one of the partners.
Shared clinical decision-making (SCDM) about HPV vaccination has been recommended for U.S. mid-adults aged 27-45 since 2019. To explore barriers and facilitators to HPV vaccination in this population, we conducted 14 virtual focus groups with 86 unvaccinated mid-adults (34 men and 52 women) in Californias medically underserved Inland Empire between September 2020 and January 2021. We systematically analyzed the focus group data using the rigorous and accelerated data reduction (RADaR) technique to identify key themes. Identified barriers included: lack of awareness, vaccine hesitancy, and perceived unaffordability (cited in 14 groups); lack of healthcare provider communication and insufficient time (13 groups); fear of moral judgment (12 groups); lack of motivation and information needs (10 groups); and lack of reliable transportation and foregone care during the COVID-19 pandemic (3 groups). Proposed facilitators included: tailored HPV vaccine information for mid-adults, cost mitigation, and improved vaccine accessibility (12 groups); healthcare provider-initiated conversations (6 groups); and vaccine reminders (4 groups). These findings highlight challenges to HPV vaccination among U.S. mid-adults eligible for SCDM and point to actionable strategies for improvement. Specifically, tailored educational interventions, decision-making tools for pharmacists, and integrating HPV vaccination into other healthcare encounters may enhance vaccination efforts in areas with limited primary care resources.
Candida parapsilosis is known to cause severe and persistent outbreaks in clinical settings. Patients infected with multidrug-resistant C. parapsilosis (MDR Cp) isolates were identified in a large Turkish hospital from 2017-2020. We subsequently identified three additional patients infected with MDR Cp isolates in 2022 from the same hospital and two echinocandin-resistant (ECR) isolates from a single patient in another hospital. The increasing number of MDR and ECR isolates contradicts the general principle that the severe fitness cost associated with these phenotypes could prevent their dominance in clinical settings. Here, we employed a multidimensional approach to systematically assess the fitness costs of MDR and ECR C. parapsilosis isolates. Whole-genome sequencing revealed a novel MDR genotype infecting two patients in 2022. Despite severe in vitro defects, the levels and tolerances of the biofilms of our ECR and MDR isolates were generally comparable to those of susceptible wild-type isolates. Surprisingly, the MDR and ECR isolates showed major alterations in their cell wall components, and some of the MDR isolates consistently displayed increased tolerance to the fungicidal activities of primary human neutrophils and were more immunoevasive during exposure to primary human macrophages. Our systemic infection mouse model showed that MDR and ECR C. parapsilosis isolates had comparable fungal burden in most organs relative to susceptible isolates. Overall, we observed a notable increase in the genotypic diversity and frequency of MDR isolates and identified MDR and ECR isolates potentially capable of causing persistent outbreaks in the future.
The interplay of conductivity and friction in layered materials such as graphite is an open area of investigation. Here, we measure local conductivity and friction on terraces of freshly cleaved highly oriented pyrolytic graphite via atomic force microscopy under ambient conditions. The graphite surface is found to exhibit a rich electrical landscape, with different terraces exhibiting different levels of conductivity. A peculiar dependency of conductivity on scan direction is observed on some terraces. The terraces that exhibit this dependency are also found to show enhanced friction values. A hypothesis based on tip asymmetry and the puckering effect is proposed to explain the findings. Our results highlight the non-triviality of the electrical and tribological properties of graphite on the nanoscale, as well as their interplay.
The standard redox potentials of metal nanoparticles are important for understanding their chemical properties. Traditionally, these redox potentials are measured by using voltammetry. Although voltammetry is fast and cost-effective, loading or landing the nanoparticles on electrodes alters their electrochemical properties, posing a challenge for accurately determining their intrinsic redox potentials. Here, a contactless method was developed utilizing chemical assays and the Nernst equation to measure the standard reduction potentials of gold nanoparticles in their colloidal state. To showcase the versatility and accuracy of this Nernstian approach, the reduction potentials were measured for a size range of 5.0-73 nm, revealing their scaling law and dependence on the nanoparticle surface energy.
Dynamin 1 (Dyn1) GTPase, a principal driver of membrane fission during synaptic endocytosis, self-assembles into short mechanoactive helices cleaving the necks of endocytic vesicles. While structural information about Dyn1 helix is abundant, little is known about the nanoscale dynamics of the helical scaffolding at the moment of fission, complicating mechanistic understanding of Dyn1 action. To address the role of the helix dynamics in fission, we used High-Speed Atomic Force Microscopy (HS-AFM) and fluorescence microscopy to track and compare the spatiotemporal characteristics of the helices formed by wild-type Dyn1 and its K44A mutant impaired in GTP hydrolysis on minimal lipid membrane templates. In the absence of nucleotide, membrane-bound WTDyn1 and K44ADyn1 self-assembled into tubular protein scaffolding of similar diameter encaging the lipid bilayer. In both cases, the GTP addition caused scaffold constriction coupled with formation of 20 to 30 nm nanogaps in the protein coverage. While both proteins reached scaffold diameters characteristic for membrane superconstriction causing fission, the fission was detected only with WTDyn1. We associated the fission activity with the dynamic evolution of the nanogaps: K44ADyn1 gaps were static, while WTDyn1 gaps actively evolved via repetitive nonaxisymmetric constriction-bending deformations caused by localized GTP hydrolysis. Modeling of the deformations implicated filament twist as an additional deformation mode which combines with superconstriction to facilitate membrane fission. Our results thus show that the dynamics of the Dyn1 helical scaffold goes beyond radial constriction and involves nonaxisymmetric deformations, where filament twist emerges as a critical driver of membrane fission.
