Emily P. Fox

An expanded regulatory network temporally controls Candida albicans biofilm formation

Candida albicans biofilms are composed of highly adherent and densely arranged cells with properties distinct from those of free‐floating (planktonic) cells. These biofilms are a significant medical problem because they commonly form on implanted medical devices, are drug resistant and are difficult to remove. C. albicans biofilms are not static structures; rather they are dynamic and develop over time. Here we characterize gene expression in biofilms during their development, and by comparing them to multiple planktonic reference states, we identify patterns of gene expression relevant to biofilm formation. In particular, we document time‐dependent changes in genes involved in adhesion and metabolism, both of which are at the core of biofilm development. Additionally, we identify three new regulators of biofilm formation, Flo8, Gal4, and Rfx2, which play distinct roles during biofilm development over time. Flo8 is required for biofilm formation at all time points, and Gal4 and Rfx2 are needed for proper biofilm formation at intermediate time points.

Genetic control of conventional and pheromone-stimulated biofilm formation in Candida albicans.

Candida albicans can stochastically switch between two phenotypes, white and opaque. Opaque cells are the sexually competent form of C. albicans and therefore undergo efficient polarized growth and mating in the presence of pheromone. In contrast, white cells cannot mate, but are induced – under a specialized set of conditions – to form biofilms in response to pheromone. In this work, we compare the genetic regulation of such “pheromone-stimulated” biofilms with that of “conventional” C. albicans biofilms. In particular, we examined a network of six transcriptional regulators (Bcr1, Brg1, Efg1, Tec1, Ndt80, and Rob1) that mediate conventional biofilm formation for their potential roles in pheromone-stimulated biofilm formation. We show that four of the six transcription factors (Bcr1, Brg1, Rob1, and Tec1) promote formation of both conventional and pheromone-stimulated biofilms, indicating they play general roles in cell cohesion and biofilm development. In addition, we identify the master transcriptional regulator of pheromone-stimulated biofilms as C. albicans Cph1, ortholog of Saccharomyces cerevisiae Ste12. Cph1 regulates mating in C. albicans opaque cells, and here we show that Cph1 is also essential for pheromone-stimulated biofilm formation in white cells. In contrast, Cph1 is dispensable for the formation of conventional biofilms. The regulation of pheromone- stimulated biofilm formation was further investigated by transcriptional profiling and genetic analyses. These studies identified 196 genes that are induced by pheromone signaling during biofilm formation. One of these genes, HGC1, is shown to be required for both conventional and pheromone-stimulated biofilm formation. Taken together, these observations compare and contrast the regulation of conventional and pheromone-stimulated biofilm formation in C. albicans, and demonstrate that Cph1 is required for the latter, but not the former.

A sticky situation: Untangling the transcriptional network controlling biofilm development in Candida albicans

Candida albicans is a commensal microorganism of the human microbiome; it is also the most prevalent fungal pathogen of humans. Many infections caused by C. albicans are a direct consequence of its proclivity to form biofilms—resilient, surface-associated communities of cells where individual cells acquire specialized properties that are distinct from those observed in suspension cultures. We recently identified the transcriptional network that orchestrates the formation of biofilms in C. albicans. These results set the stage for understanding how biofilms are formed and, once formed, how the specialized properties of biofilms are elaborated. This information will provide new insight for understanding biofilms in more detail and may lead to improvements in preventing and treating biofilm-based infections in the future.

A Histone Deacetylase Complex Mediates Biofilm Dispersal and Drug Resistance in Candida albicans

ABSTRACT Biofilms are resilient, surface-associated communities of cells with specialized properties (e.g., resistance to drugs and mechanical forces) that are distinct from those of suspension (planktonic) cultures. Biofilm formation by the opportunistic human fungal pathogen Candida albicans is medically relevant because C. albicans infections are highly correlated with implanted medical devices, which provide efficient substrates for biofilm formation; moreover, biofilms are inherently resistant to antifungal drugs. Biofilms are also important for C. albicans to colonize diverse niches of the human host. Here, we describe four core members of a conserved histone deacetylase complex in C. albicans (Set3, Hos2, Snt1, and Sif2) and explore the effects of their mutation on biofilm formation. We find that these histone deacetylase complex members are needed for proper biofilm formation, including dispersal of cells from biofilms and multifactorial drug resistance. Our results underscore the importance of the physical properties of biofilms in contributing to drug resistance and dispersal and lay a foundation for new strategies to target biofilm dispersal as a potential antifungal intervention. IMPORTANCE Through the formation of biofilms—surface-associated communities of cells—microorganisms can establish infections, become drug resistant, and evade the host immune system. Here we investigate how four core members of a conserved histone deacetylase complex mediate biofilm formation by Candida albicans, the major fungal pathogen of humans. We show that this histone deacetylase complex is required for biofilm dispersal, a process through which cells leave the biofilm to establish new infections. We also show that the deacetylase complex mediates biofilm drug resistance. This work provides new insight into how the physical properties of biofilms affect dispersal and drug resistance and suggests new potential antifungal strategies that could be effective against biofilms. Through the formation of biofilms—surface-associated communities of cells—microorganisms can establish infections, become drug resistant, and evade the host immune system. Here we investigate how four core members of a conserved histone deacetylase complex mediate biofilm formation by Candida albicans, the major fungal pathogen of humans. We show that this histone deacetylase complex is required for biofilm dispersal, a process through which cells leave the biofilm to establish new infections. We also show that the deacetylase complex mediates biofilm drug resistance. This work provides new insight into how the physical properties of biofilms affect dispersal and drug resistance and suggests new potential antifungal strategies that could be effective against biofilms.

A comparison of biofluid cytokine markers across platform technologies: Correspondence or divergence?

Background Quantification of biofluid cytokines is a rapidly growing area of translational research. However, comparability across the expanding number of available assay platforms for detection of the same proteins remains to be determined. We aimed to directly compare a panel of commonly measured cytokines in plasma of typically aging adults across two high sensitivity quantification platforms, Meso Scale Discovery high performance electrochemiluminiscence (HPE) and single‐molecule immunosorbent assays (Simoa) by Quanterix. Methods 57 community‐dwelling older adults completed a blood draw, neuropsychological assessment, and brain MRI as part of a healthy brain aging study. Plasma samples from the same draw dates were analyzed for IL‐10, IP‐10, IL‐6, TNF&agr;, and IL‐1&bgr; on HPE and Simoa, separately. Reliable detectability (coefficient of variance (CV) < 20% and outliers 3 interquartiles above the median removed), intra‐assay precision, absolute concentrations, reproducibility across platforms, and concurrent associations with external variables of interest (e.g., demographics, peripheral markers of vascular health, and brain health) were examined. Results The proportion of cytokines reliably measured on HPE (87.7–93.0%) and Simoa (75.4–93.0%) did not differ (ps > 0.32), with the exception of IL‐1&bgr; which was only reliably measured using Simoa (68.4%). On average, CVs were acceptable at <8% across both platforms. Absolute measured concentrations were higher using Simoa for IL‐10, IL‐6, and TNF&agr; (ps < 0.05). HPE and Simoa shared only small‐to‐moderate proportions of variance with one another on the same cytokine proteins (range: r = 0.26 for IL‐10 to r = 0.64 for IL‐6), though platform agreement did not dependent on cytokine concentrations. Cytokine ratios within each platform demonstrated similar relative patterns of up‐ and down‐regulation across HPE and Simoa, though still significantly differed (ps < 0.001). Supporting concurrent validity, all 95% confidence intervals of the correlations between cytokines and external variables overlapped between the two platforms. Moreover, most associations were in expected directions and consistently so across platforms (e.g., IL‐6 and TNF&agr;), though with several notable exceptions for IP‐10 and IL‐10. Conclusions HPE and Simoa showed comparable detectability and intra‐assay precision measuring a panel of commonly examined cytokine proteins, with the exception of IL‐1&bgr; which was not reliably detected on HPE. However, Simoa demonstrated overall higher concentrations and the two platforms did not show agreement when directly compared against one another. Relative cytokine ratios and associations demonstrated similar patterns across platforms. Absolute cytokine concentrations may not be directly comparable across platforms, may be analyte dependent, and interpretation may be best limited to discussion of relative associations. HighlightsMSD HPE and Quanterix Simoa are analytic platforms to detect biofluid cytokines.Detectability and reliability of the same cytokines did not differ across platforms.Absolute cytokine concentrations were higher using Simoa vs. HPE.Cross‐platform correlations were low, but cytokine ratios showed similar patterns.Cytokine measurement is platform‐specific and is best interpreted in relative terms.

Perceived Stress is Associated with Accelerated Monocyte/Macrophage Aging Trajectories in Clinically Normal Adults

OBJECTIVES Chronic stress is associated with poorer age-related cognition, but the mechanisms of this relationship are not well understood. Aging increases expression of activated macrophages, leading to exacerbated immune responses to stressors. We examined the impact of stress and aging on macrophage-related inflammation and cognition in clinically normal adults. METHODS Three hundred eighty clinically normal adults were followed longitudinally (age M = 73 years; visit range: 1-8; M = 2.5 visits). Participants completed the Perceived Stress Scale, a neuropsychological battery, and blood draws. Plasma was analyzed for cytokines related to macrophage function (interleukin 6, tumor necrosis factor alpha, macrophage inflammatory protein-1 alpha, macrophage inflammatory protein-1 beta). Linear mixed-effects examined the effects of age, baseline stress, and their interaction predicting macrophage cytokines, adjusting for sex, education, and depressive symptoms. Latent growth curve models assessed the mediating role of macrophage cytokines in the relationship between age and cognition in high or low stress. RESULTS Baseline perceived stress interacted with age to predict macrophage cytokines longitudinally. Specifically, high-stress adults demonstrated accelerated age-related elevations in macrophage cytokines across time. Macrophage cytokines negatively tracked with executive functioning longitudinally. Macrophage cytokines mediated 19% of the relationship between age and executive function in high-stress, but not low-stress, adults. CONCLUSIONS Our data provide evidence of accelerated immune aging among individuals with high stress. Elevated macrophage cytokine trajectories mediated the effect of age on executive function only in individuals with high stress, suggesting these constructs may be more tightly linked in elevated stress contexts. Stress interventions are warranted to optimize immune aging, with possible downstream cognitive benefits among even clinically normal adults.

The transcriptional network controlling biofilm development in Candida albicans

Candida albicans is a natural component of the human microbiome, but it is also the major fungal pathogen of humans. This fungus asymptomatically colonizes many areas of the human body, especially the gastrointestinal and genitourinary tracts of healthy individuals. Alterations in host immunity, stress, resident microbiota and other factors can lead to C. albicans overgrowth, causing a wide range of infections from superficial mucosal to hematogenously disseminated candidiasis. Candida infections are especially serious in immunocompromised individuals, such as AIDS patients, patients undergoing anticancer therapies, and transplantation patients receiving immunosuppression therapy, as well as immunocompetent patients with implanted medical devices. To date, most studies of C. albicans have been performed in suspension cultures; however, the medical impact of C. albicans (like that of many other microorganisms) depends on its ability to form surface-associated communities called biofilms. Biofilms are notorious for forming on various implanted medical devices, including catheters, pacemakers, heart valves, dentures and prosthetic joints, which provide a surface and sanctuary for biofilm