Biology subjects

Markey, L.

Publications and source records attributed to Markey, L..

6 recordsLinked to original sources

bioRxiv2025

Microbiome diversity of low biomass skin sites is captured by metagenomics but not 16S amplicon sequencing

Established workflows for microbiome analysis work well for high microbial biomass samples, like stool, but often fail to accurately define microbial communities when applied to low microbial biomass samples. Here, we systemically compare microbiome analysis methods --16S rRNA sequencing, shallow metagenomics, and qPCR PMP panels--as well as extraction methods across skin swab samples and mock community dilutions. While extraction method minimally impacted results, with no significant signal for method-specific contamination or bias, we observed critical differences in inferred composition across analysis methods for low biomass samples. Metagenomic sequencing and qPCR revealed concordant, diverse microbial communities on low biomass leg skin samples, whereas 16S amplicon sequencing exhibited extreme bias toward the most abundant taxon. Both qPCR and metagenomics showed that female genital tract bacteria dominated the leg skin microbiome in about half of female subjects. Metagenomics also enabled sub-species analysis, which demonstrated that individuals have consistent within-species diversity across high-biomass forehead and low-biomass leg skin sites. This work illustrates that shallow metagenomics provides the necessary sensitivity and taxonomic resolution to characterize species and strain-level diversity in extremely low biomass samples, opening possibilities for microbiome discovery in previously unexplored niches. ImportanceDespite the importance of the skin microbiome in health and disease, there have been far fewer studies characterizing the microbiome of skin compared to that of the gut. In part, this is because microbiome methods were initially developed for bacteria-rich samples like stool and these methods perform poorly on bacteria-poor samples like skin swabs. The perceived difficulty of getting reliable data from such low biomass sites has limited the scope and types of analyses performed. Here we demonstrate that shotgun, whole-genome, metagenomic sequencing recovers the full input from even very dilute control community samples, and reveals a highly diverse population even on very low biomass skin sites. We describe a streamlined sample processing and analysis pipeline which can be applied broadly to characterize low biomass microbiome samples and reveal new host-microbiome interactions.

microbiology↗

bioRxiv2025

Intraspecies associations from strain-rich metagenome samples

Genetically distinct strains of a species can vary widely in phenotype, reducing the utility of species-resolved microbiome measurements for detecting associations with health or disease. While metagenomics theoretically provides information on all strains in a sample, current strain-resolved analysis methods face a tradeoff: de novo genotyping approaches can detect novel strains but struggle when applied to strain-rich or low-coverage samples, while reference database methods work robustly across sample types but are insensitive to novel diversity. We present PHLAME, a method that bridges this divide by combining the advantages of reference-based approaches with novelty awareness. PHLAME explicitly defines clades at multiple phylogenetic levels and introduces a probabilistic, mutation-based, framework to accurately quantify novelty from the nearest reference. By applying PHLAME to publicly available human skin and vaginal metagenomes, we uncover previously undetected clade associations with coexisting species, geography, and host age. The ability to characterize intraspecies associations and dynamics in previously inaccessible environments will propel new mechanistic insights from accumulating metagenomic data.

microbiology↗

bioRxiv2023

Commensal skin bacteria exacerbate inflammation and delay skin healing

The skin microbiome can both trigger beneficial immune stimulation and pose a potential infection threat. Previous studies have shown that colonization of mouse skin with the model human skin commensal Staphylococcus epidermidis is protective against subsequent excisional wound or pathogen challenge. However, less is known about concurrent skin damage and exposure to commensal microbes, despite growing interest in interventional probiotic therapy. Here, we address this open question by applying commensal skin bacteria at a high dose to abraded skin. While depletion of the skin microbiome via antibiotics delayed repair from damage, application of commensals-- including the mouse commensal Staphylococcus xylosus, three distinct isolates of S. epidermidis, and all other tested human skin commensals-- also significantly delayed barrier repair. Increased inflammation was observed within four hours of S. epidermidis exposure and persisted through day four, at which point the skin displayed a chronic-wound-like inflammatory state with increased neutrophil infiltration, increased fibroblast activity, and decreased monocyte differentiation. Transcriptomic analysis suggested that the prolonged upregulation of early canonical proliferative pathways inhibited the progression of barrier repair. These results highlight the nuanced role of members of the skin microbiome in modulating barrier integrity and indicate the need for caution in their development as probiotics.

microbiology↗

bioRxiv2020

Colonization with the commensal fungus Candida albicans perturbs the gut-brain axis through dysregulation of endocannabinoid signaling

Anxiety disorders are the most prevalent mental health disorder worldwide, with a lifetime prevalence of 5-7% of the human population. Although the etiology of anxiety disorders is incompletely understood, one aspect of host health that affects anxiety disorders is the gut-brain axis. We used a mouse model of gastrointestinal (GI) colonization to demonstrate that the commensal fungus Candida albicans affects host health via the gut-brain axis. In mice, bacterial members of the gut microbiota can influence the host gut-brain axis, affecting anxiety-like behavior and the hypothalamus-pituitary-adrenal (HPA) axis which produces the stress hormone corticosterone (CORT). Here we showed that mice colonized with C. albicans demonstrated increased anxiety-like behavior and increased basal production of CORT as well as dysregulation of CORT production following acute stress. The HPA axis and anxiety-like behavior are negatively regulated by the endocannabinoid anandamide (AEA). C. albicans-colonized mice exhibited systemic changes in the endocannabinoidome, within the GI tract and the brain, and showed a negative correlation between brain AEA levels and serum CORT. Further, increasing AEA levels using the well-characterized fatty acid amide hydrolase (FAAH) inhibitor URB597 was sufficient to reverse both neuroendocrine phenotypes in C. albicans-colonized mice. Thus, a commensal fungus that is a common colonizer of humans had widespread effects on the physiology of its host. To our knowledge, this is the first report of microbial manipulation of the endocannabinoid (eCB) system that resulted in neuroendocrine changes contributing to anxiety-like behavior.

microbiology↗

bioRxiv2019

A novel genetic circuitry governing hypoxic metabolic flexibility, commensalism and virulence in the fungal pathogen Candida albicans

Inside the human host, the pathogenic yeast Candida albicans colonizes predominantly oxygen-poor niches such as the gastrointestinal and vaginal tracts, but also oxygen-rich environments such as cutaneous epithelial cells and oral mucosa. This suppleness requires an effective mechanism to reprogram reversibly the primary metabolism in response to oxygen variation. Here, we have uncovered that Snf5, a subunit of SWI/SNF chromatin remodeling complex, is a major transcriptional regulator that links oxygen status to the metabolic capacity of C. albicans. Snf5 and other subunits of SWI/SNF complex were required to activate genes of alternative carbon utilization and other carbohydrates related process specifically under hypoxia. snf5 mutant exhibited an altered metabolome reflecting that SWI/SNF plays an essential role in maintaining metabolic homeostasis and carbon flux in C. albicans under hypoxia. Snf5 was necessary to activate the transcriptional program linked to both commensal and invasive growth. Accordingly, snf5 was unable to maintain its growth in the stomach, the cecum and the colon of mice. snf5 was also avirulent as it was unable to invade Galleria larvae or to cause damage to human enterocytes and murine macrophages. Among candidates of signaling pathways in which Snf5 might operate, phenotypic analysis revealed that mutants of Ras1-cAMP-PKA pathway, as well as mutants of Yak1 and Yck2 kinases exhibited a similar carbon flexibility phenotype as did snf5 under hypoxia. Genetic interaction analysis indicated that the adenylate cyclase Cyr1, a key component of the Ras1-cAMP pathway, but not Ras1, interacted genetically with Snf5. Our study yielded unprecedented insight into the oxygen-sensitive regulatory circuit that control metabolic flexibility, stress, commensalism and virulence in C. albicans.\n\nAuthor SummaryA critical aspect of eukaryotic cell fitness is the ability to sense and adapt to variations in oxygen concentrations in their local environment. Hypoxia leads to a substantial remodeling of cell metabolism and energy homeostasis, and thus, organisms must develop an effective regulatory mechanism to cope with oxygen depletion. Candida albicans is an opportunistic yeast that is the most prevalent human fungal pathogens. This yeast colonizes diverse niches inside the human host with contrasting carbon sources and oxygen concentrations. While hypoxia is the predominant conditions that C. albicans encounters inside most of the niches, the impact of this condition on metabolic flexibility, a major determinant of fungal virulence, was completely neglected. Here, we uncovered that the chromatin remodelling complex SWI/SNF is master regulatory circuit that links oxygen status to a broad spectrum of carbon utilization routes. Snf5 was essential for the maintenance of C. albicans as a commensal and also for the expression of its virulence. The oxygen-sensitive regulators identified in this work provide a framework to comprehensively understand the virulence of human fungal pathogens and represent a therapeutic value to fight fungal infections.

microbiology↗

bioRxiv2018

A synthetic system that combats fungal infections

Due to a limited set of antifungals available and problems in early diagnosis invasive fungal infections caused by Candida species are among the most common hospital-acquired infections with staggering mortality rates. Here, we describe an engineered system able to sense and respond to the fungal pathogen Candida albicans, the most common cause of candidemia. In doing so, we identified hydroxyphenylacetic acid (HPA) as a novel molecule secreted by C. albicans. Furthermore, we engineered E. coli to be able to sense HPA produced by C. albicans. Finally, we constructed a sense-and-respond system by coupling the C. albicans sensor to the production of an inhibitor of hypha formation thereby reducing filamentation, virulence factor expression and fungal-induced epithelial damage. This system could be used as a basis for the development of novel prophylactic approaches to prevent fungal infections.

synthetic biology↗