Comparison of Healthy and Dandruff Scalp Microbiome Reveals the Role of Commensals in Scalp Health

Several scalp microbiome studies from different populations have revealed the association of dandruff with bacterial and fungal dysbiosis. However, the functional role of scalp microbiota in scalp disorders and health remains scarcely explored. Here, we examined the bacterial and fungal diversity of the scalp microbiome and their potential functional role in the healthy and dandruff scalp of 140 Indian women. Propionibacterium acnes and Staphylococcus epidermidis emerged as the core bacterial species, where the former was associated with a healthy scalp and the latter with dandruff scalp. Along with the commonly occurring Malassezia species (M. restricta and M. globosa) on the scalp, a strikingly high association of dandruff with yet uncharacterized Malassezia species was observed in the core mycobiome. Functional analysis showed that the fungal microbiome was enriched in pathways majorly implicated in cell-host adhesion in the dandruff scalp, while the bacterial microbiome showed a conspicuous enrichment of pathways related to the synthesis and metabolism of amino acids, biotin, and other B-vitamins, which are reported as essential nutrients for hair growth. A systematic measurement of scalp clinical and physiological parameters was also carried out, which showed significant correlations with the microbiome and their associated functional pathways. The results point toward a new potential role of bacterial commensals in maintaining the scalp nutrient homoeostasis and highlights an important and yet unknown role of the scalp microbiome, similar to the gut microbiome. This study, therefore, provides new perspectives on the better understanding of the pathophysiology of dandruff

The Dual Activity Responsible for the Elongation and Branching of β-(1,3)-Glucan in the Fungal Cell Wall

International audienceβ-(1,3)-Glucan, the major fungal cell wall component, ramifies through β-(1,6)-glycosidic linkages, which facilitates its binding with other cell wall components contributing to proper cell wall assembly. Using Saccharomyces cerevisiae as a model, we developed a protocol to quantify β-(1,6)-branching on β-(1,3)-glucan. Permeabilized S. cerevisiae and radiolabeled substrate UDP-(14C)glucose allowed us to determine branching kinetics. A screening aimed at identifying deletion mutants with reduced branching among them revealed only two, the bgl2Δ and gas1Δ mutants, showing 15% and 70% reductions in the branching, respectively, compared to the wild-type strain. Interestingly, a recombinant Gas1p introduced β-(1,6)-branching on the β-(1,3)-oligomers following its β-(1,3)-elongase activity. Sequential elongation and branching activity of Gas1p occurred on linear β-(1,3)-oligomers as well as Bgl2p-catalyzed products [short β-(1,3)-oligomers linked by a linear β-(1,6)-linkage]. The double S. cerevisiae gas1Δ bgl2Δ mutant showed a drastically sick phenotype. An ScGas1p ortholog, Gel4p from Aspergillus fumigatus, also showed dual β-(1,3)-glucan elongating and branching activity. Both ScGas1p and A. fumigatus Gel4p sequences are endowed with a carbohydrate binding module (CBM), CBM43, which was required for the dual β-(1,3)-glucan elongating and branching activity. Our report unravels the β-(1,3)-glucan branching mechanism, a phenomenon occurring during construction of the cell wall which is essential for fungal life.IMPORTANCE The fungal cell wall is essential for growth, morphogenesis, protection, and survival. In spite of being essential, cell wall biogenesis, especially the core β-(1,3)-glucan ramification, is poorly understood; the ramified β-(1,3)-glucan interconnects other cell wall components. Once linear β-(1,3)-glucan is synthesized by plasma membrane-bound glucan synthase, the subsequent event is its branching event in the cell wall space. Using Saccharomyces cerevisiae as a model, we identified GH72 and GH17 family glycosyltransferases, Gas1p and Bgl2p, respectively, involved in the β-(1,3)-glucan branching. The sick phenotype of the double Scgas1Δ bgl2Δ mutant suggested that β-(1,3)-glucan branching is essential. In addition to ScGas1p, GH72 family ScGas2p and Aspergillus fumigatus Gel4p, having CBM43 in their sequences, showed dual β-(1,3)-glucan elongating and branching activity. Our report identifies the fungal cell wall β-(1,3)-glucan branching mechanism. The essentiality of β-(1,3)-glucan branching suggests that enzymes involved in the glucan branching could be exploited as antifungal targets

The N-terminal Domain of Drosophila Gram-negative Binding Protein 3 (GNBP3) Defines a Novel Family of Fungal Pattern Recognition Receptors

International audienceGram-negative binding protein 3 (GNBP3), a pattern recognition receptor that circulates in the hemolymph of Drosophila, is responsible for sensing fungal infection and triggering Toll pathway activation. Here, we report that GNBP3 N-terminal domain binds to fungi upon identifying long chains of beta-1,3-glucans in the fungal cell wall as a major ligand. Interestingly, this domain fails to interact strongly with short oligosaccharides. The crystal structure of GNBP3-Nter reveals an immunoglobulin-like fold in which the glucan binding site is masked by a loop that is highly conserved among glucan-binding proteins identified in several insect orders. Structure-based mutagenesis experiments reveal an essential role for this occluding loop in discriminating between short and long polysaccharides. The displacement of the occluding loop is necessary for binding and could explain the specificity of the interaction with long chain structured polysaccharides. This represents a novel mechanism for beta-glucan recognition

Body Site Is a More Determinant Factor than Human Population Diversity in the Healthy Skin Microbiome

<div><p>We studied skin microbiota present in three skin sites (forearm, axilla, scalp) in men from six ethnic groups living in New York City. <b>Methods.</b> Samples were obtained at baseline and after four days following use of neutral soap and stopping regular hygiene products, including shampoos and deodorants. DNA was extracted using the MoBio Power Lyzer kit and 16S rRNA gene sequences determined on the IIlumina MiSeq platform, using QIIME for analysis. <b>Results.</b> Our analysis confirmed skin swabbing as a useful method for sampling different areas of the skin because DNA concentrations and number of sequences obtained across subject libraries were similar. We confirmed that skin location was the main factor determining the composition of bacterial communities. Alpha diversity, expressed as number of species observed, was greater in arm than on scalp or axilla in all studied groups. We observed an unexpected increase in α-diversity on arm, with similar tendency on scalp, in the South Asian group after subjects stopped using their regular shampoos and deodorants. Significant differences at phylum and genus levels were observed between subjects of the different ethnic origins at all skin sites. <b>Conclusions.</b> We conclude that ethnicity and particular soap and shampoo practices are secondary factors compared to the ecological zone of the human body in determining cutaneous microbiota composition.</p></div

Comparison of microbiota differences by ethnic group, analyzed by LEfSe using baseline samples.

<p>Taxa with relative abundance ≥ 0.1% present in at least one sample in each location were included. The cladograms indicate the phylogenetic distribution of the microbial lineages associated with ethnic group, with lineages with Linear Discriminant Analysis (LDA) score ≥2.0 displayed. Significance differences for each ethnic group of the most abundant class are indicated by color, as indicated in the key. Each node’s diameter is proportional to the taxon’s abundance. The strategy of multiclass analysis is non-strict (≥ one class differential). Nodes represent phylogenetic levels from domain to genus (from inside out) (AA, n = 18 subjects; AC, n = 11; CA, n = 16; EA, n = 25; LA, n = 19; SA, n = 20).</p

Taxonomic analysis of cutaneous microbiota from 110 subjects in six different subject groups.

<p><b>Panel A.</b> By site (arm, axilla, scalp) at the phylum level. A total of 41 phyla were found. The sequences for the dominant 9 phyla (>0.1% in any group) accounted for >99.7% of total sequences in all ethnic groups. Baseline: Samples collected before special soap wash used. Follow-up: Samples collected after special soap wash used. <b>Panel B:</b> Genus level. A total 726 genera were detected; only predominant genera (Mean>0.01%) are shown.</p

Comparison of diversity of the cutaneous microbiota over time, by ethnic group.

<p><b>Panel A.</b> Rarefaction analysis of cutaneous microbiota in relation to sampling time in samples from arms (Brown), axilla (Black) and scalp (Yellow), African-American (AA, n = 108 samples); African-Continental (AC, n = 66); Caucasian-American (CA, n = 96); East-Asian (EA, n = 150); Latin-American (LA, n = 114); South-Asian (SA, n = 96). Rarefaction analysis represented by phylogenetic distance. The solid and dashed lines indicate samples collected before (T1) and after (T2) a special soap wash was used, respectively. Only a significant difference between time-points was found for the SA group. <b>Panel B.</b> Intra- and inter-group beta-diversity over time. Mean (±SD) pairwise unweighted UniFrac distances are shown. The ethnic groups are colored as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0151990#pone.0151990.g001" target="_blank">Fig 1</a>. Significance was determined by one-way ANOVA with the Tukey method for correction for multiple comparisons (*<0.05; **<0.01; ***<0.001; ****<0.0001).</p

Staphylococcus epidermidis isolates from atopic or healthy skin have opposite effect on skin cells: potential implication of the AHR pathway modulation

International audienceIntroduction S taphylococcus epidermidis is a commensal bacterium ubiquitously present on human skin. This species is considered as a key member of the healthy skin microbiota, involved in the defense against pathogens, modulating the immune system, and involved in wound repair. Simultaneously, S. epidermidis is the second cause of nosocomial infections and an overgrowth of S. epidermidis has been described in skin disorders such as atopic dermatitis. Diverse isolates of S. epidermidis co-exist on the skin. Elucidating the genetic and phenotypic specificities of these species in skin health and disease is key to better understand their role in various skin conditions. Additionally, the exact mechanisms by which commensals interact with host cells is partially understood. We hypothesized that S. epidermidis isolates identified from different skin origins could play distinct roles on skin differentiation and that these effects could be mediated by the aryl hydrocarbon receptor (AhR) pathway. Methods For this purpose, a library of 12 strains originated from healthy skin (non-hyperseborrheic (NH) and hyperseborrheic (H) skin types) and disease skin (atopic (AD) skin type) was characterized at the genomic and phenotypic levels. Results and discussion Here we showed that strains from atopic lesional skin alter the epidermis structure of a 3D reconstructed skin model whereas strains from NH healthy skin do not. All strains from NH healthy skin induced AhR/OVOL1 path and produced high quantities of indole metabolites in co-culture with NHEK; especially indole-3-aldehyde (IAld) and indole-3-lactic acid (ILA); while AD strains did not induce AhR/OVOL1 path but its inhibitor STAT6 and produced the lowest levels of indoles as compared to the other strains. As a consequence, strains from AD skin altered the differentiation markers FLG and DSG1. The results presented here, on a library of 12 strains, showed that S. epidermidis originated from NH healthy skin and atopic skin have opposite effects on the epidermal cohesion and structure and that these differences could be linked to their capacity to produce metabolites, which in turn could activate AHR pathway. Our results on a specific library of strains provide new insights into how S. epidermidis may interact with the skin to promote health or disease