Detection of Staphylococcal Species Diversity within Human Skin Microbiome Using a Simple PCR-SSCP Technique

Objective. The aim of this study was to detect the diversity of Staphylococcus species of a healthy human skin using a simple techniquevPCR-SSCP. Methods: Blood samples, saliva, and skin swaps samples were collected from 50 persons from Hilla City - Iraq. The genomic DNA was extracted from these samples using the Bacteria Genomic DNA Kit. The concentrations and purity of DNA extract estimated by NanoDrop spectrophotometer. Polymerase chain reaction – single-strand conformational polymorphism (PCR–SSCP) technique was performed to detect the diversity between Staphylococcal species in the human skin microbiome using a specific primer of the 16SrRNA gene. Results: The PCR results, indicated that the Staphylococcal species were found within the ski community, but it's not infected blood and mouth of test healthy individuals. SSCP-heteroduplex patterns of PCR products appeared the presence Staphylococcal species diversity within skin microbiome of test healthy individuals. Conclusion: In spite of the PCR-SSCP, heteroduplex method was simple and cheap and appeared the diversity betweenvStaphylococcalspeciesvin the human skin microbiome, but it's not diagnosed the bacterial strains. So these results required to confirm by DNAvsequencingvtechnique.

Download Full-text

Temporal Stability of the Healthy Human Skin Microbiome Following Dead Sea Climatotherapy

Acta Dermato Venereologica ◽

10.2340/00015555-2769 ◽

2018 ◽

Vol 98 (2) ◽

pp. 256-261 ◽

Cited By ~ 8

Author(s):

M Brandwein ◽

G Fuks ◽

A Israel ◽

A Al-Ashhab ◽

D Nejman ◽

...

Keyword(s):

Human Skin ◽

Temporal Stability ◽

Dead Sea ◽

Skin Microbiome ◽

Healthy Human

Download Full-text

A Mixed Community of Skin Microbiome Representatives Influences Cutaneous Processes more than Individual Members

10.21203/rs.2.24334/v1 ◽

2020 ◽

Author(s):

Kristin H Loomis ◽

Susan K Wu ◽

Amanda Ernlund ◽

Kristina Zudock ◽

Allison Reno ◽

...

Keyword(s):

Microbial Community ◽

Human Skin ◽

Skin Microbiome ◽

Colonization Resistance ◽

Proliferating Cells ◽

Healthy Human ◽

Host Interactions ◽

The Individual ◽

The Impact ◽

Mixed Community

Abstract Background Skin, the largest organ of the human body by weight, hosts a diversity of microorganisms that can influence health. The microbial residents of the skin are now appreciated for their roles in host immune interactions, wound healing, colonization resistance, and various skin disorders. Still, much remains to be discovered in terms of the host pathways influenced by skin microorganisms, as well as the higher-level skin properties impacted through these microbe-host interactions. Towards this direction, recent efforts using mouse models pointed to pronounced changes in the transcriptional profiles of the skin in response to the presence of a microbial community. However, there is a need to quantify the roles of microorganisms at both the individual and community-level in healthy human skin. In this study, we utilize human skin equivalents to study the effects of individual taxa and a microbial community in a precisely controlled context. Through transcriptomics analysis, we identify key genes and pathways influenced by skin microbes, and we also characterize higher-level impacts on skin processes and properties through histological analyses. Results The presence of a microbiome on a 3D skin tissue model led to significantly altered patterns of gene expression, influencing genes involved in the regulation of apoptosis, proliferation, and the extracellular matrix (among others). Moreover, microbiome treatment influenced the thickness of the epidermal layer, reduced the number of actively proliferating cells, and increased filaggrin expression. Many of these findings were evident upon treatment with the mixed community, but either not detected or less pronounced in treatments by single microorganisms, underscoring the impact that a diverse skin microbiome has on the host. Conclusions This work contributes to the understanding of how microbiome constituents individually and collectively influence human skin processes and properties. The results show that, while it is important to understand the effect of individual microbes on the host, a full community of microbes has unique and pronounced effects on the skin. Thus, in its impacts on the host, the skin microbiome is more than the sum of its parts.

Download Full-text

Staphylococcus aureus-Specific Tissue-Resident Memory CD4+ T Cells Are Abundant in Healthy Human Skin

Frontiers in Immunology ◽

10.3389/fimmu.2021.642711 ◽

2021 ◽

Vol 12 ◽

Author(s):

Astrid Hendriks ◽

Malgorzata Ewa Mnich ◽

Bruna Clemente ◽

Ana Rita Cruz ◽

Simona Tavarini ◽

...

Keyword(s):

Staphylococcus Aureus ◽

T Cells ◽

Human Skin ◽

Ex Vivo ◽

Functional Characterization ◽

T Cell Response ◽

Skin Microbiome ◽

Entry Site ◽

Healthy Human ◽

Skin Explants

The skin is an immunocompetent tissue that harbors several kinds of immune cells and a plethora of commensal microbes constituting the skin microbiome. Staphylococcus aureus is a prominent skin pathogen that colonizes a large proportion of the human population. We currently have an incomplete understanding of the correlates of protection against S. aureus infection, however genetic and experimental evidence has shown that CD4+ T cells play a key role in orchestrating a protective anti-S. aureus immune response. A high S. aureus-specific memory CD4+ T cell response has been reported in the blood of healthy subjects. Since T cells are more abundant in the skin than in blood, we hypothesized that S. aureus-specific CD4+ T cells could be present in the skin of healthy individuals. Indeed, we observed proliferation of tissue-resident memory CD4+ T cells and production of IL-17A, IL-22, IFN-γ and TNF-β by cells isolated from abdominal skin explants in response to heat-killed S. aureus. Remarkably, these cytokines were produced also during an ex vivo epicutaneous S. aureus infection of human skin explants. These findings highlight the importance of tissue-resident memory CD4+ T cells present at barrier sites such as the skin, a primary entry site for S. aureus. Further phenotypical and functional characterization of these cells will ultimately aid in the development of novel vaccine strategies against this elusive pathogen.

Download Full-text

Staphylococcus saccharolyticus: An Overlooked Human Skin Colonizer

Microorganisms ◽

10.3390/microorganisms8081105 ◽

2020 ◽

Vol 8 (8) ◽

pp. 1105

Author(s):

Charlotte M. Ahle ◽

Kristian Stødkilde ◽

Mastaneh Afshar ◽

Anja Poehlein ◽

Lesley A. Ogilvie ◽

...

Keyword(s):

Human Skin ◽

Staphylococcus Epidermidis ◽

Amplicon Sequencing ◽

Growth Conditions ◽

Skin Microbiota ◽

Healthy Human ◽

Staphylococcal Species ◽

Staphylococcus Capitis ◽

Culture Independent ◽

Slow Growing

Coagulase-negative staphylococcal species constitute an important part of the human skin microbiota. In particular, facultative anaerobic species such as Staphylococcus epidermidis and Staphylococcus capitis can be found on the skin of virtually every human being. Here, we applied a culture-independent amplicon sequencing approach to identify staphylococcal species on the skin of healthy human individuals. While S. epidermidis and S. capitis were found as primary residents of back skin, surprisingly, the third most abundant member was Staphylococcus saccharolyticus, a relatively unstudied species. A search of skin metagenomic datasets detected sequences identical to the genome of S. saccharolyticus in diverse skin sites, including the back, forehead, and elbow pit. Although described as a slow-growing anaerobic species, a re-evaluation of its growth behavior showed that S. saccharolyticus can grow under oxic conditions, and, in particular, in a CO2-rich atmosphere. We argue here that S. saccharolyticus was largely overlooked in previous culture-dependent and -independent studies, due to its requirement for fastidious growth conditions and the lack of reference genome sequences, respectively. Future studies are needed to unravel the microbiology and host-interacting properties of S. saccharolyticus and its role as a prevalent skin colonizer.

Download Full-text

An enhanced characterization of the human skin microbiome: a new biodiversity of microbial interactions

10.1101/2020.01.21.914820 ◽

2020 ◽

Author(s):

Akintunde Emiola ◽

Wei Zhou ◽

Julia Oh

Keyword(s):

Human Skin ◽

De Novo ◽

Microbial Interactions ◽

Skin Microbiome ◽

Metagenomic Sequencing ◽

Interspecies Interactions ◽

Microbial Composition ◽

Healthy Human ◽

Skin Site ◽

Cutibacterium Acnes

ABSTRACTThe healthy human skin microbiome is shaped by skin site physiology, individual-specific factors, and is largely stable over time despite significant environmental perturbation. Studies identifying these characteristics used shotgun metagenomic sequencing for high resolution reconstruction of the bacteria, fungi, and viruses in the community. However, these conclusions were drawn from a relatively small proportion of the total sequence reads analyzable by mapping to known reference genomes. ‘Reference-free’ approaches, based on de novo assembly of reads into genome fragments, are also limited in their ability to capture low abundance species, small genomes, and to discriminate between more similar genomes. To account for the large fraction of non-human unmapped reads on the skin—referred to as microbial ‘dark matter’—we used a hybrid de novo and reference-based approach to annotate a metagenomic dataset of 698 healthy human skin samples. This approach reduced the overall proportion of uncharacterized reads from 42% to 17%. With our refined characterization, we revisited assumptions about the skin microbiome, and demonstrated higher biodiversity and lower stability, particularly in dry and moist skin sites. To investigate hypotheses underlying stability, we examined growth dynamics and interspecies interactions in these communities. Surprisingly, even though most skin sites were relatively stable, many dominant skin microbes, including Cutibacterium acnes and staphylococci, were actively growing in the skin, with poor or no relationship between growth rate and relative abundance, suggesting that host selection or interspecies competition may be important factors maintaining community homeostasis. To investigate other mechanisms facilitating adaptation to a specific skin site, we identified Staphylococcus epidermidis genes that are likely involved in stress response and provide mechanisms essential for growth in oily sites. Finally, horizontal gene transfer—another mechanism of competition by which strains may swap antagonistic or virulent coding regions—was relatively limited in healthy skin, but suggested exchange of different metabolic and environmental tolerance pathways. Altogether, our findings underscore the value of a combined reference-based and de novo approach to provide significant new insights into microbial composition, physiology, and interspecies interactions to maintain community homeostasis in the healthy human skin microbiome.

Download Full-text

The Human Skin Double-Stranded DNA Virome: Topographical and Temporal Diversity, Genetic Enrichment, and Dynamic Associations with the Host Microbiome

mBio ◽

10.1128/mbio.01578-15 ◽

2015 ◽

Vol 6 (5) ◽

Cited By ~ 110

Author(s):

Geoffrey D. Hannigan ◽

Jacquelyn S. Meisel ◽

Amanda S. Tyldsley ◽

Qi Zheng ◽

Brendan P. Hodkinson ◽

...

Keyword(s):

Human Skin ◽

Dna Sequences ◽

Skin Microbiome ◽

Metagenomic Sequencing ◽

Healthy Human ◽

Cutaneous Manifestations ◽

Virus Like Particles ◽

Double Stranded Dna ◽

Viral Communities ◽

Health And Disease

ABSTRACT Viruses make up a major component of the human microbiota but are poorly understood in the skin, our primary barrier to the external environment. Viral communities have the potential to modulate states of cutaneous health and disease. Bacteriophages are known to influence the structure and function of microbial communities through predation and genetic exchange. Human viruses are associated with skin cancers and a multitude of cutaneous manifestations. Despite these important roles, little is known regarding the human skin virome and its interactions with the host microbiome. Here we evaluated the human cutaneous double-stranded DNA virome by metagenomic sequencing of DNA from purified virus-like particles (VLPs). In parallel, we employed metagenomic sequencing of the total skin microbiome to assess covariation and infer interactions with the virome. Samples were collected from 16 subjects at eight body sites over 1 month. In addition to the microenviroment, which is known to partition the bacterial and fungal microbiota, natural skin occlusion was strongly associated with skin virome community composition. Viral contigs were enriched for genes indicative of a temperate phage replication style and also maintained genes encoding potential antibiotic resistance and virulence factors. CRISPR spacers identified in the bacterial DNA sequences provided a record of phage predation and suggest a mechanism to explain spatial partitioning of skin phage communities. Finally, we modeled the structure of bacterial and phage communities together to reveal a complex microbial environment with a Corynebacterium hub. These results reveal the previously underappreciated diversity, encoded functions, and viral-microbial dynamic unique to the human skin virome. IMPORTANCE To date, most cutaneous microbiome studies have focused on bacterial and fungal communities. Skin viral communities and their relationships with their hosts remain poorly understood despite their potential to modulate states of cutaneous health and disease. Previous studies employing whole-metagenome sequencing without purification for virus-like particles (VLPs) have provided some insight into the viral component of the skin microbiome but have not completely characterized these communities or analyzed interactions with the host microbiome. Here we present an optimized virus purification technique and corresponding analysis tools for gaining novel insights into the skin virome, including viral “dark matter,” and its potential interactions with the host microbiome. The work presented here establishes a baseline of the healthy human skin virome and is a necessary foundation for future studies examining viral perturbations in skin health and disease.

Download Full-text

A mixed community of skin microbiome representatives influences cutaneous processes more than individual members

Microbiome ◽

10.1186/s40168-020-00963-1 ◽

2021 ◽

Vol 9 (1) ◽

Author(s):

Kristin H. Loomis ◽

Susan K. Wu ◽

Amanda Ernlund ◽

Kristina Zudock ◽

Allison Reno ◽

...

Keyword(s):

Microbial Community ◽

Human Skin ◽

Skin Microbiome ◽

Colonization Resistance ◽

Proliferating Cells ◽

Healthy Human ◽

Host Interactions ◽

The Individual ◽

The Impact ◽

Mixed Community

Abstract Background Skin, the largest organ of the human body by weight, hosts a diversity of microorganisms that can influence health. The microbial residents of the skin are now appreciated for their roles in host immune interactions, wound healing, colonization resistance, and various skin disorders. Still, much remains to be discovered in terms of the host pathways influenced by skin microorganisms, as well as the higher-level skin properties impacted through these microbe-host interactions. Towards this direction, recent efforts using mouse models point to pronounced changes in the transcriptional profiles of the skin in response to the presence of a microbial community. However, there is a need to quantify the roles of microorganisms at both the individual and community-level in healthy human skin. In this study, we utilize human skin equivalents to study the effects of individual taxa and a microbial community in a precisely controlled context. Through transcriptomics analysis, we identify key genes and pathways influenced by skin microbes, and we also characterize higher-level impacts on skin processes and properties through histological analyses. Results The presence of a microbiome on a 3D skin tissue model led to significantly altered patterns of gene expression, influencing genes involved in the regulation of apoptosis, proliferation, and the extracellular matrix (among others). Moreover, microbiome treatment influenced the thickness of the epidermal layer, reduced the number of actively proliferating cells, and increased filaggrin expression. Many of these findings were evident upon treatment with the mixed community, but either not detected or less pronounced in treatments by single microorganisms, underscoring the impact that a diverse skin microbiome has on the host. Conclusions This work contributes to the understanding of how microbiome constituents individually and collectively influence human skin processes and properties. The results show that, while it is important to understand the effect of individual microbes on the host, a full community of microbes has unique and pronounced effects on the skin. Thus, in its impacts on the host, the skin microbiome is more than the sum of its parts.

Download Full-text

Draft Genome Sequence of a New Staphylococcal Species Isolated from Human Skin

Microbiology Resource Announcements ◽

10.1128/mra.01499-19 ◽

2020 ◽

Vol 9 (6) ◽

Author(s):

Kristian Stødkilde ◽

Anja Poehlein ◽

Holger Brüggemann

Keyword(s):

Human Skin ◽

Genome Sequencing ◽

Genome Sequence ◽

Draft Genome ◽

Average Nucleotide Identity ◽

Healthy Human ◽

Separate Species ◽

Content Type ◽

Staphylococcal Species ◽

Staphylococcus Haemolyticus

A hemolytic staphylococcal strain, Staphylococcus sp. strain 170179, was isolated from healthy human skin. Genome sequencing and comparison of strain 170179 to other staphylococci revealed a relatedness to Staphylococcus haemolyticus with an average nucleotide identity of 87.5%, indicating that Staphylococcus sp. 170179 belongs to a separate species.

Download Full-text

Comparison of three amplicon sequencing approaches to determine staphylococcal populations on human skin

BMC Microbiology ◽

10.1186/s12866-021-02284-1 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Charlotte Marie Ahle ◽

Kristian Stødkilde-Jørgensen ◽

Anja Poehlein ◽

Wolfgang R. Streit ◽

Jennifer Hüpeden ◽

...

Keyword(s):

Human Skin ◽

Skin Diseases ◽

Cost Effective ◽

Amplicon Sequencing ◽

Species Level ◽

Skin Microbiome ◽

Staphylococcal Species ◽

Culture Independent ◽

Skin Samples ◽

Mock Communities

Abstract Background Staphylococci are important members of the human skin microbiome. Many staphylococcal species and strains are commensals of the healthy skin microbiota, while few play essential roles in skin diseases such as atopic dermatitis. To study the involvement of staphylococci in health and disease, it is essential to determine staphylococcal populations in skin samples beyond the genus and species level. Culture-independent approaches such as amplicon next-generation sequencing (NGS) are time- and cost-effective options. However, their suitability depends on the power of resolution. Results Here we compare three amplicon NGS schemes that rely on different targets within the genes tuf and rpsK, designated tuf1, tuf2 and rpsK schemes. The schemes were tested on mock communities and on human skin samples. To obtain skin samples and build mock communities, skin swab samples of healthy volunteers were taken. In total, 254 staphylococcal strains were isolated and identified to the species level by MALDI-TOF mass spectrometry. A subset of ten strains belonging to different staphylococcal species were genome-sequenced. Two mock communities with nine and eighteen strains, respectively, as well as eight randomly selected skin samples were analysed with the three amplicon NGS methods. Our results imply that all three methods are suitable for species-level determination of staphylococcal populations. However, the novel tuf2-NGS scheme was superior in resolution power. It unambiguously allowed identification of Staphylococcus saccharolyticus and distinguish phylogenetically distinct clusters of Staphylococcus epidermidis. Conclusions Powerful amplicon NGS approaches for the detection and relative quantification of staphylococci in human samples exist that can resolve populations to the species and, to some extent, to the subspecies level. Our study highlights strengths, weaknesses and pitfalls of three currently available amplicon NGS approaches to determine staphylococcal populations. Applied to the analysis of healthy and diseased skin, these approaches can be useful to attribute host-beneficial and -detrimental roles to skin-resident staphylococcal species and subspecies.

Download Full-text