- Email: [email protected]
422 Establishing and validating a new flow cytometric method for diagnosis of Lyme borreliosis
ABSTRACTS | Inflammation, Immunity and Infection 418
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Identification of new cell surface targets on human T helper cells A Graessel1, S Hauck2, C von Toerne2, E Kloppmann3, T Goldberg3, H Koppensteiner4, M Schindler5, L Krause1, C Schmidt-Weber1 and S Blank1 1 Technische Universita¨t and Helmholtz Zentrum Mu¨nchen, Center of Allergy and Environment (ZAUM), Munich, Germany, 2 Helmholtz Zentrum Mu¨nchen, Research Unit Protein Science, Neuherberg, Germany, 3 Technische Universita¨t Mu¨nchen, Department of Informatics, Bioinformatics & Computational Biology i12, Munich, Germany, 4 Helmholtz Zentrum Mu¨nchen, Institute of Virology, Neuherberg, Germany and 5 University Clinic Tu¨bingen, Institute of Medical Virology and Epidemiology of Viral Diseases, Tu¨bingen, Germany Skin, a physical component of the immune system, together with the underlying cellular network including T cells as key players, forms an important protection compartment. Abnormal differentiation or overshooting reactions of Th cells often contribute to skin diseases. Therefore, discovery of new easy accessible therapeutic targets on T cells, such as surface proteins, would help to modulate a pathologic T cell response. Human naive CD4+ T cells were isolated from healthy blood donors and stimulated with aCD3/aCD28 in a time course experiment. Cell surface proteins were identified and quantified via PAL-qLC-MS/MS technique, a flow cytometry-based surface screening and a microarray expression analysis coupled to bioinformatics. 229 cell surface proteins were identified on human naive CD4+ T cells and more than 900 additional transcripts, encoding cell surface proteins, were detected. Comparison of the transcriptomic and proteomic level highlighted a set of 32 differentially regulated targets. Interestingly, by inspecting the background of the 229 identified cell surface proteins, 24 of them were not described to be present on the surface of naive or activated CD4+ T cells before. The combination of transcriptomic and proteomic datasets led to a large compilation of surface proteins on naive and activated CD4+ T cells, representing a rich source of proteins, which were not described on naive T cells before and which might serve as novel therapeutic targets in the context of T cell driven diseases.
RNase 7 promotes uptake and sensing of self-DNA by human keratinocytes and reduces HSV-1 infection of human keratinocytes V Kopfnagel1, K Baumert1, S Wagenknecht1, J Harder2, M Kleine3 and T Werfel1 1 Division of Immunodermatology and Allergy Research, Hannover Medical School, Hannover, Germany, 2 Department of Dermatology, University Hospital Schleswig-Holstein, Kiel, Germany and 3 PLANTON GmbH, Kiel, Germany RNase 7 is one of the major antimicrobial peptides (AMPs) secreted by keratinocytes and has been found to be upregulated in chronic inflammatory skin diseases such as atopic dermatitis and psoriasis. The keratinocyte derived AMPs hBD-2 and LL-37 have been described to promote TLR9 mediated activation of human pDCs by human self-DNA. Recently we observed a similar strong effect of RNase 7 in pDCs. Keratinocytes express several TLRs including a functional TLR9 receptor and activation of TLR9 has been shown to induce the production of the chemokine IP-10. Therefore, we investigated here the activation of keratinocytes by RNase 7 in combination with human DNA using IP-10 as experimental readout. Stimulation of keratinocytes with RNase 7 and human DNA induced a strong increase of IP10 production which was dependent on TLR9 activation. Of note, stimulation of keratinocytes with hBD-2 and LL-37 in combination with DNA failed to induce IP-10 production. The production of IP-10 was mediated by the induction of the type I interferon IFNb. We detected a fast increase of IFNb mRNA expression after stimulation with RNase 7 and DNA and IP-10 production was significantly downregulated by blocking of the interferon-a/b receptor. Furthermore, stimulation with RNase 7 and DNA led to an IFNb dependent upregulation of TLR3 and IFIT 1 mRNA expression, two proteins which are involved in antiviral defense. Importantly, pretreatment of keratinocytes with RNase 7 and DNA significantly reduced HSV1 infection of human keratinocytes. The reduction of HSV-1 infection by RNase 7 and DNA was time dependent and inhibited by blocking of TLR9 and the interferon-a/b receptor. Our study demonstrates for the first time that RNase 7 functions as an alarmin in human keratinocytes by converting self- DNA released by dying host cells into a danger signal that activates an antiviral immune response.
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Cytochrome P450s are deactivators of the aryl hydrocarbon receptor in human immune cells R Effner1, J Hiller2, S Eyerich1, C Traidl-Hoffmann2, K Brockow3, M Triggiani4, H Behrendt1, C Schmidt-Weber1 and JT Buters1 1 Center of Allergy and Environment (ZAUM), Technical University of Munich/Helmholtz Center Munich, Munich, Germany, 2 Institute of Environmental Medicine (UNIKA-T), Technical University of Munich/Helmholtz Center Munich, Munich, Germany, 3 Department of Dermatology and Allergy, Technical University of Munich, Munich, Germany and 4 Division of Allergy and Clinical Immunology, Department of Medicine, University of Salerno, Salerno, Italy The aryl hydrocarbon receptor (AhR) is increasingly recognized to have immune-modulatory functions by regulating T cell differentiation and cytokine expression. AhR also controls cytochrome P450s (CYPs) that metabolize environmental and endogenous AhR ligands. However, it is unclear whether the enzymatic CYP activity is important for immunity. The objective of the study was to investigate the impact of CYP1 activity on AhR-regulated interleukin (IL)-22, IL-17 and on the stem cell factor receptor (c-Kit) in human immune cells. Peripheral blood mononuclear cells (PBMCs) were incubated with 1-(1-propynyl)-pyrene (1PP), a suicide inhibitor for CYP1A1 in the presence of low concentrations of the AhR ligand 6formylindolo[3,2-b]carbazole (FICZ) alone or in combination with the AhR antagonist CH223191. Cytokines and c-Kit expressions were measured by qRT-PCR, ELISA and flow cytometry. Expression of xenobiotica-metabolizing enzymes and transporters in purified immune cells was determined by qRT-PCR arrays. CYP1 inhibition elevated c-Kit and IL-22 expression but down-regulated IL-17 in human PBMCs. All effects were inverted by the addition of CH-223191. Human lymphatic immune cells responded to CYP1 inhibition with c-Kit induction. To map this pathway, transcription of CYP-coding genes fingerprinted human CD4+, CD4+CD45RO+, CD8+ T cells and B cells, CD14+ cells, human primary foreskin mast cells and basophils. CYP1-mediated AhR feedback pathway was active in a range of human immune cells indicating that CYP enzymes could bridge environment and immunity via tissue regulating factors such as c-Kit and IL-22.
Immediate allergic hypersensitivity to beta-lactam antibiotics: Definition of cross-reactivity patterns using comparative molecular field analysis J Chemelle1, F Delcroix2, A Roziere3, M Vocanson3, F Hacard2, F Be´rard2, J Nicolas2, R Terreux1 and A Nosbaum3 1 Institute of Biology and Chemistry of Proteins, Lyon, France, 2 Allergy and Clinical Immunology, Lyon Sud University Hospital, Lyon, France and 3 INSERM U1111, CIRI, Lyon, France Prospective identification of cross-reactive molecules in patients with reported IgE-mediated hypersensitivity to beta-lactam (BL) antibiotics is key to establish therapeutic alternatives and tolerated BLs. In this study, we assessed the validity of molecular modelling to determine cross-reactivity patterns in BL-allergic patients. Forty-two BLs were clustered using comparative molecular field analysis (CoMFA) and 3D clustering to build a hierarchal dendrogram. Dendrogram clusters were then compared to skin test results of 171 BL-allergic patients to establish molecular cross-reactivity patterns. Six clusters of BLs were defined according to CoMFA analyses, based on the presence of side chains on the BL moiety, their hydrophilic properties and steric interactions. The correlation of BL clusters with the clinical data showed that 34 of 171 (20%) patients had positive tests to molecules from cluster 1, 151 patients (88%) to clusters 2-3, 36 patients (21%) to cluster 4 and 12 patients (7%) to cluster 6. The cluster 5 was not routinely tested. Hence, 77% of allergic patients (n¼132) turned out to be positive to one or several molecules from one unique cluster (mono cross-reactive pattern), whereas 23% of patients (n¼39) had positive tests for at least 2 different clusters (poly crossreactive pattern). Thus, for a large majority of patients, a simple evaluation of BL clusters could guide the alternative BL choice. Finally, logical algorithm determined that eight BLs were good BL cluster representatives. In conclusion, using in silico modelling, two major patterns of molecular cross-reactivity have been identified among a large series of patients with IgE-mediated hypersensitivity to BL. We propose that a simplified BL-allergy workup, testing only 8 BLs, could robustly classify patients, and facilitate the management of such reactions.
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Establishing and validating a new flow cytometric method for diagnosis of Lyme borreliosis N Bounas-Pyrros1, A Todorova1, T Biedermann2, H Hofmann2 and C Traidl-Hoffmann1 1 Chair and Institute of Environmental Medicine, UNIKA-T, Technical University of Munich and Helmholtz Zentrum Mu¨nchen, Augsburg, Germany and 2 Dept. of Dermatology and Allergy Biederstein, Technical University of Munich, Munich, Germany Serological antibody detection tests are standard diagnostic procedure for Lyme disease. Indirect detection of pathogens is performed by ELISA and Immunoblot technique. A new flow cytometric method was developed, which detects human antibodies in serum and provides quantitative results. Our goal was to test out this method and to compare the results with those of the established methods. 101 sera from patients, who presented at the Department of Dermatology and Allergy Biederstein with a clinical diagnosis of Lyme disease were analysed for Borrelia antibodies using both the Immunoblot method (Borrelia ViraStripeÒ IgG, IgM Test) and the flow cytometric method (SERION MultianalytÔ Borrelia burgdorferi IgG, IgM test). The patients were divided in three groups according to their medical history and clinical symptoms: early stage infection, late stage infection and atypical diagnoses. The results from both diagnostic methods were then statistically evaluated and compared. Overall the MultianalytÔ showed a lower sensitivity than the Immunoblot method, both in the IgG and IgM tests. Out of 74 early and late stage infection sera 67 were rated positive in the Immunoblot IgG test and 54 were classified positive by the MultianalytÔ IgG. Thus the IgG sensitivity of the MultianalytÔ was with 73% lower than that of the Immunoblot with 90.5%. Out of 39 early stage infection sera 21 were rated positive in the Immunoblot IgM and 14 were classified positive in the MultianalytÔ IgM. Regarding IgM testing the MultianalytÔ reached a lower sensitivity with 35.9%, compared to the Immunoblot with 53.9%. - The new flow cytometric method exhibited a lower sensitivity than the standardly used Immunoblot Antibody titers can be determined by the quantitative results of the MultianalytÔ - The lower sensitivity score can partially be explained by the different evaluation criteria of the MultianalytÔ
S232 Journal of Investigative Dermatology (2016), Volume 136
Isotretinoin and lymecycline treatments modify the skin microbiota in acne H Kelha¨la¨1, V Aho2, N Fyhrquist3, P Pereira2, M Kubin1, L Paulin2, R Palatsi1, P Auvinen2, K Tasanen1 and A Lauerma4 1 Department of Dermatology, University of Oulu/Oulu University Hospital, Oulu, Finland, 2 Institute of Biotechnology, DNA Sequencing and Genomics Laboratory, University of Helsinki, Helsinki, Finland, 3 Department of Bacteriology and Immunology, University of Helsinki, Helsinki, Finland and 4 Department of Dermatology, University of Helsinki/Helsinki University Central Hospital, Helsinki, Finland Oral retinoids and tetracyclines have a major role in acne treatment. Here, we report for the first time the effect of isotretinoin and lymecycline therapy on the skin microbiota in cheek, back and armpit swab samples of acne vulgaris patients using 16S ribosomal RNA (16S rRNA) gene amplicon sequencing. Propionibacterium acnes was the most common in sebaceous areas of healthy and untreated acne skin and more abundant in back than cheek samples. Five taxa including a Streptococcus differed significantly between the cheek samples of healthy controls and acne patients, and acne severity was positively correlated with the abundance of Propionibacterium. Both treatments reduced clinical acne grades and the abundance of Propionibacterium, while the abundance of several other taxa was significantly higher in treated cheek samples compared with untreated ones. Less variation was observed in back samples and none in armpit samples. There were no differences in alpha diversity between control and acne patients in any of the sampled skin areas, but the diversity of the microbiota on the cheek and the back was significantly increased after acne treatments. This study provides insight into the skin microbiota in acne and how it is modulated by systemic acne treatment.
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Identification of new cell surface targets on human T helper cells A Graessel1, S Hauck2, C von Toerne2, E Kloppmann3, T Goldberg3, H Koppensteiner4, M Schindler5, L Krause1, C Schmidt-Weber1 and S Blank1 1 Technische Universita¨t and Helmholtz Zentrum Mu¨nchen, Center of Allergy and Environment (ZAUM), Munich, Germany, 2 Helmholtz Zentrum Mu¨nchen, Research Unit Protein Science, Neuherberg, Germany, 3 Technische Universita¨t Mu¨nchen, Department of Informatics, Bioinformatics & Computational Biology i12, Munich, Germany, 4 Helmholtz Zentrum Mu¨nchen, Institute of Virology, Neuherberg, Germany and 5 University Clinic Tu¨bingen, Institute of Medical Virology and Epidemiology of Viral Diseases, Tu¨bingen, Germany Skin, a physical component of the immune system, together with the underlying cellular network including T cells as key players, forms an important protection compartment. Abnormal differentiation or overshooting reactions of Th cells often contribute to skin diseases. Therefore, discovery of new easy accessible therapeutic targets on T cells, such as surface proteins, would help to modulate a pathologic T cell response. Human naive CD4+ T cells were isolated from healthy blood donors and stimulated with aCD3/aCD28 in a time course experiment. Cell surface proteins were identified and quantified via PAL-qLC-MS/MS technique, a flow cytometry-based surface screening and a microarray expression analysis coupled to bioinformatics. 229 cell surface proteins were identified on human naive CD4+ T cells and more than 900 additional transcripts, encoding cell surface proteins, were detected. Comparison of the transcriptomic and proteomic level highlighted a set of 32 differentially regulated targets. Interestingly, by inspecting the background of the 229 identified cell surface proteins, 24 of them were not described to be present on the surface of naive or activated CD4+ T cells before. The combination of transcriptomic and proteomic datasets led to a large compilation of surface proteins on naive and activated CD4+ T cells, representing a rich source of proteins, which were not described on naive T cells before and which might serve as novel therapeutic targets in the context of T cell driven diseases.
RNase 7 promotes uptake and sensing of self-DNA by human keratinocytes and reduces HSV-1 infection of human keratinocytes V Kopfnagel1, K Baumert1, S Wagenknecht1, J Harder2, M Kleine3 and T Werfel1 1 Division of Immunodermatology and Allergy Research, Hannover Medical School, Hannover, Germany, 2 Department of Dermatology, University Hospital Schleswig-Holstein, Kiel, Germany and 3 PLANTON GmbH, Kiel, Germany RNase 7 is one of the major antimicrobial peptides (AMPs) secreted by keratinocytes and has been found to be upregulated in chronic inflammatory skin diseases such as atopic dermatitis and psoriasis. The keratinocyte derived AMPs hBD-2 and LL-37 have been described to promote TLR9 mediated activation of human pDCs by human self-DNA. Recently we observed a similar strong effect of RNase 7 in pDCs. Keratinocytes express several TLRs including a functional TLR9 receptor and activation of TLR9 has been shown to induce the production of the chemokine IP-10. Therefore, we investigated here the activation of keratinocytes by RNase 7 in combination with human DNA using IP-10 as experimental readout. Stimulation of keratinocytes with RNase 7 and human DNA induced a strong increase of IP10 production which was dependent on TLR9 activation. Of note, stimulation of keratinocytes with hBD-2 and LL-37 in combination with DNA failed to induce IP-10 production. The production of IP-10 was mediated by the induction of the type I interferon IFNb. We detected a fast increase of IFNb mRNA expression after stimulation with RNase 7 and DNA and IP-10 production was significantly downregulated by blocking of the interferon-a/b receptor. Furthermore, stimulation with RNase 7 and DNA led to an IFNb dependent upregulation of TLR3 and IFIT 1 mRNA expression, two proteins which are involved in antiviral defense. Importantly, pretreatment of keratinocytes with RNase 7 and DNA significantly reduced HSV1 infection of human keratinocytes. The reduction of HSV-1 infection by RNase 7 and DNA was time dependent and inhibited by blocking of TLR9 and the interferon-a/b receptor. Our study demonstrates for the first time that RNase 7 functions as an alarmin in human keratinocytes by converting self- DNA released by dying host cells into a danger signal that activates an antiviral immune response.
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Cytochrome P450s are deactivators of the aryl hydrocarbon receptor in human immune cells R Effner1, J Hiller2, S Eyerich1, C Traidl-Hoffmann2, K Brockow3, M Triggiani4, H Behrendt1, C Schmidt-Weber1 and JT Buters1 1 Center of Allergy and Environment (ZAUM), Technical University of Munich/Helmholtz Center Munich, Munich, Germany, 2 Institute of Environmental Medicine (UNIKA-T), Technical University of Munich/Helmholtz Center Munich, Munich, Germany, 3 Department of Dermatology and Allergy, Technical University of Munich, Munich, Germany and 4 Division of Allergy and Clinical Immunology, Department of Medicine, University of Salerno, Salerno, Italy The aryl hydrocarbon receptor (AhR) is increasingly recognized to have immune-modulatory functions by regulating T cell differentiation and cytokine expression. AhR also controls cytochrome P450s (CYPs) that metabolize environmental and endogenous AhR ligands. However, it is unclear whether the enzymatic CYP activity is important for immunity. The objective of the study was to investigate the impact of CYP1 activity on AhR-regulated interleukin (IL)-22, IL-17 and on the stem cell factor receptor (c-Kit) in human immune cells. Peripheral blood mononuclear cells (PBMCs) were incubated with 1-(1-propynyl)-pyrene (1PP), a suicide inhibitor for CYP1A1 in the presence of low concentrations of the AhR ligand 6formylindolo[3,2-b]carbazole (FICZ) alone or in combination with the AhR antagonist CH223191. Cytokines and c-Kit expressions were measured by qRT-PCR, ELISA and flow cytometry. Expression of xenobiotica-metabolizing enzymes and transporters in purified immune cells was determined by qRT-PCR arrays. CYP1 inhibition elevated c-Kit and IL-22 expression but down-regulated IL-17 in human PBMCs. All effects were inverted by the addition of CH-223191. Human lymphatic immune cells responded to CYP1 inhibition with c-Kit induction. To map this pathway, transcription of CYP-coding genes fingerprinted human CD4+, CD4+CD45RO+, CD8+ T cells and B cells, CD14+ cells, human primary foreskin mast cells and basophils. CYP1-mediated AhR feedback pathway was active in a range of human immune cells indicating that CYP enzymes could bridge environment and immunity via tissue regulating factors such as c-Kit and IL-22.
Immediate allergic hypersensitivity to beta-lactam antibiotics: Definition of cross-reactivity patterns using comparative molecular field analysis J Chemelle1, F Delcroix2, A Roziere3, M Vocanson3, F Hacard2, F Be´rard2, J Nicolas2, R Terreux1 and A Nosbaum3 1 Institute of Biology and Chemistry of Proteins, Lyon, France, 2 Allergy and Clinical Immunology, Lyon Sud University Hospital, Lyon, France and 3 INSERM U1111, CIRI, Lyon, France Prospective identification of cross-reactive molecules in patients with reported IgE-mediated hypersensitivity to beta-lactam (BL) antibiotics is key to establish therapeutic alternatives and tolerated BLs. In this study, we assessed the validity of molecular modelling to determine cross-reactivity patterns in BL-allergic patients. Forty-two BLs were clustered using comparative molecular field analysis (CoMFA) and 3D clustering to build a hierarchal dendrogram. Dendrogram clusters were then compared to skin test results of 171 BL-allergic patients to establish molecular cross-reactivity patterns. Six clusters of BLs were defined according to CoMFA analyses, based on the presence of side chains on the BL moiety, their hydrophilic properties and steric interactions. The correlation of BL clusters with the clinical data showed that 34 of 171 (20%) patients had positive tests to molecules from cluster 1, 151 patients (88%) to clusters 2-3, 36 patients (21%) to cluster 4 and 12 patients (7%) to cluster 6. The cluster 5 was not routinely tested. Hence, 77% of allergic patients (n¼132) turned out to be positive to one or several molecules from one unique cluster (mono cross-reactive pattern), whereas 23% of patients (n¼39) had positive tests for at least 2 different clusters (poly crossreactive pattern). Thus, for a large majority of patients, a simple evaluation of BL clusters could guide the alternative BL choice. Finally, logical algorithm determined that eight BLs were good BL cluster representatives. In conclusion, using in silico modelling, two major patterns of molecular cross-reactivity have been identified among a large series of patients with IgE-mediated hypersensitivity to BL. We propose that a simplified BL-allergy workup, testing only 8 BLs, could robustly classify patients, and facilitate the management of such reactions.
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Establishing and validating a new flow cytometric method for diagnosis of Lyme borreliosis N Bounas-Pyrros1, A Todorova1, T Biedermann2, H Hofmann2 and C Traidl-Hoffmann1 1 Chair and Institute of Environmental Medicine, UNIKA-T, Technical University of Munich and Helmholtz Zentrum Mu¨nchen, Augsburg, Germany and 2 Dept. of Dermatology and Allergy Biederstein, Technical University of Munich, Munich, Germany Serological antibody detection tests are standard diagnostic procedure for Lyme disease. Indirect detection of pathogens is performed by ELISA and Immunoblot technique. A new flow cytometric method was developed, which detects human antibodies in serum and provides quantitative results. Our goal was to test out this method and to compare the results with those of the established methods. 101 sera from patients, who presented at the Department of Dermatology and Allergy Biederstein with a clinical diagnosis of Lyme disease were analysed for Borrelia antibodies using both the Immunoblot method (Borrelia ViraStripeÒ IgG, IgM Test) and the flow cytometric method (SERION MultianalytÔ Borrelia burgdorferi IgG, IgM test). The patients were divided in three groups according to their medical history and clinical symptoms: early stage infection, late stage infection and atypical diagnoses. The results from both diagnostic methods were then statistically evaluated and compared. Overall the MultianalytÔ showed a lower sensitivity than the Immunoblot method, both in the IgG and IgM tests. Out of 74 early and late stage infection sera 67 were rated positive in the Immunoblot IgG test and 54 were classified positive by the MultianalytÔ IgG. Thus the IgG sensitivity of the MultianalytÔ was with 73% lower than that of the Immunoblot with 90.5%. Out of 39 early stage infection sera 21 were rated positive in the Immunoblot IgM and 14 were classified positive in the MultianalytÔ IgM. Regarding IgM testing the MultianalytÔ reached a lower sensitivity with 35.9%, compared to the Immunoblot with 53.9%. - The new flow cytometric method exhibited a lower sensitivity than the standardly used Immunoblot Antibody titers can be determined by the quantitative results of the MultianalytÔ - The lower sensitivity score can partially be explained by the different evaluation criteria of the MultianalytÔ
S232 Journal of Investigative Dermatology (2016), Volume 136
Isotretinoin and lymecycline treatments modify the skin microbiota in acne H Kelha¨la¨1, V Aho2, N Fyhrquist3, P Pereira2, M Kubin1, L Paulin2, R Palatsi1, P Auvinen2, K Tasanen1 and A Lauerma4 1 Department of Dermatology, University of Oulu/Oulu University Hospital, Oulu, Finland, 2 Institute of Biotechnology, DNA Sequencing and Genomics Laboratory, University of Helsinki, Helsinki, Finland, 3 Department of Bacteriology and Immunology, University of Helsinki, Helsinki, Finland and 4 Department of Dermatology, University of Helsinki/Helsinki University Central Hospital, Helsinki, Finland Oral retinoids and tetracyclines have a major role in acne treatment. Here, we report for the first time the effect of isotretinoin and lymecycline therapy on the skin microbiota in cheek, back and armpit swab samples of acne vulgaris patients using 16S ribosomal RNA (16S rRNA) gene amplicon sequencing. Propionibacterium acnes was the most common in sebaceous areas of healthy and untreated acne skin and more abundant in back than cheek samples. Five taxa including a Streptococcus differed significantly between the cheek samples of healthy controls and acne patients, and acne severity was positively correlated with the abundance of Propionibacterium. Both treatments reduced clinical acne grades and the abundance of Propionibacterium, while the abundance of several other taxa was significantly higher in treated cheek samples compared with untreated ones. Less variation was observed in back samples and none in armpit samples. There were no differences in alpha diversity between control and acne patients in any of the sampled skin areas, but the diversity of the microbiota on the cheek and the back was significantly increased after acne treatments. This study provides insight into the skin microbiota in acne and how it is modulated by systemic acne treatment.