Predicting Alu exonization in the human genome with a deep learning model.

Alu exonization, or the recruitment of intronic Alu elements into gene sequences, has contributed to functional diversification; however, its extent and the ways in which it influences gene regulation are not fully understood. We developed an unbiased approach to predict Alu exonization events from genomic sequences implemented in a deep learning model, eXAlu, that overcomes the limitations of tissue or condition specificity and the computational burden of RNA-seq analysis. The model captures previously reported characteristics of exonized Alu sequences and can predict sequence elements important for Alu exonization. Using eXAlu, we estimate the number of Alu elements in the human genome undergoing exonization to be between 55-110K, 11-21 fold more than represented in the GENCODE gene database. Using RT-PCR we were able to validate selected predicted Alu exonization events, supporting the accuracy of our method. Lastly, we highlight a potential application of our method to identify polymorphic Alu insertion exonizations in individuals and in the population from whole genome sequencing data.

MntJULiP and Jutils: Differential splicing analysis of RNA-seq data with covariates.

Differences in alternative splicing patterns can reveal important markers of phenotypic differentiation, including biomarkers of disease. Emerging large and complex RNA-seq datasets from disease and population studies include multiple confounders such as sex, age, ethnicity and clinical attributes, which demand highly specialized data analysis tools. However, few methods are equipped to handle the new challenges. We describe an implementation of our programs MntJULiP and Jutils for differential splicing detection and visualization from RNA-seq data that takes into account covariates. MntJULiP detects intron-level differences in alternative splicing from RNA-seq data using a Bayesian mixture model. Jutils visualizes alternative splicing variation with heatmaps, PCA and sashimi plots, and Venn diagrams. Our tools are scalable and can process thousands of samples within hours. We applied our methods to the collection of GTEx brain RNA-seq samples to deconvolute the effects of sex and age at death on the splicing patterns. In particular, clustering of covariate adjusted data identifies a subgroup of individuals undergoing a distinct splicing program during aging. MntJULiP and Jutils are implemented in Python and are available from https://github.com/splicebox/.

Regulation of human interferon signaling by transposon exonization.

Innate immune signaling is essential for clearing pathogens and damaged cells, and must be tightly regulated to avoid excessive inflammation or autoimmunity. Here, we found that the alternative splicing of exons derived from transposable elements is a key mechanism controlling immune signaling in human cells. By analyzing long-read transcriptome datasets, we identified numerous transposon exonization events predicted to generate functional protein variants of immune genes, including the type I interferon receptor IFNAR2. We demonstrated that the transposon-derived isoform of IFNAR2 is more highly expressed than the canonical isoform in almost all tissues, and functions as a decoy receptor that potently inhibits interferon signaling including in cells infected with SARS-CoV-2. Our findings uncover a primate-specific axis controlling interferon signaling and show how a transposon exonization event can be co-opted for immune regulation.

The Interaction of Borrelia burgdorferi with Human Dendritic Cells: Functional Implications.

Dendritic cells bridge the innate and adaptive immune responses by serving as sensors of infection and as the primary APCs responsible for the initiation of the T cell response against invading pathogens. The naive T cell activation requires the following three key signals to be delivered from dendritic cells: engagement of the TCR by peptide Ags bound to MHC molecules (signal 1), engagement of costimulatory molecules on both cell types (signal 2), and expression of polarizing cytokines (signal 3). Initial interactions between Borrelia burgdorferi, the causative agent of Lyme disease, and dendritic cells remain largely unexplored. To address this gap in knowledge, we cultured live B. burgdorferi with monocyte-derived dendritic cells (mo-DCs) from healthy donors to examine the bacterial immunopeptidome associated with HLA-DR. In parallel, we examined changes in the expression of key costimulatory and regulatory molecules as well as profiled the cytokines released by dendritic cells when exposed to live spirochetes. RNA-sequencing studies on B. burgdorferi-pulsed dendritic cells show a unique gene expression signature associated with B. burgdorferi stimulation that differs from stimulation with lipoteichoic acid, a TLR2 agonist. These studies revealed that exposure of mo-DCs to live B. burgdorferi drives the expression of both pro- and anti-inflammatory cytokines as well as immunoregulatory molecules (e.g., PD-L1, IDO1, Tim3). Collectively, these studies indicate that the interaction of live B. burgdorferi with mo-DCs promotes a unique mature DC phenotype that likely impacts the nature of the adaptive T cell response generated in human Lyme disease.

Interleukin-6 Drives Mitochondrial Dysregulation and Accelerates Physical Decline: Insights From an Inducible Humanized IL-6 Knock-In Mouse Model.

Chronic activation of inflammatory pathways (CI) and mitochondrial dysfunction are independently linked to age-related functional decline and early mortality. Interleukin 6 (IL-6) is among the most consistently elevated chronic activation of inflammatory pathways markers, but whether IL-6 plays a causative role in this mitochondrial dysfunction and physical deterioration remains unclear. To characterize the role of IL-6 in age-related mitochondrial dysregulation and physical decline, we have developed an inducible human IL-6 (hIL-6) knock-in mouse (TetO-hIL-6mitoQC) that also contains a mitochondrial-quality control reporter. Six weeks of hIL-6 induction resulted in upregulation of proinflammatory markers, cell proliferation and metabolic pathways, and dysregulated energy utilization. Decreased grip strength, increased falls off the treadmill, and increased frailty index were also observed. Further characterization of skeletal muscles postinduction revealed an increase in mitophagy, downregulation of mitochondrial biogenesis genes, and an overall decrease in total mitochondrial numbers. This study highlights the contribution of IL-6 to mitochondrial dysregulation and supports a causal role of hIL-6 in physical decline and frailty.

Overexpression screen of chromosome 21 genes reveals modulators of Sonic hedgehog signaling relevant to Down syndrome.

Trisomy 21 and mutations in the Sonic hedgehog (SHH) signaling pathway cause overlapping and pleiotropic phenotypes including cerebellar hypoplasia, craniofacial abnormalities, congenital heart defects and Hirschsprung disease. Trisomic cells derived from individuals with Down syndrome possess deficits in SHH signaling, suggesting that overexpression of human chromosome 21 genes may contribute to SHH-associated phenotypes by disrupting normal SHH signaling during development. However, chromosome 21 does not encode any known components of the canonical SHH pathway. Here, we sought to identify chromosome 21 genes that modulate SHH signaling by overexpressing 163 chromosome 21 cDNAs in a series of SHH-responsive mouse cell lines. We confirmed overexpression of trisomic candidate genes using RNA sequencing in the cerebella of Ts65Dn and TcMAC21 mice, model systems for Down syndrome. Our findings indicate that some human chromosome 21 genes, including DYRK1A, upregulate SHH signaling, whereas others, such as HMGN1, inhibit SHH signaling. Individual overexpression of four genes (B3GALT5, ETS2, HMGN1 and MIS18A) inhibits the SHH-dependent proliferation of primary granule cell precursors. Our study prioritizes dosage-sensitive chromosome 21 genes for future mechanistic studies. Identification of the genes that modulate SHH signaling may suggest new therapeutic avenues for ameliorating Down syndrome phenotypes.

Comprehensive and scalable quantification of splicing differences with MntJULiP.

Tools for differential splicing detection have failed to provide a comprehensive and consistent view of splicing variation. We present MntJULiP, a novel method for comprehensive and accurate quantification of splicing differences between two or more conditions. MntJULiP detects both changes in intron splicing ratios and changes in absolute splicing levels with high accuracy, and can find classes of variation overlooked by other tools. MntJULiP identifies over 29,000 differentially spliced introns in 1398 GTEx brain samples, including 11,242 novel introns discovered in this dataset. Highly scalable, MntJULiP can process thousands of samples within hours to reveal splicing constituents of phenotypic differentiation.

Expression of HLA and Autoimmune Pathway Genes in Liver Biopsies of Young Subjects With Autoimmune Hepatitis Type 1.

OBJECTIVES: To test the hypothesis that autoimmune hepatitis (AIH type I) in young subjects is due to genetic differences in proinflammatory genes responding to viral triggers in patients and controls. METHODS: Intrahepatic gene expression was compared between AIH type I (n = 24, age 9-30 years) patients (hereafter referred to as the AIH group) and controls (n = 21, age 4-25 years). RNA sequencing was performed on complementary DNA (cDNA) libraries made from total RNA extracted from formalin-fixed paraffin-embedded (FFPE) liver biopsy samples. Gene expression levels were quantified, and differentially expressed genes were functionally analyzed. Pathway analysis was performed using the databases Kyoto Encyclopedia of Genes and Genomes (KEGG) and PANTHER. The remaining sequences were mapped to the RefSeq complete set of viral genomes. RESULTS: Differential gene analysis identified 181 genes that were significantly differentially expressed (136 upregulated in the AIH group). Autoimmune pathway genes such as CD19 and CD20 which are important in B cell regulation and maturation as well as, CD8 and LY9 , which are T-cell related, were upregulated in our AIH group. Genes implicated in AIH pathogenesis including CXCL10 , which is thought to be associated with AIH severity and progression, complement genes ( C1QA, C1QB , and C1QC ), and human leucocyte antigen ( HLA ) genes ( HLA-DRB1, HLA-DRA, HLA-B , and HLA-C ) were upregulated in samples from the AIH group. Specific viral etiologies were not found. CONCLUSIONS: Unbiased next-generation sequencing and differential gene expression analysis of the AIH group has not only added support for the role of B cells in the pathogenesis and treatment of AIH but also has introduced potential new therapeutic targets: CXCL10 (anti- CXCL10 ) and several complement system-related genes.

Detection of Alu Exonization Events in Human Frontal Cortex From RNA-Seq Data.

Alu exonization events functionally diversify the transcriptome, creating alternative mRNA isoforms and accounting for an estimated 5% of the alternatively spliced (skipped) exons in the human genome. We developed computational methods, implemented into a software called Alubaster, for detecting incorporation of Alu sequences in mRNA transcripts from large scale RNA-seq data sets. The approach detects Alu sequences derived from both fixed and polymorphic Alu elements, including Alu insertions missing from the reference genome. We applied our methods to 117 GTEx human frontal cortex samples to build and characterize a collection of Alu-containing mRNAs. In particular, we detected and characterized Alu exonizations occurring at 870 fixed Alu loci, of which 237 were novel, as well as hundreds of putative events involving Alu elements that are polymorphic variants or rare alleles not present in the reference genome. These methods and annotations represent a unique and valuable resource that can be used to understand the characteristics of Alu-containing mRNAs and their tissue-specific expression patterns.

Somatic reversion impacts myelodysplastic syndromes and acute myeloid leukemia evolution in the short telomere disorders.

BACKGROUNDGermline mutations in telomerase and other telomere maintenance genes manifest in the premature aging short telomere syndromes. Myelodysplastic syndromes and acute myeloid leukemia (MDS/AML) account for 75% of associated malignancies, but how these cancers overcome the inherited telomere defect is unknown.METHODSWe used ultra-deep targeted sequencing to detect somatic reversion mutations in 17 candidate telomere lengthening genes among controls and patients with short telomere syndromes with and without MDS/AML, and we tested the functional significance of these mutations.RESULTSWhile no controls carried somatic mutations in telomere maintenance genes, 29% (16 of 56) of adults with germline telomere maintenance defects carried at least 1 (P < 0.001), and 13% (7 of 56) had 2 or more. In addition to TERT promoter mutations, which were present in 19%, another 13% of patients carried a mutation in POT1 or TERF2IP. POT1 mutations impaired telomere binding in vitro and some mutations were identical to ones seen in familial melanoma associated with longer telomere length. Exclusively in patients with germline defects in telomerase RNA (TR), we identified somatic mutations in nuclear RNA exosome genes RBM7, SKIV2L2, and DIS3, where loss-of-function upregulates mature TR levels. Somatic reversion events in 6 telomere-related genes were more prevalent in patients who were MDS/AML-free (P = 0.02, RR 4.4, 95% CI 1.2-16.7), and no patient with MDS/AML had more than 1 reversion mutation.CONCLUSIONOur data indicate that diverse adaptive somatic mutations arise in the short telomere syndromes. Their presence may alleviate the telomere crisis that promotes transformation to MDS/AML.FUNDINGThis work was supported by the NIH, the Commonwealth Foundation, the S&R Foundation Kuno Award, the Williams Foundation, the Vera and Joseph Dresner Foundation, the MacMillan Pathway to Independence Award, the American Society of Hematology Scholar Award, the Johns Hopkins Research Program for Medical Students, and the Turock Scholars Fund.

CaMKII oxidation is a critical performance/disease trade-off acquired at the dawn of vertebrate evolution.

Antagonistic pleiotropy is a foundational theory that predicts aging-related diseases are the result of evolved genetic traits conferring advantages early in life. Here we examine CaMKII, a pluripotent signaling molecule that contributes to common aging-related diseases, and find that its activation by reactive oxygen species (ROS) was acquired more than half-a-billion years ago along the vertebrate stem lineage. Functional experiments using genetically engineered mice and flies reveal ancestral vertebrates were poised to benefit from the union of ROS and CaMKII, which conferred physiological advantage by allowing ROS to increase intracellular Ca2+ and activate transcriptional programs important for exercise and immunity. Enhanced sensitivity to the adverse effects of ROS in diseases and aging is thus a trade-off for positive traits that facilitated the early and continued evolutionary success of vertebrates.

Jutils: a visualization toolkit for differential alternative splicing events.

MOTIVATION: Gene alternative splicing plays an important role in development, tissue specialization and disease and differences in splicing patterns can reveal important factors for phenotypic differentiation. While multiple computational methods exist to determine splicing differences, there is a need for user-friendly visualizations that present an intuitive view of the data and work across methods. RESULTS: We developed a toolkit, Jutils, for visualizing differential splicing events at the intron (splice junction) level. Jutils is method-agnostic, converting individual tools' output into a unified representation and using it to create visualizations. Jutils creates three types of visualizations, namely heatmaps of absolute and Z-score normalized splice ratios, sashimi plots and Venn diagrams of results from multiple comparisons. Jutils is lightweight, relying solely on the unified data file for visualizations. AVAILABILITY AND IMPLEMENTATION: Jutils is implemented in Python and is available from https://github.com/Splicebox/Jutils.

Excessive O-GlcNAcylation Causes Heart Failure and Sudden Death.

BACKGROUND: Heart failure is a leading cause of death worldwide and is associated with the rising prevalence of obesity, hypertension, and diabetes. O-GlcNAcylation (the attachment of O-linked ß-N-acetylglucosamine [O-GlcNAc] moieties to cytoplasmic, nuclear, and mitochondrial proteins) is a posttranslational modification of intracellular proteins and serves as a metabolic rheostat for cellular stress. Total levels of O-GlcNAcylation are determined by nutrient and metabolic flux, in addition to the net activity of 2 enzymes: O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). Failing myocardium is marked by increased O-GlcNAcylation, but whether excessive O-GlcNAcylation contributes to cardiomyopathy and heart failure is unknown. METHODS: We developed 2 new transgenic mouse models with myocardial overexpression of OGT and OGA to control O-GlcNAcylation independent of pathologic stress. RESULTS: We found that OGT transgenic hearts showed increased O-GlcNAcylation and developed severe dilated cardiomyopathy, ventricular arrhythmias, and premature death. In contrast, OGA transgenic hearts had lower O-GlcNAcylation but identical cardiac function to wild-type littermate controls. OGA transgenic hearts were resistant to pathologic stress induced by pressure overload with attenuated myocardial O-GlcNAcylation levels after stress and decreased pathologic hypertrophy compared with wild-type controls. Interbreeding OGT with OGA transgenic mice rescued cardiomyopathy and premature death, despite persistent elevation of myocardial OGT. Transcriptomic and functional studies revealed disrupted mitochondrial energetics with impairment of complex I activity in hearts from OGT transgenic mice. Complex I activity was rescued by OGA transgenic interbreeding, suggesting an important role for mitochondrial complex I in O-GlcNAc-mediated cardiac pathology. CONCLUSIONS: Our data provide evidence that excessive O-GlcNAcylation causes cardiomyopathy, at least in part, attributable to defective energetics. Enhanced OGA activity is well tolerated and attenuation of O-GlcNAcylation is beneficial against pressure overload-induced pathologic remodeling and heart failure. These findings suggest that attenuation of excessive O-GlcNAcylation may represent a novel therapeutic approach for cardiomyopathy.

Development of Methods to Extract RNA From Archived Pediatric Needle Liver Biopsies to Produce Sequencing Data.

ABSTRACT: Genetic susceptibility has been proposed as etiopathogenic in several pediatric liver diseases including autoimmune hepatitis (AIH). High throughput sequencing (HTPS) has been applied to archived needle liver biopsies obtained from adults but rarely to pediatric biopsies. For conclusive diagnosis of AIH, most subjects have an initial formalin-fixed paraffin embedded (FFPE) needle liver biopsy that is eventually archived and may be stored for decades. OBJECTIVE: Our goal was to develop methods to utilize tissue from archived needle liver biopsies for extraction of RNA sufficient to produce HTPS data. METHODS: We extracted total RNA from 45 FFPE needle liver biopsy samples (24 AIH type 1 patients and 21 controls [ages 15_11 and 19_10]; biopsy storage time 0.5-20 years) and constructed cDNA libraries that were then sequenced on an Illumina HiSeq2000 platform. RESULTS: Forty (89%) of the libraries produced high-quality sequences for further analyses. The average number of sequences obtained per library from HTPS was 55,136,519 (range 14,914,291-184,027,499). There was a significant inverse relationship between the number of human reads obtained and the age of the specimen (P < 2_10_7). It was possible to classify more than 90% of the reads as known genes in samples that had been stored for less than 10 years. CONCLUSIONS: Archived needle liver biopsies can be used for sequence based interrogation of the etiologic origins of complex liver diseases of young subjects, such as AIH.

Comparison of the skin microbiota in acne and rosacea.

Acne and rosacea, despite their similar clinical presentations, follow distinct clinical courses, suggesting that fundamental differences exist in their pathophysiology. We performed a case-control study profiling the skin microbiota in rosacea and acne patients compared to matched controls. Nineteen rosacea and eight acne patients were matched to controls by age ± 5 years, sex and race. DNA was extracted from facial skin swabs. The V3V4 region of the bacterial 16S rRNA gene was sequenced using Illumina MiSeq and analysed using QIIME/Metastats 2.0 software. The mean relative abundance of Cutibacterium acnes in rosacea with inflammatory papules and pustules (20.454% ±16.943%) was more similar to that of acne (19.055% ±15.469%) than that of rosacea without inflammatory papules and pustules (30.419% ±21.862%). C acnes (P = .048) and Serratia marcescens (P = .038) were significantly enriched in individuals with rosacea compared to acne. Investigating the differences between the skin microbiota in acne and rosacea can provide important clues towards understanding the disease progression in each condition.

Study of Tofacitinib in Refractory Dermatomyositis: An Open-Label Pilot Study of Ten Patients.

OBJECTIVE: This open-label 12-week study was conducted to evaluate the efficacy and safety of tofacitinib, a JAK inhibitor, in treatment-refractory active dermatomyositis (DM). METHODS: Tofacitinib in extended-release doses of 11 mg was administered daily to 10 subjects with DM. Prior to treatment, a complete washout of all steroid-sparing agents was performed. The primary outcome measure was assessment of disease activity improvement based on the International Myositis Assessment and Clinical Studies group definition of improvement. Response rate was measured as the total improvement score according to the 2016 American College of Rheumatology (ACR)/European League Against Rheumatism (EULAR) myositis response criteria. Secondary outcome measures included Cutaneous Dermatomyositis Disease Area and Severity Index (CDASI) scores, chemokine levels, immunohistochemical analysis of STAT1 expression in the skin, RNA sequencing analysis, and safety. RESULTS: At 12 weeks, the primary outcome was met in all 10 subjects. Five (50%) of 10 subjects experienced moderate improvement in disease activity, and the other 50% experienced minimal improvement according to the 2016 ACR/EULAR myositis response criteria. The secondary outcome of the mean change in the CDASI activity score over 12 weeks was statistically significant (mean ± SD 28 ± 15.4 at baseline versus 9.5 ± 8.5 at 12 weeks) (P = 0.0005). Serum chemokine levels of CXCL9/CXCL10 showed a statistically significant change from baseline. A marked decrease in STAT1 signaling in association with suppression of interferon target gene expression was demonstrated in 3 of 9 skin biopsy samples from subjects with dermatomyositis. The mean ± SD level of creatine kinase in the 10 subjects at baseline was 82 ± 34.8 IU/liter, highlighting that disease activity was predominantly located in the skin. CONCLUSION: This is the first prospective, open-label clinical trial of tofacitinib in DM that demonstrates strong clinical efficacy of a pan-JAK inhibitor, as measured by validated myositis response criteria. Future randomized controlled trials using JAK inhibitors should be considered for treating DM.

A non-mosaic transchromosomic mouse model of down syndrome carrying the long arm of human chromosome 21.

Animal models of Down syndrome (DS), trisomic for human chromosome 21 (HSA21) genes or orthologs, provide insights into better understanding and treatment options. The only existing transchromosomic (Tc) mouse DS model, Tc1, carries a HSA21 with over 50 protein coding genes (PCGs) disrupted. Tc1 is mosaic, compromising interpretation of results. Here, we "clone" the 34 MB long arm of HSA21 (HSA21q) as a mouse artificial chromosome (MAC). Through multiple steps of microcell-mediated chromosome transfer, we created a new Tc DS mouse model, Tc(HSA21q;MAC)1Yakaz ("TcMAC21"). TcMAC21 is not mosaic and contains 93% of HSA21q PCGs that are expressed and regulatable. TcMAC21 recapitulates many DS phenotypes including anomalies in heart, craniofacial skeleton and brain, molecular/cellular pathologies, and impairments in learning, memory and synaptic plasticity. TcMAC21 is the most complete genetic mouse model of DS extant and has potential for supporting a wide range of basic and preclinical research.

Chromatin structure regulates cancer-specific alternative splicing events in primary HPV-related oropharyngeal squamous cell carcinoma.

Human papillomavirus-related oropharyngeal squamous cell carcinoma (HPV+ OPSCC) represents a unique disease entity within head and neck cancer with rising incidence. Previous work has shown that alternative splicing events (ASEs) are prevalent in HPV+ OPSCC, but further validation is needed to understand the regulation of this process and its role in these tumours. In this study, eleven ASEs (GIT2, CTNNB1, MKNK2, MRPL33, SIPA1L3, SNHG6, SYCP2, TPRG1, ZHX2, ZNF331, and ELOVL1) were selected for validation from 109 previously published candidate ASEs to elucidate the post-transcriptional mechanisms of oncogenesis in HPV+ disease. In vitro qRT-PCR confirmed differential expression of 9 of 11 ASE candidates, and in silico analysis within the TCGA cohort confirmed 8 of 11 candidates. Six ASEs (MRPL33, SIPA1L3, SNHG6, TPRG1, ZHX2, and ELOVL1) showed significant differential expression across both methods. Further evaluation of chromatin modification revealed that ASEs strongly correlated with cancer-specific distribution of acetylated lysine 27 of histone 3 (H3K27ac). Subsequent epigenetic treatment of HPV+ HNSCC cell lines (UM-SCC-047 and UPCI-SCC-090) with JQ1 not only induced downregulation of cancer-specific ASE isoforms, but also growth inhibition in both cell lines. The UPCI-SCC-090 cell line, with greater ASE expression, also showed more significant growth inhibition after JQ1 treatment. This study confirms several novel cancer-specific ASEs in HPV+OPSCC and provides evidence for the role of chromatin modifications in regulation of alternative splicing in HPV+OPSCC. This highlights the role of epigenetic changes in the oncogenesis of HPV+OPSCC, which represents a unique, unexplored target for therapeutics that can alter the global post-transcriptional landscape.

Minocycline and Its Impact on Microbial Dysbiosis in the Skin and Gastrointestinal Tract of Acne Patients.

BACKGROUND: Associations between acne and gastrointestinal comorbidities suggest that microbial dysbiosis and intestinal permeability may promote inflammatory acne, a condition often managed with oral antibiotics. OBJECTIVE: We performed a case-control study to investigate the skin and gut microbiota in 8 acne patients before and after receiving oral minocycline compared to controls matched by age ±5 years, sex, and race. METHODS: DNA was extracted from stool samples and facial skin swabs. Sequencing of the V3V4 region of the bacterial 16S rRNA gene was performed using Illumina MiSeq and analyzed using QIIME/MetaStats 2.0 software. RESULTS: Acne patients included 7 female and 1 male, ages 20~32. Shannon diversity was not significantly different between the skin (p=0.153) or gut (p<0.999) microbiota of acne patients before and after antibiotics. The gut microbiota in pre-antibiotic acne patients compared to acne-free controls was depleted in probiotics Lactobacillus iners (p=0.001), Lactobacillus zeae (p=0.001), and Bifidobacterium animalis (p=0.026). After antibiotics, the gut microbiota of acne patients was depleted in Lactobacillus salivarius (p=0.001), Bifidobacterium adolescentis (p=0.002), Bifidobacterium pseudolongum (p=0.010), and Bifidobacterium breve (p=0.042), while the skin microbiota was enriched in probiotics Bifidobacterium longum (p=0.028) and Leuconostoc mesenteroides (p=0.029) and depleted in Staphylococcus epidermidis (p=0.009) and Prevotella nigrescens (p=0.028). At the phylum level, significant enrichment of Bacteroidetes in stool of acne patients following antibiotic treatment (p=0.033) led to a decreased Firmicutes to Bacteroidetes ratio. CONCLUSION: Minocycline produces significant derangements in the microbiota of the skin and gut, including many probiotic species, highlighting the potential for more targeted antimicrobial treatments for acne.