1 2273 169 EPIGENETIC REGULATION ALTERS BIOFILM ARCHITECTURE AND COMPOSITION IN MULTIPLE CLINICAL ISOLATES OF NONTYPEABLE HAEMOPHILUS INFLUENZAE. BIOFILMS PLAY A CRITICAL ROLE IN THE COLONIZATION, PERSISTENCE, AND PATHOGENESIS OF MANY HUMAN PATHOGENS. MULTIPLE MUCOSA-ASSOCIATED PATHOGENS HAVE EVOLVED A MECHANISM OF RAPID ADAPTATION, TERMED THE PHASEVARION, WHICH FACILITATES A COORDINATED REGULATION OF NUMEROUS GENES THROUGHOUT THE BACTERIAL GENOME. THIS EPIGENETIC REGULATION OCCURS VIA PHASE VARIATION OF A DNA METHYLTRANSFERASE, MOD. THE PHASEVARION OF NONTYPEABLE HAEMOPHILUS INFLUENZAE (NTHI) SIGNIFICANTLY AFFECTS THE SEVERITY OF EXPERIMENTAL OTITIS MEDIA AND REGULATES SEVERAL DISEASE-RELATED PROCESSES. HOWEVER, THE ROLE OF THE NTHI PHASEVARION IN BIOFILM FORMATION IS UNCLEAR. THE PRESENT STUDY SHOWS THAT THE PHASEVARIONS OF MULTIPLE NTHI CLINICAL ISOLATES REGULATE IN VITRO BIOFILM FORMATION UNDER DISEASE-SPECIFIC MICROENVIRONMENTAL CONDITIONS. THE IMPACT OF PHASEVARION REGULATION WAS GREATEST UNDER ALKALINE CONDITIONS THAT MIMIC THOSE KNOWN TO OCCUR IN THE MIDDLE EAR DURING DISEASE. UNDER ALKALINE CONDITIONS, NTHI STRAINS THAT EXPRESS THE MODA2 METHYLTRANSFERASE FORMED BIOFILMS WITH SIGNIFICANTLY GREATER BIOMASS AND LESS DISTINCT ARCHITECTURE THAN THOSE FORMED BY A MODA2-DEFICIENT POPULATION. THE BIOFILMS FORMED BY NTHI STRAINS THAT EXPRESS MODA2 ALSO CONTAINED LESS EXTRACELLULAR DNA (EDNA) AND SIGNIFICANTLY LESS EXTRACELLULAR HU, A DNABII DNA-BINDING PROTEIN CRITICAL FOR BIOFILM STRUCTURAL STABILITY. STABLE BIOFILM STRUCTURE IS CRITICAL FOR BACTERIAL PATHOGENESIS AND PERSISTENCE IN MULTIPLE EXPERIMENTAL MODELS OF DISEASE. THESE RESULTS IDENTIFY A ROLE FOR THE PHASEVARION IN REGULATION OF BIOFILM FORMATION, A PROCESS INTEGRAL TO THE CHRONIC NATURE OF MANY INFECTIONS. UNDERSTANDING THE ROLE OF THE PHASEVARION IN BIOFILM FORMATION IS CRITICAL TO THE DEVELOPMENT OF PREVENTION AND TREATMENT STRATEGIES FOR THESE CHRONIC DISEASES.IMPORTANCE UPPER RESPIRATORY TRACT INFECTIONS ARE THE NUMBER ONE REASON FOR A CHILD TO VISIT THE EMERGENCY DEPARTMENT, AND OTITIS MEDIA (MIDDLE EAR INFECTION) RANKS THIRD OVERALL. BIOFILMS CONTRIBUTE SIGNIFICANTLY TO THE CHRONIC NATURE OF BACTERIAL RESPIRATORY TRACT INFECTIONS, INCLUDING OTITIS MEDIA, AND MAKE THESE DISEASES PARTICULARLY DIFFICULT TO TREAT. SEVERAL MUCOSA-ASSOCIATED HUMAN PATHOGENS UTILIZE A MECHANISM OF RAPID ADAPTATION TERMED THE PHASEVARION, OR PHASEVARIABLE REGULON, TO RESIST ENVIRONMENTAL AND HOST IMMUNE PRESSURES. IN THIS STUDY, WE ASSESSED THE ROLE OF THE PHASEVARION IN REGULATION OF BIOFILM FORMATION BY NONTYPEABLE HAEMOPHILUS INFLUENZAE (NTHI), WHICH CAUSES NUMEROUS RESPIRATORY TRACT DISEASES. WE FOUND THAT THE NTHI PHASEVARION REGULATES BIOFILM STRUCTURE AND CRITICAL BIOFILM MATRIX COMPONENTS UNDER DISEASE-SPECIFIC CONDITIONS. THE FINDINGS OF THIS WORK COULD BE SIGNIFICANT IN THE DESIGN OF IMPROVED STRATEGIES AGAINST NTHI INFECTIONS, AS WELL AS DISEASES DUE TO OTHER PATHOGENS THAT UTILIZE A PHASEVARION. 2018 2 5016 40 PERSISTENT BACTERIAL COINFECTION OF A COVID-19 PATIENT CAUSED BY A GENETICALLY ADAPTED PSEUDOMONAS AERUGINOSA CHRONIC COLONIZER. PSEUDOMONAS AERUGINOSA IS A BIOFILM-FORMING OPPORTUNISTIC PATHOGEN WHICH CAUSES CHRONIC INFECTIONS IN IMMUNOCOMPROMISED PATIENTS AND LEADS TO HIGH MORTALITY RATE. IT IS IDENTIFIED AS A COMMON COINFECTING PATHOGEN IN COVID-19 PATIENTS CAUSING EXACERBATION OF ILLNESS. IN OUR HOSPITAL, P. AERUGINOSA IS ONE OF THE TOP COINFECTING BACTERIA IDENTIFIED AMONG COVID-19 PATIENTS. WE COLLECTED A STRONG BIOFILM-FORMING P. AERUGINOSA STRAIN DISPLAYING SMALL COLONY VARIANT MORPHOLOGY FROM A SEVERE COVID-19 PATIENT. GENOMIC AND TRANSCRIPTOMIC SEQUENCING ANALYSES WERE PERFORMED WITH PHENOTYPIC VALIDATION TO INVESTIGATE ITS ADAPTATION IN SARS-COV-2 INFECTED ENVIRONMENT. GENOMIC CHARACTERIZATION PREDICTED SPECIFIC GENOMIC ISLANDS HIGHLY ASSOCIATED WITH VIRULENCE, TRANSCRIPTIONAL REGULATION, AND DNA RESTRICTION-MODIFICATION SYSTEMS. EPIGENETIC ANALYSIS REVEALED A SPECIFIC N(6)-METHYL ADENINE (M(6)A) METHYLATING PATTERN INCLUDING METHYLATION OF ALGINATE, FLAGELLAR AND QUORUM SENSING ASSOCIATED GENES. DIFFERENTIAL GENE EXPRESSION ANALYSIS INDICATED THAT THIS ISOLATE FORMED EXCESSIVE BIOFILM BY REDUCING FLAGELLAR FORMATION (7.4 TO 1,624.1 FOLDS) AND OVERPRODUCING EXTRACELLULAR MATRIX COMPONENTS INCLUDING CDRA (4.4 FOLDS), ALGINATE (5.2 TO 29.1 FOLDS) AND PEL (4.8-5.5 FOLDS). IN SUMMARY, WE DEMONSTRATED THAT P. AEUGINOSA CLINICAL ISOLATES WITH NOVEL EPIGENETIC MARKERS COULD FORM EXCESSIVE BIOFILM, WHICH MIGHT ENHANCE ITS ANTIBIOTIC RESISTANCE AND IN VIVO COLONIZATION IN COVID-19 PATIENTS. 2021 3 1080 25 CLOSED COMPLETE GENOME SEQUENCES OF TWO NONTYPEABLE HAEMOPHILUS INFLUENZAE STRAINS CONTAINING NOVEL MODA ALLELES FROM THE SPUTUM OF PATIENTS WITH CHRONIC OBSTRUCTIVE PULMONARY DISEASE. NONTYPEABLE HAEMOPHILUS INFLUENZAE (NTHI) IS AN IMPORTANT BACTERIAL PATHOGEN THAT CAUSES OTITIS MEDIA AND EXACERBATIONS OF CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD). HERE, WE REPORT THE COMPLETE GENOME SEQUENCES OF NTHI STRAINS 10P129H1 AND 84P36H1, ISOLATED FROM COPD PATIENTS, WHICH CONTAIN THE PHASE-VARIABLE EPIGENETIC REGULATORS MODA15 AND MODA18, RESPECTIVELY. 2018 4 333 23 ALTERATION OF PTGS2 PROMOTER METHYLATION IN CHRONIC PERIODONTITIS. LEVELS OF PROSTAGLANDIN E(2) AND THE PROSTAGLANDIN-ENDOPEROXIDE SYNTHASE-2 (PTGS2, OR COX-2) INCREASE IN ACTIVELY PROGRESSING PERIODONTAL LESIONS, BUT DECREASE IN CHRONIC DISEASE. WE HYPOTHESIZED THAT CHRONIC INFLAMMATION IS ASSOCIATED WITH ALTERED DNA METHYLATION LEVELS WITHIN THE PTGS2 PROMOTER, WITH EFFECTS ON COX-2 MRNA EXPRESSION. PTGS2 PROMOTER METHYLATION LEVELS FROM PERIODONTALLY INFLAMED GINGIVAL BIOPSIES SHOWED A 5.06-FOLD INCREASE AS COMPARED WITH NON-INFLAMED SAMPLES (P = 0.03), AND THE ODDS OF METHYLATION IN A CPG SITE IN THE INFLAMED GINGIVAL GROUP IS 4.46 TIMES HIGHER THAN IN THE SAME SITE IN THE NON-INFLAMED GROUP (P = 0.016). THE LEVEL OF METHYLATION AT -458 BP WAS INVERSELY ASSOCIATED WITH TRANSCRIPTIONAL LEVELS OF PTGS2 (RT-PCR) (P = 0.01). ANALYSIS OF THE DATA SUGGESTS THAT, IN CHRONICALLY INFLAMED TISSUES, THERE IS A HYPERMETHYLATION PATTERN OF THE PTGS2 PROMOTER IN ASSOCIATION WITH A LOWER LEVEL OF PTGS2 TRANSCRIPTION, CONSISTENT WITH A DAMPENING OF COX-2 EXPRESSION IN CHRONIC PERIODONTITIS. THESE FINDINGS SUGGEST THAT THE CHRONIC PERSISTENCE OF THE BIOFILM AND INFLAMMATION MAY BE ASSOCIATED WITH EPIGENETIC CHANGES IN LOCAL TISSUES AT THE BIOFILM-GINGIVAL INTERFACE. 2010 5 2374 42 EPIGENETIC REGULATION OF TNFA EXPRESSION IN PERIODONTAL DISEASE. BACKGROUND: TUMOR NECROSIS FACTOR-ALPHA (TNF-ALPHA) PLAYS A CENTRAL ROLE IN THE MOLECULAR PATHOGENESIS OF PERIODONTAL DISEASE. HOWEVER, THE EPIGENETIC REGULATION ATTRIBUTABLE TO MICROBIAL AND INFLAMMATORY SIGNALS AT THE BIOFILM-GINGIVAL INTERFACE ARE POORLY UNDERSTOOD. IN THIS STUDY, THE DNA METHYLATION ALTERATION WITHIN THE TNFA PROMOTER IN HUMAN GINGIVAL BIOPSIES FROM DIFFERENT STAGES OF PERIODONTAL DISEASE IS INVESTIGATED AND THE REGULATORY MECHANISM OF TNFA TRANSCRIPTION BY DNA METHYLATION IS EXPLORED. METHODS: GINGIVAL BIOPSIES WERE OBTAINED FROM 17 PATIENTS WITH CHRONIC PERIODONTITIS (CP) AND 18 PERIODONTALLY HEALTHY INDIVIDUALS. ANOTHER 11 INDIVIDUALS PARTICIPATED IN AN EXPERIMENTALLY INDUCED GINGIVITIS STUDY, AND GINGIVAL BIOPSIES WERE COLLECTED AT THE BASELINE, INDUCTION, AND RESOLUTION PHASE. TO CONFIRM THAT TNFA PROMOTER METHYLATION MODULATED TNFA TRANSCRIPTION, THP.1 CELLS WERE TREATED WITH A DNA METHYLTRANSFERASE INHIBITOR, 5-AZA-2-DEOXYCYTIDINE (5-AZA-2DC), AND AN RAW294.7 CELL LINE TRANSFECTED WITH A TNFA PROMOTER-SPECIFIC LUCIFERASE REPORTER SYSTEM WITH OR WITHOUT METHYLATION WAS USED. RESULTS: IN GINGIVAL BIOPSIES FROM INDIVIDUALS WITH SEVERE CP, TWO INDIVIDUAL CYTOSINE-GUANINE DINUCLEOTIDES (CPG SITES) WITHIN THE TNFA PROMOTER (AT -163 AND -161 BP) DISPLAYED INCREASED METHYLATION IN CP SAMPLES COMPARED TO THOSE WITH GINGIVAL HEALTH (16.1% +/- 5.1% VERSUS 11.0% +/- 4.6%, P = 0.02 AND 19.8% +/- 4.1% VERSUS 15.4% +/- 3.6%, P = 0.04, RESPECTIVELY). THE METHYLATION LEVEL AT -163 BP WAS INVERSELY ASSOCIATED WITH THE TRANSCRIPTION LEVEL OF TNFA (P = 0.018). HOWEVER, NO SIGNIFICANT DIFFERENCE IN THE TNFA PROMOTER METHYLATION PATTERN WAS OBSERVED IN SAMPLES BIOPSIED DURING THE INDUCTION OR RESOLUTION PHASE OF EXPERIMENTALLY INDUCED GINGIVITIS, WHICH REPRESENTED A REVERSIBLE PERIODONTAL LESION. THP.1 CELLS TREATED WITH 5-AZA-2DC DEMONSTRATED A TIME-DEPENDENT INCREASE IN TNFA MESSENGER LEVEL. IT WAS ALSO FOUND THAT THE LUCIFERASE ACTIVITY DECREASED 2.6-FOLD IN A CONSTRUCT CONTAINING AN IN VITRO METHYLATED TNFA PROMOTER WHEN COMPARED TO THE UNMETHYLATED INSERT (P = 0.03). CONCLUSION: ALTHOUGH THE BIOPSY SAMPLES REPRESENTED A MIXED CELL POPULATION, THE CHANGE IN PROMOTER METHYLATION STATUS IN CHRONIC PERIODONTAL DISEASE SUGGESTED THAT DNA METHYLATION MAY BE AN IMPORTANT REGULATORY MECHANISM IN CONTROLLING TNFA TRANSCRIPTIONAL EXPRESSION IN PERIODONTAL DISEASE. 2013 6 4858 31 ORAL ANTIBIOTIC USE AND CHRONIC DISEASE: LONG-TERM HEALTH IMPACT BEYOND ANTIMICROBIAL RESISTANCE AND CLOSTRIDIOIDES DIFFICILE. WE RECENTLY REPORTED AN INCREASED COLON CANCER RISK ASSOCIATED WITH ORAL ANTIBIOTIC USE IN A LARGE UNITED KINGDOM POPULATION. THIS ASSOCIATION BETWEEN ANTIBIOTIC EXPOSURE AND CANCER RISK ADDS TO A GROWING BODY OF EVIDENCE THAT ANTIBIOTIC USE HAS UNINTENDED OFF-TARGET LONG-TERM HEALTH CONSEQUENCES. THIS ADDENDUM HIGHLIGHTS MAJOR STUDIES LINKING ANTIBIOTIC USE AND CHRONIC DISEASE IN PEDIATRIC AND ADULT POPULATIONS. MICROBIOTA DYSBIOSIS IS THE KEY PROPOSED MECHANISM UNDERLYING ANTIBIOTIC:DISEASE ASSOCIATIONS, RESULTING IN ALTERATIONS IN GENE EXPRESSION, EPIGENETIC MODIFICATION, COLONIZATION BY PATHOGENIC BACTERIA, INSTIGATION OF BIOFILMS, AND IMMUNE REGULATION AND INFLAMMATION. THESE ADVERSE OUTCOMES OF ANTIBIOTIC EXPOSURE UNDERSCORE THE NEED FOR DIAGNOSTIC AND ANTIBIOTIC STEWARDSHIP, AS WELL AS THE URGENCY FOR FURTHER DEVELOPMENT OF NON-ANTIBIOTIC THERAPIES FOR BACTERIAL INFECTIONS. 2020 7 3705 32 INFLUENCE OF EPIGENETICS ON PERIODONTITIS AND PERI-IMPLANTITIS PATHOGENESIS. PERIODONTITIS IS A DISEASE CHARACTERIZED BY TOOTH-ASSOCIATED MICROBIAL BIOFILMS THAT DRIVE CHRONIC INFLAMMATION AND DESTRUCTION OF PERIODONTAL-SUPPORTING TISSUES. IN SOME INDIVIDUALS, DISEASE PROGRESSION CAN LEAD TO TOOTH LOSS. A SIMILAR CONDITION CAN OCCUR AROUND DENTAL IMPLANTS IN THE FORM OF PERI-IMPLANTITIS. THE IMMUNE RESPONSE TO BACTERIAL CHALLENGES IS NOT ONLY INFLUENCED BY GENETIC FACTORS, BUT ALSO BY ENVIRONMENTAL FACTORS. EPIGENETICS INVOLVES THE STUDY OF GENE FUNCTION INDEPENDENT OF CHANGES TO THE DNA SEQUENCE AND ITS ASSOCIATED PROTEINS, AND REPRESENTS A CRITICAL LINK BETWEEN GENETIC AND ENVIRONMENTAL FACTORS. EPIGENETIC MODIFICATIONS HAVE BEEN SHOWN TO CONTRIBUTE TO THE PROGRESSION OF SEVERAL DISEASES, INCLUDING CHRONIC INFLAMMATORY DISEASES LIKE PERIODONTITIS AND PERI-IMPLANTITIS. THIS REVIEW AIMS TO PRESENT THE LATEST FINDINGS ON EPIGENETIC INFLUENCES ON PERIODONTITIS AND TO DISCUSS POTENTIAL MECHANISMS THAT MAY INFLUENCE PERI-IMPLANTITIS, GIVEN THE PAUCITY OF INFORMATION CURRENTLY AVAILABLE. 2022 8 4718 45 NON-TYPEABLE HAEMOPHILUS INFLUENZAE ISOLATES FROM PATIENTS WITH CHRONIC OBSTRUCTIVE PULMONARY DISEASE CONTAIN NEW PHASE-VARIABLE MODA METHYLTRANSFERASE ALLELES CONTROLLING PHASEVARIONS. PHASEVARIONS (PHASE-VARIABLE REGULONS) ARE EMERGING AS AN IMPORTANT AREA OF BACTERIAL GENE REGULATION. MANY BACTERIAL PATHOGENS CONTAIN PHASEVARIONS, WITH GENE EXPRESSION CONTROLLED BY THE PHASE-VARIABLE EXPRESSION OF DNA METHYLTRANSFERASES VIA EPIGENETIC MECHANISMS. NON-TYPEABLE HAEMOPHILUS INFLUENZAE (NTHI) CONTAINS THE PHASE-VARIABLE METHYLTRANSFERASE MODA, OF WHICH MULTIPLE ALLELIC VARIANTS EXIST (MODA1-21). WE HAVE PREVIOUSLY DEMONSTRATED 5 OF 21 THESE MODA ALLELES ARE OVERREPRESENTED IN NTHI STRAINS ISOLATED FROM CHILDREN WITH MIDDLE EAR INFECTIONS. IN THIS STUDY WE INVESTIGATED THE MODA ALLELE DISTRIBUTION IN NTHI STRAINS ISOLATED FROM PATIENTS WITH CHRONIC OBSTRUCTIVE PULMONARY DISEASE, COPD. WE DEMONSTRATE THAT THE DISTRIBUTION OF MODA ALLELES IN A LARGE PANEL OF COPD ISOLATES IS DIFFERENT TO THE DISTRIBUTION SEEN IN MIDDLE EAR INFECTIONS, SUGGESTING DIFFERENT MODA ALLELES MAY PROVIDE DISTINCT ADVANTAGES IN THE DIFFERING NICHES OF THE MIDDLE EAR AND COPD AIRWAYS. WE ALSO IDENTIFIED TWO NEW PHASE-VARIABLE MODA ALLELES - MODA15 AND MODA18 - AND DEMONSTRATE THAT THESE ALLELES METHYLATE DISTINCT DNA SEQUENCES AND CONTROL UNIQUE PHASEVARIONS. THE MODA15 AND MODA18 ALLELES HAVE ONLY BEEN OBSERVED IN COPD ISOLATES, INDICATING THAT THESE TWO ALLELES MAY BE MARKERS FOR ISOLATES LIKELY TO CAUSE EXACERBATIONS OF COPD. 2019 9 4395 43 MODM DNA METHYLTRANSFERASE METHYLOME ANALYSIS REVEALS A POTENTIAL ROLE FOR MORAXELLA CATARRHALIS PHASEVARIONS IN OTITIS MEDIA. MORAXELLA CATARRHALIS IS A SIGNIFICANT CAUSE OF OTITIS MEDIA AND EXACERBATIONS OF CHRONIC OBSTRUCTIVE PULMONARY DISEASE. HERE, WE CHARACTERIZE A PHASE-VARIABLE DNA METHYLTRANSFERASE (MODM), WHICH CONTAINS 5'-CAAC-3' REPEATS IN ITS OPEN READING FRAME THAT MEDIATE HIGH-FREQUENCY MUTATION RESULTING IN REVERSIBLE ON/OFF SWITCHING OF MODM EXPRESSION. THREE MODM ALLELES HAVE BEEN IDENTIFIED (MODM1-3), WITH MODM2 BEING THE MOST COMMONLY FOUND ALLELE. USING SINGLE-MOLECULE, REAL-TIME (SMRT) GENOME SEQUENCING AND METHYLOME ANALYSIS, WE HAVE DETERMINED THAT THE MODM2 METHYLATION TARGET IS 5'-GAR(M6)AC-3', AND 100% OF THESE SITES ARE METHYLATED IN THE GENOME OF THE M. CATARRHALIS 25239 MODM2 ON STRAIN. PROTEOMIC ANALYSIS OF MODM2 ON AND OFF VARIANTS REVEALED THAT MODM2 REGULATES EXPRESSION OF MULTIPLE GENES THAT HAVE POTENTIAL ROLES IN COLONIZATION, INFECTION, AND PROTECTION AGAINST HOST DEFENSES. INVESTIGATION OF THE DISTRIBUTION OF MODM ALLELES IN A PANEL OF M. CATARRHALIS STRAINS, ISOLATED FROM THE NASOPHARYNX OF HEALTHY CHILDREN OR MIDDLE EAR EFFUSIONS FROM PATIENTS WITH OTITIS MEDIA, REVEALED A STATISTICALLY SIGNIFICANT ASSOCIATION OF MODM3 WITH OTITIS MEDIA ISOLATES. THE MODULATION OF GENE EXPRESSION VIA THE MODM PHASE-VARIABLE REGULON (PHASEVARION), AND THE SIGNIFICANT ASSOCIATION OF THE MODM3 ALLELE WITH OTITIS MEDIA, SUGGESTS A KEY ROLE FOR MODM PHASEVARIONS IN THE PATHOGENESIS OF THIS ORGANISM. 2014 10 6261 49 THE MORAXELLA CATARRHALIS PHASE-VARIABLE DNA METHYLTRANSFERASE MODM3 IS AN EPIGENETIC REGULATOR THAT AFFECTS BACTERIAL SURVIVAL IN AN IN VIVO MODEL OF OTITIS MEDIA. BACKGROUND: MORAXELLA CATARRHALIS IS A LEADING CAUSE OF OTITIS MEDIA (OM) AND CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD). M. CATARRHALIS CONTAINS A TYPE III DNA ADENINE METHYLTRANSFERASE (MODM) THAT IS PHASE-VARIABLY EXPRESSED (I.E., ITS EXPRESSION IS SUBJECT TO RANDOM, REVERSIBLE ON/OFF SWITCHING). MODM HAS SIX TARGET RECOGNITION DOMAIN ALLELES (MODM1-6), AND WE HAVE PREVIOUSLY SHOWN THAT MODM2 IS THE PREDOMINANT ALLELE, WHILE MODM3 IS ASSOCIATED WITH OM. PHASE-VARIABLE DNA METHYLTRANSFERASES MEDIATE EPIGENETIC REGULATION AND MODULATE PATHOGENESIS IN SEVERAL BACTERIA. MODM2 OF M. CATARRHALIS REGULATES THE EXPRESSION OF A PHASEVARION CONTAINING GENES IMPORTANT FOR COLONIZATION AND INFECTION. HERE WE DESCRIBE THE PHASE-VARIABLE EXPRESSION OF MODM3, THE MODM3 METHYLATION SITE AND THE SUITE OF GENES REGULATED WITHIN THE MODM3 PHASEVARION. RESULTS: PHASE-VARIABLE EXPRESSION OF MODM3, MEDIATED BY VARIATION IN LENGTH OF A 5'-(CAAC)(N)-3' TETRANUCLEOTIDE REPEAT TRACT IN THE OPEN READING FRAME WAS DEMONSTRATED IN M. CATARRHALIS STRAIN CCRI-195ME WITH GENESCAN FRAGMENT LENGTH ANALYSIS AND WESTERN IMMUNOBLOT. WE DETERMINED THAT MODM3 IS AN ACTIVE N6-ADENINE METHYLTRANSFERASE THAT METHYLATES THE SEQUENCE 5'-AC(M6)ATC-3'. METHYLATION WAS DETECTED AT ALL 4446 5'-ACATC-3' SITES IN THE GENOME WHEN MODM3 IS EXPRESSED. RNASEQ ANALYSIS IDENTIFIED 31 GENES THAT ARE DIFFERENTIALLY EXPRESSED BETWEEN MODM3 ON AND OFF VARIANTS, INCLUDING FIVE GENES THAT ARE INVOLVED IN THE RESPONSE TO OXIDATIVE AND NITROSATIVE STRESS, WITH POTENTIAL ROLES IN BIOFILM FORMATION AND SURVIVAL IN ANAEROBIC ENVIRONMENTS. AN IN VIVO CHINCHILLA (CHINCHILLA LANIGERA) MODEL OF OTITIS MEDIA DEMONSTRATED THAT TRANSBULLAR CHALLENGE WITH THE MODM3 OFF VARIANT RESULTED IN AN INCREASED MIDDLE EAR BACTERIAL LOAD COMPARED TO A MODM3 ON VARIANT. IN ADDITION, CO-INFECTION EXPERIMENTS WITH NTHI AND M. CATARRHALIS MODM3 ON OR MODM3 OFF VARIANTS REVEALED THAT PHASE VARIATION OF MODM3 ALTERED SURVIVAL OF NTHI IN THE MIDDLE EAR DURING EARLY AND LATE STAGE INFECTION. CONCLUSIONS: PHASE VARIATION OF MODM3 EPIGENETICALLY REGULATES THE EXPRESSION OF A PHASEVARION CONTAINING MULTIPLE GENES THAT ARE POTENTIALLY IMPORTANT IN THE PROGRESSION OF OTITIS MEDIA. 2019 11 3235 40 HEMIN AVAILABILITY INDUCES COORDINATED DNA METHYLATION AND GENE EXPRESSION CHANGES IN PORPHYROMONAS GINGIVALIS. PERIODONTAL DISEASE IS A CHRONIC INFLAMMATORY DISEASE IN WHICH THE ORAL PATHOGEN PORPHYROMONAS GINGIVALIS PLAYS AN IMPORTANT ROLE. PORPHYROMONAS GINGIVALIS EXPRESSES VIRULENCE DETERMINANTS IN RESPONSE TO HIGHER HEMIN CONCENTRATIONS, BUT THE UNDERLYING REGULATORY PROCESSES REMAIN UNCLEAR. BACTERIAL DNA METHYLATION HAS THE POTENTIAL TO FULFIL THIS MECHANISTIC ROLE. WE CHARACTERIZED THE METHYLOME OF P. GINGIVALIS, AND COMPARED ITS VARIATION TO TRANSCRIPTOME CHANGES IN RESPONSE TO HEMIN AVAILABILITY. PORPHYROMONAS GINGIVALIS W50 WAS GROWN IN CHEMOSTAT CONTINUOUS CULTURE WITH EXCESS OR LIMITED HEMIN, PRIOR TO WHOLE-METHYLOME AND TRANSCRIPTOME PROFILING USING NANOPORE AND ILLUMINA RNA-SEQ. DNA METHYLATION WAS QUANTIFIED FOR DAM/DCM MOTIFS AND ALL-CONTEXT N6-METHYLADENINE (6MA) AND 5-METHYLCYTOSINE (5MC). OF ALL 1,992 GENES ANALYZED, 161 AND 268 WERE RESPECTIVELY OVER- AND UNDER-EXPRESSED WITH EXCESS HEMIN. NOTABLY, WE DETECTED DIFFERENTIAL DNA METHYLATION SIGNATURES FOR THE DAM "GATC" MOTIF AND BOTH ALL-CONTEXT 6MA AND 5MC IN RESPONSE TO HEMIN AVAILABILITY. JOINT ANALYSES IDENTIFIED A SUBSET OF COORDINATED CHANGES IN GENE EXPRESSION, 6MA, AND 5MC METHYLATION THAT TARGET GENES INVOLVED IN LACTATE UTILIZATION AND ABC TRANSPORTERS. THE RESULTS IDENTIFY ALTERED METHYLATION AND EXPRESSION RESPONSES TO HEMIN AVAILABILITY IN P. GINGIVALIS, WITH INSIGHTS INTO MECHANISMS REGULATING ITS VIRULENCE IN PERIODONTAL DISEASE. IMPORTANCE DNA METHYLATION HAS IMPORTANT ROLES IN BACTERIA, INCLUDING IN THE REGULATION OF TRANSCRIPTION. PORPHYROMONAS GINGIVALIS, AN ORAL PATHOGEN IN PERIODONTITIS, EXHIBITS WELL-ESTABLISHED GENE EXPRESSION CHANGES IN RESPONSE TO HEMIN AVAILABILITY. HOWEVER, THE REGULATORY PROCESSES UNDERLYING THESE EFFECTS REMAIN UNKNOWN. WE PROFILED THE NOVEL P. GINGIVALIS EPIGENOME, AND ASSESSED EPIGENETIC AND TRANSCRIPTOME VARIATION UNDER LIMITED AND EXCESS HEMIN CONDITIONS. AS EXPECTED, MULTIPLE GENE EXPRESSION CHANGES WERE DETECTED IN RESPONSE TO LIMITED AND EXCESS HEMIN THAT REFLECT HEALTH AND DISEASE, RESPECTIVELY. NOTABLY, WE ALSO DETECTED DIFFERENTIAL DNA METHYLATION SIGNATURES FOR THE DAM "GATC" MOTIF AND BOTH ALL-CONTEXT 6MA AND 5MC IN RESPONSE TO HEMIN. JOINT ANALYSES IDENTIFIED COORDINATED CHANGES IN GENE EXPRESSION, 6MA, AND 5MC METHYLATION THAT TARGET GENES INVOLVED IN LACTATE UTILIZATION AND ABC TRANSPORTERS. THE RESULTS IDENTIFY NOVEL REGULATORY PROCESSES UNDERLYING THE MECHANISM OF HEMIN REGULATED GENE EXPRESSION IN P. GINGIVALIS, WITH PHENOTYPIC IMPACTS ON ITS VIRULENCE IN PERIODONTAL DISEASE. 2023 12 4494 43 MORAXELLA CATARRHALIS RESTRICTION-MODIFICATION SYSTEMS ARE ASSOCIATED WITH PHYLOGENETIC LINEAGE AND DISEASE. MORAXELLA CATARRHALIS IS A HUMAN-ADAPTED PATHOGEN, AND A MAJOR CAUSE OF OTITIS MEDIA (OM) AND EXACERBATIONS OF CHRONIC OBSTRUCTIVE PULMONARY DISEASE. THE SPECIES IS COMPRISED OF TWO MAIN PHYLOGENETIC LINEAGES, RB1 AND RB2/3. RESTRICTION-MODIFICATION (R-M) SYSTEMS ARE AMONG THE FEW LINEAGE-ASSOCIATED GENES IDENTIFIED IN OTHER BACTERIAL GENERA AND HAVE MULTIPLE FUNCTIONS INCLUDING DEFENSE AGAINST FOREIGN INVADING DNA, MAINTENANCE OF SPECIATION, AND EPIGENETIC REGULATION OF GENE EXPRESSION. HERE, WE DEFINE THE REPERTOIRE OF R-M SYSTEMS IN 51 PUBLICLY AVAILABLE M. CATARRHALIS GENOMES AND REPORT THEIR DISTRIBUTION AMONG M. CATARRHALIS PHYLOGENETIC LINEAGES. AN ASSOCIATION WITH PHYLOGENETIC LINEAGE (RB1 OR RB2/3) WAS OBSERVED FOR SIX R-M SYSTEMS, WHICH MAY CONTRIBUTE TO THE EVOLUTION OF THE LINEAGES BY RESTRICTING DNA TRANSFORMATION. IN ADDITION, WE OBSERVED A RELATIONSHIP BETWEEN A MUTUALLY EXCLUSIVE TYPE I R-M SYSTEM AND A TYPE III R-M SYSTEM AT A SINGLE LOCUS CONSERVED THROUGHOUT A GEOGRAPHICALLY AND CLINICALLY DIVERSE SET OF M. CATARRHALIS ISOLATES. THE TYPE III R-M SYSTEM AT THIS LOCUS CONTAINS THE PHASE-VARIABLE TYPE III DNA METHYLTRANSFERASE, MODM, WHICH CONTROLS A PHASEVARION (PHASE-VARIABLE REGULON). WE OBSERVED AN ASSOCIATION BETWEEN MODM PRESENCE AND OM-ASSOCIATED MIDDLE EAR ISOLATES, INDICATING A POTENTIAL ROLE FOR MODM-MEDIATED EPIGENETIC REGULATION IN OM PATHOBIOLOGY. 2018 13 6332 31 THE ROLE OF CYTOMEGALOVIRUS INFECTION IN THE PATHOGENESIS OF PERIODONTAL DISEASES. THE MAIN CHARACTERISTIC OFPERIODONTAL DISEASE IS CHRONIC INFLAMMATION THAT LEADS TO PROGRESSIVE DESTRUCTION OF THE CONNECTIVE TISSUES AND BONE WITH SUBSEQUENT TOOTH MOBILITY AND FINALLY TOOTH LOSS. TRADITIONALLY, THE PATHOGENESIS OF PERIODONTITIS WAS BASED ON THE INFECTION CAUSED BY BACTERIA THAT COLONIZE TOOTH SURFACE AND GINGIVAL SULCUS. ACCUMULATED EVIDENCE SHOW THAT HOST RESPONSE FACTORS SUCH AS INFLAMMATORY REACTION AND ACTIVATION OF THE INNATE IMMUNE SYSTEM ARE CRITICAL TO THE PATHOGENESIS OF PERIODONTAL DISEASE. PERIODONTAL DISEASE HAS BEEN WIDELY RECOGNIZED AS A CHRONIC DISEASE BUT THE NATURE OF CHRONICITY REMAINS UNCLEAR. THE QUESTION IS WHETHER PERIODONTAL DISEASE IS A CONTINUOUS PROCESS OR CONSISTS OF EPISODES OF EXACERBATIONS AND REMISSIONS. MAYBE CYTOMEGALOVIRUS INFECTION OF THE PERIODONTIUM, DEPENDING ON THE LATENT OR ACTIVE PHASE OF INFECTION, CAN PARTLY EXPLAIN THE EPISODIC PROGRESSIVE NATURE OF PERIODONTAL DISEASE. CYTOMEGALOVIRUS INFECTION IMPAIRS PERIODONTAL DEFENSE AND PERMITS OVERGROWTH OF PERIODONTOPATHOGENIC BACTERIA. OWING TO ADVANCES IN NEW TECHNOLOGIES, EXPERIMENTAL EVIDENCE SHOW THE INFLUENCE AND INTERRELATEDNESS OF GENOMIC, EPIGENETIC, PROTEOMIC AND METABOLIC FACTORS IN THE PATHOGENESIS OF PERIODONTAL DISEASE. DATA ON THE PATHOGENESIS OF PERIODONTAL DISEASE ARE REVIEWED. 2011 14 2762 23 EXPRESSION OF TET2 ENZYME INDICATES ENHANCED EPIGENETIC MODIFICATION OF CELLS IN PERIODONTITIS. DNA METHYLATION IS AN IMPORTANT EPIGENETIC MECHANISM INVOLVED IN THE REGULATION OF GENE EXPRESSION, AND A REDUCTION IN DNA METHYLATION INFLUENCES CELL-CYCLE PROGRESSION AND CELL DIFFERENTIATION IN INFLAMMATORY CELLS. THE AIM OF THE PRESENT STUDY WAS TO ANALYZE THE DNA-METHYLATION PATTERN AT LOCAL AND GLOBAL/SYSTEMIC LEVELS IN PATIENTS WITH PERIODONTITIS AND GINGIVITIS. TWENTY-ONE SUBJECTS WITH GENERALIZED, SEVERE PERIODONTITIS AND 17 SUBJECTS WITH GINGIVAL INFLAMMATION BUT NO ATTACHMENT LOSS WERE RECRUITED. GINGIVAL BIOPSIES AND PERIPHERAL BLOOD SAMPLES WERE COLLECTED AND PREPARED FOR IMMUNOHISTOCHEMICAL ANALYSIS OF 5-METHYLCYTOSINE (5MC), 5-HYDROXYMETHYLCYTOSINE (5HMC), TEN-ELEVEN TRANSLOCATION 2 (TET2), AND DNA METHYLTRANSFERASE 1 (DNMT1). WHILST A SIMILAR PATTERN FOR 5MC AND 5HMC DNA METHYLATION WAS FOUND IN BOTH TYPES OF LESIONS, A SIGNIFICANTLY LARGER PROPORTION OF TET2-POSITIVE CELLS WAS FOUND IN PERIODONTITIS LESIONS THAN IN GINGIVITIS LESIONS. QUANTITATIVE REAL-TIME PCR ANALYSIS SHOWED NO DIFFERENCES BETWEEN GINGIVITIS AND PERIODONTITIS LESIONS REGARDING EXPRESSION OF TET2 AND ISOCITRATE DEHYDROGENASE (IDH) GENES, WHILE THE GLOBAL LEVEL OF 5HMC WAS SIGNIFICANTLY HIGHER IN BLOOD THAN IN TISSUE IN PATIENTS WITH PERIODONTITIS. IT IS SUGGESTED THAT EPIGENETIC CHANGES ARE MORE COMMON IN PERIODONTITIS LESIONS THAN IN GINGIVITIS LESIONS AND THAT SUCH CHANGES ARE TISSUE SPECIFIC. 2016 15 1124 21 COMPLETE GENOME SEQUENCE OF MORAXELLA CATARRHALIS STRAIN CCRI-195ME, ISOLATED FROM THE MIDDLE EAR. MORAXELLA CATARRHALIS IS AN IMPORTANT BACTERIAL PATHOGEN THAT CAUSES OTITIS MEDIA AND EXACERBATIONS OF CHRONIC OBSTRUCTIVE PULMONARY DISEASE. HERE, WE REPORT THE COMPLETE GENOME SEQUENCE OF M. CATARRHALIS STRAIN CCRI-195ME, WHICH CONTAINS THE PHASE-VARIABLE EPIGENETIC REGULATOR MODM3. 2017 16 3440 26 HYPERMETHYLATION AND LOW TRANSCRIPTION OF TLR2 GENE IN CHRONIC PERIODONTITIS. PERIODONTITIS IS AN INFLAMMATORY DISORDER CHARACTERIZED BY INTERACTIONS BETWEEN PERIODONTAL PATHOGENS AND HOST'S IMMUNE RESPONSE. EPIGENETIC MAY CONTRIBUTE TO DISEASE DEVELOPMENT AND OUTCOME BY INFLUENCING THE EXPRESSION OF GENES INVOLVED IN THE IMMUNE RESPONSE. IT HAS BEEN SHOWN THAT TOLL-LIKE RECEPTORS (TLR) PLAY AN IMPORTANT ROLE IN THE RESPONSE TO PERIODONTOPATHIC BACTERIA. THE AIM OF STUDY WAS TO EVALUATE THE METHYLATION STATUS AND THE EXPRESSION OF TLR2 GENE IN GINGIVAL SAMPLES FROM INDIVIDUALS WITH AND WITHOUT PERIODONTITIS. DNA WAS ANALYZED USING THE METHYL PROFILER DNA METHYLATION QPCR ASSAY. DNA METHYLATION AND TRANSCRIPT LEVELS WERE EVALUATED BY REAL-TIME POLYMERASE CHAIN REACTION. THE PERIODONTITIS GROUP SHOWED A HYPERMETHYLATED PROFILE AND A LOW EXPRESSION OF GENE. POSITIVE CORRELATION BETWEEN THE TLR2 METHYLATION FREQUENCY AND PROBING DEPTH WAS OBSERVED. THIS STUDY GIVES THE FIRST EVIDENCE OF METHYLATION FREQUENCY IN INFLAMED PERIODONTAL TISSUES AND OF THE POSSIBLE PARTICIPATION OF METHYLATION IN THE DEVELOPMENT OF PERIODONTITIS. 2013 17 5375 18 RECENT ADVANCES ON POSSIBLE ASSOCIATION BETWEEN THE PERIODONTAL INFECTION OF PORPHYROMONAS GINGIVALIS AND CENTRAL NERVOUS SYSTEM INJURY. CHRONIC PERIODONTITIS CAUSED BY PORPHYROMONAS GINGIVALIS (P. GINGIVALIS) INFECTION GENERALLY LASTS FOR A LIFETIME. THE LONG-TERM EXISTENCE AND DEVELOPMENT OF P. GINGIVALIS INFECTION GRADUALLY AGGRAVATE THE ACCUMULATION OF INFLAMMATORY SIGNALS AND TOXIC SUBSTANCES IN THE BODY. RECENT EVIDENCE HAS REVEALED THAT P. GINGIVALIS INFECTION MAY BE RELEVANT TO SOME CENTRAL NERVOUS SYSTEM (CNS) DISEASES. THE CURRENT WORK COLLECTS INFORMATION AND TRIES TO EXPLORE THE POSSIBLE RELATIONSHIP BETWEEN P. GINGIVALIS INFECTION AND CNS DISEASES, INCLUDING THE INTERACTION OR PATHWAYS BETWEEN PERIPHERAL INFECTION AND CNS INJURY, AND THE UNDERLYING NEUROTOXIC MECHANISMS. 2021 18 6795 14 [EFFECT OF HISTONE ACETYLATION ON OSTEOGENIC DIFFERENTIATION OF PERIODONTAL LIGAMENT STEM CELLS DERIVED FROM PERIODONTITIS TISSUE]. EPIGENETICS IS DEFINED AS A CHANGE IN GENE EXPRESSION WITHOUT THE ALTERATION OF THE GENETIC SEQUENCE. SUCH A CHANGE WOULD BE INHERITED BY OFFSPRING. HISTONE ACETYLATION IS A TYPE OF EPIGENETICS. EXISTING STUDIES PROPOSED THAT CHRONIC PERIODONTITIS IS RELATED TO EPIGENETIC MODIFICATION. IN THIS REVIEW, WE SUMMARISED THE INFLUENCE OF CHRONIC PERIODONTITIS ON PERIODONTAL LIGAMENT STEM CELLS BY HISTONE ACETYLATION. 2019 19 3783 24 INTERFERON-GAMMA PROMOTER HYPOMETHYLATION AND INCREASED EXPRESSION IN CHRONIC PERIODONTITIS. AIM: THE GOAL OF THIS INVESTIGATION WAS TO DETERMINE WHETHER EPIGENETIC MODIFICATIONS IN THE IFNG PROMOTER ARE ASSOCIATED WITH AN INCREASE OF IFNG TRANSCRIPTION IN DIFFERENT STAGES OF PERIODONTAL DISEASES. MATERIALS AND METHODS: DNA WAS EXTRACTED FROM GINGIVAL BIOPSY SAMPLES COLLECTED FROM 47 TOTAL SITES FROM 47 DIFFERENT SUBJECTS: 23 PERIODONTALLY HEALTHY SITES, 12 EXPERIMENTALLY INDUCED GINGIVITIS SITES AND 12 CHRONIC PERIODONTITIS SITES. LEVELS OF DNA METHYLATION WITHIN THE IFNG PROMOTER CONTAINING SIX CPG DINUCLEOTIDES WERE DETERMINED USING PYROSEQUENCING TECHNOLOGY. INTERFERON GAMMA MRNA EXPRESSION WAS ANALYSED BY QUANTITATIVE POLYMERASE CHAIN REACTIONS USING ISOLATED RNA FROM PART OF THE BIOLOGICAL SAMPLES MENTIONED ABOVE. RESULTS: THE METHYLATION LEVEL OF ALL SIX ANALYSED CPG SITES WITHIN THE IFNG PROMOTER REGION IN THE PERIODONTITIS BIOPSIES 52% [INTERQUARTILE RANGE, IQR (43.8%, 63%)] WAS SIGNIFICANTLY LOWER THAN PERIODONTALLY HEALTHY SAMPLES 62% [IQR (51.3%, 74%)], P=0.007 AND GINGIVITIS BIOPSIES 63% [IQR (55%, 74%)], P=0.02. THE TRANSCRIPTIONAL LEVEL OF IFNG IN PERIODONTITIS BIOPSIES WAS 1.96-FOLD AND SIGNIFICANTLY HIGHER THAN TISSUES WITH PERIODONTAL HEALTH (P=0.04). ALTHOUGH THE MRNA LEVEL FROM EXPERIMENTAL GINGIVITIS SAMPLES EXHIBITED AN 8.5-FOLD INCREASE AS COMPARED WITH PERIODONTALLY HEALTHY SAMPLES, NO SIGNIFICANT METHYLATION DIFFERENCE WAS OBSERVED IN EXPERIMENTAL GINGIVITIS SAMPLE. CONCLUSIONS: A HYPOMETHYLATION PROFILE WITHIN IFNG PROMOTER REGION IS RELATED TO AN INCREASE OF IFNG TRANSCRIPTION PRESENT IN THE CHRONIC PERIODONTITIS BIOPSIES, WHILE SUCH AN INCREASE OF IFNG IN EXPERIMENTALLY INDUCED GINGIVITIS SEEMS INDEPENDENT OF PROMOTER METHYLATION ALTERATION. 2010 20 1619 59 DNA METHYLTRANSFERASE REGULATES NITRIC OXIDE HOMEOSTASIS AND VIRULENCE IN A CHRONICALLY ADAPTED PSEUDOMONAS AERUGINOSA STRAIN. OPPORTUNISTIC PATHOGENS SUCH AS PSEUDOMONAS AERUGINOSA ADAPT THEIR GENOMES RAPIDLY DURING CHRONIC INFECTIONS. UNDERSTANDING THEIR EPIGENETIC REGULATION MAY PROVIDE BIOMARKERS FOR DIAGNOSIS AND REVEAL NOVEL REGULATORY MECHANISMS. WE PERFORMED SINGLE-MOLECULE REAL-TIME SEQUENCING (SMRT-SEQ) TO CHARACTERIZE THE METHYLOME OF A CHRONICALLY ADAPTED P. AERUGINOSA CLINICAL STRAIN, TBCF10839. TWO N(6)-METHYLADENINE (6MA) METHYLATION RECOGNITION MOTIFS (RCCANNNNNNNTGAR AND TRGANNNNNNTGC [MODIFICATION SITES ARE IN BOLD]) WERE IDENTIFIED AND PREDICTED AS NEW TYPE I METHYLATION SITES USING REBASE ANALYSIS. WE CONFIRMED THAT THE MOTIF TRGANNNNNNTGC WAS METHYLATED BY THE METHYLTRANSFERASE (MTASE) M.PAETBCFII, ACCORDING TO METHYLATION SENSITIVITY ASSAYS IN VIVO AND VITRO. TRANSCRIPTOMIC ANALYSIS SHOWED THAT A DELTAPAETBCFIIM KNOCKOUT MUTANT SIGNIFICANTLY DOWNREGULATED NITRIC OXIDE REDUCTASE (NOR) REGULATION AND EXPRESSION OF CODING GENES SUCH AS NOSR AND NORB, WHICH CONTAIN METHYLATED MOTIFS IN THEIR PROMOTERS OR CODING REGIONS. THE DELTAPAETBCFIIM STRAIN EXHIBITED REDUCED INTERCELLULAR SURVIVAL CAPACITY IN NO-PRODUCING RAW264.7 MACROPHAGES AND ATTENUATED VIRULENCE IN A GALLERIA MELLONELLA INFECTION MODEL; THE COMPLEMENTED STRAIN RECOVERED THESE DEFECTIVE PHENOTYPES. FURTHER PHYLOGENETIC ANALYSIS DEMONSTRATED THAT HOMOLOGS OF M.PAETBCFII OCCUR FREQUENTLY IN P. AERUGINOSA AS WELL AS OTHER BACTERIAL SPECIES. OUR WORK THEREFORE PROVIDED NEW INSIGHTS INTO THE RELATIONSHIP BETWEEN DNA METHYLATION, NO DETOXIFICATION, AND BACTERIAL VIRULENCE, LAYING A FOUNDATION FOR FURTHER EXPLORING THE MOLECULAR MECHANISM OF DNA METHYLTRANSFERASE IN REGULATING THE PATHOGENICITY OF P. AERUGINOSA. IMPORTANCE PSEUDOMONAS AERUGINOSA IS AN OPPORTUNISTIC PATHOGEN WHICH CAUSES ACUTE AND CHRONIC INFECTIONS THAT ARE DIFFICULT TO TREAT. COMPARATIVE GENOMIC ANALYSIS HAS SHOWED BROAD GENOME DIVERSITY AMONG P. AERUGINOSA CLINICAL STRAINS AND REVEALED THEIR DIFFERENT REGULATORY TRAITS COMPARED TO THE LABORATORY STRAINS. WHILE CURRENT INVESTIGATION OF THE EPIGENETICS OF P. AERUGINOSA IS STILL LACKING, UNDERSTANDING EPIGENETIC REGULATION MAY PROVIDE BIOMARKERS FOR DIAGNOSIS AND FACILITATE DEVELOPMENT OF NOVEL THERAPIES. DENITRIFICATION CAPABILITY IS CRITICAL FOR MICROBIAL VERSATILITY IN RESPONSE TO DIFFERENT ENVIRONMENTAL STRESS CONDITIONS, INCLUDING THE BACTERIAL INFECTION PROCESS, WHERE NITRIC OXIDE (NO) CAN BE GENERATED BY PHAGOCYTIC CELLS. THE DENITRIFICATION REGULATION MECHANISMS HAVE BEEN STUDIED INTENSIVELY AT GENETIC AND BIOCHEMICAL LEVELS. HOWEVER, THERE IS VERY LITTLE EVIDENCE ABOUT THE EPIGENETIC REGULATION OF BACTERIAL DENITRIFICATION MECHANISM. P. AERUGINOSA TBCF10839 IS A CHRONICALLY HOST-ADAPTED STRAIN ISOLATED FROM A CYSTIC FIBROSIS (CF) PATIENT WITH SPECIAL ANTIPHAGOCYTOSIS CHARACTERISTICS. HERE, WE INVESTIGATED THE REGULATORY EFFECT OF AN ORPHAN DNA MTASE, M.PAETBCFII, IN P. AERUGINOSA TBCF10839. WE DEMONSTRATED THAT THE DNA MTASE REGULATES THE TRANSCRIPTION OF DENITRIFICATION GENES REPRESENTED BY NOR AND AFFECTS ANTIPHAGOCYTIC ABILITY IN BACTERIA. IN SILICO ANALYSIS SUGGESTED THAT DNA METHYLATION MODIFICATION MAY ENHANCE GENE EXPRESSION BY AFFECTING THE BINDING OF TRANSACTING FACTORS SUCH AS DNR AND RPON. OUR FINDINGS NOT ONLY DEEPEN THE UNDERSTANDING OF THE ROLE OF DNA MTASE IN TRANSCRIPTIONAL REGULATION IN P. AERUGINOSA BUT ALSO PROVIDE A THEORETICAL FOUNDATION FOR THE IN-DEPTH STUDY OF THE MOLECULAR MECHANISM OF THE EPIGENETIC REGULATION ON DENITRIFICATION, VIRULENCE, AND HOST-PATHOGEN INTERACTION. 2022