scholarly journals Targeting the HUβ Protein PreventsPorphyromonas gingivalisfrom Entering into Preexisting Biofilms

2018 ◽  
Vol 200 (11) ◽  
pp. e00790-17 ◽  
Author(s):  
Christopher J. Rocco ◽  
Lauren O. Bakaletz ◽  
Steven D. Goodman
Keyword(s):  
Oral Cavity ◽  
Periodontal Disease ◽  
Porphyromonas Gingivalis ◽  
The United States ◽  
Extracellular Dna ◽  
Bacterial Biofilms ◽  
Oral Streptococci ◽  
Content Type ◽  
Oral Streptococcus ◽  
Streptococcal Species

ABSTRACTThe oral cavity is home to a wide variety of bacterial species, both commensal, such as various streptococcal species, and pathogenic, such asPorphyromonas gingivalis, one of the main etiological agents of periodontal disease. Our understanding of how these bacteria ultimately cause disease is highly dependent upon understanding how they coexist and interact with one another in biofilm communities and the mechanisms by which biofilms are formed. Our research has demonstrated that the DNABII family of DNA-binding proteins are important components of the extracellular DNA (eDNA)-dependent matrix of bacterial biofilms and that sequestering these proteins via protein-specific antibodies results in the collapse of the biofilm structure and release of the resident bacteria. While the high degree of similarity among the DNABII family of proteins has allowed antibodies derived against specific DNABII proteins to disrupt biofilms formed by a wide range of bacterial pathogens, the DNABII proteins ofP. gingivalishave proven to be antigenically distinct, allowing us to determine if we can use anti-P. gingivalisHUβ antibodies to specifically target this species for removal from a mixed-species biofilm. Importantly, despite forming homotypic biofilmsin vitro,P. gingivalismust enter preexisting biofilmsin vivoin order to persist within the oral cavity. The data presented here indicate that antibodies derived against theP. gingivalisDNABII protein, HUβ, reduce by half the amount ofP. gingivalisorganisms entering into preexisting biofilm formed by four oral streptococcal species. These results support our efforts to develop methods for preventing and treating periodontal disease.IMPORTANCEPeriodontitis is one of the most prevalent chronic infections, affecting 40 to 50% of the population of the United States. The root cause of periodontitis is the presence of bacterial biofilms within the gingival space, withPorphyromonas gingivalisbeing strongly associated with the development of the disease. Periodontitis also increases the risk of secondary conditions and infections such as atherosclerosis and infective endocarditis caused by oral streptococci. To induce periodontitis,P. gingivalisneeds to incorporate into preformed biofilms, with oral streptococci being important binding partners. Our research demonstrates that targeting DNABII proteins with an antibody disperses oral streptococcus biofilm and preventsP. gingivalisentry into oral streptococcus biofilm. These results suggest potential therapeutic treatments for endocarditis caused by streptococci as well as periodontitis.

scholarly journals Identification and Characterization of Porphyromonas gingivalis Client Proteins That Bind to Streptococcus oralis Glyceraldehyde-3-Phosphate Dehydrogenase

2012 ◽  
Vol 81 (3) ◽  
pp. 753-763 ◽  
Author(s):  
Kazuhiko Maeda ◽  
Hideki Nagata ◽  
Masae Kuboniwa ◽  
Miki Ojima ◽  
Tsukasa Osaki ◽  
...  
Keyword(s):  
Amino Acid ◽  
Porphyromonas Gingivalis ◽  
Interaction Analysis ◽  
Site Directed Mutagenesis ◽  
Receptor Protein ◽  
Oral Streptococci ◽  
Amino Acid Residues ◽  
Content Type ◽  
Streptococcus Oralis ◽  
Client Proteins

ABSTRACTCoaggregation ofPorphyromonas gingivalisand oral streptococci is thought to play an important role inP. gingivaliscolonization. Previously, we reported thatP. gingivalismajor fimbriae interacted withStreptococcus oralisglyceraldehyde-3-phosphate dehydrogenase (GAPDH), and that amino acid residues 166 to 183 of GAPDH exhibited strong binding activity towardP. gingivalisfimbriae (H. Nagata, M. Iwasaki, K. Maeda, M. Kuboniwa, E. Hashino, M. Toe, N. Minamino, H. Kuwahara, and S. Shizukuishi, Infect. Immun.77:5130–5138, 2009). The present study aimed to identify and characterizeP. gingivaliscomponents other than fimbriae that interact withS. oralisGAPDH. A pulldown assay was performed to detect potential interactions betweenP. gingivalisclient proteins andS. oralisrecombinant GAPDH with amino acid residues 166 to 183 deleted by site-directed mutagenesis. Seven proteins, namely,tonB-dependent receptor protein (RagA4), arginine-specific proteinase B, 4-hydroxybutyryl-coenzyme A dehydratase (AbfD), lysine-specific proteinase, GAPDH, NAD-dependent glutamate dehydrogenase (GDH), and malate dehydrogenase (MDH), were identified by two-dimensional gel electrophoresis followed by proteomic analysis using tandem mass spectrometry. Interactions between these client proteins andS. oralisGAPDH were analyzed with a biomolecular interaction analysis system.S. oralisGAPDH showed high affinity for five of the seven client proteins (RagA4, AbfD, GAPDH, GDH, and MDH). Interactions betweenP. gingivalisandS. oraliswere measured by a turbidimetric method and fluorescence microscopy. RagA4, AbfD, and GDH enhanced coaggregation, whereas GAPDH and MDH inhibited coaggregation. Furthermore, the expression ofluxSinP. gingivaliswas upregulated by RagA4, AbfD, and GDH but was downregulated by MDH. These results indicate that the fiveP. gingivalisclient proteins function as regulators inP. gingivalisbiofilm formation with oral streptococci.

scholarly journals Porphyromonas gingivalis galE Is Involved in Lipopolysaccharide O-Antigen Synthesis and Biofilm Formation

2006 ◽  
Vol 74 (11) ◽  
pp. 6145-6153 ◽  
Author(s):  
Ryoma Nakao ◽  
Hidenobu Senpuku ◽  
Haruo Watanabe
Keyword(s):  
Oral Cavity ◽  
Periodontal Disease ◽  
Porphyromonas Gingivalis ◽  
Parental Strain ◽  
Biofilm Formation ◽  
Gene Knockout ◽  
Partial Recovery ◽  
Wild Type ◽  
O Antigen ◽  
Crucial Component

ABSTRACT Porphyromonas gingivalis is a crucial component of complex plaque biofilms that form in the oral cavity, resulting in the progression of periodontal disease. To elucidate the mechanism of periodontal biofilm formation, we analyzed the involvement of several genes related to the synthesis of polysaccharides in P. gingivalis. Gene knockout P. gingivalis mutants were constructed by insertion of an ermF-ermAM cassette; among these mutants, the galE mutant showed some characteristic phenotypes involved in the loss of GalE activity. As expected, the galE mutant accumulated intracellular carbohydrates in the presence of 0.1% galactose and did not grow in the presence of galactose at a concentration greater than 1%, in contrast to the parental strain. Lipopolysaccharide (LPS) analysis indicated that the length of the O-antigen chain of the galE mutant was shorter than that of the wild type. It was also demonstrated that biofilms generated by the galE mutant had an intensity 4.5-fold greater than those of the wild type. Further, the galE mutant was found to be significantly susceptible to some antibiotics in comparison with the wild type. In addition, complementation of the galE mutation led to a partial recovery of the parental phenotypes. We concluded that the galE gene plays a pivotal role in the modification of LPS O antigen and biofilm formation in P. gingivalis and considered that our findings of a relationship between the function of the P. gingivalis galE gene and virulence phenotypes such as biofilm formation may provide clues for understanding the mechanism of pathogenicity in periodontal disease.

scholarly journals Characterization of the O-Glycoproteome of Porphyromonas gingivalis

2022 ◽  
Author(s):  
Paul D. Veith ◽  
Mikio Shoji ◽  
Nichollas E. Scott ◽  
Eric C. Reynolds
Keyword(s):  
Rheumatoid Arthritis ◽  
Cardiovascular Disease ◽  
Periodontal Disease ◽  
Porphyromonas Gingivalis ◽  
Thermodynamic Stability ◽  
Protein Function ◽  
Protein Glycosylation ◽  
Content Type ◽  
Severe Periodontal Disease

Porphyromonas gingivalis is an oral pathogen primarily associated with severe periodontal disease and further associated with rheumatoid arthritis, dementia, cardiovascular disease, and certain cancers. Protein glycosylation can be important for a variety of reasons including protein function, solubility, protease resistance, and thermodynamic stability.

scholarly journals Protein Analysis of Sapienic Acid-Treated Porphyromonas gingivalis Suggests Differential Regulation of Multiple Metabolic Pathways

2015 ◽  
Vol 198 (1) ◽  
pp. 157-167 ◽  
Author(s):  
Carol L. Fischer ◽  
Deborah V. Dawson ◽  
Derek R. Blanchette ◽  
David R. Drake ◽  
Philip W. Wertz ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Fatty Acid ◽  
Oral Cavity ◽  
Porphyromonas Gingivalis ◽  
Metabolic Pathways ◽  
Acid Treatment ◽  
Cell Protein ◽  
Content Type ◽  
Kegg Pathways ◽  
Molecular Functions

ABSTRACTLipids endogenous to skin and mucosal surfaces exhibit potent antimicrobial activity againstPorphyromonas gingivalis, an important colonizer of the oral cavity implicated in periodontitis. Our previous work demonstrated the antimicrobial activity of the fatty acid sapienic acid (C16:1Δ6) againstP. gingivalisand found that sapienic acid treatment alters both protein and lipid composition from those in controls. In this study, we further examined whole-cell protein differences between sapienic acid-treated bacteria and untreated controls, and we utilized open-source functional association and annotation programs to explore potential mechanisms for the antimicrobial activity of sapienic acid. Our analyses indicated that sapienic acid treatment induces a unique stress response inP. gingivalisresulting in differential expression of proteins involved in a variety of metabolic pathways. This network of differentially regulated proteins was enriched in protein-protein interactions (P= 2.98 × 10−8), including six KEGG pathways (Pvalue ranges, 2.30 × 10−5to 0.05) and four Gene Ontology (GO) molecular functions (Pvalue ranges, 0.02 to 0.04), with multiple suggestive enriched relationships in KEGG pathways and GO molecular functions. Upregulated metabolic pathways suggest increases in energy production, lipid metabolism, iron acquisition and processing, and respiration. Combined with a suggested preferential metabolism of serine, which is necessary for fatty acid biosynthesis, these data support our previous findings that the site of sapienic acid antimicrobial activity is likely at the bacterial membrane.IMPORTANCEP. gingivalisis an important opportunistic pathogen implicated in periodontitis. Affecting nearly 50% of the population, periodontitis is treatable, but the resulting damage is irreversible and eventually progresses to tooth loss. There is a great need for natural products that can be used to treat and/or prevent the overgrowth of periodontal pathogens and increase oral health. Sapienic acid is endogenous to the oral cavity and is a potent antimicrobial agent, suggesting a potential therapeutic or prophylactic use for this fatty acid. This study examines the effects of sapienic acid treatment onP. gingivalisand highlights the membrane as the likely site of antimicrobial activity.

scholarly journals PerR-Regulated Manganese Ion Uptake Contributes to Oxidative Stress Defense in an Oral Streptococcus

2014 ◽  
Vol 80 (8) ◽  
pp. 2351-2359 ◽  
Author(s):  
Xinhui Wang ◽  
Huichun Tong ◽  
Xiuzhu Dong
Keyword(s):  
Iron Homeostasis ◽  
Critical Role ◽  
Brain Heart Infusion ◽  
Oral Streptococci ◽  
Content Type ◽  
Manganese Ion ◽  
Gram Positive Bacteria ◽  
Wild Type Phenotype ◽  
Oral Streptococcus ◽  
Antioxidative Stress

ABSTRACTMetal homeostasis plays a critical role in antioxidative stress.Streptococcus oligofermentans, an oral commensal facultative anaerobe lacking catalase activity, produces and tolerates abundant H2O2, whereas Dpr (an Fe2+-chelating protein)-dependent H2O2protection does not confer such high tolerance. Here, we report that inactivation ofperR, a peroxide-responsive repressor that regulates zinc and iron homeostasis in Gram-positive bacteria, increased the survival of H2O2-pulsedS. oligofermentans32-fold and elevated cellular manganese 4.5-fold.perRcomplementation recovered the wild-type phenotype. When grown in 0.1 to 0.25 mM MnCl2,S. oligofermentansincreased survival after H2O2stress 2.5- to 23-fold, and even greater survival was found for theperRmutant, indicating that PerR is involved in Mn2+-mediated H2O2resistance inS. oligofermentans. Mutation ofmntAcould not be obtained in brain heart infusion (BHI) broth (containing ∼0.4 μM Mn2+) unless it was supplemented with ≥2.5 μM MnCl2and caused 82 to 95% reduction of the cellular Mn2+level, whilemntABCoverexpression increased cellular Mn2+2.1- to 4.5-fold. Thus, MntABC was identified as a high-affinity Mn2+transporter inS. oligofermentans. mntAmutation reduced the survival of H2O2-pulsedS. oligofermentans5.7-fold, whilemntABCoverexpression enhanced H2O2-challenged survival 12-fold, indicating that MntABC-mediated Mn2+uptake is pivotal to antioxidative stress inS. oligofermentans. perRmutation or H2O2pulsing upregulatedmntABC, while H2O2-induced upregulation diminished in theperRmutant. This suggests thatperRrepressesmntABCexpression but H2O2can release the suppression. In conclusion, this work demonstrates that PerR regulates manganese homeostasis inS. oligofermentans, which is critical to H2O2stress defenses and may be distributed across all oral streptococci lacking catalase.

scholarly journals Macrophage Polarization Alters Postphagocytosis Survivability of the CommensalStreptococcus gordonii

2017 ◽  
Vol 86 (3) ◽  
Author(s):  
Andrew J. Croft ◽  
Sarah Metcalfe ◽  
Kiyonobu Honma ◽  
Jason G. Kay
Keyword(s):  
Macrophage Polarization ◽  
Oral Bacteria ◽  
Ros Production ◽  
Systemic Diseases ◽  
Oxygen Species ◽  
Oral Streptococci ◽  
Phagocytic Cells ◽  
Content Type ◽  
Systemic Inflammatory Disease ◽  
Oral Streptococcus

ABSTRACTOral streptococci are generally considered commensal organisms; however, they are becoming recognized as important associate pathogens during the development of periodontal disease as well as being associated with several systemic diseases, including as a causative agent of infective endocarditis. An important virulence determinant of these bacteria is an ability to evade destruction by phagocytic cells, yet how this subversion occurs is mostly unknown. UsingStreptococcus gordoniias a model commensal oral streptococcus that is also associated with disease, we find that resistance to reactive oxygen species (ROS) with an active ability to damage phagosomes allows the bacterium to avoid destruction within macrophages. This ability to survive relies not only on the ROS resistance capabilities of the bacterium but also on ROS production by macrophages, with both being required for maximal survival of internalized bacteria. Importantly, we also show that this dependence on ROS production by macrophages for resistance has functional significance:S. gordoniiintracellular survival increases when macrophages are polarized toward an activated (M1) profile, which is known to result in prolonged phagosomal ROS production compared to that of alternatively (M2) polarized macrophages. We additionally find evidence of the bacterium being capable of both delaying the maturation of and damaging phagosomes. Taken together, these results provide essential insights regarding the mechanisms through which normally commensal oral bacteria can contribute to both local and systemic inflammatory disease.

scholarly journals Natural Competence Is a Major Mechanism for Horizontal DNA Transfer in the Oral Pathogen Porphyromonas gingivalis

mBio ◽  
2012 ◽  
Vol 3 (1) ◽  
Author(s):  
Gena D. Tribble ◽  
Todd W. Rigney ◽  
Doan-Hieu V. Dao ◽  
Cindy T. Wong ◽  
Jennifer E. Kerr ◽  
...  
Keyword(s):  
Porphyromonas Gingivalis ◽  
Dna Transfer ◽  
Extracellular Dna ◽  
Minor Role ◽  
Chromosomal Dna ◽  
Natural Competence ◽  
Content Type ◽  
A Minor ◽  
First Time

ABSTRACTPorphyromonas gingivalisis a Gram-negative anaerobe that resides exclusively in the human oral cavity. Long-term colonization byP. gingivalisrequires the bacteria to evade host immune responses while adapting to the changing host physiology and alterations in the composition of the oral microflora. The genetic diversity ofP. gingivalisappears to reflect the variability of its habitat; however, little is known about the molecular mechanisms generating this diversity. Previously, our research group established that chromosomal DNA transfer occurs betweenP. gingivalisstrains. In this study, we examine the role of putative DNA transfer genes in conjugation and transformation and demonstrate that natural competence mediated bycomFis the dominant form of chromosomal DNA transfer, with transfer by a conjugation-like mechanism playing a minor role. Our results reveal that natural competence mechanisms are present in multiple strains ofP. gingivalis, and DNA uptake is not sensitive to DNA source or modification status. Furthermore, extracellular DNA was observed for the first time inP. gingivalisbiofilms and is predicted to be the major DNA source for horizontal transfer and allelic exchange between strains. We propose that exchange of DNA in plaque biofilms by a transformation-like process is of major ecological importance in the survival and persistence ofP. gingivalisin the challenging oral environment.IMPORTANCEP. gingivaliscolonizes the oral cavities of humans worldwide. The long-term persistence of these bacteria can lead to the development of chronic periodontitis and host morbidity associated with tooth loss.P. gingivalisis a genetically diverse species, and this variability is believed to contribute to its successful colonization and survival in diverse human hosts, as well as evasion of host immune defenses and immunization strategies. We establish here that natural competence is the major driving force behindP. gingivalisDNA exchange and that conjugative DNA transfer plays a minor role. Furthermore, we reveal for the first time the presence of extracellular DNA inP. gingivalisbiofilms, which is most likely the source of DNA exchanged between strains within dental plaque. These studies expand our understanding of the mechanisms used by this important member of the human oral flora to transition its relationship with the host from a commensal to a pathogenic relationship.

scholarly journals Reduced Glutathione Mediates Resistance to H2S Toxicity in Oral Streptococci

2016 ◽  
Vol 82 (7) ◽  
pp. 2078-2085 ◽  
Author(s):  
Xi Jia Ooi ◽  
Kai Soo Tan
Keyword(s):  
Periodontal Disease ◽  
Biofilm Formation ◽  
Reduced Glutathione ◽  
Differential Susceptibility ◽  
Oral Streptococci ◽  
Content Type ◽  
Defense Systems ◽  
Oral Microflora ◽  
Antioxidant Defense Systems ◽  
Dependent Mechanism

ABSTRACTPeriodontal disease is associated with changes in the composition of the oral microflora, where health-associated oral streptococci decrease while Gram-negative anaerobes predominate in disease. A key feature of periodontal disease-associated anaerobes is their ability to produce hydrogen sulfide (H2S) abundantly as a by-product of anaerobic metabolism. So far, H2S has been reported to be either cytoprotective or cytotoxic by modulating bacterial antioxidant defense systems. Although oral anaerobes produce large amounts of H2S, the potential effects of H2S on oral streptococci are currently unknown. The aim of this study was to determine the effects of H2S on the survival and biofilm formation of oral streptococci. The growth and biofilm formation ofStreptococcus mitisandStreptococcus oraliswere inhibited by H2S. However, H2S did not significantly affect the growth ofStreptococcus gordoniiorStreptococcus sanguinis. The differential susceptibility of oral streptococci to H2S was attributed to differences in the intracellular concentrations of reduced glutathione (GSH). In the absence of GSH, H2S elicited its toxicity through an iron-dependent mechanism. Collectively, our results showed that H2S exerts antimicrobial effects on certain oral streptococci, potentially contributing to the decrease in health-associated plaque microflora.

scholarly journals A Linear Plasmid-Like Prophage of Actinomyces odontolyticus Promotes Biofilm Assembly

2018 ◽  
Vol 84 (17) ◽  
Author(s):  
Mengyu Shen ◽  
Yuhui Yang ◽  
Wei Shen ◽  
Lujia Cen ◽  
Jeffrey S. McLean ◽  
...  
Keyword(s):  
Oral Cavity ◽  
Human Microbiome ◽  
Genomic Analysis ◽  
Electron Microscopic Examination ◽  
Oral Microbiome ◽  
Extracellular Dna ◽  
Linear Plasmid ◽  
Electron Microscopic ◽  
Content Type ◽  
Human Oral Cavity

ABSTRACT The human oral cavity is home to a large number of bacteria and bacteriophages (phages). However, the biology of oral phages as members of the human microbiome is not well understood. Recently, we isolated Actinomyces odontolyticus subsp. actinosynbacter strain XH001 from the human oral cavity, and genomic analysis revealed the presence of an intact prophage named xhp1. Here, we demonstrated that xhp1 is a linear plasmid-like prophage, which is a newly identified phage of A. odontolyticus. The prophage xhp1 genome is a 35-kb linear double-stranded DNA with 10-bp single-stranded, 3’ cohesive ends. xhp1 exists extrachromosomally, with an estimated copy number of 5. Annotation of xhp1 revealed 54 open reading frames, while phylogenetic analysis suggests that it has limited similarity with other phages. xhp1 phage particles can be induced by mitomycin C and belong to the Siphoviridae family, according to transmission electron microscopic examination. The released xhp1 particles can reinfect the xhp1-cured XH001 strain and result in tiny blurry plaques. Moreover, xhp1 promotes XH001 biofilm formation through spontaneous induction and the release of host extracellular DNA (eDNA). In conclusion, we identified a linear plasmid-like prophage of A. odontolyticus, which enhances bacterial host biofilm assembly and could be beneficial to the host for its persistence in the oral cavity. IMPORTANCE The biology of phages as members of the human oral microbiome is understudied. Here, we report the characterization of xhp1, a novel linear plasmid-like prophage identified from a human oral isolate, Actinomyces odontolyticus subsp. actinosynbacter strain XH001. xhp1 can be induced and reinfect xhp1-cured XH001. The spontaneous induction of xhp1 leads to the lysis of a subpopulation of bacterial hosts and the release of eDNA that promotes biofilm assembly, thus potentially contributing to the persistence of A. odontolyticus within the oral cavity.

scholarly journals Draft Whole-Genome Sequences of Periodontal Pathobionts Porphyromonas gingivalis, Prevotella intermedia, and Tannerella forsythia Contain Phase-Variable Restriction-Modification Systems

2017 ◽  
Vol 5 (46) ◽  
Author(s):  
Richard D. Haigh ◽  
Liam A. Crawford ◽  
Joseph D. Ralph ◽  
Joseph J. Wanford ◽  
Sonia R. Vartoukian ◽  
...  
Keyword(s):  
Periodontal Disease ◽  
Porphyromonas Gingivalis ◽  
Disease Process ◽  
Oral Bacteria ◽  
Host Responses ◽  
Prevotella Intermedia ◽  
Phase Variable ◽  
Genome Sequences ◽  
Tannerella Forsythia ◽  
Content Type

ABSTRACT Periodontal disease comprises mild to severe inflammatory host responses to oral bacteria that can cause destruction of the tooth-supporting tissue. We report genome sequences for 18 clinical isolates of Porphyromonas gingivalis, Prevotella intermedia, and Tannerella forsythia, Gram-negative obligate anaerobes that play a role in the periodontal disease process.

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