-
European Journal of Oral Sciences Oct 2018The periodontal pathogen Porphyromonas gingivalis can invade host cells, a virulence trait which may contribute to the persistence of infection at subgingival sites....
The periodontal pathogen Porphyromonas gingivalis can invade host cells, a virulence trait which may contribute to the persistence of infection at subgingival sites. Whilst the antibiotic protection assay has been commonly employed to investigate and quantify P. gingivalis invasion, data obtained have varied widely and a thorough investigation of the factors influencing this is lacking. We investigated the role of a number of bacterial and host-cell factors and report that the growth phase of P. gingivalis, source (laboratory strain vs. clinical strain), host-cell identity (cell line vs. primary), host-cell lysis method, and host-cell passage number had no significant effect on bacterial invasion. However, incubation time, host-cell seeding density, method of quantification (viable count vs. DNA), and whether host cells were plated or in suspension, were shown to influence invasion. Also, cells isolated by rapid adhesion to fibronectin exhibited higher levels of P. gingivalis invasion, possibly as a result of increased levels of active α5β1 integrin. Interestingly, this may represent a population of cells with stem cell-like properties. This study provides important new information by identifying the most important factors that influence P. gingivalis invasion assays and may help to explain variations in the levels previously reported.
Topics: Bacterial Adhesion; Carcinoma, Squamous Cell; Cell Line, Tumor; Cells, Cultured; DNA, Bacterial; Fibronectins; Host-Pathogen Interactions; Humans; Periodontal Diseases; Porphyromonas gingivalis; Stem Cells
PubMed: 30070725
DOI: 10.1111/eos.12557 -
Clinical Infectious Diseases : An... Sep 1997
Topics: Fatty Acids; Porphyromonas
PubMed: 9310675
DOI: 10.1086/516223 -
Biomaterials Science Sep 2021, the pathogen of periodontal disease, is thought to be involved in various diseases throughout the body gingival tissue blood capillaries. However, the dynamic...
, the pathogen of periodontal disease, is thought to be involved in various diseases throughout the body gingival tissue blood capillaries. However, the dynamic analysis of the infection mechanism, particularly the deep invasion process of the gingival tissue, has not yet been elucidated because of the lack of both and models. In this study, we developed a vascularized three-dimensional (3D) gingival model with an epithelial barrier expressing cell-cell junctions using collagen microfibers (CMFs) to enable the dynamic analysis of the invasion process. Lipid raft disruption experiments in the gingival epithelial cell layer demonstrated that migrates into the deeper epithelium the intercellular pathway rather than intracellular routes. was shown to invade the 3D gingival model, being found inside blood capillaries during two days of culture. Notably, the number of bacteria had increased greatly at least two days later, whereas the mutant lacking the cysteine proteases, gingipains, showed a significantly lower number of survivors. The secretion of interleukin-6 (IL-6) from the gingival tissue decreased during the two days of infection with the wild type , but the opposite was found for the mutant suggesting that infection disturbs IL-6 secretion at an early stage. By allowing the dynamic observation of the invasion from the epithelial cell layer into the blood capillaries for the first time, this model will be a powerful tool for the development of novel therapeutics against periodontal infection related diseases.
Topics: Capillaries; Cells, Cultured; Epithelial Cells; Gingiva; Humans; Porphyromonas gingivalis
PubMed: 34582534
DOI: 10.1039/d1bm00831e -
Anaerobe Dec 2018Despite the wide implementation of MALDI-TOF MS for the rapid and reliable identification of most microorganisms, some taxonomic groups such as the Porphyromonas genus...
Despite the wide implementation of MALDI-TOF MS for the rapid and reliable identification of most microorganisms, some taxonomic groups such as the Porphyromonas genus remain largely untested. In this study we evaluated the performance of MALDI-TOF MS on this genus using a collection of 39 isolates sent for routine identification to our institution over a 16-year period. All of them were identified by DNA-sequencing analysis of the 16S rRNA gene plus the hsp60 gene when the previous one did not yield species-level assignment. MALDI-TOF MS provided correct identification at least at the genus level of 21/39 isolates (53.9%). Twelve isolates were correctly identified at the species level with a score value ≥ 2.0 and 9 more with score values < 2.0 and ≥ 1.7. The species most represented in the database (P. gingivalis and P. somerae) lay within this category. However, the species poorly represented in this database (P. asaccharolytica and P. uenonis) were mostly identified with lower scores (1.35-1.67) or remained unidentified by MALDI-TOF MS. The addition of two P. asaccharolytica reference spectra to our in-house library allowed 72.9% of genus-level identifications with 17/37 isolates (45.9%) identified with score values ≥ 2.0. Our results showed a high level of correlation between MALDI-TOF MS and DNA-based identification for Porphyromonas spp. strains at the species level, even with score values < 2.0. The reliability provided by MALDI-TOF MS increased when the database was fed with spectra from the species poorly represented in the commercial database.
Topics: Bacterial Typing Techniques; Bacteroidaceae Infections; DNA, Bacterial; Diagnostic Tests, Routine; Humans; Porphyromonas; RNA, Ribosomal, 16S; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 30541687
DOI: 10.1016/j.anaerobe.2018.06.017 -
Scientific Reports Jul 2017The development of antibiotics cannot keep up with the speed of resistance acquired by microorganisms. Recently, the development of antimicrobial photodynamic therapy...
The development of antibiotics cannot keep up with the speed of resistance acquired by microorganisms. Recently, the development of antimicrobial photodynamic therapy (aPDT) has been a necessary antimicrobial strategy against antibiotic resistance. Among the wide variety of bacteria found in the oral flora, Porphyromonas gingivalis (P. gingivalis) is one of the etiological agents of periodontal disease. aPDT has been studied for periodontal disease, but has risks of cytotoxicity to normal stained tissue. In this study, we performed aPDT using protoporphyrin IX (PpIX), an intracellular pigment of P. gingivalis, without an external photosensitizer. We confirmed singlet oxygen generation by PpIX in a blue-light irradiation intensity-dependent manner. We discovered that blue-light irradiation on P. gingivalis is potentially bactericidal. The sterilization mechanism seems to be oxidative DNA damage in bacterial cells. Although it is said that no resistant bacteria will emerge using aPDT, the conventional method relies on an added photosensitizer dye. PpIX in P. gingivalis is used in energy production, so aPDT applied to PpIX of P. gingivalis should limit the appearance of resistant bacteria. This approach not only has potential as an effective treatment for new periodontal diseases, but also offers potential antibacterial treatment for multiple drug resistant bacteria.
Topics: Anti-Bacterial Agents; Bacteroidaceae Infections; Humans; Light; Microbial Viability; Periodontal Diseases; Photosensitizing Agents; Porphyromonas gingivalis; Protoporphyrins; Singlet Oxygen
PubMed: 28701797
DOI: 10.1038/s41598-017-05706-1 -
Microbiology (Reading, England) Apr 2008HtrA is a heat-stress protein that functions both as a chaperone and as a serine protease. HtrA has been shown in several organisms to be involved in responses to...
HtrA is a heat-stress protein that functions both as a chaperone and as a serine protease. HtrA has been shown in several organisms to be involved in responses to stressful environmental conditions and involvement of HtrA in virulence has been reported in pathogenic species. A Porphyromonas gingivalis htrA mutant demonstrated no significant difference to the W83 parent strain when subjected to high temperature and pH values from 3 to 11. However, the htrA mutant showed increased sensitivity to H(2)O(2). Cell invasion assays indicated that the total interaction (adherence) with KB cells, human coronary artery endothelial cells and gingival epithelial cells (GEC) was the same for both the wild-type and the htrA mutant. However, the htrA mutant showed increased invasion in KB cells and GEC. Microarray experiments indicated that a total of 253 genes were differentially regulated in the htrA mutant, including a group of stress-related genes, which might be responsible for the observed decreased resistance to H(2)O(2). In animal experiments, a competition assay showed that the htrA mutant did not survive as well as the wild-type. In another in vivo assay, fewer mice infected with the htrA mutant died than mice infected with W83, suggesting that the htrA gene is virulence-related. These data indicate that the htrA gene in P. gingivalis does not relate to stress conditions such as high temperature and pH, but rather to H(2)O(2) stress. The htrA gene also appears to be important for virulence and survival in in vivo animal models.
Topics: Animals; Anti-Bacterial Agents; Bacterial Adhesion; Bacterial Proteins; Bacteroidaceae Infections; Cell Line; Gene Deletion; Gene Expression Profiling; Hot Temperature; Humans; Hydrogen Peroxide; Hydrogen-Ion Concentration; Mice; Mice, Inbred BALB C; Microbial Viability; Mutagenesis, Insertional; Oligonucleotide Array Sequence Analysis; Porphyromonas gingivalis; Survival Analysis; Virulence Factors
PubMed: 18375808
DOI: 10.1099/mic.0.2007/015131-0 -
Molecular Medicine Reports Aug 2015The Porphyromonas gingivalis bacterium is one of the most influential pathogens in oral infections. In the current study, the antimicrobial activity of α-amylase and...
The Porphyromonas gingivalis bacterium is one of the most influential pathogens in oral infections. In the current study, the antimicrobial activity of α-amylase and pentamidine against Porphyromonas gingivalis was evaluated. Their in vitro inhibitory activity was investigated with the agar overlay technique, and the minimal inhibitory and bactericidal concentrations were determined. Using the bactericidal concentration, the antimicrobial actions of the inhibitors were investigated. In the present study, multiple techniques were utilized, including scanning electron microscopy (SEM), general structural analysis and differential gene expression analysis. The results obtained from SEM and bactericidal analysis indicated a notable observation; the pentamidine and α-amylase treatment destroyed the structure of the bacterial cell membranes, which led to cell death. These results were used to further explore these inhibitors and the mechanisms by which they act. Downregulated expression levels were observed for a number of genes coding for hemagglutinins and gingipains, and various genes involved in hemin uptake, chromosome replication and energy production. However, the expression levels of genes associated with iron storage and oxidative stress were upregulated by α-amylase and pentamidine. A greater effect was noted in response to pentamidine treatment. The results of the present study demonstrate promising therapeutic potential for α-amylases and pentamidine. These molecules have the potential to be used to develop novel drugs and broaden the availability of pharmacological tools for the attenuation of oral infections caused by Porphyromonas gingivalis.
Topics: Adhesins, Bacterial; Anti-Bacterial Agents; Bacteroidaceae Infections; Cysteine Endopeptidases; Gene Expression Regulation, Bacterial; Gingipain Cysteine Endopeptidases; Hemagglutinins; Humans; Pentamidine; Porphyromonas gingivalis; alpha-Amylases
PubMed: 25846026
DOI: 10.3892/mmr.2015.3584 -
Infection and Immunity Jan 2011Porphyromonas gingivalis has been implicated in the etiology of adult periodontitis. In this study, we examined the viability of Drosophila melanogaster as a new model...
Porphyromonas gingivalis has been implicated in the etiology of adult periodontitis. In this study, we examined the viability of Drosophila melanogaster as a new model for examining P. gingivalis-host interactions. P. gingivalis (W83) infection of Drosophila resulted in a systemic infection that killed in a dose-dependent manner. Differences in the virulence of several clinically prevalent P. gingivalis strains were observed in the Drosophila killing model, and the results correlated well with studies in mammalian infection models and human epidemiologic studies. P. gingivalis pathobiology in Drosophila did not result from uncontrolled growth of the bacterium in the Drosophila hemolymph (blood) or overt damage to Drosophila tissues. P. gingivalis killing of Drosophila was multifactorial, involving several bacterial factors that are also involved in virulence in mammals. The results from this study suggest that many aspects of P. gingivalis pathogenesis in mammals are conserved in Drosophila, and thus the Drosophila killing model should be useful for characterizing P. gingivalis-host interactions and, potentially, polymicrobe-host interactions.
Topics: Animals; Disease Models, Animal; Drosophila melanogaster; Female; Porphyromonas gingivalis; Time Factors; Virulence
PubMed: 21041487
DOI: 10.1128/IAI.00784-10 -
Infection and Immunity Nov 2006Porphyromonas gingivalis is a crucial component of complex plaque biofilms that form in the oral cavity, resulting in the progression of periodontal disease. To...
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.
Topics: Biofilms; Microscopy, Electron, Scanning; Mutation; O Antigens; Penicillin Resistance; Porphyromonas gingivalis; UDPglucose 4-Epimerase
PubMed: 16954395
DOI: 10.1128/IAI.00261-06 -
Archives of Oral Biology Jan 2022To investigate the antibacterial and anti-biofilm effects of colloidal bismuth subcitrate (CBS) on Porphyromonas gingivalis (P. gingivalis) in its planktonic and biofilm...
OBJECTIVE
To investigate the antibacterial and anti-biofilm effects of colloidal bismuth subcitrate (CBS) on Porphyromonas gingivalis (P. gingivalis) in its planktonic and biofilm forms and also compare it with that of 0.2% chlorhexidine (CHX).
DESIGN
The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of CBS were determined by the microdilution method; the bacteriostatic rate of CBS was determined by the MTT assay; the effect of CBS on the membrane integrity of P. gingivalis was investigated by the flow cytometric methods. The effects of CBS on the biomass and bacterial activity of biofilm were investigated. Confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM) were used to investigate the activity and structure of biofilms.
RESULTS
The MIC and MBC values were 18.75 µg/mL and 37.5 µg/mL. CBS could damage the cell membrane of P. gingivalis. CBS effectively inhibited biofilm formation and promoted dissociation at higher concentrations of 37.5 µg/mL and 75 µg/mL, respectively. The results also indicated an altered biofilm structure and reduced biofilm thickness and bacterial aggregation.
CONCLUSIONS
CBS affected the metabolic and physiological processes of P. gingivalis, inhibited the formation of biofilm, and disrupted the mature biofilm.
Topics: Anti-Bacterial Agents; Biofilms; Microbial Sensitivity Tests; Organometallic Compounds; Porphyromonas gingivalis
PubMed: 34742000
DOI: 10.1016/j.archoralbio.2021.105300