Comparative gene expression analysis of Porphyromonas gingivalis ATCC 33277 in planktonic and biofilms states

Background and objective Porphyromonas gingivalis is a keystone pathogen in the onset and progression of periodontitis. Its pathogenicity has been related to its presence and survival within the subgingival biofilm. The aim of the present study was to compare the genome-wide transcription activities of P. gingivalis in biofilm and in planktonic growth, using microarray technology. Material and methods P. gingivalis ATCC 33277 was incubated in multi-well culture plates at 37°C for 96 hours under anaerobic conditions using an in vitro static model to develop both the planktonic and biofilm states (the latter over sterile ceramic calcium hydroxyapatite discs). The biofilm development was monitored by Confocal Laser Scanning Microscopy (CLSM) and Scanning Electron Microscopy (SEM). After incubation, the bacterial cells were harvested and total RNA was extracted and purified. Three biological replicates for each cell state were independently hybridized for transcriptomic comparisons. A linear model was used for determining differentially expressed genes and reverse transcription quantitative polymerase chain reaction (RT-qPCR) was used to confirm differential expression. The filtering criteria of ≥ ±2 change in gene expression and significance p-values of <0.05 were selected. Results A total of 92 out of 1,909 genes (4.8%) were differentially expressed by P. gingivalis growing in biofilm compared to planktonic. The 54 up-regulated genes in biofilm growth were mainly related to cell envelope, transport, and binding or outer membranes proteins. Thirty-eight showed decreased expression, mainly genes related to transposases or oxidative stress. Conclusion The adaptive response of P. gingivalis in biofilm growth demonstrated a differential gene expression.

[1]  Yong Chen,et al.  Comparative transcriptomic analysis of Clostridium acetobutylicum biofilm and planktonic cells. , 2016, Journal of biotechnology.

[2]  K. Pyrć,et al.  Functional Analysis of Porphyromonas gingivalis W83 CRISPR-Cas Systems , 2015, Journal of bacteriology.

[3]  J. Moon,et al.  Microarray analysis of the transcriptional responses of Porphyromonas gingivalis to polyphosphate , 2014, BMC Microbiology.

[4]  S. Roje,et al.  Sinorhizobium meliloti flavin secretion and bacteria-host interaction: role of the bifunctional RibBA protein. , 2014, Molecular plant-microbe interactions : MPMI.

[5]  P. Veith,et al.  Porphyromonas gingivalis outer membrane vesicles exclusively contain outer membrane and periplasmic proteins and carry a cargo enriched with virulence factors. , 2014, Journal of proteome research.

[6]  Bayly S. Wheeler Small RNAs, big impact: small RNA pathways in transposon control and their effect on the host stress response , 2013, Chromosome Research.

[7]  Yu-Yen Chen,et al.  Porphyromonas gingivalis and Treponema denticola Synergistic Polymicrobial Biofilm Development , 2013, PloS one.

[8]  M. Duncan,et al.  Role of Sodium in the RprY-Dependent Stress Response in Porphyromonas gingivalis , 2013, PloS one.

[9]  J. Kerr,et al.  Genetic diversity in the oral pathogen Porphyromonas gingivalis: molecular mechanisms and biological consequences. , 2013, Future microbiology.

[10]  David A. C. Beck,et al.  Regulon controlled by the GppX hybrid two component system in Porphyromonas gingivalis. , 2013, Molecular oral microbiology.

[11]  K. Leung,et al.  Role of rpoS in Escherichia coli O157:H7 Strain H32 Biofilm Development and Survival , 2012, Applied and Environmental Microbiology.

[12]  F. Yoshimura,et al.  Porphyromonas gingivalis FimA Fimbriae: Fimbrial Assembly by fimA Alone in the fim Gene Cluster and Differential Antigenicity among fimA Genotypes , 2012, PloS one.

[13]  F. Yoshimura,et al.  Porphyromonas gingivalis FimA fimbriae: Roles of the fim gene cluster in the fimbrial assembly and antigenic heterogeneity among fimA genotypes , 2012 .

[14]  David A. C. Beck,et al.  Deep Sequencing of Porphyromonas gingivalis and Comparative Transcriptome Analysis of a LuxS Mutant , 2012, Front. Cell. Inf. Microbio..

[15]  H. Fletcher,et al.  Nitric Oxide Stress Resistance in Porphyromonas gingivalis Is Mediated by a Putative Hydroxylamine Reductase , 2012, Journal of bacteriology.

[16]  Zhongge Zhang,et al.  Transposon-Mediated Adaptive and Directed Mutations and Their Potential Evolutionary Benefits , 2012, Journal of Molecular Microbiology and Biotechnology.

[17]  Wen-Han Yu,et al.  Comprehensive Transcriptome Analysis of the Periodontopathogenic Bacterium Porphyromonas gingivalis W83 , 2011, Journal of bacteriology.

[18]  R. Hayes,et al.  Oral Microbiome Profiles: 16S rRNA Pyrosequencing and Microarray Assay Comparison , 2011, PloS one.

[19]  Y. Noiri,et al.  Time Course of Gene Expression during Porphyromonas gingivalis Strain ATCC 33277 Biofilm Formation , 2011, Applied and Environmental Microbiology.

[20]  M. Kuehn,et al.  Biological functions and biogenesis of secreted bacterial outer membrane vesicles. , 2010, Annual review of microbiology.

[21]  M. Kuehn,et al.  Virulence and Immunomodulatory Roles of Bacterial Outer Membrane Vesicles , 2010, Microbiology and Molecular Biology Reviews.

[22]  L. Wong,et al.  Oral Biofilms: Emerging Concepts in Microbial Ecology , 2010, Journal of dental research.

[23]  P. Kolenbrander,et al.  Subgingival Biofilm Communities in Health and Disease , 2009 .

[24]  R. Lamont,et al.  Pathway analysis for intracellular Porphyromonas gingivalis using a strain ATCC 33277 specific database , 2009, BMC Microbiology.

[25]  R. Lamont,et al.  Proteomics of Porphyromonas gingivalis within a model oral microbial community , 2009, BMC Microbiology.

[26]  F. Yoshimura,et al.  Surface components of Porphyromonas gingivalis. , 2009, Journal of periodontal research.

[27]  C. Seers,et al.  Comparative transcriptomic analysis of Porphyromonas gingivalis biofilm and planktonic cells , 2009, BMC Microbiology.

[28]  M. Shemesh,et al.  DNA-microarrays identification of Streptococcus mutans genes associated with biofilm thickness , 2008, BMC Microbiology.

[29]  E. Reynolds,et al.  Porphyromonas gingivalis RgpA-Kgp Proteinase-Adhesin Complexes Penetrate Gingival Tissue and Induce Proinflammatory Cytokines or Apoptosis in a Concentration-Dependent Manner , 2008, Infection and Immunity.

[30]  R. Sayre,et al.  The vitamin riboflavin and its derivative lumichrome activate the LasR bacterial quorum-sensing receptor. , 2008, Molecular plant-microbe interactions : MPMI.

[31]  Thomas D. Schmittgen,et al.  Analyzing real-time PCR data by the comparative CT method , 2008, Nature Protocols.

[32]  D. R. Bond,et al.  Shewanella secretes flavins that mediate extracellular electron transfer , 2008, Proceedings of the National Academy of Sciences.

[33]  R. Lamont,et al.  Quantitative proteomics of intracellular Porphyromonas gingivalis , 2007, Proteomics.

[34]  D. Caugant,et al.  fimA Genotypes and Multilocus Sequence Types of Porphyromonas gingivalis from Patients with Periodontitis , 2007, Journal of Clinical Microbiology.

[35]  H. Kuramitsu,et al.  Porphyromonas gingivalis Vesicles Enhance Attachment, and the Leucine-Rich Repeat BspA Protein Is Required for Invasion of Epithelial Cells by “Tannerella forsythia” , 2006, Infection and Immunity.

[36]  M. Simionato,et al.  Porphyromonas gingivalis Genes Involved in Community Development with Streptococcus gordonii , 2006, Infection and Immunity.

[37]  R. Lamont,et al.  Oral microbial communities in sickness and in health. , 2005, Trends in microbiology.

[38]  Lian-Hui Zhang,et al.  VqsM, a novel AraC‐type global regulator of quorum‐sensing signalling and virulence in Pseudomonas aeruginosa , 2005, Molecular microbiology.

[39]  Anastasia Papakonstantinopoulou,et al.  Transcriptome Analysis of Pseudomonas aeruginosa Growth: Comparison of Gene Expression in Planktonic Cultures and Developing and Mature Biofilms , 2005, Journal of bacteriology.

[40]  J. Hillman,et al.  Microarray Analysis of Quorum-Sensing-Regulated Genes in Porphyromonas gingivalis , 2005, Infection and Immunity.

[41]  P. Marsh,et al.  Dental Plaque as a Microbial Biofilm , 2004, Caries Research.

[42]  Gordon K Smyth,et al.  Statistical Applications in Genetics and Molecular Biology Linear Models and Empirical Bayes Methods for Assessing Differential Expression in Microarray Experiments , 2011 .

[43]  Søren Molin,et al.  Global impact of mature biofilm lifestyle on Escherichia coli K‐12 gene expression , 2003, Molecular microbiology.

[44]  S. Ichiyama,et al.  Effect of Porphyromonas gingivalis Vesicles on Coaggregation of Staphylococcus aureus to Oral Microorganisms , 2003, Current Microbiology.

[45]  Terry Speed,et al.  Normalization of cDNA microarray data. , 2003, Methods.

[46]  R. Lamont,et al.  Microbial dinner-party conversations: the role of LuxS in interspecies communication. , 2003, Journal of medical microbiology.

[47]  M. Schembri,et al.  Global gene expression in Escherichia coli biofilms , 2003, Molecular microbiology.

[48]  Robert J. Palmer,et al.  Communication among Oral Bacteria , 2002, Microbiology and Molecular Biology Reviews.

[49]  W. Zumft,et al.  Nitric oxide signaling and NO dependent transcriptional control in bacterial denitrification by members of the FNR-CRP regulator family. , 2002, Journal of molecular microbiology and biotechnology.

[50]  Roger E. Bumgarner,et al.  Gene expression in Pseudomonas aeruginosa biofilms , 2001, Nature.

[51]  R. McLean,et al.  Impact of rpoS Deletion onEscherichia coli Biofilms , 1999, Applied and Environmental Microbiology.

[52]  R. Blumenthal,et al.  Mapping regulatory networks in microbial cells. , 1999, Trends in microbiology.

[53]  W. Zumft Cell biology and molecular basis of denitrification. , 1997, Microbiology and molecular biology reviews : MMBR.

[54]  A Bairoch,et al.  Arac/XylS family of transcriptional regulators , 1997, Microbiology and molecular biology reviews : MMBR.

[55]  C. M. Belton,et al.  Role of fimbriae in Porphyromonas gingivalis invasion of gingival epithelial cells , 1997, Infection and immunity.

[56]  Ross Ihaka,et al.  Gentleman R: R: A language for data analysis and graphics , 1996 .

[57]  J. Tiedje,et al.  Denitrification: production and consumption of nitric oxide , 1994, Applied and environmental microbiology.

[58]  Z Lewandowski,et al.  Biofilms, the customized microniche , 1994, Journal of bacteriology.

[59]  J. G. Fisher,et al.  Inactivation of the Porphyromonas gingivalis fimA gene blocks periodontal damage in gnotobiotic rats , 1994, Journal of bacteriology.

[60]  K. Krogfelt,et al.  Investigation of minor components of Escherichia coli type 1 fimbriae: protein chemical and immunological aspects. , 1988, Microbial pathogenesis.

[61]  P. Klemm,et al.  Three fim genes required for the regulation of length and mediation of adhesion of Escherichia coli type 1 fimbriae , 1987, Molecular and General Genetics MGG.

[62]  E. Beachey,et al.  The genetic determinant of adhesive function in type 1 fimbriae of Escherichia coli is distinct from the gene encoding the fimbrial subunit , 1986, Journal of bacteriology.

[63]  Keith Durkin,et al.  Chromosome Research : An International Journal on the Molecular, Supramolecular and Evolutionary Aspects of Chromosome Biology , 2011 .

[64]  C. Arnault,et al.  Genome and stresses: Reactions against aggressions, behavior of transposable elements , 2005, Genetica.

[65]  P. Klemm,et al.  The fimD gene required for cell surface localization of Escherichia coli type 1 fimbriae , 2004, Molecular and General Genetics MGG.

[66]  S. Socransky,et al.  Dental biofilms: difficult therapeutic targets. , 2002, Periodontology 2000.

[67]  HighWire Press,et al.  Microbiology and molecular biology reviews : MMBR. , 1997 .