Complex Host – Pathogen Interactions in the Oral Cavity

We have previously shown, using human microarrays, that Streptococcus gordonii and Porphyromonas gingivalis elicit a different transcriptional signature whether they are used in mono or mixed cultures with Human Immortalized Gingival Keratinocytes (HIGK).  Objective: In this study, we aimed to investigate if this differential signature is reflected phenotypically by the levels of adhesion and invasion of HIGK cells using direct microscopic visualization.  Methods:  P. gingivalis was stained with the live-dye Cell Tracker Green (Invitrogen, Eugene, Oregon), while S. gordonii was stained with DRAQ5 (Biostatus, UK) following the manufacturers' recommendations. HIGK cells were co-cultured for 2 hours with stained P. gingivalis alone (multiplicity of infection (MOI) 100:1), stained S. gordonii alone (MOI 2,500:1), or a mixed culture of P. gingivalis and S. gordonii at their respective MOI in LAB-TEK II 4-well microscopy chamber slides  (Nalge Nunc Internatonal, Naperville, IL). Following co-culture, infected cells were washed, immobilized, permeabilized and stained with Texas Red Phalloidin (Invitrogen), which is specific for F-actin. Host-microbe interactions were observed using confocal laser microscopy, analyzed and quantified using the Imaris software (BITPLANE AG Scientific Solutions, Saint Paul, MN).  Results:  The levels of interaction and invasion were similar (p=0.106) whether P. gingivalis or S. gordonii were in single-species or in mixed cultures with HIGK cells.  Conclusions:  The differential signature that was observed using microarrays cannot be correlated at this point with discrepancies in the levels of adhesion and invasion of P. gingivalis. Hence, member of the “good oral flora” may have evolved additional cellular means to modulate the host cell's transcriptome, in addition to known antagonistic interactions.   Acknowledgements:  This study is supported by NIH/NIDCR DE16715 (MH), and UFCD summer research fellowship, and NIH/NIDCR T32 Grant DE007200.