Differential Proteomic Analysis of a Polymicrobial Biofilm

Abstract

Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia exist in a polymicrobial biofilm associated with chronic periodontitis. The aim of this study was to culture these three species as a polymicrobial biofilm and to determine proteins important for bacterial interactions. In a flow cell all three species attached and grew as a biofilm; however, after 90 h of culture P. gingivalis and T. denticola were closely associated and dominated the polymicrobial biofilm. For comparison, planktonic cultures of P. gingivalis and T. denticola were grown separately in continuous culture. Whole cell lysates were subjected to SDS-PAGE, followed by in-gel proteolytic H2 16O/H2 18O labeling. From two replicates, 135 and 174 P. gingivalis proteins and 134 and 194 T. denticola proteins were quantified by LC–MALDI TOF/TOF MS. The results suggest a change of strategy in iron acquisition by P. gingivalis due to large increases in the abundance of HusA and HusB in the polymicrobial biofilm while HmuY and other iron/haem transport systems decreased. Significant changes in the abundance of peptidases and enzymes involved in glutamate and glycine catabolism suggest syntrophy. These data indicate an intimate association between P. gingivalis and T. denticola in a biofilm that may play a role in disease pathogenesis.

Oxantel Disrupts Polymicrobial Biofilm Development of Periodontal Pathogens

“…P. gingivalis and T. denticola coaggregation has been demonstrated (54-57), which would contribute to their colocalization and synergism in biofilm formation (20,21,37). When cultured together as part of a polymicrobial biofilm, significant changes occur in the abundance of P. gingivalis and T. denticola peptidases and enzymes involved in glutamate and glycine catabolism, supporting previous reports of syntrophy, as well as changes in abundance of proteins involved in iron/heme acquisition (22).…”

“…In the present study, these three species were used as a model dysbiotic pathogenic polymicrobial biofilm (12,22). As oxantel has no direct effect on T. denticola due to its lack of Frd, there is the possibility that T. denticola could benefit from the suppression of T. forsythia and P. gingivalis.…”

“…Polymicrobial biofilms composed of P. gingivalis, T. forsythia, and T. denticola were cultured in a static well assay in OBGM, a growth medium we have developed for the culture of these three bacterial species (19,22). The starting inoculum for each biofilm in the 2-ml well was composed of an average of 6.9 ϫ 10 6 P. gingivalis cells, 6.7 ϫ 10 6 T. forsythia cells, or 8.8 ϫ 10 6 T. denticola cells, with a combined viability of Ͼ97%, as determined by flow cytometry.…”

“…When coinoculated in small animal models of periodontitis, P. gingivalis and T. denticola exhibit a synergistic virulence (17)(18)(19). These species also display synergistic biofilm formation (20)(21)(22) and respond to each other's presence by modulating the abundance of a range of proteins (22).…”

Significances of OMV and Extracellular Vesicle Proteomics

“…Cabral et al, did the differential proteomics of multi-drug resistant Acinetobacter cultured in three diverse conditions (exponential, late stationary phase and biofilms stage) and they also examine the effects of salicylate on the biofilm formation which is a biofilm inhibitory compound. This multiple approach strategy explained unique lifestyle of A. baumannii, which are involved in both virulence and biofilms formation [148]. Differential proteomics of the Porphyromonas gingivalis, Treponema denticola suggest synergestic relationship in the polymicrobial biofilms formation [149].…”