The ability of Porphyromonas gingivalis to cause adult periodontitis is determined by its arsenal of virulence factors. Here, we investigated the importance of biofilm formation and bacterial dipeptidyl peptidase IV (DPPIV) for the pathogenicity of clinical P. gingivalis isolates. In our study, the isolates with biofilm-forming capacity also showed high DPPIV activity in vitro. Moreover, DPPIV activity increased in P. gingivalis biofilms compared to planktonic cells. In a murine subcutaneous abscess model, the biofilm-forming isolates with high DPPIV activity proved to be pathogenic, while the nonbiofilm formers with low DPPIV activity did not induce abscesses. The biofilm-forming ATCC 33277 strain with low DPPIV activity was not pathogenic in mice either. Our results suggest that biofilm formation and DPPIV activity contribute to the pathogenic potential of P. gingivalis. Furthermore, we show that biofilm formation may enhance P. gingivalis virulence through an increased DPPIV activity. Because of their importance for bacterial colonization and growth, biofilm formation and DPPIV activity could present interesting therapeutic targets to tackle periodontitis.

Since biofilms are important in many clinical, industrial, and environmental settings, reliable methods to quantify these sessile microbial populations are crucial. Most of the currently available techniques do not allow the enumeration of the viable cell fraction within the biofilm and are often time consuming. This paper proposes flow cytometry (FCM) using the single-stain viability dye TO-PRO(®)-3 iodide as a fast and precise alternative. Mature biofilms of Candida albicans and Escherichia coli were used to optimize biofilm removal and dissociation, as a single-cell suspension is needed for accurate FCM enumeration. To assess the feasibility of FCM quantification of biofilms, E. coli and C. albicans biofilms were analyzed using FCM and crystal violet staining at different time points. A combination of scraping and rinsing proved to be the most efficient technique for biofilm removal. Sonicating for 10 min eliminated the remaining aggregates, resulting in a single-cell suspension. Repeated FCM measurements of biofilm samples revealed a good intraday precision of approximately 5 %. FCM quantification and the crystal violet assay yielded similar biofilm growth curves for both microorganisms, confirming the applicability of our technique. These results show that FCM using TO-PRO(®)-3 iodide as a single-stain viability dye is a valid fast alternative for the quantification of viable cells in a biofilm.

Various clinical and research settings require fast and reliable quantification of bacterial suspensions. The viable plate count method (VPC) is generally seen as 'the gold standard' for bacterial enumeration. However, VPC-based quantification of anaerobes such as Porphyromonas gingivalis is time-consuming due to their stringent growth requirements and shows poor repeatability. Comparison of VPC, turbidity measurement and TaqMan-based qPCR demonstrated that qPCR possesses important advantages regarding speed, accuracy and repeatability.

The promising anti-adhesion in vitro results could not be confirmed in the mouse model, even after the highest attainable exposure. It is concluded that raw or defatted milk fat globule membrane fractions do not have any prophylactic or therapeutic potential against Helicobacter infection.

The proposed technique allows detection of very low quantities of H. pylori DNA in biological samples. In laboratory animal models, detailed monitoring of infection and complete clearance of infection can be demonstrated thanks to the low detection limit.