The use of adhesive poly(HEMA)-based hydrogels is standard practice in dental restorative procedures. Microorganisms, which potentially can cause oral pathologies, may colonize these polymers. In the present work, bacterial adhesion to polymers prepared with 2-hydroxyethyl methacrylate (HEMA) and to different molar ratios of 2-acrylamido-2-methylpropane-sulfonic acid (AMPS) and/or to 2-methacryloyloxyethyl-tri-methyl-ammonium chloride (METAC) co-monomers were tested. A colorimetric assay system that utilizes the Microbo revelation medium (Microbo srl, Rome, Italy) for microbial counts is shown to be capable of counting the number of adherent bacterial cells without removing them from polymer surfaces. In conditions that mimic those present in the oral cavity, similar bacterial adhesion percentages on the same polymer were observed with the different bacteria belonging to both gram-positive and gram-negative genera, such as Streptococcus sobrinus and Streptococcus oralis (resident microorganisms in the oral cavity) and Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa (transient microorganisms in the oral cavity). It is determined that the physico-chemical characteristics of poly(HEMA)-based hydrogels are the major factors promoting bacterial adhesion, which increased with increasing water content in the swollen polymers, reaching maximal values on the cationic polymers.
Streptococcus mutans, a gram-positive immobile bacterium, is an oral pathogen considered to be the principal etiologic agent of dental caries. Although some researches suggest that trace metals, including iron, can be associated with dental caries, the function of salivary iron and lactoferrin in the human oral cavity remains unclear. The data reported in this study indicates that iron-deprived saliva (Fe3+ < 0.1 microM) increases S. mutans aggregation and biofilm formation in the fluid and adherent phases as compared with saliva (Fe3+ from 0.1 to 1 microM), while iron-loaded saliva (Fe3+ > 1 microM) inhibits both phenomena. Our findings are consistent with the hypothesis that S. mutans aggregation and biofilm formation are negatively iron-modulated as confirmed by the different effect of bovine lactoferrin (bLf), added to saliva at physiological concentration (20 microg/ml) in the apo- or iron-saturated form. Even if saliva itself induces bacterial aggregation, iron binding capability of apo-bLf is responsible for the noticeable increase of bacterial aggregation and biofilm development in the fluid and adherent phases. On the contrary, iron-saturated bLf decreases aggregation and biofilm development by supplying iron to S. mutans. Therefore, the iron-withholding capability of apo-Lf or native Lf is an important signal to which S. mutans counteracts by leaving the planktonic state and entering into a new lifestyle, biofilm, to colonize and persist in the human oral cavity. In addition, another function of bLf, unrelated to its iron binding capability, is responsible for the inhibition of the adhesion of S. mutans free, aggregated or biofilm on abiotic surfaces. Both these activities of lactoferrin, related and unrelated to the iron binding capability, could have a key role in protecting the human oral cavity from S. mutans pathogenicity.
Shigella and enteroinvasive Escherichia coli (EIEC) strains secrete virulence proteins by a complex machinery called the type III secretion (TTS) apparatus. Secretion of virulence proteins is a tightly-regulated phenomenon such that the TTS system is weakly active when bacteria are grown in common laboratory media. Activation of the TTS system is triggered by contact with eukaryotic cells, or can be artificially stimulated by the addition of Congo red dye to the growth medium. Exploiting the ability of bovine lactoferrin (bLf) to bind iron we have found that the TTS of EIEC strain HN280 seems to be activated in conditions of low-iron availability, obtained by incubation of bacteria with bLf enclosed within a dialysis bag. Activation of secretion was assessed by measuring the release of IpaB and C, chosen as reporters of secreted virulence proteins. The contribution of small bLf-derived components, diffusing across the dialysis membrane, in the release of Ipa proteins has also been determined. Activation of secretion was not due to bLf-induced damage of the HN280 outer membrane and was not associated with increased transcription of the mxi operon. Thus, low-iron availability might be an environmental signal perceived by enteroinvasive micro-organisms in order to modulate secretion of virulence proteins.
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