In vitro adhesion of commensal and pathogenic bacteria to commercial titanium implants with different surfaces.

Although dental implants have undergone impressive evolution in recent years, periimplantitis still remains a relevant problem and information on the susceptibility of commercial implants to bacterial colonization is insufficient. This work evaluated the susceptibility of different commercial implants to bacterial colonization, to identify key features for good performances. Twenty-four implants, produced with different technologies, were colonized with 9 bacterial strains following pre-conditioning with culture medium, or saliva or serum proteins and adherent bacteria were enumerated by Real Time quantitative PCR. The studied implants differed significantly for susceptibility to bacterial adhesion. Pre-conditioning of surfaces affected adhesion assays in a species specific manner. Although surface topography influenced bacterial adhesiveness, implants produced by different manufacturers with comparable technologies showed great variability of results. These data demonstrate that susceptibility of implants to bacterial colonization is influenced by productive technologies (in a surface topography proportional manner) and by the productive environment. In choosing an implant the clinician should rely upon specific experimental studies, because surface characteristics alone cannot predict susceptibility to colonization by pathogenic bacteria. Tests should include assays performed in the medium of culture and in the presence of serum proteins.

Combination therapy including serratiopeptidase improves outcomes of mechanical-antibiotic treatment of periimplantitis.

This study was designed as a retrospective analysis of clinical outcomes of cases of periimplantitis treated by mechanical debridement and the administration of antibiotics combined or not with the administration of either the proteolytic enzyme serratiopeptidase (SPEP) or non-steroidal anti-inflammatory drugs (NSAIDs). Clinical charts of 544 partially edentulous patients treated for periimplantitis between June 1996 and December 2010 were analyzed to obtain clinical data of the affected implants just before the beginning of treatment and 12 months later to evaluate the outcomes of combined mechanical antibiotic treatment alone or in combination with the co-administration of the anti-inflammatory SPEP or NSAIDs. The comparative analysis revealed that therapeutic outcomes were significantly different in the three groups. Failure rate in the group that received SPEP (6 percent) was significantly lower compared to the group that received NSAIDS (16.9 percent; P less than 0.01) and to the group that received no anti-inflammatory therapy (18.9 percent; P less than 0.01). Treatment including SPEP was associated with significantly better healing also when successful treatments alone were considered. The data reported in this paper strongly support the hypothesis that SPEP is a valid addition to protocols for the combined therapy of peri-implantitis. In fact, it allows to enhance success rates significantly and also favors better tissue repair around successfully treated implants as compared to other regimens.

Adhesion and biofilm formation by periodontopathogenic bacteria on different commercial brackets.

AIM: To compare early bacterial adhesion and biofilm formation of common and uncommon periodontal pathogens on a variety of commercial brackets in vitro. MATERIALS AND METHODS: In vitro adhesion and biofilm formation of 4 bacterial strains on 15 different commercial brackets, in standard culture mediums with and without addition of either serum or human saliva was evaluated by quantitative real time PCR after extraction of bacterial DNA. RESULTS: Materials significantly influenced bacterial adhesiveness in a species-specific way. Titanium and gold brackets constantly yielded the lowest values with all tested bacteria and in all tested conditions. Bracket materials and medium of growth significantly influenced biofilm formation. CONCLUSION: Materials and environmental conditions significantly influence biofilm formation by periodontal pathogens at the surface of brackets. Whenever possible brackets should be kept far from the gingival margin and if this is not possible, brackets made of gold, titanium, and ceramic should be preferentially used.

Adhesion and biofilm formation by oral streptococci on different commercial brackets.

AIM: To compare early bacterial adhesion and biofilm formation in vitro by different oral streptococci on a variety of commercial brackets. METHODS: Adhesion and biofilm formation in vitro of 6 Streptococcus spp. on 15 different commercial brackets, in standard culture medium and in human saliva were evaluated by the MTT reduction assay. RESULTS: Significant differences were evidenced in both early adhesion and biofilm formation among the studied brackets and between the two conditions of growth. Gold brackets resulted less prone to colonisation compared to composite brackets. The growth rates of the tested species on the different tested materials were significantly different. CONCLUSION: The adopted experimental plan, dissecting the two phases of plaque formation on different brackets in different conditions, showed that composite brackets are more susceptible to adhesion and colonisation by streptococci, while the remaining tested brackets did not show differences that could be clinically relevant. Data suggest that different personal behaviours affecting the oral environment could significantly affect colonisation of brackets by oral streptococci.

Influence of oral conditions on colonization by highly toxigenic Staphylococcus aureus strains.

OBJECTIVES: As the oral cavity is regarded as a relevant site for Staphylococcus aureus colonization and interhuman transmission, this study aimed to investigate whether different oral conditions influence the rates of S. aureus oral carriage and genetic characters of S. aureus isolates. SUBJECTS AND METHODS: Staphylococcus aureus was searched in samples collected from cheek, gingival margin, and anterior nares of 45 healthy subjects, 27 periodontitis affected subjects, and 29 subjects with fixed prosthetic restorations. Isolates were screened for 17 genetic determinants, and Partial Least Square Discriminant Analysis was performed to evaluate whether specific characters correlated with oral condition or site of isolation. RESULTS: The three subject groups showed comparable nasal carriage rates but, both the periodontitis and prosthetic restoration groups showed significantly higher oral carriage rates, as compared to healthy subjects (P = 0.01 and 0.02, respectively). Moreover, periodontitis affected subjects hosted strains possessing a distinct genotypic and phenotypic background, characterized by the presence of a larger number of exotoxins encoding genes. CONCLUSIONS: These data confirm that the oral cavity is an important site of S. aureus colonization and demonstrate that conditions modifying the oral environment, as the presence of periodontitis and of fixed prosthetic restorations, promote S. aureus carriage and may favor the spread of more pathogenic strains.

Microbiological and molecular characterization of nosocomial and community Staphylococcus aureus isolates.

This study aimed to evaluate the incidence of Staphylococcus aureus infections in different departments of Belcolle Hospital in Viterbo and the surrounding area between January 2003 and June 2008. Isolates of methicillin-resistant S. aureus (MRSA) recovered in this time interval were characterized by microbiological and molecular methods to evaluate the reliability of simple criteria to distinguish between hospital-acquired and community-acquired isolates. MRSA accounted for 33% of all S. aureus, with a significantly higher prevalence in isolates from nosocomial infections. MRSA isolates were assayed by PCR for the presence of 13 genes associated with virulence, agr type and SCCmec type. Cumulative data were analysed by partial least square discriminant analysis and a clear correlation was demonstrated between genetic profiles and classification of isolates as hospital or community acquired according to simple temporal criteria. Nosocomial MRSA isolates from blood samples showed significantly higher genetic diversity than other nosocomial isolates. Our data confirm the existence of significant differences between community- and hospital-acquired MRSA isolates.

Adipocytokines and bone mineral density in adolescent female athletes.

AIM: To evaluate the relationships of visfatin, adiponectin and leptin with bone mineral density (BMD) and bone mineral content (BMC) in adolescent female athletes with different training patterns. METHODS: This study involved 170 healthy 13-15-year-old girls divided into six groups based on activity: sport games (i.e. basketball, volleyball, badminton) (n=49), track sprint (n=24), rhythmic gymnastics (n=23), swimming (n=24), cross-country skiing (n=17) and sedentary controls (n=33). BMD and BMC at femoral neck and lumbar spine (L2-L4) were measured using dual-energy X-ray absorptiometry. Visfatin, adiponectin, leptin, insulin and glucose were measured, and the insulin resistance index was calculated using homeostasis model assessment. RESULTS: There were no relationships found between visfatin concentrations and bone mineral parameters in adolescent female athletes or controls. Adiponectin was inversely correlated to BMD and BMC of femoral neck and lumbar spine (r=-0.47-0.62) in the swimmer group only, but after adjustments for age, height and body mass these associations disappeared. Leptin concentrations correlated with bone mineral parameters even after adjusting for age, height and body mass (r=0.42-0.63) in the gymnast group only. CONCLUSION: We may conclude that after adjustment, leptin is the only adipokine of those measured that correlates to femoral neck and lumbar spine BMD and femoral neck BMC in the rhythmic gymnast group.

Herpesviruses and periodontal disease: a cautionary tale.

Periodontitis is an inflammatory disease of bacterial origin, characterized by an inconstant progression of lesions affecting the tooth supporting tissues. In spite of more than half a century of research efforts, the clinician still lacks any specific molecular or microbial diagnostic tool to predict the progression of periodontal lesions. Recently, several reports have proposed a role for some herpesviruses in the etiology of destructive phases of periodontitis. This paper critically analyzes these data in the light of consolidated knowledge that was developed in the characterization of virus-bacteria cooperative interactions, and proposes new topics of investigation to clarify the role of herpesviral infections in periodontitis and their potential predictive role as markers of progression.

Microbial contamination of brand new nickel-titanium endodontic instruments.

AIMS: This study aims to estimate the microbial presence on the surface of different brand new NiTi endodontic instruments for clinical use. MATERIAL AND METHODS: Eleven different types of NiTi rotary endodontic instruments, obtained from their fresh opened original packages, were assigned to three different groups, according to packaging type and sterilization and tested for bacterial contamination. Isolated bacteria were identified by using standard microbiological methods and then counted. Differences observed in groups were analyzed statistically by using the one-way analysis of variance (ANOVA) for dependent samples and the Tukey HSD post hoc test. RESULTS: Statistical differences were found between instruments delivered in plastic boxes which bacterial count resulted higher than those obtained from instruments delivered in blisters (p<0.01). CONCLUSIONS: Some brand new endodontic instruments showed degrees of bacterial contamination that both quantitatively and qualitatively deserve to be considered in clinical procedures.

Effect of the inoculum size on susceptibility tests performed on sessily growing bacteria.

It has been clearly established that the inoculum size greatly affects the results of antibiotic susceptibility tests performed in both liquid and solid media in standard laboratory growth conditions (i.e. planktonic). Recently methods were developed to perform antibiotic susceptibility tests on bacteria growing in sessile conditions. The present study investigates the effect of the inoculum size on results obtained by these methods. Results show that the inoculum size does not affect tests performed in sessile conditions. A simple and reliable method is proposed to be applied to routine microbiological laboratory procedures.

Effect of slime production on the antibiotic susceptibility of isolates from prosthetic infections.

The antibacterial activity of 6 antibiotics towards 10 gram-positive and 6 gram-negative glycocalyx-producing strains, has been evaluated by employing a method which partially simulates the in vivo colonization of prosthetic devices. The results showed that routine antibiotic sensitivity tests are not predictive about the response of the glycocalyx-embedded bacteria, and that prophylaxis may be useful with ofloxacin and clindamycin, before placing a prosthetic device. Once bacterial colonization had already occurred, however, none of the tested antibiotics was able to eradicate the sessile bacterial form. The minimum bactericidal concentration (MBC) values, indeed, were much higher than those determined on the planktonic form, and were much higher than serum and tissue levels that can be reached in vivo.

Taxonomy and identification of periodontopathogenic bacteria and related species.

The tumultuous evolution of oral and periodontal microbiology has focussed the attention of many researchers on the necessity to clarify the taxonomic position and identification criteria of putative periodontopathogens, in order to elucidate the role played by each species in the pathogenesis of periodontal disease. Many of the most important periodontopathogens recently underwent a radical reclassification process that may create some confusion and certainly deserves an accurate analysis aimed at focussing the actual situation of these bacteria. The taxonomic evolution and identification criteria of species of the genera Bacteroides, Prevotella, Porphyromonas and Actinobacillus is here analyzed in detail to clarify this recent evolutive taxonomic process, and to explain the importance of molecular studies for both taxonomic and identification purposes in this field.

A rapid staining procedure to demonstrate glycocalyx production and bacterial biofilms.

A novel staining procedure to demonstrate glycocalyx production by clinical isolates is presented. The short times required, specificity and sensitivity suggest that the staining could be applied to routine in vitro diagnostic procedures.

Chasing bacteria within the cells using levofloxacin-loaded hyaluronic acid nanohydrogels.

In the present work, an innovative approach based on the delivery of levofloxacin (LVF) from polysaccharide nanohydrogels for the treatment of bacterial intracellular infections is described. The nanohydrogels (NHs) were obtained by self-assembling of the hyaluronic acid-cholesterol amphiphilic chains in aqueous environment. LVF, a fluoroquinolone antibiotic scarcely efficient in intracellular infections, was entrapped within such NHs by nanoprecipitation, thus forming a drug delivery system (LVF-NHs) that was tested for its activity on different bacteria strains. The MIC values of levofloxacin-loaded nanohydrogels were determined for Staphylococcus aureus and Pseudomonas aeruginosa strains and compared to those obtained using free LVF. The intracellular antimicrobial activity of LVF-NHs and free LVF was compared on HeLa epithelial cell line infected by the above mentioned bacteria, and the increase in antibacterial efficacy of LVF-NHs with respect to that of free LVF was evidenced. The obtained results allow to conclude that this new approach can be considered as really promising method for intracellular infection treatments.

Preparation and characterization of antimicrobial wound dressings based on silver, gellan, PVA and borax.

Silver-loaded dressings are designed to provide the same antimicrobial activity of topical silver, with the advantages of a sustained silver release and a reduced number of dressing changes. Moreover, such type of dressing must provide a moist environment, avoiding fiber shedding, dehydration and adherence to the wound site. Here we describe the preparation of a novel silver-loaded dressing based on a Gellan/Hyaff(®) (Ge-H) non woven, treated with a polyvinyl alcohol (PVA)/borax system capable to enhance the entrapment of silver in the dressing and to modulate its release. The new hydrophilic non woven dressings show enhanced water uptake capability and slow dehydration rates. A sustained silver release is also achieved. The antibacterial activity was confirmed on Staphylococcus aureus and Pseudomonas aeruginosa.

Diffusion of meticillin-resistant Staphylococcus aureus USA300 strains in central Italy.

Meticillin-resistant Staphylococcus aureus (MRSA) is an outstanding, clonally evolving pathogen that in recent years, under the selective pressure of antibiotics, has acquired the crucial ability to infect people outside of hospitals. MRSA USA300 has progressively become synonymous with severe community-associated staphylococcal disease worldwide. Whilst spreading worldwide, these clones have progressively acquired resistance to several antibiotics and have gained the ability to cause infections in hospital settings. Recently, USA300-related strains showing resistance to several antibiotics have been isolated from community-acquired infections in Italy. This paper reports the high frequency of isolation of USA300-related strains both from community- and hospital-acquired infections in central Italy as well as their genotypic characteristics and antibiotic susceptibility. Analysis of these characteristics by partial least squares discriminant analysis enabled it to be demonstrated that whilst moving from the community to the hospital setting these isolates underwent an adaptive process that generated clones showing distinctive characteristics. These observations further support the hypothesis that the threatening generation of strains combining both resistance and virulence is becoming a reality, and stress the necessity of constant molecular epidemiological surveillance of MRSA.

Violacein and biofilm production in Janthinobacterium lividum.

AIMS: To analyse the environmental stimuli modulating violacein and biofilm production in Janthinobacterium lividum. METHODS AND RESULTS: Violacein and biofilm production by J. lividum DSM1522(T) was assayed in different growth conditions. Our data suggest that violacein and biofilm production is controlled by the carbon source, being inhibited by glucose and enhanced by glycerol. J. lividum produced violacein also in the presence of different sub-inhibitory concentrations of ampicillin. As opposite, the production of N-acylhomoserine lactone(s), quorum sensing regulators was shown to be positively regulated by glucose. Moreover, violacein-producing cultures of J. lividum showed higher CFU counts than violacein-nonproducing ones. CONCLUSIONS: Taken together, our results suggest that violacein and biofilm production could be regulated by a common metabolic pathway and that violacein as well as biofilm could represent a response to environmental stresses and a key factor in the survival mechanisms of J. lividum. SIGNIFICANCE AND IMPACT OF THE STUDY: Although several recent studies disclosed a number of interesting biological properties of violacein, few data are reported on the physiologic function of violacein in J. lividum. This paper adds new information on the complex mechanisms allowing and regulating bacterial life in hostile environments.

Proteolytic enzymes: a new treatment strategy for prosthetic infections?

Among the different mechanisms of bacterial resistance to antimicrobial agents that have been studied, biofilm formation is one of the most widespread. This mechanism is frequently the cause of failure in the treatment of prosthetic device infections, and several attempts have been made to develop molecules and protocols that are able to inhibit biofilm-embedded bacteria. We present data suggesting the possibility that proteolytic enzymes could significantly enhance the activities of antibiotics against biofilms. Antibiotic susceptibility tests on both planktonic and sessile cultures, studies on the dynamics of colonization of 10 biofilm-forming isolates, and then bioluminescence and scanning electron microscopy under seven different experimental conditions showed that serratiopeptidase greatly enhances the activity of ofloxacin on sessile cultures and can inhibit biofilm formation.

Periodontal regeneration procedures may induce colonization by glycocalyx-producing bacteria.

Sixteen patients that underwent periodontal regeneration procedures by implantation of Goretex membranes were studied to evaluate the microbiota that colonized membranes. The microbiological follow-up showed that colonization was principally due to opportunist gram-negative glycocalyx-producing bacteria. The comparison of the microbial flora of treated and untreated sites showed the influence of surface characteristics on the quality of the resident microbiota. These findings suggest the necessity for developing efficient prohylactic protocols for these cases.

Invasion of human cultured cells by Prevotella nigrescens.

Invasion of gingival and junctional epithelial cells has been recently proposed as a potentially relevant mechanism in the pathogenesis and recurrence of periodontal disease. The gram negative anaerobe Prevotella nigrescens was shown to be involved in the development of periodontal lesions in man, suggesting a possible involvement of invasivity as a mean to circumvent the host immune surveillance and other hostile factors. Appropriately designed invasion assays demonstrated that P. nigrescens efficiently invades human epithelial cells, through a mechanism whose efficiency is influenced by the phase of growth, by the multiplicity of infection, and by the cell line used, and that requires microfilament integrity, but is not affected by an impairment of microtubule organization. Intracellular replication assays suggested that P. nigrescens probably multiplies within Kb epithelial cells, causing extensive cell alterations. Invasion of gingival epithelial cells could consequently be a basic step in the virulence mechanism of the species.