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Understanding how bacteria adapt their social behavior to environmental changes is of crucial importance from both biological and clinical perspectives. Staphylococcus aureus is among the most common infecting agents in orthopedics, but its recalcitrance to the immune system and to antimicrobial treatments in the physiological microenvironment are still poorly understood. By means of optical and confocal microscopy, image pattern analysis, and mathematical modeling, we show that planktonic biofilm-like aggregates and sessile biofilm lifestyles are two co-existing and interacting phases of the same environmentally adaptive developmental process and that they exhibit substantial differences when S. aureus is grown in physiological fluids instead of common lab media. Physicochemical properties of the physiological microenvironment are proposed to be the key determinants of these differences. Besides providing a new tool for biofilm phenotypic analysis, our results suggest new insights into the social behavior of S. aureus in physiological conditions and highlight the inadequacy of commonly used lab media for both biological and clinical studies of bacterial development. [ABSTRACT FROM AUTHOR]

Recent in vitro studies reported the exceptional ability of some bacterial species to form biofilm-like aggregates in human and animal synovial fluids (SF), but evidences from infected clinical samples are still lacking. In this study, we investigated whether this bacterial phenotype was present in infected SFs collected from joint infections and if it was maintained in in vitro settings. SFs sent for culture to the Laboratory of Microbiology of our institute were directly analyzed by means of confocal laser scanning microscopy (CLSM), and the infective agents were isolated for further in vitro tests. Moreover, sterile SF was collected from patients who did not receive previous antibiotic therapy to investigate the formation of bacterial aggregates, together with biofilm and matrix production on a titanium surface. Finally, antibiotic susceptibility studies were performed by using bovine SF. Four Staphylococcus aureus , one Staphylococcus lugdunensis , and one Prevotella bivia strain were identified in the infected SFs. The CLSM analysis showed that all staphylococci were present as a mixture of single cells and bacterial clumps surrounded by an exopolymeric substance, which comprised SF-derived fibrin, while all P. bivia cells appeared separated. Despite that, differences in the ability to aggregate between S. aureus and S. lugdunensis were observed in clinical SFs. These different phenotypes were further confirmed by in vitro growth, even though the application of such ex vivo approach lead all staphylococci to form exceptionally large microbial aggregates, which are several folds bigger than those observed in clinical samples. Planktonic aggregates challenged for antibiotic susceptibility revealed a sharp increase of recalcitrance to the treatments. Although this is still at a preliminary stage, the present work confirmed the ability of staphylococci to form free-floating biofilm-like aggregates in infected SF from patients with joint infections. Furthermore, the obtained results pointed out that future in vitro research on joint infections will benefit from the use of human- or animal-derived SF. Even though this approach should be carefully validated in further studies comprising a larger microbial population, these findings pose new challenges in the treatment of infected native and prosthetic joints and for the approach to new investigations. [ABSTRACT FROM AUTHOR]

Background: Infections of the ears, paranasal sinuses, nose and throat are very common and represent a serious issue for the healthcare system. Bacterial biofilms have been linked to upper respiratory tract infections and antibiotic resistance, raising serious concerns regarding the therapeutic management of such infections. In this context, novel strategies able to fight biofilms may be therapeutically beneficial and offer a valid alternative to conventional antimicrobials. Biofilms consist of mixed microbial communities, which interact with other species in the surroundings and communicate through signaling molecules. These interactions may result in antagonistic effects, which can be exploited in the fight against infections in a sort of "bacteria therapy". Streptococcus salivarius and Streptococcus oralis are α-hemolytic streptococci isolated from the human pharynx of healthy individuals. Several studies on otitis-prone children demonstrated that their intranasal administration is safe and well tolerated and is able to reduce the risk of acute otitis media. The aim of this research is to assess S. salivarius 24SMB and S. oralis 89a for the ability to interfere with biofilm of typical upper respiratory tract pathogens.Methods: To investigate if soluble substances secreted by the two streptococci could inhibit biofilm development of the selected pathogenic strains, co-cultures were performed with the use of transwell inserts. Mixed-species biofilms were also produced, in order to evaluate if the inhibition of biofilm formation might require direct contact. Biofilm production was investigated by means of a spectrophotometric assay and by confocal laser scanning microscopy.Results: We observed that S. salivarius 24SMB and S. oralis 89a are able to inhibit the biofilm formation capacity of selected pathogens and even to disperse their pre-formed biofilms. Diffusible molecules secreted by the two streptococci and lowered pH of the medium revealed to be implied in the mechanisms of anti-biofilm activity.Conclusions: S. salivarius 24SMB and S. oralis 89a possess desirable characteristics as probiotic for the treatment and prevention of infections of the upper airways. However, the nature of the inhibition appear to be multifactorial and additional studies are required to get further insights. [ABSTRACT FROM AUTHOR]

Background: The purpose of this in vitro study was to evaluate the antibiofilm activity of a novel air-polishing powder consisting of erythritol and chlorhexidine, assessing its ability to reduce previously grown microbial biofilm and to prevent biofilm formation on titanium surfaces.Methods: Clinical strains of Staphylococcus aureus, Pseudomonas aeruginosa, Bacteroides fragilis and Candida albicans isolated from peri-implantitis lesions were used. Biofilm was grown on sandblasted titanium discs and treated with erythritol/chlorhexidine. The antimicrobial activity was evaluated by determining the minimum inhibitory concentration and the minimum microbicidal concentration. The antibiofilm activity was assessed by semiquantitative spectrophotometric assay and by confocal laser scanning microscopy.Results: Erythritol/chlorhexidine displayed an inhibitory and a microbicidal activity against all the tested strains. The spectrophotometric analysis showed that the treatment was effective in both reducing the previously developed biofilm and decreasing biofilm formation on titanium surfaces. Confocal laser scanning microscopy analysis showed a significant reduction of the total biofilm volume, with an increase of the percentage of dead cells of all the microorganisms tested.Conclusions: Erythritol/chlorhexidine displayed significant antimicrobial and antibiofilm activity against microorganisms isolated from peri-implantitis lesions. Due to its properties, it might represent a promising approach for the prevention and treatment of peri-implant diseases associated to microbial biofilm infections. [ABSTRACT FROM AUTHOR]

Over the years, various techniques have been proposed for the quantitative evaluation of microbial biofilms. Spectrophotometry after crystal violet staining is a widespread method for biofilm evaluation, but several data indicate that it does not guarantee a good specificity, although it is rather easy to use and cost saving. Confocal laser microscopy is one of the most sensitive and specific tools to study biofilms, and it is largely used for research. However, in some cases, no quantitative measurement of the matrix thickness or of the amount of embedded microorganisms has been performed, due to limitation in availability of dedicated software. For this reason, we have developed a protocol to evaluate the microbial biofilm formed on sandblasted titanium used for orthopaedic implants, that allows measurement of biomass volume and the amount of included cells. Results indicate good reproducibility in terms of measurement of biomass and microbial cells. Moreover, this protocol has proved to be applicable for evaluation of the efficacy of different anti-biofilm treatments used in the orthopaedic setting. Summing up, the protocol here described is a valid and inexpensive method for the study of microbial biofilm on prosthetic implant materials. [ABSTRACT FROM AUTHOR]

Objective. Bacterial biofilms have been detected in biopsies of the adenoid and middle ear mucosa of otitis-prone children and children with chronic middle otitis media. However, the invasiveness of biopsy makes it unsuitable for routine clinical practice, especially in pediatrics. This study aimed to investigate nasopharyngeal biofilm-producing otopathogens (BPOs) of nasopharyngeal swabs (NPS) in children with a history of nonsevere recurrent acute otitis media (RAOM) and healthy controls.Study Design. A cross-sectional study with planned data collection.Setting. University of Milan.Subjects and Methods. Transoral NPS were taken from infants and children aged 10 months to 11 years with nonsevere RAOM or healthy controls without adenoid hypertrophy. Nasopharyngeal colonization by otopathogens was assessed by means of microbiological cultures and standard bacterial identification, as well as nasopharyngeal BPOs by means of spectrophotometric analysis.Results. The study involved 113 children (56.6% males; median age 40 months; range, 10-132 months): 58 with a history of nonsevere RAOM (51.3%) and 55 controls (48.7%). Otopathogens were significantly more frequently detected in the RAOM group (24/58, 41.4%) than in controls (8/55, 14.5%; P = .003); the main pathogens were respectively Haemophilus influenzae (12/24, 50.0%) and Streptococcus pyogenes (3/8, 37.5%). Nasopharyngeal BPOs were more frequently isolated in the RAOM group (17/58, 29.3%) than in controls (6/55, 10.9%; P = .02). H influenzae (12/17, 70.6%) was confirmed as the main pathogen in the RAOM group.Conclusion. The presence of nasopharyngeal BPOs is an important factor favoring RAOM; it is therefore useful investigating biofilms even in children with nonsevere recurrences of AOM without adenoid hypertrophy. [ABSTRACT FROM AUTHOR]

Abstract Sialolithiasis represents the most common disorders of salivary glands in middle-aged patients. It has been hypothesized that the retrograde migration of bacteria from the oral cavity to gland ducts may facilitate the formation of stones. Thus, in the present study, a microbiome characterization of salivary calculi was performed to evaluate the abundance and the potential correlations between microorganisms constituting the salivary calculi microbiota. Our data supported the presence of a core microbiota of sialoliths constituted principally by Streptococcus spp., Fusobacterium spp. and Eikenella spp., along with the presence of important pathogens commonly involved in infective sialoadenitis. [ABSTRACT FROM AUTHOR]