Real-Time Microsensor Measurement of Local Metabolic Activities in Ex Vivo Dental Biofilms Exposed to Sucrose and Treated with Chlorhexidine

Dental biofilms are characterized by structural and functional heterogeneity. Due to bacterial metabolism, gradients develop and diverse ecological microniches exist. The aims of this study were (i) to determine the metabolic activity of microorganisms in naturally grown dental biofilms ex vivo by measuring dissolved oxygen (DO) and pH profiles with microelectrodes with high spatial resolution and (ii) to analyze the impact of an antimicrobial chlorhexidine (CHX) treatment on microbial physiology during stimulation by sucrose in real time. Biofilms were cultivated on standardized human enamel surfaces in vivo. DO and pH profiles were measured in a flow cell system in sterile human saliva, after sucrose addition (10%), again after alternative treatment of the sucrose exposed biofilms with CHX (0.2%) for 1 or 10 min or after being killed with paraformaldehyde (4%). Biofilm structure was visualized by vitality staining with confocal microscopy. With saliva as the sole nutrient source oxygen consumption was high within the superficial biofilm layers rendering deeper layers (>220 μm) anoxic. Sucrose addition induced the thickness of the anaerobic zone to increase with a concurrent decrease in pH (7.1 to 4.4). CHX exposure reduced metabolic activity and microbial viability at the biofilm surface and drove metabolic activity deeper into the biofilm. CHX treatment led to a reduced viability at the biofilm surface with minor influence on overall biofilm physiology after 1 min; even after 10 min there was measurable respiration and fermentation inside the biofilm. However, the local microenvironment was more aerated, less acidogenic, and presumably less pathogenic

Chlorhexidine Substantivity on Salivary Flora and Plaque-Like Biofilm: An In Situ Model

This work was supported by project FIS PI11/01383 from Carlos III Institute of Health (Ministry of Economy and Competitiveness, Madrid, Spain

Prevalence of oral pathologic findings in an ancient pre-Columbian archeologic site in the Atacama Desert

OBJECTIVE: To determine the prevalence of oral pathologic findings in an ancient culture that inhabited the Atacama Desert. MATERIALS AND METHODS: A systematic examination was performed on the remains of 83 individuals unearthed from a prehistoric burial ground. A total of 57 skeletal remains achieved appropriate inclusion criteria, from which estimated age at death, gender, ante- and postmortem tooth loss, prevalence and location of caries, apical periodontitis sequela, alveolar bone resorption and attrition were recorded. RESULTS: From the analyzed skeletal remains (13 male, 22 female and 22 not identifiable), the mean age estimated was 29.9 +/- 13.8 years. A total of 89.4% of them presented permanent dentition with a mean ante-mortem tooth loss of 9.0 teeth and a postmortem mean tooth loss of 14.4 teeth per subject. In all, 46.4% of the postmortem remaining permanent teeth (n = 237) showed caries lesions. Interproximal caries was most frequently observed (31.5%), followed by occlusal (25.9%) and cervical caries (19.4%). Root remnants were found in 23.1% of the cases. In addition, 58.0% of the adults presented attrition, 26.0% signs of apical periodontitis and 44.0% loss of alveolar bone support >5 mm. CONCLUSION: The remains of jaws and teeth of the individuals examined in this study presented sequelae of severe oral health damage due to caries and periodontal disease

Biofilm plaque and hydrodynamic effects on mass transfer, fluoride delivery and caries

Background. The biofilm concept of dental plaque now is widely accepted in the dental clinic, particularly with respect to its importance to oral hygiene. A number of reviews have focused on the microbial ecology of biofilm with regard to oral health; however, there has been less focus on how the interaction of biofilms and hydrodynamics with mass transfer (the movement of molecules and particulates) and physiological processes may relate to caries. Types of Studies Reviewed. The authors reviewed reports in the microbiology and dental literature addressing microbiological, engineering and clinical aspects of biofilms with respect to mass transport and microbial physiology, with an emphasis on fluoride ions (F−). Conclusions and Practical Implications. These data illustrate how dental plaque biofilms may affect the delivery of cariogenic agents, such as sucrose, or anticariogenic agents, such as F−, into and out of the biofilm, with subsequent consequences for the development of physio-chemical microenvironments at the tooth surface. Increasing the flow rate in an overlying fluid (such as saliva or mouthrinse) increases transport from the fluid into and through biofilms. Increasing the delivery of anticariogenic agents such as F− into the plaque biofilm, by generating strong fluid flows, may be a useful strategy for enhancing the anticaries effects of F− in areas of the mouth where complete biofilm removal is not possible with routine daily cleaning techniques

Real time microsensor measurement of local metabolic activities in ex vivo dental biofilms treated with chlorhexidine

Dental biofilms are characterized by structural and functional heterogeneity. Due to bacterial metabolism, gradients develop and diverse ecological microniches exist. The aims of this study were (i) to determine the metabolic activity of microorganisms in naturally grown dental biofilms ex vivo by measuring dissolved oxygen (DO) and pH profiles with microelectrodes with high spatial resolution and (ii) to analyze the impact of an antimicrobial chlorhexidine (CHX) treatment on microbial physiology during stimulation by sucrose in real time. Biofilms were cultivated on standardized human enamel surfaces in vivo. DO and pH profiles were measured in a flow cell system in sterile human saliva, after sucrose addition (10%), again after alternative treatment of the sucrose exposed biofilms with CHX (0.2%) for 1 or 10 min or after being killed with paraformaldehyde (4%). Biofilm structure was visualized by vitality staining with confocal microscopy. With saliva as the sole nutrient source oxygen consumption was high within the superficial biofilm layers rendering deeper layers (&gt;220 mum) anoxic. Sucrose addition induced the thickness of the anaerobic zone to increase with a concurrent decrease in pH (7.1 to 4.4). CHX exposure reduced metabolic activity and microbial viability at the biofilm surface and drove metabolic activity deeper into the biofilm. CHX treatment led to a reduced viability at the biofilm surface with minor influence on overall biofilm physiology after 1 min; even after 10 min there was measurable respiration and fermentation inside the biofilm. However, the local microenvironment was more aerated, less acidogenic, and presumably less pathogenic.<br/

The oral microbiota is a reservoir for antimicrobial resistance: resistome and phenotypic resistance characteristics of oral biofilm in health, caries, and periodontitis

Abstract Background Antimicrobial resistance (AMR) is an ever-growing threat to modern medicine and, according to the latest reports, it causes nearly twice as many deaths globally as AIDS or malaria. Elucidating reservoirs and dissemination routes of antimicrobial resistance genes (ARGs) are essential in fighting AMR. Human commensals represent an important reservoir, which is underexplored for the oral microbiota. Here, we set out to investigate the resistome and phenotypic resistance of oral biofilm microbiota from 179 orally healthy (H), caries active (C), and periodontally diseased (P) individuals (TRN: DRKS00013119, Registration date: 22.10.2022). The samples were analysed using shotgun metagenomic sequencing combined, for the first time, with culture technique. A selection of 997 isolates was tested for resistance to relevant antibiotics. Results The shotgun metagenomics sequencing resulted in 2,069,295,923 reads classified into 4856 species-level OTUs. PERMANOVA analysis of beta-diversity revealed significant differences between the groups regarding their microbiota composition and their ARG profile. The samples were clustered into three ecotypes based on their microbial composition. The bacterial composition of H and C samples greatly overlapped and was based on ecotypes 1 and 2 whereas ecotype 3 was only detected in periodontitis. We found 64 ARGs conveying resistance to 36 antibiotics, particularly to tetracycline, macrolide-lincosamide-streptogramin, and beta-lactam antibiotics, and a correspondingly high prevalence of phenotypic resistance. Based on the microbiota composition, these ARGs cluster in different resistotypes, and a higher prevalence is found in healthy and caries active than in periodontally diseased individuals. There was a significant association between the resistotypes and the ecotypes. Although numerous associations were found between specific antibiotic resistance and bacterial taxa, only a few taxa showed matching associations with both genotypic and phenotypic analyses. Conclusions Our findings show the importance of the oral microbiota from different niches within the oral cavity as a reservoir for antibiotic resistance. Additionally, the present study showed the need for using more than one method to reveal antibiotic resistance within the total oral biofilm, as a clear mismatch between the shotgun metagenomics method and the phenotypic resistance characterization was shown