Abstract
ABSTRACTOral biofilms have not been studied using both metabolome and protein profiling concurrently. Bacteria produce proteases that lead to degradation of functional salivary proteins. The novel protocol described here allows for complete characterisation of in vitro oral biofilms, including proteolytic, metabolic, and microbiome analysis.Biofilms were grown on hydroxyapatite discs from whole mouth saliva, using sterilised saliva as a growth-medium, in different growth environments. Salivary protein degradation was assessed from spent saliva growth-medium using SDS-polyacrylamide gel electrophoresis (SDS-PAGE), and metabolic activity by nuclear magnetic resonance (NMR). Discs were assessed for depth and coverage of biofilms by confocal laser scanning microscopy (CLSM), and biofilms were collected at the end of the experiment for 16S rRNA gene sequence analysis.There was a significant difference in biofilm viability, salivary protein degradation, and metabolites identified between biofilms grown aerobically and biofilms exposed to an anaerobic environment. Bacterial 16S rRNA gene sequencing showed the predominant genus in the 7-day aerobic biofilms was Streptococcus, in aerobic-anaerobic and anaerobic 7-day biofilms Porphyromonas, and in aerobic-anaerobic and anaerobic 13-day biofilms Fusobacterium.This data suggests new growth requirements and capabilities for analysing salivary biofilms in vitro, which can be used to benefit future research into oral bacterial biofilms.
Highlights
There was a significant difference in biofilm viability, salivary protein degradation, and metabolites identified between biofilms grown aerobically and biofilms exposed to an anaerobic environment
This study has successfully developed a method for assessing the proteolytic and metabolic activity of in vitro salivary biofilms, using saliva as an inoculum and growth medium
This is the first known study to report the use of nuclear magnetic resonance (NMR) to analyse metabolites produced by oral biofilm bacteria grown in sterile whole mouth saliva
Summary
There was a significant difference in biofilm viability, salivary protein degradation, and metabolites identified between biofilms grown aerobically and biofilms exposed to an anaerobic environment. Bacterial 16S rRNA gene sequencing showed the predominant genus in the 7-day aerobic biofilms was Streptococcus, in aerobic-anaerobic and anaerobic 7-day biofilms Porphyromonas, and in aerobic-anaerobic and anaerobic 13day biofilms Fusobacterium This data suggests new growth requirements and capabilities for analysing salivary biofilms in vitro, which can be used to benefit future research into oral bacterial biofilms. This, is not an exhaustive list, and is obviously limited heavily by the methods used in coaggregation studies, as it is known that there are species of oral bacteria that cannot be cultured In contrast to these studies, there is new and conflicting evidence that suggests Corynebacterium is the most important bridging genus, and that plaque biofilms become more aerobic at the periphery as opposed to anaerobic [11]. Based on these differing coaggregation models, three different incubation environments were assessed in this study; purely aerobic, purely anaerobic and a mixed aerobicanaerobic incubation
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
