Organization of supragingival plaque at the micron scale

One of the most significant advances in oral microbiology in recent years is advanced imaging methodologies to reveal the geography of oral biofilms. Although the revolution in bacterial sequencing has left us with an unprecedented abundance of data regarding the diversity and abundance of bacteria in caries and periodontal diseases, detailed information on the organization of dental biofilms in man in micrometre scale is lacking. By using spectral imaging fluorescence in situ hybridization guided by metagenomic sequence analysis, Mark Welch et al. [1] described a multigenus, highly organized microbial consortium in human supragingival plaque. Their unique methodology (CLASI-FISH) (see reference [2] for additional details) was developed by Gary Borisy at the Forsyth Institute, Cambridge, MA, USA. It is superior to conventional FISH which has a very limited number of different target organisms that can be detected simultaneously with standard epifluorescence or confocal laser scanning microscopy [3]. CLASI-FISH allows simultaneous differentiation of up to 28 bacterial species within a biofilm using the oral cavity as a model. It allows us for the first time to visualize the geography of biofilms, i.e. to see where specific bacteria are located and with what species they most commonly exist. This can give us insight into how bacteria interact with each other and with their hosts in health and disease. The 10 bacterial probes applied in the current study targeted 96–98% of the cells in healthy supragingival plaque as suggested by rRNA tag sequencing data from the Human Microbiome Project (HMP) [4]. Each of the probes contained a specific oligo nucleotide conjugated to a unique fluorophore. This provided analysis of the organization of microbial communities through combinational labelling and spectral imaging. The oral, microbial biogeography (i.e. the micro-scale distribution of microorganisms that reside within the oral microbiome) thus established also allows us to understand more about the physiology and ecology of the biofilm community and its systems biology. Admittedly, previous literature has described bacterial coadhesion or coaggregation in great detail as well as ecological succession in dental biofilm; however, we have so far lacked the micrometre-scale resolution necessary to study the spatial organization of individual bacterial cell consortia [5]. CLASI-FISH is clearly different from nucleic acid methods where the structure of plaque is destroyed. Mark Welch et al. [1] applied spectral fluorescence imaging to examine the structures produced by nine key bacterial taxa of supragingival plaque. By using sequencing data from the HMP [4], major bacterial taxa thought to be prevalent and abundant in the overall structure and function of supragingival plaque were selected and by imaging the spatial organization of these taxa, a complex, spatially organized, multigenus consortium of bacteria was detected. Mark Welch et al. [1] reported that supragingival biofilm from 22 healthy volunteers consisted of a radially arranged, nine-taxon structure that was established around cells of filamentous corynebacteria. This bacterial consortium had a radius ranging from a few tens to a few hundreds of microns and was spatially differentiated. The authors described it as a hedgehog structure due to its filaments. The localization of the different taxa here suggested that they had specific functions in the consortium. As expected, anaerobic bacteria tended to localize in the interior where oxygen tension is low while facultative anaerobic and obligate aerobic bacteria tended to be at the periphery of the consortium (Figure 1). Consumers of sugars and oxygen such as streptococci produce metabolites like lactate, CO2, and H2O2 and tended to localize close to each other. Even if these were the most characteristic and reproducible features of the plaque structure, there was a spatial, temporal, and individual variation in the abundance of hedgehogs and other types of consortia in the supragingival plaque. The authors’ hypothesis suggested that Corynebacterium, which was abundant and the genus most characteristic of supragingival plaque, bound to the already existing dental biofilm substrate composed of Streptococcus and Actinomyces. Corynebacterium, projected in three planes, was found to structure the