Laboratory microbial culture models simulate the oral environment for cariology studies. Unlike in vivo studies, laboratory simulations do not face problems related to the uncontrollable fluctuations of the oral environment.[5,6] Two complementary microbiological approaches can be taken to generate biofilm in microbial culture models. One of these approaches investigates the evolution of a plaque microcosm from natural oral microflora.[7] The other approach is the construction of defined-species biofilm consortia with major plaque species, or a mixture of different species of the acquired oral bacteria, such as the American Type Culture Collection (ACTT) bacteria. The designs of laboratory microbial culture models vary according to the purpose of the laboratory studies – these models are classified as closed and open systems. Each system is a compromise between the reality of the in vivo ecosystem and the simplification of the system. However, a well-designed model and a study allow researchers to obtain precise and useful results.[7]
The closed system
Microbial culture models in the closed system have a finite supply of nutrients. The growth rates of the biofilm are rapid at the beginning of the cultivation cycle when there is plethora of nutrients. However, this is rarely observed in the in vivo growth of biofilm.[8,9] The growth conditions will change significantly following the consumption of nutrients and the accumulation of metabolic products. Hence, the physiological and biological properties of the biofilm are not comparable with those observed in vivo. Researchers use closed system models for their simplicity, high productivity, repeatability, controllability of the experimental conditions, less contamination, and cost-effective properties. The agar plate and microtiter biofilm models are two examples of a simple microbial culture model in closed system.
The open system
The open system can be described as a continuous culture cycle system. It enables the supply of a fresh culture medium and the removal of metabolites and spent culture liquid simultaneously. Hence, the concentration of bacteria and metabolic products remains constant.[10] Moreover, the biofilm is able to stay in a stable state or in a dynamic balance.[11] Nevertheless, the repeatability of experimental result is low because of the heterogeneity of the biofilm in the open system.
The open system simulates the in vivo environment better than the closed system, allowing better regulation of the biofilm growth rate and other variables. Common microbial culture models in the open system include the chemostat model, the flow cell biofilm model, the constant depth film fermenter model, the drip flow biofilm reactor, the multiple Sorbarod model, and the multiple artificial mouth model.
The multiple artificial mouth (MAM) is a computer-controlled, multiple station model. A MAM can accurately simulate an in vivo environment using computer-controlled facilities.[12]
It has several microstations which are relatively independent to one another. Different experimental conditions can be applied simultaneously in different microstations.
Environmental variables can be easily controlled in the MAM, thus allowing analysis of the biofilm during its development without contaminating other samples. Acidity can be monitored using a pH electrode and a micro-reference electrode.[6] These well-controlled conditions improve the standardization and flexibility of the MAM, therefore enhancing its ability to culture biofilms close to natural oral flora. Sissons et al. found that biofilms developed in this system exhibited metabolic and pH behaviour that resembled typical natural plaques.[12] The MAM has been adopted in different studies, such as biodiversity of plaque, fluoride and phosphate assay[13,14], plaque calcium level measurement[14], and the generation of consortia using major plaque species15. The biofilm samples in this model were exposed to the same temperature and gas-phase fluctuation. The MAM aims to mimic the oral environment therefore saliva substitutes play an important role in the model. Approximate laminar flows are applied to simulate the situations in the oral cavity, instead of turbulent flow in chemostat.