Antimicrobial effect of SDF has been proven by various previous studies [15–20, 25], but the effect of SDF on micro-ecology of dental plaque has seldom been explored. Based on the already known antimicrobial effect of SDF, this pioneer preliminary study aimed to further our knowledge of SDF on dental plaque from the micro-ecosystem aspect.
In this study, extracted deciduous teeth with large caries from school-age children were collected and topically treated in vitro with a 38% SDF solution. Dental plaque from the caries pre- and post-intervention were collected for microbial sequencing and the results were compared. The amount of 9 out of 15 richest bacteria genera, the microbial diversity and the microbial community composition in the dental plaque after SDF intervention were noticeably changed. The relative proportions of Pseudomonas, Fusobacterium and Pseudoramibacter were higher than before the intervention, although no significant different was found (P > 0.05). After SDF intervention, microbial association in the dental plaque became more complex with positive connections overwhelming the negative ones. Carbohydrate transportation and metabolic functions in the dental plaque were significantly reduced (P < 0.05).
The microbial diversity was reduced, the microbial richness as well as microbial community composition was shifted post 38% SDF intervention, which supported the broad spectrum bactericidal effect as shown in previous studies [25], although no statistically significant difference was found in this study. A recent study on 38% SDF, which compared pre- and one-month-post-intervention dental plaque sample from the mouth of adult patients [29], found no significant difference in the plaque microorganisms composition either. However, difference in dentition, sampling conditions and the follow-up interval should also be noted. The statistical indifference in this study may also be due to the relatively small sample size adopted in this study. Based on experiences of previous studies, a large difference in the microbial diversity before and after SDF intervention was estimated, yielding a number of five children needed in this study. And this relatively small sample size was also practical if balancing the research expenses. Clearly, a larger sample with more funding should be adopted in future studies.
Most of the genera present before the intervention (Olsenella, Streptococcus, Bifidobacterium, Prevotella_7, Lactobacillus, Actinomyces, Leptotrichia, Selenomonas_3, Veillonella) were sensitive to SDF, and the relative amounts decreased substantially. This was consistent with previous studies [15, 16, 18, 20] and supported the broad spectrum bactericidal effect of SDF [25]. Streptococcus, Lactobacillus and Actinomyces have long been recognized as classical cariogenic bacteria. Prevotella_7, Selenomonas_3, and Bifidobacterium are often found in deep dentin caries [30]. Olsenella, which could produce lactic acid, seldom mentioned in the past, have recently been found associated with dentin and root caries [31, 32]. Leptotrichia has a strong glycolysis effect [33], and its detection rate is increased in the process of caries [34, 35]. Veillonella has a unique intracellular pH control mechanism [36], which might promote the cariogenicity of Streptococcus mutans [35, 37]. By inhibiting these bacteria, cariogenicity of the dental plaque may be suppressed, creating a new microbial balance.
It was also found that Pseudomonas, Fusobacterium and Pseudoramibacter could withstand the effect of topically applied SDF. The possible role of Pseudomonas in dental caries was indicated recently. NavNeet Kaur et al. [38] detected a high level of Pseudomonas in the dentin caries. In Liang's study [39], Pseudomonas was mentioned as a potential pathogen of caries, but suggested the relationship be further studied. Pseudomonas is a non-fermenting, obligate aerobic Gram-negative bacillus. It has no special nutritional requirements and is common in soil, fresh water, seawater. A total of twenty-nine species have been identified, of which at least three are pathogenic to animals or humans, causing infections. Pseudomonas aeruginosa is part of the commensal oral flora, but can become a pathogen and cause nosocomial infection [40]. Small infective doses of the bacteria can produce local abscesses, large doses can lead to death from systemic infection. Fluorescent pseudomonas ssp. can cause spoilage of frozen meat, eggs, milk and dairy products. Pseudomonas birensis can be transmitted through the mouth, the respiratory tract or in wounds. Pseudoramibacter is a nonmotile, nonsporeforming, strictly anaerobic, Gram-positive bacillus, which was saccharolytic. The end products of its fermentation are formate, acetate, butyrate, caproate, and hydrogen [41]. It can cause periapical infection [42–44], it often found in deep dentin caries [30, 45] and infected root canals [41]. Similarly, Fusobacterium is often detected in deep dentin caries [30, 45] and infected root canals [43, 44, 46]. It is a strictly anaerobic Gram-negative bacillus living in the oral cavity and the upper digestive, intestine and urogenital tracts of humans or animals, as well as in the soil. It is most commonly seen in oral dental plaque. Most of the strains do not ferment any sugars, only a few strains are known to weakly ferment glucose and fructose. Briefly, Pseudomonas, Fusobacterium and Pseudoramibacter may not play important role in the development of dental caries, but they are more closely related to root canal or periapical infections. In this study, deciduous teeth with extensive caries or obvious dental pulp symptoms from school-age children was studied, which could explain the richness of these bacteria in the sample. The results would also provide guiding information in the prevention and control of extensive deciduous caries associated with pulp infection or periapical disease. The tolerance of these three kind of bacteria to single time SDF application in this in vitro study should be pay cautious attention to when use SDF in the clinical situation in child population. Further studies are worth conducting and should be designed according to this special concern in order to form a clear guide.
Restricted by ethic issues, extracted deciduous teeth were used in this study. As a result, intervention and sampling were all carried out in vitro. An in vitro study differs from the real clinical situation. The effect of common oral flora, saliva, food and environmental microorganisms on the formation of new microbial balance could not be reproduced or evaluated. New micro-ecological balance formation after SDF intervention in this study could only be based on the before-intervention status as well as the effect of SDF. Bacteria which were sensitive to SDF were controlled, while those with relatively high tolerance to SDF grew relatively better after the intervention. After SDF intervention, the microbial association in the dental plaque became more complex with positive connections overwhelming the negative ones, indicating that the connections and collaborations among the remaining bacterial communities were enhanced.
The functional characteristics of plaque in caries were also compared and analyzed based on the COG database for homologous classification of gene products. Carbohydrate transportation and metabolism function in plaque were significantly reduced 24 hours and 1 week after intervention (shown in yellow in Fig. 6, G), and the replication, recombination and repair function in plaque (shown in pink in Fig. 6, L) were also significantly reduced. These changes revealed from another aspect the overall inhibitory effect of SDF on the bacterial community in dental plaque, which affected the properties of the whole plaque, especially the cariogenicity within the plaque micro-ecology. The function of signal transduction mechanism was significantly increased, which could be correlated with the enhancement of synergistic effect between residual bacteria in plaque after intervention.
Most parameters in this study showed that the greatest microbial difference was found at one week after SDF intervention, indicating a possible time effect of SDF. However, the observation period in this study was relatively short which may have its limitations, and there were differences between this in vitro study and the real mouth situation, time effect of SDF application on dental plaque may not be well explained, and should be studied further in good experiment situation which can mimic the real mouth environment well or in real mouth situation if possible.