Bacterial urease activity in dental plaque and in saliva generates alkali, which can increase the plaque pH and can protect non-cariogenic oral bacteria against acidification6–11
; consequently it has been hypothesized that oral urease activity may be a caries-inhibiting factor, and that loss of this activity may lead to caries development. Recent pilot clinical studies have reported significantly higher plaque urease activity in caries-free vs. caries-active subjects, but not significant differences in saliva urease activity, suggesting a potentially significant caries protective role for plaque urease but not for urease in saliva. These were cross-sectional studies that compared adult subjects with extreme caries activity to caries-free subjects12, 13
. One recent cross-sectional study reported similar findings in a small group of children; however his study was not evaluating the relationship of urease with caries directly14
The present study is, to our knowledge, the first to examine the role of urease in caries development in children, prospectively. The design of the study involved repeated urease and caries measurements over a three-year period on a single panel of children with different caries levels at baseline, including caries-free children. Caries detection was performed using a sensitive, yet simple method, which facilitated the detection of non-cavitated lesions (“white spots”) even in small children, as well as the differentiation between “white spots” restricted to the enamel, versus those where dentin was already involved. For many investigators and clinicians, “white spot” lesions in children younger than six years of age are considered “caries”. However, in this study, these lesions were considered as “failures”, i.e., caries, only when there was a clear indication using FOTI that dentin was involved, for the reasons explained in the Methods. The study considered several known caries risk factors as potential confounders, such as age, gender, baseline caries status, mutans streptococci levels, sugar consumption and plaque amounts. We have recently reported some interesting findings regarding the complex interactions observed between urease activity and the aforementioned risk factors in this study15
. For all these reasons, it is not surprising that this study yielded some original and unexpected findings compared to the previous pilot clinical studies, which were cross-sectional, utilized adult populations with significant caries activity, and did not control for potential confounders.
The most unexpected and important finding of this study was that saliva urease activity had a significant effect on the risk of developing caries, and that this effect was not protective, but in fact caries promoting. As urease activity in saliva increased, caries incidence rates increased up to four times, and the risk of developing new caries increased progressively up to five times, even when the models were adjusted for baseline caries, age, and mutans streptococci levels. A possible explanation for this finding is that there may be competition between ureolytic bacteria in plaque and in saliva for limiting urea. Saliva is the principal source of urea in the oral cavity23
. As urease activity in saliva increases, salivary urea is more rapidly hydrolyzed and the amount of urea available to plaque bacteria may become limited, leading to a more acidic plaque. Another explanation for the association of saliva urease with increased caries risk can be the positive relationship between urease activity in saliva and salivary mutans streptococci levels in this study15
. This observation was attributed to the knowledge that subjects with high salivary mutans levels had more acidic salivary pH, which is known to de-repress urease expression in Streptococcus salivarius
. Interestingly, this relationship was observed in non-fasting subjects, but not in fasting subjects, and this may be the reason why it was not observed in the previous cross-sectional studies that used fasting subjects only.
The findings of the present study support the proposed protective effect of plaque urease on caries development, evidenced by progressively decreasing caries incidence rates and hazard ratios as plaque urease activity levels increased. However, the association of plaque urease activity with caries development in this study was not as significant as previously suggested in the cross-sectional studies, especially when only one measurement was used to define urease categories, or when urease was analyzed as a time-dependent variable in the Cox Proportional Hazards models. The magnitude and significance of this effect increased as multiple measurements were used to define baseline urease activity, such as the average of the first two, or first three visits. That was true for both plaque and saliva urease activity. The reason for this observation is most likely the poor reproducibility of urease activity at the six-month recall visits in this study, especially in plaque urease15
. A possible reason for the lower reproducibility of plaque urease was that plaque samples were pooled from different surfaces; therefore, their composition may have been different at each visit. Another source of variability in urease measurements in this study was whether the children had eaten prior to sample collection or not. Fasting prior to sample collection was difficult to control at the level of the study design due to the young age of the children, but it was measured during the study and controlled for in the analysis. Fasting had a strong impact on the reproducibility of plaque urease, compared with urease in saliva15
. Notably, subjects in the prior cross-sectional studies that found a significant difference in plaque urease activity between caries-free and caries-active had fasted prior to sampling. It appears, therefore, that it may be possible to observe a much stronger effect of plaque urease on caries development if a more standardized protocol for plaque and saliva collection is employed, such as collecting plaque from specific tooth sites, and under fasting conditions.
In conclusion, the findings of this prospective study suggest that increased urease activity in the saliva of small children may be associated with progressively increased caries risk. This finding may be attributed to a competition between ureolytic bacteria in the plaque and in saliva for salivary urea, or it may reflect the positive relationship between urease and mutans levels in saliva. Work in progress may clarify the mechanism of action. However, in either case, this observation could have significant clinical implications, as this study suggests that urease activity in saliva could be an equal or better predictor of caries risk in children than salivary mutans levels. Measuring urease activity in saliva is much faster and can be cheaper compared to the microbiological tests for mutans detection in saliva. Increased urease activity in dental plaque could be associated with decreased caries risk in children, but in order to demonstrate this association clearly and to apply it in clinical practice it will be necessary to improve the reproducibility of urease measurements in plaque. Using plaque urease as a risk indicator for caries would also require that the measurements be performed under fasting conditions, which may be a difficult requirement for children, but apparently the use of salivary urease does not have this requirement. An attractive and interesting approach that we are currently exploring is the possibility of combining low plaque urease activity and increased saliva urease activity into a single predictor for caries risk in children.