Analysis of Urease Expression in Actinomyces naeslundii WVU45

Abstract

The hydrolysis of urea by ureases of oral bacteria in dental plaque can cause a considerable increase in plaque pH, which can inhibit the development of dental caries. There is also indirect evidence that urea metabolism may promote the formation of calculus and that ammonia release from urea could exacerbate periodontal diseases. Actinomyces naeslundii, an early colonizer of the oral cavity and a numerically significant plaque constituent, demonstrates comparatively low levels of urease activity on isolation, so this organism has not been considered a major contributor to total oral urease activity. In this study it was observed that urease activity and urease-specific mRNA levels in A. naeslundii WVU45 can increase up to 50-fold during growth under nitrogen-limiting conditions. Using primer extension analysis, a putative, proximal, nitrogen-regulated promoter of the A. naeslundii urease gene cluster was identified. The functionality and nitrogen responsiveness of this promoter were confirmed using reporter gene fusions and 5 deletion analysis. The data indicated that regulation of urease expression by nitrogen availability in A. naeslundii may require a positive transcriptional activator. Plaque bacteria may experience nitrogen limitation when carbohydrates are present in excess. Therefore, based on the results of this study and in contrast to previous beliefs, strains of A. naeslundii may have the potential to be significant contributors to total plaque ureolysis, particularly during periods when there is an increased risk for caries development.

The effect of sucrose on plaque and saliva urease levels in vivo

“…The best so far characterized urease enzymes in oral bacteria are the ones produced by S. salivarius and by Actinomyces naeslundii . The expression of ureases in these oral bacteria can be increased 50 to 300 –fold under increased carbohydrate concentrations or acidic pH, conditions that are clinically analogous to a cariogenic challenge (5, 6, 16, 21). To determine if urease activity in plaque and/or in saliva is induced during a cariogenic exposure in vivo , we measured urease activity and S. salivarius ureC mRNA during fasting conditions and after a short rinse with a sucrose solution, which is known to induce an immediate and reversible drop in plaque pH (24).…”

“…First, in vitro studies have been conducted on single species cultures of S. salivarius and A. naeslundii (5, 6, 16, 21). In contrast, the natural plaque and saliva samples used here represent multi-species bacterial populations, which may exhibit high variability with respect to the types and proportions of ureolytic species they contain.…”

“…For example, in Streptococcus salivarius urease expression can be increased up to 300-fold in acidic pH and during excess of carbohydrate (5, 6, 21). Urease expression in Actinomyces naeslundii can increase up to 50-fold during growth under nitrogen-limitation, a condition that oral bacteria may experience when carbohydrates are present in excess (16). Based on these in vitro observations, ongoing clinical studies at the University of Puerto Rico are examining the long-term effects of sugar consumption on urease activity levels in vivo , as it relates to caries development in children (3).…”

Alkali production associated with malolactic fermentation by oral streptococci and protection against acid, oxidative, or starvation damage

“…Another major source of microbially produced alkali in the mouth involves cytoplasmic ureases, such as those of Streptococcus salivarius (Chen and Burne 1996) or Actinomyces naeslundii (Morou-Bermudez and Burne 2000), which can catalyze hydrolysis of urea normally present in saliva to CO 2 and NH 3 , but without concomitant ATP production. Urease production appears to be regulated mainly in response to nitrogen starvation rather than acidification, so protection against acidification can be a secondary function for the enzyme.…”

Oral Bacterial Acid–Base Metabolism in Caries Screening: A Proof-Of-Concept Study

“…Using a more streamlined technology could also improve the low repeatability observed between the two visits. The variability in the results between the two visits could also be explained by the fact that expression of the genes involved in sugar and urea metabolism in oral bacteria is highly regulated in response to carbohydrate availability, nitrogen availability, and pH (Morou-Bermudez and Burne, 2000; Burne and Marquis 2000; Lemos et al, 2005; Li et al, 2008). To address this issue we utilized a standardized sample collection protocol that was performed under fasting conditions; however, it is possible that the participants followed the fasting requirement in the first visit only, but not the second one.…”