Biosensor
In process of tooth decay, two kinds of bacteria dominate in different time. In our daily life, after we wash tooth, some kind of primary bacteria whose don’t harm our tooth, will start to adhesion on our tooth. In the early stage of tooth decay, adhesion of mutans streptococci will form plaque and start to destroy enamel. One of them , Streptococcus mutans is the main bacteria.[1] In the later period, Lactobacilli[8] will play a leading role at destroy the dentin.[3]
CSP (Competence-Stimulating Peptide)
- Streptococcus mutans
Streptococcus .mutans is kind of gram negative bacteria that the main reason of cavity. After we brush teeth, some bacteria will again attach on tooth and become primary colonizers, S.mutans adhere to the primary colonizers by cell-cell interaction.[5] To survive and growth the colony, S.mutans will secret some short-length peptide, CSP. (Competence-Stimulating Peptide)[1] This kind of peptide will direct S.mutans to secret biotoxin, driving other bacteria out, and form biofilm. This situation we called it quorum sensing. When biofilm become more thick but soft, we call it plaque. Therefore, we detected the amount of CSP to presumably form biofilm.[4,7]
- How did we detect CSP?
We used CSP-comDE two-component system from S.mutans and transformed into our host bacteria B.subtilis (Bacillus subtilis strain MW12). It is coming to the concrete measures.
Using two different plasmids for transformation. One of them, we called it plasmid comDE, carry genes encoding comD and comE. It will product membrane-bound histidine kinase receptor, comD, and cytoplasmic response regulator comE. When CSP from the other S.mutans triggered comD on our host B.subtilis, comD will autophosphorylate the receptor comE.[9] Activated comE directly activates the other plasmid which carry gene encoding GFP(green fluorescent protein)to produce our protein. Using the detector to measure the strength of green light, we will know the amount of CSP.
Lactate
- Lactobacilli
Lactobacilli will become dominated bacteria[3] when tooth decay got serious and damaged dentin. The main reason is that Lactobacilli secret lactic acid by doing fermentation process.[9] Lactic acid make an environment of tooth more acid. More acid surroundings let calcium which is the main component of tooth, more soluble in saliva. This called decalcification. In the early stage of tooth cavity, S.mutans is also using lactic acid to damage tooth. In comparison with detecting S.mutans, detecting Lactobacilli is more difficult because Lactobacilli is not a specific bacteria but group of bacteria.[9] Therefore, we changed our mine. We decided to detect lactic acid rather than detect Lactobacilli. Another reason is that lactic acid is more directly reason for tooth decay.
- How do we detect lactic acid?
We used LldPRD operon from Escherichia coli and did some modification. Transformed into our host E.coli. (Escherichia coli strain DH5α)
We used only one plasmid for transformation. This plasmid carry genes encoding Lldr and GFP(green fluorescent protein),which is having LldPRD operon. The Lldr gene will continue to produce Lldr until it reaches equilibrium. Lldr bound on the O1 binding site and O2 binding site, the LldPRD operon will be closed and cause GFP can’t be produced. When L-lactate come into our host E.coli and trigger on LldPRD operon, Lldr will be removed and cause LldPRD operon can work temporarily. GFP will be produced until the operon be closed again.[1]
References
Picture resources
(1) http://www.dimensionsofdentalhygiene.com/ddhright.aspx?id=434
(2) https://www.flickr.com/photos/ajc1/8344600413
References
(Aguilera et al., 2008; Ambatipudi et al., 2010; Badet & Thebaud, 2008; Cvitkovitch, Li, & Ellen, 2003; Forssten, Björklund, & Ouwehand, 2010; Karpiński & Szkaradkiewicz, 2013; Lemme, Gröbe, Reck, Tomasch, & Wagner-Döbler, 2011; Leung, Dufour, & Lévesque, 2015; Liu, Xue, & Wang, 2015; Senadheera & Cvitkovitch, 2008)
[1] Aguilera, L., Campos, E., Giménez, R., Badía, J., Aguilar, J., & Baldoma, L. (2008). Dual role of LldR in regulation of the lldPRD operon, involved in L-lactate metabolism in Escherichia coli. Journal of bacteriology, 190(8), 2997-3005.
[2] Ambatipudi, K. S., Hagen, F. K., Delahunty, C. M., Han, X., Shafi, R., Hryhorenko, J., . . . Koo, H. (2010). Human common salivary protein 1 (CSP-1) promotes binding of Streptococcus mutans to experimental salivary pellicle and glucans formed on hydroxyapatite surface. Journal of proteome research, 9(12), 6605.
[3] Badet, C., & Thebaud, N. (2008). Ecology of lactobacilli in the oral cavity: a review of literature. The open microbiology journal, 2, 38.
[4] Cvitkovitch, D. G., Li, Y.-H., & Ellen, R. P. (2003). Quorum sensing and biofilm formation in Streptococcal infections. Journal of Clinical Investigation, 112(11), 1626.
[5] Forssten, S. D., Björklund, M., & Ouwehand, A. C. (2010). Streptococcus mutans, caries and simulation models. Nutrients, 2(3), 290-298.
[6] Karpiński, T. M., & Szkaradkiewicz, A. K. (2013). Microbiology of dental caries. Journal of Biology, 3(1), M21-M24.
[7] Lemme, A., Gröbe, L., Reck, M., Tomasch, J., & Wagner-Döbler, I. (2011). Subpopulation-specific transcriptome analysis of competence-stimulating-peptide-induced Streptococcus mutans. Journal of bacteriology, 193(8), 1863-1877.
[8] Leung, V., Dufour, D., & Lévesque, C. M. (2015). Death and survival in Streptococcus mutans: differing outcomes of a quorum-sensing signaling peptide. Frontiers in microbiology, 6.
[9] Liu, T., Xue, S., & Wang, L. (2015). ABC Transporter CslAB, a Stabilizer of ComCDE Signal in Streptococcus mutans. Jundishapur journal of microbiology, 8(8).
[10] Senadheera, D., & Cvitkovitch, D. G. (2008). Quorum sensing and biofilm formation by Streptococcus mutans Bacterial Signal Transduction: Networks and Drug Targets (pp. 178-188): Springer.
