To exert anticaries effects, probiotics are described to inhibit growth and biofilm formation of cariogenic bacteria such as Streptococcus mutans (SM). We screened 8 probiotics and assessed how SM growth or biofilm formation inhibition affects cariogenicity of probiotic-SM mixed-species biofilms in vitro. Growth inhibition was assessed by cocultivating probiotics and 2 SM strains (ATCC 20532/25175) on agar. Probiotics were either precultured before SM cultivation (exclusion), or SM precultured prior to probiotic cultivation (displacement). Inhibition of SM culture growth was assessed visually. Inhibition of SM biofilm formation on bovine enamel was assessed using a continuous-flow short-term biofilm model, again in exclusion or displacement mode. The cariogenicity of mixed-species biofilms of SM with the most promising growth and biofilm formation inhibiting probiotic strains was assessed using an artificial mouth model, and enamel mineral loss (ΔZ) was measured microradiographically. We found limited differences in SM growth inhibition in exclusion versus displacement mode, and in inhibition of SM 20532 versus 25175. Results were therefore pooled. Lactobacillus acidophilus LA-5 inhibited significantly more SM culture growth than most other probiotics. L. casei LC-11 inhibited SM biofilm formation similarly to other alternatives but showed the highest retention of probiotics in the biofilms (p < 0.05). Mineral loss from SM monospecies biofilms (ΔZ = 9,772, 25th/75th percentiles: 6,277/13,558 vol% × µm) was significantly lower than from mixed-species SM × LA-5 biofilms (ΔZ = 24,578, 25th/75th percentiles: 19,081/28,768 vol% × µm; p < 0.01) but significantly higher than from SM × LC-11 biofilms (ΔZ = 4,835, 25th/75th percentiles: 263/7,865 vol% × µm; p < 0.05). Probiotics inhibiting SM culture growth do not necessarily reduce the cariogenicity of SM-probiotic biofilms. Nevertheless, SM biofilm formation inhibition may be relevant in the reduction of cariogenicity.

Keywords:
Bacteria, Biofilm(s), Caries, Microbial ecology, Probiotics
1.
Aas JA, Griffen AL, Dardis SR, Lee AM, Olsen I, Dewhirst FE, Leys EJ, Paster BJ: Bacteria of dental caries in primary and permanent teeth in children and young adults. J Clin Microbiol 2008;46:1407-1417.
2.
Adami A, Cavazzoni V: Occurrence of selected bacterial groups in the faeces of piglets fed with Bacillus coagulans as probiotic. J Basic Microbiol 1999;39:3-9.
3.
Ahola AJ, Yli-Knuuttila H, Suomalainen T, Poussa T, Ahlstrom A, Meurman JH, Korpela R: Short-term consumption of probiotic-containing cheese and its effect on dental caries risk factors. Arch Oral Biol 2002;47:799-804.
4.
Anderson P, Levinkind M, Elliot JC: Scanning microradiographic studies of rates of in vitro demineralization in human and bovine dental enamel. Arch Oral Biol 1998;43:649-656.
5.
Arioli S, Ragg E, Scaglioni L, et al: Alkalizing reactions streamline cellular metabolism in acidogenic microorganisms. PLoS One 2010;5:e15520.
6.
Badet C, Thebaud NB: Ecology of lactobacilli in the oral cavity: a review of literature. Open Microbiol J 2008;2:38-48.
7.
Beighton D, Al-Haboubi M, Mantzourani M, Gilbert SC, Clark D, Zoitopoulos L, et al: Oral bifidobacteria: Caries-associated bacteria in older adults. J Dent Res 2010;89:970-974.
8.
Caglar E, Kuscu OO, Selvi Kuvvetli S, Kavaloglu Cildir S, Sandalli N, Twetman S: Short-term effect of ice-cream containing Bifidobacterium lactis BB-12 on the number of salivary mutans streptococci and lactobacilli. Acta Odont Scand 2008;66:154-158.
9.
Caglar E, Sandalli N, Twetman S, Kavaloglu S, Ergeneli S, Selvi S: Effect of yogurt with bifidobacterium dn-173 010 on salivary mutans streptococci and lactobacilli in young adults. Acta Odont Scand 2005;63:317-320.
10.
Cheikhyoussef A, Pogori N, Chen W, Zhang H: Antimicrobial proteinaceous compounds obtained from bifidobacteria: from production to their application. Int J Food Microbiol 2008;125:215-222.
11.
Cotter PD, Hill C: Surviving the acid test: responses of gram-positive bacteria to low pH. Microbiol Mol Biol Rev 2003;67:429-453.
12.
Dibdin GH, Shellis RP, Wilson CM: An apparatus for the continuous culture of micro-organisms on solid surfaces with special reference to dental plaque. J Appl Bacteriol 1976;40:261-268.
13.
Endres JR, Clewell A, Jade KA, Farber T, Hauswirth J, Schauss AG: Safety assessment of a proprietary preparation of a novel probiotic, Bacillus coagulans, as a food ingredient. Food Chem Toxicol 2009;47:1231-1238.
14.
Glavina D, Gorseta K, Skrinjaric I, Vranic DN, Mehulic K, Kozul K: Effect of lgg yoghurt on Streptococcus mutans and Lactobacillus spp. salivary counts in children. Coll Antropol 2012;36:129-132.
15.
Gruner D, Paris S, Schwendicke F: Probiotics for managing caries and periodontitis: Systematic review and meta-analysis. J Dent 2016
https://doi.org/10.1016/j.jdent.2016.03.002
.
16.
Hamon Y, Peron Y: La sérologie des bactériocines. Importance de cette méthode. CR Acad Sci 1965;260:6730.
17.
Hasslöf P, West CE, Karlsson Videhult F, Brandelius C, Stecksén-Blicks C: Early intervention with probiotic Lactobacillus paracasei F19 has no long-term effect on caries experience. Caries Res 2013;47:559-565.
18.
Haukioja A, Soderling E, Tenovuo J: Acid production from sugars and sugar alcohols by probiotic lactobacilli and bifidobacteria in vitro. Caries Res 2008;42:449-453.
19.
Hedberg M, Hasslöf P, Sjöström I, Twetman S, Stecksén-Blicks C: Sugar fermentation in probiotic bacteria - an in vitro study. Oral Microbiol Immunol 2008;23:482-485.
20.
Kalman DS, Schwartz HI, Alvarez P, Feldman S, Pezzullo JC, Krieger DR: A prospective, randomized, double-blind, placebo-controlled parallel-group dual site trial to evaluate the effects of a Bacillus coagulans-based product on functional intestinal gas symptoms. BMC Gastroenterol 2009;9:85.
21.
Keller MK, Hasslöf P, Dahlén G, Stecksén-Blicks C, Twetman S: Probiotic supplements (Lactobacillus reuteri DSM 17938 and ATCC PTA 5289) do not affect regrowth of mutans streptococci after full-mouth disinfection with chlorhexidine: a randomized controlled multicenter trial. Caries Res 2012;46:140-146.
22.
Keller MK, Twetman S: Acid production in dental plaque after exposure to probiotic bacteria. BMC Oral Health 2012;12:44.
23.
Kneist S, Schmidt F, Callaway A, Willershausen B, Rupf S, Wicht M, Thiede B: Diversity of lactobacillus species in deep carious lesions of primary molars. Eur Arch Paediatr Dent 2010;11:181-186.
24.
Laleman I, Detailleur V, Slot DE, Slomka V, Quirynen M, Teughels W: Probiotics reduce mutans streptococci counts in humans: a systematic review and meta-analysis. Clin Oral Invest 2014;18:1539-1552.
25.
Lang C, Böttner M, Holz C, Veen M, Ryser M, Reindl A, Pompejus M, Tanzer JM: Specific lactobacillus/mutans streptococcus co-aggregation. J Dent Res 2010;89:175-179.
26.
Mandel DR, Eichas K, Holmes J: Bacillus coagulans: a viable adjunct therapy for relieving symptoms of rheumatoid arthritis according to a randomized, controlled trial. BMC Compl Altern Med 2010;10:1.
27.
Marsh PD: Dental plaque as a biofilm and a microbial community - implications for health and disease. BMC Oral Health 2006;6:S14.
28.
Martinez FAC, Balciunas EM, Converti A, Cotter PD, de Souza Oliveira RP: Bacteriocin production by Bifidobacterium spp. A review. Biotech Adv 2013;31:482-488.
29.
Marttinen A, Haukioja A, Karjalainen S, Nylund L, Satokari R, Öhman C, Holgerson P, Twetman S, Söderling E: Short-term consumption of probiotic lactobacilli has no effect on acid production of supragingival plaque. Clin Oral Invest 2012;16:797-803.
30.
Meurman JH: Probiotics: do they have a role in oral medicine and dentistry? Eur J Oral Sci 2005;113:188-196.
31.
Meurman JH, Antila H, Korhonen A, Salminen S: Effect of Lactobacillus rhamnosus strain GG (ATCC 53103) on the growth of Streptococcus sobrinus in vitro. Eur J Oral Sci 1995;103:253-258.
32.
Näse L, Hatakka K, Savilahti E, Saxelin M, Pönkä A, Poussa T, Korpela R, Meurman JH: Effect of long-term consumption of a probiotic bacterium, Lactobacillus rhamnosus GG, in milk on dental caries and caries risk in children. Caries Res 2001;35:412-420.
33.
Russell DA, Ross RP, Fitzgerald GF, Stanton C: Metabolic activities and probiotic potential of bifidobacteria. Int J Food Microbiol 2011;149:88-105.
34.
Schlundt J: Health and nutritional properties of probiotics in food including powder milk with live lactic acid bacteria. Report of a Joint FAO/WHO expert consultation on evaluation of health and nutritional properties of probiotics in food including powder milk with live lactic acid bacteria. 2001. http://www.who.int/foodsafety/publications/fs_management/en/probiotics.pdf.
35.
Schwendicke F, Dorfer C, Kneist S, Meyer-Lueckel H, Paris S: Cariogenic effects of probiotic Lactobacillus rhamnosus GG in a dental biofilm model. Caries Res 2014a;48:186-192.
36.
Schwendicke F, Horb K, Kneist S, Dörfer C, Paris S: Effects of heat-inactivated bifidobacterium BB12 on cariogenicity of Streptococcus mutans in vitro. Arch Oral Biol 2014b;59:1384-1390.
37.
Singh R, Damle SG, Chawla A: Salivary mutans streptococci and lactobacilli modulations in young children on consumption of probiotic ice-cream containing Bifidobacterium lactis BB12 and Lactobacillus acidophilus LA5. Acta Odont Scand 2011;69:389-394.
38.
Sissons CH, Cutress TW, Hoffman MP, Wakefield JSJ: A multi-station dental plaque microcosm (artificial mouth) for the study of plaque growth, metabolism, pH, and mineralization. J Dent Res 1991;70:1409-1416.
39.
Stecksén-Blicks C, Sjöström I, Twetman S: Effect of long-term consumption of milk supplemented with probiotic lactobacilli and fluoride on dental caries and general health in preschool children: a cluster-randomized study. Caries Res 2009;43:374-381.
40.
Tabasco R, Garcia-Cayuela T, Pelaez C, Requena T: Lactobacillus acidophilus LA-5 increases lactacin B production when it senses live target bacteria. Int J Food Microbiol 2009;132:109-116.
41.
Taipale T, Pienihäkkinen K, Alanen P, Jokela J, Söderling E: Administration of Bifidobacterium animalis subsp. lactis BB-12 in early childhood: a post-trial effect on caries occurrence at four years of age. Caries Res 2013;47:364-372.
42.
Taipale T, Pienihäkkinen K, Salminen S, Jokela J, Söderling E: Bifidobacterium animalis subsp. lactis BB-12 administration in early childhood: a randomized clinical trial of effects on oral colonization by mutans streptococci and the probiotic. Caries Res 2012;46:69-77.
43.
Terai T, Okumura T, Imai S, Nakao M, Yamaji K, Ito M, Nagata T, Kaneko K, Miyazaki K, Okada A, Nomura Y, Hanada N: Screening of probiotic candidates in human oral bacteria for the prevention of dental disease. PLoS One 2015;0:e0128657.
44.
Twetman L, Larsen U, Fiehn N-E, Stecksén-Blicks C, Twetman S: Coaggregation between probiotic bacteria and caries-associated strains: an in vitro study. Acta Odont Scand 2009;67:284-288.
45.
Twetman S, Keller MK: Probiotics for caries prevention and control. Adv Dent Res 2012;24:98-102.
46.
Wong L, Sissons CH: A comparison of human dental plaque microcosm biofilms grown in an undefined medium and a chemically defined artificial saliva. Arch Oral Biol 2001;46:477-486.
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