In vitro evaluation of the bactericidal effect of human breast milk against Streptococcus mutans at three time periods

Background: Human breast milk is a dynamic and multi-faceted fluid that contains essential nutrients for infant health and development. Its composition changes throughout the stages of lactation, and besides providing the newborns with plenty of nutrients, it also protects them against infections such as sepsis, pneumonia, and enteritis, especially in premature infants. The question of whether breastfeeding is cariogenic or not remains unresolved when comparing the results of different researchers. The objective of this study was to measure the bactericidal effect of breast milk against Streptococcus mutans (S. mutans), which has a high cariogenic potential. Methods: Breast milk samples were collected from 9 donors and divided into 3 groups: Group 1 (fresh milk), Group 2 (milk stored at −18 ◦C for 24 hours) and Group 3 (milk stored at −18 ◦C for 72 hours). From each group, 3 disks soaked in milk were placed on an agar culture of S. mutans. In the same culture a disk soaked with chlorhexidine was placed as a control. Inhibition halos were measured after 24, 48 and 72 hours. Results: Only Group 1 (fresh milk) incubated for 24 hours showed a statistically significant difference, presenting the lowest inhibition (p < 0.05). In the other eight data sets, there was no significant difference showing higher or lower inhibition (p > 0.05). However, there was a trend toward reduced inhibitory capacity when the milk was frozen for 72 hours. Conclusions: Human breast milk showed a bactericidal effect against S. mutans, which has a high cariogenic potential. Given this, we recommend exclusive breastfeeding during the first 6 months of life and thereafter combining breastfeeding and complementary feeding for as long as desired by the child and mother.

Human breast milk; Streptococcus mutans; Bactericidal effect; Cariogenic

[1] Yi DY, Kim SY. Human breast milk composition and function in human health: from nutritional components to Microbiome and MicroRNAs. Nutrients. 2021; 13: 3094.

[2] Saeed A, Ali H, Yasmin A, Baig M, Ullah A, Kazmi A, et al. Unveiling the antibiotic susceptibility and antimicrobial potential of bacteria from human breast milk of Pakistani women: an exploratory study. BioMed Research International. 2023; 2023: 6399699.

[3] Dallas DC, Guerrero A, Khaldi N, Castillo PA, Martin WF, Smilowitz JT, et al. Extensive in vivo human milk peptidomics reveals specific proteolysis yielding protective antimicrobial peptides. Journal Proteome Research. 2013; 12: 2295–2304.

[4] Fu Y, Ji C, Chen X, Cui X, Wang X, Feng J, et al. Investigation into the antimicrobial action and mechanism of a novel endogenous peptide β-casein 197 from human milk. AMB Express. 2017; 7: 119.

[5] Sharma C, Singh BP, Thakur N, Gulati S, Gupta S, Mishra SK, et al. Antibacterial effects of Lactobacillus isolates of curd and human milk origin against food-borne and human pathogens. 3 Biotech. 2017; 7: 31.

[6] Tesfay MY, Delbiso TD. Prevalence of dental carries and its association with breastfeeding duration among young children in Addis Ababa, Ethiopia. BMC Public Health. 2024; 24: 1525.

[7] Sritangsirikul S, Kitsahawong K, Matangkasombut O, Seminario AL, Pitiphat W. A longitudinal study on the impact of breastfeeding with or without formula milk on dental caries. Scientific Reports. 2024; 14: 10384.

[8] Momeni SS, Beno SM, Baker JL, Edlund A, Ghazal T, Childers NK, et al. Caries-associated biosynthetic gene clusters in Streptococcus mutans. Journal of Dental Research. 2020; 99: 969–976.

[9] Spatafora G, Li Y, He X, Cowan A, Tanner ACR. The evolving microbiome of dental caries. Microorganisms. 2024; 12: 121.

[10] Chen DR, Lin HC. Research updates: cariogenic mechanism of Streptococcus mutans. Journal of Sichuan University. Medical Science Edition. 2022; 53: 208–213. (In Chinese)

[11] Agarwal N, Khodke S, Naik S. Infant dietary pattern and its association with early childhood caries in preschool children: a cross-sectional study. International Journal of Clinical Pediatric Dentistry. 2023; 16: 421–425.

[12] Łubiech K, Twarużek M. Lactobacillus bacteria in breast milk. Nutrients. 2020; 12: 3783.

[13] Duangthip D, Chen KJ, Gao SS, Lo ECM, Chu CH. Managing early childhood caries with atraumatic restorative treatment and topical silver and fluoride agents. International Journal of Environmental Research and Public Health. 2017; 14: 1204.

[14] Meek JY, Noble L; Section on Breastfeeding. Policy statement: breastfeeding and the use of human milk. Pediatrics. 2022; 150: e2022057988.

[15] Shrestha SK, Arora A, Manohar N, Ekanayake K, Foster J. Association of breastfeeding and early childhood caries: a systematic review and meta-analysis. Nutrients. 2024; 16: 1355.

[16] Kato T, Yorifuji T, Yamakawa M, Inoue S, Saito K, Doi H, et al. Association of breast feeding with early childhood dental caries: Japanese population-based study. BMJ Open. 2015; 5: e006982.

[17] Abanto J, Maruyama JM, Pinheiro E, Matijasevich A, Antunes JLF, Bönecker M, et al. Prolonged breastfeeding, sugar consumption and dental caries at 2 years of age: a birth cohort study. Community Dentistry and Oral Epidemiology. 2023; 51: 575–582.

[18] Chaffee BW, Feldens CA, Vítolo MR. Association of long-duration breastfeeding and dental caries estimated with marginal structural models. Annals of Epidemiology. 2014; 24: 448–454.

[19] Marwah N, Nigam AG, Yadav SP, Meghpara M, Godhani S, Chalana S. Association of early childhood caries with feeding, dietary habits, and oral hygiene practices among rural and urban school children of Jaipur. International Journal of Clinical Pediatric Dentistry. 2022; 15: 273–279.

[20] Panchanadikar NT, S A, Muthu MS, H S, Jayakumar P, Agarwal A. breastfeeding and its association with early childhood caries—an umbrella review. Journal of Clinical Pediatric Dentistry. 2022; 46: 75–85.

[21] Gomersall JC, Slack-Smith L, Kilpatrick N, Muthu MS, Riggs E. Interventions with pregnant women, new mothers and other primary caregivers for preventing early childhood caries. Cochrane Database of Systematic Reviews. 2024; 5: CD012155.

[22] Bernabé E, MacRitchie H, Longbottom C, Pitts NB, Sabbah W. Birth weight, breastfeeding, maternal smoking and caries trajectories. Journal of Dental Research. 2017; 96: 171–178.

[23] Salli K, Söderling E, Hirvonen J, Gürsoy UK, Ouwehand AC. Influence of 2′-fucosyllactose and galacto-oligosaccharides on the growth and adhesion of Streptococcus mutans. British Journal of Nutrition. 2020; 124: 824–831.

[24] Human Milk Banking Association of North America. HMBANA standards for donor human milk banking: an overview. 2024. Available at: https://www.hmbana.org/file_download/inline/c4bd9e2e-4257-4441-a93a-94cc7647b304 (Accessed: 29 July 2025).

[25] Ye L, Cao L, Song W, Yang C, Tang Q, Yuan Z. Interaction between apical periodontitis and systemic disease (Review). International Journal of Molecular Medicine. 2023; 52: 60.

[26] Isola G, Polizzi A, Santagati M, Alibrandi A, Iorio-Siciliano V, Ramaglia L. Effect of nonsurgical mechanical debridement with or without chlorhexidine formulations in the treatment of peri-implant mucositis. A randomized placebo-controlled clinical trial. Clinical Oral Implants Research. 2025; 36: 566–577.