Identification of Non- Streptococcus mutans Bacteria from Predente Infant Saliva Grown on Mitis-Salivarius-Bacitracin Agar

Abstract

Objective: Although mitis-salivarius-bacitracin (MSB) agar is a commonly used selective medium for detecting Streptococcus mutans in clinical studies, non-S. mutans microorganisms are cultivatable on MSB agar. Since few studies have identified non-S. mutans bacteria grown on MSB, this study aimed to identify and differentiate MSB-grown non-S. mutans bacteria from predente infants' oral cavity. Study design: The saliva from 51 predente infants were plated on MSB agars. Bacteria colonies were characterized based on their morphology under direct visualization and light microscopic observation. Colony PCR targeting S. mutans htrA locus and 16S rRNA DNA sequencing were used for further bacteria identification. Results: Overall, 80% of the predente infants had oral bacteria grown on the MSB agar. Nine bacteria were identified, including S. mutans, Staphylococcus epidermidis, Klebsiella quasi-pneumoniae, Klebsiella pneumoniae, Enterobacter kobei, Enterococcus faecalis, Staphylococcus hominis, Streptococcus anginosus and Phytobacter. The most frequently detected bacteria were S. epidermidis (41.5%), followed by E. kobei (24.4%), K. pneumoniae (17.1%) and S. mutans (9.8%.) Conclusions: Multiple non-S. mutans bacteria from infants' oral cavity could grow on MSB agar. Caution should be exercised in counting the colony forming units of S. mutans from oral samples on MSB agar to avoid overestimation by assuming that all colonies on the MSB agar are S. mutans. Using the colony morphological guide we summarized, these non-S. mutans bacteria could be distinguished from S. mutans. Our study provides a key reference to pediatric cariology clinical-epidemiological studies that commonly use MSB to identify/quantify S. mutans in infants and young children.

Keywords: Colony morphology; Mitis-salivarius-bacitracin agar; Oral bacteria; Predente infants; Saliva; Streptococcus mutans.

Figure 1.. Colony morphology of bacteria grown on MSB agar

*Nine bacteria strains (A-1 to A-9 and B-1 to B-9) from predente infants’ saliva sample grew on MSB agar. A-10 and B-10 were Streptococcus sobrinus (ATCC® 33478™) wild strain.