Genotypic Diversity of Mutans Streptococci in Brazilian Nursery Children Suggests Horizontal Transmission

Abstract

Streptococcus mutans strains were isolated from cohorts of Brazilian nursery school children and genotyped by arbitrarily primed PCR and restriction fragment length polymorphism analysis. Of 24 children with two to five S. mutans isolates, 29% carried two or more genotypes. The presence of matching genotypes of S. mutans among children attending one nursery suggests horizontal transmission.Dental caries is a transmissible infectious disease in which mutans streptococci (MS) play the major role. Infective strains of Streptococcus mutans, the most prevalent species of the MS group, may persist for many years in the mouths of preschool children (1,6,22). Early colonization is related to high caries activity during childhood (3,7,11). The mechanisms by which MS colonize and accumulate on tooth surfaces are not completely clear. In addition to environmental and host factors, specific genotypes of S. mutans may be more aggressive colonizers. This is suggested by previous findings of positive relationships between the production of water-insoluble glucan from sucrose by glucosyltransferase (GTF) activities and the intensity of MS oral colonization and caries incidence (19).Essential to the development of strategies for the prevention of dental caries is the identification of sources of MS transmission. DNA fingerprinting studies indicate that vertical transmission of bacteria from mother to child is the major route for early acquisition (2,13,15,18,21). However, detection of genotypes in children that are not found in their mothers or other family members indicates that MS may also be acquired from other sources (13,21,22). Since the spread of infectious agents is likely to occur in a nursery environment, we investigated the genetic similarity of MS strains isolated from Brazilian children attending nursery schools. The production of GTF isozymes was also examined to validate the genotypic similarities that we identified.The study group included 35 MS-infected children between 12 and 30 months of age (mean Ϯ standard deviation ϭ 23 Ϯ 5 months). This group accounted for 49% of the MS-colonized children previously described in a larger population (20), and it was primarily selected for the study of MS virulence factors. These children attended nine nursery schools in the city of Piracicaba, São Paulo, Brazil, for 5 days per week, 10 h per day. A total of four sucrose-rich meals were provided daily in the nurseries. Clinical exams were performed to record the number of erupted teeth and manifest caries lesions as previously described (20). Written informed consent was obtained from the parents, and all consent and experimental procedures were approved by the institutional Ethical Committee of the University of São Paulo School of Dentistry.One to five isolates of MS were recovered from each of the 35 children. As previously described (19), oral samples were collected with tongue blades which were then pressed on the surface of mitis salivarius agar contact plates (Difco) (12) containing 2 IU of bacitracin (Sigma)/ml and 1...

Aleitamento materno e cárie do lactente e do pré-escolar: uma revisão crítica

“…7,10,[29][30][31] Horizontal transmission also has been described in nursery facilities of day-care centers and within families. 31,32 The severity of ECC is directly related to the early establishment of mutans streptococci in the infant. 10,30 These bacteria admittedly need nondesquamative surfaces to colonize because their positivity increases with the number of erupted teeth and with age.…”

Genotypic diversity of S. mutans in dental biofilm formed in situ under sugar stress exposure

“…Page W. Caufield, New York University, NY, USA) and distilled and deionized water were applied in all PCR baths, as positive and negative controls. Products of AP-PCR were electrophoretically resolved in 1.5% agarose gels that were run at 3V/cm during 3 h in TBE running buffer (16). The gel was stained with a 5 μg/mL of ethidium bromide solution (Invitrogen) for 10 min and their images captured by a digital imaging system (Gel logic 100 Imaging System, Kodak, Japan).…”

“…AP-PCR assays were performed with the arbitrary primer OPA 02 (5'-TGCCGAGCTG-3´) (16). The amplifications occurred under the following conditions: 95ºC for 2 min, for initial denaturation, and 45 cycles of 94ºC for 30 s (denaturation), 36ºC for 30 s (annealing) and 72ºC for 1 min (extension) and a final extension at 72ºC for 5 min.…”

“…The colonies from BHI agar were inoculated into Todd Hewitt Broth (Difco) and incubated for 18 h, at 37ºC and 10% pCO 2 . Cultures were then centrifuged (Jouan, France) at 10,000 g, 4ºC for 15 min, genomic DNA was extracted from the cell pellet, using the Master Pure DNA purification kit (Epicentre Technologies, Madison, WI, USA) (16), and stored at -20ºC. Integrities of the genomic DNA samples were checked in samples electrophoretically resolved in 1% agarose gel (Invitrogen, Barcelona, Spain) and stained with ethidium bromide (5 μg/mL).…”

“…The exclusion criteria were antibiotic use for the last 2 months before starting the study, use of any form of medication that modifies salivary secretion, use of fixed or removable orthodontic appliance, periodontal disease or general/systemic illness. Eight representative morphological types of S. mutans colonies were collected from each saliva sample of the selected volunteers, subcultured on MSA and Brain Heart Infusion (BHI) agar (Difco), and pure cultures stored at -70ºC in 10% skim milk medium (Difco) (16) for further genotypic analysis. The purity and identity of the isolates were checked by Gram's staining and colonial morphology on MSA.…”