Mônica Moreira

Methodological variations in the isolation of genomic DNA from Streptococcus bacteria

O genero Streptococcus encontra-se amplamente distribuido na natureza e algumas especies constituem a microbiota humana da cavidade bucal, como Streptococcus pyogenes, que pode estar associado a importantes doencas humanas, Streptococcus mutans e Streptococcus sobrinus, relacionados a carie dental. O DNA genomico destas tres especies foi isolado utilizando-se dois metodos, o primeiro utilizando o detergente brometo de cetiltrimetilamonio (CTAB) a 65oC e outro associando ultra-som a uma mistura de silica e celite em CTAB. O metodo que possibilitou a extracao do DNA genomico das bacterias Gram positivas, com qualidade, boa reprodutibilidade facil execucao foi aquele que utilizou ultra-som associado a silica e celite em CTAB.

Genetic variability of Streptococcus mutans isolated from low-income families, as shown by RAPD markers

The detection of Streptococcus mutans isolates with high genetic variability in different individuals indicates the occurrence of transmissibility. In this context, nine low-income families (40 individuals in total) with similar social conditions were assessed to identify the bioserotypes of S. mutans using both biochemical and RAPD markers and to establish the degree of similarity among the intra-familial isolates. The polymorphism analysis used the coefficient of Jaccard in both a Principal Coordinates Analysis (PCO) and in a UPGMA clustering method. A total of 157 isolates were obtained from salivary samples, using the morphology of colonies recovered using MSB agar as indicators. From those, 64 were characterized biochemically as S. mutans and 10 as S. sombrinus. Genetic variability among isolates based on RAPD markers was not consistent with their intra-familial distribution. In particular, some individuals might have experienced multiple infections given the high genetic variability among their isolates. The occurrence of 4 isolates with 100% genetic similarity is indicative of intra-familial transmission.

Detection of Streptococcus mutans using padlock probe based on Rolling Circle Amplification (RCA)

The aim of this study was to develop and evaluate a padlock probe based on the Rolling Circle Amplific ation (RCA), which targeted to 16S-23S rDNA region of S. mutans . The specificity of developed padlock probe was te sted for DNA within a panel strains, including S. mutans isolated from the saliva and reference strains of the genus Streptococcus , as well as total DNA samples of biofilm and saliv a. The results were positive either for DNA samples of S. mutans or DNA samples recovered from the biofilm and sali va revealing the specificity of designed padlock probe. The padlock probe based on the RCA was prove d to be an effective, reproducible method for S. mutans detection and demonstrated the possibility of a ra pid detection and accurate identification of S. mutans infection.