Bacterial analysis of biofilms from tooth root surfaces presenting different caries activity / Análise bacteriológica de biofilmes de superfícies dentárias radiculares apresentando diferentes estágios de atividade de cárie

This study evaluated the numbers and determined the proportion of mutans streptococci and Lactobacillus spp., which are possible relevant cariogenic organisms, in biofilms recovered from lesions at root surfaces with active caries lesions (ARC), inactive caries lesions, and sound root surfaces (SRS). Samples were cultured in MSB agar for mutans streptococci counts, Rogosa agar for Lactobacillus spp. counts, and brain-heart infusion agar for total viable anaerobic counts. After incubation, the number of colony-forming units (CFUs) was determined and compared between groups by the Mann-Whitney U test with a significance level set at 95%. The proportion of counts of mutans streptococci and Lactobacillus spp. in the total viable microorganisms was also analyzed by Chi-square test. Ninety samples (30 from each surface) from 37 patients were cultured and analyzed. The CFU was similar between mutans streptococci and Lactobacillus spp.These species were present in at least half of the samples and no difference was found in the frequency of isolation of these species. Only 6 samples showed a proportion of more than 10% of mutans streptococci; 4 of the samples were from ARC. Most (93%) SRS samples did not contain viable Lactobacillus spp. The data indicate the low counts of mutans streptococci and Lactobacillus spp. in root sur-faces, regardless of the activity of caries lesions.

O estudo analisou contagens e proporções de mutans strep-tococci e Lactobacillus spp., que podem ser microorganismos importantes em lesões de cárie radicular com diferentes atividades. Biofilmes foram coletados em três locais: ARC ­ superfície radicular com lesão ativa de cárie; IRC ­ superfícies radiculares com lesão inativa de cárie; SRS ­ superfícies de raizes hígidas. As amostras foram cultivadas em agar MSB para contagens de mutans streptococci; agar Rogosa para Lactobacillus spp., e agar BHI para contagens de microrganis-mos viáveis anaeróbicos totais. Após a incubação, o número de unidades formadoras de colônias (UFCs) foi determinado e comparado entre os grupos pelo teste de Mann-Whitney U test. O nível de significância foi estabelecido em 95%. A pro-porção de contagem de mutans streptococci e Lactobacillusspp. no total de microrganismos viáveis também foi analisado através do teste de qui-quadrado. Um total de 90 amostras de 37 pacientes foram cultivadas e analisadas: 30 amostras de ARC, 30 de IRC e 30 de SRS. Números de UFC foram seme-lhantes entre os grupos para ambos, mutans streptococci e Lactobacillus spp. Estas espécies estavam presentes em pelo menos metade de todas as amostras e nenhuma diferença foi encontrada na frequência de isolamento dessas espécies dentro dos grupos. Apenas 6 amostras apresentaram mais de 10% de mutans streptococci e 4 foram de ARC. Em relação aos Lactobacillus spp., 93% das amostras não apresentaram proporção dessas bactérias nas SRS. Mutans streptococci e Lactobacillus spp. estão presentes em baixa proporção nas superfícies radiculares, independentemente da atividade das lesões de cárie.

The role of Candida albicans in root caries biofilms: an RNA-seq analysis

Abstract Objective This study sought to analyze the gene expression of Candida albicans in sound root surface and root caries lesions, exploring its role in root caries pathogenesis. Methodology The differential gene expression of C. albicans and the specific genes related to cariogenic traits were studied in association with samples of biofilm collected from exposed sound root surface (SRS, n=10) and from biofilm and carious dentin of active root carious lesions (RC, n=9). The total microbial RNA was extracted, and the cDNA libraries were prepared and sequenced on the Illumina Hi-Seq2500. Unique reads were mapped to 163 oral microbial reference genomes including two chromosomes of C. albicans SC5314 (14,217 genes). The putative presence of C. albicans was estimated (sum of reads/total number of genes≥1) in each sample. Count data were normalized (using the DESeq method package) to analyze differential gene expression (using the DESeq2R package) applying the Benjamini-Hochberg correction (FDR<0.05). Results Two genes (CaO19.610, FDR=0.009; CaO19.2506, FDR=0.018) were up-regulated on SRS, and their functions are related to biofilm formation. Seven genes ( UTP20 , FDR=0.018; ITR1 , FDR=0.036; DHN6 , FDR=0.046; CaO19.7197 , FDR=0.046; CaO19.7838 , FDR=0.046; STT4 , FDR=0.046; GUT1 , FDR=0.046) were up-regulated on RC and their functions are related to metabolic activity, sugar transport, stress tolerance, invasion and pH regulation. The use of alternative carbon sources, including lactate, and the ability to form hypha may be a unique trait of C. albicans influencing biofilm virulence. Conclusions C. albicans is metabolically active in SRS and RC biofilm, with different roles in health and disease.

Gene expression profile of Scardovia spp. in the metatranscriptome of root caries

Abstract A few investigations of caries biofilms have identified Scardovia spp.; however, little is known about its involvement in caries pathogenesis. The purpose of this study was to assess the gene expression profile of Scardovia spp. in root caries, and compare it with other microorganisms. Clinical samples from active root caries lesions were collected. Microbial mRNA was isolated and cDNA sequenced. The function and composition of the Scardovia were investigated using two methods: a) de novo assembly of the read data and mapping to contigs, and b) reads mapping to reference genomes. Pearson correlation was performed (p < 0.05). Proportion of Scardovia inopinata and Scardovia wiggsiae sequences ranged from 0-6% in the root caries metatranscriptome. There was a positive correlation between the transcriptome of Lactobacillus spp. and Scardovia spp. (r = 0.70; p = 0.03), as well as with other Bifidobacteriaceae (r = 0.91; p = 0.0006). Genes that code for fructose 6-phosphate phosphoketolase (the key enzyme for "Bifid shunt"), as well as ABC transporters and glycosyl-hydrolases were highly expressed. In conclusion, "Bifid shunt" and starch metabolism are involved in carbohydrate metabolism of S. inopinata and S. wiggsiae in root caries. There is a positive correlation between the metabolism abundance of Lactobacillus spp., Bifidobacteriaceae members, and Scardovia in root caries.

Oral colonization by Streptococcus mutans and its association with the severity of periodontal disease in adults

Background: Streptococcus mutans (S. mutans) is associated with the onset of caries. Since root exposure in patients affected by periodontitis leads to higher caries rates, progressively more severe forms of periodontal disease might associate with elevated counts of S. mutans. Aim: To determine whether increasingly destructive forms of periodontal disease are associated with higher counts of S. mutans in untreated patients. Methods: 206 subjects aged 20-75 were classified into three groups according to the severity of periodontal disease: 1) gingivitis, 2) chronic slight periodontitis and 3) chronic moderate or chronic severe periodontitis. S. mutans counts (cfu/mL) were obtained by direct counting on selective agar plates from saliva samples. A cumulative proportional logistic regression model was adjusted for S. mutans counts. Results: The model failed to show differences by gender, but periodontal diagnosis had a significant effect on S. mutans counts depending on age. While in the group with moderate and severe periodontitis the probability of having high counts of S. mutans significantly increased with age, the probability remained unchanged in individuals with chronic slight periodontitis or gingivitis. Conclusion: High S. mutans levels appear directly co-associated with increased severity of periodontal disease at older ages in untreated patients.