Abstract Objective This study aimed to analyze the functional profile of supragingival biofilm from sound (CAs), active (CAa), and inactive (CAi) enamel caries lesions from caries-active individuals to provide insights into the diversity of biological processes regarding biofilm dysbiosis. Methodology A metatranscriptome analysis was performed in biofilm samples collected from five caries-active individuals. Total RNA was extracted, and the microbial cDNAs were obtained and sequenced (Illumina HiSeq3000). Trimmed data were submitted to the SqueezeMeta pipeline in the co-assembly mode for functional analysis and further differential gene expression analysis (DESeq2). Results Bioinformatics analysis of mRNAs revealed a similar functional profile related to all analyzed conditions (CAa, CAi, and CAs). However, active and inactive surfaces share up-regulated genes (gtsA; qrtT; tqsA; pimB; EPHX1) related to virulence traits that were not overrepresented in sound surfaces. From a functional perspective, what matters most is the individual carious status rather than the surface condition. Therefore, pooling samples from various sites can be carried out using naturally developed oral biofilms but should preferably include carious surfaces. Conclusion Metatranscriptome data from subjects with caries activity have shown that biofilms from sound, arrested, and active lesions are similar in composition and function. CAs, CAs , (CAs) CAa, CAa (CAa) CAi (CAi cariesactive dysbiosis extracted Illumina HiSeq3000. HiSeq3000 HiSeq . HiSeq3000) coassembly co assembly DESeq2. DESeq2 DESeq (DESeq2) (CAa CAs. CAs) However upregulated up regulated gtsA (gtsA qrtT tqsA pimB EPHX1 EPHX perspective condition Therefore arrested function (CAs HiSeq300 (DESeq2 HiSeq30 (DESeq HiSeq3

Abstract The high concentration of glucose in the blood in Type 2 diabetes (T2D) may be related to either insulin resistance or insulin deficiency. Moreover, the literature points to periodontitis as the main oral disease caused by glycemia imbalance. The quantification of inflammatory markers in blood or saliva samples of T2D patients may represent a valuable tool in revealing how well an individual’s immune system can respond to injuries and periodontal treatment. In addition, an evaluation of the cytokine expression is extremely relevant to help understand the connection between periodontitis and T2D. This systematic review and meta-analysis aimed to evaluate the expression of inflammatory markers in T2D patients with periodontitis, compared with non-diabetic patients with periodontitis. A total of 3,894 studies were retrieved after a systematic literature search, 15 of which were included in the systematic review, and 4 of these 15, in the meta-analysis. The results did not indicate any statistical difference between the groups regarding TNF-α and IL-6 markers. T2D patients with periodontitis had increased levels of IL-10, compared with non-diabetic individuals with periodontitis (p = 0.003). On the other hand, the IL-4 concentration in non-diabetic individuals with periodontitis was high, compared with the T2D group (p< 0.001). Several studies did not include quantitative results and were excluded from the meta-analysis. The high IL-10 expression and low IL-4 expression in the T2D group suggest an association between the level of these markers and the impairment of the immune response in T2D patients with periodontitis.

Aims: This pilot study aimed to describe the majority and minority microbiota of saliva from individuals with advanced periodontitis, with or without type 2 diabetes mellitus (T2D), correlating the relative abundance of microorganisms with clinical parameters. Material/Methods: Six individuals diagnosed with periodontitis were included and classified according to their diagnosis of T2D. Salivary pH, number of teeth with active caries lesions, number of remaining teeth, periodontal and glycemic parameters were evaluated. V4 region amplicons of the 16S rRNA from salivary DNA were sequenced at Ion PGM. Amplicon sequence variants (ASVs) were compared according to the clinical parameters. Results: Correlation showed eight low-abundant bacteria significantly correlated with co-variables, either positively or negatively. The periodontitis-associated bacteria followed the increasing pH and number of remaining teeth. Conclusions: This survey provided potential minority microbiota correlations with clinical parameters, such as number of remaining teeth, FBG, and salivary pH. The ubiquity of some low abundant microorganisms in individuals with advanced periodontitis, exposed or not to type 2 DM, can reveal microbial signatures not yet explored.

Abstract This study aimed to assess the prevalence, extent, and associated factors of underlying dentin shadows (UDS) in the occlusal surfaces of permanent posterior teeth among 12-year-old schoolchildren from southern Brazil. A population-based cross-sectional study that included a representative sample of 1,528 schoolchildren was conducted in Porto Alegre, southern Brazil (participation rate: 83.2%). A questionnaire on sociodemographic characteristics, behavioral habits, and access to dental services was sent to parents/legal guardians of each student selected. Caries examination included the recording of non-cavitated and cavitated lesions, including the presence of UDS, as defined by the ICDAS Group (i.e., a shadow of discolored dentin visible through an apparently intact enamel surface that may or may not show signs of localized breakdown – ICDAS Code 4). The association between predictor variables and UDS prevalence was assessed using survey Poisson regression models. The prevalence of UDS was 6.3% (95%CI = 1.2 –11.3). The mean number of affected teeth among the schoolchildren presenting UDS was 1.51 (95%CI = 1.40 –1.61), ranging from 1 to 6. Type of school (public, PR = 2.23, 95%CI = 1.22 –4.07) and caries experience (DMFT 1-2, PR = 2.41, 95%CI = 1.15 –5.04; DMFT ≥ 3, PR = 3.09, 95%CI = 1.52 –6.27) were significantly associated with UDS prevalence. In conclusion, this population-based cross-sectional study found a low prevalence of UDS in the occlusal surfaces of the permanent posterior teeth of 12-year-old southern Brazilian schoolchildren. Overall caries experience and type of school were associated with UDS in this population.

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.

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.

Resumo O objetivo deste ensaio clínico randomizado foi comparar os microrganismos remanescentes após tratamentos baseados em remoção total de tecido cariado e selamento e a remoção seletiva de tecido cariado e selamento. Pacientes com lesões de cárie ativas em molares permanentes foram divididos aleatoriamente em dois grupos: grupo teste (remoção seletiva de tecido cariado-SCR; n=18), e grupo de controle (remoção total de tecido cariado-CCR; n=18). Amostras de dentina foram obtidas após a remoção da tecido cariado e após 3 meses de selamento das cavidades. Streptococcus spp., Streptococcus mutans, Lactobacillus spp. e microrganismos viáveis totais foram cultivados para contagem de células e frequência de isolamento de espécies. CCR resultou em menores contagens totais de microorganismos viáveis (p≤0,001), Streptococcus spp. (p≤0,001) e Lactobacillus spp. (p≤0,001) inicialmente. Entretanto, após o selamento, uma redução significativa nas contagens totais de microrganismos viáveis, Streptococcus spp. e Lactobacillus spp. resultou em nenhuma diferença entre os grupos após 3 meses. Conclui-se que a remoção seletiva de cárie é tão seletiva quanto a remoção completa de cárie na redução da infecção dentinária após três meses com selamento da lesão.

Abstract The aim of this randomized clinical trial was to compare the remaining microbial load after treatments based on complete and selective caries removal and sealing. Patients with active carious lesions in a permanent molar were randomly allocated into 2 groups: a test group (selective caries removal-SCR; n=18) and a control group (complete caries removal - CCR; n=18). Dentin samples were collected following the excavation and three months after sealing. Streptococcus species, Streptococcus mutans, Lactobacillus species, and total viable microorganisms were cultured to count the viable cells and frequency of species isolation. CCR resulted in significant lower total viable microorganisms counts (p≤0.001), Streptococcus species (p≤0.001) and Lactobacillus species (p≤0.001) initially. However, after sealing, a decrease in total viable microorganisms, Streptococcus species, and Lactobacillus species in the SCR resulted in no difference between the groups after 3 months. In conclusion, selective caries removal is as effective as complete caries removal in reducing dentin bacterial load 3 months after sealing.