Biofilm dysbiosis and caries activity: a surface or an individual issue?

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.

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.

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.

Microbial load after selective and complete caries removal in permanent molars: a randomized clinical trial

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.

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.

Estudo dos lactobacilos no biofilme dental / Study of lactobacilli in dental biofilm

Lactobacilos são um grupo de bactérias relacionadas com cárie dental. Há falta de estudos sobre a biologia populacional dos lactobacilos na cárie dental. O objetivo do estudo foi avaliar o efeito das modificações ambientais na composição, diversidade genética dos lactobacilos e identificar a filogenia dos Lactobacillus paracasei isolados do biofilme. Lactobacilos foram isolados em um modelo de formação de biofilme in situ antes e depois de 28 dias de exposição à solução de sacarose 20%. As colônias foram randomicamente selecionadas do meio Rogosa Ágar e subcultivadas (n=222, 31 antes e 191 após período de exposição à sacarose). Os isolados foram identificados usando o seqüenciamento parcial dos genes pheS ou rpoA. As espécies de lactobacilos predominantes encontradas foram L. paracasei, L. fermentum e L. rhamnosus. A diferença na composição e na diversidade genética de lactobacilos associada às modificações ambientais no biofilme foi analisada através de PCR utilizando palíndromes repetitivos extragênicos (REP-PCR). Após a fase com sacarose, um maior número de lactobacilos pode ser encontrado no biofilme...

Lactobacilli are a group of bactéria related to dental caries. There is a lack of studies on the population biology of this organisms in dental caries. The aim of the study was to evaluate the effect of the environmental changes in the composition, genetic diversity and identify the filogeny of Lactobacillus paracasei isolated from biofilm. Lactobacilli were isolated from a biofilm model, formed in situ prior to and during a 28-day period of exposure to 20% sucrose solution. The lactobacillus colonies were randomly selected from Rogosa Agar medium and subcultured (n=222, 31 prior to and 191 following a sucrose exposure period). The isolates were identified using pheS or rpoA gene sequence analysis. The predominant lactobacilli were L. paracasei, L. fermentum and L. rhamnosus. The difference in composition...

Estudo dos lactobacilos no biofilme dental / Study of lactobacillus in oral film

Lactobacilos são um grupo de bactérias relacionadas com cárie dental. Há falta de estudos sobre a biologia populacional dos lactobacilos na cárie dental. O objetivo do estudo foi avaliar o efeito das modificações ambientais na composição, diversidade genética dos lactobacilos e identificar a filogenia dos Lactobacillus paracasei isolados do biofilme. Lactobacilos foram isolados em um modelo de formação de biofilme in situ antes e depois de 28 dias de exposição à solução de sacarose 20%. As colônias foram randomicamente selecionadas do meio Rogosa Ágar e subcultivadas (n=222, 31 antes e 191 após período de exposição à sacarose). Os isolados foram identificados usando o seqüenciamento parcial dos genes pheS ou rpoA. As espécies de lactobacilos predominantes encontradas foram L. paracasei, L. fermentum e L. rhamnosus. A diferença na composição e na diversidade genética de lactobacilos associada às modificações ambientais no biofilme foi analisada através de PCR utilizando palíndromes repetitivos extragênicos (REP-PCR). Após a fase com sacarose, um maior número de lactobacilos pode ser encontrado no biofilme...

Lactobacilli are a group of bactéria related to dental caries. There is a lack of studies on the population biology of this organisms in dental caries. The aim of the study was to evaluate the effect of the environmental changes in the composition, genetic diversity and identify the filogeny of Lactobacillus paracasei isolated from biofilm. Lactobacilli were isolated from a biofilm model, formed in situ prior to and during a 28-day period of exposure to 20% sucrose solution. The lactobacillus colonies were randomly selected from Rogosa Agar medium and subcultured (n=222, 31 prior to and 191 following a sucrose exposure period). The isolates were identified using pheS or rpoA gene sequence analysis. The predominant lactobacilli were L. paracasei, L. fermentum and L. rhamnosus. The difference in composition...

Comparative analysis of the effect of two chlorhexidine mouthrinses on plaque accumulation and gingival bleeding

The aim of the present study was to evaluate the effect of two chlorhexidine rinsing solutions (0.12 percent and 0.2 percent) on plaque and gingival bleeding. Ten dental students participated in this double-blind, cross-over study, rinsing twice a day, for one minute, with each one of the tested solutions for fourteen days. A wash-out period of one week between treatments was observed. In order to assess gingival bleeding, the van der Weijden et al.¹ (1994) index was used. The plaque indexes used were those of Quigley, Hein² (1962) and Silness, Löe³ (1964). In the pre-experimental period, subjects received oral hygiene instructions and dental prophylaxis. The results revealed no significant differences between both concentrations in relation to plaque and gingival bleeding. Mean values (± standard deviation) of the Quigley & Hein index were 0.25 ± 0.16 for the 0.12 percent solution and 0.23 ± 0.26 for the 0.2 percent solution (p = 0.4838). Mean values (± standard deviation) of the Silness-Löe index were 0.12 ± 0.10 for the 0.12 percent solution and 0.11 ± 0.11 for the 0.2 percent solution (p = 0.7592). The bleeding index mean values at the end of the study were not different for both concentrations with mean values (± standard deviation) of 14.93 percent ± 6.68 percent and 13.95 ± 9.24 percent for the 0.12 percent and 0.2 percent solutions, respectively. Although an increase in gingival bleeding was observed, both concentrations were able to control dental plaque.