Microbial ecology and predicted metabolic pathways in various oral environments from patients with acute endodontic infections.

AIM: To assess in a cross-sectional clinical study the effect of antibiotics on the diversity, structure and metabolic pathways of bacterial communities in various oral environments in patients with acute primary infections. METHODOLOGY: Samples of saliva (SA), supragingival biofilm (SB) and from the pulp cavity (PC) were collected from teeth with acute primary infections and then grouped according to previous use of antibiotics (NoAtb = no antibiotics [n = 6]; Atb = antibiotics [n = 6]). DNA sequencing was conducted using MiSeq (Illumina, San Diego, CA, USA). The V1-V3 hyper-variable region of the 16S rRNA gene was amplified. A custom Mothur pipeline was used for 16S rRNA processing. Subsequent analyses of the sequence dataset were performed in R (using vegan, phyloseq and ggplot2 packages) or QIIME. RESULTS: Twelve patients aged from 22 to 56 years were recruited. Participants in the Atb group had taken the beta-lactamics amoxicillin (5/6) or cephalexin (1/6) for 2-3 days. A total of 332 bacterial taxa (OTUs) were identified, belonging to 120 genera, 60 families and nine phyla. Firmicutes (41%) and Bacteroidetes (38%) were the most abundant phyla in all samples. Taxa clustered significantly by oral site (PCoA analysis; P < 0.05, ANOSIM). Use of antibiotics had little effect on this clustering. However, SA, SB and PC had different degrees of richness, diversity and evenness. The greatest diversity was observed in SB samples and the least diversity was observed in PC samples. Metabolic prediction identified 163 pathways and previous use of antibiotics had a major effect on the estimated functional clustering in SA and PC samples. CONCLUSION: The ecological niche had a strong influence on the bacterial content of samples from various oral sites. Previous exposure to antibiotics may exert an effect on the phylogenetic composition of SA. Metabolic pathways appear to be modulated by antimicrobial agents in SA and PC samples. The dynamics of host/microbial interactions in the apical region and the functional ecology of the infected pulp cavity should be revisited.

Prevotella strains and lactamic resistance gene distribution in different oral environments of children with pulp necrosis.

AIM: To determine the presence of Prevotella species, the cfxA/cfxA2, blaZ and blaTEM genes associated with resistance to lactamic agents in different oral niches of children with pulp necrosis. METHODOLOGY: Children with pulp necrosis in primary teeth had samples of saliva, supragingival, pulp chamber and root canal biofilms collected and tested for Prevotella species (P. intermedia, P. nigrescens, P. tannerae) and for beta-lactam resistance genes (cfxA/cfxA2, blaZ and blaTEM). The presence of bacterial DNA was examined through PCR, with a specific primer directed to the 16S rRNA gene. Specific primers were used to detect the Prevotella species and beta-lactam resistance genes. The chi-square test was used to analyse associations between the presence of bacteria and clinical variables. The Cochran's Q test was used to assess whether the proportion of gene detection is the same between different sites. RESULTS: Thirty-two teeth were sampled from 27 patients with a mean age of 5.5 years (±1.76). The total detection rate of Prevotella strains was 29.1%, 25%, 21.8% and 32.29% in saliva, supragingival, pulp chamber and root canal samples, respectively. P. nigrescens was the most commonly detected species in all oral niches. The previous use of antibiotics was associated with detection of P. nigrescens in saliva (P = 0.03). Pain was associated with the presence of P. nigrescens (P = 0.04) and P. tannerae (P = 0.01) in pulp chamber biofilm. blaTEM was detected in the four oral niches, being more frequent (23.8%) in supragingival biofilm (Cochran's Q test, P = 0.04). The presence of P. intermedia in SB and PC was associated with the detection of blaTEM in saliva (P = 0.04). The cfxA/cfxA2 and blaZ genes were not detected in any of the four oral niches. CONCLUSIONS: The oral cavity of children with pulp necrosis had a variable distribution of Prevotella strains in different niches. Saliva, supragingival biofilm, pulp chamber and root canals of primary teeth with necrotic pulps can harbour resistance genes to beta-lactams agents.