A New Method to Extract Dental Pulp DNA: Application to Universal Detection of Bacteria

BACKGROUND: Dental pulp is used for PCR-based detection of DNA derived from host and bacteremic microorganims. Current protocols require odontology expertise for proper recovery of the dental pulp. Dental pulp specimen exposed to laboratory environment yields contaminants detected using universal 16S rDNA-based detection of bacteria. METHODOLOGY/PRINCIPAL FINDINGS: We developed a new protocol by encasing decontaminated tooth into sterile resin, extracting DNA into the dental pulp chamber itself and decontaminating PCR reagents by filtration and double restriction enzyme digestion. Application to 16S rDNA-based detection of bacteria in 144 teeth collected in 86 healthy people yielded a unique sequence in only 14 teeth (9.7%) from 12 individuals (14%). Each individual yielded a unique 16S rDNA sequence in 1-2 teeth per individual. Negative controls remained negative. Bacterial identifications were all confirmed by amplification and sequencing of specific rpoB sequence. CONCLUSIONS/SIGNIFICANCE: The new protocol prevented laboratory contamination of the dental pulp. It allowed the detection of bacteria responsible for dental pulp colonization from blood and periodontal tissue. Only 10% such samples contained 16S rDNA. It provides a new tool for the retrospective diagnostic of bacteremia by allowing the universal detection of bacterial DNA in animal and human, contemporary or ancient tooth. It could be further applied to identification of host DNA in forensic medicine and anthropology

Genetic variation in a tropical tree species influences the associated epiphytic plant and invertebrate communities in a complex forest ecosystem

Genetic differences among tree species, their hybrids and within tree species are known to influence associated ecological communities and ecosystem processes in areas of limited species diversity. The extent to which this same phenomenon occurs based on genetic variation within a single tree species, in a diverse complex ecosystem such as a tropical forest, is unknown. The level of biodiversity and complexity of the ecosystem may reduce the impact of a single tree species on associated communities. We assessed the influence of within-species genetic variation in the tree Brosimum alicastrum (Moraceae) on associated epiphytic and invertebrate communities in a neotropical rainforest. We found a significant positive association between genetic distance of trees and community difference of the epiphytic plants growing on the tree, the invertebrates living among the leaf litter around the base of the tree, and the invertebrates found on the tree trunk. This means that the more genetically similar trees are host to more similar epiphyte and invertebrate communities. Our work has implications for whole ecosystem conservation management, since maintaining sufficient genetic diversity at the primary producer level will enhance species diversity of other plants and animals

Comparison of orange oil and chloroform as gutta- percha solvents in endodontic retreatment

Objective: To compare the efficacy of gutta-percha removal achieved with simple mechanical technique using two different solvents.MATERIALS AND Methods: It was an in vitro experimental study conducted at the Aga Khan University Hospital, Karachi, Pakistan on 27 extracted human molars divided into two groups by alternate sampling technique. The teeth were prepared by manual filing and obturated with gutta-percha and calcium hydroxide sealer. Two drops of the assigned solvent were placed on the orifice of the obturated canal and Gates Glidden drills #1, 2 and 3 were used for removal of coronal gutta-percha. Manual filing was done for removal of remaining gutta-percha and the solvent was used in drops until needed. Postoperative periapical radiographs were taken to visualize the remaining gutta-percha left in the canals. Descriptive statistics (mean and SD) of the variables such as tooth type, obturation length and canal curvatures were determined. Chi-square and independent sample t-tests were applied and level of significance was set at 0.05.Results: There was 5.19 (3.8) mm of remaining gutta-percha in the orange oil group and 5.37 (4.2) mm in the chloroform group (p=0.90).CONCLUSION: There is no statistically significant difference between the orange oil and chloroform when used as solvent for removing gutta-percha