IADR Abstract Archives
Natural Competence is Common Among Clinical Isolates of Veillonella parvula and is Mediated by a Widely Conserved Competence Locus
Objectives: The six Veillonella species commonly found in the human oral cavity are among the most abundant members of the oral flora, occurring in both supra- and subgingival dental plaque as well as being present in periodontal disease. Veillonellae also play critical roles in the microbial ecology of the oral flora via their ability to facilitate the growth of numerous other species. Despite their high abundance and ecological significance, surprisingly little is known about Veillonella biology, largely due to the difficulties associated with their genetic manipulation. Our goal was to determine whether natural competence exists in this genus and if this ability can be used for the genetic manipulation of these species.
Methods: Clinical samples of buccal molar plaque were collected from a pediatric cohort. Veillonella clinical strains were isolated through a newly developed Veillonella enrichment protocol. Isolates were identified by sequencing their 16S rRNA genes and/or rpoB alleles. Confirmed Veillonella isolates were tested for natural competence by transforming either an E. coli-Veillonella shuttle vector or PCR products using a novel agar plate transformation protocol developed for this study. Natural competence was used for allelic exchange mutagenesis and assays of host specificity for DNA uptake.
Results: Out of 67 clinical plaque samples, 43 confirmed Veillonella isolates were obtained and 6 of these were naturally competent. Natural competence was found to be strongly influenced by growth medium composition. Natural competence was also selective against foreign DNA. Using host methylated DNA, a large percentage of clinical isolates of Veillonella were found to be naturally competent. Genomic comparison suggests that natural competence is widely distributed among the Veillonellaceae family.
Conclusions: To the best of our knowledge, this study is the first demonstration of natural competence and targeted allelic exchange mutagenesis within the entire Veillonellaceae family and demonstrates a simple and rapid approach to study Veillonella genetics.
Methods: Clinical samples of buccal molar plaque were collected from a pediatric cohort. Veillonella clinical strains were isolated through a newly developed Veillonella enrichment protocol. Isolates were identified by sequencing their 16S rRNA genes and/or rpoB alleles. Confirmed Veillonella isolates were tested for natural competence by transforming either an E. coli-Veillonella shuttle vector or PCR products using a novel agar plate transformation protocol developed for this study. Natural competence was used for allelic exchange mutagenesis and assays of host specificity for DNA uptake.
Results: Out of 67 clinical plaque samples, 43 confirmed Veillonella isolates were obtained and 6 of these were naturally competent. Natural competence was found to be strongly influenced by growth medium composition. Natural competence was also selective against foreign DNA. Using host methylated DNA, a large percentage of clinical isolates of Veillonella were found to be naturally competent. Genomic comparison suggests that natural competence is widely distributed among the Veillonellaceae family.
Conclusions: To the best of our knowledge, this study is the first demonstration of natural competence and targeted allelic exchange mutagenesis within the entire Veillonellaceae family and demonstrates a simple and rapid approach to study Veillonella genetics.