Analysis of interspecies adherence of oral bacteria using a membrane binding assay coupled with polymerase chain reaction‐denaturing gradient gel electrophoresis profiling

Abstract

Information on co-adherence of different oral bacterial species is important for understanding interspecies interactions within oral microbial community. Current knowledge on this topic is heavily based on pariwise coaggregation of known, cultivable species. In this study, we employed a membrane binding assay coupled with polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) to systematically analyze the co-adherence profiles of oral bacterial species, and achieved a more profound knowledge beyond pairwise coaggregation. Two oral bacterial species were selected to serve as "bait": Fusobacterium nucleatum (F. nucleatum) whose ability to adhere to a multitude of oral bacterial species has been extensively studied for pairwise interactions and Streptococcus mutans (S. mutans) whose interacting partners are largely unknown. To enable screening of interacting partner species within bacterial mixtures, cells of the "bait" oral bacterium were immobilized on nitrocellulose membranes which were washed and blocked to prevent unspecific binding. The "prey" bacterial mixtures (including known species or natural saliva samples) were added, unbound cells were washed off after the incubation period and the remaining cells were eluted using 0.2 mol· L -1 glycine. Genomic DNA was extracted, subjected to 16S rRNA PCR amplification and separation of the resulting PCR products by DGGE. Selected bands were recovered from the gel, sequenced and identified via Nucleotide BLAST searches against different databases. While few bacterial species bound to S. mutans, consistent with previous findings F. nucleatum adhered to a variety of bacterial species including uncultivable and uncharacterized ones. This new approach can more effectively analyze the co-adherence profiles of oral bacteria, and could facilitate the systematic study of interbacterial binding of oral microbial species.

Fusobacterium Species and Subspecies Differentially Affect the Composition and Architecture of Supra- and Subgingival Biofilms Models

“…polymorphum adhesin RadD by the S. mutans adhesin SpaP (Guo et al, 2017). Thus, the RaD/SpaP adhesin pair serves as a binding mechanism for fusobacteria and implies their high virulence potential, since S. mutans interacts with only a limited number of other bacterial species (Wang et al, 2011).…”

Inhibition of Streptococcus mutans biofilm formation, extracellular polysaccharide production, and virulence by an oxazole derivative

“…In addition, as dental biofilm formation is a dynamic process (Wang et al 2011), and with more than 600 bacterial species identified from the oral cavity, further study is required to determine whether the inhibitor can affect the complicated bacterial interactions between dental biofilm microorganisms. The impact of compound 5H6 on biofilm biology also requires further investigation.…”

Streptococcus mutans SpaP binds to RadD of Fusobacterium nucleatum ssp. polymorphum

“…This is the first time a cognate adhesin pair was identified for attachment of a fusobacterial species to another organism. Streptococcus mutans does not interact with a large number of other bacterial species, but it is an interesting phenomenon that the ubiquitous fusobacteria, which are known to bind to numerous other microorganisms, recognize at least two different surface structures on S. mutans for attachment. This could possibly expand their own ability as well as that of S. mutans to effectively colonize available oral surfaces.…”