DNase Activity of <i>Prevotella intermedia</i> Impairs Biofilm Development and Neutrophil Extracellular Trap Formation

Chronic periodotitis is caused by the formation of biofilms. Prevotella intermedia, a gram-negative obligate anaerobic bacterium residing in periodontal pockets is involved in the formation of biofilms and secrets a highly potent DNA-degrading activity. Biofilm contains extracellular DNA as a structural component, suggesting that DNase activity may influence P. intermedia's own biofilm development. Neutrophil extracellular traps (NETs) have mesh-like structures and composed of DNA, histone and antibacterial proteins. NETs play an important role in protecting against infection, but it is possible that DNase of P. intermedia disrupts NETs. The lack of established genetic manipulation has significantly delayed the analysis of DNase pathogenic factors. Recently, we have succeeded in establishing a genetic manipulation technique for P. intermedia. In this study, we created strains lacking two DNase candidate genes, nucA (PIOMA14_I_0621) and nucD (PIOMA14_II_0624), that were highly conserved among P. intermedia strains. We examined biochemical analysis of DNase activity, their effection on biofilm formation, and their evasion of NETs. Here, we showed both of them possessed DNase activities which appeared to account all of DNase activities of the bacterium. The mutant analysis further demonstrated that NucA and NucD destroyed biofilm and NETs formations. Neither one was perfectly responsible for DNase activity, but rather they take turns depending on the conditions. In conclusion, the nucA and nucD genes encode DNases that cooperatively function on biofilm formation and suppress NETs formation in P. intermedia.