Identification of a Novel Two-Component System inStreptococcus gordoniiV288 Involved in Biofilm Formation

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Abstract
Streptococcus gordoniiis a pioneer colonizer of the teeth, contributing to the initiation of the oral biofilm called dental plaque. To identify genes that may be important in biofilm formation, a plasmid integration library ofS. gordoniiV288 was used. After screening for in vitro biofilm formation on polystyrene, a putative biofilm-defective mutant was isolated. In this mutant, pAK36 was inserted into a locus encoding a novel two-component system (bfr[biofilm formation related]) with two cotranscribed genes that form an operon.bfrAencodes a putative response regulator, whilebfrBencodes a receptor histidine kinase. Thebfrmutant and wild-type strain V288 showed similar growth rates in Todd-Hewitt broth (THB). Abfr-catfusion strain was constructed. During growth in THB, the reporter activity (chloramphenicol acetyltransferase) was first detected in mid-log phase and reached a maximum in stationary phase, suggesting that transcription ofbfrwas growth stage dependent. After being harvested from THB, thebfrmutant adhered less effectively than did wild-type strain V288 to saliva-coated hydroxyapatite (sHA). To simulate pioneer colonization of teeth,S. gordoniiV288 was incubated with sHA for 4 h in THB with 10% saliva to develop biofilms. RNA was isolated, and expression ofbfrABwas estimated. In comparison to that of cells grown in suspension (free-growing cells),bfrmRNA expression by sessile cells on sHA was 1.8-fold greater and that by surrounding planktonic cells was 3.5-fold greater. Therefore,bfrABis a novel two-component system regulated in association withS. gordoniibiofilm formation in vitro.