The Fusobacterium nucleatum outer membrane protein RadD is an arginine‐inhibitable adhesin required for inter‐species adherence and the structured architecture of multispecies biofilm

Abstract

SummaryA defining characteristic of the suspected periodontal pathogen Fusobacterium nucleatum is its ability to adhere to a plethora of oral bacteria. This distinguishing feature is suggested to play an important role in oral biofilm formation and pathogenesis, with fusobacteria proposed to serve as central "bridging organisms" in the architecture of the oral biofilm bringing together species which would not interact otherwise. Previous studies indicate that these bacterial interactions are mediated by galactose-or arginine-inhibitable adhesins although genetic evidence for the role and nature of these proposed adhesins remains elusive. To characterize these adhesins at the molecular level, the genetically transformable F. nucleatum strain ATCC 23726 was screened for adherence properties, and arginine inhibitable adhesion was evident, while galactose-inhibitable adhesion was not detected. Six potential arginine binding proteins were isolated from the membrane fraction of F. nucleatum ATCC 23726 and identified via mass spectroscopy as members of the outer membrane family of proteins in F. nucleatum. Inactivation of the genes encoding these six candidates for arginineinhibitable adhesion and two additional homologues revealed that only a mutant derivative carrying an insertion in Fn1526 (now designated as radD) demonstrated significantly decreased coaggregation with representatives of the Gram-positive "early oral colonizers". Lack of the 350 kDa outer membrane protein encoded by radD resulted in the failure to form the extensive structured biofilm observed with the parent strain when grown in the presence of Streptococcus sanguinis ATCC 10556. These findings indicate that radD is responsible for arginine-inhibitable adherence of F. nucleatum and provides definitive molecular evidence that F. nucleatum adhesins play a vital role in inter-species adherence and multispecies biofilm formation.

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

“…E. coli was unable to interact with any of the oral species present in the defined community. Previous study indicated that coaggregation of F. nucleatum with certain oral bacterial species could be inhibited by arginine [24]. Our coaggregation assay confirmed this observation, showing that the presence of 50 mmol·L -1 arginine completely abolished the coaggregation between F. nucleatum and the test organisms (Table 2).…”

Virulence Factors of the Oral Spirochete Treponema denticola

“…T. denticola binds weakly to fusobacteria in an interaction mediated by the carbohydrate moiety of the T. denticola major sheath protein (Msp, see below) and the galactose-binding lectin receptor of the fusobacteria (Kolenbrander et al, 1995;Rosen et al, 2008;Kaplan et al, 2009). T. denticola also binds to the commensal, early-colonizing Streptococcus crista (Yao et al, 1996).…”

Stick to Your Gums

“…A spontaneous F. nucleatum ATCC 10953 mutant was isolated that was defective for co-aggregation with S. cristatus, S. gordonii, and S. sanguinis, but not with P. gingivalis (Edwards et al, 2007). The mutant lacked an approximately 360-kDa protein that was identified as a member of the auto-transporter family, RadD, confirmed as an arginine-inhibitable adhesin responsible for co-aggregation with streptococci (Kaplan et al, 2009). Dual-species biofilm formation with S. sanguinis was impaired in a radD mutant.…”

“…Dual-species biofilm formation with S. sanguinis was impaired in a radD mutant. In addition, RadD was shown to mediate arginine-sensitive interactions with secretory IgA and with human lymphocytes (Edwards et al, 2007;Kaplan et al, 2009).…”