Human oral bacteria interact with their environment by attaching to surfaces and establishing mixed-species communities. As each bacterial cell attaches, it forms a new surface to which other cells can adhere. Adherence and community development are spatiotemporal; such order requires communication. The discovery of soluble signals, such as autoinducer-2, that may be exchanged within multispecies communities to convey information between organisms has emerged as a new research direction. Direct-contact signals, such as adhesins and receptors, that elicit changes in gene expression after cell-cell contact and biofilm growth are also an active research area. Considering that the majority of oral bacteria are organized in dense three-dimensional biofilms on teeth, confocal microscopy and fluorescently labeled probes provide valuable approaches for investigating the architecture of these organized communities in situ. Oral biofilms are readily accessible to microbiologists and are excellent model systems for studies of microbial communication. One attractive model system is a saliva-coated flowcell with oral bacterial biofilms growing on saliva as the sole nutrient source; an intergeneric mutualism is discussed. Several oral bacterial species are amenable to genetic manipulation for molecular characterization of communication both among bacteria and between bacteria and the host. A successful search for genes critical for mixed-species community organization will be accomplished only when it is conducted with mixed-species communities

Twenty-eight strains of Fusobacterium nucleatum and 41 Selenomonas strains, including S. sputigena (24 strains), S. flueggei (10 strains), S. infelix (5 strains), and S. noxia (2 strains), were tested for their ability to coaggregate with each other and with 49 other strains of oral bacteria representing Actinobacillus, Actinomyces, Bacteroides, Capnocytophaga, Gemella, Peptostreptococcus, Porphyromonas, Propionibacterium, Rothia, Streptococcus, and Veillonella species. Selenomonads coaggregated with fusobacteria and with Actinomyces naeslundii PK984 but not with any of the other bacteria, including other selenomonads. In contrast, fusobacteria coaggregated with members of all genera, although not with all strains of each species tested. Each fusobacterium strain appeared to have its own set of partners and coaggregation properties, unlike their partners, whose coaggregation properties in earlier surveys delineated distinct coaggregation groups. Coaggregations of fusobacteria with the 63 gram-negative strains were usually inhibited by EDTA, whereas those with the 27 gram-positive strains were usually not inhibited. Likewise, lactose-inhibitable coaggregations were common among some strains of fusobacteria and some strains from each of the genera containing gramnegative partners but were rarely observed with gram-positive partners. Heating the fusobacteria at 85°C for 30 min completely prevented coaggregation with most partners, suggesting the involvement of a protein on the fusobacteria. Heat treatment of many of the gram-negative partners not only enhanced their coaggregation with the fusobacteria but also changed lactose-sensitive coaggregations to lactose-insensitive coaggregations. Although fusobacteria coaggregated with a broader variety of oral partner strains than any other group of oral bacteria tested to date, each fusobacterium exhibited coaggregation with only a certain set of partner strains, and none of the fusobacteria adhered to other strains of fusobacteria, indicating that recognition of partner cell surfaces is selective. The strains of F. nucleatum are heterogeneous and cannot be clustered into distinct coaggregation groups. Collectively, these results indicate that coaggregation between fusobacteria and many gram-negative partners is signfficantly different from their coaggregation with gram-positive partners. The contrasting variety of partners for fusobacteria and selenomonads supports the concept of coaggregation partner specificity that has been observed with every genus of oral bacteria so far examined. * Corresponding author. whereas others were inhibited by treating their partner. Some pairs were inhibited by 0.02 M EDTA, but others were not. A brief report of the properties of coaggregation of F. nucleatum with Streptococcus sanguis, Streptococcus mitis, Bacteroides melaninogenicus, or Staphylococcus aureus indicated that the ability of fusobacteria to coaggregate was destroyed by heat (90°C for 10 min) or protease, whereas the ability of S. sanguis to coaggregate with fusobacteria was...

The coaggregation of Fusobacterium nucleatum PK1594 and Porphyromonas (Bacteroides) gingivalis PK1924 was inhibited equally well by lactose, N-acetyl-D-galactosamine, and D-galactose, which caused 50% inhibition of coaggregation at 2 mM sugar concentration. Other sugars such as D-galactosamine, D-fucose (6-deoxy-D-galactose), and a-methyland P-methyl-D-galactosides also inhibited coaggregation. Sugar specificity was apparent, since neither L-fucose, L-rhamnose, N-acetyl-D-glucosamine, nor N-acetylneuraminic acid was an inhibitor. Protease treatment of the fusobacterium completely abolished coaggregation, whereas it had no effect on the coaggregating activity of the porphyromonad. Although numerous lactose-inhibitable coaggregating pairs are known to occur among gram-positive bacteria, this report and the accompanying survey (P. E.

A radioactivity-based assay was developed to define the participation of radioactively labeled cell types within the milieu of unlabeled partners in multigeneric aggregates. The cell types in these multigeneric aggregations consisted of various combinations of 21 strains representing five genera of human oral bacteria. The coaggregation properties of each cell type, when paired individually with various strains, were delineated and were unchanged when the microbes took part in the more complex multigeneric aggregations. Competition between homologous labeled and unlabeled cells for binding to a partner cell type was achieved only when the homologous cells were mixed together before the addition of their partner cells. Attempts to displace a labeled cell type from an aggregate by subsequent addition of a large excess of the same unlabeled cell type were unsuccessful, which suggested that the forces that bound different cell types together were very strong and the cell-to-cell interactions were stable. However, a cell type that exhibited only lactose-reversible coaggregations with partners was easily and selectively released by the addition of lactose to multigeneric aggregates otherwise consisting solely of lactose-nonreversible cell-to-cell interactions. This not only indicates the independent nature of individual coaggregations but also suggests the involvement of lectinlike adhesins in these sugar-inhibitable coaggregations. Although the molecular mechanisms responsible for multigeneric aggregations are unknown, the principle of a common partner cell type serving as a bridge between two otherwise noncoaggregating cell types was firmly established by the observation of sequential addition of one cell type to another. Thus, competition, bridging, coaggregate stability, independent nature of interactions, and partner specificity are the key principles of adherence that form the framework for continued studies of multigeneric aggregates. While the human oral cavity is a prime example of a complex microbial community, collectively the community appears to consist of simple and testable individual interactions.Intergeneric coaggregations among pairs of human oral bacteria have been investigated in several laboratories, and the results of these surveys all indicate that interactions between coaggregating partners are not random (7,9,15,18,24,35,38). On the contrary, specific coaggregating pairs were observed, and on the basis of this specificity, coaggregation groups of certain oral streptococci (Streptococcus sanguis and S. morbillorum) and actinomyces (Actinomyces naeslundii and A. viscosus) were delineated (4,(20)(21)(22). Over 100 fresh isolates and stock culture strains of both streptococci and actinomyces have been examined. The coaggregation patterns (a combination of partner specificity, reversibility of coaggregation by simple sugars and chelating agents, and inactivation of partners by prior treatment with heat or protease digestion) of about 95% of each of the two bacterial types are represented by six str...

Of the 122 human oral bacterial strains tested from 11 genera, only streptococci and a few actinomyces exhibited coaggregation among the strains within their respective genera. Eight of the ten streptococci showed multiple intrageneric coaggregations, all of which were inhibited by galactosides. The widespread intrageneric coaggregation among the streptococci and the less extensive coaggregation among the actinomyces offers an explanation for their accretion on cleaned tooth surfaces and their dominance as primary colonizers.

ScaA lipoprotein in Streptococcus gordonii is a member of the LraI family of homologous polypeptides found among streptococci, pneumococci, and enterococci. It is the product of the third gene within the scaCBA operon encoding the components of an ATP-binding cassette (ABC) transporter system. Inactivation ofscaC (ATP-binding protein) or scaA(substrate-binding protein) genes resulted in both impaired growth of cells and >70% inhibition of 54Mn2+ uptake in media containing <0.5 μM Mn2+. In wild-type andscaC mutant cells, production of ScaA was induced at low concentrations of extracellular Mn2+ (<0.5 μM) and by the addition of ≥20 μM Zn2+. Sca permease-mediated uptake of 54Mn2+ was inhibited by Zn2+ but not by Ca2+, Mg2+, Fe2+, or Cu2+. Reduced uptake of54Mn2+ by sca mutants and by wild-type cells in the presence of Zn2+ was abrogated by the uncoupler carbonylcyanide m-chlorophenylhydrazone, suggesting that Mn2+ uptake under these conditions was proton motive force dependent. The frequency of DNA-mediated transformation was reduced >20-fold in sca mutants. The addition of 0.1 mM Mn2+ to the transformation medium restored only partly the transformability of mutant cells, implying an alternate role for Sca proteins in the transformation process. Cells ofsca mutants were unaffected in other binding properties tested and were unaffected in sensitivity to oxidants. The results show that Sca permease is a high-affinity mechanism for the acquisition of Mn2+ and is essential for growth of streptococci under Mn2+-limiting conditions.

Infect. Immun. 61:981-987, 1993). The nucleotide sequence of the 6,125-bp EcoRI insert of pRA1, containing scaA, the gene encoding ScaA, was determined. Six open reading frames (ORFs) were identified. The orientation of four ORFs, two upstream (ORF 1 and ORF 2) and one downstream (ORF 4) ofscaA (ORF 3), indicated transcription in one direction, whereas ORF 5 and ORF 6 were transcribed divergently. Computer analysis of the deduced amino acid sequences identified a consensus binding site for ATP (GxxGxGKS) in the putative 28,054-Da protein encoded by ORF 1. ORF 2 potentially encoded a hydrophobic protein of 29,705 Da with six potential membrane-spanning regions. ScaA was 310 amino acids, 34,787 Da, and contained the lipoprotein consensus sequence LxxC, also reported for the ScaA-related proteins SsaB, FimA, and PsaA from Streptococcus sanguis 12, Streptococcus parasanguis FW213, and Streptococcus pneumoniae R36A, respectively. ORF 4 potentially encoded a 163-amino-acid protein of 17,912 Da, which was nearly identical to the downstream adjacent gene products of ssaB, fimA, and psaA. No significant homology with other proteins was found with the putative ORF 5 gene product, a 229-amino-acid protein of 25,107 Da. ORF 6 was incomplete and encoded a protein larger than 564 amino acids. This putative protein had a consensus Zn2+ binding motif, HExxH, found among bacterial thermolysins and mammalian neutral endopeptidases and was 40% identical to a homologous 210-amino-acid region of human enkephalinase. The genetic organization of ORFs 1, 2, and 3 was similar to those of the bacterial periplasmic-binding protein-dependent transport systems of gram-negative bacteria and binding-lipoprotein-dependent transport systems of gram-positive bacteria, and these genes appeared to encode ABC (ATP-binding cassette) proteins. This report describes a cell-to-cell adherence function associated with an ATP-binding cassette.

A total of 22 strains of Treponema spp. including members of all four named human oral species were tested for coaggregation with 7 strains of oral fusobacteria, 2 strains of nonoral fusobacteria, and 45 strains of other oral bacteria, which included actinobacilli, actinomyces, capnocytophagae, eubacteria, porphyromonads, prevotellae, selenomonads, streptococci, and veillonellae. None of the treponemes coaggregated with any of the latter 45 oral strains or with the two nonoral fusobacteria. All treponemes, eight Treponema denticola strains, eight T. socranskii strains, four oral pectinolytic treponemes, one T. pectinovorum strain, and one T. vincentii strain coaggregated with at least one strain of the fusobacteria tested as partners. The partners consisted of one strain of Fusobacterium periodonticum, five F. nucleatum strains including all four subspecies of F. nucleatum, and a strain of F. simiae obtained from the dental plaque of a monkey. In the more than 100 coaggregations observed, the fusobacterial partner was heat inactivated (85؇C for 30 min), while the treponemes were unaffected by the heat treatment. Furthermore, the fusobacteria were usually inactivated by proteinase K treatment, and the treponemes were not affected. Only the T. denticola coaggregations were inhibited by lactose and D-galactosamine. None were inhibited by any of 23 other different sugars or L-arginine. Intrageneric coaggregations were seen among the subspecies of F. nucleatum and with F. periodonticum, and none were inhibited by any of the sugars tested or by L-arginine. No intrageneric coaggregations were observed among the treponemes. These data indicate that the human oral treponemes show a specificity for oral fusobacteria as coaggregation partners. Such cell-to-cell contact may facilitate efficient metabolic communication and enhance the proliferation of each cell in the progressively more severe stages of periodontal disease.