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. 2015 Oct;94(10):1432–1438. doi: 10.1177/0022034515593706

Cellular Components Mediating Coadherence of Candida albicans and Fusobacterium nucleatum

T Wu 1, L Cen 1, C Kaplan 2, X Zhou 3, R Lux 1, W Shi 1, X He 1,✉
PMCID: PMC4577983  PMID: 26152186

Abstract

Candida albicans is an opportunistic fungal pathogen found as part of the normal oral flora. It can be coisolated with Fusobacterium nucleatum, an opportunistic bacterial pathogen, from oral disease sites, such as those involved in refractory periodontitis and pulp necrosis. The physical coadherence between these 2 clinically important microbes has been well documented and suggested to play a role in facilitating their oral colonization and colocalization and contributing to polymicrobial pathogenesis. Previous studies indicated that the physical interaction between C. albicans and F. nucleatum was mediated by the carbohydrate components on the surface of C. albicans and the protein components on the Fusobaterium cell surface. However, the identities of the components involved still remain elusive. This study was aimed at identifying the genetic determinants involved in coaggregation between the 2 species. By screening a C. albicans SN152 mutant library and a panel of F. nucleatum 23726 outer membrane protein mutants, we identified FLO9, which encodes a putative adhesin-like cell wall mannoprotein of C. albicans and radD, an arginine-inhibitable adhesin-encoding gene in F. nucleatum that is involved in interspecies coadherence. Consistent with these findings, we demonstrated that the strong coaggregation between wild-type F. nucleatum 23726 and C. albicans SN152 in an in vitro assay could be greatly inhibited by arginine and mannose. Our study also suggested a complex multifaceted mechanism underlying physical interaction between C. albicans and F. nucleatum and for the first time revealed the identity of major genetic components involved in mediating the coaggregation. These observations provide useful knowledge for developing new targeted treatments for disrupting interactions between these 2 clinically relevant pathogens.

Keywords: coaggregation, interspecies interaction, oral microbiota, opportunistic pathogen, adhesin, mannoprotein

Introduction

Candida albicans is a commensal fungal species commonly colonizing human mucosal surfaces, which can be isolated from the oral cavity of as many as 60% of healthy adult subjects (Odds et al. 1988; Glick and Siegel 1999). It coexists with diverse oral microbial species and has long been adapted to the human host (Morales and Hogan 2010). In healthy individuals, C. albicans is usually a minor component of their oral microbial flora. However, under conditions of immune dysfunction or local predisposing factors such as poor oral hygiene or ill-fitted dentures, colonizing C. albicans strains can become opportunistic pathogens. In these patients, C. albicans becomes more predominant and invasive, causing recurrent mucosal infection, such as denture stomatitis or life-threatening disseminated infections (Boerlin et al. 1995; Coleman et al. 1997; Perlroth et al. 2007; Azie et al. 2012).

The colonization of C. albicans in the human oral cavity can be achieved through its adherence to a variety of salivary pellicle components, including proline-rich proteins and statherin (Calderone and Braun 1991). Moreover, it is able to directly bind to the oral epithelial cells through adhesion/ligand interactions (Salgado et al. 2011). In addition to oral tissue surfaces, increasing evidence suggests that C. albicans could establish themselves by incorporation into existing biofilms through adherence to a variety of oral bacterial species including streptococci, actinomyces, and fusobacteria (Shirtliff et al. 2009).

Fusobacterium nucleatum is another opportunistic oral pathogen that has been implicated in periodontal disease (Signat et al. 2011). Best known for its ability to adhere to a diverse range of oral bacteria, it is considered a “bridging” organism that links early commensal colonizers and late colonizing species, many of which are periodontal pathogens; therefore, it plays an important role in the succession of genera in oral polymicrobial communities (Kolenbrander 2000; Rickard et al. 2003). Other than interacting with bacterial species, fusobacteria can establish direct physical association with eukaryotic microbes such as fungi. The coadherence between F. nucleatum and C. albicans has been well documented and has been implicated in facilitating the colonization of C. albicans within the oral cavity (Grimaudo and Nesbitt 1997; Jabra-Rizk et al. 1999). Likewise, the coisolation of C. albicans with F. nucleatum from a variety of oral disease sites—including refractory periodontitis, such as therapy-resistant apical periodontitis (Waltimo et al. 1997) and pulp necrosis (Lana et al. 2001)—further suggested their interaction as well as involvement in polymicrobial pathogenesis.

Previous studies suggested that the coadherence between C. albicans and F. nucleatum was mediated by the carbohydrate components on the surface of C. albicans and the protein components on the Fusobacterium cell surface (Grimaudo and Nesbitt 1997; Jabra-Rizk et al. 1999). However, the identities of the components involved remain elusive. In the current study, by screening a C. albicans mutant library and a panel of F. nucleatum outer membrane protein mutants, we aimed to identify the genetic components involved in coadherence between these 2 clinically relevant opportunistic pathogens.

Materials and Methods

Strains and Growth Conditions

Wild-type strain F. nucleatum 23726 and its mutant derivatives lacking one of the large fusobacterial outer membrane proteins—including Fn0254, Fn0387, Fn1449 (Fap2), Fn1526 (RadD), Fn1554, Fn1893, Fn2047, or Fn2058 (Aim1; Kaplan et al. 2009; Kaplan et al. 2010; Gur et al. 2015)—were cultivated in Columbia broth (Difco, Detroit, MI, USA) at 37 °C anaerobically (nitrogen 90%, carbon dioxide 5%, hydrogen 5%).

A C. albicans SN152 mutant library was purchased from the University of California, San Francisco (Noble et al. 2010). The library contains 674 homozygous deletion mutants covering roughly 11% of the C. albicans genome. C. albicans wild-type and mutants were precultured in yeast peptone dextrose (YPD) medium (yeast extract, 10 g/L; peptone, 20 g/L; glucose, 20 g/L) at 30 °C in an atmosphere of 5% CO2 in air. Hyphal growth of C. albicans was induced by inoculating overnight cultures in fresh YPD medium supplemented with 20% fetal bovine serum and aerobically incubated at 37 °C until the OD600 reached 0.2.

Coaggregation Assay

Coaggregation assays were preformed in modified coaggregation buffer (CAB) as previously described (He et al. 2012). Since hyphae-displaying C. albicans SN152 cells tend to precipitate even in the absence of coaggregating bacterial partner species, direct visual microscopic observation of coaggregation between C. albicans and F. nucleatum cells was performed. Briefly, exponential bacterial cells and hypha-displaying candida cells were collected and resuspended in CAB to an OD600 of 1 and 2, respectively. Equal volumes (250 µL) of bacterial and candida cells were mixed in reaction tubes. The tubes were vigorously vortexed for 30 s and allowed to stand at room temperature for 30 min. Coaggregating samples were taken and observed using light microscopy; images were taken from 10 random fields of view. For each coaggregation sample, 10 random candida hypha-displaying cells were chosen, and the attached fusobacterial cells were counted for each cell. Based on the average number (n) of attached bacterial cells per 20 µm of hyphal body length, coaggregation was graded on a 0 to 4 scale: score 4, n ≥ 20; score 3, 10 ≤ n < 20; score 2, 5 ≤ n < 10; score 1, 1 ≤ n < 5; score 0, when no attached bacterial cells could be observed.

Fluorescence-based Coaggregation Assay

To achieve a more quantitative and sensitive readout, a fluorescence-based coaggregation assay was developed (see Appendix for detailed experimental procedure). Briefly, Syto 9 green fluorescent nucleic acid dye–stained fusobacterial cells were used to set up coaggregation assays with Candida. Coaggregation mixtures were then washed to remove unattached fusobacterial cells. The coaggregation between Candida and Fusobacterium was quantified by measuring the intensity of the florescence signal retained on the Candida cell surface.

Screening a C. albicans SN152 Library for Mutants Defective in Coaggregating with F. nucleatum 23726

Two microliters of each mutant from a C. albicans SN152 mutant library stored in 96-well plates was transferred to plates containing 200 µL of fresh YPD medium in each well. Plates were incubated at 30 °C in an atmosphere of 5% CO2 in air. After overnight growth, 60 mutant strains showing growth defects (with cell density <0.1 at OD600) were excluded while the rest of the mutants were spun down and resuspended in fresh YPD medium supplemented with 20% fetal bovine serum. The plates were next further incubated aerobically at 37 °C for 4 h to induce hyphal growth. C. albicans cells were resuspended in CAB to a final OD600 of 0.5 to 1. Meanwhile, exponential-phase F. nucleatum cells were resuspended in CAB to a final OD600 of 1. One hundred microliters of bacterial cells were then added into each well of 96-well plates containing C. albicans mutants and mixed by pipetting. Plates were incubated at room temperature for 30 min before samples were taken and aggregation was observed under a light microscope. Mutants defective in coaggregating with wild-type F. nucleatum were confirmed by fluorescence-based coaggregation assays.

Inhibition of Coaggregation between F. nucleatum and C. albicans

By using the coaggregation assay described above, the following sugars or amino acids at a final concentration of 100 mM (Grimaudo and Nesbitt 1997) were tested for their ability to inhibit coaggregation between C. albicans and F. nucleatum: D-galactose, D-lactose, D-mannose, L-leucine, N-acetyl-glucosamine, and L-arginine.

Recruitment Assay

Recruitment assays were designed and performed to test the ability of coaggregation-defective C. albicans and F. nucleatum mutants identified in this study to bind to monospecies biofilms of their wild-type interacting partner. Detailed procedures are provided in the Appendix. Briefly, planktonic cells of C. albicans (wild type and mutants) or F. nucleatum (wild type and mutants) were added to the monospecies biofilm of their wild-type interacting partner, and their ability to bind to the biofilm cells was determined by cell counting.

Results

C. albicans SN152 Mutant FLO9 Exhibits Reduced Ability in Coaggregating with F. nucleatum

The coaggregating ability between C. albicans SN152 and F. nucleatum 23726 was tested using cell suspensions in CAB. Coaggregation was monitored and recorded by light microscopy and fluorescence-based assays. Wild-type C. albicans SN152 hyphal cells and F. nucleatum 23726 displayed a strong coaggregation phenotype (Fig. 1, panel 2), while no aggregation was detected between the yeast form of C. albicans and F. nucleatum under the same coaggregating conditions tested (data not shown). To identify the genetic components of C. albicans involved in its coaggregation with F. nucleatum, a homozygous C. albicans SN152 gene deletion library (Noble et al. 2010) was screened for mutants that had normal growth but were defective in coadherence with F. nucleatum. The library contains 674 homozygous deletion mutants covering roughly 11% of the C. albicans genome. The screen led to the discovery of 1 mutant, C. albicans SN152 FLO9, which displayed the most drastically reduced ability in coaggregating with F. nucleatum. As revealed by microscopic observation, the surface of the FLO9 mutant was only sparsely decorated with F. nucleatum (Fig. 1, panel 7), as compared with wild-type C. albicans, whose hyphal surface was extensively covered by the coaggregating bacterial partner. Fluorescence-based coaggregation assay showed that, when compared with the wild type, the fluorescence signal retained decreased >60%, suggesting a severe reduction in the number of attached F. nucleatum cells (Fig. 2). FLO9 encodes a putative adhesin-like cell wall mannoprotein (Noble et al. 2010), and our data suggested its involvement in mediating physical interaction with F. nucleatum.

Figure 1.

Figure 1.

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Microscopic observation of coaggregation between Candida albicans (Ca) and Fusobacterium nucleatum (Fn) under different treatments. For each interacting pair, the pictures of 10 random Ca cells were taken and 1 representative image shown. The scale bar is 10 µm. wt, wild type.

Figure 2.

Figure 2.

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Coaggregation between Candida albicans (Ca) and Fusobacterium nucleatum (Fn). Fluorescence-based coaggregation assay (as described in Materials and Methods) was performed for Fn and Ca, both wild type and mutants. The result was represented as the percentage of fluorescence signal retained compared with the control group containing wild-type Ca and Fn as a coaggregating pair. Average values ± SD are shown. Asterisk indicates the significant difference between the testing group and the control group (wild-type Fn/Ca; P < 0.05, t test). Note: The absolute number of relative fluorescence units represented by 100% was 2,500, corresponding to about 2 × 106 colony-forming units per milliliter of Fn based on the experimental procedure described in the Materials and Methods.

F. nucleatum RadD Mutant Is Defective in Coaggregating with C. albicans SN152

A previous study indicated the involvement of the protein components on the Fusobaterium cell surface in its coaggregation with C. albicans (Grimaudo and Nesbitt 1997). In this study, a panel of F. nucleatum outer membrane protein mutants generated in our laboratory (Kaplan et al. 2009; Kaplan et al. 2010) was tested for their coaggregating ability with C. albicans SN152. The results showed that among the 8 mutants tested, RadD was the only mutant defective in binding to the hypha-displaying C. albicans cells (Fig. 1, panel 6), with a >60% reduction in fluorescence signal recovered from the C. albicans–bound fusobacterial cells as compared with the wild type (Fig. 2). RadD encodes an arginine-inhibitable adhesin that has been shown to be required for interspecies coadherence between fusobacteria and oral streptococci (Edwards et al. 2007; Kaplan et al. 2009). Our data suggested that it also plays a role in mediating interaction with C. albicans.

We further tested the coaggregation phenotype between F. nucleatum RadD and C. albicans FLO9. Microscopic observation and fluorescence-based assay did not reveal any significant difference in coaggregation between the interacting pairs with either one or both mutant partners. In addition, there was visible residual coaggregation between fusobacterial and Candida cells when RadD and FLO9 mutants were used as interacting pair (Fig. 1, panel 9).

Mannose- and Arginine-inhibitable Coaggregation between C. albicans and F. nucleatum

Mutant screening revealed that the mannoprotein FLO9 from C. albicans and arginine-inhibitable RadD from F. nucleatum are involved in the interspecies coaggregation. These findings suggested that the observed strong coadherence phenotype between C. albicans and F. nucleatum might be mannose/arginine inhibitable. To confirm this, mannose and arginine, among other carbohydrates and amino acids, were tested for their ability in inhibiting the coadherence. The results showed that the presence of 0.1M mannose or arginine resulted in a drastic reduction in the coaggregation between the Candida and Fusobacterium pair, while other carbohydrates or amino acids tested did not have a significant effect on the coaggregation (Table). This was revealed by microscopic observation as well as the more quantitative fluorescence-based coaggregation assay showing about 70% reduction in the signal retention vs. the control (Fig. 1, panels 3, 5; Fig. 2), indicating the reduced number of F. nucleatum cells attached to Candida hyphal cells. Moreover, the addition of other monosaccharides (e.g., D-galactose) and amino acids (including L-leucine) had no apparent inhibitory effects on the coaggregation (Fig. 1, panel 4; Table).

Table.

Effect of Treatments on Coaggregation between Candida albicans and Fusobacterium nucleatum.

Coaggregating Pair No Treatment D-lactose D-galactose D-mannose L-leucine L-arginine N-acetyl- glucosamine
Ca wt / Fn wt 4 4 4 2 4 2 4
Ca FLO9 / Fn wt 2 2 2 2 2 2 2
Ca wt / Fn RadD 2 2 2 2 2 2 2
Ca FLO9 / Fn  RadD 2 2 2 2 2 2 2
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For each coaggregation sample, 10 random hyphae-displaying C. albicans (Ca) cells were chosen; the attached F. nucleatum (Fn) cells were counted. Based on the average number (n) of attached bacterial cells per 20 µm of hyphal body length, coaggregation was graded on a 0 to 4 scale: score 4, n ≥ 20; score 3, 10 ≤ n < 20; score 2, 5 ≤ n < 10; score 1, 1 ≤ n < 5; score 0, no attached bacterial cells could be found.

C. albicans SN152 FLO9 and F. nucleatum RadD Displayed Reduced Ability to Adhere to the Surface Attached Biofilm Cells of Their Respective Interacting Partners

Our data showed that C. albicans and F. nucleatum were able to coaggregate under planktonic conditions, while mutation of FLO9 in C. albicans and radD in F. nucleatum rendered the mutants defective in interspecies binding. To further investigate the observed FLO9- and RadD-dependent coadherence property between C. albicans and F. nucleatum under biofilm conditions, C. albicans SN152 FLO9 and F. nucleatum RadD mutants were tested for their ability to adhere to monobiofilms of wild-type F. nucleatum and C. albicans, respectively. The results showed that, compared with its wild-type counterparts, F. nucleatum RadD and C. albicans SN152 FLO9 displayed significantly reduced capabilities (with about 50% and 70% reductions, respectively) to adhere to monospecies biofilms of their respective wild-type interacting partners (Fig. 3).

Figure 3.

Figure 3.

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Binding of Candida albicans (Ca) and Fusobacterium nucleatum (Fn) to monospecies biofilm of their interacting partner species. Recruitment assay (as described in Materials and Methods) was performed to test the ability of Fn RadD and Ca FLO9 mutant to adhere to monospecies biofilm of wild type (WT) Ca (A) and Fn (B), respectively. The result was represented as the percentage of mutant cells adhered to biofilm of its coaggregating partner compared with the WT control. Average values ± SD are shown. Asterisk indicates significant difference between the 2 sample sets (P < 0.05, t test). Note: The absolute number of cells represented by 100% was about 4 × 107 colony-forming units per milliliter for Fn (A) and 2 × 106 colony-forming units per milliliter for Ca (B) based on the experimental procedure described in Materials and Methods.

Discussion

Coadhesion between different microbial species is a common feature of the oral microbiome. It plays a crucial role in colonization of the oral cavity, contributes to the formation of highly structured multispecies communities, and ultimately affects polymicrobial pathogenesis (Kolenbrander 1988, 1989; Kolenbrander et al. 1999; Kolenbrander et al. 2006; Peters et al. 2012). Such physical interspecies association not only exists among bacterial species but has also been observed for 2 clinically relevant opportunistic pathogens of different kingdoms, C. albicans and F. nucleatum (Grimaudo and Nesbitt 1997; Jabra-Rizk et al. 1999). Knowledge of the major genetic determinants involved in coadherence between these 2 microbes may help shed light on Candida succession and possible polymicrobial disease processes involving these 2 species.

Microscopic observations and fluorescence-based quantitative coaggregation assays clearly demonstrated the strong coadherence phenotype between the bacterial species F. nucleatum 23726 and the eukaryotic microbe C. albicans SN152 in the hyphal form (Fig. 1, panel 2; Fig. 2). Screening of a C. albicans SN152 mutant library resulted in the identification of the FLO9 gene, which is critical in establishing physical interaction with F. nucleatum. FLO9 encodes a putative adhesin-like cell wall mannoprotein in C. albicans (Boisrame et al. 2011). Mutation in this gene does not affect its colony morphology or vegetative growth, nor does it affect competitive fitness in a mouse intravenous infection model (Noble et al. 2010). However, this mutant displayed a drastically reduced ability in coadherence with F. nucleatum as revealed by coaggregation assays, suggesting its role in mediating the observed duo-species interaction (Fig. 1, panel 7; Fig. 2). The identification of the mannoprotein FLO9 as the surface component mediating the coadherence of C. albicans with F. nucleatum was further confirmed by our observation that the coaggregation phenotype was mannose inhibitable (Fig. 1, panel 3). This is consistent with previous reports showing that the binding of F. nucleatum strains to Candida species, including C. albicans and C. dubliniensis, could be inhibited by addition of mannose but not other carbohydrates tested (Grimaudo and Nesbitt 1997; Jabra-Rizk et al. 1999).

C. albicans has a multilayered cell wall composed of an outer layer of proteins glycosylated with N- or O-linked mannosyl residues and an inner skeletal layer of beta-glucans and chitin (Gow and Hube 2012). The mannoproteins have been shown to play important roles in diverse cellular functions, such as acting as specific immunodeterminants (Gow and Hube 2012), binding to host-secreted macromolecules (Tronchin et al. 1988), as well as functioning as adhesins to mediate the direct Candida-bacteria and Candida–host cell recognitions (Salgado et al. 2011; Dutton et al. 2014). In C. albicans, mnt1 and mnt2 encode partially redundant α-1, 2-mannosyltransferases that catalyze the addition of the second and third mannose residues in an O-linked mannose pentamer, while och1 encodes a α-1, 6-mannosyltransferase that is involved in N-glycan outer chain branch addition. In a recent study, Dutton et al. (2014) demonstrated that O-mannosylation via Mnt, specifically Mnt1, is critical for hyphal adhesion functions required for coadherence between C. albicans and Streptococcus gordonii. In our study, a mnt1 mutant displayed a normal coaggregation phenotype with F. nucleatum, while the och1 mutant showed drastically reduced coaggregation with F. nucleatum (Appendix Fig. 1) even though this mutant was not initially identified via library screening due to its growth defects. Our data suggested that N-linked glycosylation could be more important for FLO9 in determining its binding ability with its counterpart in F. nucleatum. Furthermore, FLO9 was predicted to be hyphally regulated and expressed during hyphal growth (http://www.candidagenome.org). This is in agreement with our data showing that the coadherence can be observed only between F. nucleatum and C. albicans hyphal, but not yeast cells.

Our study further revealed that RadD, an outer member protein in F. nucleatum 23726, is also involved in mediating the coadherence between fusobacterial and candidal cells. RadD is an arginine-binding protein (Edwards et al. 2007) and has been shown to be an adhesin required for the coadherence of F. nucleatum with gram-positive oral bacterial species, such as streptococci (Kaplan et al. 2009). More important, our data showed that no additive effects in the reduction of coaggregation can be observed when F. nucleatum RadD and C. albicans FLO9 were tested as a pair compared with interacting pairs containing either mutant (Fig. 1, panels 6 to 9). These data strongly suggested that RadD is acting as the counterpart component for FLO9 in C. albicans and that these 2 components could directly interact with each other and mediate coadherence. Consistent with this finding, exogenous arginine was able to achieve a similar inhibitory effect on the coaggregation between F. nucleatum and C. albicans compared with mannose (Fig. 1, panels 3 and 5; Table). Previous studies also suggested that other than a mannan-containing receptor on the C. albicans, the coadherence between F. nucleatum and C. albicans involves a heat-liable component on F. nucleatum (Grimaudo and Nesbitt 1997). Meanwhile, sequence analysis of RadD revealed a domain close to the C-terminus (1513-1707aa) with homology to a sequence encoding a bacteriophage tail fiber. Previous studies showed that the adsorption of bacteriophage can be achieved through the binding of tail fibers to the polymannose O-antigens on the Escherichia coli cell surface (Heller and Braun 1982). These observations implied that RadD potentially has binding sites for both mannose and arginine. It could act as a receptor and bind to the specific mannan residue on the FLO9 mannoprotein on the surface of C. albicans to mediate physical interaction. Meanwhile, the binding of arginine might induce a conformational change in the binding pocket for mannose and result in the disruption of the coadherence between the 2 species. Further studies are needed to determine the respective binding sites in RadD for mannose and arginine.

It is worthwhile to point out that, in a study by Jabra-Rizk et al. (1999), the coaggregation between C. albicans and F. nucleatum was mannose inhibitable, while arginine had no such effects. In that study, F. nucleatum ATCC 49256 was tested—a F. nucleatum subsp. vincentii strain instead of a F. nucleatum subsp. nucleatum strain. Furthermore, the C. albicans strains used by Jabra-Rizk et al. were all clinical isolates, and they displayed different phenotypes when coaggregating with F. nucleatum ATCC 49256. Although FLO9 is conserved among many Candida species and C. albicans subspecies, RadD homologues have been found in different Fusobacterium species and subspecies, including F. nucleatum subsp. vincentii. It remains to be determined if these homologues could display differential adhesion/ligand specificities. Our recent study showed that different F. nucleatum subspecies display significantly differential coaggregation abilities with the same Streptococcus mutans strain (unpublished data), suggesting the involvement of different adhesins or the differential binding specificity of homologous adhesins in different subspecies when interacting with their partner species. Thus, it is likely that the observed difference in arginine inhibitability of the coaggregation could be due to the different strains used. The observed strain specificity in interspecies coadhesion could have an impact on host susceptibility to disease and/or the effectiveness of coadherence preventive measures.

In addition, residual coaggregation was observed even when F. nucleatum RadD and C. albicans FLO9 were used as interacting pairs (Fig. 1, panel 9), which suggested the presence of other receptor/ligand pairs playing minor roles in mediating the observed interspecies coadherence. This is consistent with the multimodal nature of adhesion of C. albicans in its coadherence with other oral species, such as S. gordonii (Holmes et al. 1996; Silverman et al. 2010) as well as host cells (Calderone and Braun 1991).

We also demonstrated that compared with their wild-type counterparts, both F. nucleatum RadD and C. albicans FLO9 were defective in coadhering to the monospecies biofilms of wild-type C. albicans and F. nucleatum, respectively (Fig. 3; Appendix Fig. 2). Oral colonization is a crucial step in bacterial/fungal infections, which can be achieved via directly binding to host tissue surfaces or coadhering to the microbes that have already been established within surface-attached biofilm. Increasing evidence suggests that the coadherence between Candida and bacterial species could promote the formation of mix-species biofilms and affect polymicrobial pathogenicity (Adam et al. 2002; Fox et al. 2014; Xu et al. 2014). Thus, intergenic interaction between C. albicans and F. nucleatum, 2 clinically relevant oral species, not only contributes to their succession in the oral cavity but could also ultimately affect their pathogenicity during polymicrobial infections and so warrants further investigation. In addition, further characterization of the interacting cellular components could help develop new targeted treatments to interfere with the coadherence, a process that could play an important role in polymicrobial pathogenesis involving these 2 microbes.

Author Contributions

T. Wu, contributed to design, data acquisition, analysis, and interpretation, drafted the manuscript; L. Cen, contributed to data acquisition and interpretation, critically revised the manuscript; C. Kaplan, contributed to data acquisition, critically revised the manuscript; X. Zhou, contributed to data analysis and interpretation, critically revised the manuscript; R. Lux, contributed to design, data analysis, and interpretation, critically revised the manuscript; W. Shi, X. He, contributed to conception, design, data analysis, and interpretation, critically revised the manuscript. All authors gave final approval and agree to be accountable for all aspects of the work.

Supplementary Material

Supplementary material

Footnotes

A supplemental appendix to this article is published electronically only at http://jdr.sagepub.com/supplemental.

This work was supported by grants from the National Institutes of Health (NIH-1-R01-DE020102 and NIH-1-R01-DE023810-01), a grant from the Natural Sciences Foundation of China (30672322), and a grant from the International Science and Technology Cooperation Program of China (2011DFA30940).

The authors declare that W.S. and C.K. are employees of C3 Jian, Inc., which has licensed technologies from UC Regents that could be indirectly related to this research project. The authors declare no other potential conflicts of interest with respect to the authorship and/or publication of this article.

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