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J Dent Res 86(2):175-180, 2007
© 2007 International and American Associations for Dental Research

RESEARCH REPORT
Biological

Human Cytomegalovirus Enhances A. actinomycetemcomitans Adherence to Cells

W. Teughels1,*, I. Sliepen1, M. Quirynen1, S. Kinder Haake2, J. Van Eldere3, P. Fives-Taylor4, and M. Van Ranst5

1 Catholic University Leuven, Research Group for Microbial Adhesion, Department of Periodontology, Kapucijnenvoer 7, 3000 Leuven, Belgium;
2 UCLA, School of Dentistry, 10833 Le Conte Avenue, Los Angeles, CA, USA;
3 Catholic University Leuven, Centre for Molecular Diagnostics, Herestraat 49, 3000 Leuven, Belgium;
4 University of Vermont, Department of Microbiology and Molecular Genetics, Stafford Hall, 95 Carrigan Drive, Burlington, VT, USA; and
5 Catholic University Leuven, Laboratory of Clinical Virology, Rega Institute for Medical Research, Minderbroedersstraat 10, 3000 Leuven, Belgium

* corresponding author, Wim.Teughels{at}med.kuleuven.be


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS & METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Adherence of Actinobacillus actinomycetemcomitans to epithelial cells is an important step in periodontal disease pathogenesis. Recent publications describe the subgingival presence of a wide array of viruses [e.g., human cytomegalo-virus (hCMV)]. Since viruses can increase cellular susceptibility for bacterial adherence, we investigated whether hCMV renders epithelial cells more prone to adherence by Actinobacillus actinomycetemcomitans. Cultivated HeLa and primary epithelial cells were shown to be semi-permissive for hCMV infection, which resulted in increased bacterial adherence. This increase correlated with viral concentrations, was evident in all Actinobacillus actinomycetemcomitans strains examined, and increased during the first 24 hrs, followed by a slight decrease. Immediate early antigen expression was not correlated with the increased adherence of Actinobacillus actinomycetemcomitans. The results confirmed our hypothesis that the adherence of Actinobacillus actinomycetemcomitans is influenced by hCMV in vitro.

KEY WORDS: Actinobacillus actinomycetemcomitans • human cytomegalovirus • adherence • epithelial cells • polymicrobial


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS & METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Many diseases, such as periodontitis, originate from polymicrobial infections, involving more than one etiological agent. Several primary periodontopathogens are known (e.g., Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, and Tannerella forsythensis) (American Academy of Periodontology, 1996), and their virulence factors have been characterized (Kuramitsu, 2003). Emergence of these endogenous pathogens, rather than super-infection with exogenous species, induces the clinical changes associated with periodontitis (Rudney et al., 2005). An understanding of the conditions that support the emergence of pathogens is critical to an understanding of the disease process and the development of new treatment strategies.

Viral-bacterial co-infections have been documented, but mechanisms underlying synergistic pathogenesis are only partially understood (Brogden and Guthmiller, 2002). Viral infections can be a pre-requisite for or can aggravate subsequent bacterial infections, especially in respiratory disease, otitis media, or gastroenteritis. Although herpes virus-bacterial co-infections have been described in periodontitis, the mechanisms of interaction remain hypothetical (Slots, 2005). Adherence is required for the establishment of bacterial populations on mucosal surfaces, an important first step in infection (Reed and Williams, 1978), and an increased susceptibility to bacterial adherence may support the emergence of pathogens. Viruses can play an important role in this process by pre-conditioning cells for bacterial infections (Selinger et al., 1981). For example, most deaths during influenza virus epidemics occur in elderly people, often a result of secondary bacterial pneumonia (Sethi, 2002). Bacterial super-infection is partially attributed to increased cellular susceptibility for bacterial adherence induced by influenza viruses (Hakansson et al., 1994). Human cytomegalovirus (hCMV), together with Actinobacillus actinomycetemcomitans, appears to constitute an important pathogenic feature of periodontitis (Slots, 2005). hCMV infections enhance the adhesiveness of gastrointestinal pathogens to cells (Holberg-Petersen et al., 1994), and a similar mechanism might facilitate Actinobacillus actinomycetemcomitans adherence in the oral cavity. The present study examined the hypothesis that hCMV infection of epithelial cells leads to increased Actinobacillus actinomycetemcomitans adherence in vitro.


   MATERIALS & METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS & METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Bacterial Growth
Actinobacillus actinomycetemcomitans strains ATCC 33384, IDH1705, and highly leukotoxic JP2 were grown on blood-agar (Blood Agar Base II; Oxoid, Basingstoke, England), supplemented with hemin (5 mg/mL), menadione (1 mg/mL), and 5% sterile horse blood. All 3 strains adhere to and invade epithelial cells (Meyer et al., 1991; Eick and Pfister, 2004; and unpublished data).

Viral Growth
HCMV strain AD169 (ATCC VR538) was propagated in E6SM fibroblasts (Rega Institute, Leuven, Belgium). When extensive cytopathogenic effects became apparent, the culture was harvested and filtered (0.22 µm pore) for removal of cellular remnants. The viral load was determined by quantitative real-time PCR (Beuselinck et al., 2005). Mock preparations were produced by heat inactivation (70°C for 1 hr) of the filtered hCMV solution.

Cell Cultures
Primary epithelial cells from pocket epithelium were obtained from diseased sites in chronic periodontitis patients receiving periodontal surgery as a part of their treatment at the Leuven University Hospital. Informed consents were obtained from the patients. The protocol was approved by the ethical committee of the Catholic University of Leuven. Tissue samples were minced and digested in pronase (Protease XIV, Sigma Chemical Co., St. Louis, MO, USA), for isolation of the epithelium, and used for culturing epithelial monolayers (Quirynen et al., 2001). The disassociated epithelial cells were grown in keratinocyte-serum-free medium (Life Technologies Ltd., Paisley, Scotland). When pre-confluent monolayers were observed, cells were seeded in 24-well tissue culture plates (Iwaki microplate, Scitech, Diu, Japan).

HeLa cells and hCMV permissive human embryonic lung fibroblasts (HEL) (Schols et al., 1989) were cultured at 37°C in a humid 5% CO2 environment. Near-confluent monolayers were passaged to 24-well tissue culture plates as described previously (Teughels et al., 2005).

For immunofluorescence assays, the cells were grown on round glass coverslips.

Immunofluorescence Microscopy
Cells were fixed with ice-cold absolute methanol for 10 min and blocked with 10% normal goat serum for 30 min. To detect hCMV immediate early antigens (IE) or early antigens (E), we incubated cells with primary mouse antibodies (1:80) recognizing IE (clone E13) or E (clone CCH2) (Argene, Varilhes, France) for 30 min. Cells were rinsed, reacted with fluorescein-conjugated goat anti-mouse IgG antibodies (Dako, Heverlee, Belgium) at 1:200 dilution for 30 min, rinsed again, and mounted. Images were made by confocal microscopy.

Adherence Assay
We used a two-stage assay system, consisting of viral or mock infection of confluent monolayers, followed by a standard bacterial adherence assay. Monolayers were washed 3 times, then treated for 1 hr with viral or mock (heat-inactivated virus) solutions, which were subsequently replaced by new cell medium. Control cells were treated in parallel with sterile cell medium. Two-hour bacterial adherence experiments were performed (Teughels et al., 2005). After lysis of the epithelial cells, serial dilutions were plated on blood agar and incubated for bacterial cultivation. We calculated the total number of colony-forming units (CFU)/well to determine the cell-associated bacterial load.

When the influence of pre-infection time was assessed, spent cell medium was removed before viral infection and put back into the same wells after the one-hour application of the viral solution. This was done to overrule possible differences due to different contact times between fresh cell medium and epithelial cells. The cells infected for 6, 12, and 24 hrs were re-inoculated with the spent medium, and the timing of pre-infection was conducted so that all cells were ready for the adherence assay at the same time.

Statistical Analysis
In experiments with primary cells, the cell lines from different individuals were used as replicates (statistical unit). For Hela and HEL cell experiments, each newly grown cell line was used as a statistical unit. The data were set up and analyzed as a randomized block design. In all cases, we performed residual analysis to check the assumptions of normality of the error terms. The level of significance was set at p < 0.05.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS & METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
hCMV Infectivity of Epithelial Cells
To test the permissiveness of epithelial cells, we infected HeLa cell monolayers with hCMV at a multiplicity of infection (MOI) of 100 viral genomes per cell. In these cells, IE antigens became apparent after 24 to 48 hrs (Fig. 1Go). The number of IE-positive cells increased until 6 days after infection, at which time 2% of the cells expressed IE antigens. No cytopathogenic effects or E antigens were detected. Infection of primary cells from 3 different chronic periodontitis patients showed the same results as HeLa cells. IE and E antigens were never detected in uninfected and mock-infected controls, including primary cells from 12 chronic periodontitis patients.


Figure 1
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  		Figure 1. hCMV antigen expression in epithelial cells. HeLa (A) and primary cells (B) were infected with hCMV for 48 hrs. Cells were fixed with ice-cold absolute methanol and blocked with 10% normal goat serum. To detect hCMV IE antigens, we incubated cells with primary mouse antibodies (1:80) recognizing IE (clone E13) (Argene, Varilhes, France), and reacted them with fluorescein-conjugated goat anti-mouse IgG antibodies (Dako, Heverlee, Belgium) at 1:200 dilution. Images were made with an Olympus IX70 confocal microscope. (1) Phase-contrast image, (2) fluorescent image, (3) overlap image showing bright IE-positive nuclei for some epithelial cells.

 
Viral Load and Bacterial Adherence
The adherence of Actinobacillus actinomycetemcomitans to HeLa cells pre-infected with various hCMV concentrations for 48 hrs was evaluated. The 48-hour pre-infection time ensured IE antigen expression. hCMV pre-infection significantly increased Actinobacillus actinomycetemcomitans adherence for all strains (Fig. 2Go). Pre-infection at an MOI of 200 resulted in relative increases of 117%, 188%, and 319% for strains JP2, ATCC33384, and IDH1705, respectively, compared with control cells not infected with hCMV. The magnitude of adherence enhancement was bacterial-strain-dependent at an MOI of 200. There was a clear relationship between the viral pre-infection concentration and the subsequent increase in bacterial adherence. A MOI of 200 resulted in the most significant adherence enhancement, but subsequent experiments were conducted with an MOI of 100, due to limitations in the yield of hCMV from cultured cells.


Figure 2
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  		Figure 2. Effect of hCMV concentration on adherence of Actinobacillus actinomycetemcomitans strains to HeLa cells. Cells were exposed to hCMV for 1 hr (MOI range: 10–200) and incubated for an additional 48 hrs. Uninfected cells served as controls. Data are expressed as the proportion of cell-associated bacteria relative to the numbers of bacteria added to the cells initially. Each datapoint represents the mean ± SEM for 4 independent experiments. *P < 0.05, hCMV-infected group vs. uninfected controls. #P < 0.05, Actinobacillus actinomycetemcomitans strain IDH1705 vs. strains JP2 and ATCC33384 at MOI 200.

 
Viral Pre-infection Time and Bacterial Adherence
HeLa cell monolayers were pre-infected with hCMV 6, 12, 24, and 48 hrs prior to Actinobacillus actinomycetemcomitans infection. Mock infections with heat-inactivated virus were carried out in parallel for 48 hrs. Uninfected cells served as controls (0 hr pre-infection). Actinobacillus actinomycetemcomitans adherence significantly increased over the first 24 hrs of hCMV infection (Fig. 3AGo), with the adherence of strains JP2, ATCC33384, and IDH1705 increased by 103%, 156%, and 102% compared with controls. Forty-eight-hour pre-infections resulted in significantly decreased adherence for all strains when compared with 24-hour hCMV-infected cells. The mean adherence to 48-hour hCMV-infected HeLa cells was still significantly higher than to control cells. When compared with control cells, strains JP2 and ATCC33384 also adhered in higher numbers to 48-hour mock-infected cells. However, the magnitude of this increase was lower then with a pre-infection with active hCMV. No statistically significant differences between the relative enhancements in epithelial colonization of the different strains were detected. Primary epithelial monolayers from eight periodontitis patients were pre-infected with either hCMV or with mock preparations at an MOI of 100 viral genomes/cell 6, 12, 24, and 48 hrs prior to infection with Actinobacillus actinomycetemcomitans strain JP2. This highly leukotoxic strain was chosen because of its enhanced pathogenicity in periodontitis (Haubek et al., 2004). For these primary cells also, a significant increase in adherence took place with increasing hCMV infection time, when compared with colonization of control cells (Fig. 3BGo). The decrease seen after 48 hrs of pre-infection when compared with 24-hour hCMV-infected cells was not statistically significant. Mock-infected primary cells showed a slightly increased adherence that did not increase over time. The colonization of hCMV-infected primary cells was significantly higher than for time-matched mock-infected cells.


Figure 3
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  		Figure 3. Viral pre-infection time and bacterial adherence. (A) Effect of hCMV pre-infection time on adherence of Actinobacillus actinomycetemcomitans strains to HeLa cells. Cells were exposed for 1 hr to hCMV (MOI 100) and incubated for an additional 6, 12, 24, or 48 hrs, or were mock-infected for 48 hrs (squares). Uninfected cells served as controls. Data are expressed as the proportion of cell-associated bacteria relative to the numbers of bacteria added to the cells initially. Each datapoint represents the mean ± SEM for 12 independent experiments. *P < 0.05, hCMV-infected group vs. uninfected control; #P < 0.05, 48-hour hCMV-infected group vs. 48-hour mock-infected group; °P < 0.05, 48-hour vs. 24-hour hCMV-infected group. (B) Effect of hCMV pre-infection time on adherence of Actinobacillus actinomycetemcomitans strain JP2 to primary cells. Cells were exposed for 1 hr to hCMV or mock preparation (MOI 100) and incubated for an additional 6, 12, 24, or 48 hrs. Uninfected cells served as controls. Data are expressed as the proportion of cell-associated bacteria relative to the numbers of bacteria added to the cells initially. Each datapoint represents the mean ± SEM for 8 independent experiments. *P < 0.05, hCMV- or mock-infected group vs. uninfected control; #P < 0.05, hCMV vs. mock-infected group.

 
IE-antigen Expression and Bacterial Adherence
To examine the relationship between IE-antigen expression and bacterial adherence, we used a four-day hCMV or mock infection of primary cells and the highly hCMV-infection-permissive HEL cells. Prolonged infection times were necessary to attain high levels (> 75%) of IE-positive HEL cells (confirmed by immunofluorescence microscopy). Mock infections did not result in IE-positive cells. Uninfected cells served as controls. Six independent adherence experiments were performed with Actinobacillus actinomycetemcomitans strain JP2, with primary cells from six different individuals. The number of IE-positive cells (i.e., hCMV permissiveness) was different for both cell types, but the enhancement in bacterial adherence did not correlate with IE-antigen expression in either cell type (Fig. 4Go). Viral pre-infection significantly increased the colonization of both HEL cells and primary cells (< 2% IE-positive), with 81% and 146%, respectively, as compared with control cells. Consistent with previous results, mock infection also increased subsequent bacterial adherence. This increase was identical, in HEL cells, to that of an active hCMV infection, whereas, for primary cells, the increase was significantly lower than for cells infected with active hCMV.


Figure 4
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  		Figure 4. Effect of IE-antigen expression on adherence of Actinobacillus actinomycetemcomitans strain JP2 to confluent monolayers. HEL and primary cells were exposed for 1 hr to hCMV or the mock preparation (MOI 100) and incubated for 96 hrs. Uninfected cells served as controls. Data are expressed as the proportion of cell-associated bacteria relative to the number of cell-associated bacteria on uninfected control cells (= 100%). Each datapoint represents the mean ± SEM for 6 independent experiments. *P < 0.05, hCMV- or mock-infected group vs. uninfected control; #P < 0.05, hCMV vs. mock-infected group.

 

   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS & METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
Viral infections can predispose to bacterial complications. Virus-induced host defense impairment, direct pathological effects, and alterations of cellular susceptibility to bacterial adherence are feasible mechanisms that may contribute to this synergism. Sanford and colleagues (1978) first described an increased adherence of bacteria to virally infected cultured cells. Similar observations have since been made for many viruses, cells, and bacteria. However, no studies are available that examine this issue relative to periodontitis. We explored the interaction between hCMV and Actinobacillus actinomycetemcomitans in an in vitro epithelial adherence assay because of their association in periodontal lesions (Ting et al., 2000). HeLa cells are an accepted model for the study of bacterial adherence (Han et al., 2000; Richardson et al., 2005). To ensure consistency between and among the different assays, we first performed experiments using HeLa cells, and secondarily verified our findings using primary cells. hCMV infects a broad spectrum of cells in vivo, but has a narrow host cell range for productive infection in vitro (Plachter et al., 1996). Therefore, the in vitro permissiveness of the cells used in this study was verified. Both HeLa cells and primary cells were semi-permissive for hCMV infection. Over 48 hrs, IE antigens, regulatory factors for cellular and viral genes, but no E antigens, structural factors for viral replication, were detected. To assess the influence of hCMV infection on Actinobacillus actinomycetemcomitans adherence, we infected HeLa cells with hCMV prior to bacterial infection. Clear effects of viral infection, evident as increased adherence, were confirmed for all 3 bacterial strains. This phenomenon has been observed for Salmonella typhimurium on semi-permissive hCMV-infected A549 epithelial cells (Holberg-Petersen et al., 1994). Similar to observations with S. typhimurium, the enhanced Actinobacillus actinomycetemcomitans adherence peaked after 24 hrs of hCMV infection and declined thereafter. In contrast to S. typhimurium, the decline was less pronounced and was not significant for primary cells. The relative adherence enhancement was similar to that reported for other viral-bacterial interactions (Brogden and Guthmiller, 2002). In our investigation, all Actinobacillus actinomycetemcomitans strains demonstrated hCMV-dependent enhancement of adherence, in contrast to findings with adenovirus (Hakansson et al., 1994) or influenza virus (Davison and Sanford, 1981). The magnitude of adherence enhancement was bacterial-strain-dependent. The Actinobacillus actinomycetemcomitans strains used were invasive, and although we did not differentiate adherent from invasive bacteria, it is possible that invasion was also affected by hCMV infection, as has been seen for other viruses (Konkel and Joens, 1990). Viral infectivity was also an important factor. Because inactivated viruses can enhance invasiveness of bacteria, mock infections were carried out (Holberg-Petersen et al., 1994). A significant increase in adherence was seen with mock-infected HeLa cells. This enhancement was significantly less then that observed with active virus. Similar interactions were reported for Coxsackie virus (Bukholm et al., 1985), and were presumed to be due to host cell membrane alterations by decreasing the Zeta potential or increasing the cellular cation concentration. Both decrease the electrical repulsive forces that act against reversible bacterial adhesiveness. Nevertheless, effects on cellular receptors and their function or on cellular metabolism cannot be excluded. Mock infections of primary cells showed a less pronounced and insignificant enhancement. The difference in response for primary and HeLa cells is unclear, but may originate from cellular surface differences. These results also indicate that the effect of hCMV is not solely dependent on the presence of infectious virus. To account for the non-infectious viruses, we expressed the viral concentrations used in this study by the number of viral genomes, instead of by the more routinely used technique of plaque-forming units. It is worthwhile pointing out the dichotomy observed between number of positive cells (± 2%) and the onset of expression of IE antigens (24–48 hrs), on the one hand, and the relative enhancement in adherence (factor 2 after 24 hrs), on the other. Similar observations were made for in vitro hCMV induced immunosuppression of mononuclear cells (Rice et al., 1984) and enhancement of S. typhimurium adherence to epithelial cells (Holberg-Petersen et al., 1994). We used a prolonged infection of primary cells and highly hCMV-permissive HEL cells to assess the role of IE antigens in adherence enhancement. The results showed less enhancement for highly permissive HEL cells. Interestingly, mock and hCMV infections of HEL cells resulted in similar enhancements of adherence, whereas, for primary cells, the difference between both remained identical to that after 48 hrs of pre-infection (Fig. 3Go). The mediators for this cell-type-specific response are unclear. Nevertheless, it was clear that IE expression had a low predictive value for enhancement in bacterial adherence. It is possible that hCMV-infected epithelial cells secrete factors that exert a paracrine effect on the adjacent epithelium, and thus augment bacterial colonization of IE-negative cells (Avadhanula et al., 2006). Additionally, this discrepancy may originate from the expression of different IE isoforms that could not be detected by the antibodies used (Rice et al., 1984), by specific factors such as cellular gene regulation by tegument proteins (Liu and Stinski, 1992), or by non-specific events such as increased lipid fluidity of the cellular membrane (Levanon et al., 1977).

The results supported our hypothesis that adherence of Actinobacillus actinomycetemcomitans is enhanced by hCMV in vitro. The significance of these findings in the development of periodontitis remains unclear. Nevertheless, it is likely that synergistic interactions between micro-organisms in the oral cavity have important implications in disease onset and progression.


   ACKNOWLEDGMENTS
 
This work was supported by research grants OT03/52 (KU Leuven), G0240.04 (nFWO), and IWT Flanders (IS); by GABA International, Basel, Switzerland; and by Dentaid Company, Barcelona, Spain. This paper is based on a thesis submitted at the Catholic University of Leuven in partial fulfillment of the requirements for the PhD degree (W. Teughels). Preliminary reports were presented at the 2003 IADR meeting in Göteborg, Sweden (IADR/Unilever Travel Award and Robert Frank Award), and at the 2004 IADR meeting in Honolulu (Hatton Award competitor representing the IADR Continental European Division).

Received November 21, 2005; Last revision October 12, 2006; Accepted October 17, 2006


   REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS & METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
American Academy of Periodontology (1996). Consensus report on periodontal diseases: pathogenesis and microbial factors. Ann Periodontol 1:926–932.[Medline]

Avadhanula V, Rodriguez CA, Devincenzo JP, Wang Y, Webby RJ, Ulett GC, et al. (2006). Respiratory viruses augment the adhesion of bacterial pathogens to respiratory epithelium in a viral species-and cell type-dependent manner. J Virol 80:1629–1636.[Abstract/Free Full Text]

Beuselinck K, Van Ranst M, Van Eldere J (2005). Automated extraction of viral-pathogen RNA and DNA for high-throughput quantitative real-time PCR. J Clin Microbiol 43:5541–5546.[Abstract/Free Full Text]

Brogden KA, Guthmiller JM (2002). Polymicrobial diseases involving viruses and bacteria. In: Polymicrobial diseases. Brogden KA, Guthmiller JM, editors. Washington, DC: American Society for Microbiology, pp. 201–334.

Bukholm G, Holberg-Petersen M, Degre M (1985). Invasiveness of Salmonella typhimurium in HEp-2 cell cultures pretreated with UV-inactivated coxsackie virus. Acta Pathol Microbiol Immunol Scand [B] 93:61–65.[ISI][Medline]

Davison VE, Sanford BA (1981). Adherence of Staphylococcus aureus to influenza A virus infected Madin-Darby canine kidney cell cultures. Infect Immun 32:118–126.[Abstract/Free Full Text]

Eick S, Pfister W (2004). Efficacy of antibiotics against periodontopathogenic bacteria within epithelial cells: an in vitro study. J Periodontol 75:1327–1334.[ISI][Medline]

Hakansson A, Kidd A, Wadell G, Sabharwal H, Svanborg C (1994). Adenovirus infection enhances in vitro adherence of Streptococcus pneumoniae. Infect Immun 62:2707–2714.[Abstract/Free Full Text]

Han YW, Shi W, Huang GT, Kinder Haake SA, Park NH, Kuramitsu HK, et al. (2000). Interactions between periodontal bacteria and human oral epithelial cells: Fusobacterium nucleatum adheres to and invades epithelial cells. Infect Immun 68:3140–3146.[Abstract/Free Full Text]

Haubek D, Ennibi OK, Poulsen K, Benzarti N, Baelum V (2004). The highly leukotoxic JP2 clone of Actinobacillus actinomycetemcomitans and progression of periodontal attachment loss. J Dent Res 83:767–770.[Abstract/Free Full Text]

Holberg-Petersen M, Bukholm G, Rollag H, Degre M (1994). Infection with human cytomegalovirus enhances bacterial adhesiveness and invasiveness in permissive and semipermissive cells. APMIS 102:703–710.[ISI][Medline]

Konkel ME, Joens LA (1990). Effect of enteroviruses on adherence to and invasion of HEp-2 cells by Campylobacter isolates. Infect Immun 58:1101–1105.[Abstract/Free Full Text]

Kuramitsu HK (2003). Molecular genetic analysis of the virulence of oral bacterial pathogens: an historical perspective. Crit Rev Oral Biol Med 14:331–344.[Abstract/Free Full Text]

Levanon A, Kohn A, Inbar M (1977). Increase in lipid fluidity of cellular membranes induced by adsorption of RNA and DNA virions. J Virol 22:353–360.[Abstract/Free Full Text]

Liu B, Stinski MF (1992). Human cytomegalovirus contains a tegument protein that enhances transcription from promoters with upstream ATF and AP-1 cis-acting elements. J Virol 66:4434–4444.[Abstract/Free Full Text]

Meyer DH, Sreenivasan PK, Fives-Taylor PM (1991). Evidence for invasion of a human oral cell line by Actinobacillus actinomycetemcomitans. Infect Immun 59:2719–2726.[Abstract/Free Full Text]

Plachter B, Sinzger C, Jahn G (1996). Cell types involved in replication and distribution of human cytomegalovirus. Adv Virus Res 46:195–261.[ISI][Medline]

Quirynen M, Papaioannou W, Van Steenbergen TJ, Dierickx K, Cassiman JJ, van Steenberghe D (2001). Adhesion of Porphyromonas gingivalis strains to cultured epithelial cells from patients with a history of chronic adult periodontitis or from patients less susceptible to periodontitis. J Periodontol 72:626–633.[ISI][Medline]

Reed WP, Williams RC Jr (1978). Bacterial adherence: first step in pathogenesis of certain infections. J Chronic Dis 31:67–72.[ISI][Medline]

Rice GP, Schrier RD, Oldstone MB (1984). Cytomegalovirus infects human lymphocytes and monocytes: virus expression is restricted to immediate-early gene products. Proc Natl Acad Sci USA 81:6134–6138.[Abstract/Free Full Text]

Richardson J, Craighead JC, Cao SL, Handfield M (2005). Concurrence between the gene expression pattern of Actinobacillus actinomycetemcomitans in localized aggressive periodontitis and in human epithelial cells. J Med Microbiol 54(Pt 5):497–504.[Abstract/Free Full Text]

Rudney JD, Chen R, Sedgewick GJ (2005). Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, and Tannerella forsythensis are components of a polymicrobial intracellular flora within human buccal cells. J Dent Res 84:59–63.[Abstract/Free Full Text]

Sanford BA, Shelokov A, Ramsay MA (1978). Bacterial adherence to virus-infected cells: a cell culture model of bacterial superinfection. J Infect Dis 137:176–181.[ISI][Medline]

Schols D, Snoeck R, Neyts J, De Clercq E (1989). Detection of immediate early, early and late antigens of human cytomegalovirus by flow cytometry. J Virol Methods 26:247–254.[ISI][Medline]

Selinger DS, Reed WP, McLaren LC (1981). Model for studying bacterial adherence to epithelial cells infected with viruses. Infect Immun 32:941–944.[Abstract/Free Full Text]

Sethi S (2002). Bacterial pneumonia. Managing a deadly complication of influenza in older adults with comorbid disease. Geriatrics 57:56–61.[ISI][Medline]

Slots J (2005). Herpesviruses in periodontal diseases. Periodontol 2000 38:33–62.

Teughels W, Van Eldere J, van Steenberghe D, Cassiman JJ, Fives-Taylor PM, Quirynen M (2005). Influence of nicotine and cotinine on epithelial colonization by periodontopathogens. J Periodontol 76:1315–1322.[ISI][Medline]

Ting M, Contreras A, Slots J (2000). Herpesvirus in localized juvenile periodontitis. J Periodontal Res 35:17–25.[ISI][Medline]





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