J. Med. Microbiol. - Vol. 40 (1994), 48-54 0 1994 The Pathological Society of Great Britain and Ireland The role of HSV-induced Fc- and C3b(i)-receptors in bacterial adherence LIA A. M. DE GRAAF-MILTENBURG, KARIN E. VAN VLIET, T. L. M. TEN HAGEN, J. VERHOEF and J. A. G. VAN STRIJP" Eukman- Winkler Laboratory of Medical Microbiology, Utrecht University, Utrecht 3584 CX, The Netherlands Summary. Herpes simplex virus type- 1 (HSV- 1) induces Fc- and C3b(i)-receptors on infected cells. The role of these receptors in bacterial superinfection was studied by comparing the adherence of non-opsonised and opsonised bacteria to HSV-infected and non-infected HEp-2 cells. A flow cytometric adherence assay, based on the fluorescent quantitation of FITC-labelled bacteria, was developed. Opsonisation of Staphylococcus epidermidis with human serum, resulted in a marked increase in adherence to HSV-infected cells and revealed a role for C3b(i)R- and FcR-mediated adhesion. However, the enhanced adherence never exceeded the level of attachment to non-infected cells. Increased adherence of other pathogenic bacteria, including Escherichia coli, Streptococcus pneumoniae, Haemophilus influenzae and Pseudomonas aeruginosa was not observed, indicating that the HSV-receptors play a minor role in secondary infections. Bacterial adhesion factors such as the fimbriae of E. coli played a more dominant role in the adherence of bacteria to HSV-infected cells. Introduction A number of herpes viruses can induce the expression of opsonin-receptors on the surface of cultured fibroblasts, endothelial and epithelial cells. The viral proteins associated with these receptors have been identified for herpes simplex virus type-1 (HSV-1). A complex formed by the glycoproteins E and I (gE/gI) has been described as the Fc-receptor (FcR), and glycoprotein C (gC) has been shown to act as a receptor for C3b and C3bi (C3b(i)R).',2 The biological significance of the HSV-receptors in vivo is unknown. Since these glycoproteins are not essential for the actual replication of HSV-1 in cultured cells, they may play an important role in path~genicity.~ It is possible that these receptors could lead to the attachment of partially opsonised bacteria, thus enhancing the possibility of secondary infections. These secondary infections can in turn lower the local host resistance and thereby benefit the virus. This hypothesis is supported by the fact that opsonised particles such as erythrocytes adhere more strongly to HSV-infected cells than to non-infected cells and more strongly than non-opsonised erythrocytes.2- The concept that certain viral infections predispose the host to bacterial infections has long been recognised and is well documented. A study in which the history of genital herpes was associated with the increased risk of vaginal Staphylococcus aureus infection has been rep~rted.~ Studies performed with influenza A virus- infected cells have demonstrated that bacterial super- infections are correlated with the increased attachment of staphylococci to the virus-infected cells.6 The adherence of streptococci and staphylococci to HSV-infected cells has been studied in the past by electronmicroscopy.7However, the role of the Fc- and C3b(i)-receptors induced on the HSV-infected cells was not taken into account in this study. FcR and C3b(i)R may create alternative attachment sites for opsonised bacteria as shown by Mackowiak et al., who demonstrated an increased antibody-mediated Escherichia coli adhesion to cytomegalovirus-induced Fc-receptors.* Moreover, Friedman also showed that the HSV-gC was responsible for the adherence of complement-coated Salmonella typhi to HSV-infected endothelial cells. Therefore, we examined the hypothesis that HSV- induced FcR and C3b(i)R may create alternative attachment sites for opsonised bacteria to HSV- infected cells. The adherence of various pathogenic bacterial strains, including S. epidermidis, E. coli, Haemophilus injluenzae, Pseudomonas aeruginosa and Streptococcus pneumoniae to HSV-infected and non- infected HEp-2 cells was determined by a flow cyto- metric bacterial adherence assay. Materials and methods Cells and viruses Received 24 March 1993; accepted 7 June 1993. * Correspondence should be sent to Dr J. A. G. Van Strijp. HEp-2 cells (Flow Laboratories, Irvine) were cul- tured in RPMI 1640 (Gibco, Paisley, Renfrewshire), 48