Antibody-Independent Neutralization of Vesicular Stomatitis Virus by Human Complement

Abstract

Abstract Normal human serum neutralized up to 5 logs10/ml of vesicular stomatitis virus (VSV) grown in hamster (BHK), mouse (L), or human (HFF) cell lines when equal volumes of virus (∼105 PFU/ml) and serum were incubated together for 30 min at 37°C. The neutralizing activity of fresh serum was heat labile, thus distinguishing this activity from that of immune human serum which was heat stable. The lack of an antibody requirement for VSV neutralization by normal human serum was demonstrated in three ways: first, isolated human immunoglobulins failed to neutralize VSV or potentiate neutralization by fresh serum; second, absorption of fresh serum with uninfected cells or cells bearing VSV antigens did not reduce neutralizing activity; third, agammaglobulinemic serum mediated C3b deposition on the viral surface. Neutralization by normal human serum was complement (C) dependent and required the early reacting components of the classical C pathway (C1q, C2, C4) but not the late reacting components (C5, C6, C7, C8) or the alternative C pathway (factor B). Incubation of serum with concentrated VSV reduced the CH50, C2, C3, and C4 titers by 23.5, 21, 22, and 31%, respectively, and was accompanied by deposition of C3b on the viral surface. Purified VSV and the isolated external glycoprotein of VSV, G protein, bound 125I-C1q, and purified VSV also bound C1̄. However, VSV did not activate purified reconstituted C1, and VSV was not neutralized by isolated C1, C2, C3, C4, and C5. Therefore, an additional serum factor is apparently required for neutralization. These studies indicate that neutralization of VSV by normal human serum involves the components of the classical C pathway through C3b and a nonimmunoglobulin serum factor. The reaction is initiated by the direct attachment of C1 to the G protein, which leads to C activation and the sequential deposition of the C components through C3b on the viral envelope. The mechanism of action of the nonimmunoglobulin serum factor remains to be established.