Evaluation of Streptococcus pneumoniae Type XIV Opsonins by Phagocytosis-Associated Chemiluminescence and a Bactericidal Assay

Abstract

The relative roles of serum factors required for opsonization of type XIV Streptococcus pneumoniae were investigated by means of luminol-enhanced chemiluminescence (CL), bactericidal, and immunofluorescence assays employing adult sera containing high (>1,000 ng of antibody nitrogen per ml) or low (<200 ng of antibody nitrogen per ml) antibody concentrations as determined by radioimmunoassay. Specific antibody concentration correlated directly with both total and heat-labile CL activity (P < 0.005) and with the bactericidal index (P < 0.05) at a serum concentration of 10%. The importance of specific antibody as an opsonin was confirmed by the abolition of CL activity and immunoglobulin immunofluorescence observed after absorption of heated sera with type XIV pneumococcal cells and by the dose response in CL and bactericidal activity observed with the addition of immunoglobulin G to hypogammaglobulinemic serum. A role for the classical complement pathway in opsonization was indicated by significantly greater CL integrals for high-antibody sera than for low-antibody sera depleted of factor D and by the bactericidal activity noted for untreated, but not magnesium ethylene glycol-bis(3-aminoethyl ether)-N,N-tetraacetic acidchelated low-antibody sera. The alternative pathway contributed more than half of the CL activity of both highand low-antibody sera. However, after magnesium ethylene glycol-bis(P-aminoethyl ether)-N,N-tetraacetic acid chelation, only sera with high antibody concentrations or agammaglobulinemic serum reconstituted with immunoglobulin G with high specific antibody levels supported significant bactericidal activity. Therefore, type-specific antibody and complement promote opsonization of type XIV S. pneumoniae, and this may occur via either complement pathway. These results suggest that CL is a suitable tool to delineate serum factors and their contribution to opsonization, but results must be related to other functional assays.

Factors affecting the chemiluminescent response of fish phagocytes

“…Zymosan opsonized with normal pooled bass serum elicited a CL response which increased at a faster rate and reached a peak approximately five times greater than with nonopsonized zymosan. An effect of concentration of opsonized zymosan was also indicated as 1 mgml-I elicited a peak CL response approximately twice that of 0.5 mg ml-l. Opsonization of bacteria has been shown to enhance both the CL response and bacterial killing by phagocytes (Madonna & Allen, 1981;Gardner et al, 1982;MacGowan et al, 1983). Aerornonas hydrophila opsonized with normal pooled bass serum elicited a greater peak CL response from striped bass phagocytes than nonopsonized A .…”

“…Opsonization has been shown to enhance the CL response of phagocytes stimulated by living and killed bacteria. The degree of enhancement has been used to evaluate different opsonic requirements for phagocytosis (Allen, 1977;Madonna & Allen, 1981;Gardner et al, 1982;MacGowan et al, 1983). Opsonization of A .…”

Effect of C-reactive protein on the complement-mediated stimulated of human neutrophils by Streptococcus pneumoniae serotypes 3 and 6

“…Activated polymorphonuclear leukocytes (PMN) produce a chemiluminescence (CL) that originates from the relaxation of electronically excited molecules generated during the destruction of phagocytized organisms (2). This CL response has been used to evaluate the opsonophagocytic response of PMN to a number of bacterial species, including S. pneumoniae (5,11,18). We have recently shown a CRPand complement-dependent enhancement of the PMN CL response to S. pneumoniae serotype 27 (lOa).…”

Granulocyte phagocytosis of Streptococcus pneumoniae in properdin-deficient serum

“…It is also known that phagocytes possess receptors for immunoglobulin G (IgG)-Fc and for C3b, the activated form of C3 (7). The main role of antibody in the opsonization of S. pneumoniae is probably to mediate the fixation of opsonic C3b to the bacterial surface by classical, and possibly also by alternative, pathway activation of complement (4,12). Many serotypes of S. pneumoniae activate the alternative pathway (9,23,25), obviously contributing to opsonization (12,13,17).…”

“…In attempts to investigate the relative roles of the two complement pathways for opsonization of different serotypes of S. pneumoniae, studies have been performed with serum reagents that provide a nonfunctional classical pathway (12,13,17). The opsonic requirements for S. pneumoniae serotype 14 have been examined with sera depleted of factor D (12). Not much is known, however, of the influence on opsonization of an impaired alternative pathway.…”