Fc-dependent polyclonal antibodies and antibodies to outer membrane proteins A and B, but not to lipopolysaccharide, protect SCID mice against fatal Rickettsia conorii infection

Abstract

An emphasis on cellular immunity against Rickettsia has led to neglect of analysis of the role of antibody. The availability of an excellent mouse model of spotted fever rickettsiosis enabled investigation of a potential role of antibody in immunity to Rickettsia conorii. C3H severe combined immunodeficiency (SCID) mice were passively transfused with monoclonal antibodies against rickettsial outer membrane protein A (OmpA), OmpB, or lipopolysaccharide (LPS), polyclonal anti-R. conorii serum, Fab fragments of polyclonal antiserum, or no antibodies and then challenged 48 h later with 10 50% lethal doses (LD(50)) of R. conorii. All mice that received monoclonal antibodies against OmpA and two of four mice that received monoclonal antibodies against OmpB or polyclonal antisera were completely protected, but the recipients of anti-LPS antibodies or the Fab fragments were not protected. Polyclonal antibody treatment of C3H SCID mice that had been infected with 10 LD(50) of R. conorii 4 or 5 days earlier prolonged the life of the infected mice from 10.4 to 22.5 days and resulted in decreased levels of infectious rickettsiae in the spleen and liver 24 and 48 h later. Treatment with protective antibodies resulted in the development of large aggregates of R. conorii antigens in splenic macrophages and intraphagolysosomal rickettsial death and digestion. The kinetics of development of antibodies to R. conorii determined by immunoblotting revealed antibodies to LPS on day 6 and antibodies to OmpA and OmpB on day 12, when recovery from the infection had already occurred. Antibodies to particular epitopes of OmpA and OmpB may protect against reinfection, but they may not play a key role in immunity against primary infection. Antibodies might be useful for treating infections with antibiotic-resistant organisms, and some B-cell epitopes should be included in a subunit vaccine.

FIG. 1.

Immunohistochemical staining of R. conorii in the spleens of mice 32 h after treatment with polyclonal mouse anti-R. conorii serum (A and C) or normal mouse serum (B and D). The antibody-treated mice (A) contained a greater quantity of rickettsial antigen in their spleens than the normal serum-treated controls contained (B). The rickettsial antigen in splenic macrophages of antibody-treated mice (C) comprised amorphous rickettsial antigen and fragmented particles smaller than rickettsiae (short and long arrows). Rickettsial antigen in the spleens of normal serum-treated mice (D) included individual, morphologically typical rickettsiae (arrowhead). (A and B) Magnification, ×200; (C and D) magnification, ×1,000.

FIG. 2.

Electron photomicrograph of the liver of a mouse infected with R. conorii 32 h after treatment with nonimmune serum. Several viable rickettsiae (arrows) are free in the cytosol. The arrowhead indicates a rickettsia enlarged in the inset. Bar = 0.5 μm. (Inset) Viable rickettsia with typical bacillary morphology, including normal ultrastructure of the cytoplasm, nucleoid, cell wall (arrowhead), and cytoplasmic membrane (arrow). Bar = 0.5 μm.

FIG. 3.

Electron photomicrograph of three morphologically nonviable rickettsiae (large arrows) within a phagolysosome (asterisk) of a splenic macrophage in an R. conorii-infected SCID mouse 32 h after polyclonal immune antibody treatment. There are distortions of the cell wall (small arrows), dilated periplasmic space (p), loss of the typical bacillary morphology, and vesicles shed from the cell wall (arrowheads). Bar = 0.5 μm.

FIG. 4.

Western immunoblots of R. conorii antigens reactive with sera collected serially from mice on days 6 (lane 2), 12 (lane 3), 18 (lane 4), and 30 (lane 5) after infection (left panel) and with monoclonal antibodies to OmpA (lane 2), OmpB (lane 3), and LPS (lane 4) (right panel). Lanes 1 contains molecular size standards (37 to 150 kDa).