Specificity of three anti-complement factor 3 monoclonal antibodies.

Monoclonal antibodies which recognize specific C3 fragments may be used to distinguish C3 cleavage products bound to organisms. We defined the specificity of three commercially available monoclonal antibodies by Western immunoblot analysis, enzyme-linked immunosorbent assay, and a quantitative flow cytometric technique. Two monoclonal antibodies with specificity for (i) an erythrocyte-bound C3d epitope or (ii) an erythrocyte-bound C3c epitope retained their specificity in all assays. However, the third monoclonal antibody with selectivity for erythrocyte-bound C3bi failed to retain specificity for C3bi bound to non-erythrocyte surfaces in each of our assays; binding instead to all C3 fragments containing the C3g domain. We postulate that erythrocyte C3b-binding surface proteins may alter the availability of certain C3b epitopes and influence observed anti-C3 monoclonal antibody specificity. We conclude that the specificity of monoclonal antibodies for C3 fragments should be confirmed with assays which do not employ erythrocytes or other surfaces bearing C3 receptors. Also, our quantitative flow cytometric technique is a potentially valuable tool for the enumeration of particle-bound C3 fragments.

Complement binding to Aspergillus conidia correlates with pathogenicity.

Complement has significant effects on the phagocytosis of Aspergillus organisms. We examined the amount and type of complement component C3 bound to the resting conidia of 29 isolates from nine Aspergillus species. The highly pathogenic species A. fumigatus and A. flavus bound fewer C3 molecules per unit of conidial surface area than did the less pathogenic species A. glaucus, A. nidulans, A. niger, A. ochraceus, A. terreus, A. versicolor, and A. wentii, as determined by quantitative flow cytometry. Immunoblot analysis of C3 fragments bound to conidia demonstrated that for all species most C3b was apparently converted to iC3b. For seven species, iC3b was clearly the major C3 product recognized by immunoblotting. However, A. niger and A. nidulans appear to promote further breakdown of opsonic C3 fragments to C3dg. We found significant variations in size and C3 binding among isolates within the same species. Intraspecies variation may contribute to seemingly discrepant results obtained in studies of Aspergillus phagocytosis.

Complications of bacteremia due to Stomatococcus mucilaginosus in neutropenic children.

Stomatococcus mucilaginosus, a normal inhabitant of the human oral cavity and upper respiratory tract, can cause fatal sepsis and meningitis in neutropenic patients. We identified eight cases of bacteremia due to S. mucilaginosus in children with cancer, of whom five developed complications despite receiving appropriate antibiotics. At the time cultures were positive, seven patients had profound neutropenia (< 100 neutrophils and band forms/mm3) and four had mucositis; five had central venous catheters. In two cases, there was unequivocal evidence of catheter-related sepsis. Bacteremia was eradicated in all patients within 48 hours after initiation of antibiotics. Despite prompt instigation of effective antibiotic therapy, the complication rates in this series were high: septic shock (50%), pneumonia (50%), dermatologic manifestations (38%), altered neurological status (25%), meningitis (13%), and adult respiratory distress syndrome (13%). No fatalities were attributable to S. mucilaginosus infection. These cases illustrate the virulence of S. mucilaginosus organisms in neutropenic children and suggest a substantial risk of sequelae even when adequate antibiotic therapy is given.

Monoclonal antibodies against a 97-kilodalton antigen from Aspergillus flavus.

We prepared a panel of five monoclonal antibodies (MAbs) directed against Aspergillus flavus that all reacted against one 97-kDa antigen by western blot (immunoblot). Flow cytometry demonstrated that these antibodies bound (in increasing degrees) to all morphologic stages of A. flavus growth: conidia, swollen conidia, and hyphae. Cross-reactivity among species was examined by enzyme-linked immunosorbent assay of fungal culture filtrates. Four MAbs reacted with 10 of 11 A. flavus isolates, and the fifth one reacted with 9 of them. One MAb also reacted with A. fumigatus, two reacted with A. niger, A. wentii, and A. nidulans, and all five reacted with A. ochraceus. None reacted with A. terreus, A. glaucus, A. versicolor, or a Penicillium species. Each MAb bound to A. flavus hyphae in formalin-fixed paraffin sections of a muscle biopsy from a confirmed human case of invasive aspergillosis. In summary, these MAbs identified a 97-kDa antigen found on A. flavus that is both surface bound and an exoantigen. Either the same or a cross-reacting antigen is present in A. fumigatus and other Aspergillus species.

Comparative virulence of Staphylococcus epidermidis isolates in a murine catheter model.

Among coagulase-negative staphylococci, Staphylococcus epidermidis is the species most commonly implicated in catheter-related infections. Whether some staphylococcal organisms are inherently more virulent than others, or whether their ability to infect relates more to the sheer numbers of organisms at the catheter site, remains unclear. We therefore compared eight S. epidermidis isolates and two other coagulase-negative staphylococci using a murine model that allowed us to quantify catheter colonization and abscess formation in the same animal. The organisms were isolated from different clinically relevant settings and were classified according to their slime phenotype. The ability to evoke abscesses or colonize catheters in half of the animals (ID50) was assessed. ID50 inoculum titers (log10 data +/- SD) ranged widely, from 8.5 +/- 0.3 to 10.2 +/- 0.2 for abscess formation (p < 0.005) and from 7.5 +/- 0.5 to 10.3 +/- 1.0 for catheter colonization (p < 0.005). ID50 values by statistical criteria suggested variability among organisms in the ability to induce abscess formation. High slime production correlated with both parameters, but not with the clinical source of the isolate. Our findings demonstrate impressive heterogeneity in the ability of a representative group of S. epidermidis isolates to colonize catheters and to evoke abscess formation and implicate slime productivity as a major virulence factor. The murine model used permitted simultaneous analysis of multiple factors involved in pathogenesis and should be useful in establishing the basis of S. epidermidis pathogenicity.