Biological Correlates of Capsular (Quellung) Reactions ofCryptococcus neoformans

Abstract

The capsular swelling or quellung reaction was reported almost 100 years ago and described the effect of Abs on the appearance of microbial capsules. Despite widespread use to assess Ab binding to capsules, relatively little is known as to the mechanism of this effect or its biological consequences. The fungus Cryptococcus neoformans is an attractive system to study capsule reactions because it has a large polysaccharide capsule that is readily visible by light microscopy. When viewed by differential interference contrast microscopy, binding of mAb to C. neoformans cells produced two distinct capsular reactions that depended on the Ab epitope specificity and the yeast serotype. In the first pattern, termed “rim,” the capsule appears transparent with a highly refractive outer edge. In the second pattern, termed “puffy,” the capsule appears opaque and lacks a highly refractive outer rim. mAbs that bind with a rim pattern suppress the overall rate of C3 deposition on the yeast via the classical and alternative complement pathways. In contrast, mAbs that bind with a puffy pattern do not affect C3 deposition. Protective and nonprotective IgM mAbs produce rim and puffy patterns, respectively. These results indicate that: 1) capsule reactions are a consequence of Ab-induced changes in capsular refractive index; 2) the type of capsule reaction depends on the Ab specificity; and 3) Ab-induced changes in refractive index correlate with biological activities important for host defense against C. neoformans. Our results provide the first evidence associating distinct capsule reaction patterns with Ab biological activity.

Immunoglobulin M Efficacy AgainstCryptococcus neoformans: Mechanism, Dose Dependence, and Prozone-Like Effects in Passive Protection Experiments

“…The interaction of each mAb with the capsule also produces qualitatively different types of capsular reactions (16). Both mAbs activate the classical complement pathway, but mAb 12A1 suppresses capsular activation of the alternative complement pathway (16). The differences in immunofluorescence-binding pattern, capsular reaction, competition studies, and peptide mimotope reactivity indicate that these two mAbs bind to different epitopes in GXM (12,13,17).…”

“…These patterns reflect the formation of different types of Ag-Ab complexes in the C. neoformans capsule (15). The interaction of each mAb with the capsule also produces qualitatively different types of capsular reactions (16). Both mAbs activate the classical complement pathway, but mAb 12A1 suppresses capsular activation of the alternative complement pathway (16).…”

Constant domains influence binding of mouse–human chimeric antibodies to the capsular polypeptide ofBacillus anthracis

“…These reactions show that antibody binding to capsular structures may produce a variety of patterns based on the capsule architecture, epitope accessibility, and sometimes antibody isotype/subclass. 11,18 Our current understanding of these patterns largely comes from studying antibodies that react with the GXM capsule of C. neoformans. These studies separate antibody-capsule reactions into two distinct classes.…”

“…the K D of each antibody was determined using the steady-state equilibrium model in BiAevaluation software, where the theoretical rU max is determined and the apparent K D is equal to the antibody concentration that is necessary to reach ½ rU max . 11,14,18 Two phenotypes make up the most commonly observed patterns; these are categorized as either "rim"-type or "puffy"-type, depending on the distribution of the antibody over and through the intact capsule.…”

Opsonin-free, real-time imaging of Cryptococcus neoformans capsule during budding

“…C. neoformans capsules labelled with monoclonal antibodies [1] or complement [2] were incubated in capsule-inducing conditions and at specific time points during induction the probes were detected using fluorescence microscopy. More recently, the Quellung reaction observed on differential interference microscopy (DIC), which produces a change in refractive index when antibodies bind to the capsule [3,4], was used to monitor capsule thickening in C. neoformans cells in real time [5,6]. These reports yielded important insights into the kinetics of capsule synthesis, but all suffered from an important caveat: because antibodies and complement proteins interact directly with capsule components, they may change its physical properties, especially at the reasonably high concentrations needed for the observations.…”