scholarly journals Capsular Localization of the Cryptococcus neoformans Polysaccharide Component Galactoxylomannan

2008 ◽  
Vol 8 (1) ◽  
pp. 96-103 ◽  
Author(s):  
Magdia De Jesus ◽  
André Moraes Nicola ◽  
Marcio L. Rodrigues ◽  
Guilhem Janbon ◽  
Arturo Casadevall
Keyword(s):  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Protective Antigen ◽  
Immune Serum ◽  
Single Chain ◽  
Encapsulated Cells ◽  
Daughter Cells ◽  
The Poor ◽  
Polysaccharide Capsule ◽  
Chain Fraction

ABSTRACT Cryptococcus neoformans capsular polysaccharide is composed of at least two components, glucuronoxylomannan (GXM) and galactoxylomannans (GalXM). Although GXM has been extensively studied, little is known about the location of GalXM in the C. neoformans capsule, in part because there are no serological reagents specific to this antigen. To circumvent the poor immunogenicity of GalXM, this antigen was conjugated to protective antigen from Bacillus anthracis as a protein carrier. The resulting conjugate elicited antibodies that reacted with GalXM in mice and yielded an immune serum that proved useful for studying GalXM in the polysaccharide capsule. In acapsular cells, immune serum localized GalXM to the cell wall. In capsulated cells, immune serum localized GalXM to discrete pockets near the capsule edge. GalXM was abundant on the nascent capsules of budding daughter cells. The constituent sugars of GalXM were found in vesicle fractions consistent with vesicular transport for this polysaccharide. In addition, we generated a single-chain fraction variable fragment antibody with specificity to oxidized carbohydrates that also produced punctate immunofluorescence on encapsulated cells that partially colocalized with GalXM. The results are interpreted to mean that GalXM is a transient component of the polysaccharide capsule of mature cells during the process of secretion. Hence, the function of GalXM appears to be more consistent with that of an exopolysaccharide than a structural component of the cryptococcal capsule.

scholarly journals The capsule ofCryptococcus neoformansmodulates phagosomal pH through its acid-base properties

2018 ◽  
Author(s):  
Carlos M. De Leon-Rodriguez ◽  
Man Shun Fu ◽  
M. Osman Corbali ◽  
Radames J.B. Cordero ◽  
Arturo Casadevall
Keyword(s):  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Glucuronic Acid ◽  
Pathogenic Fungus ◽  
Weak Acid ◽  
Phagocytic Cells ◽  
Acid Base ◽  
Encapsulated Cells ◽  
Human Pathogenic Fungus ◽  
Base Properties

AbstractPhagosomal acidification is a critical cellular mechanism for the inhibition and killing of ingested microbes by phagocytic cells. The acidic environment activates microbicidal proteins and creates an unfavorable environment for the growth of many microbes. Consequently, numerous pathogenic microbes have developed strategies for countering phagosomal acidification through various mechanisms that include interference with phagosome maturation. The human pathogenic fungusCryptococcus neoformansresides in acidic phagosome after macrophage ingestion that actually provides a favorable environment for replication since the fungus replicates faster at acidic pH. We hypothesized that the glucuronic acid residues in the capsular polysaccharide had the capacity to affect phagosome acidity through their acid-base properties. A ratiometric fluorescence comparison of imaged phagosomes containingC. neoformansto those containing beads showed that the latter were significantly more acidic. Similarly, phagosomes containing non-encapsulatedC. neoformanscells were more acidic than those containing encapsulated cells. Acid-base titrations of isolatedC. neoformanspolysaccharide revealed that it behaves as a weak acid with maximal buffering capacity around pH 4-5. We interpret these results as indicating that the glucuronic acid residues in theC. neoformanscapsular polysaccharide can buffer phagosomal acidification. Interference with phagosomal acidification represents a new function for the cryptococcal capsule in virulence and suggests the importance of considering the acid-base properties of microbial capsules in the host-microbe interaction for other microbes with charged residues in their capsules.ImportanceCryptococcus neoformansis the causative agent of cryptococcosis, a devastating fungal disease that affects thousands of individuals worldwide. This fungus has the capacity to survive inside phagocytic cells, which contributes to persistence of infection and dissemination. One of the major mechanisms of host phagocytes is to acidify the phagosomal compartment after ingestion of microbes. This study shows that the capsule ofC. neoformanscan interfere with full phagosomal acidification by serving as a buffer.

scholarly journals Cryptococcus neoformans capsule regrowth experiments reveal dynamics of enlargement and architecture

2021 ◽  
Author(s):  
Maggie P. Wear ◽  
Ella Jacobs ◽  
Siqing Wang ◽  
Scott McConnell ◽  
Anthony Bowen ◽  
...  
Keyword(s):  
Cell Wall ◽  
Cryptococcus Neoformans ◽  
Virulence Factor ◽  
Fungal Pathogen ◽  
Capsular Polysaccharide ◽  
Wall Surface ◽  
Polysaccharide Capsule ◽  
Capsular Material ◽  
Enzymatic Methods

The polysaccharide capsule of fungal pathogen Cryptococcus neoformans is a critical virulence factor that has historically evaded characterization. Polysaccharides remain attached to the cell as capsular polysaccharide (CPS) or are shed into the surroundings in the form of exopolysaccharide (EPS). While a great deal of study has been done examining the properties of EPS, far less is known about CPS. In this work, we detail the development of new physical and enzymatic methods for the isolation of CPS which can be used to explore the architecture of the capsule and removed capsular material. Sonication and glucanex digestion yield soluble CPS preparations, while French Press and modified glucanex digestion plus vortexing remove the capsule and cell wall producing polysaccharide aggregates that we call capsule ghosts. The existence of capsule ghosts implies an inherent organization that allows it to exist independent of the cell wall surface. As sonication and glucanex digestion were noncytotoxic, it was possible to observe the cryptococcal cells rebuilding their capsule, revealing new insights into capsule architecture and synthesis consistent with a model in which the capsule is assembled from smaller polymers, which are then assemble into larger ones.

scholarly journals The Capsule ofCryptococcus neoformansModulates Phagosomal pH through Its Acid-Base Properties

mSphere ◽  
2018 ◽  
Vol 3 (5) ◽  
Author(s):  
Carlos M. De Leon-Rodriguez ◽  
Man Shun Fu ◽  
M. Osman Çorbali ◽  
Radames J. B. Cordero ◽  
Arturo Casadevall
Keyword(s):  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Glucuronic Acid ◽  
Pathogenic Fungus ◽  
Weak Acid ◽  
Phagocytic Cells ◽  
Acid Base ◽  
Content Type ◽  
Encapsulated Cells ◽  
Base Properties

ABSTRACTPhagosomal acidification is a critical cellular mechanism for the inhibition and killing of ingested microbes by phagocytic cells. The acidic environment activates microbicidal proteins and creates an unfavorable environment for the growth of many microbes. Consequently, numerous pathogenic microbes have developed strategies for countering phagosomal acidification through various mechanisms that include interference with phagosome maturation. The human-pathogenic fungusCryptococcus neoformansresides in acidic phagosomes after macrophage ingestion that actually provides a favorable environment for replication, since the fungus replicates faster at acidic pH. We hypothesized that the glucuronic acid residues in the capsular polysaccharide had the capacity to affect phagosomal acidity through their acid-base properties. A ratiometric fluorescence comparison of imaged phagosomes containingC. neoformansto phagosomes containing beads showed that the latter were significantly more acidic. Similarly, phagosomes containing nonencapsulatedC. neoformanscells were more acidic than those containing encapsulated cells. Acid-base titrations of isolatedC. neoformanspolysaccharide revealed that it behaves as a weak acid with maximal buffering capacity around pH 4 to 5. We interpret these results as indicating that the glucuronic acid residues in theC. neoformanscapsular polysaccharide can buffer phagosomal acidification. Interference with phagosomal acidification represents a new function for the cryptococcal capsule in virulence and suggests the importance of considering the acid-base properties of microbial capsules in the host-microbe interaction for other microbes with charged residues in their capsules.IMPORTANCECryptococcus neoformansis the causative agent of cryptococcosis, a devastating fungal disease that affects thousands of individuals worldwide. This fungus has the capacity to survive inside phagocytic cells, which contributes to persistence of infection and dissemination. One of the major antimicrobial mechanisms of host phagocytes is to acidify the phagosomal compartment after ingestion of microbes. This study shows that the capsule ofC. neoformanscan interfere with full phagosomal acidification by serving as a buffer.

scholarly journals Let it bud: an ultrastructural study of Cryptococcus neoformans surface during budding events

2020 ◽  
Author(s):  
Glauber R. de S. Araújo ◽  
Carolina de L. Alcantara ◽  
Noêmia Rodrigues ◽  
Wanderley de Souza ◽  
Bruno Pontes ◽  
...  
Keyword(s):  
Cell Wall ◽  
Plasma Membrane ◽  
Cryptococcus Neoformans ◽  
Drug Targets ◽  
Spatial Organization ◽  
Capsular Polysaccharide ◽  
Light And Electron Microscopy ◽  
Ultrastructural Organization ◽  
Membrane Structures ◽  
Daughter Cells

AbstractCryptococcus neoformans is a fungal pathogen that causes life-threatening infections in immunocompromised individuals. It is surrounded by three concentric structures that separate the cell from the extracellular space: the plasma membrane, the cell wall and the polysaccharide capsule. Although several studies have revealed the chemical composition of these structures, little is known about their ultrastructural organization and remodeling during C. neoformans budding event. Here, by combining the state-of-the-art in light and electron microscopy techniques we describe the morphological remodeling that occurs synergistically among the capsule, cell wall and plasma membrane during budding in C. neoformans. Our results show that the cell wall deforms to generate a specialized budding region at one of the cell’s poles. This region subsequently begins to break into layers that are slightly separated from each other and with thick tips. We also observe a reduction in density of the capsular polysaccharide around these specialized regions. Daughter cells present a distinct spatial organization, with polysaccharide fibers aligned in the direction of budding. In addition, to control the continuous openings between mother and daughter cells, the latter developed a strategy to shield themselves by forming multilamellar membrane structures in conjunction with their capsules. Together, our findings provide compelling ultrastructural evidence for a dynamic C. neoformans surface remodeling during budding and may have important implications for future studies exploring these remodeled specialized regions as drug-targets against cryptococcosis.

scholarly journals Antiidiotypic DNA vaccination induces serum bactericidal activity and protection against group B meningococci

2006 ◽  
Vol 203 (1) ◽  
pp. 111-118 ◽  
Author(s):  
Concetta Beninati ◽  
Angelina Midiri ◽  
Giuseppe Mancuso ◽  
Carmelo Biondo ◽  
Milena Arigò ◽  
...  
Keyword(s):  
Bactericidal Activity ◽  
Capsular Polysaccharide ◽  
Signal Sequence ◽  
Developed Countries ◽  
Single Chain ◽  
Serum Bactericidal Activity ◽  
Scfv Gene ◽  
Genes Encoding ◽  
Encapsulated Bacteria ◽  
Group B

No vaccine is available for preventing infections by serogroup B Neisseria meningitidis (MenB), which accounts for a major portion of meningococcal cases in developed countries, because of the poor immunogenicity of the capsular polysaccharide (CP) even after protein conjugation. We have previously induced anticapsular antibodies by immunization with a single chain variable fragment (scFv), which mimics a protective CP epitope. This surrogate antigen, however, was ineffective at inducing serum bactericidal activity, an accepted marker of protection in humans. Serum bactericidal activity was consistently achieved by immunizing mice with the scFv-encoding gene. Immunization with vectors without a secretory signal sequence before the scFv resulted in markedly higher bactericidal activity relative to those with such a sequence. The induced antibodies were capsule specific, as shown by complete inhibition of bactericidal activity by purified MenB CP and by resistance to killing of MenA or MenC. Moreover, these antibodies were predominantly of the IgG2a isotype, reflecting a T helper type 1 response. Administration of sera from scFv gene–vaccinated animals protected infant rats against MenB bacteremia. These data illustrate the potential of vaccination with genes encoding capsular mimics in providing protection against MenB and other encapsulated bacteria.

UDP Glucuronate Decarboxylase and Synthesis of Capsular Polysaccharide in Cryptococcus neoformans

1982 ◽  
Vol 152 (2) ◽  
pp. 932-934
Author(s):  
Eric S. Jacobson ◽  
William R. Payne
Keyword(s):  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Decarboxylase Activity

UDP glucuronate decarboxylase activity was comparable in encapsulated and non-encapsulated strains of Cryptococcus neoformans , required NAD ( K a = 0.2 mM), and was inhibited by NADH ( K i = 0.1 mM) and UDP xylose.

scholarly journals Monoclonal Antibodies Reactive with Immunorecessive Epitopes of Glucuronoxylomannan, the Major Capsular Polysaccharide of Cryptococcus neoformans

2003 ◽  
Vol 10 (5) ◽  
pp. 903-909 ◽  
Author(s):  
Suzanne Brandt ◽  
Peter Thorkildson ◽  
Thomas R. Kozel
Keyword(s):  
Monoclonal Antibody ◽  
Immune Response ◽  
Monoclonal Antibodies ◽  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Passive Immunization ◽  
The Other ◽  
Serotype A ◽  
Humoral Immune ◽  
New Information

ABSTRACT Cryptococcus neoformans is surrounded by an antiphagocytic capsule whose primary constituent is glucuronoxylomannan (GXM). An epitope shared by GXM serotypes A, B, C, and D is immunodominant when mice are immunized with serotype A GXM. In contrast, an epitope shared only by serotypes A and D is immunodominant when mice are immunized with serotype D. Hybridomas secreting antibodies reactive with subdominant epitopes were identified through a positive-negative screening procedure in which antibody-secreting colonies were characterized by reactivity with both the immunizing polysaccharide and GXMs from each of the four major serotypes. In this manner, a monoclonal antibody (MAb) that was reactive with an epitope shared only by serotypes A and B was identified and designated F10F5. Such an epitope has not been described previously. Immunization of mice with de-O-acetylated serotype A GXM generated a hybridoma that secreted an antibody, designated F12D2, that was reactive with all four serotypes. Unlike previously described monoclonal and polyclonal panspecific antibodies, the reactivity of MAb F12D2 was not altered by de-O-acetylation of GXM. These results indicate that there are at least two panspecific GXM epitopes; one epitope is dependent on O acetylation for antibody reactivity, and the other is independent of O acetylation. This study identifies strategies for production of MAbs that are reactive with subdominant or cryptic GXM epitopes and provides new information regarding the antigenic makeup and the humoral immune response to GXM, an essential virulence factor that is a target for active and passive immunization.

SPECIFIC DEGRADATION OF CRYPTOCOCCUS NEOFORMANS 3723 CAPSULAR POLYSACCHARIDE BY A MICROBIAL ENZYME: I. ISOLATION, PARTIAL PURIFICATION, AND PROPERTIES OF THE ENZYME

1961 ◽  
Vol 7 (1) ◽  
pp. 53-60 ◽  
Author(s):  
Hans H. Gadebusch ◽  
John D. Johnson
Keyword(s):  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Enzyme Reaction ◽  
Partial Purification ◽  
Intracellular Enzyme ◽  
Purification And Properties ◽  
Microbial Enzyme ◽  
Specific Degradation ◽  
Kinetics Of ◽  
Enzyme I

A partially purified intracellular enzyme from a species of Alcaligenes is described which specifically initiates the degradation of the he heteropolysaccharide of Cryptococcus neoformans, isolate 3723, The enzyme is active in the presence of serum and can be inactivated by heating at 45 °C for 10 minutes, The kinetics of the enzyme reaction are similar to those of other enzymes. Recovery and identification of the four known monosaccharides from enzymatic hydrolyzates suggest the presence of a number of other enzymes in these preparations.

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