Cryptococcus neoformans capsule regrowth experiments reveal dynamics of enlargement and architecture

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.

Carbohydrate based meningococcal vaccines: past and present overview

Neisseria meningitidis is a major cause of bacterial meningitidis worldwide. Children less than five years and adolescents are particularly affected. Nearly all invasive strains are surrounded by a polysaccharide capsule, based on which, 12 N. meningitidis serogroups are differentiated. Six of them, A, B, C, W, X, and Y, cause the vast majority of infections in humans. Mono- and multi-valent carbohydrate-based vaccines against meningococcal infections have been licensed or are currently in clinical development. In this mini-review, an overview of the past and present approaches for producing meningococcal glycoconjugate vaccines is provided.

Correlates of Nonrandom Patterns of Serotype Switching in Pneumococcus

Background Pneumococcus is a diverse pathogen, with >90 serotypes, each of which has a distinct polysaccharide capsule. Pneumococci can switch capsules, evading vaccine pressure. Certain serotype pairs are more likely to occur on the same genetic background as a results of serotype switching, but the drivers of these patterns are not well understood. Methods We used the PubMLST and Global Pneumococcal Sequencing Project databases to quantify the number of genetic lineages on which different serotype pairs occur together. We also quantified the genetic diversity of each serotype. Regression model were used to evaluate the relationship between shared polysaccharide components and the frequency of serotype co-occurrence and diversity. Results A number of serotype pairs occurred together on the same genetic lineage more commonly than expected. Co-occurrence of between-serogroup pairs was more common when both serotypes had glucose as a component of the capsule (and, potentially, glucuronic acid, any-N-acetylated sugar, or ribitol). Diversity also varied markedly by serotype and was associated with the presence of specific sugars in the capsule. Conclusions Certain pairs of serotypes are more likely to co-occur on the same genetic background. These patterns were correlated with shared polysaccharide components. This might reflect adaptation of strains to produce capsules with specific characteristics.

Uji pigmen dan deteksi kapsul polisakarida pada Staphylococcus aureus isolat asal broiler PIGMENT TEST AND DETECTION OF POLISAKARIDA CAPSULES ON Staphylococcus aureus ISOLAT BROILER

Staphylococcus aureus is a pathogenic bacterium causing disease in humans and animals. In broilers it cause septicemia, tendosinovitis, dermatitis, endocarditis, wound infections and arthritis and bumblefoot. The ability of Staphylococcus aureus to cause disease depends on the virulence factors they bear. The purpose of this research is to investigate the distribution of pigment production type and the existence of genes of polysaccharide capsule phenotype and genotype as determinant factor of virulence of bacteria on 15 isolates of Staphylococcus aureus from broiler. Pigment production test showed that 86.7% of isolates producing yellow pigment and 13.3% isolates produce orange pigment. The detection of polysaccharide capsules was phenotypically performed with hydrophobicity test with serum soft agar medium (SSA) showed 53.3% isolate grow compact and 46,7% isolate grown difuse and hydrophobicity test by salt aggregation test method (SAT) showed 66,7% hydrophil and 33.3% are hydrophobic. Genotype detection of polysaccharide capsule genes by polymerase chain reaction (PCR) showed 66.7% detected cap5 (amplicon 361bp) and 33.3% detected cap8 (173bp ampliole). The type of pigment production and the presence of polysaccharide capsules are some of the virulent factors in Staphylococcus aureus.

Correlates of non-random patterns of capsule switching in pneumococcus

ABSTRACTBackgroundPneumococcus is a diverse pathogen, with >90 serotypes, each of which has a distinct polysaccharide capsule. Pneumococci can switch capsules, evading vaccine pressure. Certain serotype pairs are more likely to switch, but the drivers of these patterns are not well understood.MethodsWe used the PubMLST and Global Pneumococcal Sequencing (GPS) databases to quantify the number of genetic lineages on which different serotype pairs co-occur. We also quantified the genetic diversity of each serotype. Regression models evaluated the relationship between shared polysaccharide structural components and the frequency of serotype switching and diversity.ResultsA number of serotype pairs co-occurred on the same genetic lineage more commonly than expected. Co-occurrence of between-serogroup pairs was more common when both serotypes had glucose as a component of the capsule (and, potentially, glucuronic acid). Diversity also varied markedly by serotype and was lower for serotypes with glucuronic acid in the capsule and higher for those with galactose in the capsule.ConclusionsCertain pairs of serotypes are more likely to occur on the same genetic background, and these patterns were correlated with shared polysaccharide components. This might indicate adaptation of strains to produce capsules with particular characteristics.

Putative novel cps loci in a large global collection of pneumococci

The pneumococcus produces a polysaccharide capsule, encoded by the cps locus, that provides protection against phagocytosis and determines serotype. Nearly 100 serotypes have been identified with new serotypes still being discovered, especially in previously understudied regions. Here we present an analysis of the cps loci of more than 18 000 genomes from the Global Pneumococcal Sequencing (GPS) project with the aim of identifying novel cps loci with the potential to produce previously unrecognized capsule structures. Serotypes were assigned using whole genome sequence data and 66 of the approximately 100 known serotypes were included in the final dataset. Closer examination of each serotype’s sequences identified nine putative novel cps loci (9X, 11X, 16X, 18X1, 18X2, 18X3, 29X, 33X and 36X) found in ~2.6 % of the genomes. The large number and global distribution of GPS genomes provided an unprecedented opportunity to identify novel cps loci and consider their phylogenetic and geographical distribution. Nine putative novel cps loci were identified and examples of each will undergo subsequent structural and immunological analysis.

Kingella kingaeSurface Polysaccharides Promote Resistance to Neutrophil Phagocytosis and Killing

ABSTRACTBacterial pathogens have evolved strategies that enable them to evade neutrophil-mediated killing. The Gram-negative coccobacillusKingella kingaeis an emerging pediatric pathogen and is increasingly recognized as a common etiological agent of osteoarticular infections and bacteremia in young children.K. kingaeproduces a polysaccharide capsule and an exopolysaccharide, both of which are important for protection against complement-mediated lysis and are required for full virulence in an infant rat model of infection. In this study, we examined the role of theK. kingaepolysaccharide capsule and exopolysaccharide in protection against neutrophil killing. In experiments with primary human neutrophils, we found that the capsule interfered with the neutrophil oxidative burst response and prevented neutrophil binding ofK. kingaebut had no effect on neutrophil internalization ofK. kingae. In contrast, the exopolysaccharide resisted the bactericidal effects of antimicrobial peptides and efficiently blocked neutrophil phagocytosis ofK. kingae. This work demonstrates that theK. kingaepolysaccharide capsule and exopolysaccharide promote evasion of neutrophil-mediated killing through distinct yet complementary mechanisms, providing additional support for theK. kingaesurface polysaccharides as potential vaccine antigens. In addition, these studies highlight a novel interplay between a bacterial capsule and a bacterial exopolysaccharide and reveal new properties for a bacterial exopolysaccharide, with potential applicability to other bacterial pathogens.IMPORTANCEKingella kingaeis a Gram-negative commensal in the oropharynx and represents a leading cause of joint and bone infections in young children. The mechanisms by whichK. kingaeevades host innate immunity during pathogenesis of disease remain poorly understood. In this study, we established that theK. kingaepolysaccharide capsule and exopolysaccharide function independently to protectK. kingaeagainst reactive oxygen species (ROS) production, neutrophil phagocytosis, and antimicrobial peptides. These results demonstrate the intricacies ofK. kingaeinnate immune evasion and provide valuable information that may facilitate development of a polysaccharide-based vaccine againstK. kingae.