scholarly journals Capsular polysaccharide synthesis in Streptococcus pneumoniae serotype 14: molecular analysis of the complete cps locus and identification of genes encoding glycosyltransferases required for the biosynthesis of the tetrasaccharide subunit

1997 ◽  
Vol 26 (1) ◽  
pp. 197-208 ◽  
Author(s):  
Marc A. B. Kolkman ◽  
Warren Wakarchuk ◽  
Piet J. M. Nuijten ◽  
Bernard A. M. Van Der Zeijst
Keyword(s):  
Streptococcus Pneumoniae ◽  
Molecular Analysis ◽  
Capsular Polysaccharide ◽  
Polysaccharide Synthesis ◽  
Genes Encoding

scholarly journals Parallel β-Helix Proteins Required for Accurate Capsule Polysaccharide Synthesis and Virulence in the Yeast Cryptococcus neoformans

2007 ◽  
Vol 6 (4) ◽  
pp. 630-640 ◽  
Author(s):  
Oliver W. Liu ◽  
Mark J. S. Kelly ◽  
Eric D. Chow ◽  
Hiten D. Madhani
Keyword(s):  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Resonance Spectroscopy ◽  
Wild Type ◽  
Repeat Proteins ◽  
Knockout Mutants ◽  
Polysaccharide Synthesis ◽  
Mutant Strains ◽  
Genes Encoding ◽  
Opportunistic Fungal Pathogen

ABSTRACT The principal capsular polysaccharide of the opportunistic fungal pathogen Cryptococcus neoformans consists of an α-1,3-linked mannose backbone decorated with a repeating pattern of glucuronyl and xylosyl side groups. This structure is critical for virulence, yet little is known about how the polymer, called glucuronoxylomannan (GXM), is faithfully synthesized and assembled. We have generated deletions in two genes encoding predicted parallel β-helix repeat proteins, which we have designated PBX1 and PBX2. Deletion of either gene results in a dry-colony morphology, clumpy cells, and decreased capsule integrity. Two-dimensional nuclear magnetic resonance spectroscopy of purified GXM from the mutants indicated that both the wild-type GXM structure and novel, aberrant linkages were present. Carbohydrate composition and linkage analysis determined that these aberrant structures are correlated with the incorporation of terminal glucose residues that are not found in wild-type capsule polysaccharide. We conclude that Pbx1 and Pbx2 are required for the fidelity of GXM synthesis and may be involved in editing incorrectly added glucose residues. PBX1 and PBX2 knockout mutants showed severely attenuated virulence in a murine inhalation model of cryptococcosis. Unlike acapsular strains, these mutant strains induced delayed symptoms of cryptococcosis, though the infected animals eventually contained the infection and recovered.

scholarly journals Genetic Basis for the Structural Difference between Streptococcus pneumoniae Serotype 15B and 15C Capsular Polysaccharides

2003 ◽  
Vol 71 (11) ◽  
pp. 6192-6198 ◽  
Author(s):  
Saskia van Selm ◽  
Lisette M. van Cann ◽  
Marc A. B. Kolkman ◽  
Bernard A. M. van der Zeijst ◽  
Jos P. M. van Putten
Keyword(s):  
Streptococcus Pneumoniae ◽  
Genetic Analysis ◽  
Genetic Basis ◽  
Capsular Polysaccharide ◽  
Structural Difference ◽  
Open Reading Frames ◽  
Structural Level ◽  
Reading Frame ◽  
Polysaccharide Synthesis ◽  
Short Tandem

ABSTRACT In a search for the genetic basis for the structural difference between the related Streptococcus pneumoniae capsular serotypes 15B and 15C and for the reported reversible switching between these serotypes, the corresponding capsular polysaccharide synthesis (cps) loci were investigated by keeping in mind that at the structural level, the capsules differ only in O acetylation. The cps locus of a serotype 15B strain was identified, partially PCR amplified with primers based on the related serotype 14 sequence, and sequenced. Sequence analysis revealed, among other open reading frames, an intact open reading frame (designated cps15bM) whose product, at the protein level, exhibited characteristics of previously identified acetyltransferases. Genetic analysis of the corresponding region in a serotype15C strain indicated that the same gene was present but had a premature stop in translation. Closer analysis indicated that the serotype 15B gene contained a short tandem TA repeat consisting of eight TA units. In serotype 15C, this gene contained nine TA units that resulted in a frameshift and a truncated product. Genetic analysis of 17 serotype 15B and 15C clinical isolates revealed a perfect correlation between the serotype and the length of the short tandem repeat in the putative O-acetyltransferase gene. The number of TA repeating units varied between seven and nine in the various isolates. Together, the data strongly suggest that the structural difference between serotypes 15B and 15C is based on variation in the short tandem TA repeat in the O-acetyltransferase gene and that the transition between serotypes is due to slipped-strand mispairing with deletion or insertion of TA units in the cps15bM gene.

scholarly journals Construction of Otherwise Isogenic Serotype 6B, 7F, 14, and 19F Capsular Variants of Streptococcus pneumoniae Strain TIGR4

2003 ◽  
Vol 69 (12) ◽  
pp. 7364-7370 ◽  
Author(s):  
Krzysztof Trzcinski ◽  
Claudette M. Thompson ◽  
Marc Lipsitch
Keyword(s):  
Streptococcus Pneumoniae ◽  
Capsular Polysaccharide ◽  
Rflp Analysis ◽  
Capsular Type ◽  
Chromosomal Dna ◽  
Subsequent Transformation ◽  
Polysaccharide Synthesis ◽  
Quellung Reaction ◽  
Flanking Regions

ABSTRACT The polysaccharide capsule is the primary virulence factor in Streptococcus pneumoniae. There are at least 90 serotypes of S. pneumoniae, identified based on the immunogenicity of different capsular sugars. The aim of this study was to construct pneumococcal strains that are isogenic except for capsular type. Serotype 4 strain TIGR4 was rendered unencapsulated by recombinational replacement of the capsular polysaccharide synthesis (cps) locus with the bicistronic Janus cassette (C. K. Sung, J. P. Claverys, and D. A. Morrison, Appl. Environ. Microbiol. 67:5190-5196, 2001). In subsequent transformation with chromosomal DNA, the cassette was replaced by the cps locus derived from a strain of a different serotype, either 6B, 7F, 14, or 19F. To minimize the risk of uncontrolled recombinational replacements in loci other than cps, the TIGRcps::Janus strain was“ backcross” transformed three times with chromosomal DNA of subsequently constructed capsular type transformants. Capsular serotypes were confirmed in all new capsule variants by the Quellung reaction. Restriction fragment length polymorphism (RFLP) analysis of the cps locus confirmed the integrity of the cps region transformed into the TIGR strain, and RFLP of the flanking regions confirmed their identities with the corresponding regions of the recipient. Transformants had in vitro growth rates greater than or equal to that of TIGR4. All four strains were able to colonize C57BL/6 mice (female, 6 weeks old) for at least 7 days when mice were intranasally inoculated with 6 × 106 to 8 × 106 CFU. The constructed capsular variants of TIGR4 are suitable for use in studies on the role of S. pneumoniae capsular polysaccharide in immunity, colonization, and pathogenesis.

scholarly journals SPD_1495 Contributes to Capsular Polysaccharide Synthesis and Virulence in Streptococcus pneumoniae

mSystems ◽  
2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Yun-Dan Zheng ◽  
Ying Pan ◽  
Ke He ◽  
Nan Li ◽  
Donghong Yang ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Capsular Polysaccharide ◽  
Deep Understanding ◽  
Bacterial Virulence ◽  
Mobility Shift ◽  
Synthesis Mechanism ◽  
Content Type ◽  
Polysaccharide Synthesis ◽  
Key Factor ◽  
Mobility Shift Assay

ABSTRACT Streptococcus pneumoniae, a Gram-positive human pathogen, causes a series of serious diseases in humans. SPD_1495 from S. pneumoniae is annotated as a hypothetical ABC sugar-binding protein in the NCBI database, but there are few reports on detailed biological functions of SPD_1495. To fully study the influence of SPD_1495 on bacterial virulence in S. pneumoniae, we constructed a deletion mutant (D39Δspd1495) and an overexpressing strain (D39spd1495+). Comparative analysis of iTRAQ-based quantitative proteomic data of the wild-type D39 strain (D39-WT) and D39Δspd1495 showed that several differentially expressed proteins that participate in capsular polysaccharide synthesis, such as Cps2M, Cps2C, Cps2L, Cps2T, Cps2E, and Cps2D, were markedly upregulated in D39Δspd1495. Subsequent transmission electron microscopy and uronic acid detection assay confirmed that capsular polysaccharide synthesis was enhanced in D39Δspd1495 compared to that in D39-WT. Moreover, knockout of spd1495 resulted in increased capsular polysaccharide synthesis, as well as increased bacterial virulence, as confirmed by the animal study. Through a coimmunoprecipitation assay, surface plasmon resonance, and electrophoretic mobility shift assay, we found that SPD_1495 negatively regulated cps promoter expression by interacting with phosphorylated ComE, a negative transcriptional regulator for capsular polysaccharide formation. Overall, this study suggested that SPD_1495 negatively regulates capsular polysaccharide formation and thereby enhances bacterial virulence in the host. These findings also provide valuable insights into understanding the biology of this clinically important bacterium. IMPORTANCE Capsular polysaccharide is a key factor underlying the virulence of Streptococcus pneumoniae in human diseases. Thus, a deep understanding of capsular polysaccharide synthesis is essential for uncovering the pathogenesis of S. pneumoniae infection. In this study, we show that protein SPD_1495 interacts with phosphorylated ComE to negatively regulate the formation of capsular polysaccharide. Deletion of spd1495 increased capsular polysaccharide synthesis and thereby enhanced bacterial virulence. These findings further reveal the synthesis mechanism of capsular polysaccharide and provide new insight into the biology of this clinically important bacterium.

scholarly journals Antibodies to Streptococcus pneumoniae Capsular Polysaccharide Enhance Pneumococcal Quorum Sensing

mBio ◽  
2011 ◽  
Vol 2 (5) ◽  
Author(s):  
Masahide Yano ◽  
Shruti Gohil ◽  
J. Robert Coleman ◽  
Catherine Manix ◽  
Liise-anne Pirofski
Keyword(s):  
Monoclonal Antibodies ◽  
Streptococcus Pneumoniae ◽  
Quorum Sensing ◽  
Direct Effect ◽  
Pneumococcal Disease ◽  
Effector Cell ◽  
Capsular Polysaccharide ◽  
Genetic Exchange ◽  
Content Type ◽  
Specific Antibodies

ABSTRACTThe use of pneumococcal capsular polysaccharide (PPS)-based vaccines has resulted in a substantial reduction in invasive pneumococcal disease. However, much remains to be learned about vaccine-mediated immunity, as seven-valent PPS-protein conjugate vaccine use in children has been associated with nonvaccine serotype replacement and 23-valent vaccine use in adults has not prevented pneumococcal pneumonia. In this report, we demonstrate that certain PPS-specific monoclonal antibodies (MAbs) enhance the transformation frequency of two differentStreptococcus pneumoniaeserotypes. This phenomenon was mediated by PPS-specific MAbs that agglutinate but do not promote opsonic effector cell killing of the homologous serotypeinvitro. Compared to the autoinducer, competence-stimulating peptide (CSP) alone, transcriptional profiling of pneumococcal gene expression after incubation with CSP and one such MAb to the PPS of serotype 3 revealed changes in the expression of competence (com)-related and bacteriocin-like peptide (blp) genes involved in pneumococcal quorum sensing. This MAb was also found to induce a nearly 2-fold increase in CSP2-mediated bacterial killing or fratricide. These observations reveal a novel, direct effect of PPS-binding MAbs on pneumococcal biology that has important implications for antibody immunity to pneumococcus in the pneumococcal vaccine era. Taken together, our data suggest heretofore unsuspected mechanisms by which PPS-specific antibodies could affect genetic exchange and bacterial viability in the absence of host cells.IMPORTANCECurrent thought holds that pneumococcal capsular polysaccharide (PPS)-binding antibodies protect against pneumococcus by inducing effector cell opsonic killing of the homologous serotype. While such antibodies are an important part of how pneumococcal vaccines protect against pneumococcal disease, PPS-specific antibodies that do not exhibit this activity but are highly protective against pneumococcus in mice have been identified. This article examines the effect of nonopsonic PPS-specific monoclonal antibodies (MAbs) on the biology ofStreptococcus pneumoniae. The results showed that in the presence of a competence-stimulating peptide (CSP), such MAbs increase the frequency of pneumococcal transformation. Further studies with one such MAb showed that it altered the expression of genes involved in quorum sensing and increased competence-induced killing or fratricide. These findings reveal a novel, previously unsuspected mechanism by which certain PPS-specific antibodies exert a direct effect on pneumococcal biology that has broad implications for bacterial clearance, genetic exchange, and antibody immunity to pneumococcus.

scholarly journals Genetic Relatedness of the Streptococcus pneumoniae Capsular Biosynthetic Loci

2007 ◽  
Vol 189 (21) ◽  
pp. 7841-7855 ◽  
Author(s):  
Angeliki Mavroidi ◽  
David M. Aanensen ◽  
Daniel Godoy ◽  
Ian C. Skovsted ◽  
Margit S. Kaltoft ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Genetic Relatedness ◽  
Pneumococcal Serotypes ◽  
Major Cluster ◽  
Genes Encoding ◽  
Pneumococcal Serotype ◽  
Streptococcal Polysaccharide ◽  
Single Cluster ◽  
Dependent Pathway ◽  
Sequence Similarities

ABSTRACT Streptococcus pneumoniae (the pneumococcus) produces 1 of 91 capsular polysaccharides (CPS) that define the serotype. The cps loci of 88 pneumococcal serotypes whose CPS is synthesized by the Wzy-dependent pathway were compared with each other and with additional streptococcal polysaccharide biosynthetic loci and were clustered according to the proportion of shared homology groups (HGs), weighted for the sequence similarities between the genes encoding the shared HGs. The cps loci of the 88 pneumococcal serotypes were distributed into eight major clusters and 21 subclusters. All serotypes within the same serogroup fell into the same major cluster, but in six cases, serotypes within the same serogroup were in different subclusters and, conversely, nine subclusters included completely different serotypes. The closely related cps loci within a subcluster were compared to the known CPS structures to relate gene content to structure. The Streptococcus oralis and Streptococcus mitis polysaccharide biosynthetic loci clustered within the pneumococcal cps loci and were in a subcluster that also included the cps locus of pneumococcal serotype 21, whereas the Streptococcus agalactiae cps loci formed a single cluster that was not closely related to any of the pneumococcal cps clusters.

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