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MBio Jun 2014The fungal pathogen Cryptococcus neoformans has several virulence factors, among which the most important is a polysaccharide capsule. The size of the capsule is...
UNLABELLED
The fungal pathogen Cryptococcus neoformans has several virulence factors, among which the most important is a polysaccharide capsule. The size of the capsule is variable and can increase significantly during infection. In this work, we investigated the relationship between capsular enlargement and the cell cycle. Capsule growth occurred primarily during the G1 phase. Real-time visualization of capsule growth demonstrated that this process occurred before the appearance of the bud and that capsule growth arrested during budding. Benomyl, which arrests the cells in G2/M, inhibited capsule growth, while sirolimus (rapamycin) addition, which induces G1 arrest, resulted in cells with larger capsule. Furthermore, we have characterized a mutant strain that lacks a putative G1/S cyclin. This mutant showed an increased capacity to enlarge the capsule, both in vivo (using Galleria mellonella as the host model) and in vitro. In the absence of Cln1, there was a significant increase in the production of extracellular vesicles. Proteomic assays suggest that in the cln1 mutant strain, there is an upregulation of the glyoxylate acid cycle. Besides, this cyclin mutant is avirulent at 37°C, which correlates with growth defects at this temperature in rich medium. In addition, the cln1 mutant showed lower intracellular replication rates in murine macrophages. We conclude that cell cycle regulatory elements are involved in the modulation of the expression of the main virulence factor in C. neoformans.
IMPORTANCE
Cryptococcus neoformans is a pathogenic fungus that has significant incidence worldwide. Its main virulence factor is a polysaccharide capsule that can increase in size during infection. In this work, we demonstrate that this process occurs in a specific phase of the cell cycle, in particular, in G1. In agreement, mutants that have an abnormal longer G1 phase show larger capsule sizes. We believe that our findings are relevant because they provide a link between capsule growth, cell cycle progression, and virulence in C. neoformans that reveals new aspects about the pathogenicity of this fungus. Moreover, our findings indicate that cell cycle elements could be used as antifungal targets in C. neoformans by affecting both the growth of the cells and the expression of the main virulence factor of this pathogenic yeast.
Topics: Animals; Bacterial Capsules; Cell Cycle; Cryptococcosis; Cryptococcus neoformans; Fungal Proteins; Humans; Macrophages; Mice; Moths; Virulence Factors
PubMed: 24939886
DOI: 10.1128/mBio.00945-14 -
Infection and Immunity Dec 2019Virulence genes are regulated by a complex regulatory network in Some of the regulators are global in nature and affect many downstream genes. MgrA is a multiple-gene...
Virulence genes are regulated by a complex regulatory network in Some of the regulators are global in nature and affect many downstream genes. MgrA is a multiple-gene regulator that has been shown to activate genes involved in capsule biosynthesis and repress surface protein genes. The goal of this study was to demonstrate the biological significance of MgrA regulation of capsule and surface proteins. We found that strain Becker possessed one fibronectin-binding protein, FnbA, and that FnbA was the predominant protein involved in invasion of nonphagocytic HeLa cells. By genetic analysis of strains with different amounts of capsule, we demonstrated that capsule impeded invasion of HeLa cells by masking the bacterial cell wall-anchored protein FnbA. Using variants with different levels of transcription, we further demonstrated that MgrA negatively impacted invasion by activating the genes involved in capsule biosynthesis and repressing the gene. Thus, we conclude that MgrA negatively impacts cell invasion of Becker by promoting capsule and repressing FnbA.
Topics: Adhesins, Bacterial; Bacterial Capsules; Cell Line, Tumor; Gene Expression Regulation, Bacterial; HeLa Cells; Humans; Polysaccharides, Bacterial; Staphylococcal Infections; Staphylococcus aureus; Virulence
PubMed: 31591167
DOI: 10.1128/IAI.00590-19 -
Scientific Reports Dec 2018Streptococcus pneumoniae's polysaccharide capsule is an important virulence factor; vaccine-induced immunity to specific capsular polysaccharide effectively prevents...
Streptococcus pneumoniae's polysaccharide capsule is an important virulence factor; vaccine-induced immunity to specific capsular polysaccharide effectively prevents disease. Serotype 1 S. pneumoniae is rarely found in healthy persons, but is highly invasive and a common cause of meningitis outbreaks and invasive disease outside of the United States. Here we show that genes for polysaccharide capsule similar to those expressed by pneumococci were commonly detected by polymerase chain reaction among upper respiratory tract samples from older US adults not carrying pneumococci. Serotype 1-specific genes were predominantly detected. In five oropharyngeal samples tested, serotype 1 gene belonging to S. mitis expressed capsules immunologically indistinct from pneumococcal capsules. Whole genome sequencing revealed three distinct S. mitis clones, each representing a cps1 operon highly similar to the pneumococcal cps1 reference operon. These findings raise important questions about the contribution of commensal streptococci to natural immunity against pneumococci, a leading cause of mortality worldwide.
Topics: Bacterial Capsules; Cross Reactions; Cross-Sectional Studies; Gene Expression; Gene Order; Genes, Bacterial; Humans; Phylogeny; Pneumococcal Infections; Serogroup; Streptococcus mitis; Streptococcus pneumoniae; Virulence Factors
PubMed: 30568178
DOI: 10.1038/s41598-018-35921-3 -
MBio Mar 2019The polysaccharide capsule is an essential virulence factor for in both community-acquired hypervirulent strains as well as health care-associated classical strains...
The polysaccharide capsule is an essential virulence factor for in both community-acquired hypervirulent strains as well as health care-associated classical strains that are posing significant challenges due to multidrug resistance. Capsule production is known to be transcriptionally regulated by a number of proteins, but very little is known about how these proteins collectively control capsule production. RmpA and RcsB are two known regulators of capsule gene expression, and RmpA is required for the hypermucoviscous (HMV) phenotype in hypervirulent strains. In this report, we confirmed that these regulators performed their anticipated functions in the ATCC 43816 derivative, KPPR1S: and mutants are HMV negative and have reduced capsule gene expression. We also identified a novel transcriptional regulator, RmpC, encoded by a gene near The Δ strain has reduced capsule gene expression but retains the HMV phenotype. We further showed that a regulatory cascade exists in which KvrA and KvrB, the recently characterized MarR-like regulators, and RcsB contribute to capsule regulation through regulation of the promoter and through additional mechanisms. In a murine pneumonia model, the regulator mutants have a range of colonization defects, suggesting that they regulate virulence factors in addition to capsule. Further testing of the and mutants revealed that they have distinct and overlapping functions and provide evidence that HMV is not dependent on overproduction of capsule. This distinction will facilitate a better understanding of HMV and how it contributes to enhanced virulence of hypervirulent strains. continues to be a substantial public health threat due to its ability to cause health care-associated and community-acquired infections combined with its ability to acquire antibiotic resistance. Novel therapeutics are needed to combat this pathogen, and a greater understanding of its virulence factors is required for the development of new drugs. A key virulence factor for is the capsule, and community-acquired hypervirulent strains produce a capsule that causes hypermucoidy. We report here a novel capsule regulator, RmpC, and provide evidence that capsule production and the hypermucoviscosity phenotype are distinct processes. Infection studies showing that this and other capsule regulator mutants have a range of phenotypes indicate that additional virulence factors are in their regulons. These results shed new light on the mechanisms controlling capsule production and introduce targets that may prove useful for the development of novel therapeutics for the treatment of this increasingly problematic pathogen.
Topics: Animals; Bacterial Capsules; Bacterial Proteins; Disease Models, Animal; Gene Deletion; Gene Expression Regulation, Bacterial; Gene Regulatory Networks; Klebsiella Infections; Klebsiella pneumoniae; Mice; Mutation; Phenotype; Pneumonia, Bacterial; Virulence; Viscosity
PubMed: 30914502
DOI: 10.1128/mBio.00089-19 -
Genes Aug 2020is the causative agent of a multitude of diseases, and further study into its pathogenies is vital. The pneumococcus is genetically malleable, and several tools are...
is the causative agent of a multitude of diseases, and further study into its pathogenies is vital. The pneumococcus is genetically malleable, and several tools are available to manipulate this pathogen. In this study, we attempted to utilize one such tool, the Sweet Janus cassette, to replace the capsule locus with other capsule loci in our strain background and found that the efficiency of allelic replacement was low and the number of revertant false-positive colonies was high. We determined that the capacity to recombine capsule varied by the initial isolated colony, suggesting that frequency of reversion is dependent on the bacterial clone. Alternative selection markers may further expand the application of Sweet Janus. We created novel cassettes that utilized chlorinated phenylalanine as an alternative counter-selection agent in conjunction with the Janus or Sweet Janus cassette, providing a new dual or triple selection marker. Moreover, we created cassettes that do not require engineered resistance in the background strain, including both single and dual selection markers. We were able to utilize all constructs in allelic replacement of the capsule loci. These novel constructs provide a new means for generating gene deletions in that expand experimental applications.
Topics: Alleles; Bacterial Capsules; Drug Resistance, Bacterial; Gene Conversion; Gene Deletion; Genetics, Microbial; Plasmids; Recombination, Genetic; Selection, Genetic; Streptococcus pneumoniae
PubMed: 32825523
DOI: 10.3390/genes11090965 -
Microbiology (Reading, England) Jul 2021Capsular polysaccharides (CPSs) protect bacteria from host and environmental factors. Many bacteria can express different CPSs and these CPSs are phase variable. For...
Capsular polysaccharides (CPSs) protect bacteria from host and environmental factors. Many bacteria can express different CPSs and these CPSs are phase variable. For example, ) is a prominent member of the human gut microbiome and expresses eight different capsular polysaccharides. Bacteria, including , have been shown to change their CPSs to adapt to various niches such as immune, bacteriophage, and antibiotic perturbations. However, there are limited tools to study CPSs and fundamental questions regarding phase variance, including if gut bacteria can express more than one capsule at the same time, remain unanswered. To better understand the roles of different CPSs, we generated a CPS1-specific antibody and a flow cytometry assay to detect CPS expression in individual bacteria in the gut microbiota. Using these novel tools, we report for the first time that bacteria can simultaneously express multiple CPSs. We also observed that nutrients such as glucose and salts had no effect on CPS expression. The ability to express multiple CPSs at the same time may provide bacteria with an adaptive advantage to thrive amid changing host and environmental conditions, especially in the intestine.
Topics: Bacterial Capsules; Bacteroides thetaiotaomicron; Gastrointestinal Microbiome; Humans; Polysaccharides, Bacterial
PubMed: 34224345
DOI: 10.1099/mic.0.001066 -
Clinical Infectious Diseases : An... Apr 2008The virulence of group A streptococci (GAS) correlates closely with expression of its surface antigen, M protein, and its hyaluronic acid capsule. In studies of human... (Review)
Review
The virulence of group A streptococci (GAS) correlates closely with expression of its surface antigen, M protein, and its hyaluronic acid capsule. In studies of human GAS infection, the former has received considerable attention. For several decades, however, systematic identification of encapsulated virulent strains by the mucoid colonies they produce has been neglected in clinical studies. In part, this may be due to the capsule's evanescent expression on artificial media, its repression during convalescent carriage, lack of expertise in recognizing its colonial morphology, and the growing tendency for clinical laboratories to eschew throat cultures in favor of rapid laboratory tests for group A polysaccharide. Older and more recent studies are reviewed here that emphasize the capsule's basic role in infection. We believe that it is time to refocus newer clinical studies and techniques on achieving early recognition of potentially dangerous, heavily encapsulated strains of GAS for which spread may be prevented.
Topics: Bacterial Capsules; History, 20th Century; History, 21st Century; Humans; Streptococcal Infections; Streptococcus pyogenes; Virulence; Virulence Factors
PubMed: 18444821
DOI: 10.1086/529194 -
Current Topics in Microbiology and... 2006Biofilm formation is a main virulence determinant in many bacterial infections. It significantly increases bacterial resistance to antibiotics and innate host defense.... (Review)
Review
Biofilm formation is a main virulence determinant in many bacterial infections. It significantly increases bacterial resistance to antibiotics and innate host defense. In general, the specific physiology of biofilms and the barrier function of the extracellular biofilm matrix determine resistance to antibacterials. However, resistance to antimicrobial peptides appears to be mainly based on the interaction with biofilm and capsule exopolymers. These polymers may work by electrostatic repulsion and/or sequestration of antibacterial substances. As biofilm polymers play an eminent role in biofilm structuring and resistance, their destruction by dedicated enzymes is a promising attempt to prevent colonization and develop treatment for biofilm-associated infections.
Topics: Adhesins, Bacterial; Animals; Antimicrobial Cationic Peptides; Bacterial Capsules; Bacterial Infections; Biofilms; Drug Resistance, Bacterial; Humans; Polysaccharides, Bacterial
PubMed: 16909925
DOI: 10.1007/3-540-29916-5_10 -
Research in Microbiology 1994
Review
Topics: Bacterial Capsules; Bacterial Proteins; Escherichia coli; Escherichia coli Proteins; Gene Expression Regulation, Bacterial; In Vitro Techniques; Rec A Recombinases; Transcription Factors
PubMed: 7855424
DOI: 10.1016/0923-2508(94)90086-8 -
Future Microbiology Sep 2009Streptococcus pneumoniae remains an important pathogen despite licensure of a seven-valent pneumococcal protein conjugate vaccine. As a result, serotyping strains... (Review)
Review
Streptococcus pneumoniae remains an important pathogen despite licensure of a seven-valent pneumococcal protein conjugate vaccine. As a result, serotyping strains remains of paramount importance to both assess the effectiveness of current vaccines and closely monitor for the emergence of nonvaccine strains. Given the limitations of the quellung reaction, both molecular and immunology-based serotyping methods have been pursued. Currently, the most promising assay combines an immunologic assay with multiplex PCR of serotype-specific genes. The key limitation with a molecular-based assay is the plasticity of the pneumococcus, as capsular transformation or point mutations could easily result in serotype misclassification. Based on the currently available techniques, a comprehensive immunology-based assay appears to be the most promising alternative to the quellung reaction. In the future, assays that utilize high-throughput sequencing technology and/or matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) could lead to a novel pneumococcal serotyping method.
Topics: Bacterial Capsules; Bacterial Typing Techniques; Genetic Variation; Genotype; Humans; Mass Spectrometry; Pneumococcal Infections; Polymerase Chain Reaction; Sequence Analysis, DNA; Serotyping; Streptococcus pneumoniae
PubMed: 19722839
DOI: 10.2217/fmb.09.58