scholarly journals Identification of colonisation and virulence determinants of Streptococcus pneumoniae via experimental evolution in mouse infection models

2020 ◽  
Author(s):  
Angharad E Green ◽  
Deborah Howarth ◽  
Chrispin Chaguza ◽  
Haley Echlin ◽  
R Frèdi Langendonk ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Experimental Evolution ◽  
Parallel Evolution ◽  
Serial Passage ◽  
Invasive Disease ◽  
Virulence Determinants ◽  
Upper Respiratory Tract ◽  
Single Nucleotide ◽  
Single Nucleotide Change ◽  
Infection Models

AbstractStreptococcus pneumoniae is a commensal of the human nasopharynx and a major cause of respiratory and invasive disease. Pneumococcus stimulates upper respiratory tract inflammation that promotes shedding from mucosal surfaces and transmission to new hosts. Colonisation and transmission are partially antagonistic processes. Adhesion to surfaces and evasion of host responses favours the former, whilst detachment, promoted by inflammation, is necessary for the latter. We sought to determine how adaptation and evolution of pneumococcus within its nasopharyngeal niche might progress when selective pressures associated with transmission were removed. This was achieved by serial passage of pneumococci in mouse models of nasopharyngeal carriage, manually transferring bacteria between mice. To assess the role of host environmental factors on pneumococcal evolution, we also performed analogous experimental evolution in a mouse pneumonia model, passaging pneumococci through lungs. Nasopharynx-passaged pneumococci became more effective colonisers, whilst those evolved within lungs showed reduced virulence. We observed selection of mutations in genes associated with cell wall biogenesis and metabolism in both nasopharynx and lung lineages, but identified prominent examples of parallel evolution that were niche specific. We focussed on gpsA, a gene in which the same single nucleotide polymorphism arose in two independently evolved nasopharynx-passaged lineages. We identified a single nucleotide change conferring resistance to oxidative stress and enhanced nasopharyngeal colonisation potential. We show that gpsA is also a frequent target of mutation during human colonisation. These findings highlight the role played by the host environment in determining trajectories of bacterial evolution and the potential of experimental evolution in animal infection models for identification of novel pathogen virulence and colonisation factors.

scholarly journals Pneumococcal Colonization and Virulence Factors Identified Via Experimental Evolution in Infection Models

2021 ◽  
Author(s):  
Angharad E Green ◽  
Deborah Howarth ◽  
Chrispin Chaguza ◽  
Haley Echlin ◽  
R Frèdi Langendonk ◽  
...  
Keyword(s):  
Experimental Evolution ◽  
Parallel Evolution ◽  
Experimental System ◽  
Serial Passage ◽  
Invasive Disease ◽  
Vaccine Antigen ◽  
Nasopharyngeal Carriage ◽  
Single Nucleotide ◽  
Single Nucleotide Change ◽  
Infection Models

Abstract Streptococcus pneumoniae is a commensal of the human nasopharynx and a major cause of respiratory and invasive disease. We examined adaptation and evolution of pneumococcus, within nasopharynx and lungs, in an experimental system where the selective pressures associated with transmission were removed. This was achieved by serial passage of pneumococci, separately, in mouse models of nasopharyngeal carriage or pneumonia. Passaged pneumococci became more effective colonizers of the respiratory tract and we observed several examples of potential parallel evolution. The cell wall-modifying glycosyltransferase LafA was under strong selection during lung passage, whereas the surface expressed pneumococcal vaccine antigen gene pvaA and the glycerol-3-phosphate dehydrogenase gene gpsA were frequent targets of mutation in nasopharynx-passaged pneumococci. These mutations were not identified in pneumococci that were separately evolved by serial passage on laboratory agar. We focused on gpsA, in which the same single nucleotide polymorphism arose in two independently evolved nasopharynx-passaged lineages. We describe a new role for this gene in nasopharyngeal carriage and show that the identified single nucleotide change confers resistance to oxidative stress and enhanced nasopharyngeal colonization potential. We demonstrate that polymorphisms in gpsA arise and are retained during human colonization. These findings highlight how within-host environmental conditions can determine trajectories of bacterial evolution. Relative invasiveness or attack rate of pneumococcal lineages may be defined by genes that make niche-specific contributions to bacterial fitness. Experimental evolution in animal infection models is a powerful tool to investigate the relative roles played by pathogen virulence and colonization factors within different host niches.

scholarly journals Variation in the Presence of Neuraminidase Genes among Streptococcus pneumoniae Isolates with Identical Sequence Types

2006 ◽  
Vol 74 (6) ◽  
pp. 3360-3365 ◽  
Author(s):  
Melinda M. Pettigrew ◽  
Kristopher P. Fennie ◽  
Matthew P. York ◽  
Janeen Daniels ◽  
Faryal Ghaffar
Keyword(s):  
Streptococcus Pneumoniae ◽  
Young Children ◽  
Invasive Disease ◽  
Virulence Determinants ◽  
Upper Respiratory Tract ◽  
Capsule Type ◽  
Virulence Potential ◽  
The Relationship ◽  
Sequence Types ◽  
Specific Virulence

ABSTRACT Streptococcus pneumoniae frequently colonizes the upper respiratory tract of young children and is an important cause of otitis media and invasive disease. Carriage is more common than disease, yet the genetic factors that predispose a given clone for disease are not known. The relationship between capsule type, genetic background, and virulence is complex, and important questions remain regarding how pneumococcal clones differ in their ability to cause disease. Pneumococcal neuraminidase cleaves sialic acid-containing substrates and is thought to be important for pneumococcal virulence. We describe the distribution of multilocus sequence types (ST), capsule type, and neuraminidase genes among 342 carriage, middle ear, blood, and cerebrospinal fluid (CSF) pneumococcal strains from young children. We found 149 STs among our S. pneumoniae isolates. nanA was present in all strains, while nanB and nanC were present in 96% and 51% of isolates, respectively. The distribution of nanC varied among the strain collections from different tissue sources (P = 0.03). The prevalence of nanC was 1.41 (95% confidence interval, 1.11, 1.79) times higher among CSF isolates than among carriage isolates. We identified isolates of the same ST that differed in the presence of nanB and nanC. These studies demonstrate that virulence determinants, other than capsule loci, vary among strains of identical ST. Our studies suggest that the presence of nanC may be important for tissue-specific virulence. Studies that both incorporate MLST and take into account additional virulence determinants will provide a greater understanding of the pneumococcal virulence potential.

scholarly journals Mutations in PneumococcalcpsEGenerated viaIn VitroSerial Passaging Reveal a Potential Mechanism of Reduced Encapsulation Utilized by a Conjunctival Isolate

2015 ◽  
Vol 197 (10) ◽  
pp. 1781-1791 ◽  
Author(s):  
Mara G. Shainheit ◽  
Michael D. Valentino ◽  
Michael S. Gilmore ◽  
Andrew Camilli
Keyword(s):  
Streptococcus Pneumoniae ◽  
Invasive Disease ◽  
Adaptive Mutation ◽  
Specific Gene ◽  
High Dose ◽  
Wild Type ◽  
Single Nucleotide ◽  
Content Type ◽  
Sepsis Model

ABSTRACTThe polysaccharide capsule ofStreptococcus pneumoniaeis required for nasopharyngeal colonization and for invasive disease in the lungs, blood, and meninges. In contrast, the vast majority of conjunctival isolates are acapsular. The first serotype-specific gene in the capsule operon,cpsE, encodes the initiating glycosyltransferase and is one of the few serotype-specific genes that can tolerate null mutations. This report characterizes a spontaneously arising TIGR4 mutant exhibiting a reduced capsule, caused by a 6-nucleotide duplication incpsEwhich results in duplication of Ala and Ile at positions 45 and 46. This strain (AI45dup) possessed more exposed phosphorylcholine and was hypersusceptible to C3 complement deposition compared to the wild type. Accordingly, the mutant was significantly better at forming abiotic biofilms and binding epithelial cellsin vitrobut was avirulent in a sepsis model.In vitroserial passaging of the wild-type strain failed to reproduce the AI45dup mutation but instead led to a variety of mutants with reduced capsule harboring single nucleotide polymorphisms (SNPs) incpsE. A single passage in the sepsis model after high-dose inoculation readily yielded revertants of AI45dup with restored wild-type capsule level, but the majority of SNP alleles ofcpsEcould not revert, suppress, or bypass. Analysis ofcpsEin conjunctival isolates revealed a strain with a single missense mutation at amino acid position 377, which was responsible for reduced encapsulation. This study supports the hypothesis that spontaneous, nonreverting mutations incpsEserve as a form of adaptive mutation by providing a selective advantage toS. pneumoniaein niches where expression of capsule is detrimental.IMPORTANCEWhile the capsule ofStreptococcus pneumoniaeis required for colonization and invasive disease, most conjunctival isolates are acapsular by virtue of deletion of the entire capsular operon. We show that spontaneous acapsular mutants isolatedin vitroharbor mostly nonrevertible single nucleotide polymorphism (SNP) null mutations incpsE, encoding the initiating glycosyltransferase. From a small collection of acapsular conjunctival isolates, we identified one strain with a complete capsular operon but containing a SNP incpsEthat we show is responsible for the acapsular phenotype. We propose that acapsular conjunctival isolates may arise initially from such nonreverting SNP null mutations incpsE, which can be followed later by deletion of portions or all of thecpsoperon.

scholarly journals Contributions of Pneumolysin, Pneumococcal Surface Protein A (PspA), and PspC to Pathogenicity of Streptococcus pneumoniae D39 in a Mouse Model

2007 ◽  
Vol 75 (4) ◽  
pp. 1843-1851 ◽  
Author(s):  
Abiodun D. Ogunniyi ◽  
Kim S. LeMessurier ◽  
Rikki M. A. Graham ◽  
James M. Watt ◽  
David E. Briles ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Systemic Disease ◽  
Protein A ◽  
Virulence Gene ◽  
Surface Protein ◽  
Invasive Disease ◽  
Upper Respiratory Tract ◽  
Virulence Proteins ◽  
Pneumococcal Surface Protein A ◽  
Genes Encoding

ABSTRACTSuccessful colonization of the upper respiratory tract byStreptococcus pneumoniaeis an essential first step in the pathogenesis of pneumococcal disease. However, the bacterial and host factors that provoke the progression from asymptomatic colonization to invasive disease are yet to be fully defined. In this study, we investigated the effects of single and combined mutations in genes encoding pneumolysin (Ply), pneumococcal surface protein A (PspA), and pneumococcal surface protein C (PspC, also known as choline-binding protein A) on the pathogenicity ofStreptococcus pneumoniaeserotype 2 (D39) in mice. Following intranasal challenge with D39, stable colonization of the nasopharynx was maintained over a 7-day period at a level of approximately 105bacteria per mouse. The abilities of the mutant deficient in PspA to colonize the nasopharynx and to cause lung infection and bacteremia were significantly reduced. Likewise, the PspC mutant and, to a lesser extent, the Ply mutant also had reduced abilities to colonize the nasopharynx. As expected, the double mutants colonized less well than the parent to various degrees and had difficulty translocating to the lungs and blood. A significant additive attenuation was observed for the double and triple mutants in pneumonia and systemic disease models. Surprisingly, the colonization profile of the derivative lacking all three proteins was similar to that of the wild type, indicating virulence gene compensation. These findings further demonstrate that the mechanism of pneumococcal pathogenesis is highly complex and multifactorial but ascribes a role for each of these virulence proteins, alone or in combination, in the process.

scholarly journals The Underlying Structure of Adaptation under Strong Selection in 12 Experimental Yeast Populations

2014 ◽  
Vol 13 (9) ◽  
pp. 1200-1206 ◽  
Author(s):  
Linda M. Kohn ◽  
James B. Anderson
Keyword(s):  
Sexual Reproduction ◽  
Asexual Reproduction ◽  
Experimental Evolution ◽  
Parallel Evolution ◽  
High Salt ◽  
The Other ◽  
Underlying Structure ◽  
Single Nucleotide ◽  
Strong Selection ◽  
Low Glucose

ABSTRACTThe aims of this study were to determine (i) whether adaptation under strong selection occurred through mutations in a narrow target of one or a few nucleotide sites or a broad target of numerous sites and (ii) whether the programs of adaptation previously observed from three experimental populations were unique or shared among populations that underwent parallel evolution. We used archived population samples from a previous study, representing 500 generations of experimental evolution in 12 populations under strong selection, 6 populations in a high-salt environment and 6 populations in a low-glucose environment. Each set of six populations included four with sexual reproduction and two with exclusively asexual reproduction. Populations were sampled as resequenced genomes of 115 individuals and as bulk samples from which frequencies of mutant alleles were estimated. In a high-salt environment, a broad target of 11 mutations within the proton exporter,PMA1, was observed among the six populations, in addition to expansions of theENAgene cluster. This pattern was shared among populations that underwent parallel evolution. In a low-glucose environment, two programs of adaptation were observed. The originally observed pattern of mutation inMDS3/MKT1in population M8 was a narrow target of a single nucleotide, unique to this population. Among the other five populations, the three mutations were shared in a broad target, sensing/signaling genesRAS1andRAS2. RAS1/RAS2mutations were not observed in the high-salt populations;PMA1mutations were observed only in a high-salt environment.

Anatomical site-specific carbohydrate availability impacts Streptococcus pneumoniae virulence and fitness during colonization and disease

2021 ◽  
Author(s):  
Hansol Im ◽  
Katherine L. Kruckow ◽  
Adonis D’Mello ◽  
Feroze Ganaie ◽  
Eriel Martinez ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Disease Model ◽  
Invasive Disease ◽  
Relative Fitness ◽  
Anatomical Site ◽  
Virulence Determinants ◽  
Acid Fermentation ◽  
Altered Expression ◽  
Site Specific ◽  
Carbohydrate Availability

Streptococcus pneumoniae ( Spn ) colonizes the nasopharynx asymptomatically but can also cause severe life-threatening disease. Importantly, stark differences in carbohydrate availability exist between the nasopharynx and invasive disease sites, such as the bloodstream, which most likely impact Spn ’s behavior. Herein, using chemically-defined media (CDM) supplemented with physiological levels of carbohydrates, we examined how anatomical-site specific carbohydrate availability impacted Spn physiology and virulence. Spn grown in CDM modeling the nasopharynx (CDM-N) had reduced metabolic activity, slower growth rate, demonstrated mixed acid fermentation with marked H 2 O 2 production, and were in a carbon-catabolite repression (CCR)-derepressed state versus Spn grown in CDM modeling blood (CDM-B). Using RNA-seq, we determined the transcriptome for Spn WT and its isogenic CCR deficient mutant in CDM-N and CDM-B. Genes with altered expression as a result of changes in carbohydrate availability or catabolite control protein deficiency, respectively, were primarily involved in carbohydrate metabolism, but also encoded for established virulence determinants such polysaccharide capsule and surface adhesins. We confirmed that anatomical site-specific carbohydrate availability directly influenced established Spn virulence traits. Spn grown in CDM-B formed shorter chains, produced more capsule, were less adhesive, and were more resistant to macrophage killing in an opsonophagocytosis assay. Moreover, growth of Spn in CDM-N or CDM-B prior to the challenge of mice impacted relative fitness in a colonization and invasive disease model, respectively. Thus, anatomical site-specific carbohydrate availability alters Spn physiology and virulence, in turn promoting anatomical-site specific fitness.

scholarly journals Live Attenuated Streptococcus pneumoniae Strains Induce Serotype-Independent Mucosal and Systemic Protection in Mice

2007 ◽  
Vol 75 (5) ◽  
pp. 2469-2475 ◽  
Author(s):  
Aoife M. Roche ◽  
Samantha J. King ◽  
Jeffrey N. Weiser
Keyword(s):  
Streptococcus Pneumoniae ◽  
Pneumococcal Disease ◽  
Capsular Polysaccharide ◽  
Protein A ◽  
Surface Protein ◽  
High Dose ◽  
Virulence Determinants ◽  
Upper Respiratory Tract ◽  
Vaccine Strategy ◽  
Live Attenuated Vaccine

ABSTRACT Streptococcus pneumoniae is an important human pathogen causing both mucosal (otitis media and pneumonia) and systemic (sepsis and meningitis) diseases. Due to increasing rates of antibiotic resistance, there is an urgent need to improve prevention of pneumococcal disease. Two currently licensed vaccines have been successful in reducing pneumococcal disease, but there are limitations with their use and effectiveness. Another approach for prevention is the use of live attenuated vaccines. Here we investigate the safety and protection induced by live attenuated strains of S. pneumoniae containing combinations of deletions in genes encoding three of its major virulence determinants: capsular polysaccharide (cps), pneumolysin (ply), and pneumococcal surface protein A (pspA). Both the cps and ply/pspA mutants of a virulent type 6A isolate were significantly attenuated in a mouse model of sepsis. These attenuated strains retained the ability to colonize the upper respiratory tract. A single intranasal administration of live attenuated vaccine without adjuvant was sufficient to induce both systemic and mucosal protection from challenge with a high dose of the parent strain. Immunization with cps mutants demonstrated cross-protective immunity following challenge with a distantly related isolate. Serum and mucosal antibody titers were significantly increased in mice immunized with the vaccine strains, and this antibody is required for full protection, as μMT mice, which do not make functional, specific antibody, were not protected by immunization with vaccine strains. Thus, colonization by live attenuated S. pneumoniae is a potentially safe and less complex vaccine strategy that may offer broad protection.

scholarly journals Experimental Human Challenge Reveals Distinct Mechanisms of Acquisition or Protection Against Pneumococcal Colonization

2018 ◽  
Author(s):  
Elissavet Nikolaou ◽  
Simon P. Jochems ◽  
Elena Mitsi ◽  
Sherin Pojar ◽  
Edessa Negera ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Immune Responses ◽  
Sampling Method ◽  
Invasive Disease ◽  
Upper Respiratory Tract ◽  
Post Challenge ◽  
Upper Respiratory ◽  
Density Dynamics ◽  
Human Immune ◽  
Pneumococcal Colonization

AbstractColonization of the upper respiratory tract with Streptococcus pneumoniae is the precursor of pneumococcal pneumonia and invasive disease. Following exposure, however, it is unclear which human immune mechanisms determine whether a pathogen will colonize. We used a human challenge model to investigate host-pathogen interactions in the first hours and days following intranasal exposure to Streptococcus pneumoniae. Using a novel home sampling method, we measured early immune responses and bacterial density dynamics in the nose and saliva after pneumococcal exposure. We found that nasal colonization can take up to 24 hours to become established. Also, two distinct bacterial clearance profiles were associated with protection: nasal clearers with immediate clearance of bacteria in the nose by the activity of pre-existent mucosal neutrophils and saliva clearers with detectable pneumococcus in saliva at one-hour post challenge and delayed clearance mediated by an inflammatory response and increased neutrophil activity 24 hours post bacterial encounter.

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